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https://openalex.org/W4390650199
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https://jurnal.iaii.or.id/index.php/RESTI/article/download/5558/883
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English
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The Image Extraction Using the HSV Method to Determine the Maturity Level of Palm Oil Fruit with the k-nearest Neighbor Algorithm
|
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi)
| 2,023
|
cc-by
| 7,515
|
1. Introduction goes to the processing section will not meet the maturity
criteria for palm oil fruit. Of course, this will reduce the
quality of the fruit oil and be detrimental to the
company. Oil palm (Elaeis guineensis Jacq.) is an oil-producing
monocotyledonous plant, Indonesia's main commodity
product. Indonesia, as the largest exporter of Crude
Palm Oil (CPO) in the world, always maintains and
follows international palm oil fruit standards so that
Indonesian CPO products are much sought after by the
international food and non-food industry. To be able to
compete in domestic and global markets, farmers and
palm oil mills (POM) must pay attention to several
aspects, including maintenance, cultivation techniques,
and post-harvest handling [1]. There are systems that can help sorters identify ripeness
of palm fruit, one of which is by utilizing digital image
processing and pattern recognition. The image of the oil
palm fruit will later be processed on a computer in the
form of a digital image. One of the most widely used and successfully used
methods is the Hue Saturation value (HSV) [2]–[6]. In
this study, lime maturity classification was carried out
based on HSV values to obtain lime maturity values [7]. Oil palm cultivation is also widespread on several
islands in Indonesia. One of the palm oil companies in
West Sulawesi is PT. Unggul Widya Lestari
Technologi. In the company's plantation section, the
harvested palm oil fruit will be sorted first before being
taken to the processing section. Sorting is carried out to
distinguish ripe palm fruit from immature ones by
looking objectively at the color of the palm fruit. This
technique is considered to be very manual because it
will take quite a long time considering that the
company's plantation area is not small. This also allows
sorting errors to occur when looking at the color of ripe
and unripe palm fruit. If this happens, then the fruit that Digital image processing techniques are used to
simplify and speed up the process of testing the maturity
level of oil palm fruit which will then be classified using
a classification method, namely the K-Nearest
Neighbor (KNN) algorithm [8]–[10]. K – Nearest
Neighbor (KNN) is a method for classifying objects or
data based on learning data taken from k nearest
neighbors. In research that uses the KNN algorithm to classify lime
ripeness based on color. From the test results, the
accuracy value is 92%. The Image Extraction Using the HSV Method to Determine the Maturity
Level of Palm Oil Fruit with the k-nearest Neighbor Algorithm Mohammad Yazdi Pusadan1, Indah Safitri2, Wirdayanti3
1,2,3Department of Information Technology, Faculty of Engineering, University of Tadulako, Palu, Indonesia
1yazdi.diyanara@gmail.com, 2indahmarvino@gmail.com, 3wirdaida28@gmail.com Abstract The oil palm is one of the monocot oil-producing plants in Indonesia. Sorting errors in oil palm fruit is caused by a sorter error
when distinguishing the color of ripe and immature oil palm fruit. In addition to inefficient time, the area of oil palm plantations
is also a factor causing the sorter to make mistakes in sorting. This study aims to produce a system that can classify oil palm
maturity based on feature extraction of hue, saturation, and value (HSV) color features. The HSV method is used to produce
color characteristics from the image of the oil palm fruit. The classification of oil palm fruit maturity is classified using the K-
Nearest Neighbor (KNN) algorithm with a dataset of 400 oil palm fruit image data with a data sharing ratio of 70% training
data and 30% test data. 280 image data were used as training data which is divided into 140 image data of ripe oil palm fruit
140 image data of immature oil palm fruit and 120 image data of oil palm used as test data which is divided into 60 image data
of ripe oil palm and 45 image data unripe palm oil. Based on the result of tests that have been carried out using a confusion
matrix with varied k values, namely, 5 and 7, the average accuracy is 94.16% Keywords: oil palm; maturity classification; HSV; k-NN; confusion matrix g
Decree of the Director General of Higher Education, Research and Technology, No. 158/E/KPT/2021
Validity period from Volume 5 Number 2 of 2021 to Volume 10 Number 1 of 2026 g
Decree of the Director General of Higher Education, Research and Technology, No. 158/E/KPT/2021
Validity period from Volume 5 Number 2 of 2021 to Volume 10 Number 1 of 2026 1. Introduction For this reason, researchers Accepted: 20-11-2023 | Received in revised: 25-12-2023 | Published: 29-12-2023 Accepted: 20-11-2023 | Received in revised: 25-12-2023 | Published: 29-12-2023 1448 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) conducted research with the title "Image Extraction
Using the HSV Method to Determine the Maturity
Level of Palm Oil Fruit Using the KNN Algorithm at
PT. Unggul Widya Lestari Technologi”. conducted research with the title "Image Extraction
Using the HSV Method to Determine the Maturity
Level of Palm Oil Fruit Using the KNN Algorithm at
PT. Unggul Widya Lestari Technologi”. The aim of this research is to identify oil palm fruit. Research location at PT. Unggul Widya Teknologi
Lestari. 2.4. Software Development In general, the stages of the image extraction system
using the HSV method to determine the maturity level
of oil palm fruit using the k-NN algorithm can be seen
in Figure 1. The urgency of this research is how to implement digital
image processing and the k-Nearest Neighbor (k-NN)
algorithm to classify the maturity of oil palm fruit. Image acquisition is the process of retrieving image data
where the data will be stored in a database. Image
acquisition is carried out by taking pictures using a
camera at a distance of approximately 20-30 cm. From
taking these images, 400 data were obtained, with 200
images of raw coconut fruit and 200 images of ripe oil
palm fruit. The research gaps determined in this research are: i)
Identifying Types of Diseases in Cocoa Using Digital
Image Processing (HSV) & k-NN; ii) Improved
accuracy; iii) determination of the threshold/range; and
iv) framework for a method for detecting the maturity
level of oil palm fruit based on digital image processing. Furthermore, the goal achieved is to have a system for
classifying the maturity of oil palm fruit into ripe and
immature categories using the k-NN algorithm based on
HSV color features. Preprocessing, at this stage image processing is carried
out to produce a better image to be processed to the next
stage which consists of several processes including
cropping, resizing and normalizing the RGB image. Cropping is the process of cutting an image, to remove
parts of the image that are not needed. Resize is used to
normalize the image size so that it has the same size. Meanwhile, RGB image normalization aims to ensure
that the values of each color component can be
compared with each other because the images were
taken under different intensity conditions. 2.2. Data types and sources This research is based on the results of several studies
that have been carried out, namely as follows:
identifying the level of maturity of oil palm fruit. A
method based on thermal imaging using an IR camera. Feature extraction uses color features, namely
Dominant Color Descriptor (DWD) and color moment. The results of the DWD and Color Moment features
become input for the classification process using the K-
Nearest Neighbor (KNN) method[1], [11], [12]. The type of data in this research is primary data. Primary data is data obtained directly from parties who
need the data. The data source in this research is a
picture of oil palm fruit taken directly by the researcher. 2.3. Data collection technique Data collection is a stage that aims to obtain
information or data related to research. Three data
collection methods were used in data collection,
namely: image acquisition. Image acquisition is carried
out by taking pictures using a camera at a distance of
approximately 20-30 cm. From taking these images,
400 data were obtained, with 200 images of raw coconut
fruit and 200 images of ripe oil palm fruit. Classification of palm oil maturity can be done using
the Artificial Neural Network (ANN) method[13]. To
extract images using RGB and HSV where the results
of image extraction will become image weight values
which are used as a database for creating ANN which
will then classify oil palm maturity. After image acquisition is obtained, the dataset will
then be divided into training data and test data, then the
Hold out method is used to divide the data into training
data and test data, namely: 70% and 30%. Where the
training data used is 280, consisting of 140 images of
unripe fruit and 140 images of ripe fruit. The test data
used was 120 data consisting of 60 images of unripe
fruit and 60 images of ripe fruit. The k-Nearest Neighbor algorithm can be used to
classify the ripeness level of orange fruit based on color
features[7]. The resulting accuracy[11], [14]–[16] is
92%. Hue Saturation Value (HSV) [2], [12], [15], [17]–[22]
in classifying Philodendron leaf ornamental plants is
used to extract images through color features. Apart
from that, other classification models can be used using
the Naïve Bayes method. The resulting accuracy
reached 90.48%. 2. Research Methods This research uses a qualitative research design. This
research emphasizes the aspect of in-depth data in order
to obtain quality research results. This research relies on
descriptive descriptions of words or sentences that are
arranged carefully and systematically starting from data
collection to reporting research results through
interviews and observations. Then it is hoped that it will
be able to produce a presentation of accuracy values
above 75%. Feature extraction, at this stage extracts object features
that can differentiate them from other objects. In this
research, HSV color feature extraction is used, the
resulting values of which become input values for the
image classification process. 2.1. Object and locations DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0) 1449 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) image acquisition
Preprocessing
color extraction feature
Classifcation
Image cropping
Resize
RGB Image
Normalization
RGB Image
Normalization
Hue Saturation Value
(HSV) Feature
Taking pictures of
palm fruit using a
smartphone
K-Nearest Neighbor
Algorithm
Figure 1. Stages of software development fo color features using HSDV Preprocessing Classifcation K-Nearest Neighbor
Algorithm Resize Figure 1. Stages of software development fo color features using HSDV To obtain HSV feature values, previously the RGB
image was transformed to HSV using the Formula (1 to
7). The feature extraction stages can be seen in Figure 2. RGB
Normalization
Tranformation
RGB to HSV
HSV Features
Figure 1. Feature Extraction Stages The feature extraction stages can be seen in Figure 2. RGB
Normalization
Tranformation
RGB to HSV
HSV Features
Figure 1. Feature Extraction Stages The feature extraction stages can be seen in Figure 2. RGB
Normalization
Tranformation
RGB to HSV
HSV Features
Figure 1. Feature Extraction Stages To get color feature values, start by converting an RGB
image to an HSV image. Before an RGB image is
converted into an HSV image, the RGB value needs to
be normalized first using Formula (1 to 3). Figure 1. Feature Extraction Stages Classification, at this stage the algorithm is able to
classify objects into existing classes based on the
characteristics of the object's features using the k-NN
algorithm. The class categories are raw and cooked. 2. Research Methods 𝑟 =
𝑅
𝑅+𝐺+𝐵
(1)
𝑔=
𝐺
𝑅+𝐺+𝐵
(2)
𝑏=
𝐵
𝑅+𝐺+𝐵
(3) (1) Classification, at this stage the algorithm is able to
classify objects into existing classes based on the
characteristics of the object's features using the k-NN
algorithm. The class categories are raw and cooked. Testing, implementation of the K-NN algorithm was
carried out using testing data and direct testing on
images of oil palm fruit located on PT's oil palm
plantations PT. Unggul Widya Teknologi Lestari. Testing was carried out by comparing the results of
system implementation and test results from the
Plantation and Livestock Service of Pasangkayu
Regency. The accuracy of this system will be tested
using a confusion matrix [23]–[25]. The confusion
matrix can be seen in Table 1. (3) After normalization, the representation of the RGB to
HSV conversion calculation can be seen in Formula (4
to 7). After normalization, the representation of the RGB to
HSV conversion calculation can be seen in Formula (4
to 7). 𝑉= max(𝑟, 𝑔, 𝑏)
(4)
𝑆= {
0, 𝑗𝑖𝑘𝑎 𝑉= 0
1 −
min(𝑟,𝑔,𝑏)
𝑉
, 𝑉> 0
(5)
𝐻=
{
0, 𝑗𝑖𝑘𝑎 𝑆= 0
60° ∗
(𝑔−𝑏)
𝑆∗𝑉, 𝑗𝑖𝑘𝑎 𝑉= 𝑟
60°∗[2+𝑏−𝑟
𝑆∗𝑉],𝑗𝑖𝑘𝑎 𝑉=𝑔
60°∗[4+𝑟−𝑔
𝑆∗𝑉],𝑗𝑖𝑘𝑎 𝑉=𝑏
}
(6)
𝐻= 𝐻+ 360 𝑗𝑖𝑘𝑎 𝐻< 0
(7) 𝑉= max(𝑟, 𝑔, 𝑏) 𝑉= max(𝑟, 𝑔, 𝑏) (4) (6) Table 1. Confusion matrix DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0)
1450
𝐻=
{
𝑆∗𝑉
𝑗
60°∗[2+𝑏−𝑟
𝑆∗𝑉],𝑗𝑖𝑘𝑎 𝑉=𝑔
60°∗[4+𝑟−𝑔
𝑆∗𝑉],𝑗𝑖𝑘𝑎 𝑉=𝑏
}
(6)
𝐻= 𝐻+ 360 𝑗𝑖𝑘𝑎 𝐻< 0
(7)
V is the Maximum value (r,g,b), S is the Saturation
value, H = Hue value. Table 1. Confusion matrix
Confusion Matrix
2 Class
Classification
Positif
Negatif
Class
Target
Positif
True Positif (TP)
False Positif (FP)
Negatif
False Negatif (FN)
True Negatif (TN)
Literature Review
Collecting Data
Design
Implementation
Input
Testing
Clasification
success
Result
START
END
Figure 2. Research stages 𝐻= 𝐻+ 360 𝑗𝑖𝑘𝑎 𝐻< 0 (7) V is the Maximum value (r,g,b), S is the Saturation
value, H = Hue value. V is the Maximum value (r,g,b), S is the Saturation
value, H = Hue value. V is the Maximum value (r,g,b), S is the Saturation
value, H = Hue value. DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0)
1450
Literature Review
Collecting Data
Design
Implementation
Input
Testing
Clasification
success
Result
START
END
Figure 2. 2. Research Methods Research stages Clasification
success DOI: https://doi.org/10.29207/resti.v7i6.5558 DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0) p
g
Creative Commons Attribution 4.0 International License (CC BY 4.0) 1450 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Var reader = new FileReader();
Reader.onload = function€ {
$uploadCrop.croppie(‘bind’, {
url: e.target.result
}).then(function(){
Console.log(‘jQuery bind complete’);
});
} Var reader = new FileReader();
Reader.onload = function€ {
$uploadCrop.croppie(‘bind’, {
url: e.target.result
}).then(function(){
Console.log(‘jQuery bind complete’);
});
} Result, in this stage, conclusions are drawn in the form
of final results which are expected to answer the
research objectives, namely an image extraction system
using the HSV method to determine the level of
maturity of oil palm fruit using the k-NN algorithm. Based on the research stages above, the following
research flow diagram is in Figure 3. These stages are a
series of processes that take place sequentially. The
accuracy value obtained in the test results affects the
amount (n) of data used in the training data and test data. }
Reader.readAsDataURL(this.files[0]);
$(‘#uploadimageModal’).modal(‘show’); }); Cropping is the process of cutting an image at certain
coordinates in the image area. To cut part of an image,
two coordinates are used, namely the initial coordinate
which is the initial coordinate of the image being cut,
and the final coordinate which is the final coordinate of
the image being cut. So, it will form a rectangular shape
where each pixel in a certain coordinate area will be
saved in the new image. The results of pruning before
and after images of oil palm fruit can be seen in Figure
5. 3. Results and Discussions Process of inputting training data for the Palm Oil
classification system DOI: https://doi.org/10.29207/resti.v7i6.5558
e Commons Attribution 4.0 International License (CC BY 4.0)
1451
aining data for the Palm Oil
n system
mages that will be used as
rocess can be seen in the
function() {
public function post()
{
$start = microtime(true);
ini_set('max_execution_time', 300);
$this->load->
model('tambah_data_latih_model');
$file_gambar = $_POST['image'];
list($type,$file_gambar)=explode(';',
$file_gambar); .29207/resti.v7i6.5558
International License (CC BY 4.0)
51
public function post()
{
$start = microtime(true);
ini_set('max_execution_time', 300);
$this->load->
model('tambah_data_latih_model');
$file_gambar = $_POST['image'];
list($type,$file_gambar)=explode(';',
$file_gambar); public function post()
{ 3. Results and Discussions The data used are 400 images of mature and immature
oil palm fruit which will be divided into training data
and test data. Oil palm image data collection was carried
out directly at the oil palm plantation using a
smartphone camera in a portrait position. Training Data, in the training process the researcher
used images of oil palm fruit taken using a smartphone
camera. After that, a cropping process is carried out
which only focuses on the image of the oil palm fruit. Next, a resizing process will be carried out to make the
image size uniform. After
that, RGB image
normalization is carried out so that the values for each
color component can be compared with each other
because the images were taken in different intensity
conditions, after that the RGB image transformation
process to HSV is carried out, then the Hue, Saturation,
Value feature extraction is carried out, then it will be
continued to K-NN algorithm calculation stage. Figure 4. Image Cropping Process Data input, the first process carried out is inputting
training data. Where the data is obtained using a
smartphone camera, then the image is transferred to the
computer to be uploaded via the selected file. The data
used in training training data is 280 image data
consisting of 140 images of ripe fruit and 140 images of
immature fruit. The data input process can be seen in
Figure 4. Figure 4. Image Cropping Process Feature extraction, plays an important role in
distinguishing object types. The characteristics that are
processed are in the form of values that can be used to
differentiate
one
object
from
other
objects. Characteristics are expressed in a series of numbers
which can be used to identify the characteristics of the
object to be processed using color feature extraction
obtained from the resize results using the Hue,
Saturation, Value feature extraction method. The
procedure for extracting HSV features which will be
used as identical parameters for each oil palm fruit
image can be seen in the algorithm. Figure 3. 3.2. Database Implementation The tables contained in the "kelapa_sawit" database
used in the oil palm fruit maturity classification system
are as follows. The training data table is shown in Table
2. Table 2. Table of data training
field
type
length/ values
index
id
integer
4
primary key
image
varchar
64
-
H
float
-
-
S
float
-
-
V
float
-
-
Euclidean
float
-
-
category
varchar
-
-
Next, the results data table is shown in Table 3. Table 2. Table of data training
field
type
length/ values
index
id
integer
4
primary key
image
varchar
64
-
H
float
-
-
S
float
-
-
V
float
-
-
Euclidean
float
-
-
category
varchar
-
-
Next, the results data table is shown in Table 3. Next, the results data table is shown in Table 3. Table 3. Table of result
field
type
length/ values
index
id
integer
4
primary key
image
varchar
64
-
h
float
5
-
s
float
5
-
v
float
5
-
Euclidean
float
5
-
category
varchar
4
- }
$time_elapsed_secs = microtime(true) - $start;
echo json_encode(array(
"yuhui" => mime_content_type($file_gambar),
"yuhui" => $imageName,
"h" => $H, "s" => $S, 'v' => $V, "waktu"=>
$time_elapsed_secs
));
} The k-NN algorithm uses the k value parameter to
determine the decision result. Then calculate the
distance between the new data and all the data in the
training data and then determine the nearest neighbors
based on the k-th minimum distance, which uses the
category that has the highest frequency of nearest
neighbors, as the prediction value or classification
result of the new data. The K-NN algorithm procedure
used in the oil palm fruit maturity classification system
can be seen in the following figure. From the tables above, the relationship between entities
can be described as Entity Relationship (E-R) Diagram,
in Figure 6. E-R Diagram is needed to map the
relationships that occur in each entity. There are several
attributes and key attributes involved in each entity and
entity relationship. From the tables above, the relationship between entities
can be described as Entity Relationship (E-R) Diagram,
in Figure 6. E-R Diagram is needed to map the
relationships that occur in each entity. There are several
attributes and key attributes involved in each entity and
entity relationship. DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0)
an be seen in the following figure. Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) list(,$file_gambar)=explode(',',
$file_gambar);
$file_gambar = base64_decode($file_gambar);
$imageName = time().'.png';
file_put_contents('upload/'.$imageName,
$file_gambar);
$file_gambar = realpath('upload/'.$imageName);
$this->resize_img($file_gambar);
$im = imagecreatefromjpeg($file_gambar);
list($width,$height)=
getimagesize($file_gambar);
$mtx = array();
for ($i=0; $i < $width; $i++) {
for ($j=0; $j < $height; $j++) {
$rgb = imagecolorat($im, $i, $j);
$colors = imagecolorsforindex($im, $rgb);
$mtx[$i][$j]=$this-
>Tambah_data_latih_model-
>rgb_to_hsv($colors['red'],
$colors['green'],$colors['blue']);
}
}
$H = 0;
for ($i=0; $i < count($mtx); $i++) {
for ($j=0; $j < count($mtx); $j++) {
$H += $mtx[$i][$j]['H'];
}
}
$S = 0;
for ($i=0; $i < count($mtx); $i++) {
for ($j=0; $j < count($mtx); $j++) {
$S += $mtx[$i][$j]['S'];
}
}
$V = 0;
for ($i=0; $i < count($mtx); $i++) {
for ($j=0; $j < count($mtx); $j++) {
$V += $mtx[$i][$j]['V'];
}
}
$time_elapsed_secs = microtime(true) - $start;
echo json_encode(array(
"yuhui" => mime_content_type($file_gambar),
"yuhui" => $imageName,
"h" => $H, "s" => $S, 'v' => $V, "waktu"=>
$time_elapsed_secs
));
} 3.1. System Implementation System implementation is divided into two, namely:
hardware and software implementation. The hardware
implementation for this oil palm fruit maturity
classification system can run on smartphones with Quad
Core 1.4 GHz specifications, 2 GB RAM, and
computers/laptops
with
hardware
specifications,
namely Intel Core i3 2.00 processor, 1.5 GB RAM. For
a software implementation, it can run on the Google
Chrome web browser as application media. public function post()
{ public function post()
{ Figure 3. Process of inputting training data for the Palm Oil
classification system {
$start = microtime(true);
ini_set('max_execution_time', 300);
$this->load->
model('tambah_data_latih_model');
$file_gambar = $_POST['image']; {
$start = microtime(true);
ini_set('max_execution_time', 300);
$this->load->
model('tambah_data_latih_model');
$file_gambar = $_POST['image']; The procedure for uploading images that will be used as
input for the training data process can be seen in the
algorithm. list($type,$file_gambar)=explode(';',
$file_gambar); Prosedur Upload Citra
$(‘#upload’).on(‘change’, function() { DOI: https://doi.org/10.29207/resti.v7i6.5558
ative Commons Attribution 4.0 International License (CC BY 4.0) 1451 3.4. System Testing The test is divided into several parts, namely testing the
system function using black box testing, then the second
test is carried out to measure the accuracy of the palm
oil maturity classification system using the colour
feature extraction method with the K-NN algorithm. Next, testing is carried out on the testing data to
calculate the confusion matrix so that the accuracy and
error of the system can be determined. The following
are several stages of system testing. Figure 7. The form to add data test 3.3. Software Implementation The software implementation of the oil palm fruit
maturity classification system has several forms,
including: The main form (login), this palm oil maturity
classification system will be used in several stages, the
first step is that the sorter will log in first. In the login
form the sorter will enter the username and password
that the sorter already knows. The login form displays
on the laptop/computer browser and the login form
displays on the smartphone and can be seen in Figure 7. Figure 8. The form to view data training Figure 8. The form to view data training Detection Form, this form is used to detect images of
oil palm fruit that will be uploaded and get detection
results whether they are in the ripe or immature
category. The display of the detection form on the
laptop/computer and the display on the smartphone can
be seen in Figure 10. Username and password authentication is carried out by
validating session security—login checking based on
the login table contained in the admin database. The
admin can add several users who will function as
system admins. Figure 6. System login Figure 6. System login Figure 9. The menu of detection Figure 6. System login Figure 6. System login Figure 6. System login Figure 9. The menu of detection Add test data form, this form is used to add test data for
images of ripe and immature oil palm fruit by uploading
images via the browser gallery on a laptop or via
smartphone. The display of the form for adding test data
on the laptop/computer browser and the display of the
form on the smartphone can be seen in Figure 8. Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Figure 8. The form to view data training Figure 8. The form to view data training 3.2. Database Implementation data_training
result
produce
s
v
euclidean
category
id_training
image
h
id_result
id_training
s
v
euclidean
category
image
h
id_result
Figure 5. E-R Diagram classification system of palm oil id_result id_training image image category data_training produce result id_training id_result euclidean Figure 5. E-R Diagram classification system of palm oil DOI: https://doi.org/10.29207/resti.v7i6.5558 DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0) 1452 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) 3.4.1. Blackbox testing The range of feature values for the H image
of oil palm fruit ranges from a minimum value of
6599.3 to a maximum value of 50410.3. The S feature
value range ranges from 10207.2 to a maximum value
of 50401. And the range of feature V values ranges from
17418.9 to a maximum value of 47576.5. The range of
values resulting from oil palm fruit image extraction
and the overall value range for the Ripe and Unripe
categories can be seen in Table 5. Table 5. Range of Extraction Values for Palm Oil Image Color
Features
Colour Features
Min
Max
H
6599.3
50410.3
S
10207.2
50401
V`
17418.9
47576.5
Based on category and HSV value, it is shown in Table
6. Table 6. Overall Value Range for Mature and Immature Categories Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Figure 10. Matrix Confusion Matrix Test Results No. Tested
Functions
System Expectations
Test result
(Training
Data Form)
3. Test Data
Input (Test
Data Form)
It can be entered test data
Successfully
4. Detailing,
updating,
deleting
training data
(view Data
Form)
It can be display details,
can updating and delete
training data
Successfully
5. Oil palm
fruit
detection
process
(Detection
System
Form)
It can be found out the
results of detecting oil
palm fruit
Successfully
6. Palm Oil
Fruit Image
Capture
Function
It can be capture images
of oil palm fruit
Successfully
7. Function for
Uploading
Images of
Palm Oil
Fruit
It can be uploading images
of oil palm fruit
Successfully
8. Cropping
process
It can be cut images of oil
palm fruit
Successfully
3.4.2. Confusion matrix testing Figure 10. Matrix Confusion Matrix Test Results 3.5. Extraction results of oil palm fruit colour
characteristics Extraction
of
Hue
Saturation
Value
colour
characteristics in oil palm fruit images based on
equations (1) to (7) produces feature values from the
colour characteristics of oil palm fruit, both ripe and
immature. The range of feature values for the H image
of oil palm fruit ranges from a minimum value of
6599.3 to a maximum value of 50410.3. The S feature
value range ranges from 10207.2 to a maximum value
of 50401. And the range of feature V values ranges from
17418.9 to a maximum value of 47576.5. 3.4.1. Blackbox testing System function testing is carried out using black box
testing. To test system functions, see table 4. Table 4. Test Results with Black box
No. Tested
Functions
System Expectations
Test result
1. Login
Verification
(Login Form
It can be verifying the
login, that is, if the
username and password
entered are correct then
the sorter can enter the
main menu, if incorrect
then a message will appear
to log in again. Successfully
2. Input
Training
Data
It can be input training
data
Successfully Figure 7. The form to add data test Training data view form, this form is used to view palm
oil fruit image data that has been previously uploaded
in the training data form by the sorter and stored in the
“kelapa_sawit” database and table of training data. In
this form, you can also see details of image data that has
been uploaded, change and delete image data and can
be seen in Figure 9. 2. Input
Training
Data DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0) 1453 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023)
No. Tested
Functions
System Expectations
Test result
(Training
Data Form)
3. Test Data
Input (Test
Data Form)
It can be entered test data
Successfully
4. Detailing,
updating,
deleting
training data
(view Data
Form)
It can be display details,
can updating and delete
training data
Successfully
5. Oil palm
fruit
detection
process
(Detection
System
Form)
It can be found out the
results of detecting oil
palm fruit
Successfully
6. Palm Oil
Fruit Image
Capture
Function
It can be capture images
of oil palm fruit
Successfully
7. Function for
Uploading
Images of
Palm Oil
Fruit
It can be uploading images
of oil palm fruit
Successfully
8. Cropping
process
It can be cut images of oil
palm fruit
Successfully
3.4.2. Confusion matrix testing
This test is carried out to obtain accuracy and error
rate. Accuracy aims to add up the results of incorrect
predictions. The Confusion Matrix test results can be
seen in Figure 11. Figure 10. Matrix Confusion Matrix Test Results
3.5. Extraction results of oil palm fruit colour
characteristics
Extraction
of
Hue
Saturation
Value
colour
characteristics in oil palm fruit images based on
equations (1) to (7) produces feature values from the
colour characteristics of oil palm fruit, both ripe and
immature. 3.4.1. Blackbox testing The range of
values resulting from oil palm fruit image extraction
and the overall value range for the Ripe and Unripe
categories can be seen in Table 5. Table 5. Range of Extraction Values for Palm Oil Image Color
Features
Colour Features
Min
Max
H
6599.3
50410.3
S
10207.2
50401
V`
17418.9
47576.5
Based on category and HSV value, it is shown in Table
6. Table 5. Range of Extraction Values for Palm Oil Image Color
Features 3.4.2. Confusion matrix testing This test is carried out to obtain accuracy and error
rate. Accuracy aims to add up the results of incorrect
predictions. The Confusion Matrix test results can be
seen in Figure 11. Based on category and HSV value, it is shown in Table
6. Table 6. Overall Value Range for Mature and Immature Categories Table 6. Overall Value Range for Mature and Immature Categories
Category
H
S
V
MIN
MAX
MIN
MAX
MIN
MAX
Ripe
6599
50410
11749
50401
17418
47576
Immature
14005
47178
10207
27011
17434
36892
Min
6599.3
10207.2
17418.9
Max
50410.3
50401
47576.5 Table 7. Confusion Matrix with value k = 3 Table 7. Confusion Matrix with value k = 3
Confusion Matrix
Palm oil
Classification
Ripe
Immature
Class
Target
Ripe
TP =60
FP = 0
Immature
FN =0
TN = 60
To show the value of the confusion matrix with a value 3.6. Test results of the k-NN algorithm with various
values Confusion Matrix
Palm oil
Classification
Ripe
Immature
Class
Target
Ripe
TP =60
FP = 0
Immature
FN =0
TN = 60 This test uses 120 test data, where the test data is
divided into 60 images of mature oil palm and 60
images of immature oil palm. Tests were carried out at
the Pasangkayu Regency Plantation Service, where the
original test data images were validated by experts on
the ripeness of oil palm fruit, and then the results were
compared with the results of implementing the k-NN
algorithm. The k values used are k=3, k=5 and k=7. The
following is a table of confusion matrix test results with
various k. To show the value of the confusion matrix with a value
of k=5, it is in Table 8. Table 8. Confusion Matrix with value k = 5 Table 8. 4. Conclusion Based on the results of testing and analysis of the palm
oil fruit maturity classification system, it can be
concluded that the palm oil fruit maturity classification
system is capable or capable of classifying oil palm fruit
into mature and immature categories. This was done
using the K-NN algorithm based on HSV color features
with an average accuracy value of 94.16%. From the
research results, it can be seen that there are still
development needs, namely: i) the need for additional
data for training data and test data. So that the system
can be more accurate in classifying the maturity of oil
palm fruit; ii) other algorithms besides the K-NN
algorithm can be used because in this study it cannot
handle missing values; iii) there is a need for additional
object recognition methods that can differentiate one
object from other objects; and iv) there is a need for
testing in image extraction using only H, V and S
values. [9] [9]
S. Subudhiray, H. K. Palo, and N. Das, “K-nearest neighbor
based facial emotion recognition using effective features,”
IAES Int. J. Artif. Intell., vol. 12, no. 1, p. 57, 2023, doi:
10.11591/ijai.v12.i1.pp57-65. [10] A. Bokhare and P. Jha, “Machine learning models applied in
analyzing breast cancer classification accuracy,” IAES Int. J. Artif. Intell., vol. 12, no. 3, pp. 1370–1377, 2023, doi:
10.11591/ijai.v12.i3.pp1370-1377. [11] M. H. Hanafi, N. Fadillah, and A. Insan, “Optimasi Algoritma
K-Nearest Neighbor untuk Klasifikasi Tingkat Kematangan
Buah Alpukat Berdasarkan Warna,” It J. Res. Dev., vol. 4, no. 1, pp. 10–18, 2019, doi: 10.25299/itjrd.2019.vol4(1).2477. [12] Z. D. Lestari, N. Nafi’iyah, and P. H. Susilo, “Sistem
Klasifikasi Jenis Pisang Berdasarkan Ciri Warna HSV
Menggunakan Metode K-NN,” Semin. Nas. Teknol. Inf. dan
Komun., pp. 11–15, 2019. [13] S. Restiana and A. S. Rini, “Komunikasi Fisika Indonesia
PEROVSKITE MENGGUNAKAN VESTA,” vol. 15, no. 01,
pp. 46–50, 2018. [14] D. Yulianto, R. N. Whidhiasih, and M. Maimunah, “Klasifikasi
Tahap Kematangan Pisang Ambon Berdasarkan Warna
Menggunakan Naive Bayes,” PIKSEL Penelit. Ilmu Komput. Sist. Embed. Log., vol. 5, no. 2, pp. 60–67, 2018, doi:
10.33558/piksel.v5i2.268. y
Table 10. Percentage of confusion matrix test results y
Table 10. Percentage of confusion matrix test results
confusion matrices
Value K
K =3
K =5
K = 7
Accuracy
100%
92.50%
90.00%
Error Rate
0%
15.56%
10.00%
Average Accuracy: 94.16%
Average Error Rate: 8.52%
[
[ I. Transformation, W. Lung, I. Thresholding, and S. Method,
“Jurnal resti,” vol. 5, no. 158, pp. 278–285, 2023. I. Transformation, W. Lung, I. Thresholding, and S. Method,
“Jurnal resti,” vol. 5, no. 158, pp. 278–285, 2023. I. Ilhamsyah, A. Y. Rahman, and I. Istiadi, “Klasifikasi
Kualitas Biji Kopi Menggunakan MultilayerPerceptron
Berbasis Fitur Warna LCH,” J. RESTI (Rekayasa Sist. dan
Teknol. Informasi), vol. 5, no. 6, pp. 1008–1017, 2021, doi:
10.29207/resti.v5i6.3438. Based on the research results, this palm oil fruit
maturity classification system can classify oil palm fruit
into ripe and immature categories using the K-NN
algorithm based on HSV color features with an average
accuracy value of 94.16%. [7]
C. Paramita, E. Hari Rachmawanto, C. Atika Sari, and D. R. Ignatius Moses Setiadi, “Klasifikasi Jeruk Nipis Terhadap
Tingkat Kematangan Buah Berdasarkan Fitur Warna
Menggunakan K-Nearest Neighbor,” J. Inform. J. Pengemb. IT, vol. 4, no. 1, pp. 1–6, 2019, doi: 10.30591/jpit.v4i1.1267. [8] pp
jp
[8]
Mohammad Yazdi Pusadan, Syahrullah, Merry, and Ahmad
Imam Abdullah, “k-Nearest Neighbor and Feature Extraction
on Detection of Pest and Diseases of Cocoa,” J. RESTI
(Rekayasa Sist. dan Teknol. Informasi), vol. 6, no. 3, pp. 471–
480, 2022, doi: 10.29207/resti.v6i3.4064. 3.4.1. Blackbox testing Confusion Matrix with value k = 5
Confusion Matrix
Palm oil
Classification
Ripe
Ripe
Class
Target
Ripe
TP = 52
FP = 8
Immature
FN =1
TN = 59
Next, the confusion matrix value with k=7 is in table 9. To show the value of the confusion matrix with a value
of k=3, it is in Table 7. Next, the confusion matrix value with k=7 is in table 9. DOI: https://doi.org/10.29207/resti.v7i6.5558
Creative Commons Attribution 4.0 International License (CC BY 4.0) 1454 Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Table 9. Confusion Matrix with value k =7
Classification
Ripe
Classification
Ripe
Ripe
Class
Target
Ripe
TP = 50
FP = 10
Immature
FN =2
TN = 58 Table 9. Confusion Matrix with value k =7 infrastructure as well as several supporting devices. The
author also thanks several parties who supported the
availability of data during research activities. The party
involved is PT. Unggul Widya Teknologi Lestari, as
one of the Palm Oil companies in West Sulawesi
Province. TP (True Positif) indicates that palm oil in the mature
category is detected as mature, TN (True Negatif)
indicates that palm oil in the immature category is
detected as immature, FP (False Positive) indicates
mature palm oil that is detected as immature, FN False
Negative) indicates immature palm oil that is detected
as mature Reference Hanafi, N. Fadillah, and A. Insan, “Optimasi Algoritma
K-Nearest Neighbor untuk Klasifikasi Tingkat Kematangan
Buah Alpukat Berdasarkan Warna,” It J. Res. Dev., vol. 4, no. 1, pp. 10–18, 2019, doi: 10.25299/itjrd.2019.vol4(1).2477. [12] Z. D. Lestari, N. Nafi’iyah, and P. H. Susilo, “Sistem
Klasifikasi Jenis Pisang Berdasarkan Ciri Warna HSV
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Komun., pp. 11–15, 2019. [13] S. Restiana and A. S. Rini, “Komunikasi Fisika Indonesia
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h
i
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k [2]
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j p
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A. O. Salau, S. Jain, and J. N. Eneh, “A review of various
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TP+TN
TP+TN+FP+FN X 100%
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TP+TN+FP+FN X 100%
Error rate = 100 −Accuracy Accuracy =
TP+TN
TP+TN+FP+FN X 100%
Error rate = 100 −Accuracy Accuracy =
TP+TN
TP+TN+FP+FN X 100%
Error rate = 100 −Accuracy j
pp
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assisted
in
providing
laboratory
facilities
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Jurnal RESTI (Rekayasa Sistem dan Teknologi Informasi) Vol. 7 No. 6 (2023) Mohammad Yazdi Pusadan, Indah Safitri, Wirdayanti
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Satisfying Product Features of a Fall Prevention Smartphone App and Potential Users� Willingness to Pay: Web-Based Survey Among Older Adults
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JMIR MHEALTH AND UHEALTH
Rasche et al
Original Paper
Satisfying Product Features of a Fall Prevention Smartphone App
and Potential Users’ Willingness to Pay: Web-Based Survey
Among Older Adults
Peter Rasche1, MSc; Alexander Mertens1, Dr Ing, Dr rer medic; Christopher Brandl1, Dr Ing; Shan Liu2, SD, MD;
Benjamin Buecking3, Dr med; Christopher Bliemel3, Priv Doz, Dr med; Klemens Horst4, Dr med, MHBA; Christian
David Weber4, Dr med; Philipp Lichte4, Priv Doz, Dr med; Matthias Knobe4, MME, MHBA, Priv Doz, Dr med
1
Institute of Industrial Engineering and Ergonomics, Department of Mechanical Engineering, RWTH Aachen University, Aachen, Germany
2
Department of Emergency Medicine, Massachusetts General Hospital, Boston, MA, United States
3
Hand and Reconstructive Surgery, Department of Trauma, University Hospital of Giessen and Marburg, Marburg, Germany
4
Department of Orthopaedic Trauma, University of Aachen Medical Center, RWTH Aachen University, Aachen, Germany
Corresponding Author:
Peter Rasche, MSc
Institute of Industrial Engineering and Ergonomics
Department of Mechanical Engineering
RWTH Aachen University
Bergdriesch 27
Aachen, 52062
Germany
Phone: 49 0241 80 99 ext 477
Fax: 49 2418092131
Email: p.rasche@iaw.rwth-aachen.de
Abstract
Background: Prohibiting falls and fall-related injuries is a major challenge for health care systems worldwide, as a substantial
proportion of falls occur in older adults who are previously known to be either frail or at high risk for falls. Hence, preventive
measures are needed to educate and minimize the risk for falls rather than just minimize older adults’ fall risk. Health apps have
the potential to address this problem, as they enable users to self-assess their individual fall risk.
Objective: The objective of this study was to identify product features of a fall prevention smartphone app, which increase or
decrease users’ satisfaction. In addition, willingness to pay (WTP) was assessed to explore how much revenue such an app could
generate.
Methods: A total of 96 participants completed an open self-selected Web-based survey. Participants answered various questions
regarding health status, subjective and objective fall risk, and technical readiness. Seventeen predefined product features of a fall
prevention smartphone app were evaluated twice: first, according to a functional (product feature is implemented in the app), and
subsequently by a dysfunctional (product feature is not implemented in the app) question. On the basis of the combination of
answers from these 2 questions, the product feature was assigned to a certain category (must-be, attractive, one-dimensional,
indifferent, or questionable product feature). This method is widely used in user-oriented product development and captures users’
expectations of a product and how their satisfaction is influenced by the availability of individual product features.
Results: Five product features were identified to increase users’ acceptance, including (1) a checklist of typical tripping hazards,
(2) an emergency guideline in case of a fall, (3) description of exercises and integrated workout plans that decrease the risk of
falling, (4) inclusion of a continuous workout program, and (5) cost coverage by health insurer. Participants’ WTP was assessed
after all 17 product features were rated and revealed a median monthly payment WTP rate of €5.00 (interquartile range 10.00).
Conclusions: The results show various motivating product features that should be incorporated into a fall prevention smartphone
app. Results reveal aspects that fall prevention and intervention designers should keep in mind to encourage individuals to start
joining their program and facilitate long-term user engagement, resulting in a greater interest in fall risk prevention.
(JMIR Mhealth Uhealth 2018;6(3):e75) doi: 10.2196/mhealth.9467
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KEYWORDS
prevention; cell phone; accidents
Introduction
Background
Falls and fall-related injuries pose a major threat to older adults’
health and are associated with increased morbidity and mortality
[1-3]. Indication from the literature [4,5] suggests that older
adults tend not to be sufficiently aware of their potential for
falls or their fall risk. In addition, the literature [6-9] suggests
that client outcomes vary with the type of treatment prescribed,
the equipment of the clinic, and the health professional’s
abilities. Hence, preventive measures are important for fall
prevention and reduction of associated injuries over time.
One method is enabling older adults to self-assess their possible
fall risk and thereby enable them to become aware of their
potential fall risk [4,10,11]. A promising attempt is to
incorporate health apps in this context, given that the use of
health apps is rising among older adults [12-18]. A health app
is an unobtrusive way to offer potential support in terms of
prevention activities [19-21]. The sensor technology built into
smartphones is precise enough to allow extensive data collection
to record the user's state of health [14,22,23]. Different research
projects have already addressed the topic of fall prevention
using varied approaches [14,24]. The FARSEEING project,
funded by the European Union (EU), developed a smartphone
app to measure users’ fall risk based on daily activities as it
incorporated an adapted version of the Timed-Up and Go test
[14,25]. Users of this product were able to get real-time feedback
regarding their individual fall risk. An intervention or decision
about treatment was not included in this app. The question on
how to decide about the right treatment was investigated by the
ProFouND project, also funded by the EU [26]. Within this
project, an app for health care professionals was developed,
which can help the decision process regarding a certain patient
[27]. A different approach was undertaken by the FallCheck
project at Coventry University [28]. They developed an app
that helps older adults identify typical tripping hazards within
their home [24]. The question on how to motivate and instruct
physical exercises for older adults in their home was investigated
by the iStoppFalls project [29]. This project showed that a
continuous exercise plan could decrease older adults’ fall risk.
Within this project, older adults performed physical exercises
on their own. They were instructed and motivated by exergames
on a television. The correct performance of the exercises was
supervised using information and communication technology
such as activity tracker [29].
However, there has been no app that combines these approaches
into a single product. To design such a fall app, it is necessary
to determine potential users’ expectations regarding such a fall
prevention smartphone app.
Aim of This Study
This study investigates which product features potential users
expect of a fall prevention app and how these features would
contribute to users’ satisfaction with such an app. A Web-based
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survey was performed questioning older adults who were not
participants in a prior fall prevention–related study conducted
by the authors [21,30].
Research Questions
In summary, the main research questions of this study were as
follows:
1.
2.
Which product features should a fall prevention app have,
to increase the likelihood of acceptance by the elderly and
consequently reduce the risk of falls in the elderly
population?
Which product features increase or decrease use of such an
app?
This study aims to provide guidance on how to design a
user-friendly and acceptable fall prevention app for older adults.
Methods
Design
An open, self-selected, Web-based survey was designed to
investigate the research questions. The survey was designed in
German and provided for German-speaking Internet users. A
Web-based survey was used, given it is a suitable method to
reach individuals with particular characteristics or interests in
a short period without any limitations on physical space [31,32].
On the basis of research questions, the aim of this survey was
to collect data regarding expected product features of a fall
prevention app. Expectation was measured using the Kano
technique [33]. This is a preference classification technique to
identify user requirement and expectation during the early
product development stage [34]. Within the health care sector,
this technique is not a well studied and less used approach but
potentially a suitable one to design health care interventions
and services according to users’ needs [35-37].
Investigated Product Features of a Fall Prevention App
For this study, features were identified based on literature of
former fall prevention projects and expert interviews. These
product features are related to 6 different topics, including
detection of a fall risk, decision making about a treatment or
intervention, comfort functions, advice and support functions,
physical exercise advice, and cost coverage by health insurance
companies (refer to Table 1).
Detection
Related to the topic detection, 2 product features were identified
to be relevant: (1) the automatic detection of the risk of falling
through the app during general everyday activities and (2) the
detection during the execution of standardized tests. Both
product features have already been implemented in the
“FARSEEING” project [14,25]. However, it remains to be seen
whether potential users prefer continuous data collection in
everyday life, based on which a fall risk is determined or
whether they prefer to carry out explicit test procedures for
detection.
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Table 1. Investigated product features of a fall prevention app. C: criteria.
Topic
Description
Detection
C1
The app recognizes your fall risk based on a standardized test.
C2
The app automatically detects your risk of falling if you carry your smartphone with you.
Decision making
C3
The app leaves the decision to treat your fall risk to your health care professional.
C4
The app itself decides about the treatment of your fall risk.
Comfort
C5
In addition to the risk of falling, other health data such as medication can be stored in the app.
C6
You can share the results of your fall evaluation with your health care professional or friends and family by email.
C7
The following treatment appointments can be stored in the app.
Advice and support
C8
The app contains a checklist of typical tripping hazards.
C9
The app contains a guideline on how to react in the case of a fall for the falling person and the person who is helping.
Physical exercise
C10
The app includes physical exercises to reduce your risk of falling.
C11
The app includes an ongoing workout program to reduce the risk of falling.
C12
The training integrated into the app is supervised by a therapist.
C13
The training integrated in the app can be adapted to your personal needs (scope of training, exercises, time expenditure, and so on).
C14
Within the integrated training, individual goals can be defined.
C15
New social contacts can be made while using the app.
C16
The training within the app includes playful elements such as awards, rankings, and so on.
Cost coverage
C17
The costs of the app are covered by the health insurance company.
Decision Making
After the detection of a certain fall risk, the second question is
how to manage this risk and which interventions or treatments
could be applied [26,27]. Two potential product features were
included in the study: (1) a health professional decides on the
possible treatment measures based on data collected by the app
and (2) the app itself makes a recommendation for the treatment
of a possible fall risk. Here too, the question arose as to what
would be preferred by potential users.
Comfort
Several product features related to certain comfort of an app
were derived from literature. Product features include additional
data storage, sharing data, and setting reminder for medical
appointments. Mendiola et al identified these features to be
valuable features of health apps [19]. Other health apps included
such functions to increase users’ adherence to the app [38,39].
Advice and Support
Another question within this study was whether potential users
would appreciate a checklist of typical tripping hazards as what
the FallCHeck website offers [24]. Furthermore, the option of
an emergency guideline to guide users’ actions after a fall
incident was included in this investigation [38].
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Physical Exercise
Physical exercise is known to reduce a potential fall risk
[4,40-43]. Hence, the integration of physical exercises in the
fall prevention app seemed to be an important aspect. In this
context, 7 different product features were investigated. First,
including physical exercises itself was questioned. Second,
participants were asked whether they prefer a continuous
exercise plan or not. Third, participants were asked whether
they want to have a therapist to oversee their training such as
what the Otago program includes [42,43]. It was further
questioned whether the training should be adaptable to personal
needs as, for example, types of exercise or time spent on
training. Whether potential users want to set individual training
goals was asked as fifth product feature. This was included as
a study by Schlomann et al implicates older adults to exceed
their abilities in physical training by missing individual training
goals and exercises [44]. Lastly, 2 product features, one
regarding making new social contacts and another regarding
gamification were included in this investigation as both features
were recommended by Mendiola et al to be valued ones in health
apps [19].
Cost Coverage
The last product feature investigated was cost coverage by a
health insurance company. With this characteristic, it should
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be examined whether the general customary assumption of costs
by the health insurance company is desired or presupposed in
the case of a fall prevention app [45].
Kano Technique
Each of the 17 predefined product features was evaluated twice:
first, according to a functional (product feature is implemented
in the app), and subsequently by a dysfunctional (product feature
is not implemented in the app) question. This technique is based
on the Kano model, widely used in the user-oriented product
development realm [35-37].
Both types of questions were asked in succession. Five possible
answers were available for both questions:
•
•
•
•
•
I would be very happy
I take that for granted
I don’t care
I barely accept this
That would annoy me
Through the combination of answers of functional and
dysfunctional questions, the classification of a product feature
was derived, as defined earlier in the section [29]. This technique
differentiates 7 categories.
•
•
•
•
•
•
Must-be (M): These product features are taken for granted
when fulfilled but result in dissatisfaction if they are not
fulfilled.
One-dimensional (O): These product features result in
satisfaction when fulfilled and dissatisfaction when not
fulfilled. These are product features that are spoken and the
ones in which companies compete.
Attractive (A): These product features provide satisfaction
when achieved fully but do not cause dissatisfaction when
not fulfilled. These product features are not expected by a
normal customer and thereby have the potential to please
the customer.
Indifferent (I): These product features refer to aspects that
are neither good nor bad, and they do not result in either
customer satisfaction or customer dissatisfaction.
Reverse (R): These product features refer to a high degree
of achievement, resulting in dissatisfaction and to the fact
that not all customers are alike. For example, some
customers prefer high-tech products, whereas others prefer
the basic model of a product and will be dissatisfied if a
product has too many extra features.
Questionable (Q): Product features in this category should
be reviewed. It is most likely that the questions for this
product feature were not appropriate for the app of the Kano
technique.
Generally, a product feature is assigned to the category most
frequently rated [33]. To verify the results of the encoding by
the Kano technique, 2 different decision rules are available: (1)
category and total strength [46] and (2) the Fong test, if category
and total strength led to no clear categorization [47].
Furthermore, customer satisfaction (CS) coefficients were
calculated for all investigated products. This coefficient is a
measure of whether a product feature can explicitly increase
the satisfaction of the potential user or whether the existing
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product characteristic can only prevent the potential user from
being dissatisfied with the overall product [48,49]. According
to this definition, the CS coefficient is divided into 2
components. One component has a positive sign and describes
whether the satisfaction of the potential user can be increased
beyond an expected level by fulfilling the product characteristic
(CS+). The second component of the CS coefficient has a
negative sign and thus indicates to what extent the satisfaction
of the potential user would fall below an expected level if this
product characteristic is not taken into account in the overall
product (CS−). If the individual component of the CS coefficient
(CS+ or CS−) has an absolute value greater than .5, this
component and thus the CS coefficient of the associated product
characteristic is assumed to be significant [48,49].
Willingness to Pay for a Fall Prevention App
Willingness to pay (WTP) was assessed as monthly payment
[50,51]. A potential fall prevention app should be available in
the common app stores for users to explore use without having
to purchase it (Freemium Business model) [13,50]. Hence,
necessary revenues to develop and maintain the app need to be
generated afterward, meaning monthly payments by a
subscription model or in-app purchases. WTP was studied to
explore how much money potential users would spend on in-app
purchases so that developers could estimate potential revenues
[50]. This topic was addressed after participants rated the 17
product features. Participants were able to enter an amount
between 0 and several hundred euros, including 2 decimals.
Characterizing Participants of This Survey
Measuring Health Status
Participants self-reported known medical conditions and chronic
diseases. Health competency of participants was measured using
an adapted version of the European Health Literacy Scale with
16 items [52]. Corresponding statements were evaluated on a
4-point Likert scale (1=not correct and 4=fully correct).
Subsequently, final score was calculated according to Röthlin
et al [53]. Final score ranges between 0 points and 16 points,
with a high score indicating a high health competency [52].
Quality of life was assessed with the EuroQol Questionnaire
(EQ5D-3L) [54], which is a validated tool for measuring general
health-related quality of life. It consists of 5 items (mobility,
self-care, usual activities, pain or discomfort, and anxiety or
depression), each of which is rated as causing “no problems,”
“some problems,” or “extreme problems.” The EQ5D-3L thus
distinguishes 243 unique health states. Each unique health state
has a utility score which lies within a range between 0 (poor
health) and 1 (perfect health). This single EQ5D-3L summary
index score was used in this study [54].
Measuring Fall Risk
Given that purpose of this study was to investigate the desired
functions of a fall prevention app, measurements to access
participants’ objective and subjective risk of falling were
included as measured by the individual’s history of falls in the
past year [55], the Aachen Falls Prevention Scale (AFPS) [10]
and the short Falls Efficacy Scale-International (FES-I) [56].
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Objective fall risk was determined retrospectively based on the
individual’s history of falls in the past year [55]. Using fall risk
screening criteria, participants reporting ≥2 noninjury falls in
the past year or ≥1 injury fall were categorized as “fallers”;
participants reporting no falls were categorized as “nonfallers”;
the remaining subjects were defined as indifferent [55,57]. On
the basis of the answer whether participants have fallen or not,
detailed questions about the falls and their circumstances were
asked.
participants performed a self-assessment of their subjective fall
risk and their FOF. In addition, participants fulfilled a
standardized questionnaire regarding their technological
readiness. Then participants evaluated the 17 product features
regarding described measurements and entered an amount of
money they would spend to use such an app. Finally, participants
fulfilled the AFRIS questionnaire measuring whether participant
would engage with a prevention program or not.
Subjective fall risk was accessed by 2 aspects: the AFPS and
the FES-I. The AFPS is a self-assessment test containing 3 steps
participants had to perform in this survey [10]. First, participants
answered a self-test containing 10 standardized yes or no
questions (positive criterion≥5 “Yes”). Questions addressed
relevant risk factors derived from several fall risk assessment
tools [10]. Second, participants performed a balance test on
their own. During this test, participants had to position their
feet next to each other and hold this position for at least 10 s
without compensatory movement (positive criterion:
compensatory movement). In the third and final step,
participants rated their “subjective risk of falling” on a 10-point
Likert scale based on the results of the first 2 steps. A score of
more than 5 points on this scale indicates a certain fall risk
(cutoff score >5 points).
Data were collected between September 1 and October 31, 2017.
The questionnaire was programmed and made available on a
website hosted using the Unipark software (QuestBack GmbH,
Cologne, Germany) [62]. The survey was introduced as a study
examining the desired functions of a fall prevention smartphone
app (see Multimedia Appendix 1).
The short FES-I questionnaire was used to assess participants’
Fear of Falling (FOF) [56,58] to investigate whether certain
product features are related to this psychological aspect of
patients’ fall risk. This questionnaire contains 7 items rated on
a 4-point Likert scale (1=not at all concerned to 4=very
concerned). The results of all 7 items are added into a final
score, ranging from 7 (no concern about falling) to 28 (severe
concern about falling) [56].
Measuring Technology Readiness
Technology readiness was included as it might influence the
use of modern information and communication technology as
well as the engagement with these products [59]. It is calculated
based on 12 standardized items which are rated on a 5-point
Likert scale (1=not correct and 5=fully correct). For negatively
formulated items, the scale is converted so that a high point
value corresponds to high technology readiness. Subsequently,
final score is calculated by mean value over all 12 items; thus,
the score ranges between 1 and 5 points [59].
Measuring Attitude Toward a Fall-Related Intervention
Participants’ attitude toward the fictive fall prevention app was
accessed using the Attitudes Falls Related Intervention Scale
(AFRIS) [60,61]. Hence, it was possible to evaluate whether a
participant is generally interested in a fall intervention program
or not. The questionnaire consists of 6 items rated on a 7-point
Likert scale (1=I totally disagree to 7=I totally agree). The
results of all 6 items are summed up to a final score, ranging
from 6 points (no intention) to 42points (absolute intention)
[60,61].
Questionnaire
The questionnaire started by presenting a short description of
the context, followed by demographic questions regarding
participants’ age, educational level, and health status. Next, the
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Data Collection
All participants were informed about the duration of the survey,
data storage, and the leading investigator. Each participant
decided to take part in this survey voluntarily by following the
designated link to the survey. A monetary incentive of €3 per
participant was offered for participation.
The survey was tested properly by 2 independent examiners
with regard to wording and technical functionality. The survey
included 63 items, distributed over 16 different pages.
Participants were able to review their entries per page before
moving on.
Recruitment
Different channels of recruitment were applied to reach a broad
range of potential participants in this open survey. It was avoided
to address existing users of the Aachen Fall Prevention App or
participants of a different fall prevention−related study of the
authors as these participants might have a different opinion
about the design and necessity of features of a fall prevention
smartphone app [21,30]. Further exclusion criteria or screening
questionnaires were not applied. The sampling procedure was
nonprobabilistic, and respondents were selected based on their
voluntary willingness to participate [31].
The Web-based survey was promoted by a Clickworker
advertisement, targeting persons aged older than 60 years [63].
This method of recruitment was chosen because this platform
offers the possibility of providing monetary incentives. Finally,
the link to the open Web-based survey was distributed in a
mailing list for elderly who are regularly taking part in studies
at the Institute of Industrial Engineering and Ergonomics of
RWTH Aachen University, Germany. In all cases, the
recruitment was based on the same text as shown in Multimedia
Appendix 1.
In total, 157 unique individuals visited the website of the
Web-based survey. The identification of different individuals
was performed using the Unipark software based on Internet
Protocol address and cookie function. Of 157, 49 visitors never
started the survey. Nine discontinued completing the survey.
In total, 99 visitors finally participated in the survey and
completed the whole questionnaire. Three of these were
excluded for analysis as attention checkmarks within the
questionnaire showed inappropriate data quality. The
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participation rate was thus 68.8% (108/157), and the completion
rate was 63.1% (99/157). The average duration of completing
the survey was 17 min and 12 s, with a median of 15 min and
13 s.
Statistical Analysis
Data were analyzed with SPSS 22 (IBM, USA) and
MatlabR2017b (The MathWorks, USA). Investigated product
features were assigned to the corresponding category according
to the Kano technique. Furthermore, the category strength and
total strength of each product feature are provided. In case that
applying category and total strength rule resulted in an
indifferent categorization, Fong test was performed . In addition,
CS coefficients were calculated to analyze and prioritize
investigated product features in terms of their contribution to
users’ satisfaction with a fall prevention app.
Ethics Statement
The Ethics Committee at RWTH Aachen Faculty of Medicine
authorized this study and its ethical and legal implications in
its statement EK236/16.
Results
Participants
In total, 96 participants took part in this study. The mean age
was 63.8 years (SD 7.02), and 51% (49/96) were female. All
participants lived autonomously in a flat or house. In all, 29%
(19/96) lived together with their family, 56% (54/96) with their
marriage partner or companion, and 29% (28/96) lived alone.
The level of education varied from minor educational degree
to postsecondary degree.
About 78% (75/96) of all participants stated to use a smartphone;
however, none of the participants had any experience at all with
a smartphone app aiming to prevent falls, and 19% (19/96)
stated to already use health apps.
Health Status
About 64% (62/96) of all participants suffered from a chronic
disease such as high blood pressure (37%, 36/96), back pain
(20%, 20/96), cardiovascular disease (16%, 16/96), or diabetes
(12%, 12/96).
Health literacy varied a median score of 15.00 points
(interquartile range, IQR 4) on a range from 0 to 16 points,
indicating a high qualification and interest in managing personal
health.
Median score of quality of life as measured by the EQ5D-3L
was 0.716 (IQR 0.365) ranging from 0 to 1 and thereby
indicating a good quality of life within the sample.
Fall Risk
Fifty-eight (60%, 58/96) participants stated that they had fallen
within the last year. Furthermore, 31 (32%, 31/96) reported to
have fallen at least once within the last year, and finally, 7
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participants (7%, 7/96) indicated to have fallen between 2 and
3 times within the last year. Seven of these 38 participants, who
had fallen, needed to visit the hospital for medical care as a
direct result of their fall. Hence, 84 participants (87%, 84/96)
were classified as “nonfallers,” and 12 participants (12%, 12/96)
were classified as “fallers.” Fallers are defined as participants
reporting ≥2 noninjury falls in the past year or ≥1 injury fall.
Main reasons for falling were tripping (26%, 25/96), dizziness
(4%, 4/96), and physical weakness (4%, 4/96), whereas
combination of reasons are possible as multiple answers were
allowed.
For 8 participants (8%, 8/96), the self-test (step 1 of the AFPS:
10 standardized questions, positive criterion ≥5 points) was
positive. In contrast, 6 (6%, 6/96) participants did not pass the
balance test (step 2 of the AFPS: balance test, positive criterion:
compensatory movement). After steps 1 and 2 of the AFPS had
been completed, 88 (91%, 88/96) participants estimated their
“subjective risk of falling” to be low (≤5 points), and 8 (8%,
8/96) participants rated their “subjective risk of falling” as high
(>5 points). The overall median value was 2.0 points (IQR 2.0)
on a 10-point Likert scale ranging from 0 to 10 points.
The median FOF was 8.0 points (IQR 2.5) on a scale ranging
from 7 to 28 points, suggesting a low FOF.
Technology Readiness
The median technology readiness was 4.0 points (IQR 0.917)
on a scale ranging from 1 to 5 points, indicating a high
technology readiness.
Attitude Toward a Fall-Related Intervention
Median score for the attitude toward a fall-related intervention
was 24.0 points (IQR 9.5) ranging from 6 points (no intention)
to 42points (absolute intention), indicating moderate intention
to attend a fall intervention program.
Classified Product Features According to Kano
Technique
Table 2 presents the investigated product features assigned to
the corresponding category according to the Kano technique.
Furthermore, the category strength and total strength of each
product feature are provided. The Fong test was performed in
case that category and total strength rule did not lead to a clear
categorization. According to these rules, all product features
were valid categorized.
Figure 1 provides CS coefficients for each product feature. Both
components of the CS coefficient (CS+) and (CS−) are shown
as one bar, whereas darker color indicates (CS+) values.
Willingness to Pay for a Fall Prevention App
One of the last questions asked was how much money
participants would spend per month to use a fall prevention
smartphone app. Results showed a wide variety ranging from
€0 to €80 per month with a median amount to spend €5 per
month (IQR 10; see Figure 2).
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Table 2. Investigates functions of a fall prevention app and their results. C: criteria. N/A: not applicable. Sig: significant categorization according to
Fong test.
Topic
Product feature
Category
Category strength (%)
Total strength (%)
Fong test
C1
Fall risk identification by standardized test
Must-be
60.42
95.80
N/A
C2
Automatically identification of fall risk
Must-be
17.71
94.80
C3
Decision about treatment by health care professional
Must-be
82.29
99.00
C4
Decision about treatment by app
Must-be
50.00
91.70
C5
Additional data storage
Must-be
46.88
99.00
C6
Data sharing via email
Must-be
41.67
97.90
C7
Appointment reminder
Must-be
25.00
97.90
Detection
Decision
Comfort
Advice and support
C8
Checklist of typical stumbling blocks
Attractive
11.46
99.00
Sig
C9
Guideline in case of a fall incident
Attractive
6.25
97.90
Sig
C10
Description of physical exercises to reduce fall risk
Attractive
14.58
100.00
C11
Continuous workout program
Attractive
18.75
100.00
C12
Training integrated is supervised by a therapist
Must-be
8.33
100.00
Sig
C13
Individualization of training within app
Must-be
10.42
97.90
Sig
C14
Define individual training goals
Must-be
0.00
97.90
Sig
C15
Make new social contacts
Must-be
33.33
100.00
C16
Serious gaming elements
Must-be
60.42
97.90
Cost coverage by health insurer
Attractive
33.33
97.90
Physical exercise
Cost coverage
C17
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Figure 1. Customer satisfaction (CS) coefficients of investigated product features.
Figure 2. Histogram for the willingness to pay.
Separate univariate one-way analyses of variance (ANOVA)
revealed no significant effects of the between-subject factors
“age,” “gender,” “education,” “Health Literacy Scale,” “number
of chronic diseases,” “faller or nonfaller,” “FES-I,” “technology
readiness,” or “attitude toward a fall-related intervention” for
the WTP. Separate univariate ANOVAs revealed significant
effects for the WTP if the categorization of the product features
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is treated as between-subject factor. Regarding 4 product
features, significant effects were revealed (decision about
treatment by app, F2,68=3.593, P<.05; description of physical
exercises to reduce fall risk, F1,68=8.964, P<.05; continuous
workout program, F1,68=4.87, P<.05; and make new social
contacts, F1,68=1.124, P<.05). Regarding the feature “decision
of treatment by app,” 8 participants categorized this feature as
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questionable. Within these 8% (8/96), mean amount spent per
month was higher than within the group of participants who
categorized this feature as attractive or must-be one. In case of
the other 3 features, participants who rated these as attractive
ones were also willing to spend a higher median amount of
money to use such an app.
Discussion
Principal Findings
In an exploratory approach, requirements were ascertained
which may influence users’ acceptance of a fall prevention
smartphone app. Seventeen product features were rated
according to the Kano technique. According to the calculated
category and total strengths as well as the Fong tests, all product
features have been validly categorized by the Kano technique.
In total, 12 must-be product features were identified ranging
from “automated detection of a fall risk,” over “storing
additional medical data” within the app up to “letting a health
care professional and the app make decisions about the type of
intervention treatment.” Five remaining product features were
identified as so-called attractive ones. These are features
participants do not expect a fall prevention app to have but
would be attracted to the app if it would have this function.
Product features within this group were mainly related to
offering a physical training program via the app, including a
personalized workout plan and individual goal setting. In
addition, a checklist of typical tripping hazards and an action
guide in case a fall occurs were identified as attractive product
features.
Detailed analysis using CS coefficient calculations revealed
that all except one product feature significantly increased or
prevented loss of users’ satisfaction. The exception was the
product feature “define individual training goals,” as this feature
showed no significant contribution regarding an extent of
satisfaction or dissatisfaction.
Missing fall risk detection, missing consultation of a health care
professional regarding the treatment, and missing serious gaming
aspects within the app were rated highest among negative CS
coefficients and therefore would greatly reduce users’
acceptance (CSdecision by health care professional=−0.92,
CSdetection=−0.82, and CSserious gaming=−0.81). Product features,
for example, cost coverage by health insurance companies, a
continuous workout plan, and instructions for home-based
physical exercises, both aiming to decrease a fall risk, would
significantly increase users’ acceptance and attraction to a fall
prevention smartphone app (CScost coverage=0.67, CSworkout
plan=0.59, and CSexercise instructions=0.57).
These results impressively show how difficult it is to design a
user-accepted fall prevention app as all suggested product
features of a possible app were evaluated as “must-be” or
“attractive” product features. Hence, a fall prevention app would
need numerous features to be developed and implemented. One
reason for the categorization might be that participants had not
ever used a fall prevention smartphone app and thereby desired
as many features as possible. After their first experience, they
might have a more concrete idea of the features they might need.
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Nevertheless, CS coefficients indicate a clear priority order
among investigated must-be and attractive product features.
Developers should address the topic of decision making and
fall risk detection as well as serious gaming aspects in their
potential app. In addition, clear instructions for exercises and
workouts that would decrease a fall risk, as well as cost coverage
by health insurance companies would increase users’ acceptance
as well as their attraction to use such an app.
Investigated WTP revealed a median amount of €5 per month
(IQR 10) participants would invest to use a smartphone app,
incorporating the 17 products features as they rated them. This
amount is similar to the average price of paid apps within the
Apple App Store and Google Play as measured in 2017 [64].
Different independent ANOVA indicate that participants who
rated the features “description of physical exercises to reduce
fall risk,” “continuous workout program,” and “make new social
contacts” as attractive ones were willing to pay a higher amount
of money to use such an app as participants who did not. Only
for the fourth significant product feature (decision about
treatment by app) was a reverse correlation identified.
Participants who did not rate this feature as an attractive one
were willing to pay a higher amount of money. This might be
because of the 8% (8/96) of participants who categorized this
feature (decision about treatment by app) as questionable.
Results show that attractive product features result in a higher
WTP. Features such as “description of physical exercises to
reduce fall risk,” “continuous workout program,” and “make
new social contacts” would motivate potential users to spend
significantly more money for the ability to use a fall prevention
app as the other investigated ones.
Comparison With Prior Work
Product Features
The FARSEEING demonstrated the technical possibility of
measuring users’ fall risk during daily activities by a special
device [14,25]. This study could extend this knowledge as it
shows that potential users would appreciate an automated fall
risk assessment and detection as this would be done by a fall
prevention smartphone app. The survey revealed detection of
users’ potential fall risk to be a must-be product feature, whereas
automated detection without performing standardized test as a
Timed-Up and Go test would be preferred (product features:
criteria (C)1; C2).
An intervention or decision about treatment was not included
in the app of the FARSEEING project. Nevertheless, this was
considered to be the next mandatory step to design an innovative
fall prevention app. Therefore, the product features “decision
of treatment by health care professional” (product feature: C3)
and “decision of treatment by app” (product feature: C4) were
investigated in this survey. Related results show that the decision
about a necessary intervention is a must-be product feature,
whereas “decision of treatment by health care professional”
even reached the highest negative CS coefficient, indicating
that missing this feature would reduce potential users’
satisfaction significantly. Hence, future research should try to
include such options into a fall prevention app. A recent attempt
was made by the ProFouND project [26]. Within this project,
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an app for health care professionals was developed, which
should support them during the decision process about the
treatment of a certain patient [27].
Mendiola et al identified in their content analysis 12 features a
health app might profit from [19]. In this study, 3 of these 12
features were investigated regarding their relevance for a fall
prevention app. In this survey, participants classified product
features as additional data storage, data sharing via email, and
appointment reminder as must-be features of a fall prevention
app. Hence, this survey supports the content analysis of
Mendiola et al by empirical data.
Physical exercise is known to be an important factor to reduce
a person’s fall risk [4,29,40-43]. Therefore, it was part of this
study to investigate how a physical exercise program should be
designed according to potential users’ expectations. The results
indicate that participants are aware of the positive effect physical
exercise has on a potential fall risk, as all related product
features were at least rated as must-be features. Interestingly, a
continuous workout program such as the Otago program and
the instruction of physical exercises within the app were even
classified as attractive features. On the basis of the results from
this study, physical exercise interventions within a fall
prevention app should be designed as continuous workout
programs, which are supervised by a health professional or
clinician. Nevertheless, instructions for the exercises themselves
should be available within the app; so potential users are able
to exercise on their own. Such an exercise program is also
expected to support social contact and add an element of fun or
be more satisfying as serious gaming elements were classified
as must-be features.
These results are similar to the findings of Danbjørg et al [65].
Their study revealed that older adults appreciate a personal
therapist as the therapist could motivate them through comments
and personal social contact. Furthermore, they did identify
“motivation by competition” to be a highly relevant factor to
motivate older adults to perform physical activities [65]. The
feature of defining individual training goals was rated
ambivalent. This result is quite reasonable as a study by
Schlomann et al identified diverse acceptance of fixed training
goals within fitness apps among older adults [44]. Their study
revealed that the elderly try to achieve a socially accepted goal,
as it is suggested by a fitness app, even if this is exceeding their
physical abilities [44].
Willingness to Pay
Participants’ WTP was quite small compared with the expected
product features of a fall prevention smartphone app. Cost
coverage of a fall prevention app was classified as an attractive
feature according to the Kano technique. Comparing measured
WTP for fall risk prevention to Alzheimer prevention shows
how small the amount of money is that potential patients would
invest in a fall prevention service. WTP for an Alzheimer
prevention was about $155 per month, whereas a median amount
around €5 per month was revealed for a fall risk prevention
[66]. Nevertheless, to the best of the authors’ knowledge, this
is the first survey being able to price a fall prevention
smartphone app and therefore is able to support developers in
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designing an app satisfying users’ expectations. Prior studies
primarily investigated the clinical costs of fall patients as well
as the amount of money saved by different intervention
programs [67,68]. On the basis of the results of this survey,
researchers, as well as practitioners, can better understand which
product features are necessary to design a smartphone app that
will be acceptable to potential users and also be cost-effective.
Limitations
This study has several limitations related to its methodological
design as well as the reported results. The open Web-based
study was not representative because of regional recruiting in
Germany via Clickworker. A bias in recruitment might lead to
differences in the groups in the accessed fall risk or desired
product features of a fall prevention app.
Furthermore, participants’ health status and quality of life were
good within the sample. Therefore, results might differ with a
sample of participants suffering from worse health status or
who have poorer quality of life. Just a small portion had already
experienced a fall incidence; therefore, rating of product features
might change with a sample including individuals with a higher
number of experienced fall incidents. Future research might
address this topic by in-depth focus groups to design a fall
prevention app, especially for already fallen older adults.
Conclusions
Fall incidents are severe problems among the elderly [2]. A
major problem in this context is that older adults are unaware
of their potential fall risk as it rises slowly [4,5]. It is, therefore,
necessary to offer older adults a low-threshold service to assess
their own risk of falling. In view of the increasing use of health
apps in society and especially in the group of elderly individuals,
an app appears to be a useful long-term approach to helping
older people to prevent falls [12-16].
The aim of this study was to determine potential product features
of a fall prevention app adults aged older than 60 years would
appreciate irrespective of whether they already experienced a
fall or not.
In an exploratory approach, product features were ascertained
that potential users would expect from a fall prevention
smartphone app. To the best of the authors’ knowledge, this is
the first study explicitly investigating this aspect. In total, 17
product features were investigated, which were derived from
different recent research projects about fall prevention. Twelve
aspects were determined to be “must-be” product features,
including unobtrusive fall risk detection, decision making about
necessary treatment, and offering physical exercises to reduce
the risk of falling. Attractive features of a fall prevention app
would include educational features such as a checklist for typical
tripping hazards and a guide of action in case of a fall. Hence,
the authors are of the opinion that such an app could be
successfully adapted within a common app store. This may
enable interested older adults to identify, monitor, and treat
under the supervision of a health professional their risk of falls,
albeit the effectiveness of such an app would need to be
evaluated in follow-up research studies.
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Acknowledgments
This publication is part of the research project “TECH4AGE,” financed by the Federal Ministry of Education and Research
(BMBF, under Grant No. 16SV7111) and promoted by VDI/VDE Innovation + Technik GmbH.
Conflicts of Interest
None declared.
Multimedia Appendix 1
Introduction text of survey (German/English).
[PDF File (Adobe PDF File), 20KB-Multimedia Appendix 1]
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Abbreviations
AFPS: Aachen Falls Prevention Scale
AFRIS: Attitudes Falls Related Intervention Scale
ANOVA: one-way analyses of variance
EQ5D-3L: EuroQol Questionnaire 3 Level
C: criteria
CS: customer satisfaction coefficient
EU: European Union
FES-I: short Falls Efficacy Scale-International
FOF: fear of falling
IQR: interquartile range
WTP: willingness to pay
Edited by G Eysenbach; submitted 21.11.17; peer-reviewed by W Meekes, M Stein, E Stanmore; comments to author 04.01.18; revised
version received 16.02.18; accepted 23.02.18; published 27.03.18
Please cite as:
Rasche P, Mertens A, Brandl C, Liu S, Buecking B, Bliemel C, Horst K, Weber CD, Lichte P, Knobe M
Satisfying Product Features of a Fall Prevention Smartphone App and Potential Users’ Willingness to Pay: Web-Based Survey Among
Older Adults
JMIR Mhealth Uhealth 2018;6(3):e75
URL: http://mhealth.jmir.org/2018/3/e75/
doi: 10.2196/mhealth.9467
PMID: 29588268
©Peter Rasche, Alexander Mertens, Christopher Brandl, Shan Liu, Benjamin Buecking, Christopher Bliemel, Klemens Horst,
Christian David Weber, Philipp Lichte, Matthias Knobe. Originally published in JMIR Mhealth and Uhealth
(http://mhealth.jmir.org), 27.03.2018. This is an open-access article distributed under the terms of the Creative Commons Attribution
License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any
medium, provided the original work, first published in JMIR mhealth and uhealth, is properly cited. The complete bibliographic
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https://www.nature.com/articles/s41467-022-33649-3.pdf
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English
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ChAd155-RSV vaccine is immunogenic and efficacious against bovine RSV infection-induced disease in young calves
|
Research Square (Research Square)
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cc-by
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ChAd155-RSV vaccine is immunogenic
and efficacious against bovine RSV
infection-induced disease in young calves Received: 29 September 2021
Accepted: 27 September 2022
Check for updates Rineke de Jong
1, Norbert Stockhofe-Zurwieden1, Judith Bonsing1,
Kai-Fen Wang2,5, Sarah Vandepaer3, Badiaa Bouzya4, Jean-François Toussaint4,6,
Ilse Dieussaert4, Haifeng Song
2,7
& Ann-Muriel Steff
2 Rineke de Jong
1, Norbert Stockhofe-Zurwieden1, Judith Bonsing1,
Kai-Fen Wang2,5, Sarah Vandepaer3, Badiaa Bouzya4, Jean-François Toussaint4,6,
Ilse Dieussaert4, Haifeng Song
2,7
& Ann-Muriel Steff
2 Received: 29 September 2021
Accepted: 27 September 2022
Check for updates Respiratory syncytial virus (RSV) infection causes a substantial lower-
respiratory-tract disease burden in infants, constituting a global priority for
vaccine development. We evaluated immunogenicity, safety and efficacy of a
chimpanzee adenovirus (ChAd)-based vaccine candidate, ChAd155-RSV, in a
bovine RSV (bRSV) challenge model. This model closely reproduces the
pathogenesis/clinical manifestations of severe pediatric RSV disease. In ser-
onegative calves, ChAd155-RSV elicits robust neutralizing antibody responses
against human RSV. Two doses protect calves from clinical symptoms/lung
pathological changes, and reduce nasal/lung virus loads after both a short
(4-week) and a long (16-week) interval between last immunization and sub-
sequent bRSV challenge. The one-dose regimen confers near-complete or
significant protection after short-term or long-term intervals before challenge,
respectively. The presence of pre-existing bRSV-antibodies does not affect
short-term efficacy of the two-dose regimen. Immunized calves present no
clinical signs of enhanced respiratory disease. Collectively, this supports the
development of ChAd155-RSV as an RSV vaccine candidate for infants. Respiratory syncytial virus (RSV) infection is a major cause of lower
respiratory tract disease (LRTD) in infants. Infants aged under 1 year
have the highest incidence of severe LRTD (i.e., bronchiolitis and
pneumonia)1, and the disease burden extends even beyond the second
year of life. Worldwide, there were an estimated 33.1 million new epi-
sodes of RSV-linked acute lower respiratory infection in children aged
under 5 years (in 2015), and about 10% of these cases were severe
enough to warrant hospitalization2. In the same age group, the esti-
mated RSV-attributableglobal mortality wasaround 59,600 in-hospital
deaths and 118,200 overall deaths per year, 99% of which occurred in
developing countries2,3. Due to this high incidence of severe disease and a lack of cost-
effective preventative measures, RSV represents a major priority for
vaccine development4. However, progress in this area has been largely
hindered by the possibility of vaccination-induced enhanced respira-
tory disease (ERD), which manifested after vaccination with a formalin-
inactivated RSV vaccine candidate (FI-RSV, Lot 100) in the 1960s5–7. Article https://doi.org/10.1038/s41467-022-33649-3 1Wageningen Bioveterinary Research, Wageningen University & Research, Houtribweg 39, 8221 RA Lelystad, The Netherlands. 2GSK, 14200 Shady Grove
Road, Rockville, MD 20850, USA. 3CONSULTYS Benelux S.A, 73D Rue de Namur, 1000 Brussels, Belgium. 4GSK, Rue de l’Institut 89, 1330 Rixensart, Belgium.
5Present address: Atara Biotherapeutics, Inc., 2380 Conejo Spectrum St Suite 200, Thousand Oaks, CA 91320, USA. 6Present address: Sanofi-Pasteur,
14 Espace Henry Vallée, 69007 Lyon, France. 7Present address: Suzhou Abogen Bioscience Ltd, Suzhou, Jiangsu, China.
e-mail: haifengsong2003@yahoo.com; ann-muriel.x.steff@gsk.com ChAd155-RSV vaccine is immunogenic
and efficacious against bovine RSV
infection-induced disease in young calves Consequently, RSV vaccines
that induce cellular expression of RSV antigens, such as viral vectored
or nucleic acid-based platforms, are considered particularly suited for
RSV pediatric vaccination11. These candidates may behave biologically
as a live-virus vaccination as the antigens are produced intracellularly,
thus preferentially inducing CD8+ T cell responses and skewing the
adaptive immunity towards a Th1 phenotype. y
y
Adenoviral (Ad) vectors represent an attractive vaccine platform,
as they are known to induce both cellular and humoral responses
against the expressed antigens12–14. Particularly adenoviruses of simian
origin, such as chimpanzee Ad (ChAd) strains, are commonly used as
vaccine vectors, due to their lower seroprevalence and thus decreased
vector neutralization in humans as compared to human Ad515–20. Sev-
eral ChAd-vectored candidate vaccines against infectious agents
including RSV and SARS-CoV-2 have been demonstrated to induce
favorable immunogenicity, safety and efficacy profiles in clinical
trials21–25. A pediatric RSV vaccine candidate based on a group C ChAd
155 vector, ChAd155-RSV, encodes RSV fusion protein (F) in secreted
form to induce humoral responses, and intracellularly expressed
transcription anti-termination protein M2-1 and nucleocapsid protein
N aimed to induce a CD8+ T cell response22,26. ChAd155-RSV has first
been evaluated in animal models26 and has subsequently progressed to
Phase I, I/II, and II clinical development stages22 (NCT02927873,
NCT03636906). Upon challenge after a long DOI (Fig. 2b, e, h, k), clinical symp-
toms in the controls were similar to those in the short-DOI controls. Two vaccinations completely prevented general illness, fever, and
increased respiratory rates. However, mild URTD symptoms were
observed on a limited number of days (Fig. 2h). In Study 2, performed separately, the efficacy of a 1-dose vacci-
nation for protection against bRSV challenge 16-weeks after immuni-
zation was evaluated (Fig. 2c, f, i, l). Unexpectedly, the challenge
outcome in the age-matched placebo control group in this study was
more severe than in Study 1. In Study 1, all placebo control calves
developed moderately to severely increased breathing rates from dpc
6 onwards, and recovered by dpc 10 or 11. In Study 2, the onset of
respiratory distress was observed earlier, at dpc 5, and most of the
placebo control calves developed severely increased breathing rates
(Fig. 2l). This resulted in persistent respiratory distress or non-
reversible respiratory failure in five (out of nine) placebo control
calves. ChAd155-RSV vaccine is immunogenic
and efficacious against bovine RSV
infection-induced disease in young calves On dpc 7 and 8, four calves from the placebo group were pre-
terminated as they complied to humane endpoint criteria while
another calf had died unexpectedly at dpc 8. Despite the more severe
clinical outcome of bRSV challenge in Study 2, the 1-dose vaccination
group was nearly completely protected after long-term DOI against
general illness and fever (P ≤0.001), while the breathing rate was
slightly reduced (P ≤0.05) and the URTD symptoms were comparable
to those in the placebo group (Fig. 2c, f, i, l). Nevertheless, none of the
1-dose ChAd155-RSV vaccinated calves was pre-terminated, and all of
them had nearly recovered by the end of the study (dpc 12). Preclinicalstudies in RSV infection models indicated thatno single
validated animal model can faithfully predict the clinical outcome and
efficacy
of
an
RSV
vaccine
candidate
in
the
human
target
population27–29. Bovine RSV (bRSV) infection symptoms in young
calves share many aspects of severe RSV infection in human infants
with respect to pathogenesis and clinical manifestations, including
fever, runny nose, coughing and labored breathing27,30,31. The experi-
mental calf model is thus a unique translational model in the pre-
clinical evaluation of human RSV vaccines with regard to clinical
efficacy32, conferring advantages over mice and cotton rats model
which are only semi-permissive for the human challenge virus,
requiring a high challenge dose (i.e., 5-6 log10 PFU/TCID50), and do not
show obvious respiratory symptoms27,33–36. Human and bovine RSV
proteins including the F protein have a high-level homology with >80%
amino acid identity, thus inducing cross-reactive immunity27. In addi-
tion, FI-bRSV-induced ERD in calves has been reported by several
groups30,37–39, allowing the identification of ERD signs already in the
preclinical vaccine development phases. In the current study, the
humoral immunogenicity, safety, and efficacy of the ChAd155-RSV
vaccine has been evaluated in a bovine RSV challenge model in young
calves. We report that ChAd155-RSV vaccination elicited nAb respon-
ses, and two doses protected the calves from clinical symptoms and
lung pathological changes upon bRSV challenge. The presence of pre-
existing RSV-antibodies did not affect the short-term efficacy of a two-
dose vaccination regimen in this model, and no clinical signs of ERD
were observed post-challenge. The collective data supported further
development of the ChAd155-based RSV vaccine as a candidate for
pediatric immunization. ChAd155-RSV vaccine is immunogenic
and efficacious against bovine RSV
infection-induced disease in young calves Effect of ChAd155-RSV on bRSV-induced lung pathology
At necropsy (dpc 12/13), lungs were examined macroscopically to
determine the consolidated lung area (CLA) as percentage of the total
lung area (see Supplemental Fig. 1 for representative photos, and
Fig. 3a–c). Additionally, three tissue samples per lung were examined
microscopically for histopathological changes (see Supplemental
Fig. 2 for representative images), and sections were scored for evi-
dence of bronchitis, peribronchitis/perivasculitis, interstitial pneu-
monia and alveolitis. Scores are presented as total histopathological
sum scores (Fig. 3d–f) as well as by individual pathological symptom
(Supplemental Fig. 3). In Study 1, macroscopic CLAs upon short DOI and challenge were
observed in all control calves (≤7%), but were only minimally or not
observed in vaccinated calves of the single- or two-dose groups
(Fig. 3a). All control calves displayed histopathological changes (scores
5 to 34, on a scale of 48; Fig. 3d). These changes varied from focal
bronchitis/alveolitis to multifocal or extended changes associated with
obliterating bronchitis and severe pneumonia, with an influx of neu-
trophil granulocytes and lymphomonocytic inflammatory cells in the
alveoli. By contrast, vaccinated calves displayed minimal alterations
(scores ≤5), except for one animal from the single-dose group with a ChAd155-RSV vaccine is immunogenic
and efficacious against bovine RSV
infection-induced disease in young calves In
one of these trials, FI-RSV vaccination of 31 infants resulted in 2 deaths
and 80% hospitalization (of the 20 vaccinees who became infected), as
compared to 5% hospitalization in the control group vaccinated with
parainfluenza vaccines5. Although the mechanisms underlying FI-RSV-
induced ERD are not completely clear, many studies agreed that a T Nature Communications| (2022) 13:6142 Nature Communications| (2022) 13:6142 1 Article https://doi.org/10.1038/s41467-022-33649-3 ChAd155-RSV efficacy against bRSV-induced clinical disease
In Study 1, placebo control calves developed typical clinical symptoms
of bRSV infection when challenged after a short DOI. General illness
symptoms (expressed in scores) accompanied by fever (rectal tem-
perature >39.5 °C) appeared at day 6 post challenge (dpc 6), peaked at
dpc 7/8 and resolved by dpc 10/11 (Fig. 2a, d). Simultaneously, their
upper respiratory tract disease (URTD) scores and respiratory rates
were increased (Fig. 2g, j). In contrast, calves having received 2 doses of
the vaccine were nearly completely protected from clinical symptoms,
including fever. Although these animals occasionally showed (slightly)
increased URTD scores and respiratory rates, these levels remained
significantly lower than those for the controls (P ≤0.01 and P ≤0.001,
respectively). As compared to the two-dose vaccine group, the single-
dose vaccine group exhibited slightly higher general illness and URTD
scores. However, the protective efficacy in this group was overall
comparable to that in the two-dose group, and clearly higher relative
to the control group. helper (Th) 2-biased cellular response, a high ratio of non-functional
antibodies (Abs) to neutralizing Abs (nAbs), and a lack of CD8+ cyto-
toxic T cell responses are among its major attributes8–10. An ideal RSV
candidate should therefore induce high nAb titers, a Th1-biased CD4+ T
cell response, and CD8+ T cell responses. Consequently, RSV vaccines
that induce cellular expression of RSV antigens, such as viral vectored
or nucleic acid-based platforms, are considered particularly suited for
RSV pediatric vaccination11. These candidates may behave biologically
as a live-virus vaccination as the antigens are produced intracellularly,
thus preferentially inducing CD8+ T cell responses and skewing the
adaptive immunity towards a Th1 phenotype. helper (Th) 2-biased cellular response, a high ratio of non-functional
antibodies (Abs) to neutralizing Abs (nAbs), and a lack of CD8+ cyto-
toxic T cell responses are among its major attributes8–10. An ideal RSV
candidate should therefore induce high nAb titers, a Th1-biased CD4+ T
cell response, and CD8+ T cell responses. Nature Communications| (2022) 13:6142 Results R
i
i d
l
d
i
d
i bRSV Ab
i
l
Group:
Ab–
Short DOI (2D): ChAd155-RSV / placebo
Ab–
Short DOI (1D): ChAd155-RSV
Ab–
Long DOI (2D): ChAd155-RSV / placebo
Group:
Ab–
Long DOI (Ab– / 1D): ChAd155-RSV / placebo
Ab+
Short DOI (Ab+ / 2D): ChAd155-RSV / placebo
Ab–
Short DOI (Ab– / 2D): ChAd155-RSV (bridge)
DOI 4 wk
DOI 16 wk
DOI 4 wk
DOI 16 wk
DOI 4 wk
DOI 4 wk
Study 1 in colostrum-deprived seronegative calves
Study 2 in calves fed Ab-negative or Ab-positive colostrum
4
6
18
26
27+5 d
22
10
12
0
Study week
6
18
23+5 d
22
8
14
0
Study week Study 1 in colostrum-deprived seronegative calves Study 2 in calves fed Ab-negative or Ab-positive colostrum Birth
Injection time-point
Ab–
Rearing period colostrum-deprived anti-bRSV Ab-negative calves
Ab+/–
Rearing period colostrum-fed anti-bRSV Ab-positive (Ab+) or -negative (Ab–) calves
Post-challenge evaluation of clinical symptoms, lung pathology, viral load, and immunogenicity; termination
Vaccine immunogenicity evaluation using a long or short duration of immunity (DOI)
bRSV challenge
Fig. 1 | Study designs. Study 1 was conducted in bovine RSV (bRSV)-seronegative
(Ab−) colostrum-deprived calves. Study 2 was conducted in Ab−calves fed bRSV
antibody-negative colostrum, and in bRSV-seropositive (Ab+) calves fed
Ab-positive colostrum. Animals (n = 7 to 9/group) received 1 or 2 doses (1D or 2D,
respectively) of either ChAd155-RSV vaccine or placebo (phosphate-buffered saline;
controls). Corresponding vaccine and control groups followed the same trea
challenge schedules after either a short (4 weeks) or a long (16 weeks) dura
immunity (short DOI or long DOI, respectively). Short or long DOI calves w
recruited in two separate cohorts to arrange the challenge on the same
calendar day. Ab–
Rearing period colostrum-deprived anti-bRSV Ab-negative calves Fig. 1 | Study designs. Study 1 was conducted in bovine RSV (bRSV)-seronegative
(Ab−) colostrum-deprived calves. Study 2 was conducted in Ab−calves fed bRSV
antibody-negative colostrum, and in bRSV-seropositive (Ab+) calves fed
Ab-positive colostrum. Animals (n = 7 to 9/group) received 1 or 2 doses (1D or 2D,
respectively) of either ChAd155-RSV vaccine or placebo (phosphate-buffered saline; controls). Corresponding vaccine and control groups followed the same treatment/
challenge schedules after either a short (4 weeks) or a long (16 weeks) duration of
immunity (short DOI or long DOI, respectively). Results Short or long DOI calves were
recruited in two separate cohorts to arrange the challenge on the same
calendar day. loads determined by bRSV qPCR on all samples (Supplemental Fig. 4)
were consistent with the viral infectivity data generated from selected
time points (described below). score of 10 (Fig. 3d). In parallel, all seven control calves with long DOI
showed CLAs of up to 6.37 %, whereas consolidation was absent (n = 7)
or minimal (n = 1) in calves which received two vaccine doses (Fig. 3b). All control calves showed histopathological changes (scores: 1–36),
while all except one (with score 11) in the two-dose vaccine group were
completely protected against histopathological alterations (Fig. 3e). In Study 1, virus loads in BALs from control calves of both the
short- and long-DOI groups peaked on dpc 7 (mean titer ~300
TCID50/mL), but were cleared by dpc 9 (Fig. 4a, c). One or two vac-
cinations resulted in significantly lower viral loads at dpc 5 and 7 in
both short-DOI groups (P ≤0.001) and in the two-dose long-DOI
group (P ≤0.01 at dpc 5 and P ≤0.001 at dpc 7). Kinetics in the
nasopharynx of short- or long-DOI controls were similar as for BALs,
with a gradual increase from dpc 3 onwards, peaks on dpc 6 or 7
(mean titers:1000–10,000 TCID50/mL), and no re-isolated infectious
virus on dpc 9 (Fig. 4b, d). Two vaccine doses significantly reduced
viral loads as compared to controls, in both the short- and the
long-DOI groups (on dpc 3, 5, 6 and 7, and on dpc 6 and 7, respec-
tively). In the single-dose short-DOI vaccine group, viral clearance
was also observed earlier than in controls, i.e., on dpc 6 and 7
(P ≤0.001, Fig. 4b). In line with the clinical observations, lung pathology was more
severe after the long DOI in Study 2. Indeed, compared to Study 1,
control calves had larger CLAs and more histopathological changes
(0.4–53% and scores 28–39; Fig. 3c, f, respectively). In addition, greater
CLAs were observed in the five animals that were either pre-terminated
(n = 4) or found dead (n = 1) atdpc 7/8 (Fig. 3c). The single-dose vaccine
group exhibited at most minimal (0.8%) macroscopic consolidation,
though microscopic evaluation still revealed histopathological chan-
ges (scores 2–32). Results In two separate studies (Fig. 1 and Table 1), the protective efficacy of a
single-dose or two-dose vaccination regimen with ChAd155-RSV was
assessed upon bRSV challenge. The challenge was performed after
either a short (4-week) or a long (16-week) interval after the (last)
vaccination—further referred to as a short/long duration of immunity
(DOI). In addition, the efficacy of a two-dose regimen was assessed in
the presence of pre-existing bRSV-antibodies after a short DOI. Nature Communications| (2022) 13:6142 2 https://doi.org/10.1038/s41467-022-33649-3 Article Birth
Injection time-point
Ab–
Rearing period colostrum-deprived anti-bRSV Ab-negative calves
Ab+/–
Rearing period colostrum-fed anti-bRSV Ab-positive (Ab+) or -negative (Ab–) calves
Post-challenge evaluation of clinical symptoms, lung pathology, viral load, and immunogenicity; termination
Vaccine immunogenicity evaluation using a long or short duration of immunity (DOI)
Group:
Ab–
Short DOI (2D): ChAd155-RSV / placebo
Ab–
Short DOI (1D): ChAd155-RSV
Ab–
Long DOI (2D): ChAd155-RSV / placebo
Group:
Ab–
Long DOI (Ab– / 1D): ChAd155-RSV / placebo
Ab+
Short DOI (Ab+ / 2D): ChAd155-RSV / placebo
Ab–
Short DOI (Ab– / 2D): ChAd155-RSV (bridge)
DOI 4 wk
DOI 16 wk
DOI 4 wk
DOI 16 wk
DOI 4 wk
DOI 4 wk
Study 1 in colostrum-deprived seronegative calves
Study 2 in calves fed Ab-negative or Ab-positive colostrum
4
6
18
26
27+5 d
22
10
12
0
bRSV challenge
Study week
6
18
23+5 d
22
8
14
0
Study week
g. 1 | Study designs. Study 1 was conducted in bovine RSV (bRSV)-seronegative
b−) colostrum-deprived calves. Study 2 was conducted in Ab−calves fed bRSV
tibody-negative colostrum, and in bRSV-seropositive (Ab+) calves fed
-positive colostrum. Animals (n = 7 to 9/group) received 1 or 2 doses (1D or 2D,
pectively) of either ChAd155-RSV vaccine or placebo (phosphate-buffered saline;
controls). Corresponding vaccine and control groups followed the same treatm
challenge schedules after either a short (4 weeks) or a long (16 weeks) duratio
immunity (short DOI or long DOI, respectively). Short or long DOI calves were
recruited in two separate cohorts to arrange the challenge on the same
calendar day. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs
Most infants have maternal RSV Abs transferred via the placenta. We
therefore investigated whether such pre-existing responses would
impact the vaccine’s immunogenicity and efficacy. As transplacental
transfer does not occur in the bovine species, newborn calves in Study
2 received colostrum, which was either bRSV Ab-positive (resulting in a
group mean nAb titer of 1:200 at pre-vaccination), or bRSV Ab-
negative. In both cases, colostrum administration was followed by two
vaccine doses, 4 weeks apart, and a challenge after a short DOI (see
Fig. 1 and Table 1). A control group with colostrum-derived pre-existing
bRSV Ab received two phosphate-buffered saline (PBS) injections and a
challenge following the same schedule. Upon challenge, this control
group displayed the typical bRSV-induced clinical symptoms, includ-
ing general illness, fever, increased respiratory rates and signs of URTD
(Fig. 5a–d). Additionally, increases in CLAs, overall lung pathological
scores and increased viral loads were observed in their BAL and/or
nasal samples (Fig. 5e–h). Of note, this placebo group behaved simi-
larly to the one-dose long-term DOI placebo group from Study 2, with
more severe clinical symptoms compared to that in Study 1, leading to
premature termination in 3 out of 9 animals. In contrast to the con-
trols, calves in the two-dose vaccination groups with or without pre-
existing bRSV Ab exhibited either a complete absence of symptoms,
or significantly reduced symptoms of general illness, fever, and
respiratory rates, although they still showed URTD symptoms
(Fig. 5a–d). In addition, none of these vaccinated animals were pre-
terminated. Although occasionally, clinical scores were slightly
higher in the group with versus without pre-existing Abs, these dif-
ferences were not statistically significant. Moreover, two vaccina-
tions resulted in significant reductions in the CLAs—which were
absent or minimal in all (9/9) animals with pre-existing Abs, and in
nearly all (7/9) animals without pre-existing Abs—and in reduced
pathology sum scores (P ≤0.01; Fig. 5e and P ≤0.001; Fig. 5f). The
vaccinations also resulted in reduced virus titers and faster viral
clearance from the lung and nasopharynx (Fig. 5g, h). Overall, the
presence of pre-existing Abs had minimal impact on the vaccine
efficacy in calves which received two vaccine doses and were chal-
lenged 4 weeks post-dose 2. Furthermore, the fact that all these Kinetics of vaccine-induced nAb responses were analyzed by standard
neutralization assay using hRSV A Long. ChAd155-RSV efficacy on bRSV replication in nasal and
lung fluids In Study 2, the kinetics of viral replication were slightly advanced,
with a peak at day 5 and decreased thereafter (Fig. 4e, f). This was likely Following bRSV challenge, nasopharyngeal brush samples were
obtained daily, and BAL samples on dpc 5, 7, and 9 (Fig. 4). The viral Nature Communications| (2022) 13:6142 3 https://doi.org/10.1038/s41467-022-33649-3 Article Table 1 | Detailed study designs
Study
Colostrum feedinga
Sero-statusa
Treatments
DOI (weeks) before
challenged
N
Test articleb
(i.m.; 2 mL/dose)
No. of doses
Age at vaccine administration
(weeks)c
1
No
–
ChAd155-RSV
2
6, 10
4
8
No
–
Placebo
2
6, 10
4
8
No
–
ChAd155-RSV
1
10
4
8
No
–
ChAd155-RSV
2
6, 10
16
8
No
–
Placebo
2
6, 10
16
7
2
Yes
Ab−
ChAd155-RSV
1
6
16
9
Yes
Ab−
Placebo
1
6
16
9
Yes
Ab+
ChAd155-RSV
2
6
4
9
Yes
Ab+
Placebo
2
6
4
9
Yes
Ab−
ChAd155-RSV
2
6
4
9
aIn Study 1, calves received only antibody-negative milk replacer. In Study 2, a single feeding with anti-bRSV antibody-negative/-positive (Ab−/+) colostrum was supplied followed by feeding with
antibody-negative milk replacer. bChAd155-RSV at 5 × 1010 viral particles/dose or phosphate-buffered saline, given by intramuscular (i.m.) injection. cAll animals in a given group were born within a 3–4-day period. dDuration of immunity (DOI) period(weeks since lastvaccine administration) was followed by bRSVchallenge at 3.2 (Study 1)or 3.7 (Study 2) log10 TCID50/2 mL dose. Necropsy was performed on days
12/13 post challenge for all groups. Table 1 | Detailed study designs animals (irrespective of the presence of pre-existing bRSV Ab) were
protected from bRSV disease, as observed in Study 1 with the same
two-dose short-DOI regimen (see Fig. 2), indicated that the vaccine
performed similarly well after a more virulent challenge. due to the more virulent challenge. Nonetheless, a single vaccination
resulted in earlier viral clearance compared to placebo, as observed in
the lungs at dpc 7, and in the nasopharyngeal tract at dpc 6 or 7
(all P ≤0.001). ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs In Study 1 at 4 weeks post first
vaccination, this vaccine dose had elicited low-level responses across
the single-dose short-DOI group and both two-dose groups (geometric
mean titers [GMTs]: 35.7–57.7; Fig. 6a, b and Supplemental Fig. 5a, b). Similarly, in Study 2, low titers were observed in the single-dose long-
DOI group and in the two-dose group without pre-existing Abs (GMTs:
154 and 76, respectively; Fig. 6c, d and Supplemental Fig. 5c, d). However, the titer in the latter single-dose group was sustained at
similar levels up to the time of challenge, i.e., 16 weeks after
immunization. In both studies, the second vaccination clearly increased the nAb
titers (Fig. 6a, b, d, and Supplemental Fig. 5a, b, d). At 2 weeks post-
dose 2, this was observed in the short- and long-DOI vaccine groups in
Study 1 (GMTs: 296 and 531, respectively) and in the short-DOI vaccine
group without pre-existing Abs in Study 2 (GMT: 1323). In Study 1, long-
DOI, the titer gradually decreased over time, but remained detectable
at 16 weeks post-dose 2 (GMT: 101; Fig. 6b and Supplemental Fig. 5b). In Study 2, nAb responses induced by two vaccinations were also
compared between calves with versus without pre-existing bRSV Abs
(Fig. 6d and Supplemental Fig. 5d). While at 4 weeks post-dose 1,
responses were low in vaccinated calves without pre-existing Abs,
titers in calves with pre-existing Abs were similar between the placebo
and vaccine groups. This suggest that the nAb responses in the latter
vaccine group largely comprised pre-existing Abs derived from the Ab-
positive colostrum. The titers in both vaccinated groups increased
after the second dose, but were two- to three-fold lower in the calves
which received Ab-positive colostrum (GMTs with versus without pre-
existing Abs, at 2 or 4 weeks post-dose 2: 436 versus 1323, and 382
versus 725, respectively). Our data therefore indicate that the pre-
existing bRSV Abs may have dampened the vaccine-induced nAb
response, consistent with observations made for different vaccines in
human infants with maternal Abs40,41. However, despite the potential
blunting effect of pre-existing responses on the humoral vaccine
immunogenicity, the clinical efficacy upon bRSV challenge appeared
to be uncompromised, as described above (see Fig. 5). Nature Communications| (2022) 13:6142 4 Upon challenge, titers increased across all groups in both
studies. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Of note, among the short-DOI groups in Study 1, titers
l
i
th
i
l
d
th
i
th
t
d
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groups (Fig. 6a, b and Supplemental Fig. 5a, b). ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs A similar trend
comparable post-challenge titers among groups was seen in Stu
2 I d
d th
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tit
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1 0 1 1
3 8
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PM
AM
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1 0 1 1
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0
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7
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1 0 1 1
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o
c
s
s
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a
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a
b
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Study 1 long DOI:
Study 2 long DOI:
c
f
e
)
n
i
m
/(
e
t
a
r y
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t
a
ri
p
s
e
R
(
e
r
u
t
a
r
e
p
m
e
T
°C)
Temperature (°C)
Temperature (°C)
****
***
***
General illness score
General illness score
j
l
k
i
h
Day post-challenge
Day post-challenge
Day post-challenge
e
r
o
c
s
D
T
R
U
URTD score
URTD score
Respiratory rate (/min)
Respiratory rate (/min)
**
***
**
*****
***
*
ChAd155-RSV (1D)
Placebo-L
ChAd155-RSV (2D)
Placebo-L
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S
d
g
Fig. 2 | Clinical symptoms. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Article
https://doi.org/10.1038/s41467-022-33649 Article https://doi.org/10.1038/s41467-022-33649-3 Study 2 long DOI:
ChAd155-RSV (1D)
Placebo-L b -2
0
2
4
6
8
1 0
1 2
0
1
2
3
c
***
General illness score a -2
0
2
4
6
8
1 0
1 2
0
1
2
3
b
***
General illness score c -2
0
2
4
6
8
1 0
1 2
0
1
2
3
e
r
o
c
s
s
s
e
n
lli l
a
r
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e
G
a
**** -2
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0
1
2
3
4
5
6
7
8
9
1 0 1 1
3 8
3 9
4 0
4 1
PM
AM
f
Temperature (°C)
*** -2
-1
0
1
2
3
4
5
6
7
8
9
1 0 1 1
3 8
3 9
4 0
4 1
PM
AM
(
e
r
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t
a
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p
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T
°C)
d -2
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0
1
2
3
4
5
6
7
8
9
1 0 1 1
3 8
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4 0
4 1
PM
AM
e
Temperature (°C)
** f d e -2
0
2
4
6
8
1 0
1 2
0
1
2
3
h
URTD score -2
0
2
4
6
8
1 0
1 2
0
1
2
3
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URTD score -2
0
2
4
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8
1 0
1 2
0
1
2
3
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r
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c
s
D
T
R
U
**
g i h g j l k -2
0
2
4
6
8
1 0
1 2
2 0
4 0
6 0
8 0
1 0 0
1 2 0
l
Day post-challenge
Respiratory rate (/min)
* -2
0
2
4
6
8
1 0
1 2
2 0
4 0
6 0
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1 0 0
1 2 0
)
n
i
m
/(
e
t
a
r y
r
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ri
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Day post-challenge
***** -2
0
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1 0
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4 0
6 0
8 0
1 0 0
1 2 0
k
Day post-challenge
Respiratory rate (/min)
*** Day post-challenge Day post-challenge Day post-challenge Day post-challenge Day post-challenge Fig. 2 | Clinical symptoms. Calves (n = 7–9 biologically independent animals, see
i in Supplemental Table 1 before and after the challenge at the time points indicated. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Data are presented as means with 95% confidence intervals (vertical bars). Scores of
general illness symptoms (a–c), rectal temperatures (d–f), upper respiratory tract
disease (URTD) scores (g–i), and respiratory rates (j–l) are presented. Comparisons
between the vaccine groups and the respective control groups were estimated
using two-sided t tests on individual AUC data. *P ≤0.05; **P ≤0.01; ***P ≤0.001. Source data and statistical analyses with exact P values are provided as a Source
data and Supplementary Data 1 and 2, respectively. Fig. 2 | Clinical symptoms. Calves (n = 7–9 biologically independent animals, see
Table 1) received either a single or two doses (1D or 2D, respectively) of ChAd155-
RSV vaccine or placebo, followedby a short(4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size [n = 9] of the
placebo-L group of Study 2 decreased to n = 5 on day post-challenge [dpc] 8, and to
n = 4 on dpc 9–11; see Fig. 3). Clinical symptoms in the ChAd155-RSV-immunized or
placebo-treated calves were evaluated according to the scoring system presented in Supplemental Table 1 before and after the challenge at the time points indicated. Data are presented as means with 95% confidence intervals (vertical bars). Scores of
general illness symptoms (a–c), rectal temperatures (d–f), upper respiratory tract
disease (URTD) scores (g–i), and respiratory rates (j–l) are presented. Comparisons
between the vaccine groups and the respective control groups were estimated
using two-sided t tests on individual AUC data. *P ≤0.05; **P ≤0.01; ***P ≤0.001. Source data and statistical analyses with exact P values are provided as a Source
data and Supplementary Data 1 and 2, respectively. Fig. 2 | Clinical symptoms. Calves (n = 7–9 biologically independent animals, see
Table 1) received either a single or two doses (1D or 2D, respectively) of ChAd155-
RSV vaccine or placebo, followedby a short(4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Calves (n = 7–9 biologically independent animals, see
Table 1) received either a single or two doses (1D or 2D, respectively) of ChAd155-
RSV vaccine or placebo, followedby a short(4 weeks) or long (16 weeks) duration of
mmunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size [n = 9] of the
placebo-L group of Study 2 decreased to n = 5 on day post-challenge [dpc] 8, and to
n = 4 on dpc 9–11; see Fig. 3). Clinical symptoms in the ChAd155-RSV-immunized or
placebo-treated calves were evaluated according to the scoring system presented
in Supplemental Table 1 before and after the challenge at the time points indicat
Data are presented as means with 95% confidence intervals (vertical bars). Score
general illness symptoms (a–c), rectal temperatures (d–f), upper respiratory tr
disease (URTD) scores (g–i), and respiratory rates (j–l) are presented. Compariso
between the vaccine groups and the respective control groups were estimated
using two-sided t tests on individual AUC data. *P ≤0.05; **P ≤0.01; ***P ≤0.00
Source data and statistical analyses with exact P values are provided as a Sourc
data and Supplementary Data 1 and 2, respectively. Discussion Multiple studies have demonstrated that replication-incompetent
ChAd vectors against several viruses, including RSV and SARS-CoV-2,
are capable of inducing humoral and cell-mediated responses in
humans, with acceptable safety profiles16,19,24,25,42. Activation of both
arms of adaptive immunity is considered critical for protection against
severe RSV disease, especially in infants and older adults11,43–46. Pre-
viously, ChAd155-RSV was demonstrated to induce RSV nAbs and T cell
responses in seropositive adult humans22. Here we show that twodoses
of ChAd155-RSV induced robust nAb responses that reached the esti-
mated protective titer in human infants47 (i.e., 256) and just exceeded
the threshold of 6 log2 that is associated with a reduced risk of hos-
pitalization in infants48. The immune responses observed here in
calves, which were sustained during the 4-week study period, almost
completely protected the animals from bRSV clinical disease and lung
pathology, as did the two-dose regimen after the 16-week period. The
latter is remarkable, considering that at challenge these nAb responses
had decreased to a level (geometric mean of 101) well below the above
mentioned protection threshold47. Interestingly, this data also sug-
gests that the nAb titer before challenge impacts the magnitude of the
Ab response to viral infection, such that higher titers before infection
exhibit a lower increase after infection. A likely explanation is that a
more robust nAb response will better contain viral replication, and the
resulting lower viral levels will then induce a lower increase in Ab titer. In animals receiving a single vaccine dose, the general illness, A pediatric vaccine to protect infants against RSV-linked LTRD is
urgently needed to cover the large burden of disease in infants. Vector-based RSV vaccines, such as the ChAd155-RSV candidate
vaccine based on a chimpanzee-derived Ad-vector, elicit immune
profiles that have not been associated with ERD, and may be a sui-
table solution for pediatric use. We have evaluated the immuno-
genicity, safety (in terms of ERD) and vaccine efficacy of the Chad155-
RSV pediatric vaccine in a calf model, and studied different settings
and regimens to mimic, as closely as possible, the human infant
vaccination scenarios. We demonstrate that ChAd155-RSV was
immunogenic in naive young calves, inducing low to modest RSV nAb
responses following the first dose which were further increased fol-
lowing the second dose. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size [n = 9] of the
placebo-L group of Study 2 decreased to n = 5 on day post-challenge [dpc] 8, and to
n = 4 on dpc 9–11; see Fig. 3). Clinical symptoms in the ChAd155-RSV-immunized or
placebo-treated calves were evaluated according to the scoring system presented Upon challenge, titers increased across all groups in both
studies. Of note, among the short-DOI groups in Study 1, titers
were lower in the single-dose group than in the two-dose group at
pre-challenge (GMTs: 58 versus 316, respectively), but similar
between these groups two weeks after the challenge (Fig. 6a
and Supplemental Fig. 5a). Among the two-dose groups, post-
challenge titers were comparable between the short- and long-DOI groups (Fig. 6a, b and Supplemental Fig. 5a, b). A similar trend of
comparable post-challenge titers among groups was seen in Study
2. Indeed, the low titer in the single-dose group at pre-challenge
increased by 53-fold upon challenge, whereas fold-increases post-
challenge were lower in the two-dose groups without or with pre-
existing Abs (4.3-fold and 1.5-fold, respectively; Fig. 6c, d and
Supplemental Fig. 5c, d). Nature Communications| (2022) 13:6142 5 Article https://doi.org/10.1038/s41467-022-33649-3 0
5
1 0
2 0
4 0
6 0
1 0
0
5
1 0
2 0
4 0
6 0
1 0
0
8
1 6
2 4
3 2
4 0
4 8
0
8
1 6
2 4
3 2
4 0
4 8
***
***
0
8
1 6
2 4
3 2
4 0
4 8
0
5
1 0
2 0
4 0
6 0
1 0
m
u
s
e
r
o
c
s
y
g
o
l
o
h
t
a
p
o
t
s
i
H
Histopathology score sum
Histopathology score sum
d
e
f
**
)
%
(
a
e
r
a
g
n
u
l
d
e
t
a
d
il
o
s
n
o
C
Consolidated lung area (%)
Consolidated lung area (%)
***
a
b
c
***
***
***
***
ChAd155-RSV (2D)
Placebo-L
Study 1 long DOI groups:
ChAd155-RSV (1D)
Placebo-L
Placebo-L pre-terminated
Study 2 long DOI groups:
Study 1 short DOI groups:
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S
Fig. 3 | Lung pathology. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Macroscopic analysis of the consolidated lung
areas (CLAs; a–c) and sums of lung pathology scores determined by microscopic
analysis of the lung pathological changes (d–f) are presented for the study groups
indicated in the keys above the graphs. Sums of lung pathological scores were derived by addition of the scores for alveolitis, interstitial pneumonia, peribron-
chitis and bronchitis per individual animal. Data are presented as means ± 95%
confidence intervals. Each symbol represents an individual animal. Open symbols in
c, f indicate animals that were pre-terminated or found dead on dpc 7/8. Vaccine
groups were compared by two-sided t tests with the respective control groups
using one-way ANOVA for CLAs and histopathology score sums with treatment as
fixed effect and the appropriate variance assumption. Significant differences are
indicated by asterisks (**P ≤0.01; ***P ≤0.001). Source data and statistical analyses
with exact P values are provided as a Source data and Supplementary Data 1 and 2,
respectively. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively. On day post-challenge (dpc) 12/13 ani-
mals were subjected to necropsy. Macroscopic analysis of the consolidated lung
areas (CLAs; a–c) and sums of lung pathology scores determined by microscopic
analysis of the lung pathological changes (d–f) are presented for the study groups
indicated in the keys above the graphs. Sums of lung pathological scores were ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs 0
5
1 0
2 0
4 0
6 0
1 0
0
5
1 0
2 0
4 0
6 0
1 0
0
8
1 6
2 4
3 2
4 0
4 8
0
8
1 6
2 4
3 2
4 0
4 8
***
***
0
8
1 6
2 4
3 2
4 0
4 8
0
5
1 0
2 0
4 0
6 0
1 0
m
u
s
e
r
o
c
s
y
g
o
l
o
h
t
a
p
o
t
s
i
H
Histopathology score sum
Histopathology score sum
d
e
f
**
)
%
(
a
e
r
a
g
n
u
l
d
e
t
a
d
il
o
s
n
o
C
Consolidated lung area (%)
Consolidated lung area (%)
***
a
b
c
***
***
***
***
ChAd155-RSV (2D)
Placebo-L
Study 1 long DOI groups:
ChAd155-RSV (1D)
Placebo-L
Placebo-L pre-terminated
Study 2 long DOI groups:
Study 1 short DOI groups:
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S )
%
(
a
e
r
a
g
n
u
l
d
e
t
a
d
il
o
s
n
o
C
a Consolidated lung area (%)
c Consolidated lung area (%)
b Consolidated lung area 0
8
1 6
2 4
3 2
4 0
4 8
Histopathology score sum
f
*** 0
8
1 6
2 4
3 2
4 0
4 8
Histopathology score sum
e
*** Histopathology score sum
e Histopathology score sum
f Fig. 3 | Lung pathology. Calves (n = 7 to 9 biologically independent animals, see
Table 1) received a single or two doses (1D or 2D, respectively) of ChAd155-RSV Fig. 3 | Lung pathology. Calves (n = 7 to 9 biologically independent animals, see Fig. 3 | Lung pathology. Calves (n = 7 to 9 biologically independent animals, see
Table 1) received a single or two doses (1D or 2D, respectively) of ChAd155-RSV
vaccine or placebo, followed by a short (4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively) and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively. On day post-challenge (dpc) 12/13 ani-
mals were subjected to necropsy. ChAd155-RSV efficacy in the presence of pre-existing bRSV Abs Calves (n = 7 to 9 biologically independent animals, see
Table 1) received a single or two doses (1D or 2D, respectively) of ChAd155-RSV
vaccine or placebo, followed by a short (4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively) and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively. On day post-challenge (dpc) 12/13 ani-
mals were subjected to necropsy. Macroscopic analysis of the consolidated lung
areas (CLAs; a–c) and sums of lung pathology scores determined by microscopic
analysis of the lung pathological changes (d–f) are presented for the study groups
indicated in the keys above the graphs. Sums of lung pathological scores were
derived by addition of the scores for alveolitis, interstitial pneumonia, peribron-
chitis and bronchitis per individual animal. Data are presented as means ± 95%
confidence intervals. Each symbol represents an individual animal. Open symbols in
c, f indicate animals that were pre-terminated or found dead on dpc 7/8. Vaccine
groups were compared by two-sided t tests with the respective control groups
using one-way ANOVA for CLAs and histopathology score sums with treatment as
fixed effect and the appropriate variance assumption. Significant differences are
indicated by asterisks (**P ≤0.01; ***P ≤0.001). Source data and statistical analyses
with exact P values are provided as a Source data and Supplementary Data 1 and 2,
respectively. Discussion Moreover, the fact that all calves in the single-dose groups were
strongly protected, with reduced viral titers even though their nAb
titers were low, indicated that other arms of immunity, such as T cells,
may also have contributed to this level of protection. Indeed, the M2-1
and N proteins are included in the vaccine construct to elicit T cell
immunity, and preliminary analysis in a different calf study showed
that ChAd155-RSV induced both CD4+ and CD8+ T cell responses to RSV
proteins following a single injection (unpublished data). In addition, T
cell responses were detected in the vaccinated adult humans22. While a similar effect is expected to have occurred in the vaccinated
calves, they nevertheless exhibited the typical kinetics of a B cell
response to vaccination, in which nAb titers elicited by the first dose
were boosted by the second vaccination, and further boosted by the
challenge. This suggests that the vaccination had successfully induced
plasma-cell and memory B cell responses targeting RSV A Long. Moreover, the fact that all calves in the single-dose groups were
strongly protected, with reduced viral titers even though their nAb
titers were low, indicated that other arms of immunity, such as T cells,
may also have contributed to this level of protection. Indeed, the M2-1
and N proteins are included in the vaccine construct to elicit T cell
immunity, and preliminary analysis in a different calf study showed
that ChAd155-RSV induced both CD4+ and CD8+ T cell responses to RSV
proteins following a single injection (unpublished data). In addition, T
cell responses were detected in the vaccinated adult humans22. the lung and nasopharynx, were all comparable after long- and short-
DOI. However, among these single-dose groups, respiratory rates, and
lung histopathological scores with a long DOI (Study 2), while still
being lower than the controls, were less reduced than with the short
DOI (Study 1). In the absence of a two-dose long-DOI group in the same
study, it remains inconclusive whether this slightly compromised
protection was due to the more severe challenge condition, or to a
lower efficacy of this single-dose regime. Nevertheless, the overall
results suggest that a single-dose regime also merits consideration in
clinical development (NCT03636906). Though not measured here, several lines of evidence suggest the
presence of vaccine-induced immunity beyond the detected nAb
responses. Discussion The two-dose vaccination regimen pro-
tected the calves from bRSV-induced clinical disease and lung
pathology, and significantly reduced viral loads in the nasopharynx
and lung after a DOI of either 4 or 16 weeks. The protection conferred
by a single vaccination resembled the protection following the two-
dose regimen when applying the short DOI, and was near-complete
against general illness, fever, and CLA after the long DOI. However, in
the latter case, the effects of the single dose on respiratory rates and
lung histopathology were less profound than with two doses and a
long DOI. Finally, protection after two vaccinations and a short DOI
was similar in calves with versus without pre-existing maternal
RSV Abs. In animals receiving a single vaccine dose, the general illness,
fever and lung consolidation readouts, and the virus concentrations in Nature Communications| (2022) 13:6142 6 Article https://doi.org/10.1038/s41467-022-33649-3 5
7
9
100
101
102
103
104
105
3
4
5
6
7
8
9
100
101
102
103
104
105
5
7
9
100
101
102
103
104
105
3
4
5
6
7
8
9
100
101
102
103
104
105
Day post-challenge
Viral load (TCID50/mL)
Day post-challenge
Viral load (TCID50/mL)
BAL
***
***
BAL
***
**
Day post-challenge
Nasal
a
b
c
d
Day post-challenge
Nasal
Viral load (TCID50/mL)
Viral load (TCID50/mL)
***
***
**
***
***
***
***
Day post-challenge
Viral load (TCID50/mL)
Day post-challenge
Viral load (TCID50/mL)
e
f
Nasal
BAL
***
***
***
Study 2 – Long DOI
ChAd155-RSV (1D)
Placebo-L
Study 1 – Long DOI
ChAd155-RSV (2D)
Placebo-L
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S
Study 1 – Short DOI
***
***
***
3
4
5
6
7
8
9
100
101
102
103
104
105
5
7
9
100
101
102
103
104
105
Fig. 4 | Viral loads. Calves (n = 7 to 9 biologically independent animals, see Table 1)
received either a single or two doses (1D or 2D, respectively) of ChAd155-RSV
vaccine or placebo, followed by a short (4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size in the placebo-L
group of Study 2 had decreased to n = 4 on day post-challenge 9; see Fig. 3). Discussion Calves (n = 7 to 9 biologically independent animals, see Table 1)
received either a single or two doses (1D or 2D, respectively) of ChAd155-RSV
vaccine or placebo, followed by a short (4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size in the placebo-L
group of Study 2 had decreased to n = 4 on day post-challenge 9; see Fig. 3). Virus
loads of lung broncho-alveolar lavage (BAL) samples (a, c, e) and nasopharyngeal Fig. 4 | Viral loads. Calves (n = 7 to 9 biologically independent animals, see Table 1)
received either a single or two doses (1D or 2D, respectively) of ChAd155-RSV
vaccine or placebo, followed by a short (4 weeks) or long (16 weeks) duration of
immunity (short DOI or long DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L groups indicate the control groups subjected to the short-
DOI and long-DOI regimens, respectively (note that the sample size in the placebo-L
group of Study 2 had decreased to n = 4 on day post-challenge 9; see Fig. 3). Virus
loads of lung broncho-alveolar lavage (BAL) samples (a, c, e) and nasopharyngeal samples (b, d, f) are presented as geometric mean concentrations with 95% con-
fidence intervals. Dotted lines (6.3 TCID50/mL) represent the lower limit of detec-
tion (LOD). Negative samples were assigned a value of 1.995. Vaccine groups were
compared by two-sided t tests with the respective control groups using ANOVA
mixed models for repeated measurements. Significant differences are indicated by
asterisks color-matched with the vaccine group indicated in the keys above the
figure (**P ≤0.01; ***P ≤0.001). Source data and statistical analyses with exact P
values are provided as a Source data and Supplementary Data 1 and 2, respectively. While a similar effect is expected to have occurred in the vaccinated
calves, they nevertheless exhibited the typical kinetics of a B cell
response to vaccination, in which nAb titers elicited by the first dose
were boosted by the second vaccination, and further boosted by the
challenge. This suggests that the vaccination had successfully induced
plasma-cell and memory B cell responses targeting RSV A Long. Discussion Day post-challenge
Viral load (TCID50/mL)
Day post-challenge
Viral load (TCID50/mL)
BAL
***
***
a
b
Nasal
***
**
***
***
***
***
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S
Study 1 – Short DOI
***
***
3
4
5
6
7
8
9
100
101
102
103
104
105
5
7
9
100
101
102
103
104
105 Day post-challenge
Viral load (TCID50/mL)
b
Nasal
***
**
***
***
***
***
ChAd155-RSV (2D)
ChAd155-RSV (1D)
Placebo-S
Study 1 – Short DOI
3
4
5
6
7
8
9
100
101
102
103
104
105 b a Day post-challenge 5
7
9
100
101
102
103
104
105
3
4
5
6
7
8
9
100
101
102
103
104
105
Day post challenge
Day post challenge
BAL
***
**
Day post-challenge
Nasal
c
d
Day post-challenge
Viral load (TCID50/mL)
Viral load (TCID50/mL)
***
Study 1 – Long DOI
ChAd155-RSV (2D)
Placebo-L
*** 3
4
5
6
7
8
9
100
101
102
103
104
105
Day post-challenge
Nasal
d
Viral load (TCID50/mL)
***
Study 1 – Long DOI
ChAd155-RSV (2D)
Placebo-L
*** d c c 5
7
9
3
4
5
6
7
8
9
5
7
9
100
101
102
103
104
105
3
4
5
6
7
8
9
100
101
102
103
104
105
Day post-challenge
Day post-challenge
Day post-challenge
Viral load (TCID50/mL)
Day post-challenge
Viral load (TCID50/mL)
e
f
Nasal
BAL
***
***
***
Study 2 – Long DOI
ChAd155-RSV (1D)
Placebo-L Day post-challenge Day post-challenge 3
4
5
6
7
8
9
100
101
102
103
104
105
Day post-challenge
Day post-challenge
Viral load (TCID50/mL)
f
Nasal
***
***
Study 2 – Long DOI
ChAd155-RSV (1D)
Placebo-L e 5
7
9
100
101
102
103
104
105
3
4
5
6
7
8
9
100
101
102
103
104
105
y p
g
y p
g
Day post-challenge
Viral load (TCID50/mL)
Day post-challenge
Viral load (TCID50/mL)
e
f
Nasal
BAL
***
***
***
Study
Ch
Pla Study 2 – Long DOI
ChAd155-RSV (1D)
Placebo-L Day post-challenge Fig. 4 | Viral loads. Calves (n = 7 to 9 biologically independent animals, see Table 1)
received either a single or two doses (1D or 2D, respectively) of ChAd155-RSV Fig. 4 | Viral loads. Discussion Virus
loads of lung broncho-alveolar lavage (BAL) samples (a, c, e) and nasopharyngeal
samples (b, d, f) are presented as geometric mean concentrations with 95% con-
fidence intervals. Dotted lines (6.3 TCID50/mL) represent the lower limit of detec-
tion (LOD). Negative samples were assigned a value of 1.995. Vaccine groups were
compared by two-sided t tests with the respective control groups using ANOVA
mixed models for repeated measurements. Significant differences are indicated by
asterisks color-matched with the vaccine group indicated in the keys above the
figure (**P ≤0.01; ***P ≤0.001). Source data and statistical analyses with exact P
values are provided as a Source data and Supplementary Data 1 and 2, respectively. Nature Communications| (2022) 13:6142 Discussion Indeed, in line with the intended pediatric use of ChAd155-
RSV, calves received their first injection at a young age. At this age, the
bovine immune system is considered not fully mature, resulting in
weak nAb responses due to the induction of low-frequency and short-
living plasma-cell responses49–51. In human infants, differentiation of B
cells into long-lived plasma cells after antigenic exposure is known to
be reduced due to lack of B cell survival signals in the bone marrow52–54. A large proportion of the vaccine target population of young
infants will have maternal-derived RSV Abs due to natural infection of
the mother, and possibly increased levels due to future maternal RSV
vaccination4,55. Discussion Moreover, many infants may receive one of the Nature Communications| (2022) 13:6142 7 Article https://doi.org/10.1038/s41467-022-33649-3 0
10
20
30
40
50
Sum histopathology scores
0
10
20
30
40
50
Consolidated lung area (%)
5
7
9
100
101
102
103
104
105
BAL titer (TCID50/mL)
3
4
5
6
7
8
9
100
101
102
103
104
105
Nasal titer (TCID50/mL)
-2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
0
1
2
3
G e n e ra l illn e s s s c o re
P la c e b o
C h A d 1 5 5 -R S V A b -
C h A d
-2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
0
1
2
3
U R T D s c o re
-2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
2 0
4 0
6 0
8 0
1 0 0
1 2 0
R e s p ira to ry ra te (/m in )
-2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
3 8 .0
3 9 .0
4 0 .0
4 1 .0
R e c ta l te m p e ra tu re (
oC )
a
b
**
c
d
Days post-challenge Days post-challenge
g
h
e
f
**
***
***
Day post-challenge
Day post-challenge
***
***
***
***
Ab- vs Ab+:
p = 0.020 at day 5
ChAd155-RSV
Ab+
Ab–
Placebo-S
ChAd155-RSV
Ab+
Ab–
Placebo-S
Study 2: Short DOI (2D ± Ab): ChAd155-RSV Ab–
ChAd155-RSV Ab+ Placebo-S
***
***
**
****
b -
*
***
***
***
***
***
Days post-challenge Days post-challenge
***
Impact of pre-existing bRSV Ab on ChAd155-RSV vaccine efficacy. In
2, calves (n = 9 biologically independent animals, see Table 1) with (Ab+) or
t (Ab−) pre-existing antibodies derived from bovine RSV Ab+ or Ab−colos-
eceived two doses (2D) of ChAd155-RSV or placebo, and were challenged
short (4 weeks) duration of immunity (DOI). On day post-challenge (dpc) 12/
mals were subjected to necropsy. Discussion The placebo-S group indicates the control
which included 9 calves through dpc 7, decreasing to 8 calves on dpc 8, and
o 6 calves on dpc 9 to 11 (as the other calves were pre-terminated due to
ng humane endpoints). Analyzed were general illness scores (a), rectal
ratures (b), upper respiratory tract disease (URTD) scores, (c), respiratory
d), consolidated lung areas (CLAs; e), lung pathological changes at termi-
time point (f), and nasopharyngeal and lung viral loads (g, h, respectively;
see Supplemental Table S1 for explanation on clinical scores). Differences between
vaccine groups and the respective control groups were determined by two-sided
ANOVA (with treatment as fixed effect and the appropriate variance assumption)
performed either on the calculated areas under the curve (a–d) or on the single-
timepoint data (e, f), or by ANOVA mixed models for repeated measurements (g, h). Means (a–f) or geometric means (g, h) with 95% confidence intervals (error bars;
a–h) for each group are color-coded according to the keys in each graph. Open
symbols in e, f indicate animals that were pre-terminated on dpc 7/8. Statistically
significant differences are presented with asterisks color-matched with the vaccine
group indicated in the key above the figure (*P ≤0.05; **P ≤0.01; ***P ≤0.001). Source data and statistical analyses with exact P values are provided as a Source
data and Supplementary Data 1 and 2, respectively. Discussion Analyzed were general illness scores (a), rectal
temperatures (b), upper respiratory tract disease (URTD) scores, (c), respiratory
rates (d), consolidated lung areas (CLAs; e), lung pathological changes at termi-
nation time point (f), and nasopharyngeal and lung viral loads (g, h, respectively; see Supplemental Table S1 for explanation on clinical scores). Differences between
vaccine groups and the respective control groups were determined by two-sided
ANOVA (with treatment as fixed effect and the appropriate variance assumption)
performed either on the calculated areas under the curve (a–d) or on the single-
timepoint data (e, f), or by ANOVA mixed models for repeated measurements (g, h). Means (a–f) or geometric means (g, h) with 95% confidence intervals (error bars;
a–h) for each group are color-coded according to the keys in each graph. Open
symbols in e, f indicate animals that were pre-terminated on dpc 7/8. Statistically
significant differences are presented with asterisks color-matched with the vaccine
group indicated in the key above the figure (*P ≤0.05; **P ≤0.01; ***P ≤0.001). Source data and statistical analyses with exact P values are provided as a Source
data and Supplementary Data 1 and 2, respectively. Fig. 5 | Impact of pre-existing bRSV Ab on ChAd155-RSV vaccine efficacy. In see Supplemental Table S1 for explanation on clinical scores). Differences between
vaccine groups and the respective control groups were determined by two-sided
ANOVA (with treatment as fixed effect and the appropriate variance assumption)
performed either on the calculated areas under the curve (a–d) or on the single-
timepoint data (e, f), or by ANOVA mixed models for repeated measurements (g, h). Means (a–f) or geometric means (g, h) with 95% confidence intervals (error bars;
a–h) for each group are color-coded according to the keys in each graph. Open
symbols in e, f indicate animals that were pre-terminated on dpc 7/8. Statistically
significant differences are presented with asterisks color-matched with the vaccine
group indicated in the key above the figure (*P ≤0.05; **P ≤0.01; ***P ≤0.001). Source data and statistical analyses with exact P values are provided as a Source
data and Supplementary Data 1 and 2, respectively. Fig. 5 | Impact of pre-existing bRSV Ab on ChAd155-RSV vaccine efficacy. Discussion -2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
3 8 .0
3 9 .0
4 0 .0
4 1 .0
R e c ta l te m p e ra tu re (
oC )
b
Days post-challenge
** -2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
0
1
2
3
G e n e ra l illn e s s s c o re
a
Days post-challenge
***
*** b R e s p ira to ry ra te (/m in )
d -2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
2 0
4 0
6 0
8 0
1 0 0
1 2 0
R e s p ira to ry ra te (/m in )
d
Days post challenge
****
Days post-challenge -2
-1
0
1
2
3
4
5
6
7
8
9
1 0
1 1
0
1
2
3
U R T D s c o re
c
Days post challenge
Days post-challenge d c c Days post-challenge 0
10
20
30
40
50
Consolidated lung area (%)
**
e
**
ChAd155-RSV
Ab+
Ab–
Placebo-S 0
10
20
30
40
50
Sum histopathology scores
f
***
***
ChAd155-RSV
Ab+
Ab–
Placebo-S f e 5
7
9
100
101
102
103
104
105
BAL titer (TCID50/mL)
h
Day post-challenge
***
***
***
Ab- vs Ab+:
p = 0.020 at day 5
ChAd155-RSV
*** 3
4
5
6
7
8
9
100
101
102
103
104
105
Nasal titer (TCID50/mL)
g
Day post-challenge
***
*
***
***
***
***
*** h h g Day post-challenge Fig. 5 | Impact of pre-existing bRSV Ab on ChAd155-RSV vaccine efficacy. In
Study 2, calves (n = 9 biologically independent animals, see Table 1) with (Ab+) or
without (Ab−) pre-existing antibodies derived from bovine RSV Ab+ or Ab−colos-
trum, received two doses (2D) of ChAd155-RSV or placebo, and were challenged
after a short (4 weeks) duration of immunity (DOI). On day post-challenge (dpc) 12/
13, animals were subjected to necropsy. The placebo-S group indicates the control
group, which included 9 calves through dpc 7, decreasing to 8 calves on dpc 8, and
then to 6 calves on dpc 9 to 11 (as the other calves were pre-terminated due to
reaching humane endpoints). Discussion Significant differences between vaccine
groups and the respective control groups are presented as asterisks color-matched
with the vaccine group indicated in the keys (*P ≤0.05; ***P ≤0.001). Triangles and
crosses below the x-axis denote the time points of placebo or vaccine injections and
the bRSV challenge, respectively. Individual values for all groups are presented in
Supplemental Fig. 5. Source data and statistical analyses with exact P values are
provided as a Source data and Supplementary Data 1 and 2, respectively. in d included calves with or without pre-existing bovine RSV Ab groups (Ab+ or Ab−,
respectively). Weeks of study: weeks post-dose 1 for all groups except short DOI-1D,
which was injected at week 4 (Study 1). Dotted lines represent the limit of detection
(LOD) i.e., 32 ED60. Negative samples were assigned the value of the LOD. Titers of
vaccine and control groups werecompared by two-sided t tests using ANOVAmixed
models for repeated measurements. Significant differences between vaccine
groups and the respective control groups are presented as asterisks color-matched
with the vaccine group indicated in the keys (*P ≤0.05; ***P ≤0.001). Triangles and
crosses below the x-axis denote the time points of placebo or vaccine injections and
the bRSV challenge, respectively. Individual values for all groups are presented in
Supplemental Fig. 5. Source data and statistical analyses with exact P values are
provided as a Source data and Supplementary Data 1 and 2, respectively. outcome. However, considering the (~3.2 times) higher dose of chal-
lenge virus in Study 2 versus Study 1, the more severe clinical data post-
challenge might be explained by a challenge dose-response effect. Such effect has also been observed in a (unpublished) in-house chal-
lenge-dose titration study. In that study, a (0.7 log10) higher challenge
dose clearly changed the kinetics of the infection, and resulted in an
earlier onset and more rapid development of (more) severe symptoms,
without affecting the lung viral loads. the recovery from such microscopic symptoms generally takes longer. Nonetheless, the two-dose (short-DOI) regimens in groups without
pre-existing responses displayed similar efficacies across the two stu-
dies. This indicated that the efficacy data from Study 2 can be used to
support the overall conclusion on ChAd155-RSV vaccine. Beyond its utility in RSV vaccine immunogenicity/efficacy studies,
the calf model is also suitable for evaluations of vaccine safety
including the risk of ERD, a major focus in pediatric RSV vaccine
development. Discussion In
Study 2, calves (n = 9 biologically independent animals, see Table 1) with (Ab+) or
without (Ab−) pre-existing antibodies derived from bovine RSV Ab+ or Ab−colos-
trum, received two doses (2D) of ChAd155-RSV or placebo, and were challenged
after a short (4 weeks) duration of immunity (DOI). On day post-challenge (dpc) 12/
13, animals were subjected to necropsy. The placebo-S group indicates the control
group, which included 9 calves through dpc 7, decreasing to 8 calves on dpc 8, and
then to 6 calves on dpc 9 to 11 (as the other calves were pre-terminated due to
reaching humane endpoints). Analyzed were general illness scores (a), rectal
temperatures (b), upper respiratory tract disease (URTD) scores, (c), respiratory
rates (d), consolidated lung areas (CLAs; e), lung pathological changes at termi-
nation time point (f), and nasopharyngeal and lung viral loads (g, h, respectively; forthcoming prophylactic anti-RSV monoclonal antibodies (mAbs)56,57. Importantly, although our results suggest that the presence of
maternal Ab may have had some negative impact on the levels of the
nAb response, as observed after the second dose, this did not appear
to have affected the protection against the bRSV challenge conferred
to these animals. The collective data thus suggest that the presence of
pre-existing RSV Abs might have at most minimal effects on the effi-
cacy of ChAd155-RSV in human infants. Nevertheless, future research
could help determining which RSV F epitopes are the main targets of the humoral immune response, and whether the observed negative
effect of pre-existing Ab has any impact on vaccine immunogenicity in
humans. The more severe clinical outcome in Study 2 compared to Study 1
was unexpected, considering the potential advantageous role of the
colostrum (with or without specific immunity), which was only pro-
vided to the calves in Study 2. Discussion Bacterial examination (by bacterial
culture and/or PCR) of BAL and lung tissue samples did not reveal any
evidence for an invasion of pathogenic bacteria that might explain this Nature Communications| (2022) 13:6142 8 Article https://doi.org/10.1038/s41467-022-33649-3 0
4
6
8
1 2
1 6
2 0
2 2
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -L
C h A d 1 5 5 -R S V (2 D )
0
4
8
1 6
1 8
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
C h A d 1 5 5 -R S V (1 D )
P la c e b o -L
0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V A b -
C h A d 1 5 5 -R S V A b +
Study 1: Long DOI (2D)
a
b
A
N
)
0
6
D
E
( r
e
tit
Weeks of study
c
b
Weeks of study
Study 1: Short DOI (1D/2D)
d
0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V (1 D )
C h A d 1 5 5 -R S V (2 D )
NAb titer (ED60)
*
***
Study 2: Short DOI (2D ± Ab)
Study 2: Long DOI (1D)
b
A
N
)
0
6
D
E
( r
e
tit
NAb titer (ED60)
*** *** ***
***
*** ***
***
Weeks of study
Weeks of study
*** *** ***
***
*
***
*********
***
***
*** ***
***
Ab- vs Ab+:
p < 0.0001 (wk 0, 6)
p < 0.01 (wk 8)
Fig. 6 | Human RSV A-specific nAb titers. Human RSV subtype A (hRSV A) neu-
tralizing antibody (nAb) titers are presented. Titers are expressed as the inverse of
the serum dilution causing 60% reduction in the number of plaques as compared to
the virus control wells (ED60). Geometricmeans with 95% confidenceintervals (error
bars) for each group are color coded according to the keys in each graph. Discussion 0
4
6
8
1 2
1 6
2 0
2 2
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -L
C h A d 1 5 5 -R S V (2 D )
Study 1: Long DOI (2D)
Weeks of study
b
NAb titer (ED60)
*
***
***
*********
*** 0
4
6
8
1 2
1 6
2 0
2 2
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -L
C h A d 1 5 5 -R S V (2 D )
0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V A b -
C h A d 1 5 5 -R S V A b +
Study 1: Long DOI (2D)
Weeks of study
b
d
NAb titer (ED60)
*
***
Study 2: Short DOI (2D ± Ab)
NAb titer (ED60)
*** ***
***
Weeks of study
***
*********
***
***
*** ***
***
Ab- vs Ab+:
p < 0.0001 (wk 0, 6)
p < 0.01 (wk 8)
in d included calves with or without pre-existing bovine RSV Ab groups (Ab+ or Ab−,
respectively). Weeks of study: weeks post-dose 1 for all groups except short DOI-1D,
which was injected at week 4 (Study 1). Dotted lines represent the limit of detection
(LOD) i.e., 32 ED60. Negative samples were assigned the value of the LOD. Titers of
vaccine and control groups werecompared by two-sided t tests using ANOVAmixed
models for repeated measurements. Significant differences between vaccine
groups and the respective control groups are presented as asterisks color-matched
with the vaccine group indicated in the keys (*P ≤0.05; ***P ≤0.001). Triangles and
crosses below the x-axis denote the time points of placebo or vaccine injections and
the bRSV challenge, respectively. Individual values for all groups are presented in
Supplemental Fig. 5. Source data and statistical analyses with exact P values are
provided as a Source data and Supplementary Data 1 and 2, respectively. Discussion a
b
A
N
)
0
6
D
E
( r
e
tit
Weeks of study
Study 1: Short DOI (1D/2D)
0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V (1 D )
C h A d 1 5 5 -R S V (2 D )
*** *** ***
***
* b a Study 1: Long DOI (2D) b
A
N
)
0
6
D
E
( r
e
tit NAb titer (ED60) Weeks of study Weeks of study Weeks of study 0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V A b -
C h A d 1 5 5 -R S V A b +
Weeks of study
d
Study 2: Short DOI (2D ± Ab)
NAb titer (ED60)
*** ***
***
Weeks of study
***
*** ***
***
Ab- vs Ab+:
p < 0.0001 (wk 0, 6)
p < 0.01 (wk 8) 0
4
8
1 6
1 8
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
C h A d 1 5 5 -R S V (1 D )
P la c e b o -L
c
Study 2: Long DOI (1D)
b
A
N
)
0
6
D
E
( r
e
tit
*** *** ***
***
Weeks of study 0
4
8
1 6
1 8
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
C h A d 1 5 5 -R S V (1 D )
P la c e b o -L
c
Study 2: Long DOI (1D)
b
A
N
)
0
6
D
E
( r
e
tit
*** *** ***
*** d c 0
4
6
8
1 0
1 0
1 0 0
1 ,0 0 0
1 0 ,0 0 0
P la c e b o -S
C h A d 1 5 5 -R S V A b -
C h A d 1 5 5 -R S V A b +
Study 2: Short DOI (2D ± Ab)
NAb titer (ED60)
*** ***
***
Weeks of study
***
*** ***
***
Ab- vs Ab+:
p < 0.0001 (wk 0, 6)
p < 0.01 (wk 8) Study 2: Long DOI (1D) Study 2: Short DOI (2D ± Ab) 1
NAb titer (ED60) Weeks of study Weeks of study Weeks of study Weeks of study Fig. Discussion 6 | Human RSV A-specific nAb titers. Human RSV subtype A (hRSV A) neu-
tralizing antibody (nAb) titers are presented. Titers are expressed as the inverse of
the serum dilution causing 60% reduction in the number of plaques as compared to
the virus control wells (ED60). Geometricmeans with 95% confidenceintervals (error
bars) for each group are color coded according to the keys in each graph. Calves
(n = 7–9 biologically independent animals, see Table 1) received a single or two
doses (1D or 2D, respectively) of ChAd155-RSV vaccine or placebo, followed by a
short (4 weeks, a, d) or long (16 weeks, b, c) duration of immunity (short DOI or long
DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L
groups indicate the control groups subjected to the short-DOI and long-DOI regi-
mens, respectively (note that the sample sizes of the placebo-L and placebo-S
groups in Study 2 decreased post-challenge; see Figs. 3 and 5, respectively). Groups Fig. 6 | Human RSV A-specific nAb titers. Human RSV subtype A (hRSV A) neu-
tralizing antibody (nAb) titers are presented. Titers are expressed as the inverse of
the serum dilution causing 60% reduction in the number of plaques as compared to
the virus control wells (ED60). Geometricmeans with 95% confidenceintervals (error
bars) for each group are color coded according to the keys in each graph. Calves
(n = 7–9 biologically independent animals, see Table 1) received a single or two
doses (1D or 2D, respectively) of ChAd155-RSV vaccine or placebo, followed by a
short (4 weeks, a, d) or long (16 weeks, b, c) duration of immunity (short DOI or long
DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L
groups indicate the control groups subjected to the short-DOI and long-DOI regi-
mens, respectively (note that the sample sizes of the placebo-L and placebo-S
groups in Study 2 decreased post-challenge; see Figs. 3 and 5, respectively). Groups in d included calves with or without pre-existing bovine RSV Ab groups (Ab+ or Ab−,
respectively). Weeks of study: weeks post-dose 1 for all groups except short DOI-1D,
which was injected at week 4 (Study 1). Dotted lines represent the limit of detection
(LOD) i.e., 32 ED60. Negative samples were assigned the value of the LOD. Titers of
vaccine and control groups werecompared by two-sided t tests using ANOVAmixed
models for repeated measurements. Nature Communications| (2022) 13:6142 Discussion Calves
(n = 7–9 biologically independent animals, see Table 1) received a single or two
doses (1D or 2D, respectively) of ChAd155-RSV vaccine or placebo, followed by a
short (4 weeks, a, d) or long (16 weeks, b, c) duration of immunity (short DOI or long
DOI, respectively), and then a bovine RSV challenge. Placebo-S and placebo-L
groups indicate the control groups subjected to the short-DOI and long-DOI regi-
mens, respectively (note that the sample sizes of the placebo-L and placebo-S
groups in Study 2 decreased post-challenge; see Figs. 3 and 5, respectively). Groups
in d included calves with or without pre-existing bovine RSV Ab groups (Ab+ or Ab−,
respectively). Weeks of study: weeks post-dose 1 for all groups except short DOI-1D,
which was injected at week 4 (Study 1). Dotted lines represent the limit of detection
(LOD) i.e., 32 ED60. Negative samples were assigned the value of the LOD. Titers of
vaccine and control groups werecompared by two-sided t tests using ANOVAmixed
models for repeated measurements. Significant differences between vaccine
groups and the respective control groups are presented as asterisks color-matched
with the vaccine group indicated in the keys (*P ≤0.05; ***P ≤0.001). Triangles and
crosses below the x-axis denote the time points of placebo or vaccine injections and
the bRSV challenge, respectively. Individual values for all groups are presented in
Supplemental Fig. 5. Source data and statistical analyses with exact P values are
provided as a Source data and Supplementary Data 1 and 2, respectively. Animals and husbandry Because of the endemic status of bRSV, calves were transported from
conventional dairy farms to the institute within 2 h after spontaneous
birth. Upon arrival in the facility, animals were housed in individual
pens with straw bedding (1.4 m2 (Study 1) and 2.2 m2 (Study 2) per
animal; max. 9 calves/room). The staff applied a strict clothing and
hygiene regimen. From ~5 weeks of age, the calves were housed group-
wise, first in pens on concrete floor and rubber mats, then, during the
challenge phase, in separate pens by group without physical contact
between groups. Floor space was adjusted by age (from 3.4 to 4.8 m2/
calf at 5 and 28 weeks of age, respectively). Rooms were temperature-
and humidity-controlled (10–24 °C; 30–80%) with HEPA-filtered ingo-
ing air. Animals were fed at least twice daily. During the first 2–3 weeks
of life, they received milk three times daily, and they were weaned at
~9 weeks of age. From 2 to 3 weeks of age, they also received a corn mix
followed by hay, grass pellets and concentrates. Drinking water was
supplied ad libitum. ChAd155-RSV vaccine and bRSV challenge stock preparation
The ChAd155-RSV investigational vaccine consists of a recombinant
replication-defective ChAd155 vector construct, engineered to express
the RSV F protein deleted of the transmembrane region, and the RSV N
and M2-1 proteins, as described previously for this antigen when
inserted in a different simian adenoviral vector63. The F protein
sequence is a consensus sequence, derived from the alignment of
different subgroup A isolates retrieved from the National Centre for
Biotechnology
Information
database. The
transmembrane
and
C-terminal domains of the F sequence were deleted, and no pre-fusion
stabilizing mutations were included. All antigens were codon-
optimized for expression in eukaryotic cells. For the challenge, an in
vivo-passaged bRSV (strain Odijk, subtype A, passage 6) was used,
which was originally isolated from a calf in the field suffering from
acute respiratory disease64. The in vivo passage was performed in
cesarian-derived specific-pathogen-free calves. BALs used for experi-
mental challenge contained at least 102.5 TCID50/mL bRSV, and were
tested free from major bovine respiratory viruses, mycoplasmas, and
bacteria, by evaluating three in vitro passages on sensitive cell cultures
and classical bacterial and mycoplasma culture. The calf populations consisted of 72% males and 18% females in
Study 1 (n = 39), and of 60% males and 40% females in Study 2 (n = 45). Ethical clearance Prior to the studies’ initiation, protocols were submitted for ethical
review and approved by GSK’s ethical committee (approval no:
S001698 for Study 1, S003976 for Study 2). Both studies were con-
ducted in Wageningen University and Research (WUR; Wageningen
Bioveterinary Research institute, Lelystad, The Netherlands) in accor-
dance with the Dutch Law on Animal Experiments and the European
legislations and guidelines (2010/63/EG and ETS 123). Study 1 was
authorized by the Animal Ethics Committee of the Animal Sciences
Group of WUR. Study 2 was licensed by the Dutch Central Authority for
Scientific
Procedures
on
Animals
(no. AVD401002015194)
and
approved by WUR’s Animal Welfare Body. Blood samples were collected at different time points pre- and
post-vaccination and post challenge to assess vaccine immuno-
genicity. Upon bRSV challenge, clinical vaccine efficacy was eval-
uated through observations of clinical symptoms, measurements of
rectal temperature; as well as collection of nasopharyngeal brush
samples and broncho-alveolar lavage (BAL) samples, to measure
viral loads. At dpc 12 or 13, calves were euthanized. Lungs of all
animals were subjected to post-mortem examination to evaluate
lung macroscopic and microscopic lesions, focusing on macro-
scopic CLAs and microscopic pathological changes. For the pre-
terminated control animals in Study 2, these examinations were
performed at dpc 7 or 8. Article https://doi.org/10.1038/s41467-022-33649-3 antibodies, the individual serum total protein concentrations varied
from 52–67 g/L and 48–79 g/L, respectively (diagnostic reference
value: 55–85 g/L). by technical hurdles in the preparation of these vaccines, leading to
variable vaccine formulations and inconsistent reproduction of
enhanced disease (across different studies or across bovine FI-RSV-
vaccinated calves within the same study)30,61,62. Nonetheless, we
observed that all ChAd155-RSV-vaccinated, bRSV-challenged young
calves were either free of clinical/pathological symptoms, or exhibited
symptoms that were much less severe than in the PBS-injected and
bRSV-challenged control groups, while in case of an ERD response the
symptoms in vaccinated animals would have been more severe than in
the controls. In conclusion, our observations suggest that the risk that
ChAd155-RSV will induce ERD in human infants is extremely low, sup-
porting the further clinical development of this vaccine. In Study 1, newborn calves (n = 7 or 8 per group; Table 1) tested
negative for anti-bRSV Abs were injected either once or twice (4 weeks
apart) with ChAd155-RSV or PBS. Challenge occurred either 4 weeks or
16 weeks after the last vaccine dose (referred to as the short or the long
DOI), on the same calendar day across groups. In Study 2, two groups of 9 calves each were fed bRSV Ab-positive
colostrum as described above, and received two doses (4 weeks apart)
of either ChAd155-RSV or PBS. A third group (n = 9) was fed bRSV Ab-
negative colostrum and received two doses of ChAd155-RSV, serving to
bridge Studies 1 and 2 and as a bRSV Ab-negative control. All three
groups were challenged with bRSV after the short DOI, to evaluate the
impact of pre-existing RSV Abs on vaccine immunogenicity and effi-
cacy. Two other groups of 9 calves each in Study 2 were fed Ab-
negative colostrum, and received one dose of either ChAd155-RSV or
PBS. These animals were challenged with bRSV after the long DOI. In
Study 2, a total of eight animals in both placebo groups reached
humane endpoints (as defined in “Clinical assessments” below) before
termination of the study and were euthanized due to severe clinical
signs of bRSV infection (see “Results”). Animals and husbandry They were in majority dairy (Holstein) calves. Randomization in both
studies was performed while allowing for an even distribution of
gender, age, and breed across the treatment groups. Study designs Two placebo (PBS)-controlled calf studies were performed according
to the design displayed in Fig. 1 and detailed in Table 1. Upon arrival in
the animal facilities, calves in Study 1 were kept colostrum-deprived,
and calves in Study 2 received a single feeding with either bRSV-Ab-
negative or bRSV Ab-positive colostrum within 6 h after birth. The
colostrum was pre-collected and frozen on Scandinavian (bRSV Ab-
negative) or Dutch (bRSV Ab-positive) farms, and screened for (n)Abs
and IgG content prior to feeding. The bRSV Ab-positive colostrum
used was pooled just prior to the actual recruitment of the newborn
calves. All these calves were fed by stomach tube with 3–4 L of Ab-
negative colostrum or 3.2 L of Ab-positive colostrum, directly upon
arrival. Anti-bRSV antibody levels in calf sera obtained 1 day post
colostrum feeding showed a range of 8.3–9.3 log2 ELISA antibodies and
4.0–6.0 log2 bRSV nAbs. In calves with or without maternal-derived Discussion In calves, this phenomenon has been characterized by
advanced and exacerbated clinical symptoms upon bRSV challenge
relative to control bRSV-challenged animals, and by Th2-skewed
cytokine profiles, IgE Ab production, pulmonary eosinophilia, and a
high ratio of non-functional Abs to nAbs30,37,38,58–60. Apart from the
relatively small sample size (due in part to the husbandry requirements
for cattle), the absence of a positive control vaccine (bovine FI-RSV)
inducing ERD may therefore be a potential limitation of our studies. This was done considering that the current research work was not
designed as a safety study to extensively investigate all ERD signs or
characteristics. Moreover, bovine FI-RSV studies are often hampered The higher CLA scores of controls in Study 2 may also be related
to the more severe challenge condition. However, this difference
between studies may have been exacerbated by the fact that the mean
CLA score at necropsy was calculated on data from all animals,
including those that were terminated 5 or 6 days earlier, when they
most likely have been at their disease peak. Indeed, the overall higher
CLA scores in the pre-terminated versus surviving animals is consistent
with data showing a trend of an association between disease severity
scores and CLA scores30. Similarly, histopathology scores were also
higher in Study 2, though for these scores no difference was found
between pre-terminated and recovered animals. This may be because Nature Communications| (2022) 13:6142 9 Article Vaccination and bRSV challenge In both studies, calves were vaccinated intramuscularly in the pre-
scapular area with 5×1010 viral particles of ChAd155-RSV in a 2 mL
volume. Four or 16 weeks post-last immunization, calves were
challenged by aerosolized inoculation of 2 mL bRSV (strain Odijk,
doses: 3.2 log10 TCID50 in Study 1, and 3.7 log10 TCID50 in Study 2)
using an airbrush (Harder & Steenbeck), using a method described
by Antonis et al.30. No metaphylaxis was used during the bRSV
challenge phase. Nature Communications| (2022) 13:6142 10 https://doi.org/10.1038/s41467-022-33649-3 Article course of viral replication. Subsequently, a selection of these samples
collected on various time points was also analyzed by virus titration. Infectious virus titers were determined in duplicate by end-point
titration of 10-fold sample dilutions on immortalized embryonic
bovine tracheal cells (produced at Wageningen Bioveterinary Research
[WBVR], Lelystad, The Netherlands), seeded in 96-well plates. Dupli-
cate plates were incubated for 6–7 days at 37 °C and 5% CO2. At the end
of the incubation period, virus-infected cells were detected with an
anti-F bRSV mAb (“mAb3,” produced at WBVR, diluted 1:100), followed
by rabbit anti-mouse antibody conjugated with horseradish perox-
idase (Dako P0260, 1:200) followed by chromogenic substrate AEC
(Sigma A-6926)65,66. For each duplicate plate, the virus titer was
determined, and expressed as the endpoint titer at which 50% of the
cell monolayers was infected, as calculated according to Reed and
Munch approach. The detection limit was 6.3 TCID50/mL. Samples
tested negative were assigned a value of 2 TCID50/mL. Neutralization assay Neutralization assay
Briefly, decomplemented sera were serially diluted twofold in DMEM
medium with 3% FBS, 50 µg/mL gentamicin and 200 mM L-Gluta-
mine; RSV medium). Eight serial dilutions were tested for each serum. Samples and positive control dilutions were mixed with human RSV A
Long (ATCC, VR-26) diluted to approximately 100 plaque-forming
units/well, and incubated for either 20 min at 33 oC (Study 1) or
120 min at 35 °C (Study 2). After incubation, the virus–serum mixture
was transferred to Vero (ATCC, CCL-81) cell-seeded plates, with virus-
only wells as 100% infectivity control, while two wells received no
virus or serum (cell controls). Plates were incubated for 2 h at 35 °C,
then the medium was removed and RSV medium containing 0.5%
carboxymethylcellulose (low viscosity) was added to all wells. The
plates were incubated for 36 h at 33 °C before staining. Staining was
performed with a goat anti-RSV polyclonal antibody (Meridian
#B65860G, diluted 1:400) followed by rabbit anti-goat immunoglo-
bulin G (IgG) fluorescein isothiocyanate-conjugated (FITC; Millipore
#AP106F, diluted 1:1000; study 1) or rabbit anti-goat IgG-horse radish
peroxidase
(HRP)-conjugated
(Rockland
605-403-B69,
diluted
1:1000, study 2). Briefly, cell monolayers were washed with PBS and
fixed with 1% paraformaldehyde. After blocking with 2% powdered
skim milk/PBS, RSV-positive cells were detected using the goat RSV
antiserum followed by the rabbit anti-goat IgG conjugate, diluted in
blocking buffer containing a final concentration of 0.01% Evans Blue
as counter stain. When the HRP conjugate was used, after Ab staining,
True Blue substrate (KPL #5510-0049) was added to all wells to reveal
the infectious foci. Plates were scanned using an AxioVision reader
and software. Reciprocal nAb titers were expressed in ED60, deter-
mined as the inverse of the serum dilution causing 60% reduction in
the number of plaques as compared to the control wells (virus only,
no serum). Processing of samples for viral load
Nasopharyngeal brush and BAL samples were kept on melting ice
immediately following the sampling procedure. In the laboratory,
brushes were removed from the medium. Samples were centrifuged
for 10 min at 4 °C, at 1300 × g or 250 × g for nasopharyngeal and BAL
samples, respectively. Supernatants were stored at −70 ± 10 °C until
further analysis. Statistical analyses All data were analyzed using analysis of variance models with the
appropriate variance and co-variance assumptions and without con-
sidering multiplicity of comparisons, in SAS 9.4. The models either
considered only group as fixed effect for necropsy endpoints and AUC
data or group, time, and the interaction between time and group as
fixed effects for endpoints with repeated measures (nAbs and viral
loads). Group comparisons were assessed by two-sided t tests on
estimates computed from those models with alpha level set to 0.05. Missing values due to premature deaths on study 2, day 124, were
treated as such for nAbs and viral loads endpoints. For clinical endpoints, Areas Under the Curve (AUC) were com-
puted from post challenge data for each animal. To avoid sample size
reduction, missing values due to premature deaths in placebo groups
from study 2 were replaced by the animal’s average from dpc 5 to its
death and AUC value for general illness in healthy animals (AUC = 0) Necropsy and pathological analysis On dpc 12/13 (or day 7/8 for pre-terminated animals), calves of each
treatment group were euthanized by pentobarbital overdose and
exsanguination. The lungs were removed, and dorsal and ventral
images were taken to calculate the extent of macroscopic lesions
(Image Pro Premier 64-bit software). The CLA is presented as per-
centage of the total lung area. Lung samples were collected, if
applicable, on a transition between normal and consolidated tissue,
stored in 10% neutral buffered formalin, and embedded in paraffin. For histological examination, 5 μm sections of the right apical and
cardiac lobes and the left cardiac lobe were stained with hematoxylin
and eosin. Each tissue section was scored from 0 (absent) to 4
(severe) for the following categories of histopathological changes
according to the scoring table (Supplemental Table 2): endo-
bronchi(oli)tis, peri-bronchitis/vasculitis, interstitial pneumonia and
alveolitis. CLA calculations and histological examination (by a Eur-
opean College of Veterinary Pathologists board-certified veterinary
pathologist) were performed blinded, as a single independent
assessment. Clinical assessments Upon challenge, bRSV-related symptoms were monitored daily by the
same veterinarian, who was blinded for the treatment groups. Clinical
scores for general illness (including depression and loss of appetite),
upper respiratory tract disease (URTD; nasal and ocular discharge and
cough) and LRTD (increased breathing effort) were assessed according
to the scoring system outlined in Supplemental Table 1. The LRTD
score was partially based on the actual respiratory rate, which was
counted by visual inspection. In addition, rectal temperatures were
measured twice daily after the challenge. Humane endpoint criteria
were defined as persistent severe depression (max. 24 h), persistent
severe dyspnea e.g., respiratory rate of >100 breathings/minute (max. 48 h), or non-reversible respiratory failure such as intermittent open
mouth breathing or frothing (immediately). Sample collection Blood samples for serology (8 mL) were collected at different time
points (see Fig. 6) by venipuncture of the V. jugularis. Just prior to
challenge and on dpc 1 to 9, nasopharyngeal samples were collected
using sterile nylon bristle brushes (MW126, Medical Wire and Equip-
ment Co. Ltd). Following sampling, brushes were directly agitated in
3 mL transport medium, consisting of EMEM (GIBCO), supplemented
with 2% antibiotics and 2% fetal bovine serum (FBS). BAL samples were
collected 6 days prior to challenge and on dpc 5, 7, and 9 from all calves
(non-sedated) regardless of clinical presentation. BAL fluid was
obtained from the caudal part of the lung after instillation of
approximately 100 mL D-Phosphate Buffered Saline (GIBCO) via intu-
bation of the ventral nasal meatus. Fluid recovered from the lung (50-
80% of the instilled volume) was directly supplemented with 5% FBS. On the days of BAL sampling, clinical disease scores were assessed at
least 2 h after sample collection, to prevent the interference of any
potential transient effect of the BAL procedure on the clinical
presentation. References 20. Voysey, M. et al. Safety and efficacy of the ChAdOx1 nCoV-19 vac-
cine (AZD1222) against SARS-CoV-2: an interim analysis of four
randomised controlled trials in Brazil, South Africa, and the UK. Lancet 397, 99–111 (2021). 1. Hall, C. B. et al. Respiratory syncytial virus-associated hospitaliza-
tions among children less than 24 months of age. Pediatrics 132,
e341–e348 (2013). 2. Shi, T. et al. Global, regional, and national disease burden estimates
of acute lower respiratory infections due to respiratory syncytial
virus in young children in 2015: a systematic review and modelling
study. Lancet 390, 946–958 (2017). 2. Shi, T. et al. Global, regional, and national disease burden estimates
of acute lower respiratory infections due to respiratory syncytial
virus in young children in 2015: a systematic review and modelling
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http://www.nature.com/reprints Publisher’s note Springer Nature remains neutral with regard to jur-
isdictional claims in published maps and institutional affiliations. https://doi.org/10.1038/s41467-022-33649-3 and N.S.-Z. participated in the acquisition of data. R.d.J., A.-M.S., J.-F.T.,
H.S., I.D., K.-F.W., and S.V. analyzed and interpreted the results. All
authors were involved in drafting the manuscript or revising it critically
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sponding author. Peer review information Nature Communications thanks Laurel Gersh-
win and the other anonymous reviewer(s) for their contribution to the
peer review of this work. and N.S.-Z. participated in the acquisition of data. R.d.J., A.-M.S., J.-F.T.,
H.S., I.D., K.-F.W., and S.V. analyzed and interpreted the results. All
authors were involved in drafting the manuscript or revising it critically
for important intellectual content. All authors had full access to the data
and approved the manuscript before it was submitted by the corre-
sponding author. Competing interests p
g
K.-F.W., B.B., J.-F.T., I.D., H.S., and A.-M.S. are, or were, at the time of the
study employees of the GSK group of companies. A.-M.S., H.S., J.-F.T.,
and I.D. report ownership of GSK shares and/or restricted GSK shares. S.V. is an employee of CONSULTYS Benelux S.A., contracted by Glax-
oSmithKline Biologicals SA in the context of this study. R.d.J., N.S.-Z.,
and J.B. are employees of the Wageningen Bioveterinary Research,
Lelystad, The Netherlands, contracted by GlaxoSmithKline Biologicals
SA in the context of this study. Parts of salaries of R.d.J., N.S.-Z., and J.B. were paid from the contract fee. Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
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use is not permitted by statutory regulation or exceeds the permitted
use, you will need to obtain permission directly from the copyright
holder. To view a copy of this license, visit http://creativecommons.org/
licenses/by/4.0/. Author contributions 51. Dietert, R. R. et al. Workshop to identify critical windows of exposure
for children’s health: immune and respiratory systems work group
summary. Environ. Health Perspect. 108, 483–490 (2000). A.-M.S., J.-F.T., and R.d.J. were involved in the conception and design of
the study and/or the development of the study protocol. R.d.J., J.B., B.B., Nature Communications| (2022) 13:6142 13 Article https://doi.org/10.1038/s41467-022-33649-3 Nature Communications| (2022) 13:6142 Additional information Supplementary information The online version contains Supplementary information The online version contains
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Metabolic Reprogramming in Modulating T Cell Reactive Oxygen Species Generation and Antioxidant Capacity
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Metabolic Reprogramming in
Modulating T Cell Reactive Oxygen
Species Generation and Antioxidant
Capacity Josephin N. Rashida Gnanaprakasam, Ruohan Wu and Ruoning Wang* A robust adaptive immune response requires a phase of proliferative burst which is fol-
lowed by the polarization of T cells into relevant functional subsets. Both processes are
associated with dramatically increased bioenergetics demands, biosynthetic demands,
and redox demands. T cells meet these demands by rewiring their central metabolic
pathways that generate energy and biosynthetic precursors by catabolizing and oxidiz-
ing nutrients into carbon dioxide. Simultaneously, oxidative metabolism also produces
reactive oxygen species (ROS), which are tightly controlled by antioxidants and plays
important role in regulating T cell functions. In this review, we discuss how metabolic
rewiring during T cell activation influence ROS production and antioxidant capacity. Edited by: Edited by:
Heitor Affonso Paula Neto,
Universidade Federal do
Rio de Janeiro, Brazil Reviewed by:
Martin Rottenberg,
Karolinska Institutet (KI), Sweden
Pedro Barcellos-de-Souza,
Instituto Nacional de
Câncer (INCA), Brazil Keywords: reactive oxygen species, oxidative stress, metabolism, T cell, antioxidant INTRODUCTION T cells are central orchestrators of antigen-specific adaptive immunity and tolerance. Upon stimula-
tion of antigen receptors, T cells rapidly transit from naïve to an active state followed by massive
clonal expansion. Depending on the nature of pathogens and the surrounding cytokine milieu,
proliferating T cells can differentiate into diverse phenotypic and functional subsets to elicit a robust
immune response. After the clearance of pathogens, the majority of effector T cells die through apop-
tosis and the remaining memory T (Tmem) cells are responsible for immunity upon re-exposure to the
same pathogen. Accumulating evidence suggests that a coordinated rewiring of cellular metabolism
is required for T cell activation and differentiation by fulfilling the bioenergetic, biosynthetic, and
redox demands (1–9). Importantly, different phenotypic and functional T cell subsets are character-
ized by distinct metabolic programs (Table 1), which are largely controlled by immune modulatory
signaling cascades (10–17). Naïve T (Tnai) cells, Tmem cells, and immune-suppressive regulatory T (Treg)
cells predominantly rely on fatty acid oxidation (FAO) and oxidative phosphorylation (OXPHOS)
to meet their relatively low-energy needs (14, 15, 18, 19). However, persistent aerobic glycolysis,
the pentose phosphate pathway (PPP), and glutaminolysis are required to drive cell growth, clonal
expansion, and effector functions in both CD4+ subsets and CD8+ effector T (Teff) cells (Table 1)
(10, 15, 16, 18, 20–31). Th
b l
l
d
b
d
h
d
l *Correspondence:
Ruoning Wang
ruoning.wang@
nationwidechildrens.org Specialty section:
This article was submitted to
Molecular Innate Immunity,
a section of the journal
Frontiers in Immunology Received: 16 December 2017
Accepted: 30 April 2018
Published: 16 May 2018 Review
published: 16 May 2018
doi: 10.3389/fimmu.2018.01075 MITOCHONDRIAL OXPHOS AND NADPH
OXIDASES (NOXs) IN GENERATING ROS
IN T CELLS important source of ROS in T cells (38, 39). The ROS production
by NOX is regulated at various levels including the assembly of
functional NOX complex, the availability of prosthetic group,
flavin adenine dinucleotide, the intracellular concentration of
calcium, cell surface receptor signals mediated by G protein-
coupled receptors, complement, T cell receptor (TCR), and CD28
(35–37, 40, 41). The mitochondria are the central metabolic hub and powerhouse
of all eukaryotic cells. The oxidation of acetyl-CoA to carbon
dioxide (CO2) by the tricarboxylic acid (TCA) cycle is the central
metabolic process for fueling ATP production. While glycolysis
and FAO primarily provide the OXPHOX substrate, acetyl-CoA,
for mitochondria in Tnai cells, Tmem cells, and Treg cells (14, 15, 18, 19),
heightened mitochondrial biogenesis during T cell activation
leads to higher numbers of mitochondria and likely the enhanced
mitochondrial dependent metabolic flux in Teff cells compared
with Tnai cells (23, 32, 33). In particular, a surplus of 3-, 4-, and
5-carbon metabolites (anaplerotic substrates) including pyruvate,
malate, and α-ketoglutarate (α-KG) feed into the TCA cycle during
the catabolism of glutamine and other amino acids (5, 13, 15, 34). The electron transport chain (ETC) constantly transfers electrons
from NADH and FADH2 to oxygen while allowing protons (H+)
to pass through the inner mitochondrial membrane to form
an electrochemical proton gradient that drives ATP synthesis. However, both protons and electrons can leak from the ETC
due to the uncoupling of ATP synthase from the proton gradient
and a premature exit of electron before reaching cytochrome
c oxidase, respectively. Electron leak largely occurs at the sites
of complex I (NADH–ubiquinone oxidoreductase) and complex
III (ubiquinone–cytochrome c oxidoreductase) in the ETC and
results in the partial reduction of oxygen, generating superoxide
O2−•
(
). Subsequently, mitochondrial dismutase acts to convert
superoxide to hydrogen peroxide (H2O2), which is free to diffuse
into cytosol and act as a redox signaling molecule to elicit differ-
ent cellular responses (35–37). Thus, increased ROS production
in T cells can occur as a result of metabolic reprogramming
during T cell activation. Besides mitochondria, cytoplasmic ROS
is generated by NOXs, which is also an important source of ROS
in T cell. NOX family proteins are transmembrane proteins that
transport the electrons from nicotinamide adenine dinucleotide
(phosphate), NAD(P)H, to oxygen and generate superoxide anion
as the intermediate product of oxidase and subsequently H2O2, as
the product of dismutation of the superoxide. Citation: Rashida Gnanaprakasam JN, Wu R
and Wang R (2018) Metabolic
Reprogramming in Modulating
T Cell Reactive Oxygen Species
Generation and Antioxidant Capacity. Front. Immunol. 9:1075. doi: 10.3389/fimmu.2018.01075 These metabolic programs actively support ATP production by providing mitochondrial OXPHOS
substrates, support biomass accumulation by generating metabolic precursors for the biosynthesis
of protein, lipids, and nucleic acids, and maintain redox balance through generation and elimination
of reactive oxygen species (ROS). May 2018 | Volume 9 | Article 1075 Frontiers in Immunology | www.frontiersin.org 1 Metabolism in T cell redox balance Rashida Gnanaprakasam et al. Table 1 | The metabolic profiles of T cell subsets. T cell type
Naïve
Active
Differentiated
Memory T cell (Tmem)
Metabolic profile
FAO
OXPHOS
Aerobic glycolysis
PPP
Glutaminolysis
Th1: aerobic glycolysis/some OXPHOS
FAO
OXPHOS
Th2: aerobic glycolysis
Th9: aerobic glycolysis
Th17: aerobic glycolysis, glutaminolysis
Tfh: aerobic glycolysis, OXPHOS
Treg: FAO, OXPHOS
CTL: aerobic glycolysis
CAT: oxidation, phosphorylation
FAO, fatty acid oxidation; OXPHOS, oxidative phosphorylation; PPP, pentose phosphate pathway; Th1, T helper 1 cell; Th2, T helper 2 cell; Th9, T helper 9 cell; Th17, T helper 17
cell; Tfh, follicular helper T cell; Treg, regulatory T cell; CTL, cytotoxic T lymphocyte; CAT, chronically activated T cell; Tmem, memory T. OS, oxidative phosphorylation; PPP, pentose phosphate pathway; Th1, T helper 1 cell; Th2, T helper 2 cell; Th9, T helper 9 cell; Th17, T help
reg, regulatory T cell; CTL, cytotoxic T lymphocyte; CAT, chronically activated T cell; Tmem, memory T. MITOCHONDRIAL OXPHOS AND NADPH
OXIDASES (NOXs) IN GENERATING ROS
IN T CELLS There are different
isoforms of the NOX enzyme including NOX1, NOX2, NOX3,
NOX4, NOX5, dual oxidase 1, and DUOX2, and the expression
of these subunits varies among different tissues. NOX-2 is an Frontiers in Immunology | www.frontiersin.org ROS SIGNALING IN REGULATING T CELL
ACTIVATION AND DIFFERENTIATION However, physiologically relevant levels of ROS mediate essential redox signaling through nodulation of a wide spectrum of redox-sensitive
transcription factors to drive T cell activation and function. Figure 1 | Mitochondria and NADPH oxidases (NOX)-derived reactive oxygen species (ROS) regulates T cell activation, differentiation, and metabolism. Mitochondria and NOX are the two major sources of ROS. The stimulation of T cell receptor (TCR) initiates signaling and metabolic events that drive ROS production
in cytoplasm through NOX-dependent reaction and ROS production in mitochondria via mitochondria electron transport chain (ETC). Excess ROS causes damage
and cell death. However, physiologically relevant levels of ROS mediate essential redox signaling through nodulation of a wide spectrum of redox-sensitive
transcription factors to drive T cell activation and function. can lead to uncontrolled hyper-inflammation and tissue dam-
age. Thus, a fine-tuned balance between ROS production and
antioxidant capacity ensures appropriate levels of intracellular
ROS (Figure 2) (44, 55, 62). GSH, a tripeptide of glutamine,
cysteine, and glycine, is the most abundant antioxidant capable
of providing reducing equivalents and also serves as a versatile
nucleophilic cofactor in a wide spectrum of metabolic reactions
in aerobic organisms (63, 64). Thioredoxin (TXN) is a class of
small redox proteins that are involved in modulating cell surface
receptors and confers tolerance to oxidative stress in T cells
(65–69). A reciprocal redox reaction can be coupled between
these two antioxidant systems to act as a backup for each other
under certain conditions (70–77). Supporting these findings,
the inhibition of thioredoxin reductase (TXNRD) conferred an
increased susceptibility of cancer cells to GSH depletion (78–80). Glutathione-disulfide reductase (GSR) regenerates GSH from its
oxidized form, glutathione disulfide (GSSG), whereas TXNRD
is responsible for the regeneration of TXN once it has been
oxidized. Importantly, both GSR and TXNRD require NADPH
as a reducing agent. Upon antigen stimulation, both PPP and
glutaminolysis are significantly upregulated and further enhance
T cell antioxidant capacities by generating NADPH through and FasL and mitochondrial membrane potential (43, 49–52). NOX-derived ROS modulates the function of GATA-binding
protein 3, signal transducer and activator of transcription, and
T-box transcription factor to collectively control T cell activation
and differentiation. T cells from NOX-deficient animals showed
a skewed Th17 phenotype, whereas NOX-intact cells exhibited a
preferred Th1 response (39, 53–55). In CD8 T cells, NOX-derived
ROS is involved in regulating the production of IFN-γ and CD39
expression through c-Jun N-terminal kinase and NFκB signaling
(40, 56). ROS SIGNALING IN REGULATING T CELL
ACTIVATION AND DIFFERENTIATION T cell activation requires ligation of TCR and the major histo-
compatibility complex molecules. This interaction will initiate the
signaling cascade and activation of transcriptional factors such
as nuclear factor of activated T cells (NFAT), activator protein
1 (AP-1), and nuclear factor of kappa light chain enhancer in
B cells (42). It has been reported that TCR ligation increases
the production of ROS from OXPHOS and cytoplasmic ROS
from NADPH oxidases (NOXs), a family of plasma membrane-
associated oxidases (36, 40, 41). ROS-mediated signaling events
are required for driving T cell activation, proliferation, and dif-
ferentiation (Figure 1) (36, 41). T cells with reduced production
of mitochondrial ROS display impaired production of interleu-
kin-2 and antigen-specific proliferation, indicating an essential
signaling role for mitochondrial ROS in driving optimal TCR
signaling. The proximal TCR signaling machinery, including
zeta chain-associated protein kinase 70, linker of activated T cell,
SH2 domain-containing leukocyte protein, phospholipase Cγ1,
and protein kinase Cθ, is involved in driving ROS production
upon T cell activation (36, 41, 43). Conversely, physiologically
relevant levels of ROS facilitate the activation of oxidation-
dependent transcription factors, such as NF-κB and AP-1, which
are required for driving essential signaling events to support
T cell-mediated immune responses (44–46). However, exces-
sive ROS production following ablation of de novo glutathione
(GSH) synthesis suppresses the activity of mammalian target
of rapamycin and the expression of transcription factors NFAT
and c-MYC, the latter of which control metabolic reprogram-
ming following T cell activation (15, 47, 48). Thus, T cells fail to
meet their increased energy and biosynthetic needs and display
compromised proliferation (48). In addition, uncontrolled ROS
production is involved in the activation-induced T-cell death by
affecting expression of apoptosis related genes including Bcl-2 May 2018 | Volume 9 | Article 1075 Frontiers in Immunology | www.frontiersin.org 2 Rashida Gnanaprakasam et al. Metabolism in T cell redox balance Figure 1 | Mitochondria and NADPH oxidases (NOX)-derived reactive oxygen species (ROS) regulates T cell activation, differentiation, and metabolism. Mitochondria and NOX are the two major sources of ROS. The stimulation of T cell receptor (TCR) initiates signaling and metabolic events that drive ROS production
in cytoplasm through NOX-dependent reaction and ROS production in mitochondria via mitochondria electron transport chain (ETC). Excess ROS causes damage
and cell death. ROS SIGNALING IN REGULATING T CELL
ACTIVATION AND DIFFERENTIATION Importantly, the impact of ROS on T cell activation can
be extended to the later T cell differentiation stages. Fine tun-
ing of ROS is required for polarizing T cell in part by engaging
lineage-specific transcription factors and modulating cytokine
profiles, and consequently directs T cell-mediated inflammatory
responses (39, 40, 53–55, 57–61). METABOLIC PATHWAYS IN MODULATING
ANTIOXIDANT CAPACITIES Excessive ROS production causes collateral damage to macro-
molecules, cellular organelles, and eventually necrosis, which May 2018 | Volume 9 | Article 1075 Frontiers in Immunology | www.frontiersin.org 3 Metabolism in T cell redox balance Rashida Gnanaprakasam et al. Figure 2 | T cell metabolic programs that link to reactive oxygen species (ROS) production and the de novo synthesis of GSH. Pyruvate that is derived from
glucose via glycolysis is shuttled to the mitochondria and drives the tricarboxylic acid (TCA) cycle and fuels oxidative phosphorylation (OXPHOS). Glucose-derived
glucose-6-phosphate feeds into the pentose phosphate pathway (PPP) and produces NADPH in the cytoplasm. In addition, glutamate feeds the TCA cycle through
α-ketoglutarate (α-KG) to fuel OXPHOS and generate ROS. Excessive ROS production is regulated by glutathione (GSH), a tripeptide of glutamine, cysteine, and
glycine, which is synthesized de novo by glutamate-cysteine ligase (GCL) and glutathione synthetase (GSS). In addition, NADPH, glutathione-disulfide reductase
(GSR), and glutathione peroxidase (GPX) are involved in regenerating GSH from glutathione disulfide (GSSG), whereas thioredoxin reductase (TXNRD) is responsible
for the regeneration of thioredoxin (TXN) to control oxidative stress in T cell. Figure 2 | T cell metabolic programs that link to reactive oxygen species (ROS) production and the de novo synthesis of GSH. Pyruvate that is derived from
glucose via glycolysis is shuttled to the mitochondria and drives the tricarboxylic acid (TCA) cycle and fuels oxidative phosphorylation (OXPHOS). Glucose-derived
glucose-6-phosphate feeds into the pentose phosphate pathway (PPP) and produces NADPH in the cytoplasm. In addition, glutamate feeds the TCA cycle through
α-ketoglutarate (α-KG) to fuel OXPHOS and generate ROS. Excessive ROS production is regulated by glutathione (GSH), a tripeptide of glutamine, cysteine, and
glycine, which is synthesized de novo by glutamate-cysteine ligase (GCL) and glutathione synthetase (GSS). In addition, NADPH, glutathione-disulfide reductase
(GSR), and glutathione peroxidase (GPX) are involved in regenerating GSH from glutathione disulfide (GSSG), whereas thioredoxin reductase (TXNRD) is responsible
for the regeneration of thioredoxin (TXN) to control oxidative stress in T cell. GSH, the glucose, or glutamine starvation significantly dampens
T cell activation (10, 13, 15, 20, 48). GSH, the glucose, or glutamine starvation significantly dampens
T cell activation (10, 13, 15, 20, 48). metabolic reactions that are controlled by glucose-6-phosphate
dehydrogenase, phosphoglycerate dehydrogenase, malic enzyme
1, and isocitrate dehydrogenase 1. The intracellular GSH con-
centrations are normally in a range of three orders of magnitude
higher than extracellular GSH. METABOLIC PATHWAYS IN MODULATING
ANTIOXIDANT CAPACITIES Even though some cells are able
to recycle extracellular GSH, it may only play a minor role in
maintaining intracellular GSH pool (63, 64, 81–86). By contrast,
both the regeneration of GSH from GSSG (recycling pathway)
and de novo synthesis of GSH, by glutamate-cysteine ligase
(GCL) and glutathione synthase (GS), are required to maintain
intracellular GSH levels (64, 87). The ligation of glutamate and
cysteine to form dipeptide ϒ-glutamylcysteine (ϒ-GC) is the first
and also the rate-limiting step of GSH de novo synthesis, which
is controlled by ATP-dependent ligase GCL, a heterodimer of
a catalytic subunit (GCLC) and modifier subunit (GCLM). Subsequently, GSH is formed by GS-mediated ligation of ϒ-GC
and glycine (88, 89). Thus, the supply of intracellular cysteine,
glycine, and glutamate must fulfill the need of de novo synthesis of
GSH during T cell activation. Supporting this idea, the metabolic
processes that are involved in providing three amino acids are
tightly regulated upon T cell activation (13, 15, 90–92). Upon
T cell activation, heightened glycolysis, PPP, and glutaminolysis
intersect with the de novo synthesis of GSH through promoting
cysteine uptake and providing glycine, glutamine, and NADPH
(93–95). As such, the genetic abrogation of de novo synthesis of Frontiers in Immunology | www.frontiersin.org GLUTAMINE CATABOLISM IN
COORDINATING THE PRODUCTION
OF ROS AND GSH Glutamine has been known as a key nutrient, which supports
a diverse range of cellular functions (93–102). Glutamine pro-
vides high proportions of the energy from OXPHOS, provides
precursors for various biosynthetic pathways, as a key nitrogen
and carbon donor, and also is catabolized to various intermedi-
ate metabolites that have signaling roles in modulating cellular
processes. In specialized cells, such as the cells of the nervous
system, glutamine catabolism intersects with signaling networks
to support the production of central neurotransmitters including
glutamate, GABA, and aspartate (103–106). To meet bioenergetic
and biosynthetic demand during T cell growth and proliferation,
glutaminolysis replenishes the anapleurotic substrate α-KG that
fuels OXPHOS via the TCA cycle and also provides sources
of nitrogen and carbon to support the biosynthesis of nones-
sential amino acids, lipids, nucleotides, and polyamines (13, 15,
102, 107). Similar to cancer cells, de novo synthesis of GSH in
T cells, which relies on glutamine to provide precursors, plays May 2018 | Volume 9 | Article 1075 Frontiers in Immunology | www.frontiersin.org 4 Metabolism in T cell redox balance Rashida Gnanaprakasam et al. Figure 3 | Cellular redox homeostasis is essential for mounting an effective T cell-mediated immune response. In addition to generate ATP and provide
biosynthetic precursors, T cell activation-induced metabolic reprogramming actively regulates redox homeostasis. The coordination of de novo synthesis of
glutathione (GSH) and the production of reactive oxygen species (ROS) ensures T cell redox balance and a fine-tuned T cell response. Figure 3 | Cellular redox homeostasis is essential for mounting an effective T cell-mediated immune response. In addition to generate ATP and provide
biosynthetic precursors, T cell activation-induced metabolic reprogramming actively regulates redox homeostasis. The coordination of de novo synthesis of
glutathione (GSH) and the production of reactive oxygen species (ROS) ensures T cell redox balance and a fine-tuned T cell response. synthesis and the metabolic flux shunted toward OXPHOS. Consistent with this idea, the overall high consumption rate of
glutamine in proliferative cells is suggested to provide a sensitive
and precise regulation on intermediate metabolites that can be
committed toward several metabolic branches, hence permitting
rapid responses to meet the demands for energy production or
antioxidant production (99, 108). In addition to increasing anti-
oxidant capacity, T cells may adapt by shifting glucose catabolism
from OXPHOS toward aerobic glycolysis, which could provide
biosynthetic precursors and rapidly produce ATP by the substrate
level of phosphorylation. GLUTAMINE CATABOLISM IN
COORDINATING THE PRODUCTION
OF ROS AND GSH an essential role in suppressing oxidative stress. Accordingly,
glutaminolysis is a branched pathway that consists of several
paths, enabling energy production through oxidation and
biomolecule production, including GSH through biosynthesis
(93–95). While the ATP generating capacity of glutaminolysis
is considered to be redundant with glucose oxidation and/or
FAO, the oxidation of glutamine is indispensable for driving
T cell proliferation and differentiation (13, 15, 102). However,
enhanced glutamine oxidation in the mitochondria also increases
the production of its by-product, mitochondrial ROS, the main
source of cellular ROS in T cells (35, 37). Therefore, glutamate
represents a key branch point in glutaminolysis that can be
committed toward mitochondrial oxidation to produce ATP
and ROS, or toward de novo synthesis of GSH to modulate redox
balance and suppress oxidative stress. In addition, the high rate
of glutaminolysis ensures that the capacity to supply glutamate,
the most abundant intracellular metabolite in cells, exceeds the
demand for glutamate from each of the downstream metabolic
branches. The branched pathways in glutaminolysis enable the
production of counteracting metabolites, i.e., ROS and GSH,
from a common metabolic precursor, and permit a fine-tuned
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biomolecules and cellular organelles. Under pathophysiological
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mune diseases. However, oxidative stress caused by elevated ROS May 2018 | Volume 9 | Article 1075 Frontiers in Immunology | www.frontiersin.org 5 Rashida Gnanaprakasam et al. Metabolism in T cell redox balance FUNDING This work was supported by R21AI117547 and 1R01AI114581
from National Institutes of Health, V2014-001 from the
V-Foundation and 128436-RSG-15-180-01-LIB from the
American Cancer Society (RW). AUTHOR CONTRIBUTIONS may also render key immune effector cells vulnerable to agents
that can either modulate stress response or modulate metabolic
pathways for ROS and GSH production (Figure 3). Redox sign-
aling is essential to regulate T cell metabolism. Technological
advancement in genetic models and metabolomics will allow us
to understand the key metabolic processes that dictate T cell fate
through modulation ROS and GSH production. Thus, further
research is expected to illustrate the complex interplay between
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(CC BY). REFERENCES The use, distribution or reproduction in other forums is permitted, provided
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INTRODUCTION interval that lasted up to a few days (Brashers-Krug et al., 1996;
Muellbacher et al., 2002; Walker et al., 2003a). In those studies,
using A-B-A paradigms, the first learning of task A and task B
was conducted within the same day, and task A was subsequently
retested on the next day. The interval between task A and task B
on the first day was manipulated to examine the temporal char-
acteristics of consolidation and retrograde interference. When
task A and task B were temporally apart for more than 4–6 h,
task B did not disrupt the learning of task A, hence task A was
considered as consolidated (Brashers-Krug et al., 1996; Walker
et al., 2003a). Nevertheless, immunity to interference over such a
short time period may not necessarily indicate immunity to other
forms of interference. In fact, several studies have shown that
memory might never become permanently immune to disruption
(Misanin et al., 1968; Nadel and Land, 2000; Nader et al., 2000). Using fear conditioning procedures and electroconvulsive shock
to hippocampus cells in mice, Nader et al. (2000) demonstrated
that consolidated memories can be reactivated by a conditioned
stimulus and become labile to disruption again. The observation
that reactivation can render memories labile to modifications is
termed reconsolidation (Przybyslawski and Sara, 1997). Newly learned information is often transient and thus, inaccessi-
ble during later recollection attempts. Thus, learned information
can undergo interference by other information or weakened with
the passage of time. The process by which information is stabi-
lized and becomes immune to certain types of interfering agents
is termed consolidation (Dudai, 2004; Eichenbaum, 2008). Understanding the functional and neural mechanisms that
mediate consolidation is a fundamental issue in neuroscience
(Dayan and Cohen, 2011; Censor et al., 2012). Newly learned
memories are often unstable, and thus, tend to be disrupted by
subsequently learned information. This disruption is called ret-
rograde interference. Retrograde interference can occur in several
types of learning, including declarative memory (Alvarez and
Squire, 1994), motor learning (Brashers-Krug et al., 1996; Cohen
and Robertson, 2011), and perceptual learning (Seitz et al., 2005;
Yotsumoto et al., 2009a; Hung and Seitz, 2011). Retrograde inter-
ference has attracted considerable attention, as it directly relates
to consolidation. COMPUTATIONAL NEUROSCIENCE ORIGINAL RESEARCH ARTICLE
published: 28 May 2013 published: 28 May 2013
doi: 10.3389/fncom.2013.00069 Frontiers in Computational Neuroscience Consolidated learning can be susceptible to
gradually-developing interference in prolonged
motor learning Yuko Yotsumoto1, Takeo Watanabe 2, Li-Hung Chang 2 and Yuka Sasaki 2*
1 Department of Life Sciences, The University of Tokyo, Tokyo, Japan
2 Department of Cognitive, Linguistic, and Psychological Sciences, Brown University, Providence, RI, USA When multiple items are learned in sequential order, learning for one item tends to
be disrupted by subsequently learned items. Such retrograde interference has been
studied with paradigms conducted over a relatively short term. Resistance to interference
is generally believed to be a measure of learning or consolidation. Here, we used a
finger-tapping motor sequence paradigm to examine interference in prolonged motor
learning. Three groups of nine subjects participated in training sessions for 16 days, and
practiced three different sequences in different orders and combinations. We found that a
well-trained motor sequence was subject to a gradual interference when the subsequent
learning was paired in a particular order. The results suggest that a well-learned motor
memory is still susceptible to interference, and that resistance to interference in one
condition does not necessarily imply full, permanent consolidation. Reviewed by: Reviewed by:
Avi Mendelsohn, Weizmann
Institute of Science, Israel
Nitzan Censor, NIH, USA *Correspondence:
Yuka Sasaki, Department of
Cognitive, Linguistic, and
Psychological Sciences, Brown
University, PO Box 1821, 190 Thayer
Street, Providence, RI 02912, USA. e-mail: yuka_sasaki@brown.edu Keywords: motor learning, interference, retrograde interference, finger tapping, consolidation Keywords: motor learning, interference, retrograde interference, finger tapping, consolidation Edited by: Edited by:
Lior Shmuelof, Columbia University,
USA Edited by:
Lior Shmuelof, Columbia University,
USA Lior Shmuelof, Columbia University,
USA Reviewed by:
Avi Mendelsohn, Weizmann
Institute of Science, Israel
Nitzan Censor, NIH, USA
*Correspondence:
Yuka Sasaki, Department of
Cognitive, Linguistic, and
Psychological Sciences, Brown
University, PO Box 1821, 190 Thayer
Street, Providence, RI 02912, USA. e-mail: yuka_sasaki@brown.edu Reviewed by:
Avi Mendelsohn, Weizmann
Institute of Science, Israel
Nitzan Censor, NIH, USA
*Correspondence:
Yuka Sasaki, Department of
Cognitive, Linguistic, and
Psychological Sciences, Brown
University, PO Box 1821, 190 Thayer
Street, Providence, RI 02912, USA. e-mail: yuka_sasaki@brown.edu Keywords: motor learning, interference, retrograde interference, finger tapping, consolidation METHODS
SUBJECTS sequence C. The training sequences performed on Days 9–16 were
identical for all groups, which included sequence B for the first 12
blocks followed by sequence A for the second 12 blocks. A total of 27 right-handed subjects (13 females, 14 males) par-
ticipated in the study. Subject ages ranged from 18 to 34 years
(mean, 22.0 years; standard deviation, 3.8 years). None of the
subjects played a musical instrument. All subjects gave informed
written consent. The Institutional Review Board of Massachusetts
General Hospital approved the study. Subjects were instructed to tap four keys on a keypad
(Nostromo SpeedPad, BELKIN) according to the instructed
sequence, using the fingers of their non-dominant hand. Each
of the four keys was assigned a number between 1 and 4, with
1 corresponding to the little finger, 2 to the ring finger, 3 to the
middle finger, and 4 to the index finger. Subjects were instructed
to tap the keys as quickly and accurately as possible for 30 s. A
sequence was five key presses arranged in one of the three follow-
ing orders: sequence A [4-3-1-2-4], sequence B [4-2-3-1-4], and
sequence C [4-1-3-2-4]. The assignment was counter balanced
across subjects; therefore, sequence A could require different
tapping sequences between subjects. PROCEDURE Subjects trained to perform a finger-tapping task (Karni et al.,
1995, 1998) in 16 training sessions. Subjects participated in at
least three training sessions per week, which took place on sep-
arate days with a 1- to 2-day interval between sessions. Thus, the
total training period spanned 18–26 days (mean = 22.63 ± 2.43
days). Subjects trained with a single sequence on Days 1–8, and
then trained with two sequences on Days 9–16. For Days 1–8,
training sessions were ∼12 min in duration, and consisted of
twelve 30-s blocks, with 30-s resting periods between blocks. For
Days 9–16, training sessions were ∼24 min in duration, and con-
sisted of twelve 30-s blocks for each sequence (24 blocks in total),
with 30-s resting periods between blocks. All experimental sessions were conducted in a laboratory. The
instructed tapping sequence was displayed on the center of a com-
puter screen throughout the task to eliminate the requirement for
working memory. On Days 9–16, when two sequences were per-
formed, subjects were reminded twice about the sequence change
before the training session and immediately before the second
sequence began. Both the keypad and the tapping hand were
hidden from the subject’s view during the task. Subjects were divided randomly into three groups: Group
ABA, Group CBA, and Group BBA. Figure 1 presents a schematic
representation of the training sessions for the three groups. During Days 1–8, each group performed a different train-
ing sequence. Group ABA trained with sequence A, Group
BBA trained with sequence B, and Group CBA trained with INTRODUCTION In cases where retrograde interference manipu-
lations do not disrupt the performance on acquired memories,
it can be assumed that memory consolidation has taken place
(Brashers-Krug et al., 1996; Muellbacher et al., 2002; Walker et al.,
2003a). Retrograde interference is typically evaluated with an
A-B-A paradigm (Walker et al., 2003a; Krakauer et al., 2005). In
this paradigm, task A is learned before task B, and repeated after
learning task B. If task A performance is disrupted after learning
task B, then task B is considered to have caused retrograde inter-
ference. If learning task B does not disrupt task A performance,
then task A is considered to be “consolidated.” Here, we show that human motor memories can also expe-
rience interference after it was considered consolidated. We
observed that a well-trained motor skill acquired with long-term
training was resistant to interference in some conditions, but was
subject to strong interference when an interfering motor task
was introduced together with the well-trained motor task, and
the interfering task preceded the well-trained task. Our results
indicate that motor memory might also follow the rules of consol-
idation and reconsolidation when memory reactivation is applied
after prolonged periods of motor learning. Previous motor consolidation studies largely focused on inter-
ference generated between motor skills acquired over a short time May 2013 | Volume 7 | Article 69 | 1 www.frontiersin.org www.frontiersin.org www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. GROUP CBA Group CBA first practiced sequence C, followed by sequence B,
and then sequence A. For Group CBA, the number of correctly
tapped sequences for sequence C increased gradually over train-
ing and plateaued by Day 8. During Days 9–16, the number
of correctly tapped sequences also increased gradually, both for
sequences B and A (Figure 2B). Although, sequence B was the
second sequence for all groups, Group CBA did not show signs
of interference across sequences. A Two-Way ANOVA (day vs. sequence) with repeated measures performed with Group CBA
data revealed a significant main effect for day [F(7, 56) = 68.657,
p < 0.001]. The sequence [F(2, 16) = 2.800, NS] and the interac-
tion between day and sequence, were not significant [F(14, 112) =
1.342, NS]. GROUP BBA Group BBA first practiced sequence B, repeated sequence B,
and then performed sequence A. The sequence presented in the
second part of training was identical to that for Groups ABA
and CBA. If the well-trained sequence was subject to interfer-
ence, then sequence B performance during the second training
period should worsen over several days. Our results, however,
revealed sequence B performance improvement was maintained
throughout the second training period (Figure 2C). In addition,
sequence A performance improved across days. That is, no inter-
ference was observed between sequences A and B. A Two-Way
ANOVA (day vs. sequence) with repeated measures showed sig-
nificant main effects for day [F(7, 56) = 17.838, p < 0.001] and
sequence [F(2, 16) = 16.578, p < 0.001], and a significant inter-
action between day and sequence [F(14, 112) = 8.249, p < 0.001]. The significant main effect for the sequence and the significant
interaction between day and sequence reflect that the sequence B
performance in the second period sustained while the sequence A
performance improved. FIGURE 2 | Performance during a finger-tapping motor sequence task
for three training sequences (three groups). The mean number of
correctly typed responses as a function of the training day. In the first part
of the training session (Days 1–8), a single sequence was trained; two
sequences (sequences B and A) were practiced in all groups in the second
training period (Days 9–16). (A) Group ABA; (B) Group CBA; (C) Group BBA. FIGURE 2 | Performance during a finger-tapping motor sequence task
for three training sequences (three groups). The mean number of
correctly typed responses as a function of the training day. In the first part
of the training session (Days 1–8), a single sequence was trained; two
sequences (sequences B and A) were practiced in all groups in the second
training period (Days 9–16). (A) Group ABA; (B) Group CBA; (C) Group BBA. correctly typed responses as a function of the training day. In the first part
of the training session (Days 1–8), a single sequence was trained; two
sequences (sequences B and A) were practiced in all groups in the second
training period (Days 9–16). (A) Group ABA; (B) Group CBA; (C) Group BBA. RESULTS
GROUP ABA Group ABA first practiced sequence A, then sequence B, followed
by sequence A. The number of correctly tapped sequences for
sequence A increased gradually during training and plateaued
by Day 8 (Figure 2A). Notably, sequence B performance also
improved gradually at a rate similar to that observed for sequence
A. In contrast, performance in the second training instance for
sequence A was diminished when compared to the first sequence
A training session. That is, we found interference for sequence A
after sequence B was learned. Sequence A performance improved
again once sequence B performance plateaued (see Days 14–16 in
Figure 2A). We conducted a Two-Way ANOVA (day vs. sequence)
with repeated measures to compare the two instances of sequence
A performance, and between sequence A and sequence B per-
formance. This revealed significant effects for “day” [F(7, 56) =
7.654, p < 0.001], and a significant interaction between “day”
and “sequence” [F(14, 112) = 5.112, p < 0.001]. No significant
effect was observed for sequence [F(2, 16) = 1.470, NS]. These
findings support our hypothesis that learning sequence B intro-
duces interference on the previously learned sequence A. DATA ANALYSIS The number of correctly tapped sequences was averaged across 12
blocks for each subject. The resulting mean was used as an index
of motor learning. FIGURE 1 | Schematic representation of the experimental procedure. FIGURE 1 | Schematic representation of the experimental procedure. Frontiers in Computational Neuroscience May 2013 | Volume 7 | Article 69 | 2 www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. FIGURE 2 | Performance during a finger-tapping motor sequence task
for three training sequences (three groups). The mean number of
correctly typed responses as a function of the training day. In the first part
of the training session (Days 1–8), a single sequence was trained; two
sequences (sequences B and A) were practiced in all groups in the second
training period (Days 9–16). (A) Group ABA; (B) Group CBA; (C) Group BBA. Frontiers in Computational Neuroscience ACROSS-SESSION LEARNING vs. WITHIN-SESSION LEARNING To further, examine the nature of within-session learning, we
calculated the difference in the number of correctly tapped
sequences between the first and last blocks of each session. These
values are presented in Figure 5. These values are an index of
within-session learning, with positive values indicating within-
session learning, and negative values indicating within-session
deterioration. RELATIVE PERFORMANCE IMPROVEMENT Among the three experimental groups, performance decline was
observed only in Group ABA. To assure that such decline did
not result from differences in motor learning skills, we compared
the relative performance improvements during the first 8 train-
ing days. Had the improvement in the first 8 days differed across
groups, the differences observed in the second training period
could have been attributed to other factors, such as individual
differences in motor skills. May 2013 | Volume 7 | Article 69 | 3 Frontiers in Computational Neuroscience www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. one-sample t-tests, Bonferroni correction]. After, the second
training session, subjects showed no within-session learning until
a new tapping sequence was introduced on Day 9. In Group BBA,
subjects showed a negative value for the within-session learn-
ing on Day 4, which might be explained by fatigue [t(8) = −4.9,
p < 0.05, Bonferroni correction]. The relative improvement was computed by subtracting the
number of correctly tapped responses on Day 1 from those
observed on Days 2 through 8. Figure 3 shows the averaged rel-
ative performance improvement for all three groups in the first
part of the training. A Two-Way ANOVA with repeated measures,
with group and days as factors, indicated no significant effect
for group [F(2, 24) = 1.048, NS] or interaction between day and
group [F(12, 144) = 0.954, NS], though, a significant main effect
was observed for day [F(6, 144) = 21.783, p < 0.0001]. This mea-
sure indicates that overall learning performance for all groups was
comparable. The across-session learning (Figure 2) was calculated by aver-
aging the numbers of correctly tapped sequences across the 12
blocks. In contrast, within-session learning (Figure 5) was cal-
culated from the difference between the first and last blocks of
a session. These two types of learning showed distinct pattern
changes as the training progressed. While the average number of
correctly tapped sequences across 12 blocks continually increased
from the first to the eighth session, within-session learning was
evident only in the first training session. This difference implies
there are two types of motor learning, one characterized by grad-
ual training increases across days, and one characterized by quick
changes within a session that disappears in relatively early training
phases. WITHIN-SESSION LEARNING The number of correctly tapped sequences shown in the previ-
ous figures were averaged across 12 blocks within a session. In the
present motor task, one session consisted of twelve 30-s blocks. It is well-known that motor learning performance improves not
only across sessions but also within a session, especially during the
initial learning session (Karni et al., 1998). To examine within-
session learning, we counted the number of correctly tapped
sequences for each block and for each session, and plotted the
counts in Figure 4. For all three groups, the newly introduced
sequences exhibited within-session improvement during the ini-
tial training session (Day 1 and 9). For other training sessions
(days), the task performance was consistent across the 12 blocks. For the second half of the learning phase, Group ABA
showed interference in across-session learning, and the pat-
tern of within-session learning for sequence B differed from
that for sequence A. A Two-Way ANOVA (day vs. sequence)
with repeated measures confirmed a significant interaction
between days and the trained sequence [F(7, 56) = 2.56, p =
0.023] during the same training period. For Group ABA,
within-session learning for sequence A during Days 9–16 was
insignificant (p > 0.25 for all sessions), indicating no signs of
interference. The difference between the across-session and the
within-session learning suggests that interference from learn-
ing other sequences primarily affected long-term across-session
learning. DISCUSSION Our present findings indicate that performance on a well-trained
motor skill is subject to interference when learning of a new
sequence is inserted before performing the well-trained skill. Such
interference was not observed if the order of learning is reversed
for the new and the well-trained skills. Can diminished performance of sequence A in the second part
of training for Group ABA be attributed to a factor other than
interference from sequence B? First, it is unlikely that diminished
performance in sequence A was attributed to a limited capacity of
motor memories, since improved performance on both sequences
A and B was observed in the second part of training in Groups
CBA and BBA. Second, it is unlikely that poorer performance on
sequence A for Group ABA was due simply to switching from one
sequence to another (Imamizu et al., 2007). If there was a cost for
switching sequences within a session for Group ABA, we should
have observed decline in the earlier blocks of the interfered ses-
sions. However, the block-wise analysis shown in Figure 4, and
the within-session improvement shown in Figure 5, both indicate
performance was consistently interfered within a session. Third, it
is unlikely that extended practice of the same sequence decreased
subject motivation for typing the correct responses, which would
have led to poorer performance on the sequence performed last. We believe this because improved performance on sequence B
did not disappear in Group BBA in the second part of training. If prolonged practice bored the subjects, then performance on
sequence B would have been lower in the second part of training
in Group BBA. Fourth, it could be argued that subjects in Group
ABA showed diminished performance because of poor learning. This, however, is unlikely. While, average performance on Day 1
was slightly different across the three groups (Group CBA was
the lowest in initial performance), average improvement over the
8 daily sessions in the first part of training was almost identical
across groups (see the slope for Days 1–8 for the three groups in
Figure 2 and also Figure 3). Finally, one may argue that retrieval
of presumably consolidated information alone makes the mem-
ory once again susceptible for interference, as was shown in a
previous study (Walker et al., 2003a). Nevertheless, this may not
be the case. CORRELATIONS BETWEEN INTERFERENCE AND INDIVIDUAL
PERFORMANCE For Days 1–8, within-session learning was evident only in
the first or second sessions. For all three groups, the first train-
ing session significantly differed from zero [ABA: t(8) = 4.09,
p < 0.05; CBA: t(8) = 8.81, p < 0.05; BBA: t(8) = 5.49, p = 0.05; In the previous analyses, we describe interference by pooling the
number of correctly tapped sequences across subjects. One might
wonder if any individual differences might affect such interference
effects. For example, an individual who performs well in their
first training session might have some resistance to interference,
thus, causing less interference in the second part of the training. To examine the effects of individual performance on interference,
we calculated the performance in the first training session, and
the extent of interference for all subjects in Group ABA. We then
examined correlations between these measures. FIGURE 3 | Relative performance improvement compared to the initial
day performance by group. Performance in the first training session was quantified by
the number of correctly tapped sequences (A). To determine the
extent of interference, we first selected for each subject the ses-
sion in which the largest number of correct sequence A trials were
performed during Days 1–8. In addition, we also selected for each
subject the session with the fewest completed correct sequence
A trials during Days 9–16. Assuming the session with the worst
performance was the session with largest interference, we sub-
tracted the worst performance during Days 9–16 from the best
performance during Days 1–8. This value indicates how much
deterioration was caused in sequence A tapping by introducing
sequence B. FIGURE 3 | Relative performance improvement compared to the initial
day performance by group. May 2013 | Volume 7 | Article 69 | 4 Frontiers in Computational Neuroscience www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. eraged number of correctly tapped sequences (±s.e.m.) plotted for each block and for each session. For each group, the mean numbers
ped sequences are plotted for each learned sequence. mputational Neuroscience
www.frontiersin.org
May 2013 | Volume 7 | Article 69 | 5 FIGURE 4 | Averaged number of correctly tapped sequences (±s.e.m.) plotted for each block and for each session. For each group, the mean numbers
of correctly tapped sequences are plotted for each learned sequence. CORRELATIONS BETWEEN INTERFERENCE AND INDIVIDUAL
PERFORMANCE Frontiers in Computational Neuroscience May 2013 | Volume 7 | Article 69 | 5 www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. FIGURE 5 | Within-session learning as indicated by an increase in the
number of correctly tapped sequences within a session (±s.e.m.). Asterisks indicate sessions with significant deviation from zero (p < 0.05,
Bonferroni correction). selected for each subject the session with the best performance,
and calculated the difference between the best performance and
their performance on Day 1. This value indicates the extent of
improvement made in the first part of the training. Correlations
between the amount of learning and the amount of interference
are presented in Figure 6B. As the figure indicates, a weak trend
was observed in that subjects who showed larger improvements
during Days 1–8 also showed larger interference, though, this was
insignificant (r = 0.513, p = 0.15). Frontiers in Computational Neuroscience DISCUSSION If the retrieval of memory alone made the mem-
ory fragile, then the performance of sequence B would have been
worsened in the second part of the training for Group BBA. FIGURE 5 | Within-session learning as indicated by an increase in the
number of correctly tapped sequences within a session (±s.e.m.). Asterisks indicate sessions with significant deviation from zero (p < 0.05,
Bonferroni correction). Correlations between initial performance (Day 1) and the
extent of interference (best performance in the first part—worst
performance in the second part) are presented in Figure 6A. We
observed a trend indicating that better performance on Day 1
predicted less interference (r = −0.61, p = 0.079). Although, the
results were not significant, they implied that the initial perfor-
mance level may contribute to the occurrence of interference. While, no interference was observed for learning sequence A
or B in Group CBA, results from previous studies (Brashers-
Krug et al., 1996; Muellbacher et al., 2002; Walker et al., 2003a)
indicate that retrograde and/or anterograde interference between
sequences A and B was more likely when the training of the two
new sequences was temporally close (within a few days). Those While, we did not find significant effects between initial per-
formance and interference, the extent of learning might relate to
the extent of interference. To examine this possibility, we calcu-
lated correlations between the performance improvement during
Days 1–8, and interference in Group ABA. For the Days 1–8, we May 2013 | Volume 7 | Article 69 | 6 Frontiers in Computational Neuroscience While the motor memory
has been reported to become resistant to retrograde interference
within a day (Brashers-Krug et al., 1996; Walker et al., 2003a), the
present study indicated that the neural state of the motor memory
could become vulnerable again to the retrograde interference after
the repeated learning. Such vulnerability to the interference may
lead to reactivation and reconsolidation of the motor memory. It
is implied that the interference in prolonged learning may stem
from the later stage of neural consolidation, whereas, the interfer-
ence in short interval motor learning may stem from the earlier
stage of neural consolidation. Yet, it is unclear why the order
of the two sequences learned in the second part of training was
critical for interference, and how the prolonged motor learning
is neurally consolidated. Future studies are needed to clarify the
underlying neural correlates of full consolidation in motor skill
learning. Neural representation of consolidated motor skill is different
from that of acquisition (Brashers-Krug et al., 1996; Shadmehr
and Holcomb, 1997; Muellbacher et al., 2002; Dayan and Cohen,
2011). In addition, different neural systems for consolidation are
proposed for two types of motor skill learning. These two sys-
tems are dynamic motor adaptation learning and motor sequence
learning (Doyon et al., 2003; Doyon and Benali, 2005). For both
dynamic motor adaptation learning and motor sequence learn-
ing, the condition under which retrograde interference occurs
has been identified as successive acquisition of two motor skills
presented within a short temporal interval between the first and
second skill (Muellbacher et al., 2002; Doyon and Benali, 2005;
Krakauer et al., 2005; Krakauer and Shadmehr, 2006). The present
study found that interference occurred for retrieval of well-
trained motor sequence learning: In Group ABA, sequence A was
presumably reinstated by the learning of sequence B, thus, ren-
dering it labile to interference. Because the learning of sequence
B interfered with the previously learned sequence A, the interfer-
ence should be construed as retrograde. Whether our findings can
be applied to dynamic motor learning, or whether they apply to
motor sequence learning, remains open to investigation. Although, the neural mechanisms for prolonged motor learn-
ing have yet to be revealed, prolonged visual perceptual learning
has been reported to induce transient changes in neural activ-
ity. Using a texture discrimination task (Karni and Sagi, 1991),
Yotsumoto et al. Frontiers in Computational Neuroscience Frontiers in Computational Neuroscience www.frontiersin.org www.frontiersin.org www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. FIGURE 6 | (A) Correlations between the initial performance on Day 1 and the extent of interference. (B) Correlations between the performance improvement
during the first part of the training, and the extent of interference. FIGURE 6 | (A) Correlations between the initial performance on Day 1 and the extent of interference. (B) Correlations between the performance improvement
during the first part of the training, and the extent of interference. studies, however, did not include a condition that consisted of
learning a different sequence as employed in the current study
in Group CBA. Perhaps enhanced general ability (Krakauer et al.,
2005) in motor skill learning that resulted from first learning C
in the training for Group CBA made an efficient neural represen-
tation (Yotsumoto et al., 2008) for a new motor skill, leading to
the successful acquisition of two new (sequences B and A) motor
skills. visual cortex (V1) exhibited a different pattern: the task-relevant
BOLD signal in V1 increased for about 10 days, then returned
to its baseline toward the end of the training period. This indi-
cates that even when behavioral performance seems similarly
sustained, the underlying neural mechanisms might change as
training progresses. Although, visual perceptual learning in a tex-
ture discrimination task involves different cortical areas from
motor learning with finger tapping paradigm, the tasks are anal-
ogous in terms of consolidation and interference. The learned
feature becomes consolidated during sleep after the initial learn-
ing (Mednick et al., 2002; Walker et al., 2002, 2003b; Yotsumoto
et al., 2009b), and becomes resistant to the retrograde interfer-
ence (Karni and Sagi, 1991; Walker et al., 2003a; Yotsumoto et al.,
2009a). In the present study, 16 training sessions were conducted
in 18–26 days. It is reasonable to hypothesize that prolonged
motor learning in the present study induced transient changes,
making neural states differ between the earlier phase and the later
phase of the learning. In the present study, the repeated learning
sessions during the first part of training could have enhanced neu-
ral plasticity and modified the neural state from that of the motor
learning tested with short intervals. The interference observed in
our prolonged learning could be attributed to the neural state
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Yotsumoto,
Watanabe, Chang and Sasaki. Frontiers in Computational Neuroscience (2008) examined dynamics of brain activation
in visual cortex across 14 training sessions conducted on differ-
ent days across 3–4 weeks. While, the behavioral performance
gradually increased for about a week and reached a plateau for
rest of the training period, the BOLD signal changes in primary May 2013 | Volume 7 | Article 69 | 7 www.frontiersin.org www.frontiersin.org www.frontiersin.org Interference in prolonged motor learning Yotsumoto et al. ACKNOWLEDGMENTS To conclude, a well-learned motor memory can be suscepti-
ble to retrograde interference, and resistance to interference in
the earlier stages of learning does not necessarily indicate full,
permanent consolidation for that skill. This work was supported by the NIH grants (R01EY015980 and
R01MH091801), the National Science Foundation, the Human
Frontier Foundation, Grants-in-Aid for Scientific Research
(KAKENHI-23680028, 24330208), and the ERATO Shimojo
Implicit Brain Function Project. This study was conducted
under a protocol approved by the Institutional Review Board of
Massachusetts General Hospital, where the authors were affiliated
at the time of conducting the experiments. We thank Shih-Chi
Hsu for her help in conducting the experiments. AUTHOR CONTRIBUTIONS Yuko Yotsumoto, Takeo Watanabe, and Yuka Sasaki designed the
experiment. Yuko Yotsumoto and Li-Hung Chang collected data. Yuko Yotsumoto and Yuka Sasaki analyzed data. Yuko Yotsumoto,
Takeo Watanabe, and Yuka Sasaki wrote the paper. REFERENCES This is
an open-access article distributed under
the terms of the Creative Commons
Attribution License, which permits use,
distribution and reproduction in other
forums, provided the original authors
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English
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Utilization of a Wheat660K SNP array-derived high-density genetic map for high-resolution mapping of a major QTL for kernel number
|
Scientific reports
| 2,017
|
cc-by
| 10,670
|
Utilization of a Wheat660K SNP
array-derived high-density genetic
map for high-resolution mapping of
a major QTL for kernel number Received: 1 February 2017
Accepted: 5 May 2017
Published: xx xx xxxx Received: 1 February 2017
Accepted: 5 May 2017
Published: xx xx xxxx Fa Cui1,2,7, Na Zhang1,6, Xiao-li Fan1,3, Wei Zhang1,7, Chun-hua Zhao2, Li-juan Yang4, Rui-qing
Pan1,6, Mei Chen1,6, Jie Han1,6, Xue-qiang Zhao7, Jun Ji1,7, Yi-ping Tong7, Hong-xia Zhang2,
Ji-zeng Jia5, Guang-yao Zhao5 & Jun-ming Li1,7 In crop plants, a high-density genetic linkage map is essential for both genetic and genomic researches. The complexity and the large size of wheat genome have hampered the acquisition of a high-resolution
genetic map. In this study, we report a high-density genetic map based on an individual mapping
population using the Affymetrix Wheat660K single-nucleotide polymorphism (SNP) array as a probe
in hexaploid wheat. The resultant genetic map consisted of 119 566 loci spanning 4424.4 cM, and 119
001 of those loci were SNP markers. This genetic map showed good collinearity with the 90 K and 820 K
consensus genetic maps and was also in accordance with the recently released wheat whole genome
assembly. The high-density wheat genetic map will provide a major resource for future genetic and
genomic research in wheat. Moreover, a comparative genomics analysis among gramineous plant
genomes was conducted based on the high-density wheat genetic map, providing an overview of the
structural relationships among theses gramineous plant genomes. A major stable quantitative trait
locus (QTL) for kernel number per spike was characterized, providing a solid foundation for the future
high-resolution mapping and map-based cloning of the targeted QTL. High-density genetic linkage maps are essential for genetic and genomic research in crops1–4. Molecular breed-
ing is more effective if the molecular map is dense to provide more choices in the quality and type of mark-
ers and to increase the probability of detecting polymorphic markers in important chromosomal intervals. In
wheat, the large genome size (17 gigabases), hexaploid nature (AABBDD), high percentage of repetitive regions
and low level of polymorphism have complicated the acquisition of high-resolution genetic maps by molecular
markers1–4. To date, several kinds of molecular markers, including restriction fragment length polymorphism
(RFLP)5, 6, amplified fragment length polymorphism (AFLP)7, simple sequence repeats (SSRs)8, 9, and diversity
array technology (DArT)3, 4, 10, 11 have been used to construct genetic linkage maps in wheat. Information regard-
ing wheat molecular markers and genetic maps is available in some datasets such as GrainGenes 2.0 (https://
wheat.pw.usda.gov/GG3/), URGI (https://urgi.versailles.inra.fr/), etc. www.nature.com/scientificreports www.nature.com/scientificreports www.nature.com/scientificreports Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 Received: 1 February 2017
Accepted: 5 May 2017
Published: xx xx xxxx Utilization of a Wheat660K SNP
array-derived high-density genetic
map for high-resolution mapping of
a major QTL for kernel number Cavanagh et al.12 released a hexaploid wheat consensus genetic map with 7504 SNP markers from the Wheat9K
SNP array using a combination of seven mapping populations. Wang et al.16 mapped 46977 SNPs from the
Wheat90K array to the hexaploid wheat genetic map using a combination of eight mapping populations. Using
both Wheat9K and Wheat90K arrays, Maccaferri et al.17 released a high-density tetraploid wheat consensus
genetic map with 30144 markers (including 26626 SNPs and 791 SSRs) by integrating 13 data sets from inde-
pendent biparental mapping populations. Recently, Winfield et al.18 documented a hexaploid wheat consensus
map with 56 505 SNP markers from the Wheat820K array, spanning 3739 cM in length, using three independent
biparental populations. However, although a high-density hexaploid wheat genetic map (>100 000 markers)
based on an individual biparental mapping population would be valuable for further genetic research, such as
high-resolution mapping and map-based cloning of a targeted major quantitative trait locus (QTL), no such map
has been released. A new Whole Genome Shotgun (WGS) assembly of the Chinese Spring (CS) reference wheat genome is now
available (http://plants.ensembl.org/index.html; https://urgi.versailles.inra.fr/download/iwgsc/IWGSC-WGA_
Sequences/). However, genetic and genomic studies in wheat continue to lag behind the research in other mem-
bers of the grass family (Gramineae), such as rice and maize. The gradual enrichment of SNP markers and the
sequences released for CS (https://urgi.versailles.inra.fr/download/iwgsc/IWGSC-WGA_Sequences/), Triticum
urartu2 and Aegilops tauschii1 have facilitated comparative genomic analysis in wheat. Comparative genomic
analysis with species whose genomes have been well characterized has been used as an effective method for the
construction of high-resolution genetic linkage maps of target wheat genes and for the prediction of candidate
genes in regions of interest. For instance, the construction of high-density genetic maps has facilitated the map-
ping of the gene grain protein content-B1 (Gpc-B1)19, the yellow rust resistance gene Yr3620 and the powdery
mildew resistance gene Pm4121. Comparative genomics studies have also furthered the understanding of the
basic processes of genome evolution and the transfer of information from model species to related organisms and
facilitated the cross-referencing of various types of information, such as QTLs, mutants, and gene expression22, 23. These correlations and integrations will take full advantage of the collective intellectual contributions from scien-
tists across many disciplines22.f y
p
Wheat660K, the Affymetrix® Axiom® Wheat660, was designed by the Chinese Academy of Agricultural
Sciences and synthesized by Affymetrix. Results
SNP SNP scores in the 265 accessions. An F8:9 recombinant inbred line (RIL) population including 188 lines
derived from a cross between Kenong 9204 (KN9204) and Jing 411 (J411) (denoted as KJ-RILs), 65 KN9204-
derived advanced lines/authorized varieties, three parental lines of KN9204, three control varieties from the
Winter Wheat Performance Trial of the Northern Huang-Huai Regional Nursery of China, and Chinese Spring
(CS) were genotyped using the 630517 SNPs on the Wheat660K SNP array as probes. The sample call rates ranged
from 18.6% to 100.0%, with an average of 98.9% for the 265 accessions (data not shown). The scores for the
probes were classified into one of the following six categories according to the cluster patterns produced by the
Affymetrix software (Table S1): (i) Poly High Resolution (PHR) (188040; 29.8%); (ii) No Minor Homozygote
(NMH) (133246; 21.1%); (iii) Off-Target Variant (OTV) (18471; 3.0%); (iv) Mono High Resolution (MHR)
(163308; 25.9%); (v) Call Rate Below Threshold (CRBT) (22635; 3.6%); and (vi) Other (91425, 14.5%). Only
the first three groups (PHR, NMH and OTV) were considered useful, and they accounted for 53.9% of the
Wheat660K SNP array probes. y p
Of the 339757 functional SNPs, 136973 (40.3%) were polymorphic between KN9204 and J411. Of these, 8407
had more than 10% missing data in the 188 KJ-RILs, and were removed from the linkage analysis. Among the
remaining 128566 SNPs, 90567 (70.4%) were transitions, and 37999 (29.6%) were transversions. The 128566
functional SNPs and the previously reported 591 loci3 were used for linkage analysis and map construction. The
129157 markers fell into 5175 bins, and to create a chromosome frame, only one marker was selected as a repre-
sentative from each bin. Overview of the high-density wheat genetic map. The 5175 bin markers were used for linkage anal-
ysis based on their scores in the 188 KJ-RILs. In total, 4959 bin markers were mapped to the wheat genetic map. The co-segregated markers (redundant markers) were then added to the high-density genetic map based on
information of bin serial number and the genetic information of the corresponding bin markers. A high-density
genetic map with 119566 loci spanning 4424.4 cM was constructed (Table S2). Of these loci, 119001 were SNP
markers derived from the Wheat660K SNP array, and the remaining 565 markers were reported previously by
Cui et al.3. Of the 119001 SNPs, 83953 (70.5%) were transitions, and 35048 (29.5%) were transversions (data not
shown). Utilization of a Wheat660K SNP
array-derived high-density genetic
map for high-resolution mapping of
a major QTL for kernel number This Wheat660K SNP array is genome-specific with high density and is
highly efficient with a wide range of potential applications (http://wheat.pw.usda.gov/ggpages/topics/Wheat660_
SNP_array_developed_by_CAAS.pdf). However, genetic position in relation to Wheat660K SNPs has not yet
been documented. In this work, for the first time, we report a high-density map for wheat constructed from this
Wheat660K SNP Array. Based on SNP flanking sequences, we assigned SNPs to the genome assembly of T. aes-
tivum cv. Chinese Spring (CS) (https://urgi.versailles.inra.fr/download/iwgsc/IWGSC-WGA_Sequences/). We
also compared our high-density genetic map with the consensus genetic maps of Wheat90K and Wheat820K
based on the common contigs assembled in the chromosome survey sequencing (CSS) project. Comparative
genomic analyses based on the mapped SNP flanking sequences and the corresponding contig sequences were
also performed with the genomes of Brachypodium distachyon, Oryza sativa, Zea mays, and Sorghum bicolor. Using this mapping resource along with the phenotypic data, we identified important QTLs for yield-related trait. A major stable QTL for kernel number was identified and then characterized in detail based on the high-density
genetic map and comparative genomic analysis. Utilization of a Wheat660K SNP
array-derived high-density genetic
map for high-resolution mapping of
a major QTL for kernel number Most of these markers are mapped on the
telomeric regions, and there is very limited map resolution in proximal part of the chromosomes3. Therefore, the
density and coverage of the current genetic maps are less than satisfactory. y
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Single-nucleotide polymorphisms (SNPs) are the most abundant type of molecular marker. Accurate and
reliable methods have been developed to perform high-throughput genotyping based on SNPs12. With the devel-
opment of new sequencing technologies, increasing numbers of SNPs have been discovered in wheat1, 2, 13–15. 1Center for Agricultural Resources Research, Institute of Genetics and Developmental Biology, Chinese Academy
of Sciences, Shijiazhuang, 050022, China. 2Genetic Improvement Centre of Agricultural and Forest Crops, College
of Agriculture, Ludong Unversity, Yan’tai, 264025, China. 3Chengdu Institute of Biology, Chinese Academy of
Sciences, Chengdu, 610041, China. 4Xinxiang Academy of Agricultural Sciences, Xinxiang, 453000, China. 5Institute
of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100081, China. 6University of Chinese Academy
of Sciences, Beijing, 10049, China. 7State Key Laboratory of Plant Cell and Chromosomal Engineering, Chinese
Academy of Sciences, Beijing, 100101, China. Fa Cui, Na Zhang and Xiao-li Fan contributed equally to this work. Correspondence and requests for materials should be addressed to W.Z. (email: edithor@126.com) or G.-y.Z. (email:
zhaoguangyao@caas.cn) or J.-m.L. (email: ljm@sjziam.ac.cn) Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 1 www.nature.com/scientificreports/ Cavanagh et al.12 released a hexaploid wheat consensus genetic map with 7504 SNP markers from the Wheat9K
SNP array using a combination of seven mapping populations. Wang et al.16 mapped 46977 SNPs from the
Wheat90K array to the hexaploid wheat genetic map using a combination of eight mapping populations. Using
both Wheat9K and Wheat90K arrays, Maccaferri et al.17 released a high-density tetraploid wheat consensus
genetic map with 30144 markers (including 26626 SNPs and 791 SSRs) by integrating 13 data sets from inde-
pendent biparental mapping populations. Recently, Winfield et al.18 documented a hexaploid wheat consensus
map with 56 505 SNP markers from the Wheat820K array, spanning 3739 cM in length, using three independent
biparental populations. However, although a high-density hexaploid wheat genetic map (>100 000 markers)
based on an individual biparental mapping population would be valuable for further genetic research, such as
high-resolution mapping and map-based cloning of a targeted major quantitative trait locus (QTL), no such map
has been released. Results
SNP Most markers were mapped to the B (44.6%) and A genomes (43.7%), and only 11.7% markers were Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 2 www.nature.com/scientificreports/ Figure 1. Distribution of the 119 566 loci on the 21 wheat chromosomes. The 21 circles indicate the 21 wheat
chromosomes, with 7D to 1D, 7B to 1B, and 7A to 1A from inside to outside, respectively. Each chromosome
was evenly divided into 100 segments based on the map length, which are shown with different colours. The arc lengths indicate the percentage of markers on each segment (The total number of markers on the
corresponding chromosome divided by the number of markers on the segments). The arc in red colour indicates
the chromosomal bins near the centromere. The upper left histogram in the circle indicates the distribution of
markers on each chromosome; the upper right histogram in the circle indicates the distribution of markers near
the centromere on each chromosome; the bottom right histogram in the circle indicates the distribution of map
length on each chromosome; the bottom left histogram in the circle indicates the distribution of markers in/
near the coding sequences. Figure 1. Distribution of the 119 566 loci on the 21 wheat chromosomes. The 21 circles indicate the 21 wheat
chromosomes, with 7D to 1D, 7B to 1B, and 7A to 1A from inside to outside, respectively. Each chromosome
was evenly divided into 100 segments based on the map length, which are shown with different colours. The arc lengths indicate the percentage of markers on each segment (The total number of markers on the
corresponding chromosome divided by the number of markers on the segments). The arc in red colour indicates
the chromosomal bins near the centromere. The upper left histogram in the circle indicates the distribution of
markers on each chromosome; the upper right histogram in the circle indicates the distribution of markers near
the centromere on each chromosome; the bottom right histogram in the circle indicates the distribution of map
length on each chromosome; the bottom left histogram in the circle indicates the distribution of markers in/
near the coding sequences. mapped to the D genome. For the map lengths, the A, B, and D genomes covered 36.4%, 27.7%, and 35.8% of
the total map length, respectively. The chromosome sizes ranged from 84.4 cM (chromosome 1BL) to 289.1 cM
(chromosome 5D), averaging 210.7 cM per chromosome. Results
SNP The number of markers on each chromosome ranged
from 78 (chromosome 1BL) to 13898 (chromosome 3B), with a mean of 5693.6 loci per chromosome. Due to
the 1BL/1RS translocation of KN9204, the 1RS- or 1BS-specific markers not only showed co-segregation but
also exhibited distorted segregation in the KJ-RILs as shown by Cui et al.3. These markers were excluded from
the linkage analysis and map construction, which reduced the genetic maps for analysis to 1BL only. In addi-
tion, no markers on chromosome 5BS were polymorphic between KN9204 and J411, resulting in the release
of the 5BL genetic map only. Of the 119566 loci, 33598 (28.1%) were distributed on chromosomal regions near
the centromeres. Marker density per genetic distance unit peaked at the centromeric regions, possibly due to a
combination of low recombination rate in the centromeric regions and even gene distribution along the chro-
mosomes (Fig. 1). The 4 959 bin markers are shown in the genetic map (Fig. 2). The following mapping-bin sets
were observed: approximately 3.7% and 3.2% of the markers were unique for genomes A and B, respectively, and
approximately 9.6% of the markers for the D genome showed unique segregation patterns. mapped to the D genome. For the map lengths, the A, B, and D genomes covered 36.4%, 27.7%, and 35.8% of
the total map length, respectively. The chromosome sizes ranged from 84.4 cM (chromosome 1BL) to 289.1 cM
(chromosome 5D), averaging 210.7 cM per chromosome. The number of markers on each chromosome ranged
from 78 (chromosome 1BL) to 13898 (chromosome 3B), with a mean of 5693.6 loci per chromosome. Due to
the 1BL/1RS translocation of KN9204, the 1RS- or 1BS-specific markers not only showed co-segregation but
also exhibited distorted segregation in the KJ-RILs as shown by Cui et al.3. These markers were excluded from
the linkage analysis and map construction, which reduced the genetic maps for analysis to 1BL only. In addi-
tion, no markers on chromosome 5BS were polymorphic between KN9204 and J411, resulting in the release
of the 5BL genetic map only. Of the 119566 loci, 33598 (28.1%) were distributed on chromosomal regions near
the centromeres. Marker density per genetic distance unit peaked at the centromeric regions, possibly due to a
combination of low recombination rate in the centromeric regions and even gene distribution along the chro-
mosomes (Fig. 1). The 4 959 bin markers are shown in the genetic map (Fig. 2). Results
SNP Contigs from chromosome 3B were not separated into short/long arm bins, as individual arm datasets were not
generated for this chromosome in the Chromosome Survey Sequence (CSS) project. Figure 2. The high-density genetic map of wheat developed using an RIL population derived from the cross
of cultivars KN9204 and J411. For the redundant loci that showed co-segregation in the 188 KJ-RILs, only one
unique informative marker is shown in this figure. The approximate positions of the centromeres are indicated
by pink. Short arms are at the top. The positions of the marker loci are indicated using a ruler on the left side of
this figure. The names of the marker loci are listed to the right of the corresponding chromosomes. Loci in red
were best hits to Chinese Spring (CS) contigs of the short arm of the corresponding chromosomes. Loci in green
were best hits to CS contigs of the long arm of the corresponding chromosomes. Loci in black were unknown. Contigs from chromosome 3B were not separated into short/long arm bins, as individual arm datasets were not
generated for this chromosome in the Chromosome Survey Sequence (CSS) project. 2B (10.6 cM), 2D (10.5 cM), 3 A (19.9 cM, 19.2 cM), 3D (12.1 cM), 4D (11.5 cM, 11.2 cM, 11.0 cM, 11.0 cM), 5 A
(12.1 cM), 5D (13.2 cM), 6 A (19.2 cM), 6B (13.8 cM, 13.2 cM, 12.7 cM), 6D (14.6 cM, 13.6 cM, 10.9 cM, 10.7 cM),
7B (16.3 cM, 10.1 cM), and 7D (18.8 cM) (Fig. 2). Comparative genomic analysis. Of the 119566 loci, 118998 (99.5%) were best hits to 57036 CSS contigs,
with 2.1 polymorphic markers per contig. In total, 93.0% contigs had coincident physical and genetic positions,
4.6% were mapped to homoeologous chromosomes such as 1 A in physical position and 1B in the KJ-RIL genetic
map, and 2.4% were disordered (Fig. 2; Figure S1; Table S3). Based on the SNP flanking sequences, we assigned
116 261 SNPs to the recently released wheat genome assembly. SNP order in the present genetic map was in good
agreement with that in the wheat genome assembly, with the exception of chromosome 7DL, in which a segment
inversion was identified (Fig. Results
SNP The following mapping-bin sets
were observed: approximately 3.7% and 3.2% of the markers were unique for genomes A and B, respectively, and
approximately 9.6% of the markers for the D genome showed unique segregation patterns. y
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Considering the unique markers (the 4 959 bin markers), the highest marker saturation was found in genome
A (39.3%), followed by genomes B (33.6%) and D (27.1%). The average distance between adjacent bin mark-
ers ranged from 0.6 cM for 6B to 1.5 cM for 4D, with an average of 0.9 cM per marker pair. Gaps greater than
20.0 cM but less than 30.0 cM were present in chromosomes 3 A (24.9 cM), 3B (21.9 cM) and 4D (20.4 cM); gaps
greater than 10 cM but less than 20.0 cM were present in chromosomes 1 A (13.9 cM, 11.7 cM), 2 A (14.6 cM), Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 3 www.nature.com/scientificreports/ www.nature.com/scientificreports/ Figure 2. The high-density genetic map of wheat developed using an RIL population derived from the cross
of cultivars KN9204 and J411. For the redundant loci that showed co-segregation in the 188 KJ-RILs, only one
unique informative marker is shown in this figure. The approximate positions of the centromeres are indicated
by pink. Short arms are at the top. The positions of the marker loci are indicated using a ruler on the left side of
this figure The names of the marker loci are listed to the right of the corresponding chromosomes Loci in red Figure 2. The high-density genetic map of wheat developed using an RIL population derived from the cross
of cultivars KN9204 and J411. For the redundant loci that showed co-segregation in the 188 KJ-RILs, only one
unique informative marker is shown in this figure. The approximate positions of the centromeres are indicated
by pink. Short arms are at the top. The positions of the marker loci are indicated using a ruler on the left side of
this figure. The names of the marker loci are listed to the right of the corresponding chromosomes. Loci in red
were best hits to Chinese Spring (CS) contigs of the short arm of the corresponding chromosomes. Loci in green
were best hits to CS contigs of the long arm of the corresponding chromosomes. Loci in black were unknown. Results
SNP qKnps-4A as detected by MapQTL 6.0, IciMapping 4.1, and QTLNetwork 2.0. Software
LOD value
Position (cM)
Additive effect
PVE %
MapQTL 6.0
3.4 to 9.7
149.8 to 155.0
−1.5 to −2.4
8.0 to
21.2
IciMapping 4.1
2.8 to 16.8
149.7 to 154.5
−1.2 to −2.4
9.5 to
20.3
QTLNetwork 2.0
P value:
0.000000
149.8
−1.9
10.9 Table 1. qKnps-4A as detected by MapQTL 6.0, IciMapping 4.1, and QTLNetwork 2.0. chromosomes 3 A/3D corresponded to Bd2 and Bd3, wheat chromosomes 4 A/4B/4D corresponded to Os3, and
wheat chromosomes 7 A/7B/7D corresponded to Sb10. However, for most synteny blocks, the chromosomes were
more fragmented and scattered, with a high frequency of breakdown. chromosomes 3 A/3D corresponded to Bd2 and Bd3, wheat chromosomes 4 A/4B/4D corresponded to Os3, and
wheat chromosomes 7 A/7B/7D corresponded to Sb10. However, for most synteny blocks, the chromosomes were
more fragmented and scattered, with a high frequency of breakdown. g
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To specify the coding-region SNPs (cSNPs), perigenic SNPs (pSNPs), and intergenic SNPs (iSNPs) among
the SNP markers mapped in the KJ-RIL genetic map, a BLASTX search was performed against the CDS of T. aestivum, using SNP flanking sequences and the corresponding contig sequences as queries. Using SNP flanking
sequences as the query, 8.9% (10104 SNPs) were best hits to the CDSs of the T. aestivum, and these were consid-
ered to be the cSNPs. When using the contig sequences where the markers were best hits to as the query, approx-
imately 36.8% (41689 SNPs) were best hits for the CDSs of the T. aestivum, indicating that 27.9% (36.8–8.9%) of
SNPs were pSNPs. The remaining 63.2% are likely iSNPs (Table S5). QTLs for KNPS and the prediction of candidate genes. Using the mapping resource along with
the phenotypic data we identified important QTLs for yield-related trait. A major stable QTL for kernel num-
ber per spike (KNPS; qKnps-4A) was verified in 10 environments by using MapQTL 6.0, IciMapping 4.1, and
QTLNetwork 2.0 (Table 1; Fig. 4; Figures S5, S6). qKnps-4A contributed to 8.0–21.2% of the KNPS phenotypic
variation in the 188 KJ-RILs, with alleles from J411 increasing kernels per spike by 1.2–2.4. The QTL peaks
of qKnps-4A were in 149.8–155.0 cM, 149.7–154.5 cM, and 149.8 cM, as detected by MapQTL 6.0, IciMapping
4.1, and QTLNetwork 2.0, respectively, with Ax-109844107–Ax-110540586 in 149.6–150.3 cM in the overlapping
confidence intervals. Results
SNP 3; Figure S2; Table S3).i i
An overview (from the wheat genetic map perspective) of the relationships between the five grass family
genomes at the resolution of the genetic map in centimorgans is provided in Figures S3 and S4. There were 113288
markers that corresponded to the CDS of Brachypodium (65357 markers) (Table S4; Figure S3a, S4a), rice (58825
markers) (Table S4; Figures S3b, S4b), maize (53745 markers) (Table S4; Figures S3c, S4c), and sorghum (59994
markers) (Table S4; Figures S3d, S4d). In general, chromosomes belonging to the same homoeologous groups
showed correspondence with the same grass chromosomes, but some differences were observed. Large and espe-
cially small synteny blocks across wheat and grass family chromosomes were observed, indicating the complexity
of the wheat genome among the grass family genomes. The structural relationships among the genomes indi-
cate that for some individual wheat chromosomes, there is a preponderance of corresponding grass genes from
one or two certain linkage groups. For example, wheat chromosomes 2 A/2B/2D corresponded to Bd5, wheat Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 4 www.nature.com/scientificreports/ Figure 3. Schematic representation of the syntenic relationships between any one marker in wheat genetic and
physical maps. Gen-1A to Gen-7D represent the 21 wheat chromosomal genetic maps released in this paper;
Phy-1A to Phy-7D represent the 21 wheat chromosomal physical maps, which were constructed by assigning
116 261 SNPs to the wheat genome assembly using SNP flanking sequences as the query. For the redundant loci
that showed co-segregation in the 188 KJ-RILs, only one unique informative marker is shown in this figure. Figure 3. Schematic representation of the syntenic relationships between any one marker in wheat genetic and
physical maps. Gen-1A to Gen-7D represent the 21 wheat chromosomal genetic maps released in this paper;
Phy-1A to Phy-7D represent the 21 wheat chromosomal physical maps, which were constructed by assigning
116 261 SNPs to the wheat genome assembly using SNP flanking sequences as the query. For the redundant loci
that showed co-segregation in the 188 KJ-RILs, only one unique informative marker is shown in this figure. Software
LOD value
Position (cM)
Additive effect
PVE %
MapQTL 6.0
3.4 to 9.7
149.8 to 155.0
−1.5 to −2.4
8.0 to
21.2
IciMapping 4.1
2.8 to 16.8
149.7 to 154.5
−1.2 to −2.4
9.5 to
20.3
QTLNetwork 2.0
P value:
0.000000
149.8
−1.9
10.9
Table 1. Results
SNP Thus, we predicted that the candidate genes for qKnps-4A might be within the overlapping Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 5 www.nature.com/scientificreports/ Figure 4. Overlapping confidence intervals of QTLs for kernel number per spike (KNPS) based on MapQTL
6.0, IciMapping 4.1 and QTLNetwork 2.0. The segments in red are the overlapping confidence intervals of
qKnps-4A, and the overlapping flanking markers of qKnps-4A are indicated by triangles in red, which were
detected by QTLNetwork 2.0 based on the combined 10 environmental phenotypic values. Figure 4. Overlapping confidence intervals of QTLs for kernel number per spike (KNPS) based on MapQTL
6.0, IciMapping 4.1 and QTLNetwork 2.0. The segments in red are the overlapping confidence intervals of
qKnps-4A, and the overlapping flanking markers of qKnps-4A are indicated by triangles in red, which were
detected by QTLNetwork 2.0 based on the combined 10 environmental phenotypic values. confidence intervals of 0.7 cM. Based on the genome assembly of T. aestivum cv. CS (https://urgi.versailles. inra.fr/download/iwgsc/IWGSC-WGA_Sequences/), the overlapping confidence intervals of Ax-109844107–
Ax-110540586 spanned 3.23 Mb (4 A:680398739–4 A:683638403) in physical position (Figure S7). This region
harbours 65 predicted genes in wheat (Figure S8), which might include the candidate gene for qKnps-4A. Discussionh The high-density SNP map developed in the present study first documented the genetic positions of 119 001
SNPs from the Wheat660K SNP array. Based on the SNP flanking sequences, we assigned 118 785 SNPs to 56
904 CSS-assembled contigs (Table S3). The physical positions of the corresponding CSS-assembled contigs could
be used to validate genetic position (chromosome and chromosomal arms assignment). As shown in Fig. 2 and
Table S3, the physical and genetic positions of these mapped markers were generally in agreement. SNPs from the Wheat660K SNP array. Based on the SNP flanking sequences, we assigned 118 785 SNPs to 56
904 CSS-assembled contigs (Table S3). The physical positions of the corresponding CSS-assembled contigs could
be used to validate genetic position (chromosome and chromosomal arms assignment). As shown in Fig. 2 and
Table S3, the physical and genetic positions of these mapped markers were generally in agreement. In previous studies, 7504 SNPs from the Wheat9K SNP array12, 46977 SNPs from the Wheat90K array16,
and 56505 SNPs from the Wheat820K SNP array18 were genetically mapped to the hexaploid wheat genome. These SNPs have also been physically assigned to the corresponding CSS-assembled contigs. Based on the com-
mon CSS contigs, we analysed the synteny of the mapped SNPs (Wheat660K SNP array vs. Wheat90K array and
Wheat660K SNP array vs. Wheat820K SNP array) across different mapping populations (Figs 5 and 6). These ,
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In previous studies, 7504 SNPs from the Wheat9K SNP array12, 46977 SNPs from the Wheat90K array16,
and 56505 SNPs from the Wheat820K SNP array18 were genetically mapped to the hexaploid wheat genome. These SNPs have also been physically assigned to the corresponding CSS-assembled contigs. Based on the com-
mon CSS contigs, we analysed the synteny of the mapped SNPs (Wheat660K SNP array vs. Wheat90K array and
Wheat660K SNP array vs. Wheat820K SNP array) across different mapping populations (Figs 5 and 6). These Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 6 www.nature.com/scientificreports/ www.nature.com/scientificreports/ Figure 5. Synteny of the mapped SNPs from the Wheat660K SNP array (in blue) with that from the Wheat90K
array (in red) based on their common CSS-assembled contigs. Figure 5. Synteny of the mapped SNPs from the Wheat660K SNP array (in blue) with that from the Wheat90K
array (in red) based on their common CSS-assembled contigs. Figure 6. Discussionh Synteny of the mapped SNPs from the Wheat660K SNP array (in blue) with that from the Wheat820K
array (in green) based on their common CSS-assembled contigs. Notes: The genetic maps of chromosomes 3A,
4B, 5D, 6D and 7B derived from the Wheat820K SNP array are shown in inverted form, with long arms at the
top and short arms at the bottom18. In this figure, these inversions were corrected. Figure 6. Synteny of the mapped SNPs from the Wheat660K SNP array (in blue) with that from the Wheat820K
array (in green) based on their common CSS-assembled contigs. Notes: The genetic maps of chromosomes 3A,
4B, 5D, 6D and 7B derived from the Wheat820K SNP array are shown in inverted form, with long arms at the
top and short arms at the bottom18. In this figure, these inversions were corrected. common contigs were generally aligned with the chromosomes in a consistent order across different mapping
populations, verifying the accuracy and credibility of our high-density genetic map. Regarding the total map
length, the Wheat90K/820 SNP consensus integrative genetic map is 2.4/1.9 times longer (data not shown) than
that of the present map. The increased genetic map length is proportionate to the increased mapping population
size16, 18. A relatively small mapping population size resulted in limited identification of recombination events
and lower resolution of the genetic map, contributing to the short map length in this study24, 25. In addition,
comparative analysis among these three SNP maps revealed that genetic maps of chromosomes 3 A, 4B, 5D, 6D,
and 7B derived from the wheat 820 K SNP array were inverted, with long arms at the top and short arms at the
bottom (data not shown)18. Chromosome 4 A, derived from the Wheat820K SNP array, might be involved in
chromosomal rearrangement compared with the genetic maps derived from the Wheat90K SNP array and our common contigs were generally aligned with the chromosomes in a consistent order across different mapping
populations, verifying the accuracy and credibility of our high-density genetic map. Regarding the total map
length, the Wheat90K/820 SNP consensus integrative genetic map is 2.4/1.9 times longer (data not shown) than
that of the present map. The increased genetic map length is proportionate to the increased mapping population
size16, 18. Discussionh Based on genetic marker sequence flanking for these QTLs and the recently released WGS
assembly, we compared physical positions with qKnps-4A to determine whether they were common interacting
QTLs or regions across genetic backgrounds. The results are shown in Supplementary Table S6. qKnps-4A from
our previous research shared a confidence interval (4 A:664209916–4 A:736771450) with qKnps-4A in this study
(4 A:680398739–4 A:683638403). Interestingly, qKnps-4A has been detected in both the WY and WJ populations
(two related RIL populations sharing one common parental line of Weimai 8)4. QKNS.caas-4AL detected by Gao
et al.54, also shared a confidence interval (4 A:632 864 778–4 A:688 093 018) with qKnps-4A in our study. These
coincidences confirmed the authenticity of qKnps-4A, which should be subjected to fine mapping and map-based
cloning in the future. In fact, this work is being conducted based on the secondary mapping population of qKnps-
4A’s (data not shown). (
)
QTL mapping based on the primary mapping population could precisely characterize and locate genes under-
lying specific agronomic traits, which was also true for both major and moderate/minor QTLs55. Previous stud-
ies have confirmed that tagged or cloned genes were near their original QTL positions (logarithm of the odds
(LOD) peak)56–67. In this study, qKnps-4A was repeatedly identified using MapQTL 6.0, IciMapping 4.1, and
QTLNetwork 2.0, based on mathematical models of composite interval mapping (CIM), inclusive composite
interval mapping (ICIM) and the mixed linear model approach (MLMA), respectively. In addition, the KNPS
values of the 188 KJ-RILs and their parental lines were evaluated in 10 different environments. Based on the 10
environmental phenotypic values along with our high-density genetic map, we confirmed QTL peak position
with the aforementioned QTL mapping software and found the Ax-109844107–Ax-110540586 overlapping con-
fidence interval, which spanned 3.23 Mb (4 A:680 398 739–4 A:683 638 403) in physical position (Table 1; Fig. 4;
Figures S5, S6, S7). In addition, qKnps-4A has been mapped to 4 A:664 209 916–4 A:736 771 450 in both the WY
and WJ populations4 and to 4 A:632 864 778–4 A:688 093 018 in the Zhou 8425B/Chinese Spring population54
(Supplementary Table 7). These coincidences across different genetic backgrounds, multiple environments and
diverse QTL models strongly supported the hypothesis that the genes underlying qKnps-4A are likely located
within 4 A:680398739–4 A:683638403. One of the 65 predicated genes within this interval might be the candi-
date gene for qKnps-4A (Figure S8). Discussionh Conservation of gene identity and collinearity among gramineous plants will
depend on the rates of genome/gene evolution and rearrangement in the investigated species22. There is a high
level of genome-synteny among gramineous plants, especially wheat, Brachypodium and rice, with wheat being
more closely related to Brachypodium than to rice22, 23, 30–32. Wheat improvement programs can benefit from the
use of comparative genetics to transfer information about genes from model species to wheat, to help identify
genes controlling traits of interest, and to assess within-species allelic diversity so that the best alleles can be iden-
tified and assembled in superior varieties. i
p
In this study, synteny analyses among common wheat, Brachypodium, rice, sorghum and maize genomes
were performed based on the collinearity of the corresponding orthologous genes (best hits of CDSs) (Table S4;
Figures S3 and S4). Features of the wheat-grass genome relationships revealed in this study included a high
frequency of breakdown in microcollinearity throughout the genomes compared to the previous RFLP-based
maps22, 31, 33–36. Both large and especially small synteny blocks across wheat chromosomes and grass family chro-
mosomes were observed in this study. These features might be attributed to the higher number of markers used in
this study. More recently, Russo et al.32 conducted collinearity analysis across durum wheat, Brachypodium, and
rice based on a high-density durum wheat genetic map derived from the Wheat90K SNP assay. Both large and
especially small synteny blocks across wheat chromosomes and grass family chromosomes were also observed
in that study. Based on a high-density genetic map, we documented a wheat genome perspective of homologous
sorghum and maize genome locations based on comparative sequence analysis. A wheat genome view of homol-
ogous gramineous plant genome locations based on comparative sequence analysis would considerably improve
the predictability and efficiency of information transfer, and would be benefit evolutionary studies. p
yfi
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Wheat yield is determined by three yield components: productive spikes per unit area, KNPS, and kernel
weight, determine wheat yield. Among them, the KNPS value has steadily increased, indicating a substan-
tial contribution of increased KNPS to increased wheat yield37, 38. Over the past several decades, numerous
QTLs (or genes) for wheat kernel number have been documented based on both linkage-mapping and
association-mapping analyses4, 38–54. Some of these studies have also reported QTLs for KNPS on chromosome
4 A4, 40, 44, 50, 51, 53, 54. Discussionh A relatively small mapping population size resulted in limited identification of recombination events
and lower resolution of the genetic map, contributing to the short map length in this study24, 25. In addition,
comparative analysis among these three SNP maps revealed that genetic maps of chromosomes 3 A, 4B, 5D, 6D,
and 7B derived from the wheat 820 K SNP array were inverted, with long arms at the top and short arms at the
bottom (data not shown)18. Chromosome 4 A, derived from the Wheat820K SNP array, might be involved in
chromosomal rearrangement compared with the genetic maps derived from the Wheat90K SNP array and our Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 7 www.nature.com/scientificreports/ Wheat660K SNP array. The genetic information from the common contigs and their genetic collinearity analysis
across different mapping populations will lay the foundation for obtaining a consensus integrative genetic linkage
map.h Wheat660K SNP array. The genetic information from the common contigs and their genetic collinearity analysis
across different mapping populations will lay the foundation for obtaining a consensus integrative genetic linkage p
The SNP order in the present KJ-RIL genetic map was also in good agreement with that in the physical posi-
tion, with the exception of chromosome 7DL, in which a segment inversion was identified (Fig. 3; Figure S2;
Table S3). A previous report showed that wheat chromosome 7 was likely to involve a complex interchange26. These findings prompted us to search for candidate genes of targeted major QTLs. In wheat, a majority of recom-
bination events occurred on the most distal portions of the chromosomal arms, whereas the recombination
events tend to be suppressed around the centromere3, 27, 28. These characteristics result in a low resolution of the
genetic map in the centromeric region, which was evident in the small genetic distance in the KJ-RIL genetic
map corresponding to a large physical region around the centromere compared with the most distal portions of
the chromosomal arms (Fig. 3; Figure S2; Table S3). These findings also indicate the difficulty of high-resolution
mapping and map-based cloning of a QTL around the centromere because of the low coverage of genetic markers
and the suppression of recombination events.h The genetic and genomic research of wheat has lagged behind similar research regarding other important
crops, such as rice and maize29. Experimental procedures p
p
Plant material. In this study, F8:9 KJ-RILs derived from a cross between KN9204 and J411 were used for map
construction and QTL analysis. The original KJ-RIL population contained 427 RILs. In total, 188 randomly sam-
pled lines from the 427 KJ-RILs were used for genetic linkage analysis. In addition, 65 KN9204-derived advanced
lines/authorized varieties, three parental lines of KN9204, three control varieties from the Winter Wheat
Performance Trial of the Northern Huang-Huai Regional Nursery of China, and CS (Table S7) were genotyped
to trace the key chromosomal segment harbouring the major stable QTL for kernel number per spike (KNPS). Phenotyping. KNPS values of the 188 KJ-RILs and their parental lines were evaluated in ten different
environments (five trials that included both low- and high-nitrogen treatments). The nitrogen treatments, field
arrangements and experimental designs of the ten environments were performed as described previously3, 68, 69. Genotyping. For all subjects, leaf tissues were sampled. Genomic DNA was extracted and hybridized on the
Wheat660K SNP genotyping array by Compass Biotechnology Company (Beijing, China). The DNA samples
were prepared, and the chip genotyping was performed on the Wheat660K SNP array according to the Affymetrix
Axiom 2.0 Assay Manual Workflow protocol. DNA integrity was confirmed on agarose gels, and DNA quantity
was measured spectrophotometrically. The Wheat660K chip contains 630517 markers (http://wheat.pw.usda. gov/GG2/index.shtml). Variant quality from the Wheat660K chip genotyping was initially assessed according
to Affymetrix best practices. The 188 RILs and their parents were aslo assayed using the ‘Wheat PstI (TaqI) 2.3D’
DArT array (the medium density array) (http://www.triticarte.com.au/). The PCR-based markers were genotyped
as described in our previous study3. Genetic map construction. The 188 KJ-RILs and their parental lines were genotyped with the Wheat660K
SNP array. SNPs were rejected if they showed minor allele frequency (defined as frequency <0.3) or contained
>10% missing data. Markers were binned based on their segregation patterns in the KJ-RIL population using the
BIN function in IciMapping 4.1 (http://www.isbreeding.net/) according to Winfield et al.18. Markers that shared
their segregation pattern with at least one other marker were retained. One marker was chosen to represent each
bin on the basis of the least amount of missing data or, when the percentage of missing data was equal, at random. Markers were tested for significant segregation distortion using a chi-square test. Discussionh This information is very valuable for future high-resolution mapping and
map-based cloning of qKnps-4A. p
g
q
p
Using Ax-110540586 as a probe, the 188 KJ-RILs were divided into two groups, one group with alleles f
KN9204 and the other with alleles from J411, to perform mean comparisons regarding KNPS. The positive al Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 8 www.nature.com/scientificreports/ from qKnps-4A’s increased the KNPS value from 3.0 to 4.6, indicating a tremendous potential for their applica-
tion in wheat molecular breeding programmes designed to increase yield (Figure S9). To characterize the use of
qKnps-4A’s positive alleles in wheat breeding programmes, we dissected the genotypes of 65 KN9204-derived
advanced lines/authorized varieties, three parental lines of KN9204, three control varieties from the Winter
Wheat Performance Trial of the Northern Huang-Huai Regional Nursery of China, and CS near Ax-109844107–
Ax-110540586 (Figure S10). None of KN9204′s three parental lines carried favourable alleles for increasing
KNPS, which accounted for the negative alleles of KN9204 at this QTL. Only 3 (4.6%) of the 65 authorized/
advanced lines derived from KN9204 lines carried favourable alleles for increasing KNPS. Interestingly, two
advanced lines (KN1002-13-10 and O-97) carried heterozygous alleles, which were used to develop the secondary
mapping population of Knps-4A via self-cross (data not shown). Of the three control varieties from the Winter
Wheat Performance Trial of the Northern Huang-Huai Regional Nursery of China, S4185 carried negative alleles,
decreasing KNPS; chromosomal regions of Knps-4A in LX99 and JM22 were recombinant regions with alleles that
cannot be categorized as favourable or negative alleles that increase or decrease KNPS; and CS carried favourable
alleles, increasing KNPS. These findings indicated that favourable qKnps-4A alleles have not been fully utilized in
wheat breeding programmes in the Huang-Huai winter wheat region in China. Wheat breeders should strengthen
the selection of qKnps-4A favourable alleles in molecular breeding programmes aimed at the development of
high-yield varieties. g
y
In summary, this paper reports a high-density wheat genetic map based on an individual mapping population. In total, 119 001 SNP markers derived from the Wheat660K SNP array were mapped onto the KJ-RIL genetic
map. We observed good good collinearity of our high-density genetic map with the Wheat90K and Wheat820K
consensus genetic maps, increasing the possibility of obtaining a consensus integrative higher-density wheat
genetic map in the future. This high-density genetic map is also in good accordance with the recently released
wheat genome assembly. Discussionh Our high-density wheat genetic map provides a major resource for future wheat genetic
and genomic research. Moreover, this paper provides an overview of the structural relationships between wheat
and other gramineous plant genomes based on comparative genomics analysis. Finally, a major stable QTL for
kernel number was thoroughly characterized based on this high-density genetic map and comparative genomics
analysis. Experimental procedures SNPs were sorted into groups
using the MAP function in IciMapping 4.1, with the previously mapped 591 loci serving as anchored markers3. A logarithm of the odds (LOD) score of 3.5 and a recombination fraction of 0.3 were used to sort the SNPs with
the Kosambi mapping function70. Groups were ordered with the Kosambi mapping function within the JoinMap
v. 4.0, using a LOD score ≥3 after preliminary analysis of SNPs with LOD scores ranging from 2 to 10. The long
and short arms of each chromosome were identified from the IWGSC wheat survey sequence (http://www.wheat-
genome.org/), and groups were orientated to have the short arm above the long arm. MapChart 2.2 (http://www. biometris.nl/uk/Software/MapChart/) was used to draw the genetic map. QTL mapping analysis. The QTLs for KNPS were identified using MapQTL 6.0 (based on CIM, https://
www.kyazma.nl/index.php/mc.MapQTL/), IciMapping 4.1 (based on ICIM, http://www.isbreeding.net/soft-
ware/?type=detail&id=18), and QTLNetwork 2.0 (based on MLMA, http://ibi.zju.edu.cn/software/qtlnetwork/). For MapQTL 6.0 and IciMapping 4.1, the average KNPS values of each KJ-RIL in each environment were used for
individual environment QTL mapping analysis. For QTLNetwork 2.0, the KNPS values of each KJ-RIL in each Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 9 www.nature.com/scientificreports/ replication of the 10 environments were assembled to perform combined QTL analysis across environments to
identify QTLs with additive-by-environment (A by E) interaction effects. The overlapping confidence intervals
detected with the abovementioned programs were used to predict the candidate genes based on the sequence
information of the flanking markers and the IWGSC WGA v0.4 assembly of chromosome 4A (https://urgi.ver-
sailles.inra.fr/download/iwgsc/IWGSC-WGA_Sequences/). replication of the 10 environments were assembled to perform combined QTL analysis across environments to
identify QTLs with additive-by-environment (A by E) interaction effects. The overlapping confidence intervals
detected with the abovementioned programs were used to predict the candidate genes based on the sequence
information of the flanking markers and the IWGSC WGA v0.4 assembly of chromosome 4A (https://urgi.ver-
sailles.inra.fr/download/iwgsc/IWGSC-WGA_Sequences/). Comparative genomic analysis. The SNP flanking sequences mapped in the KJ-RIL map were kindly
provided by Professor Jia JZ. We used the Basic Local Alignment Search Tool (BLAST) (ftp://ftp.ncbi.nlm.nih. gov/blast/executables/release/) to align the SNP probes to the IWGSC survey sequences (contigs). All IWGSC
survey sequences were downloaded from http://www.wheatgenome.org/. In addition, contig sequences to which
the SNPs were best hits were screened in a BLASTN search against the coding sequences (CDSs) of B. distachyon,
rice (O. sativa L.), maize (Z. Experimental procedures mays L.), and sorghum (S. vulgare L.). All CDSs were downloaded from http://plants. ensembl.org/index.html. An expectation value (E) of 1E−10 was used as the significance threshold. Synteny anal-
yses with common wheat, Brachypodium, rice, maize and sorghum genomes were performed based on the SNP
orders in the KJ genetic map and on the corresponding CDSs in the genome sequences of Brachypodium, rice,
maize and sorghum genomes. g
g
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the contig sequences were downloaded from https://wheat-urgi.versailles.inra.fr/Seq-Repository. More recently,
the genome assembly of T. aestivum cv. Chinese Spring (CS) has been released (https://urgi.versailles.inra.fr/
download/iwgsc/IWGSC-WGA_Sequences/). Based on SNP flanking sequences, we assigned SNPs to this wheat
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novel allele of CRY2. Nat. Genet. 29, 435–440 (2001). 64. Liu, J., Van Eck, J., Cong, B. & Tanksley, S. D. A new class of regulatory genes underlying the cause of pear-shaped tomato fruit. Proc. Natl. Acad. Sci. USA 99, 13302–13306 (2002). 65. Menendez, C. M. et al. Cold sweetening in diploid potato: mapping quantitative trait loci and candidate genes. Genetics 162,
1423–1434 (2002). 6. Lin, H. References A physical map of the 1-gigabase bread wheat chromosome 3B. Science 322, 101–104 (2008). h
29. Paux, E. et al. A physical map of the 1-gigabase bread wheat c 29. Paux, E. et al. A physical map of the 1 gigabase bread wheat chromosome 3B. Science 322, 101 104 (2008). 30. Huo, N. et al. Structural characterization of Brachypodium genome and its syntenic relationship with rice and wheat. Plant Mol. Biol. 70, 47–61 (2009). Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 10 Acknowledgements g
We are grateful to Compass Biotechnology Company (Beijing, China) for their generous assistance and
technical support. We also thank Miss Liu Shuai for her valuable and thoughtful service. This research has
been jointly supported by the following grants: the National Key Research and Development Program of China
(2016YFD0101802), the National Natural Science Foundation of China (31471573), the Strategic Priority Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 11 www.nature.com/scientificreports/ Research Program of the Chinese Academy of Sciences (XDA08030107), the Youth Innovation Promotion
Association (2016095), the Hebei Provincial Science and Technology Research and Development Project
(16226320D) and the China Agriculture Research System (CARS-03-01B). Author Contributions J.L., G.Z. and J.J. proposed the method and supervised the project. F.C., N.Z. and X.F. performed genotypic
identification and data analysis. W.Z., C.Z., L.Y., R.P., M.C., J.H., X.Z., and J.J. conducted the phenotypic
evaluation. F.C., N.Z. and X.F. wrote the manuscript. All authors discussed the results and reviewed the
manuscript. Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 Additional Information dd t o a
o
at o
Supplementary information accompanies this paper at doi:10.1038/s41598-017-04028-6h Supplementary information accompanies this paper at doi:10.1038/s41598-017-04028-6 Competing Interests: The authors declare that they have no competing interests. Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and
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copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. © The Author(s) 2017 Scientific Reports | 7: 3788 | DOI:10.1038/s41598-017-04028-6 12
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English
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A Novel Position Estimation Method Based on Displacement Correction in AIS
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Sensors
| 2,014
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cc-by
| 6,759
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Yi Jiang 1,2,*, Shufang Zhang 1 and Dongkai Yang 2 Yi Jiang 1,2,*, Shufang Zhang 1 and Dongkai Yang 2 1 Information Science and Technology College, Dalian Maritime University, Dalian 116026, China;
E-Mail: sfzhang@dlmu.edu.cn 1 Information Science and Technology College, Dalian Maritime University, Dalian 116026, China;
E-Mail: sfzhang@dlmu.edu.cn 2 School of Electronic and Information Engineering, Beihang University, Beijing 100083, China;
E-Mail: edkyang@buaa.edu.cn * Author to whom correspondence should be addressed; E-Mail: jiangyi.dlmu@gmail.com;
Tel.: +86-1810-4080-985; Fax: +86-411-8472-3118 (ext. 801). * Author to whom correspondence should be addressed; E-Mail: jiangyi.dlmu@gmail.com;
Tel.: +86-1810-4080-985; Fax: +86-411-8472-3118 (ext. 801). Received: 25 June 2014; in revised form: 17 August 2014 / Accepted: 21 August 2014 /
Published: 17 September 2014 Received: 25 June 2014; in revised form: 17 August 2014 / Accepted: 21 August 2014 /
Published: 17 September 2014 Abstract: A new position estimation method by using the signals from two automatic
identification system (AIS) stations is proposed in this paper. The time of arrival (TOA)
method is enhanced with the displacement correction, so that the vessel’s position can be
determined even for the situation where it can receive the signals from only two AIS base
stations. Its implementation scheme based on the mathematical model is presented. Furthermore, performance analysis is carried out to illustrate the relation between the
positioning errors and the displacement vector provided by auxiliary sensors. Finally, the
positioning method is verified and its performance is evaluated by simulation. The results
show that the positioning accuracy is acceptable. Keywords: positioning method; displacement correction; time of arrival; automatic
identification system (AIS) Sensors 2014, 14, 17376-17389; doi:10.3390/s140917376 Sensors 2014, 14, 17376-17389; doi:10.3390/s140917376 sensors
ISSN 1424-8220
www.mdpi.com/journal/sensors
OPEN ACCESS sensors
ISSN 1424-8220
www.mdpi.com/journal/sensors
OPEN ACCESS Sensors 2014, 14 Sensors 2014, 14 17377 improve the robustness of the whole positioning and navigation process [1]. In fact, various position
estimation methods and their applications in marine navigation have been widely investigated in the
literature, such as e-Loran systems [2–5], inertial navigation systems [6,7], terrain referenced
navigation systems [8], vessel traffic service and coastal surveillance systems [9,10], etc. But the
automatic identification system (AIS), which is essentially a communication system [11], is seldom
used for robust PNT. It provides position information obtained from GNSS. Obviously, it could
promote a robust PNT for marine navigation if AIS could provide PNT information by itself, since
vessels are mandated to carry AIS equipment according to the IMO requirements for its member
countries [12]. To develop positioning service for AIS with widespread distribution is helpful to make
use of the existing resources for enhancing robustness in marine navigation [13]. The main difficulty in obtaining the position information from AIS itself is that the number of
received signals from AIS base stations may be fewer than necessary to solve the vessel’s position,
since AIS is originally designed as a communication system, not a PNT system. In other words, the
geometrical distribution of AIS base stations may be poor for positioning. Actually, the placement of
base stations provides sufficient signal coverage overlap for ensuring AIS reliability and stability. This
implies that the vessel can generally receive signals from two base stations. In these cases, the
traditional time of arrival (TOA) method is hard to use. This paper presents a method based on displacement vector correction to estimate a vessel position
by measuring signals from two AIS base stations. A vessel’s position and its clock bias are estimated
by the continuous range measurements in adjacent moments. The relationship of position information
between the adjacent moments can be derived by the displacement vector. A displacement vector is
calculated according to the heading and velocity provided by auxiliary sensors. Consequently, the
position information is obtained by solving a system of four equations with three unknowns. Utilizing
the existing devices already equipped in vessels as the auxiliary sensors, such as compasses, log
indicators, will not increase the cost of the vessel navigation system. Thus the proposed method can
maximize the reuse of resources to meeting IMO requirements for robust PNT information. The rest of the paper is organized as follows: Section 2 discusses the TOA positioning method in
AIS. Sensors 2014, 14 The novel positioning method using displacement correction is presented in Section 3, including
its mathematical model, implementation scheme and performance evaluation. In order to validate the
proposed method, several simulations are provided in Section 4. Finally, some conclusions are put
forth in Section 5. 1. Introduction Robust position, navigation and timing (PNT) information is an essential foundation of
e-navigation, developed by the International Maritime Organization (IMO), and intended to enhance
marine navigation. Though the global navigation satellite system (GNSS) is the primary navigation
system in maritime applications, an alternative position system to complement the existing GNSS may 2. TOA Positioning Method in AIS Generally speaking, position estimation can be determined using the TOA technique [14,15]. Figure 1
shows the geometrical principle of the TOA method. The vessel lies on circles with radii R1 and R2
centered on the base stations (B1 and B2), whose position can be obtained precisely. By measuring the
signals from at least two base stations, the position of the vessel can be determined by the intersection
point of the circles. Though there exist two solutions M and M′, M′ can be easily excluded according
to the approximate position of the vessel. 17378 Sensors 2014, 14 Figure 1. Geometrical principle of the TOA metohd. The latitude and longitude coordinates of the vessel are denoted by (φ,λ). For convenience, the
changes in latitude and longitude are denoted by the vertical and horizontal increments (Δφ,Δω). Then the
updated latitude and longitude coordinates of the vessel (φ′,λ′) and (Δφ,Δω) satisfy the following equations: sec
ϕ
ϕ
ϕ
λ
λ
ω
ϕ
′ =
+ Δ
′ =
+ Δ
(1) (1) The position equation can be modeled as the function of the range L = L(φ,ω), that is generally nonlinear: (
)
(
)
1
cos
sin
sin
cos
cos
cos
i
i
i
i
L
ϕ
ϕ
ϕ
ϕ
λ
λ
−
=
+
−
(2) (2) where a subscript i denotes the ith AIS base station. The linear position equation using Taylor-series
keeping only terms below second order is given by: ˆ
ˆ
ˆ
i
i
i
i
L
L
L
L
δϕ
δω
ϕ
ω
∂
∂
=
+
+
∂
∂
(3) (3) where
iL and ˆ
iL are the measured and estimated ranges from the ith base station to the vessel,
respectively. (δφ,δω) are the corrections to the estimated position vector in the vertical and horizontal
directions. The terms of partial derivative in Equation (3) can be calculated by the following formulas: ˆ
ˆ
cos
ˆ
ˆ
sin
i
i
i
i
L
L
γ
ϕ
γ
ω
∂
= −
∂
∂
= −
∂
(4) (4) where γi is the estimated azimuth angle directed from the estimation of the vessel’s position to the ith
base station. Sensors 2014, 14 Sensors 2014, 14 Sensors 2014, 14 17379 where δt is the clock bias between the vessel and AIS base stations and c is the velocity of light in the
free space. Thus the clock bias augments the two-dimensional position vector forming a three-dimensional
state vector. To estimate the vessel position, more than three independent TOA measurements from
different AIS base stations are required at the same time. However, the vessel can only receive signals
from two base stations under most conditions in the existing AIS setup. Therefore, the vessel position
estimation cannot be obtained using the traditional TOA positioning method. 3.1. A Mathematical Model At time k and time k + 1, the vessel locates at (φk, ωk) and (φk+1, ωk+1), respectively, which are to be
determined. According to Equation (6), the linearized position equation at time k is: ˆ
ˆ
ˆ
+
k
k
k
k
k
k
i
i
i
i
k
k
L
L
L
L
c t
δϕ
δω
δ
ϕ
ω
∂
∂
=
+
+
∂
∂
(7) (7) Here a superscript k is used to denote time k. If measurements from two different AIS base stations
are obtained, Equation (7) can be written in matrix form as: Here a superscript k is used to denote time k. If measurements from two different AIS base stations
are obtained, Equation (7) can be written in matrix form as: ˆ
ˆ
ˆ
ˆ
k
k
k
A
A
k
k
k
k
A
k
k
k
B
B
B
k
k
L
L
c
L
L
L
L
t
c
δϕ
ϕ
ω
δ
δω
δ
δ
ϕ
ω
∂
∂
∂
∂
=
∂
∂
∂
∂
(8) ˆ
ˆ
ˆ
ˆ
k
k
k
A
A
k
k
k
k
A
k
k
k
B
B
B
k
k
L
L
c
L
L
L
L
t
c
δϕ
ϕ
ω
δ
δω
δ
δ
ϕ
ω
∂
∂
∂
∂
=
∂
∂
∂
∂
(8) (8) where
ˆ
k
k
k
i
i
i
L
L
L
δ
=
−
. The subscript A and B correspond to the two AIS base station in Figure 1,
respectively. 2. TOA Positioning Method in AIS γi can be calculated as follows: (
)
(
)
1
2
cos
sin
ˆ
tan
cos sin
sin
cos
cos
i
i
i
i
i
ϕ
λ
λ
γ
ϕ
ϕ
ϕ
ϕ
λ
λ
−
−
=
−
−
(5) (5) The above conclusions can be obtained in the situation that time can be synchronized between the
vessel and AIS base stations. However this is a difficult task to achieve in reality. For AIS, Equation (3)
can be rewritten as:
ˆ
ˆ ˆ
ˆ
ˆ
+
i
i
i
i
L
L
L
L
c t
δϕ
δω
δ
ϕ
ω
∂
∂
=
+
+
∂
∂
(6) (6) 3.1. A Mathematical Model Similarly, the position matrix at time k + 1 is given by: 1
1
1
1
1
1
1
1
1
1
1
1
ˆ
ˆ
ˆ
ˆ
k
k
k
A
A
k
k
k
k
A
k
k
k
B
B
B
k
k
L
L
c
L
L
L
L
t
c
δϕ
ϕ
ω
δ
δω
δ
δ
ϕ
ω
+
+
+
+
+
+
+
+
+
+
+
+
∂
∂
∂
∂
=
∂
∂
∂
∂
(9) (9) where (δφk+1, δωk+1) refer to the correction values between the true and estimated position of the vessel
at time k + 1. There are five unknowns in four equations, thus it is insufficient to solve the position. The estimate of relative displacement vector ΔXk = (Δφk, Δωk) relative to the location at time k can
be calculated as follows: cos
sin
k
k
k
k
k
k
v
T
v
T
ϕ
α
ω
α
Δ
=
Δ
Δ
=
Δ
(10) (10) where v and α are vessel’s true velocity and heading, respectively; ΔT is a time interval between the
adjacent moments. The real-time vessel’s velocity and heading information can be obtained by
auxiliary sensors. 3.1. A Mathematical Model Thus, the positioning relationship between time k and time k + 1 is established using
the displacement vector according to Equation (11): Sensors 2014, 14 Sensors 2014, 14 17380 1
1
ˆ
ˆ
ˆ
ˆ
k
k
k
k
k
k
ϕ
ϕ
ϕ
ω
ω
ω
+
+
=
+ Δ
=
+ Δ
(11) (11) According to the vessel’s displacement vector between time k and k + 1, calculated by the auxiliary
information, the correction values in two adjacent moments are equal, that is: 1
1
k
k
k
k
δϕ
δϕ
δω
δω
+
+
=
=
(12) (12) Then the position matrix at time k + 1 can be rewritten as: Then the position matrix at time k + 1 can be rewritten as: Then the position matrix at time k + 1 can be rewritten as: 1
1
1
1
1
1
1
1
1
1
ˆ
ˆ
ˆ
ˆ
k
k
k
A
A
k
k
k
k
A
k
k
k
B
B
B
k
k
L
L
c
L
L
L
L
t
c
δϕ
ϕ
ω
δ
δω
δ
δ
ϕ
ω
+
+
+
+
+
+
+
+
+
+
∂
∂
∂
∂
=
∂
∂
∂
∂
(13) (13) Thus, the position at time k + 1 is the function of the position at time k, eliminating two unknown
variables from five unknown variables. 3.2. Implementation Scheme The implementation scheme of the proposed position estimation method based on displacement
correction is illustrated in Figure 2. The implementation scheme of the proposed position estimation method based on displacement
correction is illustrated in Figure 2. ure 2. Implementation scheme of position estimation based on displacement correction. Figure 2. Implementation scheme of position estimation based on displacement corre Firstly, signals from AIS base stations are received by the vessel’s receiver via an antenna. Then a
radio frequency (RF) front-end intercepts the incoming RF signal with 160 MHz and converts it to an
appropriate intermediate frequency (IF) for digitization, e.g., 455 KHz. The digital IF signals are
processed in the AIS position signal processor. It is organized into functionally identical channels, each
dynamically assigned to a different AIS base station. The signals are acquired, tracked and messages
are demodulated. S
dl
di
t
th AIS i
l t
ki
lt
TOA
t
d
d t
t th Firstly, signals from AIS base stations are received by the vessel’s receiver via an antenna. Then a
radio frequency (RF) front-end intercepts the incoming RF signal with 160 MHz and converts it to an
appropriate intermediate frequency (IF) for digitization, e.g., 455 KHz. The digital IF signals are
processed in the AIS position signal processor. It is organized into functionally identical channels, each
dynamically assigned to a different AIS base station. The signals are acquired, tracked and messages
are demodulated. Firstly, signals from AIS base stations are received by the vessel’s receiver via an antenna. Then a
radio frequency (RF) front-end intercepts the incoming RF signal with 160 MHz and converts it to an
appropriate intermediate frequency (IF) for digitization, e.g., 455 KHz. The digital IF signals are
processed in the AIS position signal processor. It is organized into functionally identical channels, each
dynamically assigned to a different AIS base station. The signals are acquired, tracked and messages
are demodulated. Secondly, according to the AIS signal tracking results, TOA measurements are produced to get the
propagation time between the vessel and different AIS base stations. Then TOA measurement is used
to calculate the range
iL , which is called pseudorange, since bias exists in the vessel’s clock. At the
same time, the exact position information of each AIS base station is extracted from the
demodulated messages. 3.1. A Mathematical Model The positioning matrix is the combination of positioning
matrices in the adjacent moments, that is: 1
1
1
1
1
1
1
1
1
1
ˆ
ˆ
ˆ
ˆ
=
ˆ
ˆ
ˆ
ˆ
k
k
A
A
k
k
k
k
k
k
A
B
B
k
k
k
k
B
k
k
k
A
A
A
k
k
k
B
k
k
B
B
k
k
L
L
c
L
L
L
c
L
L
L
L
t
c
L
L
L
c
ϕ
ω
δ
δϕ
ϕ
ω
δ
δω
δ
δ
δ
ϕ
ω
ϕ
ω
+
+
+
+
+
+
+
+
+
+
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
∂
(14) (14) Four position equations at two adjacent moments can be obtained according to measurements. Moreover, there are three unknown variables to solve four equations in Equation (14). The correction
values of the vertical and horizontal components and the clock bias can be calculated according to the
above position matrix by a least squares estimation approach. Then the vessel’s latitude and longitude
at time k can be obtained according to Equation (1). Remark 1: The combination of dead reckoning and range measurements is an existing approach
which has been used in many fields such as GPS-INS [16,17], robotics [18,19], vehicular ad-hoc
networks [20–22] and pedestrian localization in indoor environments [23,24]. Dead reckoning
determines a present position from a known past position. However pure dead reckoning methods are
prone to accumulated errors over time. Positioning methods based on range measurement have better
accuracy, but are sometimes hard to implement or use due to the time synchronization issue. Many
algorithms are used to combine dead reckoning and range measurements to improve the accuracy, such
as a Kalman filter, a particle filter or a Markov method [25,26]. It should be noted that both dead
reckoning and range measurements can estimate the position by themselves, as shown in the previous Sensors 2014, 14 17381 Sensors 2014, 14 work. The combination of two methods can improve the accuracy. 3.1. A Mathematical Model However, position estimation is
impossible when a vessel receives signals from only two AIS base stations. The proposed position
estimation method is investigated for this situation. 3.2. Implementation Scheme 3.2. Implementation Scheme Thirdly, the range ˆk
iL between the vessel and the different AIS base station at time k can be
estimated, since the initial position estimation of the vessel and the exact coordinates of the AIS base
stations are known. The initial position estimation of the vessel can use the vessel’s position
information at the last moment stored by the receiver or provided from external devices. Position
information of AIS base stations can be obtained by the demodulated messages. The measured and Sensors 2014, 14 Sensors 2014, 14 17382 estimated ranges are used to calculate the measurement prediction errors
k
iL
δ
at time k in the position
matrix as Equation (8). Then, the displacement vector (δφk+1, δωk+1) of the vessel is obtained from auxiliary sensors at time
k + 1 according to Equation (10). The vessel position (
)
1
1
ˆ
ˆ
,
k
k
ϕ
ω
+
+
at time k + 1 can be estimated
according to Equation (11). Thus the ranges
1
ˆk
iL + between the vessel and the base stations at time k + 1
can be calculated. These estimated ranges and the TOA measurements at time k + 1 are used to
construct the corresponding position equations as Equation (13). Finally, the vessel’s position and
clock bias can be solved by the combination of position equations at two adjacent moments according
to Equation (14). Sensors 2014, 14 17383 As the positioning error exists, the actual vessel position is in a circle with center at O and the radius of
ρ shown in Figure 3. That is to say an error boundary of the vessel position is indicated by a
63.2%~68.3% probability circle. As the positioning error exists, the actual vessel position is in a circle with center at O and the radius of
ρ shown in Figure 3. That is to say an error boundary of the vessel position is indicated by a
63.2%~68.3% probability circle. As already mentioned, α and v denote the vessel’s true heading and velocity, respectively. The
above analysis shows that the position errors are dependent on the measurement errors of the true
heading and voyage. Considering the influence of wind and flow, errors exist in the heading and the
velocity provided by the auxiliary sensors. Thus the error of vessel position caused by the wind and
flow may come down to the heading error and the voyage error caused by auxiliary sensors. Our error
analysis method is also applicable in this scenario. 3.3. Performance Analysis Positioning accuracy is one of the important technical parameters to evaluate position estimation
method [27,28]. The positioning error is influenced by the errors of the displacement vector, including
the heading error and the voyage error, as depicted in Figure 3. Figure 3. Positioning error influenced by heading and voyage error. As shown in Figure 3, αe is the heading error of the auxiliary sensor and SL is voyage determined by
the vessel’s velocity vK and the time interval △T. The positioning error caused by the heading error is
given by: 180
57.3
e
L
e
k
S
v
T
OA
α
π
α
Δ
=
=
(15) (15) The vessel voyage error is decided by the correction rate v△L of the auxiliary sensor and SL. The
positioning error caused by voyage error is: The vessel voyage error is decided by the correction rate v△L of the auxiliary sensor and SL. The
positioning error caused by voyage error is: L
L
L k
AB
v S
v v
T
Δ
Δ
=
=
Δ
(16) (16) Thus the positioning error ρ caused by the heading error and voyage error can be calculated by
Equation (17): Thus the positioning error ρ caused by the heading error and voyage error can be calculated by
Equation (17): 2
2
2
2
(
)
(
)
57 .3
e
L
L
L
S
OA
AB
v S
α
ρ
Δ
=
+
=
+
(17) (17) Remark 2: The vessel’s position error calculated by Equation (17) is always larger than the actual error
when the voyage is long, since the above conclusion is based on the condition that random errors are
superimposed on each other, but in an actual situation, random errors may cancel each other out, so the
actual positioning error may be less than the theoretically calculated error according to Equation (17). Remark 2: The vessel’s position error calculated by Equation (17) is always larger than the actual error
when the voyage is long, since the above conclusion is based on the condition that random errors are
superimposed on each other, but in an actual situation, random errors may cancel each other out, so the
actual positioning error may be less than the theoretically calculated error according to Equation (17). Sensors 2014, 14 Sensors 2014, 14 Sensors 2014, 14 17384 4.2. Performance Simulation This section presents simulations to demonstrate the proposed method and evaluate its performance. AIS base stations named Laotieshan and Huangbaizui are marked with yellow boxes in Figure 5. Table 1
gives information about these two AIS base stations, including the maritime mobile communications
service identity (MMSI), the latitude and longitude coordinate. In the simulation scenario, the vessel’s
trajectory is a parallelogram racetrack. The initial position of the vessel is at (38°34.414'N,
121°38.187'E) with the velocity of 20 knots (10.2888 m/s) and heading of 90 degrees. After straight
line motion for 500 s, the vessel will turn 45 degrees left with a centripetal acceleration of 0.2 m/s2. The vessel’s trajectory is indicated by the red solid line in Figure 5. Figure 5. Distribution of AIS base stations and the vessel’s trajectory. Table 1. Information of the AIS base stations and mobile locations. Figure 5. Distribution of AIS base stations and the vessel’s trajectory. Table 1. Information of the AIS base stations and mobile locations. Table 1. Information of the AIS base stations and mobile locations. Name
MMSI
Latitude
Longitude
Laotieshan
4131101
38°43.6420'N
121°08.1330'E
Huangbaizui 4131104
38°54.2850'N
121°42.9500'E Firstly, the proposed method is verified by assuming that the velocity and heading provided by
auxiliary sensors are accurate. The impact of auxiliary sensor errors will be discussed later in Section 4.3. The deviations between the true and estimated vessel’s position during the vessel’s movement are
compared in Figure 6, including the longitude, latitude and positioning error. Figure 6. Error curves of vessel dynamic accuracy. (a) Latitude error. (b) Longitude error. (c) Positioning error. 4.1. Signal Coverage Area of AIS Base Stations An AIS base station network has already been established by Chinese government, including a
national AIS management center located in Tianjin, management centers for the three sea regions
(North, South and East Sea), nineteen district management centers and nearly one hundred AIS base
stations. The coastal areas in China are all within coverage areas. Let us take the North Sea region as
an example. The positioning system using AIS is destined to be a regional radio navigation system
limited to the very high frequency (VHF) range. Typical coverage is 25 nautical miles offshore. Figure 4
is the signal coverage area of the AIS base station in the North Sea region. The overlay signal coverage
area of three AIS base stations is represented by the green “*”, while the overlay area of only two AIS
base stations is represented by the red “.”. It can be seen that it is common for vessels to receive
signals from two base stations in the actual AIS. Therefore, the position estimation proposed in the
paper is very practical. Figure 4. Signal coverage area of AIS base station in the North Sea region. Figure 4. Signal coverage area of AIS base station in the North Sea region. Figure 4. Signal coverage area of AIS base station in the North Sea region. Sensors 2014, 14 Figure 6. Error curves of vessel dynamic accuracy. (a) Latitude error. (b) Longitude error.
(c) Positioning error. 4.3. Auxiliary Sensor Error Simulation Figure 6. Error curves of vessel dynamic accuracy. (a) Latitude error. (b) Longitude error.
(c) Positioning error. (a)
(b)
0
200 400 600 800 1000120014001600180020002200
-0.4
-0.3
-0.2
-0.1
0
0.1
0.2
0.3
0.4
time (s)
deviation (cm)
0
200 400 600 800 1000120014001600180020002200
-0.4
-0.3
-0.2
-0.1
0
0.1
0.2
0.3
0.4
time (s)
deviation (cm) (b)
0
200 400 600 800 1000120014001600180020002200
-0.4
-0.3
-0.2
-0.1
0
0.1
0.2
0.3
0.4
time (s)
deviation (cm) (a)
0
200 400 600 800 1000120014001600180020002200
-0.4
-0.3
-0.2
-0.1
0
0.1
0.2
0.3
0.4
time (s)
deviation (cm) (b) (a) 17385 Sensors 2014, 14 Sensors 2014, 14 Figure 6. Cont. Figure 6. Cont. (c)
It can be seen from the simulation results that the standard deviation of the latitude error is 0.0541 cm,
the standard deviation of the longitude error is 0.0574 cm and the standard deviation of the positioning
error is 0.0698 cm. The estimated errors are all at the centimeter-level. The above simulation results
confirm the validity and feasibility of the proposed method. 0
200 400 600 800 1000120014001600180020002200
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
time (s)
deviation (cm) Figure 6. Cont. (c)
0
200 400 600 800 1000120014001600180020002200
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
time (s)
deviation (cm) (c)
0
200 400 600 800 1000120014001600180020002200
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
time (s)
deviation (cm) (c) It can be seen from the simulation results that the standard deviation of the latitude error is 0.0541 cm,
the standard deviation of the longitude error is 0.0574 cm and the standard deviation of the positioning
error is 0.0698 cm. The estimated errors are all at the centimeter-level. The above simulation results
confirm the validity and feasibility of the proposed method. It can be seen from the simulation results that the standard deviation of the latitude error is 0.0541 cm,
the standard deviation of the longitude error is 0.0574 cm and the standard deviation of the positioning
error is 0.0698 cm. The estimated errors are all at the centimeter-level. The above simulation results
confirm the validity and feasibility of the proposed method. It can be seen from the simulation results that the standard deviation of the latitude error is 0.0541 cm,
the standard deviation of the longitude error is 0.0574 cm and the standard deviation of the positioning
error is 0.0698 cm. The estimated errors are all at the centimeter-level. The above simulation results
confirm the validity and feasibility of the proposed method. Sensors 2014, 14 Sensors 2014, 14 17386 Positioning errors influenced by the heading error can be calculated by Equation (15). Positioning
errors change with the latitude of the vessel and the voyage as depicted in Figure 7. It’s shown that
positioning errors increase with the increasing latitude and voyage. A vessel’s voyage is determined by
its velocity and duration of time. We take a FURUNO DS-80 as a log indicator in the simulation. The
accuracy of the velocity is the greater one of 1.0 percent of the velocity and 0.1 knots [30]. In other
words, when the vessel’s velocity is greater than 10 knots, the accuracy of the log indicator is
1.0 percent of the velocity. Table 3 gives the velocity accuracy and error correction rate of the
log indicator for a maximum velocity of 5, 10 and 26 knots, respectively. The error correction rate is
equal to the accuracy divided by its corresponding velocity. Table 3. Velocity accuracy and error correction rate. Table 3. Velocity accuracy and error correction rate. Velocity (knots)
5
10
26
Accuracy (knots)
0 1
0 1
0 26 Table 3. Velocity accuracy and error correction rate. Velocity (knots)
5
10
26
Accuracy (knots)
0.1
0.1
0.26
Error correction rate
2.0%
1.0%
1.0%
When the time interval is set at 1, 5, 10 and 60 s, the voyages are shown in Table 4. When the time interval is set at 1, 5, 10 and 60 s, the voyages are shown in Table 4. Table 4. Voyage with different time intervals. Table 4. Voyage with different time intervals. Time Intervals (s)
Velocity (knots)
5
10
26
1
2.572222 m
5.144444 m
13.3755544 m
5
12.86111 m
25.72222 m
66.877772 m
10
25.72222 m
51.44444 m
133.755544 m
30
77.16666 m
154.33332 m
401.266632 m
60
154.33332 m
308.66664 m
802.533264 m Positioning errors ρ can be calculated by Equation (17), according to the heading error in Table 2,
the log correction rate in Table 3 and the voyage in Table 4. The positioning error ρ varying with the
velocity and time interval in the case of given heading error is shown below. Figure 8. Positioning error influenced by displacement vector. (a) Heading error is 0.4°. (b) Heading error of 0.5659°. (b) Heading error of 0.5659°. 4.3. Auxiliary Sensor Error Simulation Based on the above discussion, the proposed method provides centimeter-level positioning accuracy
with the assumption that the velocity and heading are accurate. However, this assumption is hard to
realize in engineering practice. The accurate displacement vector may not be obtained due to errors of
the auxiliary sensor. The effects of the auxiliary sensor errors on the positioning accuracy are
investigated in this section. The errors of auxiliary sensor including the heading and the voyage errors
are discussed in relation to their impact on positioning accuracy. The vessel’s heading is provided by the compass. Here we use an Anschutz Gyro Compass
Standard 22 in the simulation, whose static measuring error is within ±0.1° secφ and dynamic
measuring error is within ±0.4° secφ [29]. Thus heading errors can be calculated when the vessels are
on the equator, at latitude of 45° or at the poles by referring to Table 2. Table 2. Heading accuracy. Position
Equator
45°
Pole
Static error (°)
0.1000
0.1414
∞
Dynamic error (°)
0.4000
0.5659
∞
Figure 7. Positioning error influenced by heading error. 0
10 20 30 40 50 60
0
200
400
600
800
1000
0
2
4
6
8
10
12
latitude (°)
voyage (m)
positioning error (m) Table 2. Heading accuracy. Position
Equator
45°
Pole
Static error (°)
0.1000
0.1414
∞
Dynamic error (°)
0.4000
0.5659
∞
Figure 7. Positioning error influenced by heading error. Dynamic error (°)
0.4000
0.5659
∞
Figure 7. Positioning error influenced by heading error. 0
10 20 30 40 50 60
0
200
400
600
800
1000
0
2
4
6
8
10
12
latitude (°)
voyage (m)
positioning error (m) Figure 7. Positioning error influenced by heading error. 0
10 20 30 40 50 60
0
200
400
600
800
1000
0
2
4
6
8
10
12
latitude (°)
voyage (m)
positioning error (m) 5. Conclusions As one of the widely used land-based communication systems, AIS can also provide range-mode
positioning service to promote the robustness of the traditional navigation systems equipped on vessels. However, for the situations when the received signals using the usual TOA positioning method are not
enough, the auxiliary sensors on the vessel provide additional motion information that can help to
estimate the vessel’s position. A new position estimation method based on displacement correction in
AIS is proposed to determine the vessel’s position when the signals can be received from only two AIS
base stations. The mathematical analysis and simulation results show that the positioning accuracy of
the new method is dependent on the accuracy of the sensors. Based on our implementation scheme, our
future work will focus on the realization in a mobile AIS device. This will lay the foundation for the
demonstration project using the new ranging-mode of AIS established in the coastal areas of China. Sensors 2014, 14 5
10
15
20
25
30
0
10
20
30
40
50
60
0
2
4
6
8
10
12
velocity (knot)
time interval (s)
positioning error (m) (a)
(b)
5
10
15
20
25
30
0
10
20
30
40
50
60
0
2
4
6
8
10
velocity (knot)
time interval (s)
positioning error (m)
5
10
15
20
25
30
0
10
20
30
40
50
60
0
2
4
6
8
10
12
velocity (knot)
time interval (s)
positioning error (m) (a)
5
10
15
20
25
30
0
10
20
30
40
50
60
0
2
4
6
8
10
velocity (knot)
time interval (s)
positioning error (m) (b) (a) Sensors 2014, 14 17387 The velocity varies from 5 to 26 knots and the time interval is set from 1 s to 60 s. The positioning
error when the vessel is on the equator is shown in Figure 8a. Figure 8b shows the positioning error
when the vessel locates at latitude 45°. It can be seen from the simulation results that the positioning
error ρ is on the order of magnitude of 10 m using common vessel devices such as the Anschutz Gyro
Compass Standard 22 and FURUNO DS-80 as auxiliary sensors. Acknowledgments This research was partially supported by the Chinese National Science Foundation (No. 61231006),
the National Key Technologies R&D Program of China (No. 2012BAH36B01), the Fundamental
Research Funds for the Central Universities (No. 3132014329) and the Liaoning Provincial Natural
Science Foundation (No. 2014025002). Author Contributions Yi Jiang provided the idea of algorithms, carried out the simulations and drafted the manuscript. Shufang Zhang supervised the work, arranged the architecture and contributed to the writing of the
manuscript. Dongkai Yang supervised the work and provided major direction of the research. Conflicts of Interest The authors declare no conflict of interest. The authors declare no conflict of interest. References 1. Ward, N.; Shaw, G.; Williams, P.; Grant, A. The role of GNSS in e-navigation and the need for
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distributed under the terms and conditions of the Creative Commons Attribution license
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ProductDocuments/DS80%20Operator's%20Manual%20M1%20%202-12-03.pdf (accessed on 27
August 2014). © 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/3.0/). © 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/3.0/).
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Scope and Sequence English Class 02
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Document information: Document information: Document information: Document information: Leh Wi Learn (2018). "Scope and Sequence English Class 08." A resource pro-
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https://creativecommons.org/licenses/by/4.0/. Uploaded by the EdTech Hub, https://edtechhub.org. For more information, see https://edtechhub.org/oer. Archived on Zenodo: April 2020. DOI: 10.5281/zenodo.3745232 Please attribute this document as follows: Leh Wi Learn (2018). "Scope and Sequence English Class 08." A resource pro-
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Responses to Water Deficit and Salt Stress in Silver Fir (Abies alba Mill.) Seedlings
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Responses to Water Deficit and Salt Stress in Silve
Fir (Abies alba Mill.) Seedlings Irina Maria Todea (Morar) 1, Sara González-Orenga 2, Monica Boscaiu 2, Mariola Plazas 3,
Adriana F. Sestras 1,*
, Jaime Prohens 3
, Oscar Vicente 3
and Radu E. Sestras 1 rina Maria Todea (Morar) 1, Sara González-Orenga 2, Monica Boscaiu 2, Mariola Plazas 3,
Adriana F. Sestras 1,*
, Jaime Prohens 3
, Oscar Vicente 3
and Radu E. Sestras 1 1
Faculty of Horticulture, University of Agricultural Sciences and Veterinary Medicine, 400372 Cluj-Napoca
Romania; irina.todea@usamvcluj.ro (I.M.T.); rsestras@usamvcluj.ro (R.E.S.) 1
Faculty of Horticulture, University of Agricultural Sciences and Veterinary Medicine, 400372 Cluj-Napoca,
Romania; irina.todea@usamvcluj.ro (I.M.T.); rsestras@usamvcluj.ro (R.E.S.)
2
Mediterranean Agroforestry Institute (IAM), Universitat Politècnica de València, Camino de Vera s/n,
46022 Valencia, Spain; sagonor@doctor.upv.es (S.G.-O.); mobosnea@eaf.upv.es (M.B.)
3
Institute for Conservation and Improvement of Valencian Agrodiversity (COMAV), Universitat Politècnica
de València, Camino de Vera s/n, 46022 Valencia, Spain; maplaav@btc.upv.es (M.P.);
jprohens@btc.upv.es (J.P.); ovicente@upvnet.upv.es (O.V.)
*
Correspondence: adriana.sestras@usamvcluj.ro ;
j
(
);
j
(
)
2
Mediterranean Agroforestry Institute (IAM), Universitat Politècnica de València, Camino de Vera s/n,
46022 Valencia, Spain; sagonor@doctor.upv.es (S.G.-O.); mobosnea@eaf.upv.es (M.B.) 2
Mediterranean Agroforestry Institute (IAM), Universitat Politècnica de València, Camino de Vera s/n,
46022 Valencia, Spain; sagonor@doctor.upv.es (S.G.-O.); mobosnea@eaf.upv.es (M.B.)
3
Institute for Conservation and Improvement of Valencian Agrodiversity (COMAV), Universitat Politècnica
de València, Camino de Vera s/n, 46022 Valencia, Spain; maplaav@btc.upv.es (M.P.);
jprohens@btc.upv.es (J.P.); ovicente@upvnet.upv.es (O.V.)
d
d
l 46022 Valencia, Spain; sagonor@doctor.upv.es (S.G.-O.); mobosnea@eaf.upv.es (M.B.)
3
Institute for Conservation and Improvement of Valencian Agrodiversity (COMAV), Universitat Politècnica
de València, Camino de Vera s/n, 46022 Valencia, Spain; maplaav@btc.upv.es (M.P.);
jprohens@btc.upv.es (J.P.); ovicente@upvnet.upv.es (O.V.) , p
;
g
p
(
);
p
(
)
3
Institute for Conservation and Improvement of Valencian Agrodiversity (COMAV), Universitat Politècnica
de València, Camino de Vera s/n, 46022 Valencia, Spain; maplaav@btc.upv.es (M.P.);
jprohens@btc.upv.es (J.P.); ovicente@upvnet.upv.es (O.V.) *
Correspondence: adriana.sestras@usamvcluj.ro
Received: 4 March 2020; Accepted: 30 March 2020; Published: 2 April 2020 Abstract: Forest ecosystems are frequently exposed to abiotic stress, which adversely affects their
growth, resistance and survival. For silver fir (Abies alba), the physiological and biochemical responses
to water and salt stress have not been extensively studied. Responses of one-year-old seedlings to a
30-day water stress (withholding irrigation) or salt stress (100, 200 and 300 mM NaCl) treatments were
analysed by determining stress-induced changes in growth parameters and different biochemical
markers: accumulation of ions, different osmolytes and malondialdehyde (MDA, an oxidative
stress biomarker), in the seedlings, and activation of enzymatic and non-enzymatic antioxidant
systems. Both salt and water stress caused growth inhibition. The results obtained indicated that
the most relevant responses to drought are based on the accumulation of soluble carbohydrates
as osmolytes/osmoprotectants. Responses to high salinity, on the other hand, include the active
transport of Na+, Cl−and Ca2+ to the needles, the maintenance of relatively high K+/Na+ ratios and
the accumulation of proline and soluble sugars for osmotic balance. Interestingly, relatively high Na+
concentrations were measured in the needles of A. alba seedlings at low external salinity, suggesting
that Na+ can contribute to osmotic adjustment as a ‘cheap’ osmoticum, and its accumulation may
represent a constitutive mechanism of defence against stress. These responses appear to be efficient
enough to avoid the generation of high levels of oxidative stress, in agreement with the small increase
in MDA contents and the relatively weak activation of the tested antioxidant systems. Keywords: abiotic stress; antioxidants; drought; ion homeostasis; osmolytes; salinity; silver fir 1. Introduction Silver fir ecotypes also
show remarkable variation in features such as shade tolerance, frost hardiness and drought resilience. It is expected that climate change will contribute to reduce the abundance and distribution range of
this species; therefore, improvements in forest management programmes are recommended, as they
will be required for the conservation of silver fir [15]. Water stress, which will increase under climate
change conditions, as mentioned above, is actually one of the factors most damaging for the success of
reforestation actions. On the other hand, conifers are known to have a relatively high sensitivity to
salt [16]. Consequently, silver fir—as other conifers—does not grow naturally in saline environments
but can be affected by relatively high salt concentrations in stands near mountains roads, due to the
common practice in many European countries of road de-icing in winter, using large amounts of
NaCl [17,18]. Despite the interest of these studies, the information available regarding the responses
of this species to drought and salinity is still limited. There are several published reports dealing
with the effects of drought on silver fir, mostly describing fieldwork with adult tress (see, for example
References [19–21]), but we have not found publications addressing the physiological and biochemical
mechanisms of response to stress in Abies alba seedlings, except for our preliminary study [22]. Regardless of their degree of tolerance, plants react against environmental stressors by activating
a series of conserved responses. These common responses encompass various physiological and
biochemical processes, at the cell and whole plant levels, in specific environmental contexts [23,
24]. These general responses include the inhibition of photosynthesis and plant growth [25,26],
the accumulation of different osmolytes, the activation of antioxidant systems and the expression of
specific defence proteins [3,27–30]. Besides, certain stress conditions trigger specific responses, such as
the control of ion transport and ion homeostasis in the presence of high salt concentrations [31], or the
cold-induced synthesis of antifreeze proteins [32]. Osmolytes are of particular importance since, in addition to contributing to cellular
osmotic adjustment under stress, they have essential roles as “osmoprotectants” [33], acting as
low-molecular-weight chaperones in the direct stabilisation of membranes, proteins and other
macromolecular structures under conditions of cellular dehydration, and as signalling molecules [34–36]. 1. Introduction Drought and soil salinity are considered the most adverse and critical environmental factors
for plants, causing massive losses in agricultural production worldwide and, at the same time,
substantially affecting the distribution of wild species in nature [1,2]. Drought and salinity affect
more than 10 percent of total arable land, and desertification and salinization are rapidly spreading
globally [3–5]. Accumulation of salts dissolved in irrigation water leads to the progressive ‘secondary’
salinization of irrigated cropland, especially in arid and semiarid regions, and this problem will worsen
in the near future due to the effects of the present climate change [6]. Plants can perceive abiotic stresses, such as water deficit and salt stress, and activate
appropriate physiological, biochemical and molecular responses, with altered metabolism, growth Forests 2020, 11, 395; doi:10.3390/f11040395 www.mdpi.com/journal/forests www.mdpi.com/journal/forests 2 of 21 Forests 2020, 11, 395 and development [7]. The study of these responses and the mechanisms of tolerance to drought and
salinity is currently one of the major topics of research in plant biology. and development [7]. The study of these responses and the mechanisms of tolerance to drought and
salinity is currently one of the major topics of research in plant biology. Silver fir (A. alba Mill.) is an economically and ecologically important forest tree species, especially
in lower-mountain forests. This species covers the main mountain areas in central and southern
Europe, including populations in the Mediterranean climatic zone, in the south of Italy (Calabria),
in the northeast of Spain (Catalonia) and the south of France (south of the great Alps and Pyrenees) [8]. In Pyrenees and Calabria, the populations of A. alba belong to particular ecotypes, differentiated
genetically from other populations [9,10]. Abies alba is generally associated with spruce Picea abies (L.)
H. Karst. in the upper mountain belt, and with beech Fagus sylvatica (L.) in lower and middle mountain
belts. Moreover, this species forms pure forests in the subalpine belt in some areas of the Southern
Alps [11]. A. alba has a high-altitude range, between 500 and 2000 m a.s.l., in the pre-Alps and southern
French Alps [12,13]. Regarding environmental requirements, silver fir is tolerant to a wide variety of soil types
with different nutrient content and alkalinity conditions, except hydromorphic and compact soils. This species prefers soils deep and moist with a pH from acid to neutral [14]. 1. Introduction A general reaction to abiotic stresses, which causes oxidative stress through the generation of
‘reactive oxygen species’ (ROS), is the activation of antioxidant systems to prevent or reduce
oxidative damage of proteins, membranes and DNA [37]. These antioxidant activities include
several enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX)
(and other peroxidases), monodehydroascorbate reductase (MDAR), or glutathione reductase (GR),
among others [38]. Non-enzymatic antioxidant compounds, including vitamins C and E, carotenoids,
glutathione, or phenolic compounds, especially the subclass of flavonoids, may also play an essential
role in maintaining cellular redox balance under stress conditions [39]. Forests 2020, 11, 395 3 of 21 Considering that the identification and evaluation of reliable biochemical stress markers will
significantly contribute to elucidate the mechanisms of tolerance to drought and salinity, this work
aimed to select the optimal indicators associated with silver fir’s general responses to salt and water
stress, at the seedling stage. Improving our knowledge on the mechanisms of stress tolerance of silver
fir, apart from its academic interest, may help to design and implement more efficient conservation
programmes for this economically and ecologically important species, allowing an appropriate use of
reproductive seed material on reforestation sites. For this study, A. alba seedlings were subjected to water
deficit (withholding irrigation) and salt stress (watering the plants with increasing salt concentrations)
treatments, under controlled greenhouse conditions. After the treatments, the following variables
were determined in control and stressed plants: growth parameters, levels of photosynthetic pigments,
ions concentrations in needles and roots, the leaf contents of common osmolytes, the degree of oxidative
stress, by quantification of malondialdehyde (MDA, a reliable oxidative stress marker), total phenolic
compounds and flavonoid contents, as representative non-enzymatic antioxidants, and the specific
activities of major antioxidant enzymes. 2.2. Substrate Analysis The electrical conductivity of the substrate was measured after the treatments. Soil samples were
collected from every pot, air-dried and passed through a 2 millimetre sieve. A soil:water suspension (1:5)
was prepared in deionised water, mixed at 600 rpm for one hour at room temperature and then filtered
through filter paper. Electrical conductivity was measured with a Crison 522 Conductivity-meter and
expressed in dS m−1. The gravimetric method was used to determine soil moisture, as follows: a fraction of each soil
sample was weighed (soil weight, SW), dried in an oven at 105 ◦C until constant weight and then
weighed again (dry soil weight, DSW). The soil water content was calculated as: Soil humidity (%) = [(SW −DSW)/SW] × 100 Soil humidity (%) = [(SW −DSW)/SW] × 100 2.1. Plant Growth and Stress Treatments One-year-old seedlings of silver fir from the Romanian Carpathians Mountains (Gârda Seacă
nursery, 46◦31′ N/22◦46′ E) were transferred to the greenhouse of the Institute for the Preservation and
Improvement of Valencian Agrodiversity (COMAV), Universitat Politècnica de València, Valencia, Spain. After one week of acclimatisation, 175 seedlings, at approximately the same developmental stage, were
selected, transplanted into 0.3 L individual pots containing ‘Humin-substrat N3’ (Klasmann-Deilmann,
Germany) substrate, and randomly distributed into five groups of 35 seedlings, each group of pots
placed within a plastic tray. The pots were maintained in a greenhouse with controlled temperature
(minimum of 15 ◦C and maximum of 30 ◦C) under natural light and watered twice a week with
tap water. Salt and water stress treatments started 21 days later. Salt treatments were applied by
watering the plants twice weekly with NaCl solutions of 0 (for the controls), 100, 200 or 300 mM final
concentration (in tap water), adding 1 L of solution per tray. The water stress (WS) treatment was
performed by altogether withholding irrigation. Treatments were stopped after 30 days, before any
seedling mortality was observed. Plant samples (needles and roots) were collected separately for the
measurement of growth parameters and biochemical analyses. Seven replicates, each one consisting of
a pooled sample of five seedlings, were used per treatment. 2.6. Osmolyte Quantification Two main types of osmolytes were analysed in silver fir needles, proline (Pro) and total soluble
sugars (TTS). Pro content was measured by the ninhydrin-acetic acid method [42]. Briefly, needle
extracts were prepared from fresh plant material in a 3% (w/v) sulfosalicylic acid solution, mixed
with acid ninhydrin, incubated for one hour at 95 ◦C in a water bath, cooled and extracted with
toluene. The absorbance of the organic phase was measured at 520 nm, using toluene as the blank. Pro
concentration was calculated from a standard curve, prepared with known amounts of the osmolyte,
and expressed in µmol g−1 DW. TSS contents were measured according to a published procedure [43]. Needle fresh material
(0.05–0.10 g) was ground in the presence of liquid N2 and suspended in 3 mL of 80% (v/v) methanol. The samples were vortexed, centrifuged at 12,000 rpm for 10 min, the supernatants were collected, and a
fraction of each extract was diluted 10-fold with water. The diluted sample (0.5 mL) was supplemented
with concentrated sulphuric acid (2.5 mL) and 5% phenol (0.5 mL). Finally, the absorbance of the
sample was measured at 490 nm. TSS contents were expressed as ‘mg equivalent of glucose’ (used as
standard), mg eq. glucose g−1 DW. 2.3. Plant Growth Parameters Before starting the stress treatments (time 0), the number of needles and the stem length were
determined for all A. alba seedlings. To analyse the effects of water and salt stress on A. alba, at the stage
of vegetative growth, the increases in the number of needles (Nno) and stem length (SL) with respect 4 of 21 Forests 2020, 11, 395 to the values measured at time 0, and the fresh weight (FW) of needles, were determined. Part of
the needle material was weighed (FW), dried at 65 ◦C until constant weight, and weighed again (dry
weight, DW), and the water content percentage (WC%) of needles was calculated as: WC% = [(FW −DW)/FW] × 100 2.5. Ion Contents Sodium (Na+), potassium (K+), calcium (Ca2+) and chloride (Cl−) ions were determined in roots
and needles of all replicates, after four-week treatments, according to Weimberg [41]. Dried plant
material (0.05–0.10 g) was ground to a fine powder and extracted in 15 mL of Milli-Q water, by
heating the samples for 1 h in a boiling water bath, followed by cooling on ice and filtration through
a nylon filter of 0.45 µm pore. Na+, Ca2+ and K+ were quantified with a PFP7 flame photometer
(Jenway Inc., Staffordshire, UK) and Cl−was measured with a chloride analyser (Sherwood, model
926, Cambridge, UK). 2.4. Photosynthetic Pigments Chlorophyll a (Chl a), chlorophyll b (Chl b) and total carotenoids (Caro) were determined following
a previously described method [40]. Fresh needle material (0.05–0.10 g) was ground in the presence of
liquid nitrogen. One ml of ice-cold 80% acetone was added to the sample, which was shaken overnight
in the dark, at 4 ◦C. Following a 10 min centrifugation at 12,000 rpm, at 4 ◦C, the supernatants were
collected, and the absorbance was measured at 470, 645 and 663 nm. The following equations were
used for the calculation of pigment concentrations [40]: Chl a (µg/mL) = 12.21 × (A663) −2.81 × (A646);
Chl b (µg/mL) = 20.13 × (A646) −5.03 × (A663);
Caro (µg/mL) = (1000 × A470 −3.27 × [Chl a] −104 × [Chl b])/227 Chl a (µg/mL) = 12.21 × (A663) −2.81 × (A646);
Chl b (µg/mL) = 20.13 × (A646) −5.03 × (A663); Caro (µg/mL) = (1000 × A470 −3.27 × [Chl a] −104 × [Chl b])/227 Chlorophyll and carotenoid contents were finally expressed in mg g−1 DW. 2.5. Ion Contents 2.7. Malondialdehyde (MDA) Methanol extracts (80%, v/v, in water) were prepared by grinding 0.05–0.10 g of fresh needles,
shaking the samples in a rocker shaker overnight, at room temperature, followed by centrifugation
at 12,000 rpm for 15 min. MDA was quantified in the supernatants, as previously described [44]. 5 of 21 Forests 2020, 11, 395 Each sample was mixed with 0.5% thiobarbituric acid (TBA) prepared in 20% trichloroacetic acid
(TCA)—or with 20% TCA without TBA for the controls—and then incubated at 95 ◦C for 15 min, in
a water bath. The reactions were stopped on ice, the samples were centrifuged at 12,000 rpm for 10
min, at 4 ◦C, and the absorbance of the supernatants was measured at 532 nm. After subtracting
the non-specific absorbance at 600 and 440 nm, the MDA concentration was calculated applying the
equations described by Hodges [44] based on the molar extinction coefficient of the MDA-TBA adduct
at 532 nm (ε 532 = 155 mM−1 cm−1). 2.8. Non-Enzymatic Antioxidants Concentrations of total phenolic compounds (TPC) and total flavonoids (TF) were determined in
the same 80% methanol extracts used for MDA measurements. TPC were determined according to
the protocol of Blainski [45], which is based on the reaction with the Folin–Ciocalteu reagent, in the
presence of NaHCO3. The reaction mixtures were incubated at room temperature, in the dark, for 90
min, and the absorbance was then recorded at 765 nm. TPC concentration was expressed as equivalents
of the standard, gallic acid (mg eq. GA g−1 DW). TF were determined by nitration of catechol groups with NaNO2, followed by reaction with AlCl3
under alkaline conditions [46]. The absorbance of the samples was read at 510 nm, using catechin as
the standard. TF concentration was expressed as equivalents of catechin (mg eq. C g−1 DW). 2.9. Antioxidant Enzyme Activities 2.9. Antioxidant Enzyme Activities The specific activity of four major antioxidant enzymes, namely, superoxide dismutase (SOD),
catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR), was determined at room
temperature (~25 ◦C) in crude protein extracts prepared from silver fir needles, essentially as previously
described [47]. The plant material was ground in liquid N2 and mixed with extraction buffer (20 mM
Hepes, pH = 7.5, 50 mM KCl, 1 mM EDTA, 0.1% (v/v) Triton X-100, 0.2% (w/v) polyvinylpyrrolidone,
0.2% (w/v) polyvinylpolypyrrolidone and 5% (v/v) glycerol). To improve protein extraction, 1/10
volume of ‘high salt buffer’ (225 mM Hepes, pH 7.5, 1.5 M KCl and 22.5 mM MgCl2) was added to
the samples, mixed well by vortexing and kept for 15 min on ice. After centrifugation at 13,500 rpm
for 15 min at 4 ◦C, the supernatants were collected, concentrated in U-Tube concentrators (Novagen,
Madison, WI, USA), and centrifuged again to remove precipitated material. The supernatants, referred
to as ‘protein extracts’, were frozen in liquid N2 and stored in aliquots at −75 ◦C. Protein concentration
in the extracts was measured by the method of Bradford [48], using the Bio-Rad reagent and bovine
serum albumin (BSA) as standard. SOD activity in the protein extracts was determined following, at 560 nm, the inhibition of
nitroblue tetrazolium (NBT) photoreduction in reaction mixtures containing riboflavin as the source of
superoxide radicals [49]. A SOD unit was defined as the amount of enzyme that causes 50% inhibition
of NBT photoreduction under the assay conditions. CAT activity was assessed by the decrease in absorbance at 240 nm, which parallels the consumption
of H2O2 added to the extracts [50]. A CAT unit was defined as the amount of enzyme that will decompose
one mmol of H2O2 per minute at 25 ◦C. For ascorbate peroxidase (APX), the enzyme activity was determined by the decrease in absorbance
observed at 290 nm as ascorbate becomes oxidised in the reaction [51]. One APX unit was defined as
the amount of enzyme required to consume one mmol of ascorbate per minute, at 25 ◦C. GR activity was quantified according to Reference [52], following the decrease in absorbance at
340 nm due to oxidation of NADPH—the cofactor of the GR-catalysed reduction of oxidised glutathione
(GSSG). One GR unit was defined as the amount of enzyme that will oxidise one mmol of NADPH per
minute, at 25 ◦C. 2.9. Antioxidant Enzyme Activities GR activity was quantified according to Reference [52], following the decrease in absorbance at
340 nm due to oxidation of NADPH—the cofactor of the GR-catalysed reduction of oxidised glutathione
(GSSG). One GR unit was defined as the amount of enzyme that will oxidise one mmol of NADPH per
minute, at 25 ◦C. Forests 2020, 11, 395 6 of 21 3.1. Substrate Analysis The electric conductivity (EC1:5) of the substrate increased in parallel to the increasing NaCl
concentration in the irrigation solutions, reaching almost an eight-fold higher value in the presence of
300 mM NaCl than in the control pots, confirming the high correlation between EC and the concentration
of the saline solutions used in the salt treatments. The water stress treatment also led to a significant
increase in the substrate EC, probably due to concentration of salts in the soil. However, logically,
it was lower than in the pots watered with NaCl solutions, even at the lowest concentration tested
(100 mM). As expected, the humidity of the substrate was strongly reduced, by more than 40%, in the
pots that were not irrigated (WS treatment), whereas those watered with saline solutions showed only
slight reductions, as compared to the control (Table 1). Table 1. Electric conductivity (EC) of the substrate, in a 1:5 soil suspension (EC1:5), and soil humidity
(SH) on the pots of A. alba seedlings after 30 days of water deficit (WS) and salt stress (100, 200 and 300
mM NaCl) treatments 1. Table 1. Electric conductivity (EC) of the substrate, in a 1:5 soil suspension (EC1:5), and soil humidity
(SH) on the pots of A. alba seedlings after 30 days of water deficit (WS) and salt stress (100, 200 and 300
mM NaCl) treatments 1. Soil Parameter
Control
WS
100 mM NaCl
200 mM NaCl
300 mM NaCl
EC1:5 (dS m−1)
2.70 ± 0.08 a
5.08 ± 0.63 b
9.41 ± 0.51 c
19.04 ± 0.57 d
20.66 ± 0.54 e
(SH) (%)
62.14 ± 0.90 c
35.53 ± 2.34 a
59.50 ± 0.30 b
58.48 ± 0.40 b
59.07 ± 0.88 b
1 Values shown are means ± SE (n = 7). Different letters in each row indicate significant differences between
treatments, according to Tukey’s test (α = 0.05). 2.10. Statistical Analyses Data were analysed using the program Statgraphics Centurion XVI (Statgraphics Technologies,
The Plains, VA, USA). Significant differences between treatments were tested by one-way analysis of
variance (ANOVA) at the 95% confidence level, and post hoc comparisons were made using the Tukey’s
HSD test at p < 0.05. All mean values throughout the text are based on seven biological replicates, each
one corresponding to a composite sample of five pooled individual seedlings. Hierarchical cluster
analysis (HCA) and the corresponding heatmap were performed using the ClustVis tool [53], for traits
for which significant differences (p < 0.05) had been detected in the ANOVA with the purpose to find
a signature of responses specific to the water and salinity stresses applied. Unit variance scaling for
the normalised and centred data was used. Distance measures for the HCA were based on Pearson
correlations, and the average clustering method with the higher median value first for the tree ordering
option was used. 3.2. Plant Growth Analysis Salt and water stress inhibited the growth of A. alba seedlings, as shown by the relative reductions
in stem elongation, increase of the number of needles and biomass accumulation in the stressed plants,
as compared to the non-stressed controls. For example, all stress treatments strongly reduced the mean
stem elongation measured in the control seedlings, but no statistically significant (p < 0.05) differences
were observed between the water deficit treatment and the three different NaCl concentrations
(Figure 1a). In terms of the increment in the number of needles (Figure 1b) or the needles’ FW
(Figure 1c), growth inhibition of the salt-treated seedlings was concentration-dependent, with the
strongest effects observed at the highest NaCl concentration tested. Under our experimental conditions,
the water deficit treatment resulted in inhibition of growth similar to that caused by the lowest salinity
applied to the A. alba seedlings. Increasing external salinity led to a significant, concentration-dependent
reduction in the needles water content, down to about 45% in the presence of 300 mM NaCl (Figure 1d). Nevertheless, the stress-induced dehydration of the needles was much more pronounced in the plants
subjected to water deficit, where water content dropped to 13% (Figure 1d). Therefore, the observed 7 of 21 Forests 2020, 11, 395 reduction of fresh weight in water-stressed seedlings is probably due, to a large extent, to loss of water
by the needles. Forests 2020, 11, x FOR PEER REVIEW
7 of 20 Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation
(a), increment in the number of needles (Nno) (b), needle fresh weight (FW) (c), needle water content
(WC%) (d) of one-year-old A. alba seedlings after 30 days of growth in the presence of the indicated
salt concentrations or subjected to water deficit (completely withholding irrigation) (WS). Stem length
and number of needles’ measurements were taken just before starting the treatments (time 0), and
before collecting the samples (time 30). Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation
(a), increment in the number of needles (Nno) (b), needle fresh weight (FW) (c), needle water content
(WC%) (d) of one-year-old A. 3 3 Photosynthetic Pigments
3.3. Photosynthetic Pigments o o y
e i
ig e
Photosynthetic pigments’ (chlorophylls a and b, carotenoids) concentrations decreased under
stress. Seedlings subjected to either water deficit or salinity, showed lower chlorophylls contents than
the non-stressed controls, and the differences were statistically significant in all cases, except for Chl
a in plants watered with 100 mM NaCl. The strongest relative reductions were observed in water-
stressed seedlings (Figure 2). Needle levels of carotenoids were low and, in general, did not show
significant differences between treatments except for the increase observed in response to water
Photosynthetic pigments’ (chlorophylls a and b, carotenoids) concentrations decreased under
stress. Seedlings subjected to either water deficit or salinity, showed lower chlorophylls contents than
the non-stressed controls, and the differences were statistically significant in all cases, except for Chl a in
plants watered with 100 mM NaCl. The strongest relative reductions were observed in water-stressed
seedlings (Figure 2). Needle levels of carotenoids were low and, in general, did not show significant
differences between treatments, except for the increase observed in response to water stress (Figure 2). 3.2. Plant Growth Analysis alba seedlings after 30 days of growth in the presence of the indicated salt
concentrations or subjected to water deficit (completely withholding irrigation) (WS). Stem length and
number of needles’ measurements were taken just before starting the treatments (time 0), and before
collecting the samples (time 30). Bars represent means ± SE (n = 7). Different letters above the bars
indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation
Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation
(a), increment in the number of needles (Nno) (b), needle fresh weight (FW) (c), needle water content
(WC%) (d) of one-year-old A. alba seedlings after 30 days of growth in the presence of the indicated
salt concentrations or subjected to water deficit (completely withholding irrigation) (WS). Stem length
and number of needles’ measurements were taken just before starting the treatments (time 0), and
before collecting the samples (time 30). Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 1. Effect of water and salt stress treatments on growth inhibition in A. alba. Stem elongation
(a), increment in the number of needles (Nno) (b), needle fresh weight (FW) (c), needle water content
(WC%) (d) of one-year-old A. alba seedlings after 30 days of growth in the presence of the indicated salt
concentrations or subjected to water deficit (completely withholding irrigation) (WS). Stem length and
number of needles’ measurements were taken just before starting the treatments (time 0), and before
collecting the samples (time 30). Bars represent means ± SE (n = 7). Different letters above the bars
indicate significant differences between treatments, according to Tukey’s test (α = 0.05). stress (Figure 2)
3.4. Ions Levels It is inte
he high Na+ concentration (about 400 µmol g−1 DW) measured in needles of control
sence of salt (Figure 4a). Ca2+ and K+ levels were also higher in the needles than in the roots, in the controls a
ed treatments (Figure 3c, d and Figure 4c, d). Mean K+ contents generally decreased in
f
d
dl
d
l
d Figure 2. Photosynthetic pigments levels under water and salt stress. Needle concentrations of
chlorophyll a (Chl a), chlorophyll b (Chl b) and total carotenoids (Caro) in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated salt concentrations or subjected to
water deficit (completely withholding irrigation) (WS). Bars represent means ± SE (n = 7). For each of
the three pigments, different letters above the bars indicate significant differences between
treatments, according to Tukey’s test (α = 0.05). Figure 2. Photosynthetic pigments levels under water and salt stress. Needle concentrations of
chlorophyll a (Chl a), chlorophyll b (Chl b) and total carotenoids (Caro) in one-year-old A. alba seedlings
after 30 days of growth in the presence of the indicated salt concentrations or subjected to water deficit
(completely withholding irrigation) (WS). Bars represent means ± SE (n = 7). For each of the three
pigments, different letters above the bars indicate significant differences between treatments, according
to Tukey’s test (α = 0.05). e
o
a e
e
e
ee
i g , a
i
e po
e o
a
e
i
a
o
e
a io
epe
nner. Still, the differences with the non-stressed controls were statistically significant only in ro
in the presence of 200 or 300 mM NaCl (Figures 3c and 4c). As compared to the controls, me
+ concentrations increased in parallel to external salinity, both in roots and in needles, b
nificant differences were found only in needles. This effect was not observed under water def
ditions (Figures 3d and 4d). y
Figure 3. Ion contents in roots of one-year-old A. alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 3. Ion contents in roots of one-year-old A. stress (Figure 2)
3.4. Ions Levels As it should be expected, there were no significant differences in the levels of Na+ and Cl−between
control and drought-stressed plants, neither in needles nor in roots. On the contrary, Na+ and Cl−
levels were much higher in salt-treated plants than in the non-stressed controls. In the presence of
300 mM NaCl, Na+ contents were about three-fold higher than under non-stress conditions, both in
needles and roots (Figures 3a and 4a). On the other hand, the salt treatment induced the accumulation
of Cl−to concentrations seven-fold (roots) or five-fold (needles) higher than in the corresponding
controls (Figures 3b and 4b). For all treatments (controls, water deficit and each NaCl concentration
in the irrigation water), both in roots and needles, Na+ concentrations were always higher than
those of Cl−under the same conditions. When comparing Na+ and Cl−contents between roots
and needles, they were always substantially higher in needles, for each treatment and the two ions
(compare Figure 4a with Figure 3a, and Figure 4b with Figure 3b). It is interesting to note the high Na+
concentration (about 400 µmol g−1 DW) measured in needles of control plants, in the absence of salt
(Figure 4a). 8 of 21
o water
nd Cl− Forests 2020, 11, 395
g
significant differe
As it should Figure 2. Photosynthetic pigments levels under water and salt stress. Needle concentrations of
chlorophyll a (Chl a), chlorophyll b (Chl b) and total carotenoids (Caro) in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated salt concentrations or subjected to
water deficit (completely withholding irrigation) (WS). Bars represent means ± SE (n = 7). For each of
the three pigments, different letters above the bars indicate significant differences between
treatments, according to Tukey’s test (α = 0.05). Figure 2. Photosynthetic pigments levels under water and salt stress. Needle concentrations of
chlorophyll a (Chl a), chlorophyll b (Chl b) and total carotenoids (Caro) in one-year-old A. alba seedlings
after 30 days of growth in the presence of the indicated salt concentrations or subjected to water deficit
(completely withholding irrigation) (WS). Bars represent means ± SE (n = 7). For each of the three
pigments, different letters above the bars indicate significant differences between treatments, according
to Tukey’s test (α = 0.05). g
p
,
y,
d Cl− levels were much higher in salt-treated plants than in the non-stressed controls. stress (Figure 2)
3.4. Ions Levels In t
sence of 300 mM NaCl, Na+ contents were about three-fold higher than under non-str
ditions, both in needles and roots (Figures 3a and 4a). On the other hand, the salt treatm
uced the accumulation of Cl− to concentrations seven-fold (roots) or five-fold (needles) higher th
he corresponding controls (Figures 3b and 4b). For all treatments (controls, water deficit and ea
Cl concentration in the irrigation water), both in roots and needles, Na+ concentrations w
ways higher than those of Cl− under the same conditions. When comparing Na+ and Cl− conte
ween roots and needles, they were always substantially higher in needles, for each treatment a
two ions (compare Figure 4a with Figure 3a, and Figure 4b with Figure 3b). It is interesting
e the high Na+ concentration (about 400 µmol g−1 DW) measured in needles of control plants,
absence of salt (Figure 4a). Ca2+ and K+ levels were also higher in the needles than in the roots, in the controls and for
plied treatments (Figure 3c, d and Figure 4c, d). Mean K+ contents generally decreased in roots a
dles of water-stressed seedlings, and in response to salt stress in a concentration-depend
nner. Still, the differences with the non-stressed controls were statistically significant only in ro
d in the presence of 200 or 300 mM NaCl (Figures 3c and 4c). As compared to the controls, me
+ concentrations increased in parallel to external salinity, both in roots and in needles, b
nificant differences were found only in needles. This effect was not observed under water def
ditions (Figures 3d and 4d). Cl− levels were much higher in salt-treated plants than in the non-stressed contro
nce of 300 mM NaCl, Na+ contents were about three-fold higher than under n
tions, both in needles and roots (Figures 3a and 4a). On the other hand, the salt
ed the accumulation of Cl− to concentrations seven-fold (roots) or five-fold (needles) hi
corresponding controls (Figures 3b and 4b). For all treatments (controls, water deficit
concentration in the irrigation water), both in roots and needles, Na+ concentrati
s higher than those of Cl− under the same conditions. When comparing Na+ and Cl−
en roots and needles, they were always substantially higher in needles, for each treat
wo ions (compare Figure 4a with Figure 3a, and Figure 4b with Figure 3b). stress (Figure 2)
3.4. Ions Levels alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 3. Ion contents in roots of one-year-old A. alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 3. Ion contents in roots of one-year-old A. alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 3. Ion contents in roots of one-year-old A. alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 3. Ion contents in roots of one-year-old A. alba seedlings after 30 days of growth in the presence
of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium, (b) chloride,
(c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). 9 of 21
9 of 20 Forests 2020, 11, 395
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9 o
orests 2020, 11, x FOR PEER REVIEW
9 o
Figure 4. Ion contents in needles of one-year-old A. alba seedlings after 30 days of growth in the
presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium,
(b) chloride, (c) potassium, (d) calcium. Bars represent means ± SE (n = 7). stress (Figure 2)
3.4. Ions Levels Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 4. Ion contents in needles of one-year-old A. alba seedlings after 30 days of growth in the
presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium,
(b) chloride, (c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 4. Ion contents in needles of one-year-old A. alba seedlings after 30 days of growth in the
presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium,
(b) chloride, (c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 4. Ion contents in needles of one-year-old A. alba seedlings after 30 days of growth in the
presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). (a) sodium,
(b) chloride, (c) potassium, (d) calcium. Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). 3.5. Osmolyte Contents
Osmolyte biosynthesis is a general response of all organisms, including plants, to environmental
conditions that generate osmotic stress, such as salinity or drought. The accumulation of these
compatible solutes helps maintain osmotic balance, minimising or even avoiding cell dehydration. In
this study, we measured the levels of the two main plant osmolytes: proline (Pro) and total soluble
sugars (TSS), in needles of A. alba seedlings after the water and salt stress treatments. For this species,
under our experimental conditions, Pro concentrations were low (less than 15 µmol g−1 DW) in the
control plants and did not vary significantly in the plants subjected for one month to the water deficit
Ca2+ and K+ levels were also higher in the needles than in the roots, in the controls and for all
applied treatments (Figure 3c,d and Figure 4c,d). Mean K+ contents generally decreased in roots and
needles of water-stressed seedlings, and in response to salt stress in a concentration-dependent manner. Still, the differences with the non-stressed controls were statistically significant only in roots and in
the presence of 200 or 300 mM NaCl (Figures 3c and 4c). stress (Figure 2)
3.4. Ions Levels As compared to the controls, mean Ca2+
concentrations increased in parallel to external salinity, both in roots and in needles, but significant
differences were found only in needles. This effect was not observed under water deficit conditions
(Figures 3d and 4d). 3.7. Non-Enzymatic Antioxidants
3.7. Non-Enzymatic Antioxidants No significant changes in the needle concentrations of total phenolic compounds (TPC, Figure
7a) or total flavonoids (TF, Figure 7b), were observed when comparing the control, non-stressed
plants to those subjected to the water deficit treatment or grown in the presence of low salt
concentration (100 mM NaCl). At higher salinities (200 and 300 mM NaCl), however, a significant
increase in TPC and TF needle contents was observed. In any case, the absolute levels of the
antioxidants and their relative accumulation compared to the control samples were small, with a
maximum of about two-fold in the presence of the highest salt concentration tested for TF, and even
No significant changes in the needle concentrations of total phenolic compounds (TPC, Figure 7a)
or total flavonoids (TF, Figure 7b), were observed when comparing the control, non-stressed plants to
those subjected to the water deficit treatment or grown in the presence of low salt concentration (100
mM NaCl). At higher salinities (200 and 300 mM NaCl), however, a significant increase in TPC and TF
needle contents was observed. In any case, the absolute levels of the antioxidants and their relative
accumulation compared to the control samples were small, with a maximum of about two-fold in the
presence of the highest salt concentration tested for TF, and even less for TPC (Figure 7). 3.6. Oxidative Stress
3.6. Oxidative Stress Malondialdehyde (MDA) is a product of lipid peroxidation, often employed as a reliable biomarker
of cellular oxidative stress [54]. Needle MAD concentrations were determined in silver fir seedlings
after the stress treatments. When compared to control values, mean MDA contents showed small
increases under water deficit conditions, as well as with growing external salinity. The observed
differences, however, were not statistically significant (Figure 6). Malondialdehyde (MDA) is a product of lipid peroxidation, often employed as a reliable
biomarker of cellular oxidative stress [54]. Needle MAD concentrations were determined in silver fir
seedlings after the stress treatments. When compared to control values, mean MDA contents showed
small increases under water deficit conditions, as well as with growing external salinity. The
observed differences, however, were not statistically significant (Figure 6). Figure 6. Malondialdehyde (MDA) needle levels in one-year-old A. alba seedlings after 30 days of
growth in the presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant differences
between treatments, according to Tukey’s test (α = 0.05). Figure 6. Malondialdehyde (MDA) needle levels in one-year-old A. alba seedlings after 30 days of
growth in the presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant differences
between treatments, according to Tukey’s test (α = 0.05). Figure 6. Malondialdehyde (MDA) needle levels in one-year-old A. alba seedlings after 30 days of
growth in the presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant differences
between treatments, according to Tukey’s test (α = 0.05). Figure 6. Malondialdehyde (MDA) needle levels in one-year-old A. alba seedlings after 30 days of
growth in the presence of the indicated NaCl concentrations or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant differences
between treatments, according to Tukey’s test (α = 0.05). p
conditions; however, a
3.5. Osmolyte Contents d
Forests 2020, 11, x FOR PEER REVIEW
10 of 2 10 of 21 Forests 2020, 11, 395
glucose g−1 DW 10 of 21 Figure 5. Needle contents of (a) proline (Pro) and (b) total soluble sugars (TSS), in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 5. Needle contents of (a) proline (Pro) and (b) total soluble sugars (TSS), in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). ests 2020, 11, x FOR PEER REVIEW
10 o p
conditions; however, a
3.5. Osmolyte Contents (Figure 5a). The mean TSS level in non-stressed seedlings was 20 mg eq. glucose g−1 DW,
approximately, and increased significantly, both under water deficit and salt stress conditions, in the
latter case, in a clear concentration-dependent manner, reaching a maximum value of ca. 50 mg eq. glucose g−1 DW (Figure 5b). Osmolyte biosynthesis is a general response of all organisms, including plants, to environmental
conditions that generate osmotic stress, such as salinity or drought. The accumulation of these
compatible solutes helps maintain osmotic balance, minimising or even avoiding cell dehydration. In this study, we measured the levels of the two main plant osmolytes: proline (Pro) and total soluble
sugars (TSS), in needles of A. alba seedlings after the water and salt stress treatments. For this species,
under our experimental conditions, Pro concentrations were low (less than 15 µmol g−1 DW) in the
control plants, and did not vary significantly in the plants subjected for one month to the water
deficit conditions; however, a significant increase of about two-fold was observed in the presence
of salt (Figure 5a). The mean TSS level in non-stressed seedlings was 20 mg eq. glucose g−1 DW,
approximately, and increased significantly, both under water deficit and salt stress conditions, in the
latter case, in a clear concentration-dependent manner, reaching a maximum value of ca. 50 mg eq. glucose g−1 DW (Figure 5b). Forests 2020, 11, 395
10 of 2
latter case, in a clear concentration dependent manner, reaching a maximum value of ca. 50 mg eq. glucose g−1 DW (Figure 5b). Figure 5. Needle contents of (a) proline (Pro) and (b) total soluble sugars (TSS), in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 5. Needle contents of (a) proline (Pro) and (b) total soluble sugars (TSS), in one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). less for TPC (Figure 7).
3.8. Antioxidant Enzyme Activities The specific activities for some of the most important antioxidant enzymatic systems: SOD, CAT,
APX and GR, were calculated in protein extracts prepared from all collected needle samples (Figure 8). 11 of 21 11 of 21 Forests 2020, 11, 395
between tre Forests 2020, 11, 395
between tre A slight increase in the average values of the specific activities of all tested enzymes, except GR,
was detected under water deficit conditions. However, the differences with the corresponding controls
were statistically significant only for SOD. Regarding salt stress conditions, CAT-specific activity did
not vary significantly for any of the applied treatments (Figure 8b), whereas for the other three enzymes,
a slight, but significant, increase was observed, either at all NaCl concentrations (SOD, Figure 8a) or
only at medium and high salinities (APX, Figure 8c; GR, Figure 8d). These data are also in agreement
with the relatively low level of oxidative stress induced by water deficit or salinity in A. alba seedlings
under our experimental conditions. No significant changes in the needle concentrations of total phenolic compounds (TPC, Figure
7a) or total flavonoids (TF, Figure 7b), were observed when comparing the control, non-stressed
plants to those subjected to the water deficit treatment or grown in the presence of low salt
concentration (100 mM NaCl). At higher salinities (200 and 300 mM NaCl), however, a significant
increase in TPC and TF needle contents was observed. In any case, the absolute levels of the
antioxidants and their relative accumulation compared to the control samples were small, with a
maximum of about two-fold in the presence of the highest salt concentration tested for TF, and even
less for TPC (Figure 7). Figure 7. (a) Total phenolic compounds (TPC) and (b) total flavonoids (TF) needle contents in one-
year-old A. alba seedlings after 30 days of growth in the presence of the indicated NaCl concentrations
or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 7. (a) Total phenolic compounds (TPC) and (b) total flavonoids (TF) needle contents in
one-year-old A. alba seedlings after 30 days of growth in the presence of the indicated NaCl concentrations
or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). less for TPC (Figure 7).
3.8. Antioxidant Enzyme Activities Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Forests 2020, 11, x FOR PEER REVIEW
11 of 20
3.8. Antioxidant Enzyme Activities
The specific activities for some of the most important antioxidant enzymatic systems: SOD, CAT,
APX and GR, were calculated in protein extracts prepared from all collected needle samples (Figure
8). A slight increase in the average values of the specific activities of all tested enzymes, except GR,
was detected under water deficit conditions. However, the differences with the corresponding
controls were statistically significant only for SOD. Regarding salt stress conditions, CAT-specific
activity did not vary significantly for any of the applied treatments (Figure 8b), whereas for the other
three enzymes, a slight, but significant, increase was observed, either at all NaCl concentrations (SOD,
Figure 8a) or only at medium and high salinities (APX, Figure 8c; GR, Figure 8d). These data are also
in agreement with the relatively low level of oxidative stress induced by water deficit or salinity in
A. alba seedlings under our experimental conditions. 11 of 20
rtant antioxidant enzymatic systems: SOD, CAT,
pared from all collected needle samples (Figure
cific activities of all tested enzymes, except GR,
wever, the differences with the corresponding
Regarding salt stress conditions, CAT-specific
ed treatments (Figure 8b), whereas for the other 8b), w ifican Figure 7. (a) Total phenolic compounds (TPC) and (b) total flavonoids (TF) needle contents in one-
year-old A. alba seedlings after 30 days of growth in the presence of the indicated NaCl concentrations
or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 7. (a) Total phenolic compounds (TPC) and (b) total flavonoids (TF) needle contents in
one-year-old A. alba seedlings after 30 days of growth in the presence of the indicated NaCl concentrations
or subjected to water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the
bars indicate significant differences between treatments, according to Tukey’s test (α = 0.05). ree enzymes, a slight, but significant, increase was observed, either at all NaCl concentrations (SOD
gure 8a) or only at medium and high salinities (APX, Figure 8c; GR, Figure 8d). less for TPC (Figure 7).
3.8. Antioxidant Enzyme Activities These data are als
agreement with the relatively low level of oxidative stress induced by water deficit or salinity in
alba seedlings under our experimental conditions. Figure 8. (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and
(d) glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 8. (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and (d)
glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba seedlings
after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to water deficit
stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant
differences between treatments, according to Tukey’s test (α = 0.05). Figure 8. (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and
(d) glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 8. (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and (d)
glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba seedlings
after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to water deficit
stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant
differences between treatments, according to Tukey’s test (α = 0.05). Figure 8. (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and
(d) glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba
seedlings after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to
water deficit stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate
significant differences between treatments, according to Tukey’s test (α = 0.05). Figure 8. 3.9. Hierarchical Cluster Analysis (HCA) Growth, physiological and biochemical parameters that showed significant differences (p < 0.05)
in the ANOVA were included in a hierarchical cluster analysis (HCA) of the data (Figure 9). The HCA
analysis allowed for establishing a signature of responses depending on the stress applied. Considering
the different treatments, the two main branches of the HCA separated the control, water stress and
100 mM NaCl treatments, on the one side, from the 200 and 300 mM NaCl treatments, on the other. Besides, within the first cluster, the control and the 100 mM NaCl treatments were grouped together
and separated from the water deficit conditions. In terms of the general profile of all analysed variables,
the treatments at higher salinities, 200 and 300 mM NaCl, were the most closely correlated (Figure 9). The water stress treatment had a characteristic signature in the response, which was quite similar to the
control, but with lower needle fresh weight (NFW) and needle water content (NWC), lower levels of
chlorophylls and higher of carotenoids, lower concentrations of K, and slightly higher concentrations
of osmolytes (Pro and TSS), phenolics (TPC) and flavonoids (TF), as well as of the enzymatic activities
SOD and APX. Regarding salinity, the two treatments with highest NaCl concentrations (200 and
300 mM) were characterised by a specific signature, which was displayed with greater intensity at the
highest NaCl concentration (300 mM), corresponding to high levels of concentrations of Na, Cl and Ca,
and low levels of K, as well as high levels of osmolytes and non-enzymatic and enzymatic antioxidants. The 100 mM NaCl was generally intermediate between the control and the 200 mM NaCl treatment,
except for a remarkably high content in K in the needles (Figure 9). Forests 2020, 11, x FOR PEER REVIEW
12 of 20
NaCl concentrations (200 and 300 mM) were characterised by a specific signature, which was
displayed with greater intensity at the highest NaCl concentration (300 mM), corresponding to high
levels of concentrations of Na, Cl and Ca, and low levels of K, as well as high levels of osmolytes and
non-enzymatic and enzymatic antioxidants. The 100 mM NaCl was generally intermediate between
the control and the 200 mM NaCl treatment, except for a remarkably high content in K in the needles
(Figure 9). Two major clusters were also distinguished regarding the different measured parameters. 3.9. Hierarchical Cluster Analysis (HCA) The
first one includes needle fresh weight (NFW), K+ in roots (Kr) and total carotenoids (Caro)—the two
former variables were highly correlated and displayed the highest values in the control and the
lowest in the presence of 300 mM NaCl. The rest of the traits form a second major cluster, in which
several sub-clusters were identified (Figure 9). For example, needle water content (NWC), the
chlorophylls (Chl a and Chl b) and K+ in needles (Kn) can be grouped in one of the major sub-clusters,
all displaying the lowest levels in the water stress treatment. The other major sub-cluster includes the
rest of the variables: Ca2+, Na+ and Cl− in needles and roots (Can, Car, Nan, Nar, Cln, Clr), osmolytes
(Pro, TSS), antioxidant compounds (TPC, TF) and the SOD, APX and GR activities. Generally, these
traits displayed higher levels under salt stress, in most cases increasing in parallel to increasing
salinity. SOD, however, appeared to be activated predominantly by water deficit. The highest
correlations were observed between the ion concentrations, in both roots and needles, as well as
between the ions, Pro and GR activity (Figure 9). Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests, in one-
year-old A. alba seedlings after 30 days of water or salt stress treatments. NFW, needle fresh weight;
NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total carotenoids; Nan,
sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium in needles; Nar,
sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro, proline; TSS,
total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide dismutase
activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests,
in one-year-old A. alba seedlings after 30 days of water or salt stress treatments. less for TPC (Figure 7).
3.8. Antioxidant Enzyme Activities (a) Superoxide dismutase (SOD)-, (b) catalase (CAT)-, (c) ascorbate peroxidase (APX)- and (d)
glutathione reductase (GR)-specific activities in needle protein extracts of one-year-old A. alba seedlings
after 30 days of growth in the presence of the indicated NaCl concentrations or subjected to water deficit
stress (WS). Bars represent means ± SE (n = 7). Different letters above the bars indicate significant
differences between treatments, according to Tukey’s test (α = 0.05). 12 of 21 Forests 2020, 11, 395 12 of 21 3.9. Hierarchical Cluster Analysis (HCA) NFW, needle
fresh weight; NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total
carotenoids; Nan, sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium
in needles; Nar, sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro,
proline; TSS, total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide
dismutase activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests, in one-
year-old A. alba seedlings after 30 days of water or salt stress treatments. NFW, needle fresh weight;
NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total carotenoids; Nan,
sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium in needles; Nar,
sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro, proline; TSS,
total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide dismutase
activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests,
in one-year-old A. alba seedlings after 30 days of water or salt stress treatments. NFW, needle
fresh weight; NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total
carotenoids; Nan, sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium
in needles; Nar, sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro,
proline; TSS, total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide
dismutase activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests, in one-
year-old A. alba seedlings after 30 days of water or salt stress treatments. 4.1. Effect of Salt and Water Stress on Seedlings’ Growth and Photosynthetic Pigments Growth inhibition is probably the first and most general response of plants to different types
of stress, which induce a switch from primary to secondary metabolism as energy and metabolic
precursors, normally used for biomass accumulation during the vegetative growth phase, are redirected
to the activation of defence reactions against those stress factors [55,56]. In most plant species, therefore,
determination of growth parameters is commonly used to assess the effects of stress on the plants. This approach is not so useful for slow-growing species such as conifers, including the silver fir,
especially in the first years of life [22,57,58]. Nevertheless, after one month of water and salt stress
treatments, we could detect a significant inhibition of growth in one-year-old silver fir seedlings,
indicating that at this developmental stage, this species is relatively more sensitive to drought and
high salinity than other conifers, for example, Picea abies [59]. Fresh weight and water content are
probably the most precise parameters to assess the degree of stress-induced growth inhibition, but their
measurement requires the destruction of the plants. It is, therefore, essential to identify and characterise
appropriate physiological and biochemical stress indicators that could be quantified through simple,
sensitive and minimally invasive assays requiring small amounts of plant material. Photosynthetic pigments can be included in these putative stress markers, as inhibition of
photosynthesis is also a common effect of stress [60–62], and is accompanied by a reduction of
chlorophyll contents, by inhibition of its synthesis combined with activation of its degradation [63]. Previous studies on plants of different conifer species exposed to salt or drought treatments have
also reported a decrease in chlorophylls a and b and, in some cases, in total carotenoids [59,64,65]. Following this general trend, in the present study, water and salt stress caused a significant reduction
of chlorophylls a and b contents in silver fir needles. On the contrary, carotenoid levels were very low
and did not show any significant decrease under stress. 3.9. Hierarchical Cluster Analysis (HCA) NFW, needle fresh weight;
NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total carotenoids; Nan,
sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium in needles; Nar,
sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro, proline; TSS,
total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide dismutase
activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. Figure 9. Hierarchical cluster analysis (HCA) and heatmap of growth, physiological and biochemical
parameters displaying significant differences (p < 0.05) in analysis of variance (ANOVA) tests,
in one-year-old A. alba seedlings after 30 days of water or salt stress treatments. NFW, needle
fresh weight; NWC, needle water content; Chl a, chlorophyll a; Chl b, Chlorophyll b; Caro, total
carotenoids; Nan, sodium in needles; Cln, chloride in needles; Can, calcium in needles; Kn, potassium
in needles; Nar, sodium in roots; Clr, chloride in roots; Car, calcium in roots; Kr, potassium in roots; Pro,
proline; TSS, total soluble sugars; TPC, total phenolic compounds; TF, total flavonoids; SOD, superoxide
dismutase activity; APX, ascorbate peroxidase activity; GR, glutathione reductase activity. 13 of 21 Forests 2020, 11, 395 Two major clusters were also distinguished regarding the different measured parameters. The first
one includes needle fresh weight (NFW), K+ in roots (Kr) and total carotenoids (Caro)—the two former
variables were highly correlated and displayed the highest values in the control and the lowest in
the presence of 300 mM NaCl. The rest of the traits form a second major cluster, in which several
sub-clusters were identified (Figure 9). For example, needle water content (NWC), the chlorophylls
(Chl a and Chl b) and K+ in needles (Kn) can be grouped in one of the major sub-clusters, all displaying
the lowest levels in the water stress treatment. The other major sub-cluster includes the rest of the
variables: Ca2+, Na+ and Cl−in needles and roots (Can, Car, Nan, Nar, Cln, Clr), osmolytes (Pro,
TSS), antioxidant compounds (TPC, TF) and the SOD, APX and GR activities. Generally, these traits
displayed higher levels under salt stress, in most cases increasing in parallel to increasing salinity. SOD, however, appeared to be activated predominantly by water deficit. The highest correlations
were observed between the ion concentrations, in both roots and needles, as well as between the ions,
Pro and GR activity (Figure 9). 4. Discussion 4.1. Effect of Salt and Water Stress on Seedlings’ Growth and Photosynthetic Pigments 4.2. Effect of Salt Stress on Ion Accumulation Salt-sensitive plants (glycophytes), which include all major crops, generally respond to salt stress
by trying to keep low concentrations of toxic ions in the leaves, either by exclusion mechanisms at the
root level or by blocking its transport to the aerial parts of the plant, mechanisms that are effective only
at low soil salinities [66–68]. On the contrary, in A. alba, salt stress caused the accumulation of relatively
high concentrations of Na+ and Cl−(and also Ca2+) in roots and needles, in parallel to the increasing
NaCl concentration in the irrigation water. This behaviour has also been reported for other coniferous
species [18,65,69]. What should be highlighted is that, for each NaCl concentration, the levels of
all tested ions were substantially higher in the needles than in the roots, indicating the presence of
active systems for their transport to the aerial parts of the plants. It is also worth noting the high Na+ 14 of 21 Forests 2020, 11, 395 concentration (not so much for Cl−) measured in needles of control seedlings. This result suggests that
silver fir possesses active mechanisms for uptake and transport to the needles of these ions, mainly
Na+, to be used as ‘cheap’ inorganic osmolytes, even under low salinity conditions. In terms of
energy consumption, ion transport will be less demanding than the synthesis of organic osmolytes [70]. Therefore, it can be concluded that the responses of A. alba seedlings to stress include constitutive
mechanisms based on the accumulation in needles of Na+ and other inorganic ions, without reaching
toxic levels. The increase in Ca2+ concentration in parallel to increasing soil salinity should also be
considered as a mechanism of tolerance against salt stress, as the role of calcium counteracting the
harmful effects of NaCl is well established [71]. Accumulation of Na+ in plant tissues is generally accompanied by a reduction of K+ levels, as both
ions compete for the same membrane transporters [72,73]. Maintenance of relatively high K+/Na+
ratios is considered as a relevant mechanism for salt tolerance [74]. Salt-induced reduction of K+
concentration has been detected, indeed, in roots of silver fir seedlings, but not in needles, where no
significant differences with the controls were observed. The active transport of K+ from the roots
limited the reduction of K+/Na+ ratios in the needles, thus contributing to salt tolerance. 4.3. Osmolyte Synthesis Environment stress conditions that lead to cellular dehydration, including salinity and water
stress, trigger the cytosolic accumulation of various compatible organic solutes or osmolytes. Proline
(Pro) is a common osmolyte in plants, synthesised in many species in response to different abiotic
stresses [75]. Previous studies have reported significant increases in Pro concentrations under water
deficit and/or high salinity also in conifers, such as spruce [18,59,76] or pine [77,78]. In the present
study, a significant increase in Pro concentration has been detected in response to salt stress, although
the differences with the non-stressed controls were small, as were the absolute Pro levels were reached. Therefore, it is likely that the contribution of Pro to osmotic adjustment under salt stress conditions is
also small; nevertheless, this does not exclude a role of Pro in the mechanisms of salt tolerance in A. alba,
based on its additional functions as an ‘osmoprotectant’ [79]. On the contrary, Pro does not seem to
play any role in the response of silver fir to water deficit, at least under our experimental conditions. p y
y
p
p
Soluble carbohydrates also play functional roles in the abiotic stress responses of many different
plant species, contributing to osmotic adjustment and as osmoprotectants [80]. Accumulation of total
soluble sugars (TSS) in response to salts stress has been reported, for example, in the needles, sapwood
and inner bark of some coniferous species [81]. The experiments performed in the present study
showed a significant increase in TSS levels under water deficit and salt stress conditions, suggesting
the participation of these osmolytes in the responses of silver fir seedlings to both drought and salinity. 4.4. Oxidative Stress and Antioxidant Defence Mechanisms Reactive oxygen species (ROS) are produced in plants as by-products during different metabolic
reactions, for example, photosynthesis and respiration, and play essential roles in cell signalling
and homeostasis [82]. ‘Normal’ physiological ROS levels can increase dramatically under stress
conditions—drought, salinity, high temperatures or UV irradiation, among others—breaking down the
balance between ROS production and scavenging, and causing oxidative stress [83,84]. The harmful
effects of excess ROS can be counteracted by the activation of antioxidant enzymes, such as superoxide
dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) or glutathione reductase (GR) [85,86],
and there are many published reports demonstrating the increase in the specific activity of these
enzymes under stress conditions. SOD is considered as the first line of defence against oxidative
stress, eliminating the highly reactive superoxide radicals [87]. SOD activity increases in response
to high salinity stress, for example, in pea [88], maize [89], Morus alba [90] or Brassica napus [91]. The combined effects of drought and high light irradiation significantly increased SOD activity in
spruce (Picea asperata Mast.) seedlings [92]. CATs are tetrameric heme-containing enzymes, mainly
located in the peroxisomes, which convert the SOD-generated H2O2 into O2 and H2O, thus contributing 15 of 21 Forests 2020, 11, 395 to ROS scavenging [93,94]. APXs also eliminate hydrogen peroxide, playing an essential role in the
antioxidant system of plants [95]. GR is a highly conserved enzyme localised mainly in the chloroplast
stroma, but also present in mitochondria, cytosol and peroxisomes [83]. GR activity has been shown
to increase in response to different stresses, for example, by chilling in cucumber (Cucumis sativus L.)
leaves [96], at high temperature in Triticum aestivum [97], under salt stress in cotton calli [98] or in
alfalfa nodules subjected to water deficit [99]. In addition to the antioxidant enzymes mentioned above (and other oxidoreductases),
non-enzymatic antioxidants, such as phenolic compounds, especially the subgroup of flavonoids—many
of them showing strong antioxidant activities—are also involved in ROS scavenging and maintaining
cellular redox equilibrium. Therefore, the stress-induced biosynthesis of these metabolites can
contribute to the mechanisms of defence against abiotic stress [100,101]. g
According to the results of the present study, the only antioxidant system that appears to be
substantially involved in the response of silver fir seedlings to water deficit stress, under the specific
experimental conditions used, is SOD. 4.5. Hierarchical Cluster Analysis The joint analysis by HCA of all growth and biochemical variables that showed statistically
significant changes in response to the applied treatments generally confirmed and extended the
information provided by the individual experiments and revealed specific signatures of the growth,
physiological and biochemical responses to the different stresses. Concerning the responses of A. alba
seedlings to water deficit, it clearly showed inhibition of growth (reduction of needle fresh weight),
needle dehydration (decrease in water content), the increase in carotenoid contents, decrease in
chlorophylls and the activation of SOD, but also a weaker activation of APX and the accumulation
of TSS and, to a much lesser extent, Pro. This analysis also confirmed the concentration-dependent
reduction of fresh weight in the presence of increasing external NaCl, needle dehydration—although
less intense than under water deficit stress—relatively strong activation of GR and APX and not so
strong of SOD, and the accumulation, to a greater or lesser extent, of TPC, TF and all measured ions,
except K+ in roots. As shown here, the responses of A. alba seedlings to drought and salinity partly
overlap, although quantitative differences between the two treatments have been observed. This is
to be expected considering that both environmental conditions cause osmotic and oxidative stress in
plants. There are, however, mechanisms of response specific for salt stress, based mostly on the control
of ion transport and ion homeostasis. 4.4. Oxidative Stress and Antioxidant Defence Mechanisms Neither the activities of the other three assayed enzymes (CAT,
APX and GR) nor phenolics or flavonoids needle contents, showed significant changes with respect to
the values measured in the controls. On the contrary, salt stress induced significant increases in the
specific activities of all tested enzymes, except CAT, as well as in the levels of total phenolic compounds
and flavonoids. In all cases, however, the observed stress-induced activation of antioxidant systems
was relatively weak, as shown by data that correlate with the low degree of drought- and salt-induced
oxidative stress suggested by the observed changes in MDA contents. It appears that other mechanisms,
such as the regulation of ion transport (for salt stress) and the accumulation of specific osmolytes
(for both, salinity and water deficit) are enough to avoid higher levels of oxidative stress under the
particular conditions used in our experiments. Stronger stress treatments will likely cause a stronger
degree of oxidative stress and consequently induce a more pronounced activation of the antioxidant
enzymes, and the accumulation to higher levels of phenolic compounds and flavonoids. 5. Conclusions This work provided new experimental data on silver fir (Abies alba), an economically and
ecologically important coniferous species, for which very little information is available regarding
its responses to drought and, especially, high salinity. Silver fir does not seem to be very resistant
to water deficit or salt stress, at least at the seedling stage, since one-month application of both
treatments inhibited growth. Nevertheless, the results presented here allowed for establishing the Forests 2020, 11, 395 16 of 21 most relevant mechanisms of (limited) tolerance to drought, mostly based on the accumulation of
soluble carbohydrates as osmolytes/osmoprotectants. Tolerance to salt, on the other hand, seems to
depend on the active transport to the needles of Na+, Cl−and Ca2+, the maintenance of relatively high
K+/Na+ ratios and the accumulation of Pro and soluble sugars for osmotic adjustment. Interestingly,
A. alba seedlings present relatively high Na+ concentrations in their needles, in the absence of salt. As
far as it does not reach toxic levels, Na+ can contribute to osmotic balance as a ‘cheap’ osmoticum, and
its accumulation may represent a constitutive mechanism of defence against stress. These responses
appear to be efficient enough to avoid the generation of high levels of oxidative stress; therefore, the
activation of antioxidant systems most likely play only a secondary role in the mechanisms of tolerance
to drought and salinity in silver fir seedlings. g
y
g
In addition, from a practical point of view, we would like to suggest the increase in carotenoids
contents, the decrease of chlorophylls a and b levels and the accumulation of soluble sugars as reliable
biomarkers for drought stress in this species. Salt stress, on the other hand, is associated to the decrease
in chlorophylls levels and the accumulation in the needles of TSS, Pro (as organic osmolytes) and,
especially, cations such as Na+ or Ca2+. These biochemical stress markers may be useful for the future
management of silver fir forests and reforestation programmes. Author Contributions: Conceptualization, M.B. and R.E.S.; Formal analysis, A.F.S.; Investigation, I.M.T., S.G.-O. and M.P.; Resources, I.M.T. and A.F.S.; Supervision, M.B., O.V. and R.E.S.; Validation, A.F.S. and J.P.; Visualization,
O.V.; Writing—original draft, I.M.T.; Writing—review and editing, O.V. All authors have read and agreed to the
published version of the manuscript. Funding: This research was partially funded by Doctoral School from the University of Agricultural Sciences and
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Prognostic value of neutrophil-to-lymphocyte ratio in advanced nasopharyngeal carcinoma: a large institution-based cohort study from an endemic area
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BMC cancer
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cc-by
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© The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Abstract Background: Findings remain unclear whether neutrophil-to-lymphocyte ratio (NLR) detrimentally affects advanced
nasopharyngeal carcinoma (NPC) prognosis. We aim to evaluate the prognostic value of NLR in patients with NPC
based on a large-scale cohort from an endemic area. Methods: We selected patients retrospectively from a cohort examining long-term cancer outcomes following
diagnosis. Neutrophil counts and lymphocyte counts were assessed prior to treatment. Kaplan–Meier method and
log-rank test were used to calculate and compare survival outcomes. Additionally, Cox proportional hazards model
was utilized to carry out univariate and multivariate analyses. Results: Between October 2009 and August 2012, we enrolled 1550 consecutive NPC patients staged II-IVB. The
median value of NLR was 2.27 (interquartile range [IQR], 1.71–3.12). Determined by operating characteristic curve
using overall survival (OS) as an endpoint, the cutoff value for NLR was 2.50. At 5 years, NLR > 2.50 was associated
with inferior OS (90.3% vs 82.5%; P < 0.001), distant metastasis-free survival (DMFS, 89.4% vs 85.0%; P = 0.014), and
progression-free survival (PFS, 80.9% vs 76.5%; P = 0.031) than NLR ≤2.50. In multivariate analysis, NLR was found to
be a significant prognostic factor for OS (HR, 1.72; 95% CI, 131–2.24; P < 0.001), DMFS (HR, 1.45; 95% CI, 1.10–1.92;
P = 0.009), and PFS (HR, 1.29; 95% CI, 1.04–1.59; P = 0.021). Conclusion: Pretreatment NLR independently affects survival. Our findings suggest that NLR measurements will be
of great clinical significance in the management of NPC. Keywords: Nasopharyngeal carcinoma, advanced, Neutrophil-to-lymphocyte ratio, Prognosis [1]. Tumor-Node-Metastasis (TNM) stage is currently the
primary measure to predict NPC prognosis. However,
TNM staging system is not adequate for predicting NPC
outcome, and patients that are in the same TNM stage
often have substantial clinical heterogeneity [2]. Plasma
epstein–barr virus (EBV) DNA titre remains the sole bio-
marker that has clinical utility in patients with NPC [3, 4]. Nevertheless, the high cost and great inter-laboratory vari-
ability for examination of plasma EBV DNA hinders the
ability to include in routine clinical practice [5]. For this
reason, it is important to identify inexpensive, objective, Prognostic value of neutrophil-to-
lymphocyte ratio in advanced
nasopharyngeal carcinoma: a large
institution-based cohort study from an
endemic area Ji-Jin Yao1,2†, Feng-Ting Zhu3†, Jun Dong4†, Zi-Bin Liang5, Le-Wei Yang6, Shao-Yi Chen2, Wang-Jian Zhang7,8,
Wayne R. Lawrence7,8, Fan Zhang2, Si-Yang Wang2, Ying Sun1*
and Guan-Qun Zhou1* Yao et al. BMC Cancer (2019) 19:37
https://doi.org/10.1186/s12885-018-5236-2 Yao et al. BMC Cancer (2019) 19:37
https://doi.org/10.1186/s12885-018-5236-2 RESEARCH ARTICLE
Open Access
Prognostic value of neutrophil-to-
lymphocyte ratio in advanced
nasopharyngeal carcinoma: a large
institution-based cohort study from an
endemic area
Ji-Jin Yao1,2†, Feng-Ting Zhu3†, Jun Dong4†, Zi-Bin Liang5, Le-Wei Yang6, Shao-Yi Chen2, Wang-Jian Zhang7,8,
Wayne R. Lawrence7,8, Fan Zhang2, Si-Yang Wang2, Ying Sun1*
and Guan-Qun Zhou1* Open Access Background In Southern China, nasopharyngeal carcinoma (NPC) is a
common malignancy. Previous literature reported the an-
nual incidence rate varied from 15 to 50 cases per 100,000 * Correspondence: sunying@sysucc.org.cn; zhougq@sysucc.org.cn
†Ji-Jin Yao, Feng-Ting Zhu and Jun Dong contributed equally to this work. 1Department of Radiation Oncology, Guangdong Key Laboratory of
Nasopharyngeal Carcinoma Diagnosis and Therapy; Collaborative Innovation
Center for Cancer Medicine, State Key Laboratory of Oncology in South
China, Sun Yat-sen University Cancer Center, Guangzhou 510060,
Guangdong Province, People’s Republic of China
Full list of author information is available at the end of the article Patient population NPC patients treated by radiotherapy with curative
intent from October 2009 through August 2012 were
identified. This retrospective study with prospectively
collected data included a cohort of 1550 men and
women. Patients were included if (1) histologically con-
firmed NPC; (2) had no prior history of malignancy; (3)
absence of distant metastasis; (4) stage II-IVA disease
according to the 8th edition of the American Joint
Committee on Cancer (AJCC) staging system; (5) did
not receive prior treatment for NPC; (6) complete pre-
treatment history of hematological variables; and (7) no
infection or inflammatory conditions. Treatment Intensity-modulated radiotherapy (IMRT) was used for
treating primary tumor and the upper neck area above
the caudal edge of the cricoid cartilage. Using a previ-
ously described treatment protocol by our institution,
target volumes were delineated [15] in agreement with
the
International
Commission
on
Radiation
Units
(ICRU) and Measurements reports 62 [16] and 83 [17]. Our institutional guidelines during the study period was
in accordance to the 7th edition of the AJCC staging sys-
tem which suggested concurrent chemoradiotherapy
(CCRT) for stage II disease, and CCRT +/−neoadjuvant
and adjuvant chemotherapy for stages III to IVB NPC. Neoadjuvant
or
adjuvant
chemotherapy
contained
5-fluorouracil (800 mg/m2/day over 120 h) with cisplatin
(80 mg/m2), or cisplatin (80 mg/m2) with docetaxel (80
mg/m2) administered at three week intervals for 3 cycles. Concurrent chemotherapy consisted of cisplatin (80 or
100 mg/m2) given in weeks 1, 4, and 7 of RT, or cisplatin
(40 mg/m2) given weekly during RT, beginning on the
first day of RT. To fill this gap in knowledge, we investigated the
long-term prognostic effect of NLR on the outcome of
patients with NPC using a large-scale homogenous
patient cohort. Laboratory examination Laboratory examination
Absolute
neutrophil
and
lymphocyte
counts
were
assessed before treatment and determined utilizing a
Sysmex
XE-5000
automated
hematology
analyzer
(Sysmex, Kobe, Japan). We calculated NLR as the abso-
lute
counts of neutrophil divided
by the absolute
lymphocyte counts. © The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Yao et al. BMC Cancer (2019) 19:37 Page 2 of 8 and easily detected markers to complement NPC progno-
sis TNM classification system. RDD approval number RDDA2017000386. If someone
need to access the data, he/she should obtain our
consent, and have to explain the source of the data in
their study. Prior literature has shown systemic inflammatory
response stimulates cancer metastasis and progression
by facilitating angiogenesis and inhibiting apoptosis [6]. An important biomarker is the neutrophil to lymphocyte
ratio (NLR), which could accurately show systemic in-
flammation [7]. The association between elevated NLR
on adverse prognoses was reported for multiple tumor
types [8]. In clinical settings, neutrophil and lymphocyte
counts in peripheral blood are routinely measured,
where for the calculation of NLR, additional effort are
needed. Thus, NLR is potentially a promising prognostic
biomarker for NPC. To date, several studies have
examined the prognostic value of NLR in NPC patients
[9–13]. However, the results of these studies have been
inconsistent, and the prognostic role of NLR for NPC
have not been conclusively determined. Pretreatment evaluation We selected the primary endpoint as overall survival (OS),
and secondary endpoints included distant metastasis-free
survival (DMFS), locoregional relapse-free survival (LRFS),
and progression-free survival (PFS). We calculated overall
survival from initial treatment to death. For distant and
locoregional relapse-free survival analyses, we recorded
the latencies (i.e. time from initial treatment) to the first
remote or locoregional relapse respectively. We calculated
progression-free survival from the date of initial treatment
to the date of treatment failure or death from any cause,
whichever was first. Patients were seen every three
months during the first 2 years, every six months for years
3 through 5, and annually thereafter until death. The
duration of patient follow-up was measured from the first
day of therapy to either the day of last examination or the
day of death. All
included
patents
had
undergone
routine
pre-
treatment evaluations comprising of medical history,
complete physical examination, complete blood count,
fiber-optic nasopharyngoscopy, chest radiography, ab-
dominal sonography, magnetic resonance imaging (MRI)
of the
nasopharynx and neck, and
bone
scan
or
whole-body fluorodeoxyglucose positron emission tom-
ography. Patients were restaged in accordance to the 8th
edition of the AJCC staging system for NPC [14]. The
present study was conducted in adherence with institu-
tional policies to protect confidential material including
all patients’ information, and approved by our Institu-
tional Review Board. We uploaded the key raw data onto
the
Research
Data
Deposit
(RDD)
public
platform
(http://www.researchdata.org.cn), and was assigned the Yao et al. BMC Cancer (2019) 19:37 Yao et al. BMC Cancer (2019) 19:37 Page 3 of 8 Patient characteristics Table 1 presents the characteristics of 1550 patients with
NPC that met the inclusion criteria. The median age
was 45 years (range, 14–78 years). During the median
follow-up
duration
of
54.3 months
(IQR,
1.3–85.6
months), 224 patients died, including 153 due to distant
metastases, 45 because of local and/or regional relapse,
11 as a result of non-cancer causes, 6 from secondary
malignant tumors, and 9 due to unknown causes. Additionally, 200 patients developed distant metastases,
and 104 experienced local or regional relapse. The
5-year survival rates among patients were OS, 85.3%;
DMFS, 87.7%; LRFS, 90.1%; and PFS, 78.3%. Statistical analysis y
For categorical variables, we calculated relative frequen-
cies (percentage), while for continuous variables we
calculated median (interquartile range [IQR]). Addition-
ally, categorical variables were compared using χ2 test. Receiver operating characteristic (ROC) curve analysis
was used to evaluate the cutoff point for NLR. Cumula-
tive survival rates were depicted by Kaplan–Meier
curves
and
compared
by
Log-rank
tests
for
each
dichotomized biomarker. Univariate and multivariate
analysis utilizing a Cox proportional hazards model was
used to test the independent association of different
factors by backward elimination. All statistical analysis
were 2-tailed, and P < 0.05 was determined statistically
significant. All statistical analyses were completed using
R 3.1.2. Age, yr
Median
45
Interquartile range
38–53
Sex
Male
1167 (75.3)
Female
383 (24.7)
T stage
T1
190 (12.3)
T2
265 (17.1)
T3
796 (51.4)
T4
299 (19.3)
N stage
N0
190 (12.3)
N1
928 (59.9)
N2
226 (14.6)
N3
206 (13.3)
Overall stage
II
332 (21.4)
III
754 (48.6)
IVA-B
464 (29.9)
Family history
No
444 (28.6)
Yes
1106 (71.4)
Smoking history
No
961 (62.0)
Yes
589 (38.0)
Drinking history
No
1358 (87.6)
Yes
192 (12.4)
NLR
Median
2.27
Interquartile range
1.71–3.12
Chemotherapy
RT alone
165 (10.6)
CCRT
594 (38.3)
NACT+CCRT
756 (48.8)
CCRT+AC
35 (2.3)
Abbreviation: RT radiotherapy, CCRT concurrent chemoradiotherapy,
NACT neoadjuvant chemotherapy, AC adjuvant chemotherapy,
NLR neutrophil-to-lymphocyte ratio The prognostic value of NLR in NPC The median value of NLR was 2.27 (IQR, 1.71–3.12). Using OS as the endpoint, the cutoff value for NLR was
2.50, determined using ROC curve. At 5 years, patients
with NLR > 2.50 had significantly inferior OS (90.3%
vs 82.5%; P < 0.001) (Fig. 1a), DMFS (89.4% vs 85.0%;
P = 0.014) (Fig. 1b), and PFS (80.9% vs 76.5%; P =
0.031) (Fig. 1d) than patients with NLR ≤2.50. Never-
theless, we did not observe any difference in LRFS
between patients with NLR ≤2.50 and NLR >
2.50
(90.2% vs. 89.0%, P = 0.309; Fig. 1c). Further multi-
variate analysis revealed that age, T stage, N stage, and
NLR were associated with both OS and PFS (P < 0.05 for
all). N stage (HR, 3.50; 95% CI, 2.65–4.63; P < 0.001) and
NLR (HR, 1.45; 95% CI, 1.10–1.92; P = 0.009) were also
associated with DMFS; and only age (HR, 1.40; 95% CI,
1.02–1.92; P = 0.035) was associated with LRFS (Table 2). In order to further demonstrate the predictive value of
NLR in advanced NPC, we also analyzed the prognostic
factors for advanced NPC in the multivariate models with-
out NLR. Our results showed that age, T stage, and N
stage were independent risk factors for both OS and PFS Abbreviation: RT radiotherapy, CCRT concurrent chemoradiotherapy,
NACT neoadjuvant chemotherapy, AC adjuvant chemotherapy,
NLR neutrophil-to-lymphocyte ratio (P < 0.05 for all). Additionally, N stage (HR, 3.49; 95% CI,
2.64–4.61; P < 0.001) was an independent risk factor for
DMFS; and age was a significant predictor for LRFS (HR,
1.40; 95% CI, 1.02–1.92; P = 0.035) (Table 3). Overall, the
C-index of multivariate model with or without NLR were Yao et al. BMC Cancer (2019) 19:37 Page 4 of 8 Fig. 1 According to NLR determined by ROC (≤2.50 vs. > 2.50), Kaplan-Meier survival curves for (a) overall survival, (b) distant metastasis-free
survival, (c) locoregional recurrence-free survival, and (d) progression-free survival Fig. 1 According to NLR determined by ROC (≤2.50 vs. > 2.50), Kaplan-Meier survival curves for (a) overall survival, (b) distant metastasis-free
survival, (c) locoregional recurrence-free survival, and (d) progression-free survival 0.69 (95% CI, 0.65–0.73) and 0.68 (95% CI, 0.64–0.72),
0.67 (95% CI, 0.64–0.71) and 0.67 (95% CI, 0.63–0.70),
0.57 (95% CI, 0.52–0.61) and 0.55 (95% CI, 0.51–0.60),
and 0.63 (95% CI, 0.60–0.66) and 0.63 (95% CI, 0.60–0.66)
for OS’s, DMFS’s, LRFS’s, and PFS’s multivariate model,
respectively. The prognostic value of NLR in NPC [18], the prognostic ability of NLR is not conclusively
determined for NPC. Our analysis from a large sample
indicated that patients with NLR > 2.50 were generally
associated with inferior OS, DMFS, and PFS, compared
to patients with NLR ≤2.50, except for LRFS. Further
analyses to detect interactions between NLR and clinico-
pathological characteristics found that among patients
that either smoked or had further advanced disease
(higher T stage, N stage, and overall stage) were also
more likely to have high levels of NLR. NLR is associated with known prognostic clinical indices
In the present study, ROC curve was used to evaluate
different cutoff points for NLR. As previously de-
scribed, we divided patients into two groups accord-
ing to NLR: high NLR (> 2.50) and low NLR (≤2.50). Additionally, we examined the correlations between
NLR and various clinicopathological features. Patients
that smoked generally had higher NLR (P = 0.011)
and also patients with advanced disease (higher T
stage, P = 0.001; N stage, P = 0.015; overall stage, P <
0.001). However,
no
significant
differences
were
observed between groups regarding age, sex, family
history, history of alcohol consumption, or treatment
strategy (all P > 0.05; Table 4). y
g
In an analysis by Templeton and colleagues [19] on
NLR as a prognostic biomarker, suggested that high
NLR was associated with adverse survival regardless of
the threshold for patient stratification. Recently, An et
al. [9] retrospectively reviewed 363 NPC patients, and
suggested that a high NLR > 3.73 was strongly associated
with inferior PFS, DMFS, and LRFS for NPC patients. Another study reported by Sun et al. [10] indicated that
NLR ≥2.7 was associated with shorter PFS in patients
with NPC. In the present study, multivariate analysis
showed that increasing NLR > 2.50 was mostly detri-
mental to OS, DMFS, and PFS. Consistent with our
study, Li et al. [18] prospectively analyzed the prognostic
value of inflammatory biomarkers in a cohort of NPC
patients (N = 388), and indicated that NLR > 2.50 was
significantly associated with inferior PFS. Discussion Although an elevated ratio of NLR was reported as an
inadequate prognostic indicator in numerous cancers Yao et al. BMC Cancer (2019) 19:37 Page 5 of 8 Table 2 Multivariate analysis of NLR determined by ROC for patients with advanced NPC
Endpoint
Variable
HR
95% CI for HR
P value
OS
Age (≤45 vs. > 45)
1.69
1.29–2.22
< 0.001
Sex (Male vs. Female)
1.30
0.90–1.87
0.163
Smoking history (No vs. Yes)
1.11
0.83–1.49
0.492
T stage (T1–2 vs. T3–4)
1.34
0.98–1.85
0.071
N stage (N0–1 vs. N2–3)
2.86
2.19–3.73
< 0.001
NLR (≤2.50 vs. > 2.50)
1.72
1.31–2.24
< 0.001
Treatment (RT alone vs CCRT)
1.43
0.82–2.50
0.213
Treatment (RT alone vs NACT+CCRT)
1.23
0.70–2.14
0.472
DMFS
Sex (Male vs. Female)
1.34
0.95–1.90
0.097
N stage (N0–1 vs. N2–3)
3.50
2.65–4.63
< 0.001
NLR (≤2.50 vs. > 2.50)
1.45
1.10–1.92
0.009
LRFS
Age (≤45 vs. > 45)
1.40
1.02–1.92
0.035
N stage (N0–1 vs. N2–3)
1.35
0.97–1.88
0.077
PFS
Age (≤45 vs. > 45)
1.44
1.16–1.79
0.001
Family history (No vs. Yes)
1.17
0.91–1.50
0.213
Smoking history (No vs. Yes)
1.12
0.90–1.39
0.323
T stage (T1–2 vs. T3–4)
1.27
0.99–1.62
0.063
N stage (N0–1 vs. N2–3)
2.25
1.81–2.79
< 0.001
NLR (≤2.50 vs. > 2.50)
1.29
1.04–1.59
0.021
Abbreviation: OS overall survival, DMFS distant metastasis-free survival, LRFS locoregional relapse-free survival, PFS progression-free survival, RT radiotherapy,
CCRT concurrent chemoradiotherapy, NACT neoadjuvant chemotherapy, NLR neutrophil-to-lymphocyte ratio, HR rate ratio, CI confidence interval Table 2 Multivariate analysis of NLR determined by ROC for patients with advanced NPC
Endpoint
Variable
HR
95% CI for HR
P value breviation: OS overall survival, DMFS distant metastasis-free survival, LRFS locoregional relapse-free survival, PFS progression-free surviva
RT concurrent chemoradiotherapy, NACT neoadjuvant chemotherapy, NLR neutrophil-to-lymphocyte ratio, HR rate ratio, CI confidence i Conversely, Chua et al. [11] examined the signifi-
cance of NLR prognostic in a pooled cohort of NPC
patients (N = 380) from two controlled trials, but
were unable to determine if NLR adds prognostic
value for NPC. The first potential reasoning is that
different NLR levels have different prognostic value. In the study by Chua et al., NLR was dichotomized
into binary variables using the median value of NLR
stratified patients, but failed to use ROC curve, which
is confirmed to be a central or unifying position in
the process of assessing and using diagnostic tool to
analyze the optimum cutoff point [20, 21]. Discussion Secondly,
the study was underpowered because of the long
duration of 15 years to recruit patients, potentially
leading to inter-study heterogeneity, specifically data
maturity and quality of radiotherapy techniques as
highlighted by the authors. Third, although Chua et
al. [11] found patients with high NLR tended to have
lower survival, this trend did not reach statistical sig-
nificance. This could be due to the inability to iden-
tify an effect because of small sample size. Elevated NLR was recognized as a significant risk
factor in patients with NPC. However, the mecha-
nisms underlying this observation remains largely
unclear. One possibility might be that high NLR
serves as a marker for up-regulated inflammatory
processes within the host microenvironment that po-
tentially promote the development of more aggres-
sive tumor clones [22–24]. Another potential reason
is that elevated markers for systemic inflammatory
response may increase with elevated circulating con-
centrations of several cytokines (IL-6, IL-7, IL-8,
IL-9, IL-12, IL-1ra). Of these cytokines, IL-6 in par-
ticular acts to increase the synthesis of acute-phase
proteins, and have shown to be associated with both
adverse prognosis and tumor stage in several types
of cancers [25]. The associations between NLR and TNM stage
have previously been reported. Based on findings
from prior studies that reported a positive association
of NLR with TNM stage [11], we applied a threshold
value of NLR > 2.50 for stratifying patients. This Yao et al. BMC Cancer (2019) 19:37 Yao et al. BMC Cancer (2019) 19:37 Page 6 of 8 stratification allowed us to identify that patients with
NLR > 2.50 were strongly associated with more ad-
vanced disease (higher T stage, P = 0.001; N stage,
P = 0.015; overall stage, P < 0.001). This suggests
NLR contributes to patient stratification by providing
additional
information
about
disease
burden. Of
interest, our results indicate that individuals that
smoke commonly had higher levels of NLR. It is
plausible that the variations of NLR were influenced
by smoking-related inflammation [26]. A major limitation in the present study is that we did
not collect information on other hematologic markers of
inflammation,
such
as
lymphocyte-monocyte
ratio
(LMR) and C-reactive protein [27, 28]. Another limita-
tion is that although several threshold values were used
and validated as the cutoff for NLR, not all cutoff
values were proven significant. Discussion Moreover, different re-
search institutions use varied levels of NLR, including
2 50 [18 29] 2 75 [10] and 3 00 [11] This inconsist
Conclusions
In summary, pretreatment NLR independently affects
survival for advanced NPC. Increasing NLR > 2.50 was
mostly detrimental to OS, DMFS, and PFS in patients
with advanced NPC
Additionally
pretreatment NLR
Table 3 Multivariate models without NLR for patients with
advanced NPC
Endpoint
Variable
HR
95% CI for HR
P value
OS
Age (≤45 vs. > 45)
1.64
1.25–2.14
< 0.001
Sex (Male vs. Female)
1.3
0.90–1.88
0.158
Smoking history (No vs. Yes)
1.09
0.82–1.46
0.553
T stage (T1–2 vs. T3–4)
1.42
1.04–1.94
0.029
N stage (N0–1 vs. N2–3)
2.84
2.18–3.69
< 0.001
DMFS
Sex (Male vs. Female)
1.34
0.95–1.89
0.098
N stage (N0–1 vs. N2–3)
3.49
2.64–4.61
< 0.001
LRFS
Age (≤45 vs. > 45)
1.4
1.02–1.92
0.035
N stage (N0–1 vs. N2–3)
1.35
0.97–1.88
0.077
PFS
Age (≤45 vs. > 45)
1.42
1.15–1.77
0.001
Family history (No vs. Yes)
1.17
0.92–1.50
0.205
Smoking history (No vs. Yes)
1.11
0.89–1.38
0.34
T stage (T1–2 vs. T3–4)
1.28
1.00–1.65
0.048
N stage (N0–1 vs. N2–3)
2.24
1.81–2.78
< 0.001
Abbreviation: OS overall survival, DMFS distant metastasis-free survival,
LRFS locoregional relapse-free survival, PFS progression-free survival,
RT radiotherapy, CCRT concurrent chemoradiotherapy, NACT neoadjuvant
chemotherapy, NLR neutrophil-to-lymphocyte ratio, HR rate ratio,
CI confidence interval
Table 4 Baseline characteristics of the patients with advanced
NPC stratified by NLR
No. Discussion of patients (%) stratified by NLR
Characteristic
≤2.50 (n = 813)
> 2.50 (n = 737)
P value
Age
0.222
≤45
404 (49.7)
343 (46.6)
> 45
409 (50.3)
394 (53.4)
Sex
0.034
Male
575 (70.7)
557 (75.6)
Female
238 (29.3)
180 (24.4)
T stage
0.001
T1
110 (13.5)
80 (10.9)
T2
151 (18.6)
114 (15.5)
T3
418 (51.4)
378 (51.3)
T4
134 (16.5)
165 (22.4)
N stage
0.015
N0
106 (13.0)
84 (11.4)
N1
485 (59.7)
443 (60.1)
N2
132 (16.2)
94 (12.8)
N3
90 (11.1)
116 (15.7)
Overall stage
< 0.001
II
189 (10.9)
143 (19.4)
III
316 (38.9)
341 (46.3)
IVA-B
396 (48.7)
253 (34.3)
Family history
0.574
No
575 (70.7)
531 (72.0)
Yes
238 (29.3)
206 (28.0)
Smoking history
0.402
No
496 (61.0)
465 (63.1)
Yes
317 (39.0)
272 (36.9)
Drinking history
0.247
No
720 (88.6)
638 (86.6)
Yes
93 (11.4)
99 (13.4)
Treatment strategy
0.779
RT
89 (10.9)
76 (10.3)
CCRT alone
316 (38.9)
278 (37.7)
NACT+CCRT
408 (50.2)
383 (52.0)
Abbreviation: NLR neutrophil-to-lymphocyte ratio, CCRT concurrent
chemoradiotherapy, NACT neoadjuvant chemotherapy, RT radiotherapy Table 3 Multivariate models without NLR for patients with
advanced NPC
Endpoint
Variable
HR
95% CI for HR
P value
OS
Age (≤45 vs. > 45)
1.64
1.25–2.14
< 0.001
Sex (Male vs. Female)
1.3
0.90–1.88
0.158
Smoking history (No vs. Yes)
1.09
0.82–1.46
0.553
T stage (T1–2 vs. T3–4)
1.42
1.04–1.94
0.029
N stage (N0–1 vs. N2–3)
2.84
2.18–3.69
< 0.001
DMFS
Sex (Male vs. Female)
1.34
0.95–1.89
0.098
N stage (N0–1 vs. N2–3)
3.49
2.64–4.61
< 0.001
LRFS
Age (≤45 vs. > 45)
1.4
1.02–1.92
0.035
N stage (N0–1 vs. N2–3)
1.35
0.97–1.88
0.077
PFS
Age (≤45 vs. > 45)
1.42
1.15–1.77
0.001
Family history (No vs. Yes)
1.17
0.92–1.50
0.205
Smoking history (No vs. Yes)
1.11
0.89–1.38
0.34
T stage (T1–2 vs. T3–4)
1.28
1.00–1.65
0.048
N stage (N0–1 vs. N2–3)
2.24
1.81–2.78
< 0.001
Abbreviation: OS overall survival, DMFS distant metastasis-free survival,
LRFS locoregional relapse-free survival, PFS progression-free survival,
RT radiotherapy, CCRT concurrent chemoradiotherapy, NACT neoadjuvant
chemotherapy, NLR neutrophil-to-lymphocyte ratio, HR rate ratio,
CI confidence interval Table 4 Baseline characteristics of the patients with advanced
NPC stratified by NLR stratification allowed us to identify that patients with
NLR > 2.50 were strongly associated with more ad-
vanced disease (higher T stage, P = 0.001; N stage,
P = 0.015; overall stage, P < 0.001). Discussion This suggests
NLR contributes to patient stratification by providing
additional
information
about
disease
burden. Of
interest, our results indicate that individuals that
smoke commonly had higher levels of NLR. It is
plausible that the variations of NLR were influenced
by smoking-related inflammation [26]. A major limitation in the present study is that we did
not collect information on other hematologic markers of
inflammation,
such
as
lymphocyte-monocyte
ratio
(LMR) and C-reactive protein [27, 28]. Another limita-
tion is that although several threshold values were used
and validated as the cutoff for NLR, not all cutoff
values were proven significant. Moreover, different re-
search institutions use varied levels of NLR, including
2.50 [18, 29], 2.75 [10], and 3.00 [11]. This inconsist-
ency might be due to obvious heterogeneity between
patients within these studies. However, we must note
that NLR varied significantly for T-stage and N-stage,
and overall tumor classification stage [11]. Abbreviation: NLR neutrophil-to-lymphocyte ratio, CCRT concurrent
chemoradiotherapy, NACT neoadjuvant chemotherapy, RT radiotherapy Abbreviation: NLR neutrophil-to-lymphocyte ratio, CCRT concurrent
chemoradiotherapy, NACT neoadjuvant chemotherapy, RT radiotherapy Competing interests The authors declare no competing financial interests. 12. Yin J, Qin Y, Luo YK, Feng M, Lang JY. Prognostic value of neutrophil-to-
lymphocyte ratio for nasopharyngeal carcinoma: a meta-analysis. Medicine
(Baltimore). 2017;96(29):e7577. 12. Yin J, Qin Y, Luo YK, Feng M, Lang JY. Prognostic value of neutrophil-to-
lymphocyte ratio for nasopharyngeal carcinoma: a meta-analysis. Medicine
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Y, Uno A. Prognostic role of neutrophil-lymphocyte ratio in nasopharyngeal
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Y, Uno A. Prognostic role of neutrophil-lymphocyte ratio in nasopharyngeal
carcinoma: a meta-analysis. PLoS One. 2017;12(7):e0181478. Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations. 14. Amin M. American joint committee on cancer. In: AJCC Cancer staging
manual. New York: Springer; 2016. 14. Amin M. American joint committee on cancer. In: AJCC Cancer staging
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manual. New York: Springer; 2016. Authors’ contributions 7. Motomura T, Shirabe K, Mano Y, et al. Neutrophil-lymphocyte ratio reflects
hepatocellular carcinoma recurrence after liver transplantation via
inflammatory microenvironment. J Hepatol. 2013;58(1):58–64. Data collection and writing original draft was performed by YJJ, ZBT, and DJ. Formal analysis, reviewing and editing was done by LZB, YLW, WR and ZWJ. YJJ, ZBT, CSY, ZF and WSY participated in project administration and study
design. Study design was conceived and designed by ZGQ and SY. Prior to
submission of this manuscript, all authors have reviwered and approved. 8. Viers BR, Boorjian SA, Frank I, et al. Pretreatment neutrophil-to-lymphocyte
ratio is associated with advanced pathologic tumor stage and increased
cancer-specific mortality among patients with urothelial carcinoma of the
bladder undergoing radical cystectomy. Eur Urol. 2014;66(6):1157–64. 8. Viers BR, Boorjian SA, Frank I, et al. Pretreatment neutrophil-to-lymphocyte
ratio is associated with advanced pathologic tumor stage and increased
cancer-specific mortality among patients with urothelial carcinoma of the
bladder undergoing radical cystectomy. Eur Urol. 2014;66(6):1157–64. Availability of data and materials We uploaded the key raw data onto the Research Data Deposit (RDD) public
platform (http://www.researchdata.org.cn), with the approval RDD number as
RDDA2017000386. If someone need to access the data, he/she should obtain
our consent, and have to explain the source of the data in their study. 5. Le QT, Zhang Q, Cao H, et al. An international collaboration to harmonize
the quantitative plasma Epstein–Barr virus DNA assay for future biomarker-
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nasopharyngeal carcinoma in China. Chin J Cancer. 2011;30(2):114–9. 1. Cao SM, Simons MJ, Qian CN. The prevalence and prevention of
nasopharyngeal carcinoma in China. Chin J Cancer. 2011;30(2):114–9. 2. Lee AW, Ng WT, Chan LK, et al. The strength/weakness of the AJCC/UICC
staging system (7th edition) for nasopharyngeal cancer and suggestions for
future improvement. Oral Oncol. 2012;48(10):1007–13. 2. Lee AW, Ng WT, Chan LK, et al. The strength/weakness of the AJCC/UICC
staging system (7th edition) for nasopharyngeal cancer and suggestions for
future improvement. Oral Oncol. 2012;48(10):1007–13. Conclusions In summary, pretreatment NLR independently affects
survival for advanced NPC. Increasing NLR > 2.50 was
mostly detrimental to OS, DMFS, and PFS in patients
with advanced NPC. Additionally, pretreatment NLR
may serve as a cost-effective prognostic factor in pa-
tients with NPC, and pretreatment NLR measurements
will be of great clinical significance in the management
of NPC. Page 7 of 8 Page 7 of 8 Yao et al. BMC Cancer (2019) 19:37 Page 7 of 8 Yao et al. BMC Cancer (2019) 19:37 Consent for publication
Not applicable. Consent for publication
Not applicable. 11. Chua ML, Tan SH, Kusumawidjaja G, Shwe MT, Cheah SL, Fong KW, Soong
YL, Wee JT, Tan TW. Neutrophil-to-lymphocyte ratio as a prognostic marker
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YL, Wee JT, Tan TW. Neutrophil-to-lymphocyte ratio as a prognostic marker
in locally advanced nasopharyngeal carcinoma: a pooled analysis of two
randomised controlled trials. Eur J Cancer. 2016;67:119–29. Funding
h g
Research was supported by grants from the Health and Medical
Collaborative Innovation Project of Guangzhou City, China (201604020003),
and National Natural Science Foundation of China (No. 81402532). The
funders had no role in study design, data collection and analysis, decision to
publish, or preparation of the manuscript. 3. Lin JC, Wang WY, Chen KY, et al. Quantification of plasma Epstein–Barr virus
DNA in patients with advanced nasopharyngeal carcinoma. N Engl J Med. 2004;350:2461–70. 3. Lin JC, Wang WY, Chen KY, et al. Quantification of plasma Epstein–Barr virus
DNA in patients with advanced nasopharyngeal carcinoma. N Engl J Med. 2004;350:2461–70. 4. Wang WY, Twu CW, Chen HH, et al. Long-term survival analysis of
nasopharyngeal carcinoma by plasma Epstein-Barr virus DNA levels. Cancer. 2013;119:963–70. Author details
1 1Department of Radiation Oncology, Guangdong Key Laboratory of
Nasopharyngeal Carcinoma Diagnosis and Therapy; Collaborative Innovation
Center for Cancer Medicine, State Key Laboratory of Oncology in South
China, Sun Yat-sen University Cancer Center, Guangzhou 510060,
Guangdong Province, People’s Republic of China. 2Department of Head and
Neck Oncology, The Cancer Center of the Fifth Affiliated Hospital of Sun
Yat-sen University, Zhuhai 519001, Guangdong Province, China. 3Department
of Imaging Diagnosis and Interventional Center, State Key Laboratory of
Oncology in South China, Collaborative Innovation Center for Cancer
Medicine, Sun Yat-sen University Cancer Center, Guangzhou 510060,
Guangdong Province, People’s Republic of China. 4Department of VIP
Region, State Key Laboratory of Oncology in Southern China, Collaborative
Innovation Center of Cancer Medicine, Sun Yat-sen University Cancer Center,
Guangzhou 510060, People’s Republic of China. 5Department of thoracic
oncology, the cancer center of the fifth affiliated hospital of Sun Yat-sen
University, Zhuhai 519001, Guangdong Province, China. 6Department of
abdominal oncology, the cancer center of the fifth affiliated hospital of Sun
Yat-sen University, Zhuhai 519001, Guangdong Province, China. 7Department
of Medical Statistics and Epidemiology & Health Information Research Center
& Guangdong Key Laboratory of Medicine, School of Public Health, Sun
Yat-sen University, Guangzhou 510080, Guangdong Province, China. 15. Yao JJ, Yu XL, Zhang F, et al. Radiotherapy with neoadjuvant chemotherapy
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versus concurrent chemoradiotherapy for ascending-type nasopharyngeal
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to predict prognosis of patients with nasopharyngeal carcinoma: an analysis
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et al. Prognostic role of neutrophil-tolymphocyte ratio in solid tumors: a
systematic review and metaanalysis. J Natl Cancer Inst. 2014;106:dju124. 20. Zweig MH, Campbell G. Abbreviations 8Department of Environmental Health Sciences, School of Public Health,
University at Albany, State University of New York, New York, Rensselaer
12144, USA. AJCC: American Joint Committee on Cancer; CCRT: Concurrent
chemoradiotherapy; DMFS: Distant metastasis-free survival; EBV: Epstein–barr
virus; ICRU: International Commission on Radiation Units; IMRT: Intensity-
modulated radiotherapy; IQR: Interquartile range; LRFS: Locoregional relapse-free
survival; NLR: Neutrophil-to-lymphocyte ratio; NPC: Nasopharyngeal carcinoma;
OS: overall survival; PFS: Progression-free survival; RDD: Research Data Deposit;
ROC: Receiver operating characteristic; TNM: Tumor-Node-Metastasis Received: 28 December 2017 Accepted: 20 December 2018 Ethics approval and consent to participate 9. An X, Ding PR, Wang FH, Jiang WQ, Li YH. Elevated neutrophil to
lymphocyte ratio predicts poor prognosis in nasopharyngeal carcinoma. Tumour Biol. 2011;32(2):317–24. 9. An X, Ding PR, Wang FH, Jiang WQ, Li YH. Elevated neutrophil to
lymphocyte ratio predicts poor prognosis in nasopharyngeal carcinoma. Tumour Biol. 2011;32(2):317–24. This study was conducted in compliance with institutional policy to protect
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between HPV and host immunity in head and neck squamous cell
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1 2014;134:2755–63. 26. Andersen AS, Koldjaer Solling AS, Ovesen T, Rusan M. The interplay
between HPV and host immunity in head and neck squamous cell
carcinoma. Int J Cancer. 2014;134:2755–63. 27. J S, Gerger A, Liegl–Atzwanger B, et al. The lymphocyte/monocyte ratio
predicts poor clinical outcome and improves the predictive accuracy in
patients with soft tissue sarcomas. Int J Cancer. 2014;135:362–70. 27. J S, Gerger A, Liegl–Atzwanger B, et al. The lymphocyte/monocyte ratio
predicts poor clinical outcome and improves the predictive accuracy in
patients with soft tissue sarcomas. Int J Cancer. 2014;135:362–70. 28. Thurner EM, Krenn–Pilko S, Langsenlehner U, et al. The elevated C-reactive
protein level is associated with poor prognosis in prostate cancer patients
treated with radiotherapy. Eur J Cancer. 2015;51:610–9. 28. Thurner EM, Krenn–Pilko S, Langsenlehner U, et al. The elevated C-reactive
protein level is associated with poor prognosis in prostate cancer patients
treated with radiotherapy. Eur J Cancer. 2015;51:610–9. 29. Chang H, Gao J, Xu BQ, et al. Haemoglobin, neutrophil to lymphocyte ratio
and platelet count improve prognosis prediction of the TNM staging
system in nasopharyngeal carcinoma: development and validation in 3,237
patients from a single institution. Clin Oncol. 2013;25(11):639–46.
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DOI: https://doi.org/10.2196/preprints.27020 p
DOI: https://doi.org/10.2196/preprints.27020 Shahil Kaini1*; Lucinda Zahrah Motie1* BSc 1University College London (UCL) London GB
*these authors contributed equally 1University College London (UCL) London GB
*these authors contributed equally
Corresponding Author:
Shahil Kaini
University College London (UCL)
Gower St, Bloomsbury
London
GB p
g
Shahil Kaini
University College London (UCL)
Gower St, Bloomsbury
London
GB Virtual vs Online: Insight from Medical Students.
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Corrosion of galvanic pairs of dental alloys copper base with silver amalgams in artificial saliva
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Corrosion of galvanic pairs of dental
alloys copper base with silver
amalgams in artificial saliva Corrosión de pares galvánicos de
aleaciones dentales base cobre
con amalgamas de plata en
saliva artificial Elvar Quezada-Castillo1, Wilder Aguilar-Castro1,
Bertha Quezada-Alván2 Elvar Quezada-Castillo1, Wilder Aguilar-Castro1,
Bertha Quezada-Alván2 Laboratory of Physics of Materials "Lennart Hasselgren", Dept. Academic of Physics. National University of Trujillo,
Trujillo, La Libertad, Peru. 2 Private University Antenor Orrego, Peru. Science Department., Trujillo, La Libertad, Peru. e-mail: elvarq@yahoo.es, wilderag@yahoo.com quezada_b@yahoo.es RESUMEN La corrosión galvánica es un proceso de disolución metálica que se produce cuando dos metales de distinto
potencial electroquímico se sumergen en un electrolito originando corrientes eléctricas entre los metales a
través del medio conductor. Para restaurar las piezas dentales perdidas o deterioradas se usan diferentes alea-
ciones quedando la boca expuesta a corrientes eléctricas que circulan a través de la saliva y los fluidos denta-
rios. En el presente trabajo se determinaron simultáneamente, los potenciales y densidades de corrien-tes de
corrosión de pares galvánicos de amalgamas de plata y aleaciones dentales base cobre usando méto-dos po-
tenciodinámicos, encontrando que los más resistentes a la corrosión son los pares formados por amalgamas
de plata de alto cobre y aleaciones Cu – Zn, y que los productos de corrosión liberados en ma-yor cantidad en
el medio electrolítico están formados por iones Hg, Cu, Zn y Ni. Palabras clave: Corrosión, aleación dental, curva de polarización, saliva artificial, par galvánico. ABSTRACT Galvanic corrosion is a metal dissolution process that occurs when two metals of different electrochemical
potential are immersed in an electrolyte causing electrical currents between the metals through the conduct-
ing medium. To restore lost or damaged teeth, different alloys are used, leaving the mouth exposed to elec-
trical currents that circulate through saliva and dental fluids. In the present work, the potentials and densities
of corrosion currents of galvanic pairs of silver amalgams and copper base dental alloys were determined
simultaneously using potenciodynamic methods, finding that the most resistant to corrosion are the pairs
formed by silver amalgams of high copper and Cu - Zn alloys, and that the corrosion products released in
greater quantity in the electrolytic medium are formed by Hg, Cu, Zn and Ni ions. Keywords. Corrosion, dental alloy, polarization curve, artificial saliva, galvanic pair. Autor Responsable: Elvar Quezada-Castillo 1 Laboratory of Physics of Materials "Lennart Hasselgren", Dept. Academic of Physics. National University of Trujillo,
Trujillo, La Libertad, Peru.
2 Private University Antenor Orrego, Peru. Science Department., Trujillo, La Libertad, Peru.
e-mail: elvarq@yahoo.es, wilderag@yahoo.com quezada_b@yahoo.es 2.1 Materials Three copper base dental alloys and two silver amalgams whose chemical composition are shown in table 1
have been used. Three copper base dental alloys and two silver amalgams whose chemical composition are shown in table 1
have been used. Table 1: Chemical composition of amalgams and dental alloys base copper by weight %. ALLOYS
TRADENAME
CODE
CHEMICAL COMPOSITION
Dental Mercury
UnoDent
Hg 99.99
Silver Amalgams
Low-copper dental amalgam Standalloy F
1
Ag 71.0 - Sn 25.7 - Cu 3.3
High-copper dental amalgam Duralloy S
2
Ag 45 - Sn 31 - Cu 24
Cu-Ni alloy
Aurocast
3
Cu 57.9 - Ni 22.1 - Al 15.0 - Fe 5.0
Cu-Zn alloy
Orodent
4
Cu 53.9 - Zn 45.9 - Fe 0.05 - Ni 0.02
Cu-Al alloy
Pentacast
5
Cu 77.9 - Al 12.0 - Ni 5.1 - Fe 5.0 1: Chemical composition of amalgams and dental alloys base copper by weight %. Table 1: Chemical composition of amalgams and dental alloys base copper by weight %. Table 1: Chemical composition of amalgams and dental alloys base copper by weight %. 2.2 Preparation of test pieces • The copper base samples were prepared by the lost wax method with an oxygen - butane - propane
flame and centrifugation processes. • The amalgams were crushed and condensed according to the manufacturer's instructions [14]. In both
cases the specimens were cylinders 0.5 cm high and 0.8 cm in diameter. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. physical properties, wear resistance or corrosion resistance and acceptable and stable appearance. The most
traditional are noble alloys containing not less than 75% gold and platinum group metals. The techniques of
casting, cast and finished are simple and do not suffer deterioration of their properties over time or lose their
aesthetic appearance. In contrast, they have the disadvantage of having high density, under elastic modulus
and are too expensive [3]. physical properties, wear resistance or corrosion resistance and acceptable and stable appearance. The most
traditional are noble alloys containing not less than 75% gold and platinum group metals. The techniques of
casting, cast and finished are simple and do not suffer deterioration of their properties over time or lose their
aesthetic appearance. In contrast, they have the disadvantage of having high density, under elastic modulus
and are too expensive [3]. For this reason, alloys of non-precious metals have been developed to replace high gold alloys, which
have been used since the 1940s in developed countries. Among these we have the ones of Co - Cr, Ni - Cr
and lately those of base titanium. In developing countries such as ours copper base alloys are used: Cu - Al,
Cu - Zn and Cu - Ni and amalgams of silver of low and high copper by its reduced cost, as has been men-
tioned in foreign publications [4] and manifested in presentations made by various researchers at the meet-
ings of the Argentine Branch of the International Association for Dental Research [5-7]. Two or more dissimilar metals in the mouth form batteries that generate galvanic currents, due to elec-
trolytic contact through the saliva or the root fluids; these pairs interact by direct, indirect or intermittent con-
tact [1]. Research has been carried out on galvanic pairs of alloys of Au, Co - Cr, Ni - Cr, titanium and stain-
less steel with amalgams [8-13]. In this work we will study the susceptibility to corrosion of galvanic pairs of
copper base alloys (Cu - Al, Cu - Zn, and Cu - Ni) with silver amalgams using potenciodynamic methods and
later determine the corrosion products by EDAX and SEM. 1. INTRODUCTION The dental alloys have application in the restoration and correction of lost or misaligned dental pieces to pre-
serve the correct masticatory function and the aesthetic aspect. The designs of these prostheses can take vari-
ous forms and all branches of dentistry such as restorative, orthodontic, endodontic, implantology, periodon-
tics and geriatrics apply these models [1,2]. Alloys are used for direct fillings (amalgams), crowns, bridges, inlays, partial or total prostheses, fixed
or mobile, implanted structures, or in the form of wires to correct defects in mastication using orthodontic
appliances. These alloys are usually as cast, but in certain applications welding or bonding with porcelain or
special polymers is required. To fulfill these functions the alloys must meet a variety of biocompatibility requirements, suitable Fecha de aprobación: 10/09/2018 Fecha de aprobación: 10/09/2018 10.1590/S1517-707620190001.0636 QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019.
Electrolyte - The electrolyte used in electrochemical tests is an experimental saliva that reproduces
the electrochemical behavior of natural saliva [17], whose formula is shown in Table 2. The electro-
lyte was prepared with deionized water of 18.2 MΩ-cm electrical resistivity and with reagents of an-
alytical grade. The pH of the solution was 6.5; to prevent this value from being modified, KHCO3
must be added shortly before starting the tests. Table 2: Chemical composition of experimental saliva [17]. Table 2: Chemical composition of experimental saliva [17]. 2: Chemical composition of experimental saliva [17]. REAGENT
CONCENTRATION (g/L)
NaCl
0.600
KCl
0.720
CaCl2.2H2O
0.220
KH2PO4
0.680
Na2HPO4.12H2O
0.856
KSCN
0.060
KHCO3
1.500
H8C6O7 (cítric acid)
0.030 3. RESULTS AND DISCUSSION 3.1 Electrochemical series QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. potential, it was waited 30 minutes for the potential to stabilize always under the action of the air
stream. Potentials were measured five times with different specimens. potential, it was waited 30 minutes for the potential to stabilize always under the action of the air
stream. Potentials were measured five times with different specimens. potential, it was waited 30 minutes for the potential to stabilize always under the action of the air
stream. Potentials were measured five times with different specimens. Polarization curves - The potentiodynamic polarization curves were plotted in triplicate using the
same cell and the same potentiostat as the one used to measure the electrode potential. The reference
electrode was saturated calomel and the platinum counter electrode. The scanning speed was 12 mV
/ min and the curves were recorded with an XT Princeton Applied Reserch Model REO 151 plotter. Before plotting the curves the corrosion potential was measured in the same manner as in the previ-
ous section. The global curves (anodic and cathodic) of each alloy were drawn with different speci-
mens. Once the tests were finished, the specimens were analyzed with a scanning electron micro-
scope to determine the corrosion products by EDAX. Potential corrosion of galvanic pairs - Dental alloys are corroded in the oral environment by the
action of saliva and bone fluids so that when electrically coupled both are polarized and corrode at a
new speed. When the metals A and B of equal areas are coupled, the mixed potential and the current
density (Ecupla and Icupla) of the galvanic cupla are at the intersection of the anodic curves of the
less noble metal (A) and cathodic of the metal plus noble (B) [16].
Electrolyte - The electrolyte used in electrochemical tests is an experimental saliva that reproduces
the electrochemical behavior of natural saliva [17], whose formula is shown in Table 2. The electro-
lyte was prepared with deionized water of 18.2 MΩ-cm electrical resistivity and with reagents of an-
alytical grade. The pH of the solution was 6.5; to prevent this value from being modified, KHCO3
must be added shortly before starting the tests. 2.3 Electrochemical methods To predict the corrosion of galvanic pairs in electrolytic solutions, measurements of potential, polarization
and current must be made [15]. To predict the corrosion of galvanic pairs in electrolytic solutions, measurements of potential, polarization
and current must be made [15]. Potential measurements - The electrode potential of each alloy was measured with a saturated cal-
omel electrode in a three-electrode cell and a Princeton Applied Research Potenciostat Model 173. Prior to the measurement, the specimen was introduced to the solution and at the same time air was
circulated at a rate of 80 bubbles per minute with a fish tank aerator. Before measuring the corrosion QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. Table 3: Corrosion Potentials and Average Standard Deviations. ALLOY
Vcorr(mVsce)
σ(mVsce)
Copper high amalgam
(Duralloy S)
-230
12
Orodent
-233
12
Aurocast
-252
5
Pentacast
-260
7
Copper under amalgam
(Standalloy F)
-381
18 Table 3: Corrosion Potentials and Average Standard Deviations. 3.1 Electrochemical series 3.1 Electrochemical series
Table 3 shows the average open circuit corrosion potentials of the studied alloys according to their resistance
to corrosion. The average standard deviation calculated according to G standard: 16-95 ASTM [18] is also
shown. Copper base alloys manufactured to substitute gold for their yellow color have high potential for cor-
rosion due to the large amount of copper they contain. In contrast, the Standalloy F amalgam has a low corro-
sion potential due to the small amount of copper it contains compared to the other alloys considered in this
work. Table 3 shows the average open circuit corrosion potentials of the studied alloys according to their resistance
to corrosion. The average standard deviation calculated according to G standard: 16-95 ASTM [18] is also
shown. Copper base alloys manufactured to substitute gold for their yellow color have high potential for cor-
rosion due to the large amount of copper they contain. In contrast, the Standalloy F amalgam has a low corro-
sion potential due to the small amount of copper it contains compared to the other alloys considered in this
work. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. The Cu-Al alloy presents a passive zone that extends from - 190 mVsce to 180 mVsce with an average
current density in this area of 2μA / cm2. Its breaking potential is 200 mVsce, from which dissolution of the
interdendritic structure of the alloy known as phase k occurs, formed by segregation of Fe and Ni due to its
insolubility in the α phase [23]. The Cu-Ni alloy has a passive zone extending from -160 mVsce to 200 mVsce
and a passivation current density of 3 μA / cm2. The breaking potential is 210 mVsce, from which the dissolu-
tion of the material takes place. The Cu-Zn alloy has a small passive zone that extends from -180 mVsce up to
20 mVsce and a passive current density of 1.50 μA / cm2. The reduction of this zone compared to the previous
alloys is due to its high Zn content[1]. From the rupture potentials the current densities of the copper base
alloys rapidly increase to small over potentials as shown in figure 1, due to the selective dissolution of the
material known as dealloyed. 3.2 Polarization curves Dental amalgams are galvanically corroded in the mouth due to the different phases that have: Ag-Hg, Ag-Sn
and Sn-Hg, each of which has a different electrochemical potential due to its diverse chemical composition
[19,20] Figure 1 shows the overall polarization curves of the alloys under study. The cathodic curves of these
alloys are similar; for small over cathodic potential the current density increases rapidly observing a linear
relationship between the potential envelope and the logarithm of the current. In all cases the cathodic region
corresponds to the evolution of hydrogen [21]. The low copper amalgam (Standalloy F) has a passive zone that extends from -370 mVsce to -100
mVsce and its breaking potential is -90 mVsce, from this value the current density increases rapidly. The amal-
gam of high copper (Duralloy S), not represented in all its extension, have a current density of 2.20 μA / cm2
at the height of the rupture potentials of aurocast and pentacast. The current density of the Standalloy passive
zone is 0.30 μA / cm2 obtained by averaging the current densities of its passive zone. The most corrosive phase of the low copper amalgam is the 2 or Sn-Hg phase, due to the formation of
tin oxychloride from the tin of the 2 phase and also the release of mercury [19]. The high copper quality
amalgams do not have phase 2 and the most corrosion resistant phase of these alloys is the Ag-Hg phase
[22]. Figure 1. Polarization curves of silver amalgams and copper base alloys. Figure 1. Polarization curves of silver amalgams and copper base alloys. * indicates that the copper base alloy is corroded. * indicates that the copper base alloy is corroded. QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. In Table 4 we observe that the range of the corrosion potentials of the galvanic pairs of the alloys un-
der study with Standalloy is between - 299 mVsce and -247 mVsce and the current densities between 0.244 μA
/ cm2 and 0.142 μA / cm2. For Duralloy-formed pairs the corrosion potentials are between -255 mVsce
and -232 mVsce and the current densities between 0.055 μA / cm2 and 0.004 μA / cm2. 3.3 Potentials and densities of corrosion current of galvanic pairs The potential and corrosion current densities of the galvanic pairs under study are determined from the inter-
section of the cathodic curves of the silver amalgams and the anodic curves of the copper base alloys, as
shown in figure 2. The results are presented in table 4. Figure 2: Intersection of anodic curve of Standalloy and cathodic of Duralloy with the anodic and cathodic curves of
copper base dental alloys. Figure 2: Intersection of anodic curve of Standalloy and cathodic of Duralloy with the anodic and cathodic curves of
copper base dental alloys. Table 4: Potentials and corrosion current densities of galvanic pairs of copper base dental alloys with silver amalgam : Potentials and corrosion current densities of galvanic pairs of copper base dental alloys with silver ama
GALVANIC COUPLES NOTATION
Vcorr.cupla(mVsce)
Icorr.cupla(µA/cm2)
Standalloy/Duralloy (PG1,2)
-287
0.165
Standalloy/Aurocast (PG1,3)
-299
0.142
Standalloy/Orodent (PG1,4)
-247
0.244
Standalloy/Pentacast (PG1,5)
-264
0.216
Duralloy/Aurocast (PG2,3)
-244
0.055*
Duralloy/Orodent (PG2,4)
-232
0.004*
Duralloy/Pentacast (PG2,5)
-255
0.008*
* indicates that the copper base alloy is corroded. 3.4 Corrosion products of galvanic pairs At the end of the tests the corrosion products were observed with a scanning electron microscope (Phillips
500) and by making a general sweep of the surface of the specimens by EDAX the corrosion products whose
spectra are shown in figures 3 to 7 were qualitatively determined. Figures 3 and 4 show the SEM micrographs of the corrosion products and the EDAX spectra of the
silver amalgam of low and high copper with high peaks of Hg, Sn, Ag, Cl and Cu and small peaks of P, Si
and Ca. in agreement with the results of other researchers [24,25]. Since this technique does not register oxy-
gen or hydrogen, it is assumed that oxides, hydroxides, phosphates and chlorides are formed from these ele-
ments, such as those identified by Fathi and Mortazavi in dental patients [19]. Figures 5 to 7 show the SEM micrographs of the corrosion products and the EDAX spectra of the Cu
base alloys, showing high peaks of Cu and Zn and small peaks of Al, Ni and Fe. The analysis of the spectra
and the bibliographic references [26,27], indicate that oxides, hydroxides, phosphates and copper chlorides
are formed, mainly, and aluminum, iron and nickel, in a minority form. The ions released in corrosion processes can be stored in the mucosa in the form of staining or pass
through the digestive tract to the stomach. Some of them are eliminated abroad and the rest is stored in some
organ such as liver, kidneys, lungs or brain [28]. Cu ions are released when copper amalgams and dental alloys dissolve and are stored in the gingival
tissue adjacent to the restored tooth. These ions affect the viability and proliferation of lymphocytes. Over
time the concentration of copper increases in the gums, producing inflammation, alteration of cellular im-
munity and oral homeostasis. In general, these ions contribute to a variety of immunopathological conditions
including periodontal diseases and decreased resistance to oral infections [29]. Figure 3: SEM micrograph and EDAX spectra of corrosion products on Standalloy (× 1440). Figure 3: SEM micrograph and EDAX spectra of corrosion products on Standalloy (× 1440). QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. Figure 4: SEM micrograph and EDAX spectra of corrosion products on Duralloy (× 1280). Figure 4: SEM micrograph and EDAX spectra of corrosion products on Duralloy (× 1280). 4. CONCLUSIONS From the results obtained in the present work it can be established that: From the results obtained in the present work it can be established that: 1. Galvanic pairs of copper base dental alloys with high copper silver amalgams are more resistant to corro-
sion than pairs formed with low copper silver amalgams. The most resistant pair is PG2,4 (Duralloy /
Orodent) and the less resistant the pair PG1,5 (Standalloy / Pentacast). 1. Galvanic pairs of copper base dental alloys with high copper silver amalgams are more resistant to corro-
sion than pairs formed with low copper silver amalgams. The most resistant pair is PG2,4 (Duralloy /
Orodent) and the less resistant the pair PG1,5 (Standalloy / Pentacast). 2. All dental alloys release ions into the oral environment. Some of these diffuse into the buccal mucosa and
the rest pass into the gastrointestinal system. They are then distributed and stored in different parts of the
body, such as brain, lungs, liver and kidneys affecting the health of patients, especially when the concen-
tration of corrosion products (Hg, Cu, Ni, etc.) exceeds normal tolerance limits. 2. All dental alloys release ions into the oral environment. Some of these diffuse into the buccal mucosa and
the rest pass into the gastrointestinal system. They are then distributed and stored in different parts of the
body, such as brain, lungs, liver and kidneys affecting the health of patients, especially when the concen-
tration of corrosion products (Hg, Cu, Ni, etc.) exceeds normal tolerance limits. 3.4 Corrosion products of galvanic pairs Figure 5: SEM micrograph and EDAX spectra of corrosion products on Pentacast (× 1440). Figure 5: SEM micrograph and EDAX spectra of corrosion products on Pentacast (× 1440). QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. Figure 6: SEM micrograph and EDAX spectra of corrosion products on Orodent (× 1200). Figure 6: SEM micrograph and EDAX spectra of corrosion products on Orodent (× 1200). Figure 7: SEM micrograph and EDAX spectra of corrosion products on Aurocast (× 1120). Figure 7: SEM micrograph and EDAX spectra of corrosion products on Aurocast (× 1120). The major component mercury of amalgams is highly toxic and very permeable through the cell mem-
brane, being able to pass from the gastrointestinal tract to the lungs and brain. It has been found "mercury in
the hairs, nails and a high concentration in the pituitary gland (approximately 70 times higher than in the con-
trol groups) in people who have amalgams in the mouth" [29]. Nickel is a potential allergic, and is the most
common cause of allergic contact dermatitis, it is present in most jewelry and household utensils. Therefore,
it can be considered a danger to public health increasing with the use of dental restorations. Ni ions produce
nonspecific allergies and inflammations around restorations that can take lichenoid and sometimes erosive QUEZADA-CASTILLO, E; AGUILAR-CASTRO, W; QUEZADA-ALVÁN, B. revista Matéria, v. 24, n. 1, 2019. reactions, with women being more hypersensitive to this metal because of the jewelry that they use from an
early age [30,31]. 5. ACKNOWLEDGMENT The Materials Laboratory of the National Commission for Atomic Energy (Constituents) of the Republic
Argentina, for allowing us to use their equipment in sections of Corrosion and Electron Microscopy. The Materials Laboratory of the National Commission for Atomic Energy (Constituents) of the Republic
Argentina, for allowing us to use their equipment in sections of Corrosion and Electron Microscopy. 5] BABOIAN, R., “Electrochemical techniques for predicting galvanic corrosion”. In: BOBOIAN, 5. BIBLIOGRAPHY In: BOBOIAN, R, FRANCE, W.D J.R, ROEW, L.C., et al., Galvanic and Pitting – field and Laboratory Studies. ASTM stp 556. Philadelphia: American Society for Testing and Materials, pp 6- 9, 1976. [16] JONES DENNY A. Principles and Prevention of Corrosion. 2 º Ed. Prentice Hall, Upper Saddle Rivers,
N.J., pp. 177, 1996. [17] DUFFO,G.S., QUEZADA, E., “Development of an Artificial Saliva Solution for Studying the Corrosion
Behavior of Dental Alloys”, Corrosion, v. 60, pp. 594–602, Jun. 2004. [18] ASTM Standars: G.16-95. “Standars guide for applying statistics to analysis of corrosion dat”. In, Anual
Booke of ASTM Standards, v. 03, pp.02, 1999. [19] FATHI, M., MORTAZARVI,V., “A review on Dental Amalgam Corrosion and Its Consecuences”,
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study of Ag-Hg, Ag-Sn and Sn-Hg phases”, Electrochemica Acta, v.42, n. 1, pp.3887-3893, Jan. 2001. [21] GONZALES FERNANDEZ, G.A., Control de la corrosión: estudio y medida por técnicas electroquí-
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Metalúrgicas, pp. 97, Madrid ,1989. [22] BRETT, C. M. A., TRANDAFIR, F., “The corrosion of dental amalgam in artificial salivas: an electro-
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Dental Amalgam”, J Dent Res., v. 54, n. 5, pp. 1031-1038, Sep.-Oct. 1975. [25] FINKELSTEIN, G. F., GREENER, E. H., “In vitro polarization of dental in human saliva”, Journal of
Oral Rehabilitation, v.04, n.4, pp.347-354, Oct. 1977. [26] MUELER, H., “The effects of a human salivary dialysate upon ionic release and electrochemical corro-
sion of a Copper-Aluminim alloy”, Journal Electrochemical Society, v. 134, n. 3, pp. 575-580, Mar. 1987. [27] SARKAR, N. K., “SEM and x-ray microanalysis of in vitro corrosion of a copper-zinc alloy”, Journal of
Prosthetic Dentistry, v. 45, n. 3, pp. 292-295, Mar. 1981. [28] BRUNE, D., GJERDET, N., PAULSEN, G., “Gastrointestinal and in vitro release of copper, cadmium,
indium, mercury and zinc from conventional and copper-rich amalgams”, Scand. J. Dent. Res., v. 91, n. 1, pp. 66 -71, Feb. 1983. 5. BIBLIOGRAPHY [1] MUELLER, H. J., “Tarnish and Corrosion of Dental Alloys”, In Metal Handbook, v. 13, Ohio, Ed. ASM
International Metal Park, pp. 1336 - 1366, 1992. [2] PHILLIPS, R. W., La Ciencia de los Materiales Dentales, 9° Edición, Cap. 16, Editorial Interamericana,
México, 1988. [3] Guide to Dental Materials and Device, American Dental Association, 7 th Ed., Chicago, 1974. [4] NAKAYAMA, M., MOSER, J. B. AND GREENER, E.H. “Corrosion and Tarnish of Aluminum Bronze”,
Dent. Mater. J., v.3, pp.133, 1984. [5] Proc. XXIII Reunión Anual de la División Argentina de la International Association for Dental Research
(I.A.D.R.), Buenos Aires, 1990. [6] Proc. XXIV Reunión Anual de la División Argentina de la International Association for Dental Research
(I.A.D.R.), Villa Giordano (Córdoba), 1991. [7] Proc. XXV Reunión Anual de la División Argentina de la International Association for Dental Research
(I.A.D.R.), Paraná (Entre Ríos), 1992. [8] HALLAND, R.I., “Galvanic currents between gold and amalgam”, Scand. J Dent Res. v.88, pp. 269-272,
1980. ] RAVNHOL, G., “Corrosion Current and PH rise around titanium coupled to dental alloys”, Scand. J. Re
96, pp. 466-472, 1988. 0] RAVNHOLT, G., JENSEN, J., “Corrosion investigation of two materials for implant supraconstructio
oupled to a titanium implant”, Dent. Res. v.99, pp. 181-186, 1991. [10] RAVNHOLT, G., JENSEN, J., “Corrosion investigation of two materials for implant supraconstructions
coupled to a titanium implant”, Dent. Res. v.99, pp. 181-186, 1991. [11] GRIFFIN CH
BUCHANAN R A
LEMOUS J E
“In vitro electrochemical corrosion study of cou- coupled to a titanium implant”, Dent. Res. v.99, pp. 181-186, 1991. [11] GRIFFIN, CH., BUCHANAN,R.A., LEMOUS, J.E., “In vitro electrochemical corrosion study of cou-
pled surgical implant materials” J of Biomedical Mat Res v 17 pp 489
500 1996 1] GRIFFIN, CH., BUCHANAN,R.A., LEMOUS, J.E., “In vitro electrochemical corrosion study of co
ed surgical implant materials”, J. of Biomedical Mat. Res. v.17, pp. 489 – 500, 1996. [12] CISZEWSKI, A., BARANIAK, M., URBANEK-BRYCHCZYNSKA, M., “Corrosion by Galvanic
Coupling Between Amalgam and Different Chromium-Based Alloys”, Dental Materials, v. 23, pp. 1256 –
1261, Dec. 2007. [13] CAPELO, S., PROENCA, L., FERNANDEZ, J., et al., “Galvanic Corrosion of Two Non Noble Dental
Alloys”, Int. J. Electrochem. Sci., v. 9, pp. 593 – 609, Sep. 2014. [14] COIMPA - Sociedade Industrial de Metais Preciosos da Amazonia Ltda. 11. Instrucciones para uso de
amalgamas, Brasil. [15] BABOIAN, R., “Electrochemical techniques for predicting galvanic corrosion”. 5. BIBLIOGRAPHY [29] BUMGARDNER, J.D., LUCAS, L.C., ALVERSON, M.W., et al., “Effects of copper-based casting
alloys on two lymphocyte cell lines and the secretion of interleukin 2 and Ig”, Dent. Mater.,v. 9, pp. 85 – 90,
1993. [30] BURROWS, D. “Hypersensitivity to mercury, nickel and chromium in relation to dental materials”, In-
ternational Dental Journal, v. 36,n. 1, pp. 30-34, Mar. 1986. [31] BLANCO-DALMA, L., CARRASQUILLO-ALBERT, H., SILVA PARRA, J. “A study of nickel aller-
gy”, J. Prosth Dent, v. 52, n. 1, pp. 116-119, Jul. 1984. Elvar Fortunato Quezada Castillo http://orcid.org/0000-0002-5851-6806
Wilder Máximo Aguilar Castro http://orcid.org/ 0000-0001-7919-2014
Bertha Elba Quezada Alvan http://orcid.org/ 0000-0003-0504-4566 ORCID Elvar Fortunato Quezada Castillo http://orcid.org/0000-0002-5851-68
Wilder Máximo Aguilar Castro http://orcid.org/ 0000-0001-7919-20
Bertha Elba Quezada Alvan http://orcid.org/ 0000-0003-0504-45
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http://irep.iium.edu.my/89293/1/Performance%20Comparison%20of%20the%20Single%20Axis%20and%20Two-Axis%20Solar%20System%20using%20Adaptive%20Neuro-Fuzzy%20Inference%20System%20Controls.pdf
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Performance Comparison of the Single Axis and Two-Axis Solar System using Adaptive Neuro-Fuzzy Inference System Controls
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E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 1Department of Mathematic, Sepuluh Nopember Institute of Technology, Jl. Teknik Kimia,
Surabaya 60111, East Java, Indonesia
2Department of Engineering Physics, Sepuluh Nopember Institute of Technology, Jl. Teknik Kimia
Surabaya 60111, East Java, Indonesia
3Department of Biochemical-Biotechnology Engineering, Kuliyyah of Engineering
International Islamic University Malaysia (IIUM), Jl. Gombak, 53100, Selangor, Malaysia
4Graduate School of Renewable Energy, Darma Persada University, Jl. Taman Malaka
Selatan No. 22, Pondok Kelapa, East Jakarta 13450, Indonesia 1Department of Mathematic, Sepuluh Nopember Institute of Technology, Jl. Teknik Kimia,
Surabaya 60111, East Java, Indonesia
2Department of Engineering Physics, Sepuluh Nopember Institute of Technology, Jl. Teknik Kimia,
Surabaya 60111, East Java, Indonesia
3Department of Biochemical-Biotechnology Engineering, Kuliyyah of Engineering
International Islamic University Malaysia (IIUM), Jl. Gombak, 53100, Selangor, Malaysia
4Graduate School of Renewable Energy, Darma Persada University, Jl. Taman Malaka
Selatan No. 22, Pondok Kelapa, East Jakarta 13450, Indonesia Abstract. Solar energy is one of the renewable energy that gets more
attention from many countries. Solar photo voltaic (PV) takes the right
position to get the maximum energy yield. The study was conducted by
comparison of performance with two methods of tracking the sun with one
axis and two axes by using ANFIS control (Adaptive Neuro-Fuzzy
Inference System). The solar tracking system is a system that operates on
the sun by using a light sensor and controls the photovoltaic to always
perpendicular to the sun by changing the pitch and yaw axis of the sun
tracing properties. LDR (Light Dependent Resistor) is one of the light
sensors whose resistance changes depending on the intensity of incoming
light. Direct current (DC )motor is used as a PV drive panel in a solar
tracking system. A two-axis solar tracking system has a greater power
output than a tracking system with a single photovoltaic panel that does not
use a tracking system (fixed). Keywords: Direct current motors, light sensors, performance index,
photovoltaic, pitch, yaw 1 Introduction Solar energy is one of the renewable energy that gets more attention from many countries
[1]. According to law number 30, the year 2007 article 29 paragraph 2 about the energy, the
development of energy directed to the development of renewable energy [2]. According to
data from IRENA [3] (The International Renewable Energy Agency) at 2015 the
distribution of solar photovoltaic installations in Indonesia is only 0.1 % of the total Performance Comparison of the Single Axis and
Two-Axis Solar System using Adaptive Neuro-
Fuzzy Inference System Controls
Chairul Imron1, Imam Abadi2,*, Ilham Amirul Akbar2, Jauharotul Maknunah2,
Yusilawati Ahmad Nor3, and Aep Saepul Uyun4 Chairul Imron1, Imam Abadi2,*, Ilham Amirul Akbar2, Jauharotul Maknunah2,
Yusilawati Ahmad Nor3, and Aep Saepul Uyun4 © The Authors, published by EDP Sciences. This is an open access article distributed under the terms of the Creative Commons
Attribution License 4.0 (http://creativecommons.org/licenses/by/4.0/). * Corresponding author: imamabadi02@gmail.com * Corresponding author: imamabadi02@gmail.com © The Authors, published by EDP Sciences. This is an open access article distributed under the terms of the Creative Commons
Attribution License 4.0 (http://creativecommons.org/licenses/by/4.0/). E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 installed renewable energy technology, the world alone is installed at 11.2 % of the overall
renewable energy technology installed, while Indonesia is a country that is on the equator
which is always sunlit throughout the year. In addition, the range of the year 2010 to 2015
decreased the price of solar PV modules by 75 % to 80 %. installed renewable energy technology, the world alone is installed at 11.2 % of the overall
renewable energy technology installed, while Indonesia is a country that is on the equator
which is always sunlit throughout the year. In addition, the range of the year 2010 to 2015
decreased the price of solar PV modules by 75 % to 80 %. Solar PV needs the right position to get maximum energy yield [4, 5]. The solar tracker
can implement with one-axis solar tracking if desired higher accuracy can use two-axis
solar tracking. Two-axis solar tracking has two kinds that are solar tracking and azimuth
tracking [6]. Solar tracker not always needs to point right to the sun; when it missed 10 %,
the resulting output is still 98.5 % of that produced by solar tracker that is always pointing
towards the sun. Cloudy areas, the gain is about 20 % annually, while in bright areas, it can
reach 30 % to 40 % annually [7]. ANFIS stands for an adaptive neuro-fuzzy interface
system. ANFIS is a method with a combination between the neural network and the fuzzy
system. Some data will be trained firstly then implied to the fuzzy system [8]. In previous studies, the solar tracking system is used in a tropical area. The study
conducted in 2015 showed that a single axis solar tracking system, when used in the
tropical area, resulted in a more extensive power conversion of about 25 % compared to
fixed panels [9]. Research in 2012 in Malaysia, the performance ratio of P-Si type
photovoltaics of 59.40 %, uses a solar tracker system with one axis [10]. * Corresponding author: imamabadi02@gmail.com Therefore, it is
necessary to compare the performance between two methods of tracking the sun that is with
one axis and two-axis using ANFIS (Adaptive Neuro-Fuzzy Inference System) control in
the tropical area, so it can show the difference in performance produced by the solar tracker
system with one axis and two-axis solar tracking. The research will be done by simulating one axis and two-axis solar tracking system
with ANFIS (Adaptive Neuro-Fuzzy Inference) control with active sun-tracking mode. The
single-axis solar tracking system uses two LDR, each on the east and west. Two-axis solar
tracking system uses four LDR, each on the north, south, east, and west. Use
170 mm × 250 mm paper size (W × H mm) and adjust the margins to those shown in the
Table 1. The final printed area will be 130 mm × 210 mm. Table 1. Parameters of DC motor. Table 1. Parameters of DC motor. Motor Parameters
Parameter Value Motor
Ke
12.843 69 V s rad–1
Kt
12.844 37 Nm A–1
R
22.424 5 Ω
L
0.006 386 Henry
Jm
0.723 779 Nm s2
Bm
3.434 949 Nm s rad–1 2.1. Solar tracking system The solar tracking system is a system that acts to detect the sun's position by means of a
light sensor and controls the photovoltaic so that it is always perpendicular to the sun by
changing the pitch and yaw axis of a solar tracker. The purpose of the solar tracking system
is to keep photovoltaic always perpendicular to the sun, and it can optimize the output
power of photovoltaic. The output power of the photovoltaic will produce greater energy 2 2 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 than the photovoltaic that is not always perpendicular to the sun. Sunlight comprises several
sensor components, controller, DC motor, battery, and photovoltaic [11]. than the photovoltaic that is not always perpendicular to the sun. Sunlight comprises several
sensor components, controller, DC motor, battery, and photovoltaic [11]. 2.2. Light sensor (LDR) The node function as follows in Equation (2): 𝑂𝑖
1 = 𝜇𝐴𝑖(𝑥) (2) (2) Where x is the input for node i and Ai is the language naming associated with the node
function for the Gaussian function (commonly used), which is specified in Equation (3):
2 Where x is the input for node i and Ai is the language naming associated with the node
function for the Gaussian function (commonly used), which is specified in Equation (3):
2 (3) (
y
)
p
q
( )
(3)
μ𝐴𝑖 𝑥 = exp[−0.5 𝑥−𝑐𝑖
𝜎𝑖
2
] Where 𝜎𝑖,𝑐𝑖 is set parameter. The parameters in this layer are shown as the premise
parameter. The second layer has a non-adaptive node whose parameter values are fixed. The function of this node is as a multiplication of each input signal. The function of the
node is as in Equation (4):
𝜎𝑖 (4)
𝑂𝑖
2 = 𝑤𝑖= 𝜇𝐴𝑖 𝑥 × 𝜇𝐵𝑖 𝑥 ,
𝑖= 1.2 (4) Each output node of this layer represents the degree of activation for each rule. ch output node of this layer represents the degree of activation for each rule. This third layer is a non-adaptive node that can display the function of the normalized
firing strength that is the output ratio of the i-th nodes in the previous layer to the entire
output of the previous layer. The function of the node is as in Equation (5): (5)
𝑂𝑖
3 = 𝑤 =
𝑤𝑖
𝑤1 + 𝑤2
,
𝑖= 1.2 (5) The output of this layer is called normalized firing strength. The output of this layer is called normalized firing strength. h
f
h l
ll h
d
d
i
h
li
d
i
i
d In the fourth layer, all the nodes are adaptive to the normalized activation degree of
layer 3. The function of the node is as in Equation (6): (6)
𝑂𝑖
4 = 𝑤 𝑖𝑓𝑖= 𝑤 𝑖(𝑎𝑖
1𝑥+ 𝑎𝑖
2𝑦+ 𝑎𝑖
0) (6) This fifth layer contains only one non-adaptive node with a function to sum all
inputs. The function of the node is as in Equation (7): (7)
𝑂𝑖
5 = 𝑤 𝑖𝑓𝑖
𝑖
=
𝑤𝑖𝑓𝑖
𝑖
𝑤𝑖
𝑖 (7) 2.2. Light sensor (LDR) LDR (Light Dependent Resistor) is one of the light sensors whose resistance changes
depending on the intensity of incoming light. LDR is used to convert light energy into
electrical energy. An LDR is strung together with one resistor whose value can be changed
and one resistor whose value is fixed. This series circuit has functioned as a voltage divider. The voltage divider in Equation (1): (1)
𝑉𝑜𝑢𝑡=
𝑅2
𝑅1 + 𝑟2𝑉𝑖𝑛 (1) Fig. 1. Voltage devider and LDR. 2.3. ANFIS architecture
Fig. 2. Voltage Divider and LDR. ANFIS stands for Adaptive Neuro-Fuzzy Inference System. ANFIS is also
amalgamation of an artificial neural network method with a fuzzy system. Adapti
networks are usually used in a variety of modeling, decision-making, controlling, and sig
processing applications. The adaptive network class is functionally equivalent to a fuz
system. The architecture of this is called ANFIS. The set parameters are set to apply hyb
𝑉𝑜𝑢𝑡
𝑅1 + 𝑟2𝑉𝑖𝑛 Fig. 1. Voltage devider and LDR. Fig. 1. Voltage devider and LDR. 2.3. ANFIS architecture
Fig. 2. Voltage Divider and LDR. 2.3. ANFIS architecture Fig. 2. Voltage Divider and LDR. ANFIS stands for Adaptive Neuro-Fuzzy Inference System. ANFIS is also an
amalgamation of an artificial neural network method with a fuzzy system. Adaptive
networks are usually used in a variety of modeling, decision-making, controlling, and signal
processing applications. The adaptive network class is functionally equivalent to a fuzzy
system. The architecture of this is called ANFIS. The set parameters are set to apply hybrid 3 3 3 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 learning rules for ANFIS architecture that represent both Sugeno and Tsukamoto fuzzy
models. The layers in the ANFIS architecture [8, 12] shown in Figure 2. learning rules for ANFIS architecture that represent both Sugeno and Tsukamoto fuzzy
models. The layers in the ANFIS architecture [8, 12] shown in Figure 2. The first layer has adaptive nodes whose parameter values may change with learning. 3.1 System modelling Modeling of one-axis and two-axis solar tracking system is done by using
MATLAB/Simulink to perform simulation, using component parameters to obtain the
simulation of one-axis and two-axis solar tracking system. The LDR sensor is a sensor that
converts light energy into electricity. It is obtained from changes in the resistance found in
the LDR sensor when exposed to sunlight. In the modeled photovoltaic, the output voltage
of the fixed resistor is 1 000 Ω. The relationship equation between irradiance and resistance
to LDR sensor can be explained as in Equation (8): (8)
Where:
[12]. = Resistance of LDR sensors LDR (Ω)
I
= Irradiance into the LDR sensor (W/m2)
log𝑅= −3
4 log𝐼+ 5 (8)
log𝑅= −3
4 log𝐼+ 5 (8) [12]. = Resistance of LDR sensors LDR (Ω)
I
= Irradiance into the LDR sensor (W/m2) 4 https://doi.org/10.1051/e3sconf/202019000005 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 ICoRER 2019 Fig. 3. LDR sensor modeling. 3.1.1 Modeling of DC motor The DC motor is the driving force of the PV panel to keep up with the sun. One-axis solar
tracking system uses one motor with pitch angle tracking, while the two-axis sun tracking
system uses two motors with tracking pitch and yaw angles. The pitch angle moves
between the angles of 0° and 180° and the yaw angle moves between angles 0° and 360° at
low speeds. DC motors can also be used to drive PV panels in a solar tracking system. The
advantages of this DC motor rotation speed can be set so that PV panels can follow the sun
precisely. The modeling of a DC motor, which converts the voltage value to rpm speeds is
as in Equation (9) and Equation (10), follows [13]: (
),
[
]
𝑉𝑎 𝑡 = 𝑅𝑎. 𝐼𝑎 𝑡 + 𝐿𝑎. 𝑑𝐼𝑎(𝑡)
𝑑𝑡
+ 𝐾𝑏. 𝜔(𝑡)
(9)
𝐾𝑡. 𝐼𝑎 𝑡 = 𝐽𝑚
𝑑𝜔(𝑡)
𝑑𝑡
+ 𝐵𝑚. 𝜔(𝑡)
(10) (9) (10) Then, the equations of voltage and current in DC motors are transformed in Laplace
transformation: Then, the equations of voltage and current in DC motors are transformed in Laplace
transformation: 𝑉𝑎 𝑠 = 𝑅𝑎. 𝐼𝑎 𝑠 + 𝐿𝑎. 𝐼𝑎 𝑠 𝑠+ 𝐾𝑏. 𝜔(𝑠) (11)
𝐾𝑡.𝐼𝑎 𝑠 = 𝐽𝑚.𝜔 𝑠 𝑠+ 𝐵𝑚. 3.1.1 Modeling of DC motor 𝜔(𝑠) (12) (11) (12) After that, the substitution result is: After that, the substitution result is: After that, the substitution result is: After that, the substitution result is: (13)
𝑉𝑎 𝑠 = 𝜔 𝑠 ∙1
𝐾𝑡
∙ 𝐿𝑎∙𝐽𝑚∙𝑠2 + 𝑅𝑎∙𝐽𝑚+ 𝐿𝑎∙𝐵𝑚 𝑠+ (𝑅𝑎∙𝐵𝑚+ 𝐾𝑏∙𝐾𝑡) (13) With the relationship value between angle and speed is
𝜃 𝑠 = 1
𝑠𝜔(𝑠) , the relationship
between voltage and angle is as in Equation (14) follows: With the relationship value between angle and speed is
𝜃 𝑠 = 1
𝑠𝜔(𝑠) , the relationship
between voltage and angle is as in Equation (14) follows: 𝜃(𝑠)
𝑉𝑎(𝑠) =
𝐾𝑡
𝐿𝑎∙𝐽𝑚∙𝑠3 + 𝑅𝑎∙𝐽𝑚+ 𝐿𝑎∙𝐵𝑚 𝑠2 + 𝑅𝑎∙𝐵𝑚+ 𝐾𝑏∙𝐾𝑡 𝑠
(14) (14) Where: Va = Armature Voltage (V) Va = Armature Voltage (V) = Armature Voltage (V)
= Armature Resistance (R) g ( )
Ra = Armature Resistance (R) Ra = Armature Resistance (R) Ra = Armature Resistance (R) 5 5 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 La = Armature Inductance (H)
Ia = Armature Current (A)
ω
= Angular Speed (rad s–1)
θ
= Position of Angular Shaft Rotor
(rad) Jm = Motor Inertia (Nm s2)
Bm = Viscous Friction Coefficient (Nm s rad–1)
Kt = Torque Constant (Nm A–1)
Kb = Back EMF Constant (V s rad–1)
The parameters of DC motor are obtained with the following Equation (15 to 17) [14]; (
)
The parameters of DC motor are obtained with the following Equation (15 to 17) [14]; 𝐾𝑒=
𝑉𝑜𝑙𝑡𝑎𝑔𝑒 [𝑉]
𝑉𝑒𝑙𝑜𝑐𝑖𝑡𝑦 [𝑟𝑎𝑑
𝑠]
(15)
𝐾𝑡=
𝐾𝑒
0.118 27 𝑁𝑚
𝐴
(16)
𝐽𝑚
𝑑𝜔(𝑡)
𝑑𝑡
+ 𝐵𝜔 𝑡 = 𝑇𝑚(𝑡)
(17) (15) (16) 𝐽𝑚
𝑑𝜔(𝑡)
𝑑𝑡
+ 𝐵𝜔 𝑡 = 𝑇𝑚(𝑡)
(17) (17) This DC motor has connected gearbox with ratio1:348.079 2. After the DC motor data were
taken and the calculation of DC motor parameters according to equations, we obtain the
following parameter values: This DC motor has connected gearbox with ratio1:348.079 2. After the DC motor data were
taken and the calculation of DC motor parameters according to equations, we obtain the
following parameter values: 3.1.2 Photovoltaic modeling Photovoltaic modeling is done by using photovoltaic parameters and current and voltage
output equations on photovoltaic, as for the following Equation (18) and Equation (19)
[14]: 𝑉= 𝑉𝑛−Φ T −𝑇𝑟𝑒𝑓 −𝑅𝑝𝑣(𝐼−𝐼𝑛)
𝐼= 𝑠
𝑠𝑟𝑒𝑓
𝜇 𝑇−𝑇𝑟𝑒𝑓 + 1 −𝐼𝑠𝑐 + 𝐼𝑛
(18)
(19) 𝐼= 𝑠
𝑠𝑟𝑒𝑓
𝜇 𝑇−𝑇𝑟𝑒𝑓 + 1 −𝐼𝑠𝑐 + 𝐼𝑛
(18) (18) 𝑉= 𝑉𝑛−Φ T −𝑇𝑟𝑒𝑓 −𝑅𝑝𝑣(𝐼−𝐼𝑛)
(19) (19) Table 2 Parameter of photovoltaic. Table 2 Parameter of photovoltaic. No
Parameter
Parameter Value
1
Short Circuit Current, Isc
1.25 A
2
Open Circuit Current, Voc
21.5 V
3
Temperature Coefficient of Isc, µ
0.000 05 A °C–1
4
Temperature Coefficient of Voc, ɸ
-0.003 6 V °C–1
5
Refrence Voltage, Vn
17.2 A
6
Reference Current , In
1.16 A
7
Maximum Power at STC
20 W
8
PV Internal Resistance, Rpv
1.11 Ω 6 https://doi.org/10.1051/e3sconf/202019000005 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 3.2 Design of ANFIS control ANFIS is a combination of neural networks and fuzzy logic. Artificial neural networks that
will determine the membership function and rule base of fuzzy logic. ANFIS requires a pair
of train data to be trained on artificial neural networks. The input of this training data is
error and delta error while the output of this training data is PWM. The ANFIS control uses
the membership function with the type of Gauss and the membership number of five
represents Clockwise Fast (CWF), Clockwise Slow (CWS), Stop (S), Counter Clockwise
Slow (CCWS) and Counter Clockwise Fast (CCWF). The Membership Function of the
input before being trained is as shown in Figure 4. The membership function of the input
after being trained from each ANFIS control for DC motor control on the pitch and yaw
axis as shown in Figure 5 and Figure 6. axis as shown in Figure 5 and Figure 6. Fig. 4. Membership function of error and delta error on pitch and yaw axis. (a)
(b)
Fig. 5. Membership function of (a) Error and (b) Delta error on pitch axis. Fig. 4. Membership function of error and delta error on pitch and yaw axis. Fig. 4. Membership function of error and delta error on pitch and yaw axis. Fig. 4. Membership function of error and delta error on pitch and yaw axis. (a) (a) (b)
Fig. 5. Membership function of (a) Error and (b) Delta error on pitch axis. Fig. 5. Membership function of (a) Error and (b) Delta error on pitch axis. 7 https://doi.org/10.1051/e3sconf/202019000005 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 (a)
(b)
Fig. 6. Membership function of (a) Error and (b) Delta error on yaw axis. (a) (a) (a)
(b)
Fig. 6. Membership function of (a) Error and (b) Delta error on yaw axis. (b) Fig. 6. Membership function of (a) Error and (b) Delta error on yaw axis. 4 Result and discussion In the test setpoint, use three setpoint angles of sun elevation and three setpoint angle
azimuth suns. For altitude angles the sun uses setpoint angle 30°, 60°, and 90° while for the
azimuth angle of the sun use setpoint angle 30°, 60°, and 90°. The performance index of the
pitch angle response is as shown in Table 3. Table 3. Performance index of pitch angle. Table 3. Performance index of pitch angle. No
Performance Index
Angle Value
30°
60°
90°
1
Rise Time
3.128 s
6.286 s
9.444 s
2
Error Steady State
-0.007 35
-0.007 35
-0.007 35
3
% Maximum Overshoot
2.577 %
1.531 %
0.008 889 %
4
Settling time
4.523 s
7.681 s
10.838 s Table 3. Performance index of pitch angle. The performance index of the yaw angle response is as shown in Table 4. e performance index of the yaw angle response is as shown in Table 4. Table 4. Performance index of yaw angle. Time
Efficiency ( %)
1 Axis to fixe d
2 Axis to fixe d
2 Axis to 1 Axis
1 d
23.09 %
50.16 %
21.99 %
3 d
13.63 %
36.67 %
20.28 %
5 d
16.11 %
46.04 %
25.78 %
7 d
15.98 %
46.65 %
26.44 % Table 4. Performance index of yaw angle. 8 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 Tracking test on the sun tracking system serves to determine the response of the system
when the setpoint changed. The test is done by changing the setpoint from 30° raised to 60°
then lowered to 40° then raised again to 70°. One of the important goals of the solar system
applied to photovoltaics is to increase the output power. In the performance test on
photovoltaic compared to output power output between PV that does not use the system of
tracking the sun (fixed), solar tracking system with one axis and solar tracking system with
two axes with data taking with a span of 1 d, 3 d, 5 d and 7 d. The PV power output profile is as shown in Figures 7. The efficiency of PV on one axis,
two axis, and fixed axis systems is as shown in Table 5. Table 5. Photovoltaic efficiency. 5 Conclusion Based on the research that has been done, ANFIS control with point pitch test resulted in
rising time performance index of 3.128 s, 6.28 s, and 9.44 s, maximum overshoot 2.577 %,
1.533 %, and 0.008 9 %, settling time of 4.523 s, 7.681 s and 10.838 s at each corner 30°,
60°, and 90° and error steady state -0.073 5 V. ANFIS control with setpoint yaw test
yielded rise time index of 3.203 s, 6.235 s, and 9.267 s, maximum % overshoot 1.531 %,
0.505 %, and 0.004 8 %, settling time 3.836 s, 6.867 s and 9.899 s at each corner 30°, 60°,
and 90° and error steady state -0.008 1 V. The two-axis solar tracking system has a greater
power output than the single axis solar tracking and fixed photovoltaic. The efficiency of
power (data for 7 d) from a one-axis solar tracking system to the solar tracking system with
fixed PV is 15.98 %, while for the efficiency of the two-axis sun measuring system against
the solar tracking system with fixed PV is 46.65 %, while the efficiency of the tracking
system two axes of sun to one axis sun track system is 26.442 %. In the next research, data
used to train ANFIS should represent positive values and negative values so that ANFIS
dicks can respond well when setpoints are raised or lowered. 4 Result and discussion No
Performance Index
Angle Value
30°
60°
90°
1
Rise Time
3.203 s
6.235 s
9.267 s
2
Error Steady State
-0.008 06
-0.008 06
-0.008 06
3
% Maximum Overshoot
1.531 %
0.505 %
0.004 778 %
4
Settling time
3.836 s
6.867 s
9.899 s Table 5. Photovoltaic efficiency. No
Performance Index
Angle Value
30°
60°
90°
1
Rise Time
3.203 s
6.235 s
9.267 s
2
Error Steady State
-0.008 06
-0.008 06
-0.008 06
3
% Maximum Overshoot
1.531 %
0.505 %
0.004 778 %
4
Settling time
3.836 s
6.867 s
9.899 s
(a) (b)
(c) (d)
Fig.7. PV Power output profile for (a) 1 d (b) 3 d (c) 5 d and (d) 7 d. (a) (b) (b) (a) (c) (d)
Fig.7. PV Power output profile for (a) 1 d (b) 3 d (c) 5 d and (d) 7 d. (d) (c) (d) (c) Fig.7. PV Power output profile for (a) 1 d (b) 3 d (c) 5 d and (d) 7 d. 9 https://doi.org/10.1051/e3sconf/202019000005 E3S Web of Conferences 190, 00005 (2020)
ICoRER 2019 References 1. E. Díaz-Dorado, A. Suárez-García, C.J. Carrillo, J. Cidrás, Renew. Energy. 36,6:1826–1835(2011). https://www.sciencedirect.com/science/article/pii/S0960148110005562 E. Díaz-Dorado, A. Suárez-García, C.J. Carrillo, J. Cidrás, Renew. Energy. 36,6:1826–1835(2011). 2. President of the Republic of Indonesia. Undang-Undang Republik Indonesia Nomor
30 Tahun 2007 Tentang Energi. [Republic of Indonesia Law Number 30 Year 2007
Concerning Energy] Jakarta: Republic of Indonesia. (2007) [in Bahasa Indonesia]. https://jdih.esdm.go.id/peraturan/uu-30-2007.pdf 3. The International Renewable Energy Agency. Decentralised RE Data Review,
Indonesia [Online] from http://resourceirena.irena.org/gateway/dashboard/ (2016). [Accessed on June 2, 2020]. 4. Batayneh, Wafa, A. Owais, M. Nairoukh, Automat. Constr, 29:100–106(2013). https://www.sciencedirect.com/science/article/pii/S0926580512001550 5. I. Sefa, M. Demirtas, I. Çolak., Energ. Convers. Manage. 50,11:2709–2718(2009). https://www.sciencedirect.com/science/article/pii/S0196890409002349 6. X. Li, Z. Deng, Z. Chen, Q. Fei. IEEE Transactions on Industrial Electronics,
58,2:450–464(2011). https://ieeexplore.ieee.org/document/5439835?reload=true 7. M. Hossein, K. Alizera, J. Arzhang, M. Hossein, A. Karen, S. Ahmad. Renew. Sust. Energ. Rev. 13,8:1800–1818(2009). 7. M. Hossein, K. Alizera, J. Arzhang, M. Hossein, A. Karen, S. Ahmad. Renew. Sust. Energ. Rev. 13,8:1800–1818(2009). https://ideas.repec.org/a/eee/rensus/v13y2009i8p1800-1818.html 8. I. Abadi, C. Imron, R.D. Noriyati. E3S Web Conf. 43,01014:1–8(2018). https://www.e3s-
conferences.org/articles/e3sconf/abs/2018/18/e3sconf_astechnova2017_01014/e3scon
f_astechnova2017_01014.html 9. J.S.R. Jang, C.T. Sun, E. Mizutani, Neuro-Fuzzy and soft computing: A computational
approach to learning and machine intelligence. New Jersey: Pearson (1997). p. 648. https://www.amazon.com/Neuro-Fuzzy-Soft-Computing-Computational-
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ICoRER 2019 https://doi.org/10.1051/e3sconf/202019000005 10. Y. Garcia, O. Diaz, C. Agudelo, WIT Trans. Ecol. Environ. 195:197–207(2015). https://www.researchgate.net/publication/300634197_Performance_of_a_solar_PV_tr
acking_system_on_tropic_regions 10. Y. Garcia, O. Diaz, C. Agudelo, WIT Trans. Ecol. Environ. 195:197–207(2015).
https://www.researchgate.net/publication/300634197_Performance_of_a_solar_PV_tr
acking_system_on_tropic_regions 11. M.A. Ghazali, A.M.A. Rahman, Energy Environ. Res., 2,1:235–243(2012). https://www.researchgate.net/publication/284708513_The_Performance_of_Three_Di
fferent_Solar_Panels_for_Solar_Electricity_Applying_Solar_Tracking_Device_under
_the_Malaysian_Climate_Condition 12. A. Mohanad, M.A. Al-Nimr, Y. Qaroush, Energy Convers. Manag. 26,7–8:1229–
1245(2005). https://www.sciencedirect.com/science/article/pii/S0196890404001529 13. J. Maknunah, I. Abadi, I. Abdurrahman, C. Imron, AIP Conference Proceedings,
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P1
Protective role of autophagy in mouse cecal ligation and
puncture-induced sepsis model Methods After profound anesthesia with pentothal, the Wistar rats
were killed by exsanguination. After sternotomy, the heart was taken
and connected to the Langendorf column. The apex of the heart was
hooked to a strength sensor. Biopac student laboratory software was
used to record and analyse heart contractions. Contractions were
recorded every 5 minutes during periods of 20 minutes. Control
measurements were fi rst recorded. We measured four parameters:
heart rate, contraction force, speeds of contraction and relaxation
for control, during TNFα (20 ng/ml) exposure and after removal of
TNFα. We express the variations of parameters as percentage of
the control ± SEM. A paired t test was used to compare heart rate,
contraction amplitude, speeds of contraction and relaxation with TNFα
and control measurements and after removal of TNFα. pu ctu e
duced seps s
ode
W Takahashi1, H Hatano2, H Hirasawa1, S Oda1
1Graduate School of Medicine, Chiba University, Chiba, Japan; 2Biomedical
Research Center, Chiba University, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P1 (doi: 10.1186/cc11939) Introduction Autophagy is well known as one of the biogenic
responses against various stresses, which possesses the benefi cial roles
for survival, but little is known about the dynamics and its signifi cance
during the septic condition. We hypothesized that autophagy is
induced during the septic condition, and contributes to protect from
tissue damage which subsequently leads to organ dysfunction. We
confi rm whether the autophagic process is accelerated or sustained in
an acute phase of sepsis and we also determine its physiological role. Results Eight rat hearts Wistar (weight = 325 ± 23 g) were studied. See
Table 1. y
g
Methods Sepsis was induced by cecal ligation and puncture (CLP) in
mice. We examined the kinetics of autophagosome and auto lysosome
formation which may explain the status of autophagy by western
blotting, immunohistochemistry, and electron microscopy. To investigate
a precise role of autophagy in CLP-induced sepsis, chloroquine, an
autophagy inhibitor, was administered to the CLP-operated mice, and
blood chemistry, pathology of the liver and survival were evaluated. Table 1 (abstract P2)
TNFα
Removal of TNFα
Heart rate
78 ± 6*
91 ± 5
Contractile
62 ± 8*
91 ± 4
Speed of contraction
72 ± 6*
93 ± 2
Speed of relaxation
53 ± 10*
89 ± 4
Results expressed as percentage of control ± SEM. *P <0.05. MEETING ABSTRACTS MEETING ABSTRACTS P1
Protective role of autophagy in mouse cecal ligation and
puncture-induced sepsis model y p
gy
Results Autophagy demonstrated by the ratio of LC3-II/LC3-I was
induced over the time course up to 24 hours after CLP. The ratio was
particularly increased in the liver, heart and spleen. Autophagosome
formation became maximal at 6 hours and declined by 24 hours after
CLP. Autolysosome formation as evaluated by both fusion of GFP-LC3
dots with LAMP1 immunohistochemistry and electron microscopy
was also increased after the procedure. Furthermore, inhibition of
autophagy by chloroquine during the CLP procedure resulted in
elevation of serum AST levels, and signifi cantly increased mortality in
mice. Conclusion TNFα decreases signifi cantly the heart rate, contractile
force, speeds of contraction and relaxation on isolated perfused
rat heart. TNFα probably plays a role in the pathophysiology of
cardiomyopathy during septic shock. The partial reversibility of these
eff ects could explain why left ventricular hypokinesia in patients with
septic shock is reversible. Conclusion Autophagy was induced in several organs over the time
course of the CLP sepsis model and then the process was gradually
completed to degradation of the components. Our data suggest
autophagy plays a protective role in organ dysfunction in sepsis. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P1
Protective role of autophagy in mouse cecal ligation and
puncture-induced sepsis model
W Takahashi1, H Hatano2, H Hirasawa1, S Oda1
1Graduate School of Medicine, Chiba University, Chiba, Japan; 2Biomedical
Research Center, Chiba University, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P1 (doi: 10.1186/cc11939) P1
Protective role of autophagy in mouse cecal ligation and
puncture-induced sepsis model
W Takahashi1, H Hatano2, H Hirasawa1, S Oda1
1Graduate School of Medicine, Chiba University, Chiba, Japan; 2Biomedical
Research Center, Chiba University, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P1 (doi: 10.1186/cc11939) References References
1. Vieillard-Baron A, et al.: Actual incidence of global left ventricular
hypokinesia in adult septic shock. Crit Care Med 2008, 36:1701-1706. 2. Hunter JD, et al.: Sepsis and the heart. Br J Anaesth 2010, 104:3-11. 1. Vieillard-Baron A, et al.: Actual incidence of global left ventricular
hypokinesia in adult septic shock. Crit Care Med 2008, 36:1701-1706. yp
p
2. Hunter JD, et al.: Sepsis and the heart. Br J Anaesth 2010, 104:3-11. 2. Hunter JD, et al.: Sepsis and the heart. Br J Anaesth 2010, 104:3-11. 33rd International Symposium on Intensive Care
and Emergency Medicine Brussels, Belgium, 19-22 March 2013 Published: 19 March 2013 The aim of our study was to observe the eff ects of TNFα on the model
of perfused rat heart. P3
Eff ect of cdp-choline on microcirculatory alterations during
endotoxemia BV Nguyen1, M Guillouet2, MA Giroux-Metges2, G Gueret3,
M Ould-Ahmed1, JP Pennec2
1Hôpital d’Instruction des Armées Clermont Tonnerre, Brest, France;
2Laboratory of Physiology, Faculty of Medicine, Brest, France; 3University
Hospital, Brest, France
Critical Care 2013, 17(Suppl 2):P2 (doi: 10.1186/cc11940) K Schmidt1, M Doerr1, T Brenner1, S Hofer1, A Walther2
1Universitätsklinikum Heidelberg, Germany; 2Klinikum Stuttgart, Klinik für
Anästhesiologie u. operative Intensivmedizin, Stuttgart, Germany
Critical Care 2013, 17(Suppl 2):P3 (doi: 10.1186/cc11941) K Schmidt1, M Doerr1, T Brenner1, S Hofer1, A Walther2
1Universitätsklinikum Heidelberg, Germany; 2Klinikum Stuttgart, Klinik für
Anästhesiologie u. operative Intensivmedizin, Stuttgart, Germany
Critical Care 2013, 17(Suppl 2):P3 (doi: 10.1186/cc11941) Introduction Acute myocardial depression in septic shock is common
[1]. Myocardial depression is mediated by circulating depressant
substances, which until now have been incompletely characterized [2]. Introduction The cholinergic anti-infl ammatory pathway (CAP) is
a physiological mechanism that inhibits cytokine production and
minimizes tissue injury during infl ammation. CAP-mediated anti-
infl ammatory signals in vagal eff erent nerve fi bers result in the
release of acetylcholine, which interacts with innate immune cells that
express the nicotinic acetylcholine receptor subunit α7 (α7nAChR). Introduction The cholinergic anti-infl ammatory pathway (CAP) is
a physiological mechanism that inhibits cytokine production and
minimizes tissue injury during infl ammation. CAP-mediated anti-
infl ammatory signals in vagal eff erent nerve fi bers result in the
release of acetylcholine, which interacts with innate immune cells that
express the nicotinic acetylcholine receptor subunit α7 (α7nAChR). © 2010 BioMed Central Ltd
© 2013 BioMed Central Ltd Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Endothelial dysfunction during sepsis is responsible for increased
endothelial permeability, leukocyte–endothelial interaction and
functional breakdown of microvascular perfusion. Endotoxemia-
induced endothelial dysfunction can be reduced by cholinergic CAP
activation [1]. The aim of this study was to determine the eff ects of the
α7nAChR-agonist cdp-choline on microcirculatory alterations during
experimental endotoxemia. response (a fi vefold to ninefold rise compared with other groups who
had an increase of 14-fold to 36-fold). Conclusion Severely ill patients and secondarily hemodialysis and
diabetic patients are in a proinfl ammatory state. The response of
all examined groups to provocation by LTA was suffi cient, with a
diff erential expression of severely ill patients, a fact that refl ects their
diff erent immunologic status. P5
Correlation of the oxygen radical activity and antioxidants and
severity in critically ill surgical patients: preliminary report
J Lee1, H Shim2, JY Jang1 ,
,
g
1Yonsei University College of Medicine, Seoul, South Korea; 2Wonju Severance
Christian Hospital, Yonsei University Wonju College of Medicine, Wonju, Korea
Critical Care 2013, 17(Suppl 2):P5 (doi: 10.1186/cc11943) Introduction In septic patients, the oxygen radical (OR) showed toxic
eff ect to induce infl ammation and antioxidant activity could aff ect organ
dysfunction. This study was designed to determine the relationship
between antioxidant level and severity of organ dysfunction. Results There were no signifi cant diff erences in venular wall shear
rate between the groups after 120 minutes. There was no signifi cant
diff erence in the number of adhering leukocytes between the cdp-
choline/LPS groups (iii, iv) and the LPS group after 120 minutes. Macromolecular effl ux signifi cantly increased in all groups over
120 minutes. All groups (i, ii, iii, iv) showed a signifi cantly reduced
macromolecular effl ux compared with the LPS group after 120 minutes. Conclusion Cdp-choline has no eff ect on leukocyte–endothelial
interaction and microhemodynamic alterations during endotoxemia. By activating the CAP, cdp-choline reduces capillary leakage. Thus cdp-
choline might have a prophylactic and therapeutic anti-infl ammatory
eff ect on LPS-induced endothelial permeability. These fi ndings
identify the endothelium as a target of anti-infl ammatory cholinergic
mediators and cdp-choline as a potential therapeutic substance in
sepsis treatment. Methods The medical records of adult patients managed in a surgical
ICU from August 2012 to December 2012 were reviewed prospectively. Abstracted data included age, body weight (with BMI), APACHE II
scores, SOFA scores, MODS scores, fl uid intake, fl uid output, nutritional
support, shock, antioxidant levels, OR activities, zinc and selenium
levels, complication and mortality. In addition, length of stay (LOS) in
the ICU and in hospital, and in-hospital mortality were collected. These
data were investigated on the fi rst, the third and the seventh day,
respectively. p
y
Results A total of 13 patients were enrolled. The in-hospital mortality
rate was 7.7% and mean LOS in the ICU and hospital was 6.5 and 27.6,
respectively. Mean APACHE II score was 20.2. On the fi rst day of ICU, the
mean antioxidant level and OR were 1.5 (± 0.5) mmol/l and 1.6 (± 0.5)
mmol/l, respectively. At the same time, SOFA and MODS scores were
7.3 and 5.0, respectively, and zinc and selenium were 32.6 μg/dl and
68.4 ng/ml. P5
Correlation of the oxygen radical activity and antioxidants and
severity in critically ill surgical patients: preliminary report
J Lee1, H Shim2, JY Jang1 On the third day, mean antioxidant and OR were 1.5 (± 0.4)
and 1.8 (± 0.7) respectively (SOFA 6.6, MODS 4.9, zinc 50.0, selenium
70.7). On the seventh day, mean antioxidant and OR were 1.4 (± 0.5)
and 1.9 (± 0.7), respectively (SOFA 4.3, MODS 3.1, zinc 62.8, selenium
77.3). In the correlation analysis, MODS scores and antioxidant level
had signifi cant correlations on the fi rst and seventh days of ICU (P =
0.001, P = 0.009). p
Reference 1. Peter C, et al.: Shock 2010, 33:405-411. References 1. Noveanu M, Mebazaa A, Mueller C: Cardiovascular biomarkers in the ICU. Curr Opin Crit Care 2009, 15:377-383. y
Methods Blood sample was obtained from 10 healthy volunteers, 10
hemodialysis patients with end-stage chronic renal failure (CRF), 10
patients with type II diabetes mellitus (DM) and 10 ICU patients on the
second day of hospitalization, who suff ered nonseptic SIRS and had
an APACHE II score >25. After suitable treatment the samples were
incubated with 1 mg LTA for 8 hours and maintained at –20°C until the
measurement of cytokines TNFα, IL-6, IL-1β, and IL-10, using the ELISA
method. The results are presented as mean values ± SEM. Graph Pad 4.0
was used, applying a t test to test the variation of each cytokine in each
group, and ANOVA to assess the diff erences between the four groups. Methods Blood sample was obtained from 10 healthy volunteers, 10
hemodialysis patients with end-stage chronic renal failure (CRF), 10
patients with type II diabetes mellitus (DM) and 10 ICU patients on the
second day of hospitalization, who suff ered nonseptic SIRS and had
an APACHE II score >25. After suitable treatment the samples were
incubated with 1 mg LTA for 8 hours and maintained at –20°C until the
measurement of cytokines TNFα, IL-6, IL-1β, and IL-10, using the ELISA
method. The results are presented as mean values ± SEM. Graph Pad 4.0
was used, applying a t test to test the variation of each cytokine in each
group, and ANOVA to assess the diff erences between the four groups. Results Baseline cytokine values in the three groups were increased
compared with the control group, but the diff erence was signifi cant
only for the ICU group (Table 1, data only for IL-6 and IL-10). The
quotient IL-10/IL-6 of baseline values was between 0.23 and 0.96
among healthy, ESRD and DM persons, and 1.32 among ICU patients. In all examined groups the levels of cytokines increased signifi cantly
after stimulation with LTA, although ICU patients showed a diff erential 2. Piechota M, Banach M, Irzmanski R, Barylski M, Piechota-Urbanska M, Kowalski
J, et al.: Plasma endothelin-1 levels in septic patients. J Intensive Care Med
2007, 22:232-239. 3. Kotsovolis G, Kallaras K: The role of endothelium and endogenous
vasoactive substances in sepsis. Hippokratia 2010, 14:88-93. P3
Eff ect of cdp-choline on microcirculatory alterations during
endotoxemia Methods Using fl uorescent intravital microscopy, we determined
venular wall shear rate, macromolecular effl ux and leukocyte
adhesion in mesenteric postcapillary venules of male Wistar rats. Endotoxemia was induced over 120 minutes by intravenous infusion
of lipopolysaccharide (LPS). Control groups received an equivalent
volume of saline. Cdp-choline was applied as an i.v. bolus in treatment
groups. Animals received either (i) saline alone, (ii) cdp-choline
10 minutes prior to saline administration, (iii) cdp-choline 10 minutes
prior to LPS administration, (iv) cdp-choline 30 minutes after LPS
administration or (v) LPS alone.if Immune response after stimulation with wall components of
Gram-positive bacteria S Aloizos1, E Tsigou2, P Myrianthefs2, S Gourgiotis1, A Tsakris3, G Baltopoulos2
1NIMTS Hospital, Athens, Greece; 2A. Anargiroi Hospital, Athens, Greece;
3M di
l S h
l Ath
G S Aloizos1, E Tsigou2, P Myrianthefs2, S Gourgiotis1, A Tsakris3, G Baltopoulos2
1NIMTS Hospital, Athens, Greece; 2A. Anargiroi Hospital, Athens, Greece;
3Medical School Athens Greece S Aloizos1, E Tsigou2, P Myrianthefs2, S Gourgiotis1, A Tsakris3, G Baltopoulos2
1NIMTS Hospital, Athens, Greece; 2A. Anargiroi Hospital, Athens, Greece;
3Medical School, Athens, Greece
Critical Care 2013, 17(Suppl 2):P4 (doi: 10.1186/cc11942) Critical Care 2013, 17(Suppl 2):P4 (doi: 10.1186/cc11942) Critical Care 2013, 17(Suppl 2):P4 (doi: 10.1186/cc11942) Conclusion Antioxidant level had a correlation with organ dysfunction
which might be used as a prognostic factor in critically septic patients. To prove this, large-scale data collection is required. R f Introduction The purpose of this study was to evaluate the immune
response of patients susceptible to infection by Gram-positive bacteria
after ex vivo provocation with lipoteichoic acid (LTA) and to compare
the reaction with the one of healthy adults. Prevention of sepsis by correcting apoptosis
M Puhtinskaya, V Estrin y
Research Institute of Obstetrics and Pediatrics, Rostov-on-Don, Russia
Critical Care 2013, 17(Suppl 2):P8 (doi: 10.1186/cc11946) Research Institute of Obstetrics and Pediatrics, Rostov-on-Don, Russia
Critical Care 2013, 17(Suppl 2):P8 (doi: 10.1186/cc11946) Research Institute of Obstetrics and Pediatrics, Rostov-on-Don, Russ Critical Care 2013, 17(Suppl 2):P8 (doi: 10.1186/cc11946) Introduction Activation of apoptosis in lymphocytes determines
the development of neutropenia and of sepsis [1,2]. We investigated
prevention of sepsis and correction of lymphocyte apoptosis by
recombinant human granulocyte colony-stimulating factor (hr-GCSF,
fi lgrastim) [1,2]. Methods With the permission of the ethics committee, a controlled,
randomized, blind clinical trial included 69 term newborns on
mechanical ventilation, without neutropenia and clinical signs of
infection, with a content of lymphocytes in early apoptosis (AnnexinV–
FITC+PI–) of >9.59%, and in late (AnnexinV–FITC+PI+) of 0.56%. Lymphocytes in apoptosis were detected using antibodies to AnnexinV
and propidium iodide staining method of immunophenotyping (fl ow
cytometry; Beckman Coulter Epics XL, USA). The survey was conducted
at admission, at 3 to 5 days, and 20 days. The method of random
numbers in Group I included 39 newborns who on admission (with
written parental consent) received an intravenous infusion of hr-GCSF
dose of 10 μg/kg, 3 days. Newborns of Group II (n = 30) did not receive
hr-GCSF. Power of the study was 80% (α ≤0.05).i Conclusion In this study we noticed that during sepsis some signifi cant
diff erences in the status of activation of immune cells exist between
peripheral blood and lung resident cells. The lung milieu seems to
promote activation of monocytes while neutrophil activation is more
dependent on the site of infection. However, these observations
require further studies in a larger group of patients. Acknowledgements This study was supported by the Centre of
Postgraduate Medical Education grant no 501-01-02-012 and by the
sources of the Medical University of Warsaw. P7
Anti-infl ammatory eff ects of Kupff er cells through α7-nicotinic
acetylcholine receptors
Y Li, X Shi
Changzheng Hospital, Second Military Medical University, Shanghai, China
Critical Care 2013, 17(Suppl 2):P7 (doi: 10.1186/cc141945) Results For 3 to 5 days, Group I signifi cantly decreased apoptosis of
lymphocytes in the early from 16.1% to 7.8%, and in late from 1.3%
to 0.1%. The development of sepsis and neutropenia have been
reported. We observed no clinical or laboratory signs of adverse eff ects
of the drug. Fatal outcomes (n = 4) are not associated with hr-GCSF,
which was confi rmed postmortem. Prevention of sepsis by correcting apoptosis
M Puhtinskaya, V Estrin Decreased duration of mechanical
ventilation (P <0.05). In Group II, 27 patients at 3 to 5 days developed
neutropenia and increased lymphocytes in apoptosis (P <0.05). Sepsis
was diagnosed in 19 children; eight fatal outcomes. Introduction Nicotine exerts anti-infl ammatory eff ects in several cell
types. α7-nicotinic acetylcholine receptor (α7-nAChR), which has
high permeability to calcium, is believed to contribute signifi cantly to
nicotinic anti-infl ammatory eff ects. However, the molecular mechanism
is largely unknown. Kupff er cells in the liver play an important role in
infl ammatory response to pathogens invading, but whether there is
α7-nAChR expression in Kupff er cells or cholinergic anti-infl ammatory
pathway involved in this process remains unclear.f Conclusion hr-GCSF reduces the incidence of septic complications and
one of the mechanisms of its clinical eff ectiveness is the reduction of
apoptotic factors aff ecting the development of neutropenia. References 1. Gillan ER, Christensen RD, Suen Y, et al.: A randomized, placebo-controlled
trial of recombinant human granulocyte colony-stimulating factor
administration in newborn infants with presumed sepsis: signifi cant
induction of peripheral and bone marrow neutrophilia. Blood 1994,
84:1427-1433. 1. Gillan ER, Christensen RD, Suen Y, et al.: A randomized, placebo-controlled
trial of recombinant human granulocyte colony-stimulating factor
administration in newborn infants with presumed sepsis: signifi cant
induction of peripheral and bone marrow neutrophilia. Blood 1994,
84:1427-1433. Methods (1) Kupff er cells, isolated by collagenase digestion and
diff erential centrifugation from mice and labeled with FITC-aBGT,
were observed under laser scanning confocal microscope to test the
expression of α7-nAChR. Protein level was also tested by western
blotting, with RAW264.7 as positive control; (2) 100 nM LPS was
given to Kupff er cells, with or without 1 mM nicotine. TNFα, IL-10 and
HMGB-1 were tested at 4 hours, 12 hours or 24 hours, respectively; (3)
100 BALB/c mice were randomly divided into four group: Group I (only
lethal dose of LPS was given), Group II (nicotine and LPS were given),
Group III (LPS, nicotine and GdCl3 were given), and Group IV (LPS and
nicotine were given and the left cervical vagus nerve was cut off ). The
mortality of mice was observed for 72 hours.fi 2. Pukhtinskaya MG, Estrin VV, Gulova ES: Clinical and diagnostic value of
apoptosis markers in the pathogenesis of neutropenia and bacterial
complications in newborns with respiratory distress syndrome. Cytokines
Infl amm 2011, 10:66-69. Prevention of sepsis by correcting apoptosis
M P h
k
V E g
y
Results The mortality of sepsis reached 70%. Nonsurvivors had
signifi cantly (P = 0.001) elevated expression of CD64 on neutrophils. Expression of HLA-DR was higher in monocytes from BAL than PB
GMF (1,032 vs. 342; P = 0.02) and this tendency was present in sepsis
originating from both pneumonia and peritonitis. Percentage of HLA-
DR-positive T cells was lower in PB than in BAL (2.9% vs. 6.5%; P = 0.07),
but the GMF values for HLA-DR were higher in the circulating T cells
(1,904 vs. 1,346; P = 0.004). The expression of CD64 on neutrophils was
not signifi cantly diff erent in PB and BAL, but there was a trend towards
its higher expression in BAL from patients with pneumonia while its
expression was higher in PB of patients with peritonitis.i Simultaneous analysis of the expression of CD64 and HLA-DR in the
peripheral blood and bronchoalveolar lavage fl uid in sepsis It
was verifi ed that left cervical vagus nerve was essential for the anti-
infl ammatory eff ect of nicotine and α7 acetylcholine receptors might
play a critical role. anti-infl ammatory compensatory response. However, controversies
exist regarding the status of the immune system in local tissue
compartments during sepsis. The aim of this study was to compare
selected markers of activation between the systemic circulation and
local lung environment. Methods Twenty patients with severe sepsis were included into this
study. Peripheral blood (PB) samples and bronchoalveolar lavage fl uid
(BALF) samples were obtained on the day of diagnosis (D1). BALF was
collected from 11 patients. Samples were stained with antibodies:
CD15/CD64 and CD3/CD14/HLA-DR and isotypic control. Cells were
analysed by fl ow cytometry. Expression of markers of activation was
analysed as the geometric median of fl uorescence (GMF). All values
are expressed as median values. Comparisons between groups were
performed using Mann–Whitney and Wilcoxon tests. References 1. Wang H, et al.: Nicotinic acetylcholine receptor α7 subunit is an essential
regulator of infl ammation. Nature 2003, 421:384-388. 1. Wang H, et al.: Nicotinic acetylcholine receptor α7 subunit is an essential
regulator of infl ammation. Nature 2003, 421:384-388. 2. Wang H, et al.: Cholinergic agonists inhibit HMGB1 release and improve
survival in experimental sepsis. Nat Med 2004, 10:1216-1221. 2. Wang H, et al.: Cholinergic agonists inhibit HMGB1 release and improve
survival in experimental sepsis. Nat Med 2004, 10:1216-1221. Simultaneous analysis of the expression of CD64 and HLA-DR in the
peripheral blood and bronchoalveolar lavage fl uid in sepsis f
Results Baseline cytokine values in the three groups were increased
compared with the control group, but the diff erence was signifi cant
only for the ICU group (Table 1, data only for IL-6 and IL-10). The
quotient IL-10/IL-6 of baseline values was between 0.23 and 0.96
among healthy, ESRD and DM persons, and 1.32 among ICU patients. In all examined groups the levels of cytokines increased signifi cantly
after stimulation with LTA, although ICU patients showed a diff erential l
T Skirecki1, M Mikaszewska-Sokolewicz2, G Hoser1, U Zielińska-Borkowska1
1The Centre of Postgraduate Medical Education, Warsaw, Poland; 2 Medical
University of Warsaw, Poland T Skirecki1, M Mikaszewska-Sokolewicz2, G Hoser1, U Zielińska-Borkowska1
1The Centre of Postgraduate Medical Education, Warsaw, Poland; 2 Medical
University of Warsaw, Poland y
Critical Care 2013, 17(Suppl 2):P6 (doi: 10.1186/cc11944) Introduction The core pathophysiological changes in sepsis
involve systemic activation of the immune system followed by the Table 1 (abstract P4). Levels of cytokines before and after stimulation with LTA
Control baseline LTA
ESRD baseline LTA
DM baseline LTA
ICU baseline LTA
IL-6
8.90 ± 0.76, 245.30 ± 26.68
86.60 ± 45.55, 1,310.00 ± 154.80
15.90 ± 1.89, 252.00 ± 35.52
372.40 ± 120.60, 3,659.00 ± 485.20
IL-10
3.00 ± 1.08, 40.90 ± 7.45
19.20 ± 7.14, 273.10 ± 126.50
15.30 ± 2.08, 350.50 ± 89.42
492.60 ± 66.72, 2,822.00 ± 432.70
Values in pg/ml. Table 1 (abstract P4). Levels of cytokines before and after stimulation with LTA Table 1 (abstract P4). Levels of cytokines before and after stimulation with LTA
Control baseline LTA
ESRD baseline LTA S3 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Conclusion Kupff er cells played a crucial rule in modulating
infl ammation and the anti-infl ammatory eff ect of nicotine was partially
weakened after left cervical vagus nerve cut off or aBGT was given. It
was verifi ed that left cervical vagus nerve was essential for the anti-
infl ammatory eff ect of nicotine and α7 acetylcholine receptors might
play a critical role. Conclusion Kupff er cells played a crucial rule in modulating
infl ammation and the anti-infl ammatory eff ect of nicotine was partially
weakened after left cervical vagus nerve cut off or aBGT was given. Immune paralysis in trauma patients; implications for prehospital
interventionf Results (1) Expression of α7-nAChR in Kupff er cells was confi rmed
by confocal microscope and western blotting; (2) after nicotine was
administered, the level of TNFα and HMGB-1 increased and the level
of IL-10 decreased. Given left cervical vagus nerve cut off or aBGT,
the eff ect of nicotine was weakened; (3) Group I had the highest
mortality rate, while in Group II nicotine did reduce the mortality rate
dramatically. After the left cervical vagus nerve was cut off or aBGT
was given, the eff ects of nicotine were weakened. Diff erence for the
mortality rate between Group III and Group IV was not signifi cant. M Kox, K Timmermans, M Vaneker, GJ Scheff er, P Pickkers
Radboud University Nijmegen Medical Center, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P9 (doi: 10.1186/cc11947) Introduction Multi-trauma is one of the major indications for intensive
care admission. Recovery is frequently complicated by post-injury
immunological complications, caused by a dysfunctional immune
system; for example, sepsis and multiple organ failure. In order to treat S4 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 hematologic or metastasized malignancies, cardiac arrest, and use of
bone marrow suppressive drugs. Laboratory data were collected from
blood samples, prior to in-hospital i.v. fl uid therapy. In order to detect a
diff erence in Hb concentration of 0.2 mmol/l, we anticipated a sample
size of 283 per group, based on a standard deviation (SD) of 1.2, α = 0.05
and β = 0.8. Data are expressed as mean ± SD. or prevent this immune paralysis, knowledge on the time course of
immune paralysis in vivo and the pathophysiological mechanisms of
immune paralysis is essential. The aim of this study is to determine
factors that could predict and/or induce immunological complications
in these patients to ultimately fi nd a suitable target and timeframe for
intervention. hematologic or metastasized malignancies, cardiac arrest, and use of
bone marrow suppressive drugs. Laboratory data were collected from
blood samples, prior to in-hospital i.v. fl uid therapy. In order to detect a
diff erence in Hb concentration of 0.2 mmol/l, we anticipated a sample
size of 283 per group, based on a standard deviation (SD) of 1.2, α = 0.05
and β = 0.8. Data are expressed as mean ± SD. β
p
Results We included 296 patients in the S-group and 320 in the
C-group. Do changes in red blood cell deformability in patients with septic
shock correlate with changes in SOFA scores? T Clark1, S Jewell2, M Sair1, P Petrov2, P Winlove2 T Clark1, S Jewell2, M Sair1, P Petrov2, P Winlove2
1Derriford Hospital, Plymouth, UK; 2University of Exeter, UK
C
l C
(S
l ) P
(d
/
) Derriford Hospital, Plymouth, UK; 2University of Exeter, UK
Critical Care 2013 17(Suppl 2):P11 (doi: 10 1186/cc11949) Conclusion Immune paralysis can be established within hours after
trauma. Production of anti-infl ammatory IL-10 in the prehospital phase
could play a crucial role in the pathogenesis. Patients with a higher
injury severity score are more prone to produce excessive IL-10 in this
phase. Immune stimulatory strategies applied by the HEMS or early
after hospital admission could form a potential future approach to
prevent immune paralysis in multitrauma patients in the intensive care
ward. p
,
y
,
;
y
,
Critical Care 2013, 17(Suppl 2):P11 (doi: 10.1186/cc11949) Introduction Traditional whole blood experiments suggest that sepsis
causes abnormal red blood cell (RBC) deformability. To investigate
this at the cellular level, we employed a novel biophysical method to
observe individual RBC membrane mechanics in patients with septic
shock. Methods We collected blood samples from patients with septic
shock until either death or day 5 of admission. Thermal fl uctuations
of individual RBCs were recorded allowing a complete analysis of RBC
shape variation over time. Mean elasticity of the cell membrane was
then quantifi ed for each sample collected. Is hemoglobin concentration aff ected by sepsis in the acute phase? G Jansma, H Buter, RT Gerritsen, EC Boerma
Medical Centre Leeuwarden, the Netherlands
Critical Care 2013, 17(Suppl 2):P10 (doi: 10.1186/cc11948) Immune paralysis in trauma patients; implications for prehospital
interventionf The diff erence in Hb between the S-group and C-group was
not signifi cant (8.76 ± 1.18 mmol/l vs. 8.93 ± 1.16 mmol/l, P = 0.07). After
correction for a number of confounders, using a multivariate regression
analysis, we observed a signifi cant diff erence in Hb of –0.23 mmol/l in
the S-group in comparison with the C-group (P = 0.01).il Methods Blood was drawn from adult multi-trauma patients (n = 94)
admitted to the emergency room (ER) of the Radboud University
Nijmegen Medical Center. Blood was drawn at the trauma scene by
the helicopter emergency medical services (HEMS), at arrival in the ER
and at days 1, 3, 5, 7, 10 and 14 after trauma. Plasma concentrations of
TNFα, IL-6, IL-10, IFNγ, IL-8 and MCP-1 were determined by Luminex. Ex vivo 24-hour whole blood stimulations with LPS or pam3cys were
performed and produced TNFα, IL-6 and IL-10 was measured using
ELISA to determine the level of immune paralysis. Clinical data – for
example, Injury Severity Scores, trauma mechanism, medication and
survival – were collected from electronic patient fi les. g
p
p
g
p
Conclusion At fi rst presentation, prior to in-hospital i.v. fl uid therapy,
Hb concentration in patients with sepsis is signifi cantly lower in
comparison with controls; however, the diff erence is very small,
without the existence of anemia. R f Is hemoglobin concentration aff ected by sepsis in the acute phase?
G Jansma, H Buter, RT Gerritsen, EC Boerma
Medical Centre Leeuwarden, the Netherlands
Critical Care 2013, 17(Suppl 2):P10 (doi: 10.1186/cc11948) i
Results We recruited nine patients with septic shock. Table 1 shows
mean RBC thermal fl uctuation and SOFA scores.l Introduction In the acute phase of sepsis several potential mechanisms
may change the hemoglobin (Hb) concentration. On the one hand,
endothelial activation may lead to increased vascular permeability and
fl uid sequestration to the interstitium, leading to hemoconcentration. On the other hand, degradation of the glycocalyx has been reported [1]. Shedding of this carbohydrate-rich layer with an estimated thickness of
0.2 to 0.5 μm may lead to a substantial increase of the intravascular
space, and thus to decrease of Hb concentration [2]. The aim of this
study is to determine whether there is a decrease in Hb in the acute
phase of sepsis.i l
Conclusion RBC thermal fl uctuation analysis allows variations in RBC
elasticity during sepsis to be quantifi ed at a cellular level. We could not
identify any specifi c trend between sepsis severity and erythrocyte
elasticity. Cells demonstrated both increases and decreases in fl uctua-
tion independent of SOFA score. This is contrary to current evidence
that suggests RBC deformability is reduced during sepsis. Reference 1. Piagnerelli M, et al.: Intensive Care Med 2003, 29:1052-1061. References i
Results The plasma IL-10 concentration at ER was 16.5-fold increased
in comparison with time-point HEMS (P <0.01). Similar but less
pronounced eff ects were found for IL-8 and MCP-1. A signifi cant
correlation (P = 0.03, R = 0.53) was found between injury severity
scores and IL-10 plasma concentration at time-point ER. Time-courses
of ex vivo produced cytokines suggest that LPS-induced IL-6 and TNFα
production is already decreased in the fi rst few hours after trauma
and recovering from day 5. Ex vivo IL-10 production shows an inverse
pattern. 1. Steppan J, et al.: Sepsis and major abdominal surgery lead to fl aking of the
endothelial glycocalyx. J Surg Res 2011, 165:136-141. 2. van den Berg BM, et al.: The endothelial glycocalyx protects against
myocardial edema. Circ Res 2003, 92:592-594. 2. van den Berg BM, et al.: The endothelial glycocalyx protects against
myocardial edema. Circ Res 2003, 92:592-594. Reference Reference
1. Strauss R, et al.: Thrombocytopenia in patients in the medical intensive care
unit: bleeding prevalence, transfusion requirements, and outcome. Crit
Care Med 2002, 30:1765-1771. Methods We collected blood samples from elective cardiac surgery
patients pre (at induction) and post (immediately, each day until CICU
discharge) CPB. Thermal fl uctuations of individual RBCs were recorded
using a high-frame-rate camera allowing a complete analysis of RBC
shape variation over time. Mean elasticity of the cell membrane was
then quantifi ed for each sample collected. P14
N
t i
Results Fifteen patients were recruited. Table 1 displays the results. RBC thermal fl uctuation is measured relative to pre-bypass values. An
increase in RBC fl uctuation marks a decrease in stiff ness. CPB caused
two distinct changes in RBC elasticity; pre-fi x A indicates samples
where stiff ness increases or shows no change, B those where stiff ness
decreases. Data on day 2 were not collected in patients discharged
from the CICU. CPB type or time had no apparent impact on RBC
response to CPB. Introduction Neutrophil gelatinase-associated lipocalin (NGAL)/
lipocalin2, known as a sensitive biomarker of acute kidney injury,
prevents bacterial iron uptake, resulting in the inhibition of its
overgrowth [1]. We previously demonstrated that this protein was
discharged into gut lumen from crypt cells in septic conditions, and
inhibited the growth of Escherichia coli [2]. However, it remains unclear
which pathway is associated with the upregulation of NGAL. We
therefore designed the present study to reveal whether the pattern-
recognition receptor of bacteria, the Toll-like receptor (TLR) family,
plays a pivotal role for NGAL secretion from gut crypt cells. Conclusion RBC thermal fl uctuation analysis quantifi es the impact
of CPB on erythrocyte membrane elasticity. We clearly identifi ed two
separate RBC elasticity responses to CPB. This fi nding is contrary to
traditional fl ow measurement techniques that suggest CPB impairs
whole blood fl ow and reduces RBC deformability. Reference 1. Lindmark et al.: J Thoracic Cardiovasc Surg 2002, 123:381-383. p y
p
g
yp
Methods With our institutional approval, the ileum and colon of male
C57BL/6J mice (6 to 7 weeks) were everted and washed by Ca2+ and
Mg2+ free PBS buff er fi ve times. Tissues were incubated with Ca2+ and
Mg2+ free PBS containing 30 mM EDTA for 1 hour to isolate crypt cells
of gut. Platelet-associated oxidative stress and ADAMTS-13 levels are
inversely associated with a poor prognosis in septic shock Introduction Sepsis causes widespread microvascular injury and
thrombosis. Some hemostatic factors mediate the mechanisms
involved in sepsis-related organ ischemia and failure. Oxidative stress
is also increased in sepsis and reactive oxygen species (ROS) favor
secretion of von Willebrand factor (vWF) multimers from endothelium
and inhibit vWF proteolysis by ADAMTS-13. Moreover, the enzyme
indoleamine-2,3-dioxygenase, an important immune regulator, is
activated in sepsis and, through generation of kynurenins, promotes
antioxidative and anti-infective activities. This study evaluated the
relative role of ADAMTS-13, vWF and fi brinogen in the morbidity and
mortality of patients with septic shock (SS). The above hemostatic
factors were measured together with kynurenine and plasma protein
carbonyls, marker of oxidative stress. Results More than 70 to 80% of collected cells were stained by amido
black. LPS signifi cantly upregulated the expression of NGAL and TLR4
mRNA in ileum and colon crypt cells (P <0.05). Although the CpG-DNA did
not upregulate NGAL and TLR9 mRNA in ileum crypt cells, the apparent
expression of NGAL and TLR9 mRNA was found in colon crypt cells (P <0.05). Conclusion Bacterial stimulation of TLR4 and TLR9 pathways plays a
pivotal role in the expression of NGAL mRNA in gut, suggesting that
NGAL, derived from gut crypt cells, could contribute to the regulation
of the intraluminal microfl ora in the critically ill. y
Methods One group of 12 patients with SS, defi ned using standard
criteria, was enrolled in the ICU of the ‘A. Gemelli’ Hospital (Rome,
Italy). Biochemical, hematologic and hemodynamic parameters were
measured on days 1 to 4, 7, 14 and 21. A group of 12 age-matched and
gender-matched healthy subjects was used as controls. References References
1. Nature 2004, 432:917. 2. Crit Care Med 2011, 39:46. 1. Nature 2004, 432:917. 2. Crit Care Med 2011, 39:46. Reference The cell suspension was fi ltered through a cell strainer (40 μm)
twice, and deposited the crypt cells by centrifugation at 700×g. The
isolated crypt cells were resuspended in PBS and stained with 0.25%
amido black for labeling paneth cells. The 5×105 crypt cells were
resuspended in 50 ml HBSS containing 2.5% fetal bovine serum and
1% penicillin–streptomycin. The crypt cells were incubated at 37°C with
or without TLR ligands: lipopolysaccharide (TLR4 ligand, 10 μg/ml) and
CpG-DNA (TLR9 ligand, 8 μg/ml). After a 2-hour incubation period, the
crypt cells were deposited and eluted mRNA to measure the expression
of both NGAL and TLR mRNA using real-time PCR. P13 Platelet-associated oxidative stress and ADAMTS-13 levels are
inversely associated with a poor prognosis in septic shock
L Montini1, G De Pascale1, MA Pennisi1, ES Tanzarella1, SL Cutuli1,
A Occhionero1, R De Cristofaro2, M Antonelli1
1Catholic University School of Medicine, Rome, Italy; 2Haemostasis and
Thrombosis Center, Catholic University School of Medicine, Rome, Italy
Critical Care 2013, 17(Suppl 2):P13 (doi: 10.1186/cc11951) P15 y
Results Low ADAMTS-13 activity was observed in SS patients
(268 ± 123 ng/ml vs. 760 ± 80 ng/ml in controls). vWF levels (antigen
and activity) were increased ~3-fold compared with controls. Likewise,
plasma protein carbonyls and kynurenine were globally increased in
patients (2.1 ± 1.5 nmol/mg vs. 0.3 ± 02 nmol/mg and 14.4 ± 9.7 μM
vs. 2.3 ± 1.3 μM, respectively). Intra-ICU mortality (3 of 15) was strongly
and inversely correlated with carbonyl levels (P = 0.04) and platelets
(P = 0.022). Lethal infl uenza virus A H1N1 infection in two relatives with
autosomal dominant GATA-2 defi ciency
J Sole-Violan1, I Sologuren1, E Betancor2, S Zhang3, C Pérez1,
E Herrera-Ramos1, M Martínez-Saavedra1, M López-Rodríguez1, J Pestano2,
J Ruiz-Hernández1, J Ferrer1, F Rodríguez de Castro1, J Casanova3,
C Rodríguez-Gallego1
1Hospital GC Dr Negrín, Las Palmas de GC, Spain; 2Universidad Las Palmas de
GC, Spain; 3The Rockefeller University, New York, NY, USA
Critical Care 2013, 17(Suppl 2):P15 (doi: 10.1186/cc11953) Lethal infl uenza virus A H1N1 infection in two relatives with
autosomal dominant GATA-2 defi ciency P12
Do erythrocytes subjected to cardiopulmonary bypass exhibit
changes in their membrane mechanical properties?
T Cl k1 S J
ll2 M S i 1 P P
2 P Wi l
2 p
p
Methods In this single-center retrospective analysis we identifi ed
patients with sepsis as the primary reason for non-elective ICU
admission from a standard patient database. Patients who fulfi lled the
international criteria of sepsis and organ failure during ICU admission
were included in the sepsis group (S-group). The control group was
formed by patients with other non-elective reasons for ICU admission
(C-group). Exclusion criteria were (recent) bleeding, surgery in the last
6 weeks, chronic renal failure (creat >177 μmol/l, or hemodialysis),
untreated chronic anemia, pregnancy, polytrauma, age <18, T Clark , S Jewell , M Sair , P Petrov , P Winlove
1Derriford Hospital, Plymouth, UK; 2University of Exeter, UK
Critical Care 2013, 17(Suppl 2):P12 (doi: 10.1186/cc11950) Introduction Whole blood experiments suggest that cardiopulmonary
bypass (CPB) causes red blood cell (RBC) trauma and changes in
deformability that may contribute to postoperative microcirculatory Table 1 (abstract P11). Mean RBC fl uctuation (daily SOFA score)
Day
A
B
C
D
E
F
G
H
I
1
4.8 (10)
5.2 (9)
–
4.8 (12)
4.6 (16)
4.9 (13)
5.1 (16)
4.6 (18)
5.1 (15)
2
4.9 (9)
5.0 (10)
4.6 (13)
5.1 (11)
4.8 (17)
5.1 (13)
5.0 (16)
4.9 (19)
– (16)
3
4.0 (6)
5.1 (9)
4.7 (12)
4.8 (11)
4.9 (18)
4.8 (13)
5.0 (16)
4.7 (21)
5.3 (16)
4
–
4.8 (7)
4.6 (11)
5.0 (9)
5.9 (19)
–
– (18)
–
5.0 (15)
5
–
–
–
5.1 (6)
5.2 (18)
–
– (19)
–
5.0 (10) Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S5 Table 1 (abstract P12). Change in RBC thermal fl uctuation relative to baseline: two distinct groups seen
Patient
A1
A2
A3
A4
A5
A6
A7
B8
B9
B10
B11
B12
B13
B14
B15
Post CPB
+0.1
–0.4
–0.1
–0.5
–0.1
–0.2
–0.1
+0.5
0
+0.4
+0.1
+0.2
0
+0.2
+0.1
Day 1
0
0
+0.1
–0.1
–0.1
+0.1
+0.1
0
+0.5
+0.4
+0.7
+0.4
+0.6
+0.5
+0.7
Day 2
–0.2
+0.1
0
NA
NA
0
NA
+0.5
–0.1
+0.4
NA
+0.1
NA
NA
NA Table 1 (abstract P12). Change in RBC thermal fl uctuation relative to baseline: two distinct groups seen dysfunction. We used a novel fl uctuation microscopy technique to
quantify the eff ects of CPB on RBC elasticity at a cellular level. Lethal infl uenza virus A H1N1 infection in two relatives with
autosomal dominant GATA-2 defi ciency Conclusion Hence, we hypothesize that, in the SS setting, platelets
contribute to oxidative stress that counteracts the organ failure-
associated mortality. Thus, low platelet count, irrespective of bleedings,
may favor mortality in SS patients by generating lower ROS amounts. Introduction Most individuals infected with the 2009 pandemic H1N1
infl uenza A virus (IAV) (H1N1pdm) experienced uncomplicated fl u. Introduction Most individuals infected with the 2009 pandemic H1N1
infl uenza A virus (IAV) (H1N1pdm) experienced uncomplicated fl u. S6 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 isolated and incubated for 24 hours in the presence of 10 ng/gl LPS of
Escherichia coli O55:B5 and of 5 μg/ml phytohemagglutin (PHA); TNFα
was measured in supernatants by a bioassay on L929 fi broblasts.i isolated and incubated for 24 hours in the presence of 10 ng/gl LPS of
Escherichia coli O55:B5 and of 5 μg/ml phytohemagglutin (PHA); TNFα
was measured in supernatants by a bioassay on L929 fi broblasts. Results Fifty percent of rabbits died early; that is, within the fi rst
48 hours. Mean ± SE log10 bacteria in the liver and lung of animals
that died early was 2.27 ± 0.62 and 3.16 ± 0.78 cfu/g; respective values
of rabbits that started dying late (that is, after 72 hours) were below
the limit of detection. Mean circulating LPS at 24 hours was 2.09 EU/
ml and 1.99 EU/ml respectively (P = NS). Mean LPS of the portal vein
of the sham and of the injury groups were 1.25 and 5.62 EU/ml (P =
0.047). Concentrations of TNFα in splenocyte supernatants are shown
in Figure 1. However, in a small subset of patients the infection rapidly progressed
to primary viral pneumonia (PVP) and a minority of them developed
ARDS. Inherited and acquired variability in host immune responses
may infl uence susceptibility and outcome of IAV infection. However,
the molecular nature of such human factors remains largely elusive. p
y
yi
Results Fifty percent of rabbits died early; that is, within the fi rst
48 hours. Mean ± SE log10 bacteria in the liver and lung of animals
that died early was 2.27 ± 0.62 and 3.16 ± 0.78 cfu/g; respective values
of rabbits that started dying late (that is, after 72 hours) were below
the limit of detection. Lethal infl uenza virus A H1N1 infection in two relatives with
autosomal dominant GATA-2 defi ciency Mean circulating LPS at 24 hours was 2.09 EU/
ml and 1.99 EU/ml respectively (P = NS). Mean LPS of the portal vein
of the sham and of the injury groups were 1.25 and 5.62 EU/ml (P =
0.047). Concentrations of TNFα in splenocyte supernatants are shown
in Figure 1. y
Methods We report three adult relatives with the autosomal dominant
GATA-2 defi ciency. P1 and his son P2 had a history of myelodysplastic
syndrome and a few episodes of mild respiratory infections. They
developed PVP by H1N1pdm which rapidly evolved to ARDS. They died
at the age of 54 and 31, respectively. y
Results Patients were heterozygous for a novel R396L mutation in
GATA2. Like other patients with GATA-2 defi ciency, the three relatives
had absence of peripheral NK and B cells and monocytopenia. However
a high number of plasma cells, which were found to be pauciclonal,
were observed in peripheral blood from P1 during H1N1pdm infection. P1 and P2 had normal levels of immunoglobulins and IgG antibodies
against common viruses. Microneutralization test showed that P1
produced normal titers of neutralizing antibodies against H1N1pdm
and against the previous annual H1N1 strain. Our results suggest that
a few clones of long-living memory B cells against IAV expanded in
P1; and that these cells produced cross-reactive antibodies against
H1N1pdm, similar to those recently described. During the fl u episode
P1 had a strong increase of IFNγ-producing T cells and of IFNγ
production. The Th1-related chemokines CXCL10 and CXCL9, as well as
IFNγ, MCP-1 and IL-8, were strongly elevated in serum from P1 and P2
in the course of H1N1pdm infection. g
Conclusion Early death after injury is not related to peripheral
endotoxemia and sepsis; bacterial translocation priming for enhanced
proinfl ammatory responses is a likely explanation. Eff ects of the common 34C>T variant of the AMPD1 gene on
immune function, multiorgan dysfunction and mortality in patients
with sepsis Eff ects of the common 34C>T variant of the AMPD1 gene on
immune function, multiorgan dysfunction and mortality in patients
with sepsis
B Ramakers1, E Giamarellos-Bourboulis2, M Coenen1, M Kox1,
J Van der Hoeven1, C Routsi2, A Savva2, I Perdios3, F Diamantea4,
D Sinapidis5, P Smits1, N Riksen1, P Pickkers1
1Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands;
2University of Athens, Medical School, Athens, Greece; 3Nafplion General
Hospital, Nafplio, Greece; 4‘G.Gennimatas’ General Hospital, Athens, Greece;
5Alexandra General Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P17 (doi: 10.1186/cc11955) Conclusion GATA-2 defi ciency is the fi rst described Mendelian
inborn error of immunity underlying severe IAV infection. Primary
immunodefi ciencies predisposing to severe IAV infections may debut,
even in adults without a history of previous severe infections. The
massive IFNγ-mediated cytokine storm may explain the fatal course of
H1N1pdm infection in our patients. Introduction Adenosine exerts anti-infl ammatory and tissue protective
eff ects during systemic infl ammation. While the anti-infl ammatory
properties may induce immunoparalysis and impede bacterial
clearance, the tissue protective eff ects might limit organ damage. The eff ects of a common loss-of-function variant of the adenosine
monophosphate deaminase 1 gene (AMPD1), which is associated with
increased adenosine formation, in patients with sepsis are unknown. Methods In a prospective cohort, genetic-association study, the
eff ects of the presence of the AMPD1 gene on immune function,
multiorgan dysfunction and mortality in septic patients was studied. Pneumosepsis patients (n = 402) and controls without infection (n =
101) were enrolled. P16 P16
Bacterial translocation primes proinfl ammatory responses and is
connected to early death in an experimental model of lethal injury
N Baxevanos, T Tsaganos, A Pistiki, D Droggiti, A Spyridaki,
E Giamarellos-Bourboulis
University of Athens, Medical School, Athens, Greece
Critical Care 2013, 17(Suppl 2):P16 (doi: 10.1186/cc11954) P18 P18
Exploring the translational disconnect between the murine and
human infl ammatory response: in vitro analysis of the dose–
response relationship of LPS and NFκB activation in murine and
human immune cells
EP McCarron, I Welters, D Williams, D Antoine, A Kipar
University of Liverpool, UK
Critical Care 2013, 17(Suppl 2):P18 (doi: 10.1186/cc11956) Introduction Infl ammation, as seen in sepsis and systemic infl ammatory
response, is dependent on the activation of the NFκB pathway through
Toll-like receptors (TLRs) [1]. Recreating an infl ammatory response
using lipopolysaccharide (LPS) can provide results that are diff erent to
clinical sepsis [2]. By examining NFκB activation in murine and human
cells, a species comparison can be made to investigate diff erences at
the cell level that may contribute to the translational disconnect seen
in vivo. Methods In the prospective, noninterventional study, we assessed the
correlation between the capillary leakage marker angiopoetin-2 and
serum levels of IgG and IgM in 41 patients with community-acquired
severe sepsis or septic shock on admission. Blood samples were
obtained during the fi rst 12 hours after admission to hospital. Results Mean age of patients (17 females) was 70 years. Median
APACHE II and SOFA scores at admission were 24 and 11, respectively. The mortality rate was 45%. Thirty-four percent of all patients had level
of IgG <650 mg/dl. The median concentration of angiopoietin-2 in the
hypo-IgG group was 11,958 pg/ml, which was not statistically diff erent
(Mann–Whitney; P >0.05) than in the rest of patients with normal levels
of IgG (15,688 pg/ml). The concentration of IgM <40 mg/dl was found in
only four patients (10%) and all died. Pearson’s correlation test showed
that the correlation between the concentrations of angiopoietin-2
and IgG (correlation coeffi cient 0.191) or IgM (correlation coeffi cient
0.0408), respectively, were not statistically signifi cant (P <0.05). Methods THP1 human monocytes (passages 9 to 11) and RAW 264.7
murine macrophages (passages 15 to 20) were cultured in RPMI-1640
and DMEM respectively and then challenged with LPS. After settling
for 24 hours, cells were dosed with six or seven doses of LPS. After
1 hour, nuclear extraction and proteins were separated by acrylamide
gel electrophoresis. Membranes where then immunoblotted for actin
and p65, followed by densitometric analysis in order to quantify the
amount of p65 that had translocated from the cytoplasm to the nucleus
(by subtraction from consistent nuclear actin). Bacterial translocation primes proinfl ammatory responses and is
connected to early death in an experimental model of lethal injury
N Baxevanos, T Tsaganos, A Pistiki, D Droggiti, A Spyridaki, y
,
,
,
itical Care 2013, 17(Suppl 2):P16 (doi: 10.1186/cc11954) Introduction Some cases of multiple trauma are rapidly deteriorating;
the mechanism was investigated. Introduction Some cases of multiple trauma are rapidly deteriorating;
the mechanism was investigated. Results In pneumosepsis patients and controls, a similar prevalence
of the 34C>T (rs17602729) mutation in the AMPD1 gene was found. Univariate logistic regression analysis showed a tendency of increased
mortality in patients with the CT genotype, compared with patients
with the CC genotype (OR 1.53; 95% CI 0.95 to 2.5). Moreover, carriers of
the CT genotype tended to suff er more from multiorgan dysfunction,
OR 1.4 (0.84 to 2.3) and 3.0 (0.66 to 13.8), for CT and TT, respectively
(P = 0.07). In septic carriers of the CT genotype, the ex vivo production
of TNFα by LPS-stimulated monocytes was attenuated (P = 0.005), Methods Forty-one rabbits were assigned into two groups; sham-
operated and subject to crush of the right femur. Survival was
recorded; peripheral blood was sampled for LPS measurement by the
kinetic QCL-1000 LAL assay; quantitative tissue growth was assessed
after death. Some rabbits were sacrifi ced at 48 hours; blood was
sampled from the portal vein for LPS measurement; splenocytes were Figure 1 (abstract P16). Stimulation of TNFα from isolated splenocytes. y
y
Figure 1 (abstract P16). Stimulation of TNFα from isolated splenocytes. Figure 1 (abstract P17). Kaplan–Meier curve for the 402 sepsis patients. Figure 1 (abstract P17). Kaplan–Meier curve for the 402 sepsis patients. Figure 1 (abstract P17). Kaplan–Meier curve for the 402 sepsis patients. Figure 1 (abstract P17). Kaplan–Meier curve for the 402 sepsis patients. S7 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 indicative for more pronounced immunoparalysis in these patients. See Figure 1. P18 i
Conclusion At present the hypothesis that increased microvascular
leakage is responsible for hypogammaglobulinemia in septic patients
could not be accepted. Studies on larger number of patients are
needed. In addition, it is necessary to further explore other possible
mechanisms, such as increased catabolism and consumption of
antibodies or inadequate synthesis of immunoglobulins, which could
also be responsible for hypogammaglobulinemia in sepsis. References y
Results Murine cells required higher doses of LPS compared with
human cells in order to detect p65 (human, 1 pg/ml to 100 ng/ml;
murine, 30 pg/ml to 1,000 ng/ml). THP1 cells showed a greater fold
increase in the p65:actin ratio compared with RAW 264.7 cells. Human
cells responded to lower concentrations of LPS. Murine cells appeared
to show a molecular resistance to lower doses, but their response was
very sensitive at higher doses. A dose–response relationship of LPS
dosing and NFκB activation was observed in both cell lines. P20 P20 Hypogammaglobulinemia in sepsis is not correlated to high
circulating angiopoietin-2 levels Conclusion The presence the 34C>T variant of the AMPD1gene is not
related to infection susceptibility; however, it is associated with more
pronounced immunoparalysis in patients with sepsis, and shows
a tendency towards increased mortality. Mechanistically, the anti-
infl ammatory eff ects of adenosine may account for this and apparently
overrule its tissue protective eff ects. U Kovačič1, F Starič2, M Kmet2, M Godnič2, R Kapš2
1Faculty of Medicine, University of Ljubljana, Slovenia; 2General Hospital Novo
Mesto, Slovenia
Critical Care 2013, 17(Suppl 2):P19 (doi: 10.1186/cc11957) Introduction Hypogammaglobulinemia has been frequently found
in adult patients with severe sepsis and septic shock. Furthermore, it
seems that at least a low serum level of IgM is correlated with higher
mortality in sepsis. The mechanisms of hypogammaglobulinemia in
septic shock have not yet been explained. It has been hypothesized
that outfl ow of immunoglobulins into the extravascular space due to
increased capillary permeability could reduce immunoglobulin serum
concentrations. Angiopoietin-2, which directly disrupts the endothelial
barrier, is markedly elevated in sepsis and other infl ammatory states
and its serum level has been correlated with microvascular leakage,
end-organ dysfunction and death in sepsis. TREM-1 levels are elevated in patients with liver cirrhosis
SD G
1 CR G h
2 P K ll h
2 N S
i1 M F
t
1 S Si
h 1Chelsea and Westminster Hospital, London, UK; 2Imperial College, London, UK
Critical Care 2013, 17(Suppl 2):P20 (doi: 10.1186/cc11958) 1Chelsea and Westminster Hospital, London, UK; 2Imperial College, London, UK
Critical Care 2013, 17(Suppl 2):P20 (doi: 10.1186/cc11958) References 1. Taccone FS, et al.: Gamma-globulin levels in patients with community-
acquired septic shock. Shock 2009, 32:379-385. 1. Taccone FS, et al.: Gamma-globulin levels in patients with community-
acquired septic shock. Shock 2009, 32:379-385. 2. Werdan K, et al.: Score-based immunoglobulin G therapy of patients with
sepsis: the SBITS study. Crit Care Med 2007, 35:2693-2701. g
Conclusion Immunoblotting for p65 is a reliable and reproducible
method to determine NFκB activation in cultured cells. Macrophages
are more responsive to LPS than monocytes [3] so diff erences between
cell lines would have been expected to be the reverse of what was
observed. The species diff erence in response to LPS may contribute
to the apparent disconnect between human and murine responses to
LPS and may partially explain the diffi culties of translating therapeutic
interventions into clinical human sepsis. 2. Werdan K, et al.: Score-based immunoglobulin G therapy of patients with
sepsis: the SBITS study. Crit Care Med 2007, 35:2693-2701. P22 Introduction Septic encephalopathy is a frequent complication in
severe sepsis but its pathogenesis and mechanisms are not fully
understood. Oxygen supply and utilization are critical for organ
function, especially for the brain, a tissue extremely dependent on
oxygen and glucose. Disturbances in oxygen utilization are common
in sepsis and a number of mitochondrial dysfunctions have been
described in diff erent tissues in septic animals as well as in septic
patients. Our group described mitochondrial dysfunctions in the brain
during experimental sepsis. l
Results TREM-1 expression is signifi cantly higher in the CA group
compared with HC across all monocyte subsets but not neutrophils,
even in the absence of sepsis (see Figure 1). There is no correlation
between cell surface TREM-1 expression and severity of cirrhosis by
Child–Pugh or MELD score. sTREM and sCD14 levels were elevated in
the CA group compared with HC (P = 0.0010 and 0.0016 respectively). In addition, plasma sTREM-1 levels correlated with disease severity
according to MELD score (R = 0.71, CI = 0.22 to 0.92 P = 0.012) and serum
bilirubin (R = 0.78,CI = 0.36 to 0.94, P = 0.004). There was no correlation
with either form of TREM-1 with sCD14 levels. There was no diff erence
in cell surface or soluble TREM-1 expression between blood and ascitic
fl uid monocytes in culture-negative, non-neutrophilic ascites. Methods Experimental sepsis was induced by endotoxemia (LPS 10 mg/
kg i.p.) in Sprague–Dawley rats and by polymicrobial fecal peritonitis in
Swiss mice. Brain glucose uptake was observed in vivo in endotoxemic
rats using positron emission tomography with [18F]fl uorodeoxyglucose
and autoradiography with 2-deoxy-14C-glucose. Results Mice with polymicrobial sepsis present hypoglycemia,
hyperlactatemia and long-term cognitive impairment. We observed
a rapid increase in the uptake of fl uorescent glucose analog 2-deoxy-
2-((7-nitro-2,1,3-benzoxadiazol-4-yl)amino)-D-glucose in brain slices
from septic mice in vitro. A similar increase in brain glucose uptake
was observed in vivo in endotoxemic rats. Remarkably, the increase
in glucose uptake started 2 hours after LPS injection, earlier than
other organs. The brains of mice with experimental sepsis presented
neuroinfl ammation, mitochondrial dysfunctions and oxidative stress,
but mitochondria isolated from septic brains generated less ROS in
vitro in the fi rst 24 hours. This led us to investigate the role of NADPH
oxidase, an enzyme induced during innate immune response, as a
potential source of reactive oxygen species in experimental sepsis. 1.
Olson JC, et al.: Intensive care of the patient with cirrhosis. Hepatology 2011,
54:1864-1872. P22 Inhibiting NADPH oxidase with apocynin acutely after sepsis prevented
cognitive impairment in mice. l
Conclusion Blood monocyte and soluble TREM-1 are elevated in
cirrhotic patients even in the absence of sepsis. Soluble TREM-1 levels
correlate with disease severity in cirrhosis. Further studies are ongoing
to ascertain the utility of TREM-1 as a biomarker in cirrhosis. Reference 1. Olson JC, et al.: Intensive care of the patient with cirrhosis. Hepatology 2011,
54:1864-1872. g
p
Conclusion Our data indicate that a bioenergetic imbalance and
oxidative stress is associated with the pathophysiology of septic
encephalopathy. We are observing a new metabolic phenotype in
the brain during sepsis, characterized by a rapid increase in glucose
uptake and mitochondrial dysfunctions that may be secondary to
infl ammation and hypoxia. P21
Eff ects of diff erent doses and serotypes of LPS on blood–brain
barrier permeability in Sprague–Dawley rats
E Senturk, F Esen, P Ergin Ozcan, G Orhun, N Orhan, N Arican, M Kucuk,
M Kaya
University of Istanbul, Turkey
Critical Care 2013, 17(Suppl 2):P21 (doi: 10.1186/cc11959) University of Istanbul, Turkey Introduction The blood–brain barrier (BBB) is highly restrictive of
the transport of substances between blood and the central nervous
system. Lipopolysaccharide (LPS) from Gram-negative bacteria was
reported to aff ect the permeability of the BBB. BBB disruption using a
LPS is used as a model of septic encephalopathy in mice. The present
study was designed to investigate the eff ects of diff erent doses and
serotypes of LPS on BBB integrity in Sprague–Dawley rats. Pathophysiology of sepsis-associated brain dysfunction: an
experimental study of cerebral microdialysis and mitochondrial
function
P Kurtz1, C Vargas-Lopes1, C Madeira1, I Mello1, R Panizzutti1, L C Azevedo2,
F A Bozza1
1Fiocruz, Rio de Janeiro, Brazil; 2Hospital Sirio e Libanes, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P23 (doi: 10.1186/cc11961) P22 to the ICU have an in-hospital mortality of up to 50% [1]. Microbial
translocation (MT) is the pathogenic mechanism implicated in SBP. The
triggering receptor expressed by myelocytes-1 (TREM-1) modulates
the immune response with resultant production of proinfl ammatory
cytokines and has been used as a biomarker in the diagnosis of bacterial
infection. We wish to evaluate the role of TREM-1 as a biomarker in
cirrhosis. Bioenergetic imbalance and oxidative stress in the pathophysiology
of septic encephalopathy
J D’Avila1, R Rodrigues2, H Castro-Faria-Neto1, M Oliveira3, F Bozza4
1Oswaldo Cruz Foundation – FIOCRUZ, Rio de Janeiro, Brazil; 2Federal
University of Rio de Janeiro (UFRJ) and D’Or Institute for Research and
Education (IDOR), Rio de Janeiro, Brazil; 3UFRJ, Rio de Janeiro, Brazil; 4Oswaldo
Cruz Foundation – FIOCRUZ and IDOR, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P22 (doi: 10.1186/cc11960) Bioenergetic imbalance and oxidative stress in the pathophysiology
of septic encephalopathy
J D’Avila1, R Rodrigues2, H Castro-Faria-Neto1, M Oliveira3, F Bozza4
1Oswaldo Cruz Foundation – FIOCRUZ, Rio de Janeiro, Brazil; 2Federal
University of Rio de Janeiro (UFRJ) and D’Or Institute for Research and
Education (IDOR), Rio de Janeiro, Brazil; 3UFRJ, Rio de Janeiro, Brazil; 4Oswaldo
Cruz Foundation – FIOCRUZ and IDOR, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P22 (doi: 10.1186/cc11960) Methods Blood samples were obtained from 18 healthy controls (HC)
and 29 cirrhotic patients (CA) as defi ned by clinicoradiological criteria. Disease severity was graded according to Child–Pugh class (median
10, range 5 to 13) and modifi ed end-stage liver disease (MELD) score
(median 14, range 6 to 21). Simultaneous ascitic fl uid samples were
taken from 10 patients in the CA group. Soluble TREM-1 and CD14 levels
(a surrogate marker of MT) were measured by ELISA. Flow cytometry
was used to quantify the expression of TREM-1 on monocytes and
neutrophils in blood and ascitic fl uid.i Introduction Septic encephalopathy is a frequent complication in
severe sepsis but its pathogenesis and mechanisms are not fully
understood. Oxygen supply and utilization are critical for organ
function, especially for the brain, a tissue extremely dependent on
oxygen and glucose. Disturbances in oxygen utilization are common
in sepsis and a number of mitochondrial dysfunctions have been
described in diff erent tissues in septic animals as well as in septic
patients. Our group described mitochondrial dysfunctions in the brain
during experimental sepsis. References 1. Bonizzi G, Karin M: Trends Immunol 2004, 25:280-288. 1. Bonizzi G, Karin M: Trends Immunol 2004, 25:280-288. Introduction Sepsis and spontaneous bacterial peritonitis (SBP) are
common sequelae in patients with cirrhosis. Cirrhotics admitted 2. Remick DG, Ward PA: Shock 2005, 24(Suppl 1):7-11. pp
3. Takashiba S, et al.: Infect Immun 1999, 67(11):5573-5578. Figure 1 (abstract P20). TREM-1 expression in healthy controls compared with cirrhotic patients. cirrhotic patients. Figure 1 (abstract P20). TREM-1 expression in healthy controls compared with cirrhotic patients. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S8 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Cholinergic modulation of hippocampal activity during septic
encephalopathy
2
2 Cholinergic modulation of hippocampal activity during septic
encephalopathy A Zivkovic1, CP Bengtson2, O Sedlaczek1, R Von Haken1, H Bading2, S Hofer1
1Universitätsklinikum Heidelberg, Germany; 2Universität Heidelberg, Germany
Critical Care 2013, 17(Suppl 2):P24 (doi: 10.1186/cc11962) A Zivkovic1, CP Bengtson2, O Sedlaczek1, R Von Haken1, H Bading2, S Hofer1
1Universitätsklinikum Heidelberg, Germany; 2Universität Heidelberg, Germany
Critical Care 2013, 17(Suppl 2):P24 (doi: 10.1186/cc11962) Introduction Septic encephalopathy is a sepsis-related brain dys-
function with a deterioration of cortical functions. The experimental
studies in the rat brain revealed a deranged neurotransmitter profi le
during septic encephalopathy. Glutamatergic synapses, essential in
learning and memory, undergo use-dependent changes in synaptic
strength, referred to as plasticity. Permanent strengthening of synapses
after a brief stimulus, termed long-term potentiation (LTP), was
discovered in the hippocampus, and here it has been most thoroughly
studied. Cholinergic neurotransmission plays an important role in
regulating the cognitive functions of the brain. It acts as a signal-to-
noise ratio modulator of sensory and cognitive inputs. The irregularities
in brain functions give rise to the symptoms of delirium, including
disorganized thinking and disturbances of attention and consciousness,
which in turn might aff ect learning and memory. Possible mechanisms
for cholinergic defi ciency include impairment of synaptic functions
of acetylcholine. Imbalances in the cholinergic system during sepsis
might therefore play an extensive role in the septic delirium. Results Of the 1,600 patients who gave consent to participate in the
study, 205 had an ECI. Sixty-four had a positive blood culture for E. coli. Fifty of the patients met one or more criteria for severe sepsis or septic
shock. The NLCR was signifi cantly higher (P <0.001) within the severe
sepsis group (median = 21.1 with quartiles 11.1 to 42.4) compared with
the group with no severe sepsis (median = 11.6 with quartiles 7.6 to
18.9). Conclusion The NLCR can be used as a biomarker of disease severity
even in ECIs. The biomarker reacts rapidly, is cheap and needs no extra
sampling. The higher the value, the higher the probability for severe
sepsis. A high value can even precede the development of severe
sepsis or septic shock. However, a low value never excludes neither
bacteremia nor severe sepsis. The method cannot be used in patients
with disturbances in neutrophil or lymphocyte levels due to other
causes than sepsis. Methods By using MRI imaging we identifi ed functional changes in the
hippocampal region of patients with severe sepsis. Pathophysiology of sepsis-associated brain dysfunction: an
experimental study of cerebral microdialysis and mitochondrial
function P Kurtz1, C Vargas-Lopes1, C Madeira1, I Mello1, R Panizzutti1, L C Azevedo2,
F A Bozza1 1Fiocruz, Rio de Janeiro, Brazil; 2Hospital Sirio e Libanes, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P23 (doi: 10.1186/cc11961) y
y
y
Methods Male Sprague–Dawley rats weighing 200 to 250 g were
used in the study. Rats were given two diff erent types of LPS (026:B6-
L5543 and 026:B6-L2762) in diff erent doses (3, 5, and 10 mg/kg; i.v.). Rectal temperature and arterial blood pressure measurements were
recorded for sepsis severity. The changes in the BBB permeability were
measured using the Evans blue (EB) and sodium fl uorescein (NaFl) dye
extravasation techniques 24 hours after LPS administration. Introduction Pathophysiology of brain dysfunction associated
with sepsis is still poorly understood. Potential mechanisms involve
oxidative stress, neuroinfl ammation and blood–brain barrier altera-
tions. Our purpose was to study the metabolic alterations and markers
of mitochondrial dysfunction in a clinically relevant model of septic
shock. Results Both LPS serotypes showed comparable arterial blood pressure
and rectal temperature recordings and the severity of the disease
increased with the increasing doses (5 mg/kg and 10 mg/kg) of LPS
and the mortality rates were found to be 29% and 63% respectively. The extravasated contents of EB and NaFl tracers did not signifi cantly
increase in brain parenchyma following the administration of diff erent
doses of LPS with diff erent serotypes. Methods Twelve anesthetized (midazolam/fentanyl/pancuronium),
invasively monitored, and mechanically ventilated pigs were allocated
to a sham procedure (n = 5) or sepsis (n = 7), in which peritonitis was
induced by intra-abdominal injection of autologous feces. Animals
were studied until spontaneous death or for a maximum of 24 hours. In addition to global hemodynamic and laboratory assessment,
intracranial pressure and cerebral microdialysis were assessed at
baseline, 6, 12, 18 and 24 hours after sepsis induction. After death,
brains were removed and brain homogenates were studied to assess
markers of mitochondrial dysfunction. f
Conclusion Our results showed no disruption to BBB by two diff erent
serotypes of LPS even administered in increasing doses. These result
indicate that the BBB integrity of Sprague–Dawley rats are resistant to
the eff ects of two diff erent serotypes of LPS. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S9 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P23). Pathophysiology of sepsis-associated brain dysfunction: an
experimental study of cerebral microdialysis and mitochondrial
function Results Critically ill ICU patients diagnosed with septic delirium using
the CAM-ICU method underwent diagnostic MRI scans. The serial MRI
analysis revealed increased signal intensity in the hippocampal region
in diff usion-weighted MRI (DWI). We used endotoxemia model to induce
sepsis in the rats. Electrophysiological analysis of the hippocampal
neurons in LPS-treated rats showed impaired LTP in the excitatory
synapses, as compared with controls. Application of physostigmine, a
blood–brain barrier permeable cholinesterase inhibitor, resulted in a
partial recovery of LTP in the hippocampal synapses of LPS-treated rats. Conclusion The patients with septic delirium show functional changes
in the hippocampus. Furthermore, we show that endotoxemia aff ects
synaptic plasticity in the rat hippocampus, suggesting the involvement
of this brain region in the pathophysiology of septic delirium. Moreover,
the eff ect of the cholinergic neurotransmission onto the induction
and maintenance of synaptic plasticity in the rat hippocampus during
endotoxemia suggests that cholinergic neurotransmission might play a
critical role in septic encephalopathy. Results All septic animals developed a hyperdynamic state associated
with lower arterial pressure, fever and organ dysfunction in comparison
with control animals. In the septic animals, we observed increased
brain dialysate glutamin levels at 12, 18 and 24 hours after sepsis
induction, as compared with control animals. Moreover, after analyzing
homogenates from the frontal cortex, we found higher concentrations
of glutamin and glutamate in septic as compared with control
animals (85.67 ± 14.98 vs. 28.77 ± 7.0; P = 0.01 and 132.1 ± 19.72 vs. 53.33 ± 16.83; P = 0.02, respectively). See Figure 1. Results All septic animals developed a hyperdynamic state associated
with lower arterial pressure, fever and organ dysfunction in comparison
with control animals. In the septic animals, we observed increased
brain dialysate glutamin levels at 12, 18 and 24 hours after sepsis
induction, as compared with control animals. Moreover, after analyzing
homogenates from the frontal cortex, we found higher concentrations
of glutamin and glutamate in septic as compared with control
animals (85.67 ± 14.98 vs. 28.77 ± 7.0; P = 0.01 and 132.1 ± 19.72 vs. 53.33 ± 16.83; P = 0.02, respectively). See Figure 1. P25 Cholinergic modulation of hippocampal activity during septic
encephalopathy
A Zivkovic1, CP Bengtson2, O Sedlaczek1, R Von Haken1, H Bading2, S Hofer
1Universitätsklinikum Heidelberg, Germany; 2Universität Heidelberg, Germany
Critical Care 2013, 17(Suppl 2):P24 (doi: 10.1186/cc11962) de Jager C, et al.: The neutrophil–lymphocyte count ratio in patients with
community acquired pneumonia. PLoS ONE 2012, 7:e46561. Cholinergic modulation of hippocampal activity during septic
encephalopathy
2
2 This fi nding was
further supported by the experimental recordings in the rat brains of
lipopolysaccharide (LPS)-treated rats using the electrophysiological
patch clamp technique. P25 P25
Neutrophil–lymphocyte count ratio as a biomarker of severe sepsis
in Escherichia coli infections in adults
LR Ljungstrom1, G Jacobsson1, R Andersson2
1Skaraborgs Sjukhus, Skövde, Sweden; 2Gothenburg University, Gothenburg,
Sweden
Critical Care 2013, 17(Suppl 2):P25 (doi: 10.1186/cc11963) P25
Neutrophil–lymphocyte count ratio as a biomarker of severe sepsis
in Escherichia coli infections in adults
LR Ljungstrom1, G Jacobsson1, R Andersson2
1Skaraborgs Sjukhus, Skövde, Sweden; 2Gothenburg University, Gothenburg,
Sweden
Critical Care 2013, 17(Suppl 2):P25 (doi: 10.1186/cc11963) Neutrophil–lymphocyte count ratio as a biomarker of severe sepsis
in Escherichia coli infections in adults LR Ljungstrom1, G Jacobsson1, R Andersson2 Critical Care 2013, 17(Suppl 2):P25 (doi: 10.1186/cc11963) Introduction The neutrophil–lymphocyte count ratio (NLCR) is an
easy to analyse biomarker reacting very early in the course of acute
infl ammation. It has previously been reported to correspond to
bacteremia and recently to disease severity in community-acquired
pneumonia [1]. We have looked at 205 consecutive patients with
Escherichia coli infections (ECI) and found the same to be true for
ECIs. This may be of great clinical importance since E. coli is the most
frequently isolated pathogen in patients with infections requiring in
hospital care. Conclusion We found higher concentrations of glutamate and
glutamin in brain tissues of septic animals as compared with control. Furthermore, glutamin concentrations increased over time in the
extracellular space as measured by cerebral microdialysis. These
fi ndings suggest an increased excitatory state that is potentially
associated with high energy expenditure. However, associations with
neuronal injury need further study. Methods This study is part of a 9-month consecutive study of
community-acquired severe sepsis and septic shock in adults at
Skaraborg Hospital in the western region of Sweden. The hospital
serves a population of 256,000 inhabitants and has approximately
60,000 annual visits to the ED. All patients admitted to the hospital
receiving intravenous antibiotic treatment within the fi rst 48 hours
of admission were evaluated for severe sepsis and septic shock. Upon
admission, two sets of blood cultures and other relevant cultures were
obtained from each patient as well as sampling for NLCR and venous
plasma lactate. The patient records were evaluated by one infectious
diseases specialist. Approximately 2,300 patients were diagnosed
as having a bacterial infection. From those, an informed consent to
participate in the study could be obtained from approximately 1,600
patients. Reference de Jager C, et al.: The neutrophil–lymphocyte count ratio in patients with
community acquired pneumonia. PLoS ONE 2012, 7:e46561. de Jager C, et al.: The neutrophil–lymphocyte count ratio in patients with
community acquired pneumonia. PLoS ONE 2012, 7:e46561. S10 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P26). P26
Obesity and infl ammatory markers in severe sepsis
P Simon1, D Thomas-Rüddel2, T Nemes1, K Reinhart2, F Bloos2, UX Kaisers1
1University Hospital of Leipzig, Germany; 2Center for Sepsis Control and Care,
Jena, Germany
Critical Care 2013, 17(Suppl 2):P26 (doi: 10.1186/cc11964) Introduction Chronic infl ammation has recently been recognized as
an important factor in the pathophysiology of obesity and associated
morbidities [1]. In this clinical study we aimed at identifying possible
eff ects of obesity on infl ammatory markers in severe sepsis. fl
Methods With institutional ethical committee approval, 243
consecutive patients treated for severe sepsis or septic shock in the
ICUs of two university hospitals over a period of 5 months were studied. Six patients were excluded due to cachexia, syndromal disorders or
missing clinical data. Diagnosis of sepsis was made according to SCCM
criteria. Serum levels of C-reactive protein (CRP, mg/l) and procalcitonin
(PCT, ng/ml) on day 1 of sepsis were compared among fi ve body mass
index (BMI) strata according to WHO defi nitions. Two groups (BMI <30,
normal weight, and BMI ≥30, obesity) were formed for further analysis,
and PCT was logarithmically transformed (LogPCT), resulting in normal
distribution. Statistical analysis was performed using a t test. y
Results Patients with BMI ≥30 had higher values of PCT and CRP
(Table 1). The diff erence in LogPCT was of borderline signifi cance (P =
0.052). However, patients with positive blood cultures had signifi cantly
higher LogPCT values (P = 0.017) (Figure 1). Diff erence in CRP was not
signifi cant (P = 0.09). The trends over all fi ve BMI strata (Table 1) were
not signifi cant. P27
Cytokine gene expression can predict infectious complications
following severe trauma
HD Torrance, K Brohi, CJ Hinds, MJ O’Dwyer
Bart’s Health NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P27 (doi: 10.1186/cc11965) Cytokine gene expression can predict infectious complications
following severe trauma
HD Torrance, K Brohi, CJ Hinds, MJ O’Dwyer
Bart’s Health NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P27 (doi: 10.1186/cc11965) gi
Conclusion Obesity with BMI ≥30 seems to be associated with an
increase in infl ammatory markers in patients with severe sepsis,
particularly in bacteraemia. The role of adipose tissue in severe sepsis
should therefore be studied in more detail. 1. Wellen K, et al.: Infl ammation, stress, and diabetes. J Clin Invest 2005,
115:1111-1119. 1. Wellen K, et al.: Infl ammation, stress, and diabetes. J Clin Invest 2005,
115:1111-1119. Introduction Identifying a group of patients at high risk of developing
infectious complications is the fi rst step in the introduction of eff ective
pre-emptive therapies in specifi c patient groups. Quantifying cytokine
gene expression also furthers our understanding of trauma-induced
immunosuppression. Our group has already demonstrated that a
predictive immunological signature derived from mRNA expression in
elective thoracic surgical patients accurately predicts pneumonia risk [1]. Methods In total, 121 ventilated polytrauma patients were recruited. mRNA was extracted from PaxGene tubes collected within 2 hours
of the initial insult, at 24 and 72 hours. T-helper cell subtype specifi c
cytokines and transcription factors mRNA was quantifi ed using qPCR. Ten healthy controls served as a comparator. Table 1 (abstract P26). PCT and CRP as median (IQR)
BMI
n
PCT (ng/ml)
CRP (mg/l)
18.5 to 24.9
196
4.8 (11.9)
153 (189)
≥30.0
47
7.3 (24.6)
228 (254)
18.5 to 24.9
101
4.9 (11.7)
147 (175)
25 to 29.9
95
4.4 (12.0)
157 (218)
30 to 34.9
32
6.9 (22.6)
212 (118)
35 to 39.9
7
7.4 (38.3)
241 (264)
≥40.0
8
11.4 (32)
278 (272) y
p
Results The Median Injury Severity Score (ISS) was 29. Time 0 bloods
demonstrated a reduction in TNFα†, IL-12§, IL-23‡, RORγT* and T bet§, and
an increase in IL-10* and IL-4† mRNA levels in comparison with the control
group (*P <0.0001, †P <0.001 to 0.0001, ‡P <0.01 to 0.001, §P <0.05 to 0.01). There was a positive correlation between ISS and IL-10‡ whilst both IL-23§ Figure 1 (abstract P27). Cytokine mRNA levels in trauma (time 0) and control groups. P27
Cytokine gene expression can predict infectious complications
following severe trauma
HD Torrance, K Brohi, CJ Hinds, MJ O’Dwyer
Bart’s Health NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P27 (doi: 10.1186/cc11965) S11 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods From November 2010 to November 2011 we collected clinical
data, drug administration, scores and PCT values of 420 consecutive
patients during hospitalization. Statistical analysis was made using
SPSS software. We calculated ICU mortality, 1-month mortality and
1-year mortality. Median percentage daily variation was calculated as:
(PCT day after – PCT of the date value) / PCT of the date value×100. PCT variation in the last 48 hours of hospitalization was calculated as:
(PCT at discharge – PCT at 48 hours before discharge) / PCT 48 hours
before discharge×100. We compared peak values in dead patients
versus alive patients. A logistic regression was performed in order to
assess mortality odds ratio. and RORγT‡ were negatively correlated at time 0. TNFα†, IL-10* and IL-27‡
increased and IFNγ†, IL-12*, IL-17A§, RORγT* and T bet* mRNA levels
decreased over the initial 24 hours. Subsequent bacteraemia (18/121
patients) was associated with a lower TNFα/IL-10 ratio‡ at baseline. Similarly, higher IL-10‡ and lower T bet‡ mRNA at 24 hours also predicted
later bacteraemic episodes. Development of pneumonia followed a
similar pattern. A multivariate logistical regression model proved highly
accurate in predicting infectious complications from mRNA analysis of
early blood samples. See Figure 1. y
Conclusion Cytokine gene expression patterns indicate an immediate
and sustained impairment in Th1, Th17 and innate immunity with
concurrent upregulation of the Th2 response following major trauma. The magnitude of this response predicts subsequent infectious
complications. y
Results Of the 420 patients, 63 (15%) died in the ICU, 12 (2.86%) died
1 month after ICU discharge and 16 (3.80%) died 1 year after ICU
discharge. PCT values were higher during the last day of hospitalization
in dead patients versus alive patients. PCT percentage variation during
the last 48 hours of hospitalization had a slower trend in patients who
died than in those who survived; these diff erences are even more
marked in patients who had a septic event. A slower descending trend
of daily PCT values was found in patients who died than in those who
survived. PCT peak levels during the ICU stay were higher in dead
patients with respect to alive ones. P27
Cytokine gene expression can predict infectious complications
following severe trauma
HD Torrance, K Brohi, CJ Hinds, MJ O’Dwyer
Bart’s Health NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P27 (doi: 10.1186/cc11965) At logistic regression analysis PCT
decrease in the last 48 hours <–30% (OR 3.71), PCT peak higher than
10 ng/ml (OR 2.38), and PCT last day/PCT peak ratio >50% (OR 2.064)
were ICU mortality risk factors. PCT values were a higher predictive ICU
mortality risk factor than SOFA and APACHE II scores. Other prognostic
factors were age and lactate values. Only age was a risk factor in
1-month and 1-year mortality. References 1. Jensen JU, Heslet L, Jensen TH, et al.: Procalcitonin increase in early
identifi cation of critically ill patients at high risk of mortality. Crit Care Med
2006, 34:2596-2602. 2. Fritz HG, Brandes H, Bredle DL, et al.: Post-operative hypoalbuminaemia and
procalcitonin elevation for prediction of outcome in cardiopulmonary
bypass surgery. Acta Anaesthesiol Scand 2003, 47:1276-1283. Results We analyzed 150 consecutive episodes of severe sepsis (16%)
or septic shock (84%) in the ICU. The median age of the patients was
64 (interquartile range, 48.7 to 71) years; the main sources of infection
were intra-abdomen (45%) and respiratory (38%); 70.7% had medical
diseases. The 28-day mortality was 22.7%. The profi le of death patients
were men (64.7%, n =22), with signifi cantly higher average age (63 vs. 57 years; P = 0.049), as well as clinical severity scores, APACHE II (29.8 vs. 24.1; P <0.001) and SOFA (12.1 vs. 8.9; P <0.001) and major dysfunction
organ number (4.6 vs. 3.6; P <0.001). Bilirubin was the best predictor of
28-day mortality with the largest AUC (0.71), followed by hemoglobin
(0.69) and C3 (0.67). The multivariate logistic regression was adjusted
for three risk parameters, hemoglobin (OR: 0.68; 95% CI: 0.51 to 0.94),
bilirubin (OR:1.63; 95% CI: 1.08 to 2.45) and white blood cells (OR:1.04;
95% CI: 1.01 to 1.08) and with these parameters a ROC analysis was
performed, giving an AUC of 0.77 (0.69 to 0.84). A cohort study of routinely used sepsis biomarkers and 28-day
mortality
2 M De La Torre-Prados1, A Garcia-De la Torre2, C Trujillano-Férnández1
1Hospital Virgen de la Victoria, Málaga, Spain; 2Hospital Puerto Real, Cádiz,
Spain M De La Torre-Prados1, A Garcia-De la Torre2, C Trujillano-Férnández1
1Hospital Virgen de la Victoria, Málaga, Spain; 2Hospital Puerto Real, Cádiz,
Spain p
Critical Care 2013, 17(Suppl 2):P28 (doi: 10.1186/cc11966) Introduction The evaluation of sepsis severity is complicated by the highly
variable and nonspecifi c nature of clinical signs and symptoms. We studied
routinely used biomarkers together with clinical parameters to compare
their prognostic value for severe sepsis and evaluate their usefulness. Conclusion PCT is a good prognostic marker and is strongly correlated
to the clinical status and gravity of the patients, so PCT seems to be a
useful marker in an intensive care scenario. Methods A cohort study of 150 patients >18 years with severe sepsis
according to the Surviving Sepsis Campaign, in an ICU of a university
hospital. Demographic, clinical parameters and coagulation, infection
and infl ammation parameters during the fi rst 24 hours from severe
sepsis or septic shock onset were studied. Descriptive and comparative
statistical analysis was performed using SPSS version 15.0 (SPSS Inc.,
Chicago, IL, USA). Changes in circulating procalcitonin versus C-reactive protein in
predicting evolution of infectious disease in febrile, critically ill
patients Changes in circulating procalcitonin versus C-reactive protein in
predicting evolution of infectious disease in febrile, critically ill
patients S Hoeboer1,2, J Groeneveld1,2 S Hoeboer1,2, J Groeneveld1,2
1VU University Medical Center, Amsterdam, the Netherlands; 2Erasmus
Medical Center, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P30 (doi: 10.1186/cc11968) S Hoeboer , , J Groeneveld ,
1VU University Medical Center, Amsterdam, the Netherlands; 2Erasmus
Medical Center, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P30 (doi: 10.1186/cc11968) Introduction Although absolute values for C-reactive protein (CRP) and
procalcitonin (PCT) are well known to predict sepsis in the critically ill, it
remains unclear if and how changes in CRP and PCT predict evolution
of infectious disease and how they compare in this respect. Introduction Although absolute values for C-reactive protein (CRP) and
procalcitonin (PCT) are well known to predict sepsis in the critically ill, it
remains unclear if and how changes in CRP and PCT predict evolution
of infectious disease and how they compare in this respect. Conclusion The assessment of routine biomarkers (bilirubin, white
blood cells and hemoglobin) may be a helpful tool in the decision-
making process at the bedside, for the evaluation of early ICU admission
of recoverable patients, as indicators of infl ammatory response, organ
dysfunction or catabolism level, and their signifi cant predictive value
on mortality. Methods In 72 critically ill patients with new-onset fever, CRP and
PCT were measured on day 0, 1, 2 and 7 after inclusion, and their
clinical course was documented over 1 week with follow-up to day 28. Infection was microbiologically defi ned, as was bloodstream infection;
septic shock was defi ned as infection plus shock. p
Reference 1. White M, et al.: Chest 2011, 139:626-632. y
Reference i
Results From peak at day 0 to 2 to day 7, CRP decreases most when
(bloodstream) infection and septic shock (day 0 to 2) resolve and
increases most when complications such as a new (bloodstream)
infection or septic shock (day 3 to 7) supervene (area under the receiver
operating characteristic curve 0.70 or higher, P = 0.04 or lower). PCT
decreases most when septic shock resolves (AUC 0.72, P = 0.007) and
increases most when a new bloodstream infection or septic shock
supervenes (AUC 0.82 or higher, P <0.001). The day 7 value of PCT rather
than of CRP was predictive for 28-day outcome (AUC 0.70, P = 0.005). i
Results From peak at day 0 to 2 to day 7, CRP decreases most when
(bloodstream) infection and septic shock (day 0 to 2) resolve and
increases most when complications such as a new (bloodstream)
infection or septic shock (day 3 to 7) supervene (area under the receiver
operating characteristic curve 0.70 or higher, P = 0.04 or lower). PCT
decreases most when septic shock resolves (AUC 0.72, P = 0.007) and
increases most when a new bloodstream infection or septic shock
supervenes (AUC 0.82 or higher, P <0.001). The day 7 value of PCT rather
than of CRP was predictive for 28-day outcome (AUC 0.70, P = 0.005). Conclusion The data, obtained during ICU-acquired fever and infections,
suggest that CRP and PCT changes predict the course of infectious
disease and its complications. CRP may be favoured over PCT courses
in decisions on appropriateness and duration of antibiotic treatment,
whereas PCT rather than CRP courses may help predicting complications
such as bloodstream infection, septic shock and mortality. 1. Glickman SW, Cairns CB, Otero RM, et al.: Disease progression in
hemodynamically stable patients presenting to the emergency
department with sepsis. Acad Emerg Med 2010, 17:383-390. 1. Glickman SW, Cairns CB, Otero RM, et al.: Disease progression in
hemodynamically stable patients presenting to the emergency
department with sepsis. Acad Emerg Med 2010, 17:383-390. Procalcitonin as prognostic marker of mortality Introduction Diff erentiation of acute heart failure from infection in
patients with respiratory symptoms and a history of congestive heart
failure (CHF) is challenging due to overlap of clinical symptoms and
X-ray fi ndings. The BACH study found higher mortality rates if patients
presenting with dyspnea were treated with antibiotics and their
procalcitonin (PCT) levels were low indicating absence of bacterial
infection. Yet the BACH study was observational and causal inference
cannot be drawn. Herein, we analyzed the eff ects of PCT-guided
antibiotic stewardship in CHF patients from a previous trial (ProHOSP). p
p
p
Methods This is a secondary analysis of a previous randomized trial
of adult ED patients with respiratory symptoms and a history of CHF. Patients were randomized to administration of antibiotics based on a
PCT algorithm (PCT group) or standard guidelines without knowledge
of PCT levels (control group). The primary endpoint of this analysis is
the risk of adverse outcome defi ned as death or ICU admission within
30 days after ED admission. Figure 1 (abstract P32). Pro-ADM transpulmonary gradient at diff erent
time points. Results A total of 233 patients met the inclusion criteria, with 116 in
the PCT-guided group and 117 in the control group. In the subgroup
of patients with low initial PCT levels <0.25 ng/l (n = 110), PCT-guided
patients had a signifi cant reduction in antibiotic exposure (mean 3.7
vs. 6.5 days, diff erence –2.8 (95% CI –4.4, –1.2), P <0.001). Furthermore,
PCT-guided patients had a signifi cant lower risk for death and ICU
admission (4% vs. 20%, odds ratio 6.0 (1.3, 28.2), P = 0.02). See Figure 1. Conclusion In CHF patients with suspicion of respiratory infection,
use of a PCT protocol resulted in a signifi cant decrease of antibiotic
exposure and signifi cantly improved outcomes in patients with low PCT
levels indicating absence of bacterial infection. Whether inadequate
antibiotic therapy in these CHF patients requiring diuretic treatment
explains this diff erence in clinical outcomes needs verifi cation. Methods In this prospective cohort study, 39 patients undergoing
cardiac surgery using CPB were included. Blood was collected before
surgery (T0), after induction of anesthesia (T1), after termination of CPB
(T2), at ICU arrival (T3) and 3 hours (T4), 6 hours (T5) and 18 hours (T6)
after arrival. Pro-ADM was measured with a sandwich immunoassay. Primary endpoints were length of ICU and hospital stay (ICU-LOS,
hospital-LOS). Procalcitonin as prognostic marker of mortality p
Results An increase of arterial and venous pro-ADM plasma con-
centrations was observed after surgery. Immediately after termination of
CPB the venous concentration was signifi cantly lower than arterial pro-
ADM concentration, but at T6 the venous concentration was signifi cantly
higher, indicating a switch from a negative to positive transpulmonary
gradient (Figure 1). The pro-ADM venous–arterial diff erence at T5 was a
signifi cant predictor of ICU-LOS (P = 0.032) and the diff erence at T3 was a
signifi cant predictor of hospital-LOS (P = 0.001). Figure 1 (abstract P31). Time to adverse outcome according to group
allocation. gi
Conclusion We found that the transpulmonary gradient of pro-ADM
was a predictor for ICU-LOS and hospital-LOS at T3 and T5, respectively. Pro-ADM might be a promising marker for prediction on outcome of
patients undergoing cardiac surgery on CPB. The transpulmonary shift
of pro-ADM might be caused by an infl ammatory response. References 1. Apostolakis E, et al.: J Card Surg 2010, 25:47-55. 1. Apostolakis E, et al.: J Card Surg 2010, 25:47-55. 2. Kamei M, et al.: Acta Anaesthesiol Scand 2004, 48:980-985. 3. Ueda S, et al.: Am J Respir Crit Care Med 1999, 160:132-136. 1. Apostolakis E, et al.: J Card Surg 2010, 25:47 55. 2. Kamei M, et al.: Acta Anaesthesiol Scand 2004, 48:980-985. 3. Ueda S, et al.: Am J Respir Crit Care Med 1999, 160:132-136. 2. Kamei M, et al.: Acta Anaesthesiol Scand 2004, 48: 2. Kamei M, et al.: Acta Anaesthesiol Scand 2004, 48:980 985. 3. Ueda S, et al.: Am J Respir Crit Care Med 1999, 160:132-136. Procalcitonin as prognostic marker of mortality Procalcitonin as prognostic marker of mortality
S Zampieri1, P Bettonte2, M Ortolani1, G Frison1, V Schweiger1, L Gottin3,
E Polati1
1Policlinico G.B. Rossi, Verona, Italy; 2Santa Chiara Hospital, Trento, Italy;
3Ospedale Maggiore Civile, Verona, Italy
Critical Care 2013, 17(Suppl 2):P29 (doi: 10.1186/cc11967) y
Conclusion The data, obtained during ICU-acquired fever and infections,
suggest that CRP and PCT changes predict the course of infectious
disease and its complications. CRP may be favoured over PCT courses
in decisions on appropriateness and duration of antibiotic treatment,
whereas PCT rather than CRP courses may help predicting complications
such as bloodstream infection, septic shock and mortality. Introduction We analyze procalcitonin (PCT) as a prognostic marker, in
order to assess the clinical impact of a daily PCT measure. Introduction We analyze procalcitonin (PCT) as a prognostic marker, in
order to assess the clinical impact of a daily PCT measure. S12 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P32). Pro-ADM transpulmonary gradient at diff erent
time points. P31
Procalcitonin-guided antibiotic therapy in patients with congestive
heart failure and suspicion of lower respiratory tract infection:
results from a randomized trial
P Schuetz, E Grolimund, A Kutz, S Haubitz, B Mueller
Kantonsspital Aarau, Switzerland
Critical Care 2013, 17(Suppl 2):P31 (doi: 10.1186/cc11969) Introduction Diff erentiation of acute heart failure from infection in
patients with respiratory symptoms and a history of congestive heart
failure (CHF) is challenging due to overlap of clinical symptoms and
X-ray fi ndings. The BACH study found higher mortality rates if patients
presenting with dyspnea were treated with antibiotics and their
procalcitonin (PCT) levels were low indicating absence of bacterial
infection. Yet the BACH study was observational and causal inference
cannot be drawn. Herein, we analyzed the eff ects of PCT-guided
antibiotic stewardship in CHF patients from a previous trial (ProHOSP). Methods This is a secondary analysis of a previous randomized trial
of adult ED patients with respiratory symptoms and a history of CHF. Patients were randomized to administration of antibiotics based on a
PCT algorithm (PCT group) or standard guidelines without knowledge
of PCT levels (control group). The primary endpoint of this analysis is
the risk of adverse outcome defi ned as death or ICU admission within
30 days after ED admission. P33 The median age of the patients was S13 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 been standardized in early goal-oriented therapies, which may benefi t
from circulating biomarkers for early risk stratifi cation. We aimed to
evaluate the prognostic value of presepsin (sCD14-ST), a novel marker
of bacterial infection. 64 (interquartile range, 53 to 72) years; the main sources of infection
were respiratory tract (46%) and intra-abdomen (21%). The 28-day
mortality was 32.5%. The profi le of death patients had a signifi cantly
higher average age (64.7 vs. 57.6 years; P = 0.024), as well as clinical
severity scores, APACHE II (26.6 vs. 23; P = 0.006) and SOFA (11.6 vs. 89.2;
P <0.001). Kaplan–Meier survival analysis was signifi cant. P = 0.0017 for
patients with pADM <1.2 nmol/l. Cox regression analysis also showed
statistical signifi cance (P = 0.0033) and a likelihood ratio = 1.18 per each
1 nmol/l increase in pADM. Methods We performed a nested case–control study from the
randomized controlled Albumin Italian Outcome Sepsis (ALBIOS)
trial, enrolling patients with severe sepsis or septic shock from 100
ICUs in Italy. Fifty survivors and 50 nonsurvivors at ICU discharge
were selected, matched for age, sex, center and time of enrollment
after inclusion criteria were present. EDTA-plasma samples were
collected at days 1, 2 and 7 after enrolment for presepsin (immune-
chemiluminescence assay PATHFAST Presepsin, URL 320 pg/ml, CV 5%;
Mitsubishi Chemicals) and procalcitonin assay (PCT, Elecsys BRAHMS
Cobas® PCT, URL 0.046 ng/ml, CV 8.8%; Roche Diagnostics). p
Conclusion The protein pADM is an important prognostic biomarker
of survival when measured on admission of septic patients to the ICU. References 1. Pezzilli R, Barassi A, Pigna A, et al.: Time course of proadrenomedullin in the
early phase of septic shock. A comparative study with other
proinfl ammatory proteins. Panminerva Med 2012, 54:211-217. g
g
Results Clinical characteristics were similar between the two groups,
except for a worse SOFA score at day 1 in decedents. Presepsin at
day 1 was signifi cantly higher in decedents (2,268 (1,145 to 4,305) pg/
ml, median (Q1 to Q3)) than in survivors (1,184 (855 to 2,158) pg/ml,
P = 0.001), while PCT did not diff er (18.5 (3.3 to 45.7) vs. 10.8 (2.6 to
46.4) ng/ml, P = 0.31). P33 P33
Pro-adrenomedullin as prognostic biomarker in the sepsis
M De La Torre-Prados, A Garcia-De la Torre, A Enguix, M Mayor,
N Zamboschi, C Trujillano-Fernández, A Garcia-Alcantara
Hospital Virgen de la Victoria, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P33 (doi: 10.1186/cc11971) Pro-adrenomedullin as prognostic biomarker in the sepsis
M De La Torre-Prados, A Garcia-De la Torre, A Enguix, M Mayor,
N Zamboschi, C Trujillano-Fernández, A Garcia-Alcantara
Hospital Virgen de la Victoria, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P33 (doi: 10.1186/cc11971) Figure 1 (abstract P31). Time to adverse outcome according to group
allocation. p
g
,
g , p
Critical Care 2013, 17(Suppl 2):P33 (doi: 10.1186/cc11971) Introduction Measurement of biomarkers is a potential approach to
early assessment and prediction of mortality in septic patients. The
purpose of this study was to ascertain the prognostic value of pro-
adrenomedullin (pADM), measured in all patients admitted to the ICU
of our hospital with a diagnosis of severe sepsis or septic shock during
1 year. P32
Pro-adrenomedullin as a clinical predictor after cardiac surgery
J Van Fessem, F De Graaf, J Van Paassen, S Arbous
LUMC, Leiden, the Netherlands
Critical Care 2013, 17(Suppl 2):P32 (doi: 10.1186/cc11970) y
Methods A cohort study of 117 patients >18 years with severe sepsis
according to the Surviving Sepsis Campaign, in an ICU of a university
hospital. Demographic, clinical parameters and pADM, C-reactive
protein and procalcitonin were studied during 1 year. Descriptive and
comparative statistical analysis was performed using the statistical
software packages Statistica Stat Soft Inc 7.1 and MedCalc 9.2.1.0. Results We analyzed 117 consecutive episodes of severe sepsis (15%)
or septic shock (85%) in the ICU. The median age of the patients was Introduction Pulmonary complications after cardiac surgery like ARDS
are frequent and linked to high mortality [1]. Pro-adrenomedullin
(pro-ADM) has a possible role in the development of ARDS [2] and a
positive correlation between levels of pro-ADM and infl ammation
was found [3]. In this study we investigated whether intraoperative
and postoperative pro-ADM transpulmonary gradient could predict
postoperative morbidity. g
Results We analyzed 117 consecutive episodes of severe sepsis (15%)
or septic shock (85%) in the ICU. Early IL-6 response in sepsis is correlated with mortality and
severity score P Srisangthong, A Wongsa, P Kittiworawitkul, A Wattanathum
Phramongkutklao Hospital, Bangkok, Thailand
C iti
l C
2013 17(S
l 2) P34 (d i 10 1186/
11972) P Srisangthong, A Wongsa, P Kittiworawitkul, A Wattanathum
Phramongkutklao Hospital, Bangkok, Thailand
C iti
l C
2013 17(S
l 2) P34 (d i 10 1186/
11972) g
p
g
Critical Care 2013, 17(Suppl 2):P34 (doi: 10.1186/cc11972) Introduction IL-6, a proinfl ammatory cytokine, is synthesized from fi bro-
blasts, T lymphocytes, endothelial cells and monocytes. It serves as an
important mediator during the acute phase response to infl ammation in
sepsis. We hypothesized that the plasma IL-6 is correlated with mortality
and severity scores in critically ill patients with sepsis. Methods We conducted a prospective study of plasma IL-6 level at the
initial phase of sepsis and the risk of mortality. A total of 203 patients
with sepsis, who were admitted to the medical ICU at Phramongkutklao
Hospital, Bangkok during January to December 2011, were analyzed. Serum IL-6, C-reactive protein (CRP), and lactate were measured within
the fi rst 24 hours of ICU admission. Severity scores (APACHE II, SAP II,
and SOFA scores) were measured. The primary outcome variable was
28-day all-cause mortality. Conclusion Early presepsin measurements may provide important
prognostic information in patients with severe sepsis or septic shock,
and may be of crucial importance for early risk stratifi cation. P36 References References
1. Pathan N, et al.: Crit Care Med 2005, 33:1839-1844. 2. Harbarth S, et al.: Am J Respir Crit Care Med 2001, 164:396-402. 1. Pathan N, et al.: Crit Care Med 2005, 33:1839-1844. 2. Harbarth S, et al.: Am J Respir Crit Care Med 2001, 164:396-402. p
p
y
Methods This study was conducted as a single-center retrospective
study. Blood samples were collected from patients admitted to the
emergency room at Fukuoka University Hospital between June 2010
and October 2012. We enrolled 254 patients with suspected sepsis and
other disease patients. We classifi ed the patients into an AKI group
according to the RIFLE criteria (Risk n = 52, Injury n = 39, Failure n = 41,
Loss of kidney function and End-stage kidney disease n = 7) and a non-
AKI group (n = 115). The AKI patient group was further classifi ed into a
sepsis group and a nonsepsis group in each AKI stage and we analyzed
the diagnostic accuracy of presepsin in patients with sepsis. P33 Presepsin decreased over time in survivors, but
remained elevated in decedents (974 (674 to 1,927) vs. 2,551 (1,438
to 5,624) pg/ml at day 7, P = 0.02 for time–survival interaction); PCT
decreased similarly in the two groups (P = 0.19). Patients with early
elevated presepsin had worse SOFA score, higher number of MOFs,
hemodynamic instability (lower mean arterial pressure at baseline
and after 6 hours), and mortality rate at 90 days (75% vs. 42%, log-
rank P <0.001). The association between presepsin and outcome was
more marked in patients with late enrollment (6 to 24 hours), and in
septic shock. Early presepsin had better prognostic accuracy than PCT
(AUROC 0.69 vs. 0.56, P = 0.07), and improved discrimination over SOFA
score, especially in septic shock. pl
y p
2. Wang RL, Kang FX: Prediction about severity and outcome of sepsis by
proatrial natriuretic peptide and pro-adrenomedullin. Chin J Traumatol
2010, 13:152-157. Usefulness of presepsin in the diagnosis of sepsis in acute kidney
injury patients Usefulness of presepsin in the diagnosis of sepsis in acute kidney
injury patients Usefulness of presepsin in the diagnosis of sepsis in acute kidney
injury patients
Y Nakamura, H Ishikura, T Nishida, Y Kawanno, R Yuge, R Ichiki, A Murai
Fukuoka University, Fukuoka, Japan
Critical Care 2013, 17(Suppl 2):P36 (doi: 10.1186/cc11974) y
y
Results We found that the overall 28-day mortality was 46% (93 out of 203
patients). There was a signifi cantly positive correlation between mortality
rate and plasma IL-6 (survivors vs. nonsurvivors; 74 (4.4 to 1,718) vs. 206
(19 to 5,000) pg/ml, P <0.05), lactate (survivors vs. nonsurvivors; 1.65 (0.7
to 11.61) vs. 2.47 (0.94 to 19.13) mmol/l, P <0.05), but not CRP levels (P =
0.24). Compared with the patients with plasma IL-6 <100 pg/ml, septic
patients with IL-6 levels ≥100 were associated with an increased 28-day
mortality with the odd ratio of 2.99 (95% CI 1.42 to 6.29, P <0.05). We
also found that plasma IL-6 levels were well correlated with APACHE II
(P <0.05), SAPS II (P <0.05), and SOFA (P <0.05) scores. Y Nakamura, H Ishikura, T Nishida, Y Kawanno, R Yuge, R Ichiki, A Murai
Fukuoka University, Fukuoka, Japan
Critical Care 2013, 17(Suppl 2):P36 (doi: 10.1186/cc11974) Introduction The level of presepsin is useful for diff erentiating sepsis
from noninfectious systemic infl ammatory response syndrome. It has
been reported that the presepsin levels in patients with chronic renal
failure are abnormally high. However, there are no studies investigating
the usefulness of presepsin for diagnosis of sepsis in patients with acute
kidney injury (AKI). Our purpose of this study is to clarify the diagnostic
accuracy of presepsin in patients with AKI and the relationship between
presepsin level and AKI severity. Conclusion The initial phase plasma IL-6 levels were correlated with
severity and mortality in critically ill patients with sepsis. Serum levels of presepsin refl ects the APACHE II and SOFA scores in
patients with sepsis Serum levels of presepsin refl ects the APACHE II and SOFA scores in
patients with sepsis
R Sato, Y Suzuki, M Sato, G Takahashi, M Kojika, Y Inoue, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P37 (doi: 10.1186/cc11975) R Sato, Y Suzuki, M Sato, G Takahashi, M Kojika, Y Inoue, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P37 (doi: 10.1186/cc11975) Methods We analyzed data from a large multinational database of
patients with CAP (CAPO) [2] between 2001 and 2012. We performed
logistic regression analysis and the area under the receiver operating
characteristics (AUC) curve to study the association of these variables
with the diagnosis of Legionella. Introduction As a method for the diagnosis of sepsis, we previously
reported an ELISA method for measuring the serum levels of presepsin. That method, however, took approximately 2 hours to yield results.i Introduction As a method for the diagnosis of sepsis, we previously
reported an ELISA method for measuring the serum levels of presepsin. That method, however, took approximately 2 hours to yield results. Methods To resolve this problem, we later developed a simplifi ed
assay kit making use of immunochromatography (Point of Care test;
POC test), and are currently evaluating the usefulness of this kit for
diagnosis of sepsis and evaluation of its severity. In 21 septic patients
with sepsis, we analyzed the serum levels of presepsin in relation to
APACHE II and SOFA scores. pp
y
y
Methods To resolve this problem, we later developed a simplifi ed
assay kit making use of immunochromatography (Point of Care test;
POC test), and are currently evaluating the usefulness of this kit for
diagnosis of sepsis and evaluation of its severity. In 21 septic patients
with sepsis, we analyzed the serum levels of presepsin in relation to
APACHE II and SOFA scores. g
g
Results Data for 8,278 CAP patients were analysed; the infectious
organism was known in 2,321 cases (28%), including a total of 101
patients with urinary antigen-confi rmed Legionnaires’ disease and
983 patients with confi rmed pneumococcal disease. All variables were
predictors for Legionella with odds ratios ranging from 1.002 to 5.767. Combining the variables in a joint logistic regression model showed a
high predictive accuracy with an AUC of 0.86. Results APACHE II and SOFA scores at baseline were 28.7 ± 7.5 and
10.8 ± 3.9, respectively, in these patients. Host genetic variants associated with community-acquired
pneumonia LE Salnikova1, TV Smelaya1, VV Moroz1, A Golubev1, AV Rubanovich2
1V.A. Negovsky Research Institute of General Reanimatology, Moscow, Russia;
2N.I. Vavilov Institute of General Genetics, Russian Academy of Sciences,
Moscow, Russia
Critical Care 2013, 17(Suppl 2):P40 (doi: 10.1186/cc11978) Introduction Serum nitrite/nitrate (NOx) levels in the early phase of
septic acute lung injury (ALI)/acute respiratory distress syndrome
(ARDS) have recently been reported to possibly play a key role in
pathogenesis of ALI/ARDS. p
g
Methods NOx levels in the early phase of septic ALI/ARDS were
measured by autoanalyzer (TCI-NOX 1000; Tokyo Kasei Kogyo Co., Ltd,
Tokyo, Japan). Cytokine was measured by ELISA (Medogenix, Fleurus,
Belgium). Introduction It has been shown that polymorphic variants at some
host genes can modify risk of community-acquired pneumonia (CAP),
including those critical for the host response to CAP – innate immune
system, the lung’s defense against inhaled microorganisms, and
inhibition of fi brinolysis and the renin–angiotensin system. The aim of
the study was to analyze polymorphisms in genes potentially relevant
to CAP pathogenesis mechanisms to reveal novel and confi rm reported
genetic risk factors in the general Russian population. g
Results Both NOx and TNFα levels were signifi cantly higher in the ARDS
group than in the ALI group. A negative correlation was found between
the PaO2/FIO2 (P/F) ratio and serum NOx levels. In addition, a positive
correlation was found between the TNFα and serum NOx levels. The
30-day, 60-day and 90-day mortality rates were 8.7%, 15.2% and 19.6%,
respectively, in the patients with ALI/ARDS. There were no diff erences
in the P/F ratio, serum NOx levels or TNFα levels in the early phase
of ALI/ARDS between the 30-day survival and death groups. On the
other hand, the P/F ratio, serum NOx levels and TNFα levels in the early
phase of ALI/ARDS were signifi cantly higher in the 60-day and 90-day
death groups than in the corresponding survival groups. There were
no signifi cant diff erences in the 90-day mortality rates between the ALI
and ARDS groups.i Methods Patients with CAP (n = 334), volunteers without a previous
history of CAP, control group A (n = 141) and a second control
group B (n = 314) were included in the study. P38 P38
Relationship between serum nitrite/nitrate levels in the early phase
of septic acute lung injury and prognosis
M Sato, Y Suzuki, T Masuda, G Takahashi, M Kojika, Y Inoue, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P38 (doi: 10.1186/cc11976) References 1. Fiumefreddo R, Zaborsky R, Haeuptle J, et al.: Clinical predictors for
Legionella in patients presenting with community-acquired pneumonia
to the emergency department. BMC Pulm Med 2009, 9:4. 2. Ramirez JA: Fostering international multicenter collaborative research: the
CAPO Project. Int J Tuberc Lung Dis 2007, 11:1062-1065. Serum levels of presepsin refl ects the APACHE II and SOFA scores in
patients with sepsis These were signifi cant
correlations between the serum presepsin levels and the APACHE II
score, and also between the serum presepsin levels and the SOFA score. Conclusion Furthermore, there was also a signifi cant correlation
between the results of the POC test and the serum presepsin levels. These results indicate that measurement of the serum presepsin might
be useful for evaluating the severity of sepsis. Conclusion This analysis validates the Legionella score in an inde-
pendent sample and shows high diagnostic accuracy. Interventional
trials with adapted antibiotic regimes for non-inferiority in a real live
population are warranted. R f Compared values of presepsin (sCD14-ST) and procalcitonin as early
markers of outcome in severe sepsis and septic shock: a preliminary
report from the Albumin Italian Outcome Sepsis (ALBIOS) study Compared values of presepsin (sCD14-ST) and procalcitonin as early
markers of outcome in severe sepsis and septic shock: a preliminary
report from the Albumin Italian Outcome Sepsis (ALBIOS) study
P Caironi1, S Masson2, E Spanuth3, R Thomae4, R Fumagalli5, A Pesenti5,
M Romero6, G Tognoni6, R Latini2, L Gattinoni1
1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Università
degli Studi di Milano, Milan, Italy; 2Istituto di Ricerche Farmacologiche Mario
Negri, Milan, Italy; 3Diagnostic Engineering & Research GmbH, Heildelberg,
Germany; 4Mitsubishi Chemical Europe GmbH, Munich, Germany; 5Ospedale
San Gerardo, Monza, Italy; 6Consorzio Mario Negri Sud, Santa Maria Imbaro,
Italy
Critical Care 2013, 17(Suppl 2):P35 (doi: 10.1186/cc11973) ,
g
,
,
1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Università
degli Studi di Milano, Milan, Italy; 2Istituto di Ricerche Farmacologiche Mario
Negri, Milan, Italy; 3Diagnostic Engineering & Research GmbH, Heildelberg,
Germany; 4Mitsubishi Chemical Europe GmbH, Munich, Germany; 5Ospedale
San Gerardo, Monza, Italy; 6Consorzio Mario Negri Sud, Santa Maria Imbaro,
Italy Results For the non-AKI patients, the median of presepsin in patients
with nonsepsis (n = 78) and the sepsis group (n = 37) were 406 pg/ml
(range: 86 to 4,374) and 1,065 pg/ml (range 86 to 9,960), respectively
(P <0.0001). For the Risk patients, the median of presepsin in patients
with nonsepsis (n = 25) and the sepsis group (n = 27) were 299 pg/ml
(range: 71.2 to 3,361) and 831 pg/ml (range: 233 to 16,759), respectively
(P <0.01). For the Injury patients, the median of presepsin in patients
with nonsepsis (n = 12) and the sepsis group (n = 27) were 463 pg/ml
(range: 122 to 1,197) and 1,451 pg/ml (range: 237 to 4,200), respectively
(P <0.001). For the Failure patients, the median of presepsin in patients y
Critical Care 2013, 17(Suppl 2):P35 (doi: 10.1186/cc11973) Introduction Sepsis results from complex interactions between
infecting microorganisms and host responses, often leading to
multiple organ failures and death. Over the years, its treatment has Introduction Sepsis results from complex interactions between
infecting microorganisms and host responses, often leading to
multiple organ failures and death. Over the years, its treatment has S14 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P39 with nonsepsis (n = 14) and the sepsis group (n = 27) were 1,607 pg/ml
(range: 454 to 8,516) and 1,523 pg/ml (range: 293 to 16,764), respectively
(P = 0.175). The diagnostic accuracy of presepsin in patients with sepsis
was determined by ROC analysis, the area under the curve was 0.789
for the non-AKI patient group, 0.735 for the Risk patient group, 0.855 for
the Injury patient group and 0.593 for the Failure patient group. Conclusion In Failure and more progressed AKI patients, the diagnostic
accuracy of the presepsin level was lower than the other groups. References with nonsepsis (n = 14) and the sepsis group (n = 27) were 1,607 pg/ml
(range: 454 to 8,516) and 1,523 pg/ml (range: 293 to 16,764), respectively
(P = 0.175). The diagnostic accuracy of presepsin in patients with sepsis
was determined by ROC analysis, the area under the curve was 0.789
for the non-AKI patient group, 0.735 for the Risk patient group, 0.855 for
the Injury patient group and 0.593 for the Failure patient group. Simple and fast prediction of Legionella sp. in community-acquired
pneumonia: validation of a prediction rule
S Haubitz1, F Hitz2, L Graedel2, T Wiemken3, P Peyrani Dicastelnuovo3,
J Ramirez3, M Batschwaroff 1, C Fux1, B Mueller1, P Schütz1
1Kantonsspital Aarau, Switzerland; 2University of Basel, Switzerland;
3University of Louisville School of Medicine, Louisville, KY, USA
Critical Care 2013, 17(Suppl 2):P39 (doi: 10.1186/cc11977) Conclusion In Failure and more progressed AKI patients, the diagnostic
accuracy of the presepsin level was lower than the other groups. References 1. Endo S, et al.: J Infect Chemother 2012, 18:891-897. doi:10.1007/s10156-012-0435-2
2. Bellomo R, et al.: Crit Care 2004, 8:R204-R212. doi:10.1186/cc2872 1. Endo S, et al.: J Infect Chemother 2012, 18:891-897. doi:10.1007/s10156-012-0435-2 Introduction Ruling out Legionella sp. in patients presenting with
community-acquired pneumonia (CAP) is important due to diff erences
in treatment regimens. Yet antigen tests as well as blood cultures have
low sensitivity and an important time delay, making empirical broad
spectrum coverage necessary particularly in severe cases. Fiumefreddo
and colleagues recently proposed a clinical score based on six clinical
and laboratory variables (fever, cough, sodium, lactate-dehydrogenase,
C-reactive protein, platelet count) which allowed assessing the
likelihood of Legionella [1]. Yet these variables need validation in
an independent patient cohort before implementation into clinical
routine. 2. Bellomo R, et al.: Crit Care 2004, 8:R204-R212. doi:10.1186/cc2872 2.
Bellomo R, et al.: Crit Care 2004, 8:R204-R212. doi:10.1186/cc2872 Eff ectiveness of early ureteric stenting for urosepsis associated with
urinary tract calculi S Nishiguchi , Y Tokuda
1Shonan Kamakura General Hospital, Kamakura, Japan; 2Mito Kyodo General
Hospital, Mito, Japan
Critical Care 2013, 17(Suppl 2):P43 (doi: 10.1186/cc11981) S Nishiguchi , Y Tokuda
1Shonan Kamakura General Hospital, Kamakura, Japan; 2Mito Kyodo General
Hospital, Mito, Japan
Critical Care 2013, 17(Suppl 2):P43 (doi: 10.1186/cc11981) p
Results A total of 536 (14%) of the 3,766 enrolled patients had SCAP. They were mostly male (66%) with median age 59 (29 to 82) years,
median SAPS II 44 (21 to 80) and total SOFA score 8 (3 to 16). Thirty-
seven per cent of the cases were microbiologically documented (St. pneumoniae – 24%; Enterobacteriaceae – 20%; infl uenza A (H1N1)
virus – 18%) and 45% had septic shock. Antibiotic combination was
used in 76% of the patients and 61% received a macrolide. Median
hospital length of stay was 19 (3 to 70) days and hospital mortality
was 35%. Comorbidities were present in 70% of the patients. The most
frequent were: diabetes mellitus (21%), chronic respiratory failure
(18%) and alcoholism (15%). Median Charlson’s comorbidity index
(CCI) was 4 (0 to 13). In univariate analysis, the presence of at least one
comorbidity (odds ratio (OR) 2.29; 95% CI 1.49 to 3.52), namely cancer
(OR 3.80; 95% CI 2.14 to 6.74; P <0.001), chronic renal failure (OR 3.23;
95% CI 1.53 to 6.82; P = 0.001), immunosuppression (OR 2.12; 95% CI
1.15 to 3.92; P = 0.014) and neurologic disease (OR 1.87; 95% CI 1.10 to
3.17; P = 0.02), increased the chances of dying in the hospital. Median
CCI was also signifi cantly higher in nonsurvivors (5 vs. 3; P <0.001; OR
per point 1.10 (95% CI: 1.05 to 1.15)). The only independent risk factor
for hospital mortality was the presence of at least one comorbidity (OR
2.09; 95% CI 1.13 to 3.85). Introduction Urosepsis associated with urinary tract calculi is a critical
disease. Patients with this condition occasionally require drainage,
mostly ureteric stent placement and these patients need longer
hospitalization. However, indications for timely ureteric stenting for
urosepsis associated with urinary tract calculi have not been clearly
determined. The objective of the study was to evaluate whether earlier
stent placement might lead to shorter length of stay (LOS) in hospitals. Outcome of severe community-acquired pneumonia: the impact of
comorbidities Outcome of severe community-acquired pneumonia: the impact of
comorbidities JM Pereira1, JA Paiva1, F Froes2, JP Baptista3, J Gonçalves-Pereira4
1Centro Hospitalar S. João, Porto, Portugal; 2Hospital Santa Maria-CHLN,
Lisboa, Portugal; 3Hospitais da Universidade de Coimbra – CHUC, Coimbra,
Portugal; 4Hospital São Francisco Xavier, Lisboa, Portugal
Critical Care 2013, 17(Suppl 2):P41 (doi: 10.1186/cc11979) Conclusion In early surgery IE, it appears to be associated with mortality:
age >70 years, previous heart disease, emergency surgery, antibiotics
within 7 days before surgery, preoperative MOF, and high scores on
scales of Pons and Ontario. The causal agent and echocardiography
have no relation with worse prognosis. The general syndrome debut
and the Streptococcus spp. etiology seem to have lower mortality. IE
has pathologies with high .preoperative severity scores but these are
not suffi cient to guide therapeutic decisions. Introduction Several comorbidities have been independently asso-
ciated with both predisposition to community-acquired pneumonia
and a worse outcome. The goal of this study was to evaluate the impact
of comorbidities on the outcome of patients with severe community-
acquired pneumonia (SCAP). Methods A prospective, multicentre, observational cohort study of
all patients with SCAP consecutively admitted to 14 Portuguese ICUs
during a 12-month period. Several comorbidities were evaluated:
congestive heart failure, cancer, chronic renal failure, chronic respira-
tory failure, chronic hepatic disease, alcoholism, diabetes mellitus,
neurologic disease, immunosuppression, HIV infection. To evaluate the
impact of comorbidities associated with hospital mortality in univariate
analysis, a logistic regression analysis adjusted to other variables
(clinical relevant or statistically signifi cant in univariate analysis) was
performed. P42 P42 Prognostic scores and infective endocarditis
P Fernandez Ugidos1, R Gomez Lopez1, P Vidal Cortes1,
A Ceniceros Barros1, L Seoane Quiroga1, JM Lopez Perez2
1Complejo Hospitalario Universitario Ourense, Spain; 2Complejo Hospitalario
Universitario A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P42 (doi: 10.1186/cc11980) Introduction Infective endocarditis (IE) is a high-mortality disease,
especially in the early surgery subgroup. The aim of this study was try
to identify prognostic factors of IE that will require surgery during the
same admission of their diagnosis, including evaluation of surgical
severity scores. Conclusion We have provided the fi rst experimental evidence for the
associations of genes coding detoxifi cation enzymes with the risk of
CAP. Our results also demonstrate that predisposition to CAP is strongly
attributed to the eff ects of a number of genes with low penetrance
and therefore imply that inter-locus interactions may be regarded as
an important component of polygenic and multifactorial factors of
susceptibility to CAP. Methods A retrospective study (5 years) of all patients admitted to
a tertiary hospital in northern Spain with diagnosis of IE (modifi ed
Duke criteria) who required early surgery. Demographic, clinical and
microbiology data were collected. Chi-square and Student t tests. Signifi cance: P <0.05. SPSS17. gi
Results We had 73 patients, 79.5% men, age 65 years. Eighty-two
percent had positive blood cultures. Forty-one percent of cases
required previous ICU admission. Surgery was urgent in 35%. Fifty-six
per cent of patients had postoperative shock and 58% suff ered post-
surgery ARF. Hospital mortality was 31.5%. Regarding prognostic scales:
the mean EuroSCORE was 11.38 ± 3.93 points. No patient was placed in
the low-risk group. Ninety-four per cent of cases were at high risk. The
Parsonnet mean score was 27.7 ± 11.6. The mean Beth Israel Medical
Center was 37.6 ± 10 points. The mean Ontario scale was 7.09 ± 2.7. The
mean Surgical risk scale of Roques was 6.69. And the mean by Pons
Scale was 24.02. Dead patients are older, with previous heart disease,
require urgent surgery and have previous ICU stay; usually they have
MOF and they had received less than 7 days of antibiotic treatment. The
etiology did not worsen the prognosis. Outcome of severe community-acquired pneumonia: the impact of
comorbidities
JM Pereira1, JA Paiva1, F Froes2, JP Baptista3, J Gonçalves-Pereira4
1Centro Hospitalar S. P42 João, Porto, Portugal; 2Hospital Santa Maria-CHLN,
Lisboa, Portugal; 3Hospitais da Universidade de Coimbra – CHUC, Coimbra,
Portugal; 4Hospital São Francisco Xavier, Lisboa, Portugal
Critical Care 2013, 17(Suppl 2):P41 (doi: 10.1186/cc11979) Host genetic variants associated with community-acquired
pneumonia Using allele-specifi c
tetraprimer PCR, all subjects were genotyped for 13 polymorphic
variants in the genes of xenobiotic detoxifi cation CYP1A1 (rs2606345,
rs4646903, rs1048943), GSTM1 (Ins/Del), GSTT1 (Ins/Del), ABCB1
(rs1045642); immune and infl ammation response IL-6 (rs1800795),
TNFα (rs1800629), MBL2 (rs7096206), CCR5 (rs333), NOS3 (rs1799983),
angiotensin-converting enzyme (ACE; rs4340), and occlusive vascular
disease/hyperhomocysteinemia MTHFR (rs1801133). Conclusion Our fi ndings suggested that NOx may be involved in the
pathogenesis of ALI/ARDS. S15 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Results Seven genes CYP1A1 rs2606345T/T, GSTM1 Ins/*, ABCB1
C/C, IL-6 C/C-G/G, NOS3 T/T-G/G, CCR5 Ins/Ins, and ACE Del/Del were
associated with CAP. The highest eff ect was detected for the CYP1A1
rs2606345: in comparison with the control A, P = 3.9×10–5, OR =
2.40, 95% CI: 1.59 to 3.64; and in comparison with the control B, P =
1.4×10–5, OR = 2.0, 95% CI: 1.46 to 2.74. For the two genes CYP1A1 and
GSTM1, associations remained signifi cant after correction for multiple
comparisons. Multiple analysis by the number of all risk genotypes
showed a highly signifi cant association with CAP (P = 2.4×10–7, OR =
3.03, 95% CI 1.98 to 4.64) with the threshold for three risk genotypes. Using the ROC analysis, the AUC value for multi-locus model was
estimated as 68.38, which is rather high for genetic markers.i Yamamoto Y, et al.: BMC Urol 2012, 12:4.
Yoshimura K, et al.: J Urol 2005, 173:458-462. Yamamoto Y, et al.: BMC Urol 2012, 12:4.
Yoshim ra K et al J Urol 2005 173 458 462 2.
Yoshimura K, et al.: J Urol 2005, 173:458-462. P46 Abdominal infection plays a role in the incidence of ventilator-
associated pneumonia
GB Bouroche1, S Ruckly2, B Misset3, JF Timsit4, F Philippart3
1Institut Gustave Roussy, Villejuif, France; 2Institut Albert Bonniot, La Tronche,
France; 3Hôpital Saint Joseph, Paris, France; 4CHU Grenoble, France
Critical Care 2013, 17(Suppl 2):P46 (doi: 10.1186/cc11984) g
References 1. Clec’h C, Schwebel C, Français A: Does catheter-associated urinary tract
infection increase mortality in critically ill patients. Infect Control Hosp
Epidemiol 2007, 28:1367-1373. 1. Clec’h C, Schwebel C, Français A: Does catheter-associated urinary tract
infection increase mortality in critically ill patients. Infect Control Hosp
Epidemiol 2007, 28:1367-1373. p
Methods To investigate, in humans, the potential involvement of
previous intra-abdominal infection (IIA) in preventing or promoting
VAP, we realized a prospective observational study using data from a
multicenter database (OUTCOMEREA), including all patients admitted 2. Marx G, Reinhart K: Urosepsis: from the intensive care viewpoint. Int J
Antimicrob Agents 2008, 31S:S79-S84. 2. Marx G, Reinhart K: Urosepsis: from the intensive care viewpoint. Int J
Antimicrob Agents 2008, 31S:S79-S84. 2. Marx G, Reinhart K: Urosepsis: from the intensive care viewpoint. Int J
Antimicrob Agents 2008, 31S:S79-S84. Figure 1 (abstract P46). Cumulative incidence of VAP. Eff ectiveness of early ureteric stenting for urosepsis associated with
urinary tract calculi Methods For patients who required ureteric stent procedures for
urosepsis associated with urinary tract calculi in our hospital from
July 2008 to July 2012, we compared the LOS in our hospital between
patients with earlier stenting and those without it. A linear regression
model was used for multivariable-adjusted comparison. j
Results In a total of 30 patients (mean age, 72; 13 males), the mean
days from emergency room admission to ureteric stenting was 3.5 days
(range, 1 to 14 days) and the overall mean LOS was 36 days (range,
8 to 102 days). The early-stenting group (mean LOS, 21 days) had
signifi cantly shorter LOS than the delayed-stenting group (mean LOS,
50 days) with adjusted β coeffi cient of –26 days (95% CI, –46, –6). Conclusion In patients with urosepsis associated with urinary tract calculi,
earlier stenting within 2 days of admission may reduce the hospital LOS. References Conclusion In SCAP, the presence of at least one comorbidity doubles
the chances of dying in the hospital and is an independent risk factor
for hospital mortality. Yamamoto Y, et al.: BMC Urol 2012, 12:4. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S16 (n = 13, 13.4%) and intraabdominal infections (n = 13, 13.4%). Patients
with pneumonia were treated according to knowledge in a specifi c
moment, thus diff erent antibiotics were used. The duration of antibiotic
therapy was 20 ± 15.5 days. In nine episodes of pneumonia according
to the CDC no germ was isolated in the cultures. Six of the episodes
were polymicrobial infections. The most frequent microbes isolated
were E. coli (n = 7, 22.5% of pneumonia cases), A. fumigatus (n = 7,
22.5%), S. aureus (n = 3, 9.68%), P. aeruginosa (n = 3, 9.68%), P. mirabilis,
K. pneumoniae, E. cloacae, E. faecalis, C. glabrata, and S. marcescens (one
case each, 3.22%). Pneumonia was suspected but not confi rmed in 75
patients. Despite this, antibiotic treatment was maintained for a media
of 17.35 ± 7.01 days: 56 wide-spectrum treatments and 18 targeted
therapy after knowing the antibiogram. The length of ICU stay was
38.4 ± 70.8 (3 to 264) days, of hospital stay was 66.2 ± 80.5 (3 to 304)
days and of mechanical ventilation was 27.3 ± 50.2 (3 to 264) days. The
mortality of patients with pneumonia was 32.3%. Abdominal infection plays a role in the incidence of ventilator-
associated pneumonia GB Bouroche1, S Ruckly2, B Misset3, JF Timsit4, F Philippart3
1Institut Gustave Roussy, Villejuif, France; 2Institut Albert Bonniot, La Tronche,
France; 3Hôpital Saint Joseph, Paris, France; 4CHU Grenoble, France
Critical Care 2013, 17(Suppl 2):P46 (doi: 10.1186/cc11984) Introduction Despite many therapeutic interventions, ventilator-
acquired pneumonias (VAP) are frequent in the ICU and are associated
with major morbidity and mortality. Sepsis causes a time-dependent
modifi cation of the infl ammatory response. This reprogramming
could promote the occurrence of a secondary infection and worsen
the prognosis. In animals, peritonitis is associated with an alteration
of pulmonary immunity and an increasing mortality from secondary
pneumonia. y
Conclusion Urinary sepsis mortality is associated with late-onset and/
or inappropriate antibiotic use, as well as with immunodepression and
advanced age. Introduction Analysis of mortality-related factors in urinary sepsis
patients. Methods A retrospective descriptive study of urologic sepsis patients
in the ICU from 2008 to 2010. Clinical, epidemiological and outcome
variables were analysed. Quantitative variables are expressed as
either mean and standard deviation or as median and interquartile
range for asymmetric variables. Qualitative variables are expressed as
percentages and absolute values. Mann–Whitney’s U test and Fisher’s
exact test were applied (α error was 5% in both cases), as well as binary
logistic regression for multivariate analysis. Results There was a total number of 44 patients (aged 59.39 ± 17.71;
63.8% females). APACHE II score upon admission was 18 ± 6. Out of
these patients, 27.3% showed no underlying disorder and 18.2% (no =
8) showed chronic renal failure; 25% were immunodepressed patients;
31% underwent urinary instrumentation in the previous 15 days, yet
only three of them had undergone permanent urine catheterization. Observed mortality was 25%, while sepsis-related mortality was 22.7%. The patients who died were, on average, older that those who survived
(67.9 ±7 10.2 vs. 56.8 ± 18.7; P = 0.02). Besides, the former also reported
greater delay in turning to the hospital after symptom onset (13.4 ± 6.6
vs. 6.2 ± 4.7 days; P = 0.0001). Immunodepressed patients presented
higher mortality rate: OR 8.7 (95% CI 1.7 to 42.3), as well as those who
underwent inappropriate initial antibiotic treatment: OR 10.8 (95%
CI 2.1 to 54.7). No relation was observed between germ typology or
resistance to β-lactam antibiotics and mortality. After adjustment of
mortality due to APACHE II score upon admission, delay in the onset
of appropriate antibiotic treatment was an independent predictor of
mortality in our patients: OR 1.2, 95% CI (1.02 to 1.42). Conclusion Nosocomial pneumonia is the most frequent infection
in our series. Despite when infection was not confi rmed, antibiotic
therapy was maintained in suspect cases. We found a high incidence of
aspergillosis. Limitations because of wide duration of this study should
be considered. P46 P44
Outcomes in urinary sepsis Outcomes in urinary sepsis
C Joya-Montosa, E Trujillo-Garcia, MJ Delgado-Amaya, E Curiel-Balsera
Hospital Regional Carlos Haya, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P44 (doi: 10.1186/cc11982) Outcomes in urinary sepsis y
p
C Joya-Montosa, E Trujillo-Garcia, MJ Delgado-Amaya, E Curiel-Balsera
Hospital Regional Carlos Haya, Málaga, Spain
Critical Care 2013 17(Suppl 2):P44 (doi: 10 1186/cc11982) Introduction Analysis of mortality-related factors in urinary sepsis
patients. Nosocomial pneumonia in the postoperative period after heart
transplantation transplantation
R Gómez López1, P Fernández Ugidos1, P Vidal Cortes1, M Bouza Vieiro2,
J Miñiz2, S Fojon Polanco2, M Paniagua Martin2, R Marzoa Rivas2,
E Barge Caballero2, M Crespo Leiro2
1Complexo Hospitalario Universitario de Ourense, Spain; 2Complexo
Hospitalaro Universitario de A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P45 (doi: 10.1186/cc11983) p
R Gómez López1, P Fernández Ugidos1, P Vidal Cortes1, M Bouza Vieiro2,
J Miñiz2, S Fojon Polanco2, M Paniagua Martin2, R Marzoa Rivas2,
E Barge Caballero2, M Crespo Leiro2
1Complexo Hospitalario Universitario de Ourense, Spain; 2Complexo
Hospitalaro Universitario de A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P45 (doi: 10.1186/cc11983) Hospitalaro Universitario de A Coruña, Spain p
p
Critical Care 2013, 17(Suppl 2):P45 (doi: 10.1186/cc11983) Introduction Infections are a major complication during the
postoperative period after heart transplantation (HT). In our hospital,
nosocomial pneumonia is the most frequent infection in this period. The objective of this study is to determine the epidemiological and
microbiological characteristics of this disease in our centre. Methods A descriptive retrospective study of all medical records of HT
performed in a single institution from 1991 to 2009 followed until June
2010. Clinical and microbiological variables were considered. Centre
for Diseases Control (CDC) criteria were used to defi ne nosocomial
infections. Invasive aspergillosis was considered if there were criteria
for probable aspergillosis according to IDSA criteria. Results In 594 HTs there were 97 infectious episodes in 75 patients
(12.6%). Eighty-fi ve patients (14.3%) died during hospitalization. Infection is the second cause of mortality during the postoperative
period (17.9% of dead patients). The most common locations of
infections were pneumonia (n = 31, 31.9% of infection episodes),
bloodstream (n = 24, 24.7%), urinary tract (n = 14, 14.4%), surgical site Figure 1 (abstract P46). Cumulative incidence of VAP. S17 Critical Care 2013, Volume 17 Suppl 2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 2 (abstract P46). Cumulative occurrence of death. that numbers of CVC, intubation and surgery, the use of muscle
relaxant and steroid were independent risk factors for developing VAP. Ventilator days and ICU length of stay were longer in the VAP group
(25 vs. 6 and 25 vs. 7 days, respectively). Lastly, the hospital mortality
rate was signifi cantly higher in the VAP group (33% vs. 12%, P = 0.008). Nosocomial pneumonia in the postoperative period after heart
transplantation Conclusion The incidence of VAP was 9.2% in the SICU of Siriraj Hospital,
which was comparable with previous reports. Bundles of care to
prevent VAP should include weaning from a ventilator. Muscle relaxant
and steroid should be administered according to strong indication. Meticulous care of the airway should be implemented as protocol in
order to prevent complications that can result in the development of
VAP. Hospital-acquired bloodstream infection: Indian perspective g
,
FEHI, New Delhi, India FEHI, New Delhi, India
Critical Care 2013, 17(Suppl 2):P48 (doi: 10.1186/cc11986) Figure 2 (abstract P46). Cumulative occurrence of death. Introduction This is a 1-year prospective study to determine the
incidence, source and etiology of hospital-acquired bloodstream
infection (HABSI) in the Indian context. The resistance pattern was also
reviewed. Introduction This is a 1-year prospective study to determine the
incidence, source and etiology of hospital-acquired bloodstream
infection (HABSI) in the Indian context. The resistance pattern was also
reviewed. to the ICU for severe sepsis or septic shock who required mechanical
ventilation for at least 72 hours. Methods A single-centre prospective study in a 35-bed ICU. HABSI
was defi ned according to current CDC guidelines. HCAP, catheter-
associated UTI (CAUTI) and skin-related infections causing BSI was also
defi ned according to recent guidelines and analysed. Results In total, 2,623 patients were included, of which 290 had an
IIA. A total of 862 patients (33%) developed a VAP, 56 (19%) in the IIA
group and 806 (34%) in the non-IIA group (P <0.01). VAP, after sepsis,
occurred less frequently and later in patients with IIA. The occurrence of
IIA, in comparison with another sepsis, is a protective factor against VAP
(HR = 0.643 (0.478 to 0.863), P = 0.003). There is, however, no signifi cant
diff erence between the groups in terms of ICU mortality (28% vs. 32%,
P = 0.16). See Figures 1 and 2. i
g
g
y
Results Out of 332 positive samples, 90 samples (n = 45) were HABSI. The microbiological analysis showed 60% were Gram-negative, 6%
were candida and 27% were Gram-positive. The commonest isolate
was klebsiella and MRSA was commonest in Gram-positive. The source
of HABSI showed CRBSI was the commonest cause at 69%, which
correlates with international data. Ventilator-associated pneumonia
and CAUTI caused 9.5% BSI respectively. The resistance pattern among
Gram-negative bacteria showed multidrug-resistant (MDR) and
extreme drug-resistant (XDR) isolates were highest. See Tables 1 and 2. g
Conclusion In this study, the presence of an abdominal sepsis, in a
context of severe sepsis or septic shock, was associated with a lower
incidence of later VAP. These results have to be confi rmed in other
studies, especially prospective. They open interesting new research
directions. Table 1 (abstract P48). Source
Source
Total (%)
CRBSI
69
HCAP
9.5
CAUTI
9.5
Devices
7
Skin
5
Table 2 (abstract P48). Reference 1. Werarak P, Kiratisin P, Thamlikitkul V: Hospital acquired pneumonia and
ventilator associated pneumonia in adults at Siriraj hospital, etiology,
clinical outcomes and impact of antimicrobial resistance. J Med Assoc Thai
2010, 93(Suppl 1):126-138. Hospital-acquired bloodstream infection: Indian perspective Resistance pattern
Resistance
Total (%)
Non-MDR
15
MDR
44
XDR
33
PDR
8
Conclusion The incidence of HABSI is 27%. Of this, CRBSI cause 70%
and Gram-negative bacteria were commonest with high resistance. This is in contrast to western data where Gram-positive infections are
common. Our study highlights need for stringent guidelines for CRBSI
prevention. References
1. Richard et al.: Crit Care Med 1999, 27:887-892. 2. Valles et al. : J Infect 2008, 56:27-34. P50 Analysis of risk factors for catheter-related bloodstream infection in
a parenteral nutrition population P52 Results for introduction of a new hand hygiene program in the ICU
P Vos, A Harts-Laurijsen, R Snoeren, R Ramnarain, JA Van Oers
St Elisabeth Hospital, Tilburg, the Netherlands
Critical Care 2013, 17(Suppl 2):P52 (doi: 10.1186/cc11990) Analysis of risk factors for catheter-related bloodstream infection in
a parenteral nutrition population I Conrick-Martin1, M McGovern2, K Boner1, J Bourke1, E Fitzgerald1, R Hone1,
M Lynch1, D Phelan1, C Walshe3 I Conrick-Martin1, M McGovern2, K Boner1, J Bourke1, E Fitzgerald1, R Hone1,
M Lynch1, D Phelan1, C Walshe3 1Mater Misericordiae University Hospital, Dublin, Ireland; 2Harvard School of
Public Health, Cambridge, MA, USA; 3Beaumont Hospital, Dublin, Ireland
Critical Care 2013, 17(Suppl 2):P50 (doi: 10.1186/cc11988) 1Mater Misericordiae University Hospital, Dublin, Ireland; 2Harvard School of
Public Health, Cambridge, MA, USA; 3Beaumont Hospital, Dublin, Ireland
Critical Care 2013, 17(Suppl 2):P50 (doi: 10.1186/cc11988) 1Mater Misericordiae University Hospital, Dublin, Ireland; 2Harvard School of
Public Health, Cambridge, MA, USA; 3Beaumont Hospital, Dublin, Ireland
Critical Care 2013, 17(Suppl 2):P50 (doi: 10.1186/cc11988) Conclusion A low proinfl ammatory response is a key facilitating factor
for the development of infection. Measuring serum TNFα levels before
and after operations can thus predict the outcome. Introduction Catheter-related bloodstream infection (CRBSI) is a
complication of central venous catheters (CVCs) with an attributable
morbidity, mortality and cost [1]. We examined patient risk factors for
CRBSI in an adult parenteral nutrition (PN) population. Incidence and risk factors for ventilator-associated pneumonia in
Siriraj Hospital O Chaiwat, S Nakaviroj
Siriraj Hospital Mahidol University, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P47 (doi: 10.1186/cc11985) Introduction Ventilator-associated pneumonia (VAP) is a serious illness
with substantial morbidity and mortality and increases the cost of
hospital care. Even when bundles of care to prevent VAP have been
implemented, the incidence of VAP was not dramatically improved. This study aims to determine the incidence and risk factors of VAP in
the SICU of Siriraj Hospital. Methods During a 1-year period, 228 patients admitted to the SICU
were enrolled. All patients required ventilatory support longer than
48 hours. Data were collected by reviewing patient medical records
including demographic data, onset of VAP, type of organisms,
medication used, number of central venous catheters (CVC) used and
blood transfusion. VAP outcomes were also reported. Results VAP occurred in 21 patients (9.21%) or 8.21 per 1,000 ventilator-
days. The onset of VAP was late in the majority of patients. The most
common organism was A. baumannii (66%), followed by P. aeuruginosa
(19%). Compared with non-VAP groups, patients in the VAP group had
higher APACHE II score (18 vs. 13, P <0.001), blood transfusion (95% vs. 75%, P = 0.04), numbers of CVC used (3 vs. 1, P <0.001), muscle relaxant
used (43% vs. 3%, P <0.001) and steroid used (33% vs. 4%, P <0.001). The VAP group also had a signifi cantly higher number of intubation,
reintubation and self-extubation. Multiple logistic regression showed Conclusion The incidence of HABSI is 27%. Of this, CRBSI cause 70%
and Gram-negative bacteria were commonest with high resistance. This is in contrast to western data where Gram-positive infections are
common. Our study highlights need for stringent guidelines for CRBSI
prevention. S18 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P49
Risk factors for catheter-related bloodstream infections in a surgical
ICU
A Kundakci, O Ozkalayci, P Zeyneloglu, H Arslan, A Pirat
Baskent University Hospital, Ankara, Turkey
Critical Care 2013, 17(Suppl 2):P49 (doi: 10.1186/cc11987) P49
Risk factors for catheter-related bloodstream infections in a surgical
ICU
A Kundakci, O Ozkalayci, P Zeyneloglu, H Arslan, A Pirat
Baskent University Hospital, Ankara, Turkey
Critical Care 2013, 17(Suppl 2):P49 (doi: 10.1186/cc11987) Results The study population was 1,961 patients in whom 3,213 CVCs
were utilised over 19,511 CVC days. There were 256 CRBSI episodes in
216 patients. Median (IQR) patient age was 62 (23), and 58% were male. The incidence of CRBSI decreased signifi cantly (P <0.001) during the
study period from 16% of patients in the period 1997 to 2003 to 7%
in 2004 to 2010. The corresponding rate of CRBSI infection (per 1,000
CVC days) decreased from 18 to 10. There was a signifi cant decrease
(P <0.001) in numbers of CVCs inserted per patient (from 1.87 to
1.49). Each extra CVC PN day was associated with an increased risk of
developing CRBSI of 3.4% (OR 1.034, P <0.05). The number of PN CVCs
was associated with developing CRBSI (OR 1.218, P <0.10). Patient
factors signifi cantly associated with CRBSI included perioperative PN
use (compared with medical patients) (OR 2.414, P <0.01), and male sex
(OR 1.952, P <0.01). Introduction Preventing catheter-related bloodstream infections
(CR-BSI) can reduce the duration of hospital stay, healthcare costs, and
mortality rates. Identifying the risk factors and correction of modifi able
factors should be one of the main objectives of infection control
measures. The aim of this study is to determine the risk factors of CR-BSI
in a cohort of surgical ICU patients admitted to Baskent University
Hospital. p
Methods Following Institutional Review Board approval, data for 876
patients admitted to the surgical ICU between January 2009 and July
2012 were reviewed retrospectively. After completing the review, 25
patients diagnosed with CR-BSI were compared with 66 appropriate
matches who did not have CR-BSI. Demographical features, underlying
diseases, APACHE II (Acute Physiology and Chronic Health Evaluation)
and SOFA (Sequential Organ Failure Assessment) scores, length of stay,
organ dysfunctions, laboratory values, use of vasopressors, mechanical
ventilation, nutrition, antibiotics, transfusions, and features related to
central venous catheterization were recorded. Patients who did not
have a central venous catheter and were discharged within 2 days of
ICU stay were excluded. Conclusion This prospective study demonstrated that perioperative
PN use was associated with increased risk of CRBSI. The association
between CRBSI and CVC PN days is consistent with the theory
suggesting benefi t to limiting CVC duration and changing from PN to
enteral nutrition as soon as appropriate. Can we predict the postoperative infections?
2
3
2 O’Grady NP: Guidelines for the prevention of intravascular catheter-related
infections. Clin Infect Dis 2011, 52:e162-e193. 2. Bouza E: Catheter-related infections: diagnosis and intravascular
treatment Clin Microbiol Infect 2002 8 265 274 2. Bouza E: Catheter-related infections: diagnosis and intravascular
treatment. Clin Microbiol Infect 2002, 8:265-274. Results From 22 patients, eight deep wound and urine infections
were found in 14 days and six urine and wound infections in 30 days
after surgery, all survived. All patients were bacterially contaminated,
as wound samples taken at the end of operation demonstrated. On
day 0 the circulating TNFα values were lower in infected patients. TNF started to increase from day 3 to day 9, never reaching values of
the uneventful healing group. Soluble TNF receptor I, IL-6, ACTH, and
cortisol concentrations did not demonstrate any diff erence on day 0
but from day 1 started to increase transiently, reaching higher levels in
septic patients.l Can we predict the postoperative infections?
2
3
2 y
Results Out of 91 patients included in the fi nal analysis, 25 (27%)
patients had CR-BSI. When compared with patients who did not have
CR-BSI, those who did were older (P = 0.029), required more blood
product transfusions during the fi rst 3 days of ICU (P = 0.016), had a
longer duration of catheter stay (P = 0.019), and were more frequently
catheterized via the internal jugular vein (IJV) (P = 0.022). A logistic
regression model revealed that advanced age (OR: 1.037; 95% CI: 1.001
to 1.073; P = 0.042) was an independent risk factor for CR-BSI. Fourteen-
day and 28-day mortality rates for CR-BSI were 12% (P = 0.749) and 28%
(P = 0.406), respectively. p
p
p
Mózes1, K Gornicsar2, A Grosz3, ZS Domján2, I Buzogány2 p
p
p
T Mózes1, K Gornicsar2, A Grosz3, ZS Domján2, I Buzogány2
1Ministry of Defense Health Centre, Budapest, Hungary; 2Péterfy Hospital,
Budapest, Hungary; 3University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P51 (doi: 10.1186/cc11989) T Mózes1, K Gornicsar2, A Grosz3, ZS Domján2, I Buzogány2
1Ministry of Defense Health Centre, Budapest, Hungary; 2Péterfy Hospital,
Budapest, Hungary; 3University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P51 (doi: 10.1186/cc11989) Introduction Many patients develop infections following operations. Decreased immune competence has been demonstrated in
acute neurological conditions. A strong cytokine-mediated anti-
infl ammatory response was observed in stroke patients at infection,
although infection due to the decreased proinfl ammatory mediators
can be expected as well. To investigate this question the following
experiment was performed. y
Conclusion Although age, blood product transfusion, duration of
catheter stay, and use of IJV were diff erent between patients who did
and did not have CRBSI, advanced age was the only independent risk
factor for CR-BSI. Early suspicion of CR-BSI by the other well-known risk
factors has a substantial eff ect on the treatment of CR-BSI. References Methods Twenty-two urinary bladder cancer patients with radical
cystectomy and lymphadenectomy were studied. Blood samples were
taken on day 0 (before) and days 1, 3, 6, 9 and 14 after operation as
well as on days 30, 60, 90 and 270 during follow-up. TNFα, soluble TNFα
receptor I and IL-6 levels in sera were determined by HS ELISA and/or
ELISA. Plasma ACTH and cortisol values were measured by RIA kits. 1. O’Grady NP: Guidelines for the prevention of intravascular catheter-related
infections. Clin Infect Dis 2011, 52:e162-e193. 1. Reference 1. Blot SI, et al.: Clinical and economic outcomes in critically ill patients with
nosocomial catheter-related bloodstream infection. Clin Infect Dis 2005;
41:1591-1598. Results for introduction of a new hand hygiene program in the ICU
P Vos, A Harts-Laurijsen, R Snoeren, R Ramnarain, JA Van Oers
St Elisabeth Hospital, Tilburg, the Netherlands
Critical Care 2013, 17(Suppl 2):P52 (doi: 10.1186/cc11990) Methods The study was carried out in a 525-bed tertiary-referral
teaching hospital over a 14-year study period (1997 to 2010). All in-
patients referred for PN via CVCs were included. Prospectively collected
data were recorded in a specifi c PN record. The CRBSI audit group met
quarterly to review all sepsis episodes, assigning a diagnostic category
(CRBSI or non-CRBSI). Patient risk factors for development of CRBSI
were examined using a logistic regression model to take account of
the dichotomous nature of the outcome. Odds ratios from a model
incorporating demographic and clinical data were tested for statistical
signifi cance. Introduction Although hand hygiene is known as a core element
in prevention of healthcare-associated infections, compliance to
its program is not high. With a new campaign we tried to enlarge
awareness on hand hygiene. Introduction Although hand hygiene is known as a core element
in prevention of healthcare-associated infections, compliance to
its program is not high. With a new campaign we tried to enlarge
awareness on hand hygiene. Methods We performed an experimental, before and after, study design
on our 30-bed, mixed medical–surgical ICU. We conducted a baseline S19 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 evaluation during 3 days in May 2012 including direct observation of
hand hygiene compliance by control nurses and hand cultures of 50
healthcare workers (HCW). Based on the WHO Guidelines on Hand
Hygiene in Health Care [1], cleaning of hands with alcohol-based hand
rubs (Sterillium) was prescribed before touching a patient and before
aseptic procedures, after body fl uid exposure risk and after touching
a patient and touching his/her surroundings. Promotion of the hand
hygiene program consisted of lectures and web-based self-learning,
posters located near points of care and verbal reminders by control
nurses. New observations of hand hygiene by control nurses during
3 days and hand cultures of 50 healthcare providers were performed
in September 2012. Consumption of alcohol-based hand rub (product
volume use per patient-days) was used as a surrogate marker of hand
hygiene over time. The diff erence in hand hygiene compliance during
the two periods was examined using a chi-squared test. Diff erences
in hand cultures were examined using a Student’s t test. Time trends
in the consumption of alcohol-based hand rub were examined using
linear correlation. P <0.05 was considered statistically signifi cant. Results for introduction of a new hand hygiene program in the ICU
P Vos, A Harts-Laurijsen, R Snoeren, R Ramnarain, JA Van Oers
St Elisabeth Hospital, Tilburg, the Netherlands
Critical Care 2013, 17(Suppl 2):P52 (doi: 10.1186/cc11990) The
study was approved by the institutional Ethics Review Board. from March through October 2012, and intervened to change behavior
by providing monthly feedback to specifi c provider groups and
services. We made use of the Unit Coordinator to measure compliance
of all individuals in the ICU. Results Overall compliance by physicians was 82.1%, for nonphysicians
was 84.8%. Feedback to physicians, individually and by service,
dramatically increased hand hygiene compliance, defi ned as both
on entry and exit from the patient room, over the study period. See
Figure 1. Conclusion Physician behavior is responsive to monthly feedback
that is specifi c to the individual or surgical service. Use of the Unit
Coordinator was very eff ective at gathering a very large sample size in
a short period of time. P54
Compliance with the implementation of an ICU cluster-randomized
trial assessing the benefi ts and potential harms of universal glove
and gowning
D Kett1, DJ Morgan2, L Pineles2, MJ Zervos3, LS Munoz-Price1, AD Harris2,
BUGG Investigators2
1University of Miami/Jackson Memorial Hospital, Miami, FL, USA; 2University
of Maryland/VA Maryland Healthcare System, Baltimore, MD, USA; 3Henry
Ford Health System, Detroit, MI, USA
Critical Care 2013, 17(Suppl 2):P54 (doi: 10.1186/cc11992) Reference 1. Pittet D, et al.: Infect Control Hosp Epidemiol 2009, 30:611-622. Compliance with the implementation of an ICU cluster-randomized
trial assessing the benefi ts and potential harms of universal glove
and gowning Results During the survey, in May 158 opportunities to observe hand
hygiene were presented and 286 in September. Overall compliance
improved from 34.2% (54/158) to 51% (146/286), χ2 = 11.7 (P <0.001). In
May, 50 HCW had a mean of 63.20 ± 39.37 colony-forming units (CFU)
on their hands compared with 43.0 ± 40.19 CFU on the hands of 50
HCW in September (P = 0.024). We also observed an initial increased
use of alcohol-based hand rubs from 21 ml per patient-day in May to
a maximum 72 ml per patient-day in June, but a decline to 44 ml per
patient-day in September, Pearson correlation coeffi cient = 0.31 (P =
0.61). D Kett1, DJ Morgan2, L Pineles2, MJ Zervos3, LS Munoz-Price1, AD Harris2,
BUGG Investigators2 g
1University of Miami/Jackson Memorial Hospital, Miami, FL, USA; 2University
of Maryland/VA Maryland Healthcare System, Baltimore, MD, USA; 3Henry
Ford Health System, Detroit, MI, USA
Critical Care 2013, 17(Suppl 2):P54 (doi: 10.1186/cc11992) Introduction The Benefi ts of Universal Glove and Gowning (BUGG)
study is a cluster-randomized trial to evaluate the use of wearing gloves
and gowns for all patient contact in the ICU. The primary outcome is
VRE and MRSA acquisitions; secondary outcomes include frequency of
healthcare worker visits, infection rates, hand hygiene compliance and
adverse events. Conclusion Implementation of a new hand hygiene program at our ICU
resulted in improved hand hygiene compliance and less CFU on the
hands of HCW. There was no signifi cant increased use of alcohol-based
hand rubs over time. The results indicate that constant awareness is
vital for success. Methods We enrolled 20 ICUs in 15 states. ICUs collected nasal and
perianal swabs on all patients at admission and discharge/transfer. After a 3-month baseline period, 10 units were randomized to the
intervention arm and required to wear gloves and gowns for all patient
contact. An intervention toolkit was created based on site feedback
and compliance reports. Swab collection compliance was fed back
and discussed during site conference calls on a weekly basis. Site
coordinators monitored compliance with gloves and gowns, hand
hygiene and frequency of HCW visits and reviewed patient charts for
adverse events. Sinks as a correctable source of ESBL contamination for patients in
the ICU p
yf
g
p
Methods Thirty-nine medical wards (MW) and 12 emergency depart-
ments (ED) were enrolled. Every unit was asked to fi ll in a pre-agreed
HOR Checklist focused on the main requirements suggested by the
Guidelines. I Wolf1, P Bergervoet1, W Van der Zwet1, H Van den Oever1, P Savelkoul2,
F Sebens1 Results Analysing the human resources available, we see that the bed-
to-doctor ratio signifi cantly (P <0.01) increases from the day to the night
shift: from 6 to 43 beds per doctor on the MW (median). Otherwise,
the ED staff remains roughly the same: from 3.5 to 2.5 doctors on duty
(median). The analysis of the organizational tools (Table 1) points out Introduction The incidence of patients carrying ESBL-positive
bacteria in our ICU (12 in 780 admissions in 2011) was not considered
problematic. However, routine cultures had identifi ed ESBL-negative
patients who had become colonized with ESBL strains during their
ICU stay. Self-disinfecting siphons, preventing bacterial growth by
antibacterial coating and intermittent heating, and biofi lm formation
by electromechanical vibration, were placed in all sinks in the ICU. The
aim of the present study was to evaluate the eff ect of this intervention. Methods An intervention study in a 12-bed ICU. The intervention
involved placement of 19 self-disinfecting siphons (Biorec). All patients
with an expected ICU stay of 2 days or more between January 2011
and December 2012 were studied. Samples of throat, sputum and
rectum were taken at admission and twice weekly, and cultured for
ESBLs. Between June 2011 and October 2011, sinks in patient rooms
were cultured regularly for ESBLs. After the intervention in April
2012, multiple repeat cultures were taken. Whenever the species and
antibiogram of bacteria cultured from patients and sinks matched, they
were typed by AFLP. P55
Fighting hospital sepsis
l a low percentage of hospitals having: a Diagnostic and Therapeutic
Protocol for sepsis management (8.3%), some Hospital Empirical
Antibiotic Therapy Guidelines (0%) and an Infective Source Eradication
Protocol (8.3%). Moreover, just 25% of hospitals involve an infectious
diseases expert in every case of severe sepsis or septic shock. p
y
p
p
Conclusion We guess that the poor availability of HOR showed by the
hospitals could have a role in the Guidelines implementation and in
the patient’s outcome. Only a comparison between these results and
data collected from a Clinical Checklist, focused on sepsis bundle
compliance, and from a patient’s outcome summary could confi rm our
hypothesis. This is the aim for our next part of the study. Introduction Sepsis accounts for a very high mortality. The Surviving
Sepsis Campaign recommends a fi rst 6 hours resuscitative bundle to
improve patient outcome. Despite this, the bundle is poorly performed
because of several organizational and cultural barriers. In recognition
of this, we guess that an Educational and Organizational Intervention
out of the ICUs could impact on septic patient outcome. In order to
test our hypothesis we carried out, in 12 hospitals, a pre-intervention
survey of the human and organizational resources (HOR) available
in the management of septic patients. The aim is to seek any barrier
potentially aff ecting correct Guidelines implementation. 1. Dellinger RP, et al.: Intensive Care Med 2008, 34:17-60. Improving hand hygiene performance in a surgical ICU: a novel
method for data collection to change physician behavior
SA Nasraway
Tufts Medical Center, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P53 (doi: 10.1186/cc11991) Results During the 12-month study period, 100,210 swabs were
collected. After the baseline period, we were able to achieve and
maintain swab compliance rates between 85 and 97%. Monthly
discharge compliance increased by 21% by the beginning of the
intervention period (Figure 1). Observers found 86% compliance with
universal glove and gowning over 1,242 30-minute observation periods
(Figure 1). Ninety charts at each site were reviewed for adverse events. Conclusion Over a diverse group of US hospitals, we achieved high
compliance with surveillance cultures and implementing universal
gloving and gowning was achieved quickly with high compliance. Critical Care 2013, 17(Suppl 2):P53 (doi: 10.1186/cc11991) Introduction ICU-acquired infection is directly related to hospital
mortality. Hand hygiene is an eff ective, low-cost intervention that
can prevent the spread of bacterial pathogens, including multidrug-
resistant organisms. Historical compliance with hand hygiene
guidelines by physicians, nurses and other care providers is poor. Methods Present expectations by the Infection Control Committee are
to ‘pump in, pump out’ of every room, using 63% isopropyl alcohol. We
performed 17,622 observations of hand hygiene in the surgical ICU Figure 1 (abstract P54). Swab collection and gown/glove compliance. Figure 1 (abstract P53). Figure 1 (abstract P54). Swab collection and gown/glove compliance. Figure 1 (abstract P53). Figure 1 (abstract P54). Swab collection and gown/glove compliance. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S20 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P55
Fighting hospital sepsis
AR Calini, S Vesconi, R Fumagalli, S Marchesi, L Ghezzi, G Monti
Ospedale Niguarda Ca’ Granda, Milan, Italy
Critical Care 2013, 17(Suppl 2):P55 (doi: 10.1186/cc11993) P55
Fighting hospital sepsis
AR Calini, S Vesconi, R Fumagalli, S Marchesi, L Ghezzi, G Monti
Ospedale Niguarda Ca’ Granda, Milan, Italy
Critical Care 2013, 17(Suppl 2):P55 (doi: 10.1186/cc11993) Clinical effi cacy of recombinant human soluble thrombomodulin in
patients with septic disseminated intravascular coagulation
Y Suzuki, R Sato, M Sato, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P59 (doi: 10.1186/cc11997) Results All enalapril pigs received the maximum NE dose without
reaching the target MAP. Enalapril resulted in lower MAP, higher
CO, and transiently increased regional blood fl ows (Table 1). Fluid
administration and urinary output were similar among groups. Liver
mitochondrial respiration was reduced (State 3, State 4) in the enalapril
group. One animal died in each ML and enalapril group. y
p
Critical Care 2013, 17(Suppl 2):P59 (doi: 10.1186/cc11997) Introduction Thrombomodulin is an endothelial cell cofactor and
glycoprotein for thrombin-catalyzed activation of protein C. A recom-
binant human soluble thrombomodulin (rhsTM) has been recently
developed, and this new agent has a unique amino-terminal structure
exhibiting anti-infl ammatory activity including sequestraction and
cleavage of high-mobility group box 1(HMGB-1). Conclusion Enalapril enhances cardiac output and early regional blood
fl ows in sepsis despite lower MAP, and reduces liver mitochondrial
respiration, suggesting that RAS is involved in the hemodynamic and
metabolic changes in sepsis. Methods In this study, 13 patients with septic disseminated intravascular
coagulation (DIC) were treated with rhsTM, which is Recomodulin® Inj. 12800 (Asahi Kasei Pharma Co., Tokyo, Japan). Patients with septic DIC
were treated with 130 to 380 U/kg/day.i Eff ects of intraperitoneal immunoglobulin therapy on behavior and
cognitive functions in CLP-induced sepsis model in rats
F Esen1, E Senturk1, P Ergin Ozcan1, G Orhun1, S Gümrü2, M Kaya1,
N Orhan1, N Arıcan1, F Arıcıoglu2
1University of Istanbul, Turkey; 2Marmara University Faculty of Pharmacy,
Istanbul, Turkey
Critical Care 2013, 17(Suppl 2):P58 (doi: 10.1186/cc11996) Results There were signifi cant results for improvement of APACHE
II score and DIC diagnostic criteria score for critically ill patients after
treatment using rhsTM (P <0.01). Improvement for platelet count and
D-dimer level were also observed in this study (P <0.05). Activation
of antithrombin (AT) also was signifi cantly increased after treatment
(P <0.05). Hospital mortality was 15.4% in this study. Critical Care 2013, 17(Suppl 2):P58 (doi: 10.1186/cc11996) Introduction The present study investigated the eff ects of a single
dose of intraperitoneal (i.p.) IgG and IgGAM administration on various
behavioral alterations in a cecal ligation perforation (CLP)-induced
sepsis model in rats.i Conclusion The rhsTM might be one of most important endogenous
regulators of coagulation, acting as the major inhibitor of thrombin as
well as AT III. This new agent may play an important role in treatment
for septic DIC. Sinks as a correctable source of ESBL contamination for patients in
the ICU Table 1 (abstract P55)
Availability
Resource
(%)
Diagnostic and therapeutic protocol for septic patient management
8.3
Use of early warning score for diagnosis and management
8.3
Sepsis team
25
Microbiology laboratory: open 7 days a week
66.6
Lactate dosage: 24 hours a day availability
83.3
Central venous catheter insertion (CVC) available 24 hours a day
91.6
Hospital empirical antibiotic therapy guidelines
0
Infectious diseases team advice in any case of severe sepsis septic shock
25
Infective source eradication
Infective source eradication protocols
8.3
Intervention radiology available 24 hours a day
50
Operating room available 24 hours a day
58.3
Specifi c infection management protocol (that is, CVC infection)
58.3
Imaging reporting service available 7 days a week
41.6 yp
y
Results Before intervention Multiple ESBL-forming strains were found in
sinks of all patient rooms. Eighteen patients who were ESBL-negative
on ICU admission became colonized with 11 diff erent ESBL strains,
that were present in sinks of their admission rooms (Figure 1). Four
contaminations were proven by AFLP-tying. One patient died of ESBL-
positive E. cloacae pneumonia. After intervention All sinks were negative
for ESBL strains. No further patients became ESBL colonized during the
ICU stay. y
Conclusion Wastewater sinks were the likely source of ESBL
colonization for 18 ICU patients. After placing self-disinfecting siphons Figure 1 (abstract P56). Design and results of the intervention. Figure 1 (abstract P56). Design and results of the intervention. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S21 Table 1 (abstract P57). Hemodynamic variables, among times and groups there were no new ESBL colonizations in patients. This coincided with
th di
f ESBL t
i
f
ll i k
of immunoglobulin therapy on behavioral changes in CLP-induced
i i
t
Table 1 (abstract P57). Clinical effi cacy of recombinant human soluble thrombomodulin in
patients with septic disseminated intravascular coagulation
Y Suzuki, R Sato, M Sato, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P59 (doi: 10.1186/cc11997) Methods Female Wistar albino rats (200 to 250 g) were divided into fi ve
groups (n = 8): a naive Control group, a Sham operated group receiving
conventional antibiotic treatment, a CLP group receiving CLP procedure
and conventional antibiotic treatment, and IgG and IgGAM groups
which were also applied 1 g/kg, i.p. IgG and IGAM therapy 5 minutes
after the CLP procedure. Ten, 30 and 60 days after the surgery, animals
underwent three behavioral tasks: an open fi eld test to evaluate the
locomotor activity, an elevated plus maze test to measure the level of
anxiety, and a forced swim test to assess the possible depressive state. The results acquired from these tests were used to estimate the eff ect Sinks as a correctable source of ESBL contamination for patients in
the ICU Immunoglobulin therapy prevented the occurrence of these
behavioral changes. Especially, animals in the IgGAM group conserved
the values quite near to those of the control group in measured
parameters. Sinks as a correctable source of ESBL contamination for patients in
the ICU Hemodynamic variables, among times and groups
P
BL
R0h
R12h
R24h
R36h
R48h
(time×group)
MAP (ENL/MH/ML)
66 (9)/72 (15)/
65 (11)/73 (10)/
57 (4)/82 (6)/
58 (8)/78 (6)/
56 (12)/80 (5)/
56 (11)/79 (9)/
0.003
65 (9)
81 (16)
72 (13)
67 (12)
61 (6)
56 (13)
CO (ENL/MH/ML)
106 (13)/89 (7)/
92 (17)/67 (11)/
169 (16)/95 (19)/
187 (22)/129 (28)/
161 (25)/139 (32)/
157 (37)/140 (21)/
<0.0001
105 (13)
88 (22)
104 (13)
143 (31)
145 (13)
150 (18)
Carotid fl ow (ENL/MH/ML)
6 (2)/7 (2)/7 (2)
5 (1)/4 (1)/5 (2)
10 (3)/5 (1)/6 (2)
10 (3)/8 (1)/9 (2)
8 (2)/8 (2)/8 (2)
8 (3)/8 (2)/8 (2)
<0.0001
Femoral fl ow (ENL/MH/ML)
4 (1)/3 (1)/4 (2)
2 (1)/2 (0)/2 (1)
4 (1)/2 (1)/2 (0)
4 (1)/3 (1)/3 (1)
3 (1)/3 (1)/4 (1)
3 (2)/3 (1)/4 (1)
0.004 of immunoglobulin therapy on behavioral changes in CLP-induced
sepsis in rats.ii Results In the open fi eld test, the CLP group showed a signifi cant
decrease in total squares passed on days 10 and 30. Similarly, total
numbers of rearing and grooming were dramatically decreased in the
CLP group in comparison with control and sham groups (P <0.005). In the elevated plus maze test, the number of entries to open arms
decreased in the CLP group. In the forced swim test, there was a
tendency for increase in immobility time in the CLP group, although
the data were statistically insignifi cant. All of these values which were
indicating the importance of behavioral alterations were improved on
day 60. Immunoglobulin therapy prevented the occurrence of these
behavioral changes. Especially, animals in the IgGAM group conserved
the values quite near to those of the control group in measured
parameters. Results In the open fi eld test, the CLP group showed a signifi cant
decrease in total squares passed on days 10 and 30. Similarly, total
numbers of rearing and grooming were dramatically decreased in the
CLP group in comparison with control and sham groups (P <0.005). In the elevated plus maze test, the number of entries to open arms
decreased in the CLP group. In the forced swim test, there was a
tendency for increase in immobility time in the CLP group, although
the data were statistically insignifi cant. All of these values which were
indicating the importance of behavioral alterations were improved on
day 60. P57
Hemodynamic and mitochondrial eff ects of enalapril in
experimental sepsis AJ Pereira, V Jeger, T Corrêa, M Vuda, S Djafarzadeh, J Takala, S Jakob
University Hospital (Inselspital)/University of Bern, Switzerland
Critical Care 2013, 17(Suppl 2):P57 (doi: 10.1186/cc11995) Introduction Sepsis may cause mitochondrial dysfunction. The
renin–angiotensin system (RAS) activity is increased in sepsis, and
can interfere with the mitochondrial function either directly or by
modifying hemodynamics. We studied the eff ects of ACE inhibition
by enalapril on hemodynamics and hepatic mitochondrial function in
sepsis. Two septic groups without enalapril and with diff erent blood
pressure targets served as controls. Conclusion Sepsis, even though it has been treated with conventional
antibiotics, caused a negative eff ect on behavioral parameters. In this
study, IgG and IgGAM treated animals in the presence of CLP did not
show these behavioral changes. Therefore our results suggest that
a single dose of i.p. IgG and IgGAM treatment, which was applied
immediately after the sepsis procedure, prevents behavioral defects
observed following sepsis. p
g
Methods Sepsis (fecal peritonitis) was induced in 24 anesthetized,
mechanically ventilated pigs (40.1 ± 2.1 kg), divided into three groups
(n = 8 for each): ENL (enalapril pretreatment for 7 days and at 0.005 to
0.02 mg/kg/hour during the study; MAP target 75 to 85 mmHg), ML
(target MAP 50 to 60 mmHg), and MH (target MAP 75 to 85 mmHg). Hemodynamic support with fl uids, norepinephrine (NE; maximum
dose 5,000 μg/hour), and antibiotics were started after 12 hours of
peritonitis and continued for 48 hours. P59 59
Clinical effi cacy of recombinant human soluble thrombomodulin in
patients with septic disseminated intravascular coagulation
Y Suzuki, R Sato, M Sato, S Endo
Iwate Medical University, Morioka, Japan
Critical Care 2013, 17(Suppl 2):P59 (doi: 10.1186/cc11997) P61 A clinical randomized trial on the use of atorvastatin in patients
with sepsis or septic shock: eff ects on endothelial function
K Prado, C Ribeiro, T Furian, R Pinto Ribeiro, D Silvello, L Rohde, N Clausell,
L Becker
Hospital de Clinicas de Porto Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P61 (doi: 10.1186/cc11999) Results Forty-three patients (18 treated, 25 control) completed the
study and were further analyzed. Incidence of organ dysfunction at
enrollment (treatment vs. control) was: septic shock (94% vs. 100%,
P = 0.42), acute respiratory distress syndrome (67% vs. 56%, P = 0.33),
and renal failure (39% vs. 24%, P = 0.54). During 115 treatments no
serious device-related adverse events occurred. On average, there
were no changes in hematology and other blood parameters except
for a modest reduction in platelet count (<10%) and albumin (<5%)
with treatment. Hemoperfusion decreased IL-6 blood concentration
signifi cantly (–49.1%, P = 0.01), with similar reductions of MCP-1
(–49.5%, P = 0.002), IL-1ra (–36.5%, P = 0.001), and IL-8 (–30.2%, P =
0.002). The 28-day mortality (28% vs. 24% control, P = 0.84) and 60-
day mortality (39% vs. 32% control, P = 0.75) did not diff er signifi cantly
between the two studied groups.i Introduction Sepsis and septic shock are complex infl ammatory
syndromes. Multiple cellular activation processes are involved, and
many humoral cascades are triggered. Presumably, endothelial cells
play a pivotal rule in the pathogenesis of sepsis, not only because
they may infl uence the infl ammatory cascade but also because, upon
interaction with excessive amounts of infl ammatory mediators, the
function of these cells may become impaired. It is likely that a general
dysfunction of the endothelium is a key event in the pathogenesis
of sepsis [1]. HMG-CoA-reductase inhibitors have been shown to
exhibit pronounced immunomodulatory eff ects independent of lipid
lowering. Most of these benefi cial eff ects of statins appear to involve
restoring or improving endothelial function [2]. We hypothesize that
statins can improve endothelial dysfunction in septic patients. Conclusion In this fi rst clinical study of a novel sorbent hemoadsorption
device in patients with severe sepsis and ALI, the device appeared to
be safe and decreased the blood concentration of several cytokines. Further research is needed to study the eff ect of the device on the
clinical outcome of septic patients. y
Methods A double-blinded, placebo-controlled, randomized trial
was undertaken. A multicenter randomized controlled study of an extracorporeal
cytokine hemoadsorption device in septic patients D Schädler1, C Porzelius2, A Jörres3, G Marx4, A Meier-Hellmann5,
C Putensen6, M Quintel7, C Spies8, C Engel2, N Weiler1, M Kuhlmann9
1University Medical Center Schleswig-Holstein, Campus Kiel, Germany;
2University of Leipzig, Germany; 3Charité University Hospital Campus Virchow-
Klinikum, Berlin, Germany; 4RWTH University Hospital Aachen, Germany;
5HELIOS Klinikum, Erfurt, Germany; 6University of Bonn, Germany; 7University
Hospital Göttingen, Germany; 8Campus Charité Mitte and Campus Charité
Virchow-Klinikum, Charité-University Medicine Berlin, Germany; 9Vivantes
Klinikum im Friedrichshain, Berlin, Germany i
Results There were 85 patients (52 men, 33 women; age range 19 to
92 years (mean 69.0 ± 16.9)), and the mortality rate was 27.1% and
APACHE II score was 20.1 ± 7.6. WBC, CRP, PT and SOFA score were
signifi cantly improved after AT III concentrate therapy (13,176 ± 9,400
vs. 11,693 ± 7,089, P = 0.011, 15.9 vs. 13.3, P = 0.0015, 16.5 ± 4.4 vs. 15.9 ± 5.9, P = 0.0045, and 9.7 ± 3.8 vs. 8.6 ± 4.6, P = 0.01, respectively). Platelet was signifi cantly decreased (11.7 ± 9.7 vs. 10.6 ± 9.0, P = 0.028),
while there were no signifi cant diff erences in fi brinogen, FDP and
DIC score (387.3 ± 202.6 vs. 381.8 ± 163.2, P = 0.088, 36.9 ± 52.7 vs. 28.8 ± 39.9, P = 0.059, and 2.9 ± 1.6 vs. 2.6 ± 1.8, P = 0.25, respectively)
after the therapy. One week after the therapy, platelet and DIC score
were signifi cantly improved compared with before the therapy
(15.6 ± 10.0, P = 0.0036 and 1.8 ± 1.9, P = 0.0041). ,
,
y
Critical Care 2013, 17(Suppl 2):P62 (doi: 10.1186/cc12000) Introduction A novel sorbent hemoadsorption device for cytokine
removal (CytoSorbents, USA) was developed and successfully tested
in animal models of sepsis. The experience in the clinical setting is
still limited to case reports. In this fi rst clinical trial, we tested the
hypothesis that treatment with sorbent hemoadsorption could safely
and eff ectively reduce cytokines in septic patients with acute lung
injury (ALI). Introduction A novel sorbent hemoadsorption device for cytokine
removal (CytoSorbents, USA) was developed and successfully tested
in animal models of sepsis. The experience in the clinical setting is
still limited to case reports. In this fi rst clinical trial, we tested the
hypothesis that treatment with sorbent hemoadsorption could safely
and eff ectively reduce cytokines in septic patients with acute lung
injury (ALI). References 1. Hack EC, et al.: The endothelium in sepsis: source of and a target for
infl ammation. Crit Care Med 2001, 29:S21-S27.if 1. Hack EC, et al.: The endothelium in sepsis: source of and a target for
infl ammation. Crit Care Med 2001, 29:S21-S27.if l
p
Methods From January 2009 to December 2011, adult septic patients,
whom were given AT III concentrate in our medico-surgical ICU, were
included in this study. AT III concentrate was administered 30 to 60 U/kg
intravenously every 24 hours for 3 days in the patients with DIC status
and AT III defi ciency. Between before and after the AT III concentrate
therapy, WBC (/mm3), CRP (mg/dl), platelet (×104/μl), PT (seconds),
fi brinogen (mg/dl), FDP (μg/ml), SOFA score and DIC score were
evaluated. Values are expressed as mean ± SD. Data were analyzed by
Wilcoxon signed-rank test. P <0.05 was considered signifi cant. 2. Davignon J: Benefi cial cardiovascular pleiotropic eff ects of statins. Circulation 2004, 109(Suppl III):III-39-III-43. 2. Davignon J: Benefi cial cardiovascular pleiotropic eff ects of statins. Circulation 2004, 109(Suppl III):III-39-III-43. P60 Antithrombin III concentrate therapy may have an effi cacy in sepsis
N Saito, M Takeda, T Harada, R Moroi, M Namiki, A Yaguchi
Tokyo Women’s Medical University, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P60 (doi: 10.1186/cc11998) Introduction Antithrombin III (AT III) has been known to contribute to
anti-infl ammatory response as well as its anticoagulation. Our previous S22 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Conclusion Our data showed no benefi t with the use of a potent
statin acutely in patients with sepsis or septic shock with regards to
improvement in endothelial function. study showed AT III defi ciency happened in the early stage of sepsis
with no relation to DIC status. However, whether AT III concentrate
is a benefi cial therapy or not for septic patients is controversial. Our hypothesis is that AT III concentrate may have effi cacy as anti-
infl ammation for sepsis. study showed AT III defi ciency happened in the early stage of sepsis
with no relation to DIC status. However, whether AT III concentrate
is a benefi cial therapy or not for septic patients is controversial. Our hypothesis is that AT III concentrate may have effi cacy as anti-
infl ammation for sepsis. P61 We enrolled adult patients within 24 hours of severe
sepsis or septic shock diagnosis and randomized them to placebo or
atorvastatin 80 mg/day for a short term. Endothelial dysfunction was
assessed measuring plasmatic levels of IL-6, ET-1, VCAM-1 by ELISA and
measuring fl ow-mediated vasodilatation of the brachial artery at basal,
24 and 72 hours after randomization. A multicenter randomized controlled study of an extracorporeal
cytokine hemoadsorption device in septic patients Conclusion In the patients with septic DIC, WBC, CRP and SOFA score
were immediately improved after the AT III concentrate therapy, while
platelet and DIC score were improved later. AT III concentrate may also
contribute to anti-infl ammatory for septic DIC with anticoagulation. Methods Ventilated patients fulfi lling the criteria for severe sepsis and
ALI were enrolled in this multicenter randomized, controlled, open-
label study comparing standard of care with or without hemoperfusion
treatment. Primary endpoints were safety and IL-6 reduction. Treated
patients underwent hemoperfusion at fl ow rates of ~200 to 300 ml/
minute for 6 hours per day for 7 consecutive days. The overall mean
reduction in individual plasma cytokines for the control and treatment
groups during the treatment period was calculated using a generalized
linear model. P64 Methods Adult patients with septic shock who required renal
replacement therapy (RRT), with a confi rmed endotoxemia, and
suspected Gram-negative infection were consecutively added to the
study within the fi rst 24 hours after diagnosis. All patients received
full standard treatment for septic shock. Endotoxin elimination was
performed using the membrane oXiris (Gambro, Sweden), a medical
device for continued RRT with the unique feature of endotoxin
adsorbtion. An endotoxin activity assay was used to monitor endotoxin
elimination therapy at baseline (T0), 3 hours (T1), 12 hours (T2),
24 hours (T3), 48 hours (T4), and 72 hours (T5). Our key indicators were
the improvement in hemodynamics and organ function, and decrease
of endotoxin activity (EA) in blood. Continuous variables are presented
as mean values with standard deviations. Combined use of pumpless extracorporeal lung assist system and
continuous renal replacement therapy with citrate anticoagulation
in polytrauma patients y
HK Atalan, M Dumantepe, TB Denizalti, IA Tarhan, A Ozler
Atasehir Memorial Hospital, Istanbul, Turkey
Critical Care 2013, 17(Suppl 2):P64 (doi: 10.1186/cc12002) Introduction The usefulness of a pumpless extracorporeal lung assist
system (pECLA) and continuous renal replacement therapy (CRRT) in
critically ill patients has been demonstrated in previous studies [1,2]. The aim of this report was to examine combined use of pECLA and
CRRT to improve carbon dioxide and infl ammatory mediator removal,
which allows for lung protective ventilation strategies. Results High EA level at baseline (0.74 ± 0.14 endotoxin activity units
(EAU)) signifi cantly decreased during RRT with oXiris membrane
to 0.46 ± 0.02 (T1), 0.34 ± 0.01 (T2), 0.4 ± 0.02 (T3), 0.46 ± 0.04 (T4),
0.35 ± 0.07 (T5) EAU (P <0.05). MAP increased from baseline 72 ± 14
to 81 ± 18, 76 ± 6, 77 ± 7, 83 ± 13, 87 ± 10 mmHg (P <0.05), and the
mean norepinephrine use decreased from 0.23 ± 0.04 to 0.19 ± 0.02,
0.11 ± 0.01, 0.09 ± 0.01, 0.04 ± 0.01, 0.0 μg/kg/minute (P <0.05) at T0, T1,
T2, T3, T4, T5, respectively. The SOFA score had decreased from 14 ± 4 to
12 ± 2, 9 ± 3, 7 ± 3 points (P <0.05), and the procalcitonin level declined
from 107 ± 123 to 45 ± 41, 29 ± 30, 17 ± 157 ± 1 ng/ml (P <0.05) at T0,
T3, T4, T5. Methods In our 10 patients with ARDS due to polytrauma and sepsis,
pECLA was established by insertion of cannulae to the femoral artery
and vein. CRRT cannulae were introduced by venous line of the same
vascular access (Figure 1). We preferred regional anticoagulation with
trisodium citrate for both CRRT and ILA. Figure 1 (abstract P64). ILA and CRRT connected to the patient. Conclusion RRT with oXiris membrane resulted in the eff ective
elimination of endotoxins from the blood. The therapy was associated
with an increase in blood pressure, a reduction of vasopressor
requirements, and an improvement of organ function. The application
of the endotoxin activity assay was useful for bedside monitoring of
endotoxemia in ICU patients. P63 Continuous renal replacement therapy with the adsorbent
membrane oXiris in septic patients: a clinical experience
F Turani, F Candidi, R Barchetta, E Grilli, A Belli, E Papi, A di Marzio, M Falco
Aurelia Hospital/European Hospital, Rome, Italy
Critical Care 2013, 17(Suppl 2):P63 (doi: 10.1186/cc12001) Results We studied 47 patients, 24 in the placebo group (mean age
52 ± 20 years, 29.1% male; APACHE II risk score 23.5 ± 7.3) and 23 in
the statin group (mean age 49.5 ± 18 years, 53.4% male; APACHE II risk
score 23 ± 6.9). The baseline characteristics of the placebo group were
similar to statin patients as well as the mean length of stay in the ICU
(8.6 ± 7.4 and 9.1 ± 8 days, respectively) and the time on vasopressors
(49.3 ± 47.1 and 59 ± 91.1 hours, respectively). No signifi cant diff erence
was observed on the temporal variation of biomarker levels (IL-6,
VCAM-1, ET-1) between treatment and control groups. The intrahospital
mortality rate was 26% in the statin group and 45% in the placebo
group (P = 0.17). Introduction Renal failure is an important complication of sepsis and
CRRT with adsorbing membranes may be useful in this clinical setting
[1]. The aims of the study in septic/septic shock patients are to evaluate:
the safety of a new hemofi lter membrane oXiris with adsorbing
properties and anti-endotoxin activity; the renal and hemodynamic Introduction Renal failure is an important complication of sepsis and
CRRT with adsorbing membranes may be useful in this clinical setting
[1]. The aims of the study in septic/septic shock patients are to evaluate:
the safety of a new hemofi lter membrane oXiris with adsorbing
properties and anti-endotoxin activity; the renal and hemodynamic S23 Table 1 (abstract P63)
Parameter
Units
T0
T1
Creatinine
mg/dl
1.9 ± 0.1
1.18 ± 0.1*
Diuresis
ml/24 hours
1,284 ± 78
1,573 ± 98
Norepinephrine
μg/kg/minute
0.17 ± 0.2
0.06 ± 0.1*
IL-6
pg/ml
572 ± 78
278 ± 57*
Procalcitonin
ng/ml
35 ± 7
15 ± 2*
Endotoxin
Level/U
0.64 ± 0.2
0.49 ± 0.1
*P <0.05 vs. T0. Results Mean SAPS II and APACHE II scores were 55 and 23 respectively. Mean time on mechanical ventilation was 22 days. Mean ICU stay was
30 days for survivors and 38 days for nonsurvivors. 1.
Marshall JC, Foster D, Vincent JL, Cook DJ, Cohen J, Dellinger RP, Opal S,
Abraham E, Brett SJ, Smith T, Mehta S, Derzko A: Diagnostic and prognostic
implications of endotoxemia in critical illness: results of the MEDIC Study.
J Infect Dis 2004, 190:527-534. P66
Use of extracorporeal endotoxin elimination therapy for septic
shock gi
Results Table 1 presents the main results of this study. y
Conclusion In septic/septic shock patients with renal failure, CRRT with
a new treated heparin-coated membrane (oXiris; Gambro) is clinically
feasible, and has a positive eff ect on renal function and hemodynamics. An adsorbing eff ect on proinfl ammatory mediators may have a role in
these results. These data and the trend toward a decrease of endotoxin
during the treatment warrant further investigation. Reference Introduction Endotoxin, a component of the outer membrane of Gram-
negative bacteria, is considered an important factor in pathogenesis
of septic shock [1]. The aim of our study was to determine whether
endotoxin elimination treatment added to the standard treatment
would improve organ function in patients with septic shock. 1. Rimmelé T, et al.: Nephrol Dial Transplant 2009, 24:421-427. P63 When compared
with baseline values most relevant parameters were the improvement
in tidal volumes, plateau pressures, PaCO2 levels and pH (Figure 2). Four
patients survived while six patients died from sepsis-MOF.f Results Mean SAPS II and APACHE II scores were 55 and 23 respectively. Mean time on mechanical ventilation was 22 days. Mean ICU stay was
30 days for survivors and 38 days for nonsurvivors. When compared
with baseline values most relevant parameters were the improvement
in tidal volumes, plateau pressures, PaCO2 levels and pH (Figure 2). Four
patients survived while six patients died from sepsis-MOF. Conclusion We concluded that pECLA can eff ectively address the
impaired gas exchange in ARDS and CRRT is a safe procedure with
potential therapeutic value for treating MOF. Citrate anticoagulation
was well tolerated and fi lter life was appropriate. The use of the same
vascular access for ILA and CRRT may minimize invasive procedures
and related side eff ects. Conclusion We concluded that pECLA can eff ectively address the
impaired gas exchange in ARDS and CRRT is a safe procedure with
potential therapeutic value for treating MOF. Citrate anticoagulation
was well tolerated and fi lter life was appropriate. The use of the same
vascular access for ILA and CRRT may minimize invasive procedures
and related side eff ects. References 1. Bein T, et al.: Int J Emerg Med 2010, 3:177-179. 2. Demetrios J, et al.: Kidney Int 2005, 67:2361-2367. 1. Bein T, et al.: Int J Emerg Med 2010, 3:177-179. response; and the changes of endotoxin and proinfl ammatory
molecules. response; and the changes of endotoxin and proinfl ammatory
molecules. Methods Forty septic/septic shock patients with renal failure were
enrolled in the study. All patients had preoperative endotoxin
>0.6 level/units (EAA Spectral D) and were submitted to high-volume
hemodiafi ltration (50 ml/kg/hour, Prismafl ex; Gambro) with a new
treated heparin-coated membrane (oXiris; Gambro). At T0 (pre-
treatment) and T1 (24 hours) the main clinical and biochemical data
were evaluated. All data are expressed as mean ± SD. One-way ANOVA
test with Bonferroni correction was used to evaluate the data changes. P <0.05 was considered signifi cant. P65
Abstract withdrawn Reference 1. Marshall JC, Foster D, Vincent JL, Cook DJ, Cohen J, Dellinger RP, Opal S,
Abraham E, Brett SJ, Smith T, Mehta S, Derzko A: Diagnostic and prognostic
implications of endotoxemia in critical illness: results of the MEDIC Study. J Infect Dis 2004, 190:527-534. Figure 1 (abstract P64). ILA and CRRT connected to the patient. S24 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 Figure 2 (abstract P64). Changes in tidal volume, plateau pressure, pH and PaCO2. CO Figure 2 (abstract P64). Changes in tidal volume, plateau pressure, pH and PaCO2. P67 (133; 286) to 132 (68; 155) mg/l, P = 0.015 and SOFA score from 7.0 (3,0;
8.0) to 6.0 (3,0; 7.0), P = 0.007. At 24 hours after Alteco LPS, signifi cantly
decreased PCT from 8.7 (3.0; 25.9) to 4.8 (2.1; 10.0) ng/ml. The 28-day
mortality rate was 14.2% (n = 2) in the PMX-F group and 31.3% (n = 10)
in the Alteco LPS group. P67
Selective adsorption of lipopolysaccharide in the complex
treatment of patients with severe sepsis
S Rei, I Aleksandrova, V Kiselev, M Ilynskiy, G Berdnikov, L Marchenkova,
G Bulava, N Borovkova
Hospital Research Institute for Emergency Medicine named after N.V. Sklifosovsky, Moscow, Russia
Critical Care 2013, 17(Suppl 2):P67 (doi: 10.1186/cc12005) Table 1 (abstract P67). Clinical characteristics of the groups
Characteristic
PMX-F
Alteco LPS
Sex M/F
9/5
17/15
Age (years)
41.6 ± 17.1
42.1 ± 13.5
Shock
6 (42.9%)
13 (40.6%)
APACHE II
21 (17; 25)
21 (16; 24)
SOFA
7 (3.0; 8)
7 (3.5; 11) Table 1 (abstract P67). Clinical characteristics of the groups Introduction Severe sepsis and septic shock remain the most serious
problem of critical care medicine with a mortality rate of 30 to 55%
[1]. Several studies have demonstrated positive eff ects of selective
adsorption of LPS on blood pressure, PaO2/FiO2 ratio, endotoxin
removal and mortality [2,3]. The purpose of the study was to evaluate
the effi ciency of using the selective adsorption of LPS, Toraymyxin –
PMX-F (Toray, Japan) and Alteco® LPS Adsorber (Alteco Medical AB,
Sweden), in the complex treatment of patients with severe sepsis. Methods Forty-six patients with Gram-negative sepsis in the
postoperative period were enrolled into the study. Toraymyxin – PMX-F
was used in the PMX-F group (n = 14), while Alteco LPS adsorption
was used in the Alteco LPS group (n = 32). P69 Results Survival is shown in Figure 1. Experiments in each animal were
starting at 7:00 am. At 8:00 am, respective mean free cortisol of groups
A, C and D was 1.81, 0.55 and 2.05 μg/dl; at 1:00 pm they were 0.92, 0.47
and 1.40 μg/dl. Reference The clinical characteristics
are listed in Table 1. The SOFA score, PaO2/FiO2, procalcitonin (PCT),
C-reactive protein (CRP), endotoxin activity assay (EAA) was noted
before, 24 and 48 hours after the selective adsorption of LPS. Conclusion The use of the LPS-selective adsorption (particularly
PMX-F) in patients with severe sepsis leads to improvement of systemic
infl ammation and organ dysfunction. References 1. Vincent JL, et al.: Crit Care Med 2006, 34:344-353. Results At 48 hours after PMX-F, signifi cantly decreased PCT from 17.5
(5.0; 40.9) to 7.1 (4.8; 13.0) ng/ml, P = 0.028, decreased CRP from 180 2. Cruz DN, et al.: JAMA 2009, 301:2445-2452. 3. Yaroustovsky M et al.: Blood Purif 2009, 28:227-233. S25 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 or when vasopressor infusion is terminated whilst 11% after a fi xed
duration. Withdrawal of CS also diff ered, in that 25% tapered/weaned
steroids, 31% stopped it abruptly and 44% of CCPs would base their
CS cessation pattern on the clinical context. Only 46% of CCPs believe
that CS is benefi cial whereas 44% were unsure of the benefi ts in septic
shock. Only 29 (36%) responders indicated that their critical care unit
had a written protocol for CS in septic shock. References References
1. Dellinger et al.: Crit Care Med 2008, 36:296-327. 2. Bourne et al.: Anaesthesia 2003, 58:591-596. References
1. Dellinger et al.: Crit Care Med 2008, 36:296-327. 2. Bourne et al.: Anaesthesia 2003, 58:591-596. 1. Dellinger et al.: Crit Care Med 2008, 36:296-327. 2. Bourne et al.: Anaesthesia 2003, 58:591-596. Need for restoration of cortisol serum levels for successful
antimicrobial therapy in experimental sepsis T Doulias1, A Pistiki2, P Christopoulos1, V Papaziogas1,
E Giamarellos-Bourboulis2, I Koutelidakis1
1University of Thessaloniki, Medical School, Thessaloniki, Greece; 2University of
Athens, Medical School, Athens, Greece
Critical Care 2013, 17(Suppl 2):P70 (doi: 10.1186/cc12008) T Doulias1, A Pistiki2, P Christopoulos1, V Papaziogas1,
E Giamarellos-Bourboulis2, I Koutelidakis1
1University of Thessaloniki, Medical School, Thessaloniki, Greece; 2University of
Athens, Medical School, Athens, Greece
Critical Care 2013, 17(Suppl 2):P70 (doi: 10.1186/cc12008) yg
y
y
Results This study was terminated early as interim analysis identifi ed
no signifi cant clinical benefi t. Baseline characteristics (age/APACHE
IV score) were similar between two groups of patients. No signifi cant
diff erence was noted between two groups with respect to change in
total SOFA score (+1 vs. –5.5 for AT vs. ST, P = 0.382), vasopressor score
(–29 vs. –46.6, P = 0.775), PaO2/FiO2 ratio (–26 vs. +163, P = 0.199), total
urine output from 0 to 72 hours (3,850 ml vs. 4,570 ml, P = 0.355) and
28-day mortality (14.3% vs. 37.5%, P = 0.569). No signifi cant side eff ect
was noted when using this new hemoadsorption device. Introduction It is postulated that clinical benefi t of low-dose
hydrocortisone in septic shock is related to reversal of relative adrenal
insuffi ciency [1]. This was proved in an animal model of sepsis. fi
Methods Sixty-nine Wistar male rats were assigned to the following
groups: A, sham-operation; B, sepsis; C, bilateral adrenalectomy
and sepsis; D, bilateral adrenalectomy, sepsis and hydrocortisone
treatment; E, bilateral adrenalectomy, sepsis and ertapenem treatment;
and F, bilateral adrenalectomy, sepsis, hydrocortisone and ertapenem
treatment. Sepsis was induced by the i.p. infusion of 1×106 cfu/ml of
Escherichia coli after adrenalectomy. Hydrocortisone 10 mg/kg was
infused i.v. bid starting 1 hour after bacterial challenge. Ertapenem
5 mg/kg was infused i.v. once daily starting 1 hour after bacterial
challenge. Survival was recorded. In a separate set of experiments in
18 rats, animal sacrifi ce was performed to measure the free cortisol
concentration. Conclusion This small study cannot identify any extra clinical benefi t
on addition of Alteco endotoxin hemoadsorption to standard therapy
in patients suff ering from intra-abdominal sepsis with shock due to
Gram-negative bacterial infection. g
Reference 1. Cruz DN, et al.: JAMA 2009, 301:2445-2452. 1. Cruz DN, et al.: JAMA 2009, 301:2445-2452. P68
Alteco endotoxin hemoadsorption in Gram-negative septic shock
patients HP Shum, KC Chan, WW Yan
Pamela Youde Nethersole Eastern Hospital, Hong Kong
Critical Care 2013, 17(Suppl 2):P68 (doi: 10.1186/cc12006) Introduction Septic shock is a common cause of mortality and
morbidity in the ICU. Endotoxin hemoadsorption using a Polymyxin B
fi ber column can improve patient outcome [1]. This study investigated
the therapeutic eff ect of a new endotoxin hemoadsorption device in
Gram-negative septic shock patients. Conclusion The perceptions, usage and cessation of CS in septic shock
vary but do appear to have shifted in the last decade. A UK survey in
2003 identifi ed that only 60% of ICUs used CS for septic shock and
over 22% perform a short synacthen test [2]. It appears that many
intensivists are using CS for septic shock, despite confl icting outcome
data. We all strive to practice evidence-based medicine but until we
have a robust, reliable and methodical randomised control trial that
attempts to resolve the CS debate, practice will remain diverse on this
subject, as refl ected by our survey. g
p
p
Methods An open, controlled, prospective, randomized, single-centre
trial conducted between February 2010 and June 2012. Patients
with septic shock due to intra-abdominal sepsis were recruited and
randomized to either standard therapy (ST, n = 8) or standard therapy
plus two 2 hourly sessions of Alteco endotoxin hemoperfusion (AT, n =
7). Standard therapy included infective source control, appropriate
early antibiotic, low-dose steroid, early continuous renal replacement
therapy in the presence of acute kidney injury (RIFLE Injury class or
more), hemodynamic optimization and lung-protected ventilatory
support. Primary outcome was change in organ dysfunction at 48
hours measured by Sequential Organ Failure Assessment (SOFA)
scores. Secondary outcomes were change in vasopressor requirement
(measured by vasopressor score), PaO2/FiO2 (fraction of inspired
oxygen) ratio, total urine output at 72 hours and 28-day mortality.i Effi cacy and utility of preemptive anti-Gr(+) coverage in selected
critically ill patients Effi cacy and utility of preemptive anti-Gr(+) coverage in selected
critically ill patients
VT Todorova, SM Milanov, G Georgiev, M Milanov
Emergency Hospital ‘Pirogov’, Sofi a, Bulgaria
Critical Care 2013, 17(Suppl 2):P73 (doi: 10.1186/cc12011) critically ill patients
VT Todorova, SM Milanov, G Georgiev, M Milanov
Emergency Hospital ‘Pirogov’, Sofi a, Bulgaria
Critical Care 2013, 17(Suppl 2):P73 (doi: 10.1186/cc12011) Introduction Based on the results of our previous studies [1] we have
identifi ed clinical risk factors for the emergence of Gr(+) infections in
our ICU and we have developed a new algorithm for combating them. The choice of the particular antibiotic drug is guided by additional
risk factors for severity of illness and data on the infectious focus. The
response to therapy and its duration are also stated. The aim of the
current study was to evaluate the effi cacy and safety of this preemptive
approach. Results In total, 383 patients were admitted with septic shock, of
whom 253 (66%) were included; 231(92%) had appropriate antibiotic
therapy, of whom 52 (22.5%) died at 7 days. Average time to initiation
of appropriate antibiotic therapy was 9 ± 23 hours. By bivariate
analysis, body mass index (BMI) <20, SAPS II ≥56, SOFA score ≥11 and
bacteremia were signifi cantly associated with 7-day mortality. Urinary
tract infection (UTI) was a protective factor. Age, sex, comorbidities
(particularly immunosuppression), Knaus score, nosocomial infection
and type of germ had no infl uence on 7-day mortality. By multivariate
logistic regression, BMI <20 (OR = 4.87, 95% CI = 1.36 to 17.43, P = 0.01)
and SOFA score ≥11 (OR = 7.99, 95% CI = 3.11 to 20.5, P <0.001) were
the only factors signifi cantly associated with 7-day mortality. UTI was a
signifi cant protective factor (OR = 0.30, 95% CI = 0.10 to 0.88, P = 0.03). Time to initiation of appropriate antibiotherapy was not associated
with 7-day mortality (OR = 0.99, 95% CI = 0.99 to 1.00, P = 0.48). pp
Methods A randomized prospective controlled trial was carried out
from September 2010 to September 2012. Patients were submitted
to block randomization and stratifi ed on the basis of their initial SAPS
II exp score. Antibiotic therapy was started on the day of inclusion
in the treatment group and only with proven Gr(+) pathogen in the
control group. Steroid therapy in septic shock: survey of practice amongst UK
critical care physicians G Rajendran, K Dasari, A Dhrampal j
,
,
p
Norfolk & Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P69 (doi: 10.1186/cc12007) Figure 1 (abstract P70). Survival of rats. Introduction Corticosteroid (CS) therapy in sepsis remains contro-
versial and was fi rst introduced in sepsis management for its anti-
infl ammatory property. CS has found a role in septic shock amelioration
with inconsistent outcomes. The Surviving Sepsis Campaign (SSC)
includes CS as a Level 2C recommendation in septic shock [1]. Adapting
and practicing SSC guidelines vary between critical care units. Accordingly, a survey was conducted to elucidate the usage of CS for
septic shock by UK critical care physicians (CCPs). p
y
p y
Methods Following approval by the UK Intensive Care Society (ICS), the
survey was publicised on the ICS website and its newsletter. y
Results A total of 81 intensivists responded to this online survey. Seventy-four (92.5%) CCPs prescribed CS only if the septic shock is
poorly responsive to fl uid resuscitation and vasopressor therapy. Six (7.5%) initiated CS at the same time as vasopressor therapy. None initiated CS for patients with severe sepsis. No CS other than
hydrocortisone is being used. The most commonly used intravenous
regimen is 50 mg 6 hourly (65%) followed by 50 mg 8 hourly (11%). Only 10% of CCPs would prescribe it by infusion. Less commonly used
regimens were 100 mg 8 hourly (6%) and 100 mg 6 hourly (5%). Only
5% would consider adding fl udrocortisone. Prior to initiating CS, 5% of
CCPs would perform a short synacthen test, while 94% would not. The
majority (89%) of CCPs would stop CS after resolution of shock state Figure 1 (abstract P70). Survival of rats. S26 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 cases during the outbreak. Completed questionnaires describing 27
patients, 11 nonsurvivors and 16 survivors, were returned.i cases during the outbreak. Completed questionnaires describing 27
patients, 11 nonsurvivors and 16 survivors, were returned.i Conclusion Even when eff ective antimicrobial treatment is adminis-
tered, administration of hydrocortisone at a regimen restoring normal
secretion is mandatory for survival. Milanov S, et al.: Gram positive nosocomial infections in a general ICU –
emerging new clues. Crit Care 2011, 15(Suppl 1):P224. Steroid therapy in septic shock: survey of practice amongst UK
critical care physicians p
Results In preliminary analysis of categorical data, a signifi cantly
(Fisher exact test) greater proportion of patients with compared with
without the following fi ndings did not survive; history of alcohol
use (P = 0.05); the presence of lethargy (P = 0.01), confusion (P =
0.03), nausea (P = 0.04), abdominal pain (P = 0.02), or the need for
vasopressors (P = 0.002), oxygen, mechanical ventilation, or steroids
(all P = 0.004) at presentation; and excessive bleeding at surgery (P =
0.01). Initial analysis of continuous data demonstrated that, compared
with survivors at presentation, nonsurvivors had signifi cantly (one-way
ANOVA) increased respiratory rate, percent neutrophils on complete
blood count, hemoglobin, INR, C-reactive protein, and bilirubin and
signifi cantly decreased temperature, systolic blood pressure, platelets,
sodium, albumin, calcium (corrected for albumin), base excess and
bicarbonate (all P ≤0.05).f 1. Annane D, et al.: JAMA 2002, 288:862-871. Effi cacy and utility of preemptive anti-Gr(+) coverage in selected
critically ill patients Initial data were gathered on demographics, diagnosis,
proven risk factors for sepsis-related mortality, severity of infl ammatory
response, ventilator-associated pneumonia and organ dysfunction. Dynamics of SIRS, CPIS and SOFA scores, subsequent infectious isolates,
ventilator-free days, length of ICU stay and outcome were followed for
each patient. y
y
Conclusion Prognosis at 7 days of patients with septic shock is
largely related to the number of failing organs. The majority received
appropriate antibiotic therapy although time to initiation is longer
than recommended. Further eff orts are warranted to reduce mortality
in patients with septic shock. p
Results A total of 170 patients were enrolled. No statistically signifi cant
diff erences in their basal characteristics were found. The subsequent
score values, length of ICU stay and the number of ventilator-free days
were also comparable between groups. The majority of Gr(+) pathogens
were isolated between 6 and 10 days of inclusion. No diff erences were
found regarding the concomitant Gr(–) fl ora and the related antibiotic
therapy. The new organ dysfunction severity was similar in both groups
(P = 0.37). The in-hospital mortality was 26.2% in the treatment group
versus 18.6% in the control group (P = 0.56). Signifi cant diff erences
between the Kaplan–Meier estimates of survival were also not found
(log-rank test P = 0.81). No major adverse reactions were observed. Results A total of 170 patients were enrolled. No statistically signifi cant
diff erences in their basal characteristics were found. The subsequent
score values, length of ICU stay and the number of ventilator-free days
were also comparable between groups. The majority of Gr(+) pathogens
were isolated between 6 and 10 days of inclusion. No diff erences were
found regarding the concomitant Gr(–) fl ora and the related antibiotic
therapy. The new organ dysfunction severity was similar in both groups
(P = 0.37). The in-hospital mortality was 26.2% in the treatment group
versus 18.6% in the control group (P = 0.56). Signifi cant diff erences
between the Kaplan–Meier estimates of survival were also not found
(log-rank test P = 0.81). No major adverse reactions were observed. Conclusion The implementation of this new policy failed to reduce
the degree of organ dysfunction severity and was not associated with
signifi cant survival benefi t. Moreover, even though it did not reach
statistical signifi cance, a second peak of Gr(+) isolates was observed
as a possible complication of the preemptive therapy. P71
Impact of time to initiation of appropriate antibiotic therapy on
early mortality of patients with septic shock
JP Quenot1, C Binquet2, S Vinault2, A Pavon1
1University Hospital Dijon, France; 2CHU de Dijon, Centre d’Investigation
Clinique – Epidémiologie Clinique, Dijon, France
Critical Care 2013, 17(Suppl 2):P71 (doi: 10.1186/cc12009) P71
Impact of time to initiation of appropriate antibiotic therapy on
early mortality of patients with septic shock
JP Quenot1, C Binquet2, S Vinault2, A Pavon1
1University Hospital Dijon, France; 2CHU de Dijon, Centre d’Investigation
Clinique – Epidémiologie Clinique, Dijon, France
Critical Care 2013, 17(Suppl 2):P71 (doi: 10.1186/cc12009) Impact of time to initiation of appropriate antibiotic therapy on
early mortality of patients with septic shock Impact of time to initiation of appropriate antibiotic therapy on
early mortality of patients with septic shock JP Quenot1, C Binquet2, S Vinault2, A Pavon1 Introduction The impact of appropriate antibiotic therapy on
prognosis of patients with septic shock is well established. However,
the prognostic infl uence of time to initiation of antibiotics remains
debated. We evaluated the eff ect on 7-day mortality of time to initiation
of appropriate antibiotic therapy in patients hospitalised in critical care
for septic shock. Conclusion The implications of the apparent diff erences noted
between nonsurvivors and survivors in this survey of cases from the
fi rst known outbreak of injectional anthrax require further study. However, these diff erences might inform the design of research during
future outbreaks or of methods to identify patients most in need of
anthrax-specifi c therapies such as toxin-directed antibodies. Methods Secondary analysis from the EPISS cohort. We included only
patients admitted to the University Hospital Dijon. Septic shock was
defi ned as initiation of vasopressors in a patient with suspected or
documented infection with at least one criterion of hypoperfusion
(metabolic acidosis and/or renal insuffi ciency and/or hepatic dys func-
tion). We excluded patients with no available bacteriological data. Anti-
biotherapy was considered appropriate if at least one of the antibiotics
prescribed was active against the germ identifi ed. Bivariate and
multivariate logistic regression analysis was used to assess the impact of
time to initiation of appropriate antibiotherapy on 7-day mortality. Effi cacy and utility of preemptive anti-Gr(+) coverage in selected
critically ill patients Whether this
approach could lead to vancomycin MIC creep or there could still be
a niche for it later in the course of treatment and/or in nontrauma
patients remains to be further explored. P72 P74
Gram-negative resistance and need for ICU among urinary tract
infections in the United States
M Zilberberg1, A Shorr2
1EviMed Research Group, LLC, Goshen, MA, USA; 2Washington Hospital Center
Washington, DC, USA
Critical Care 2013, 17(Suppl 2):P74 (doi: 10.1186/cc12012) t e p e a e ce o
ect o
t e t e
u t d ug es sta t (
)
o
CPE. We defi ned MDR-PA as any PA isolate resistant to ≥3 drug classes. Enterobacteriaceae were classifi ed as CPE if resistant to both a third-
generation cephalosporin and a carbapenem. We evaluated specimens
from respiratory, bloodstream, urinary tract (UTI) and complicated
intra-abdominal infections. Introduction Urinary tract infection (UTI) can lead to both hospitali-
za tion and severe sepsis. We theorized that UTI due to Gram-
negative (GN) multidrug-resistant P. aeruginosa (MDR-PA), extended-
spectrum β-lactamase (ESBL), E. coli (EC) and Klebsiella sp. (KP), and
carbapenemase-producing Enterobacteriaceae (CPE) would be
frequently isolated in the ICU. Results We identifi ed 327,912 PA (60,695 (18.5%) MDR-PA) and 279,600
Enterobacteriaceae (2,558 (0.9%) CPE) specimens. More than one-
quarter (26.1%) of all PA were recovered from ICU patients as compared
with 17.9% of all Enterobacteriaceae specimens. Of those specimens
originating in the ICU, MDR-PA represented 21.9% of all PA organisms,
while CPE represented 1.6% of all Enterobacteriaceae. Pneumonia and
UTI accounted for 92.4% of all PA and 84.0% of all Enterobacteriaceae
specimens. The proportion of both MDR-PA and CPE was highest in
pneumonia (22.0% and 1.6%, respectively) and lowest in UTI (13.7%
and 0.6%, respectively). Over the time frame of the study, CPEs emerged
and stabilized at approximately 2.8% of all Enterobacteriaceae, while
MDR-PA increased slightly from 16.0% of all PA in 2000 to 17.8% in 2009. Conclusion Although CPE organisms have emerged as an important
pathogen, MDR-PA remains an order of magnitude more prevalent
in the United States. Pneumonia patients and those in the ICU are at
an increased risk for both MDR-PA and CPE infections compared with
those outside the ICU. y
Methods We analyzed a large US-based microbiology database,
Eurofi ns TSN, between the years 2000 and 2009. We determined the
proportion of isolates caused by MDR-PA, ESBL-EC, ESBL-KP, and CPE
relative to their susceptible counterparts. We defi ned MDR-PA as any
isolate resistant to ≥3 drug classes. ESBL organisms were defi ned as
E. coli and K. pneumoniae resistant to a third-generation cephalosporin. Prevalence of multidrug resistance, extensive drug resistance
and pandrug resistance among multiple Gram-negative isolates:
experience in a tertiary-care hospital ICU in North India
S Dewan, T Sahoo, N Chandra, A Varma
Fortis Escorts Heart Institute, New Delhi, India
Critical Care 2013, 17(Suppl 2):P76 (doi: 10.1186/cc12014) Prevalence of multidrug resistance, extensive drug resistance
and pandrug resistance among multiple Gram-negative isolates:
experience in a tertiary-care hospital ICU in North India
S Dewan, T Sahoo, N Chandra, A Varma
Fortis Escorts Heart Institute, New Delhi, India
Critical Care 2013, 17(Suppl 2):P76 (doi: 10.1186/cc12014) Prevalence of multidrug resistance, extensive drug resistance
and pandrug resistance among multiple Gram-negative isolates:
experience in a tertiary-care hospital ICU in North India g
y
g
Conclusion The microbiology of GN UTI hospitalizations has shifted
over the last decade and greater antimicrobial resistance is evident. The
prevalence of MDR-PA, ESBL-EC, ESBL-KP, and CPE is higher in the ICU
than in other hospital locations. Introduction Antibiotic resistance is a major worldwide problem
in the ICU [1]. The situation in developing countries like India is
particularly serious. Since the presence of drug-resistant bacteria in
the environment is a threat for public health, up-to-date information
on local pathogens and the drug sensitivity pattern is very crucial to
treat patients. This study was carried out to evaluate the prevalence
of multidrug resistance (MDR), extensive drug resistance (XDR) and
pandrug resistance (PDR) among multiple Gram-negative isolates in a
medical–surgical ICU in a tertiary care hospital in North India. P75
Multidrug resistance among P. aeruginosa and Enterobacteriaceae
in US hospitals, 2000 to 2009
M Zilberberg1, A Shorr2
1EviMed Research Group, LLC, Goshen, MA, USA; 2Washington Hospital Center,
Washington, DC, USA
Critical Care 2013, 17(Suppl 2):P75 (doi: 10.1186/cc12013)
Introduction Gram-negative resistance remains a major challenge
in the care of critically ill patients. Traditionally, P. aeruginosa (PA) has
represented the most concerning pathogen. However, carbapenemase-
producing Enterobacteriaceae (CPE) have emerged as a challenge, but
h
id
i l
f hi
h
i
l
d
d
Figure 1 (abstract P74). Figure 1 (abstract P74). Figure 1 (abstract P74). Methods We conducted a prospective observational study. All data
were analysed using descriptive statistics. All Gram-negative culture
isolates over a period of 13 months (October 2011 to October 2012)
were included in this study. Isolation and identifi cation were performed
using the bact alert system and VITEK2, respectively. Sensitivities were
determined by Kirby Bauer disc diff usion and broth dilution using
VITEK2-AST cards and interpreted according to Clinical and Laboratory
Standards Institute criteria. P72 Enterobacteriaceae were considered CPE if resistant to both a third-
generation cephalosporin and a carbapenem. We further examined
the evolution of the frequency of resistance among GN UTIs over time. Results We identifi ed 115,201 PA (13.7% MDR-PA), 359,090 EC (5.6%
ESBL), 97,419 KP (12.9% ESBL), and 176,110 Enterobacteriaceae (0.6%
CPE) UTI specimens. The prevalence of resistance rose for each organism
of interest from 2000 through 2009: MDR-PA, 11.6 to 12.3%; ESBL-EC,
3.3 to 8.0%; ESBL-KP, 9.1 to 18.6%; CPE 0 to 2.3%. For each organism
the proportion of resistant pathogens was consistently higher among
ICU specimens than among specimens from other hospital locations,
reaching nearly 20% for MDR-PA (Figure 1). P72 Comparison of survivors and nonsurvivors in 27 confi rmed
injectional anthrax cases from the 2009 outbreak in Scotland
M Booth1, L Donaldson1, C Xizhong2, S Junfeng2, P Eichacker2
1Glasgow Royal Infi rmary, Glasgow, UK; 2National Institutes for Health,
Bethesda, MD, USA
Critical Care 2013, 17(Suppl 2):P72 (doi: 10.1186/cc12010) Critical Care 2013, 17(Suppl 2):P72 (doi: 10.1186/cc12010) ( g
)
j
Conclusion The implementation of this new policy failed to reduce
the degree of organ dysfunction severity and was not associated with
signifi cant survival benefi t. Moreover, even though it did not reach
statistical signifi cance, a second peak of Gr(+) isolates was observed
as a possible complication of the preemptive therapy. Whether this
approach could lead to vancomycin MIC creep or there could still be
a niche for it later in the course of treatment and/or in nontrauma
patients remains to be further explored. Introduction From December 2009 to December 2010, 47 patients in
Scotland presented with confi rmed anthrax infection manifested by
soft tissue disease related to heroin injection. These cases represent
the fi rst known outbreak of a recently recognized form of anthrax,
termed injectional anthrax, which appears to be associated with a high
mortality rate (28% in confi rmed cases from the UK outbreak). While
epidemiologic data from this outbreak have been published, no report
has systematically described fi ndings in patients at presentation or
compared these fi ndings in nonsurvivors and survivors. p
Reference Methods To better describe injectional anthrax, we developed a
questionnaire and sent it to clinicians who had cared for confi rmed Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S27 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P74 Methods We analyzed a large US-based microbiology database,
Eurofi ns TSN, between the years 2000 and 2009. We aimed to describe
the prevalence of infection with either multidrug-resistant (MDR) PA or
CPE. We defi ned MDR-PA as any PA isolate resistant to ≥3 drug classes. Enterobacteriaceae were classifi ed as CPE if resistant to both a third-
generation cephalosporin and a carbapenem. We evaluated specimens
from respiratory, bloodstream, urinary tract (UTI) and complicated
intra-abdominal infections. Risk factors associated with Acinetobacter baumannii septicemia
and its mortality rates in critically ill patients Risk factors associated with Acinetobacter baumannii septicemia
and its mortality rates in critically ill patients
K Kontopoulou, K Tsepanis, I Sgouropoulos, A Triantafyllidou, D Socratous,
P Tassioudis, E Chasou, E Antypa, F Renta, E Antoniadou, K Mandraveli
General Hospital G. Gennimatas, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P77 (doi: 10.1186/cc12015) Results Sixty-three patients were admitted. The mean length of stay
(MLS) was 3 ± 3 days, and the mean age was 25 ± 14 (13 patients
<15 years). Patients were hospitalized for combat-related trauma
(74%), noncombat-related trauma, medical pathologies (10%), and
postoperative care (8%). They were Afghans (92%) or westerners
(8%). Swabs were not realized for eight patients. Forty-three percent
revealed an ESBLB colonization: Escherichia coli (22 patients), Klebsiella
pneumoniae (one patient), Acinetobacter baumanii (one patient). No
patients were colonized with MRSA. Ten patients (16%) were directly
admitted to the ICU, 12 (19%) had been hospitalized before admission,
39 (62%) were transferred after resuscitative and stabilization care
in a level 2 unit. For the two last categories, the MLS (for previous
hospitalization) was respectively 14 ± 28 days and 8 ± 6 hours. Among
patients transferred after care in a level 2 unit, MLS was no diff erent
between colonized and noncolonized patients: 8 ± 7 versus 9 ± 6 hours
(P = 0.5, Mann–Whitney test). y
y
p
K Kontopoulou, K Tsepanis, I Sgouropoulos, A Triantafyllidou, D Socratous,
P Tassioudis, E Chasou, E Antypa, F Renta, E Antoniadou, K Mandraveli
General Hospital G. Gennimatas, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P77 (doi: 10.1186/cc12015) K Kontopoulou, K Tsepanis, I Sgouropoulos, A Triantafyllidou, D Socratous,
P Tassioudis, E Chasou, E Antypa, F Renta, E Antoniadou, K Mandraveli
General Hospital G. Gennimatas, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P77 (doi: 10.1186/cc12015) Introduction Acinetobacter baumannii (A. baum) is a leading cause of
septicemia of patients hospitalized in the ICU with high mortality rates. The aim of our study is to investigate the risk factors associated with
A. baum bacteremia and its mortality rates. y
Methods A total of 937 patients (457 women and 480 men, median
age 59) admitted to the ICU during the period 1 January 2009 to 30
September 2012 were enrolled in our retrospective study. Blood
cultures were obtained from all patients. The identifi cation and the
antimicrobial susceptibility testing were performed by the automated
system VITEK2 (Bio Merieux, France). Prevalence of multidrug resistance, extensive drug resistance
and pandrug resistance among multiple Gram-negative isolates:
experience in a tertiary-care hospital ICU in North India
S Dewan, T Sahoo, N Chandra, A Varma
Fortis Escorts Heart Institute, New Delhi, India
Critical Care 2013, 17(Suppl 2):P76 (doi: 10.1186/cc12014) For the purpose of this study, we used MDR
to denote isolates resistant to representatives three or more classes
of antimicrobial agents, XDR as those resistant to all but one or two
classes and PDR as those resistant to all classes of antimicrobial agents
available [2,3]. Results Out of a total 2,796 culture specimens sent over 13 months,
250 isolates were Gram-negative (8.9%). Among these 250 (n) Gram-
negative isolates, 195 (78%) were extended-spectrum β-lactamase
(ESBL) producers and the remaining 55 (22%) were non-ESBL producers. Among the ESBL producers, PDR, XDR and MDR isolates were 14
(5.6%), 113 (45.2%) and 68 (27.2%), respectively (Figure 1). Among
the XDR-positive organisms, seven (6.1%) organisms were New Delhi
metallo-β-lactamase-1 (NDM-1) producers and fi ve (4.4%) organisms
were NDM-2 producers. Among ESBL-positive isolates, the most
predominant isolate was Klebsiella pneumoniae (29.7%) followed by
Acinetobacter aeruginosa (22.5%) and Escherichia coli (20.5%) (Figure 2). Among non-ESBL-positive isolates, the most predominant isolate was
Escherichia coli (34.5%) followed by Klebsiella pneumoniae (21.8%) and
Pseudomonas aeruginosa (14.5%) (Figure 2). P75
Multidrug resistance among P. aeruginosa and Enterobacteriaceae
in US hospitals, 2000 to 2009
M Zilberberg1, A Shorr2
1EviMed Research Group, LLC, Goshen, MA, USA; 2Washington Hospital Center,
Washington, DC, USA
Critical Care 2013, 17(Suppl 2):P75 (doi: 10.1186/cc12013) Multidrug resistance among P. aeruginosa and Enterobacteriaceae
in US hospitals, 2000 to 2009 Introduction Gram-negative resistance remains a major challenge
in the care of critically ill patients. Traditionally, P. aeruginosa (PA) has
represented the most concerning pathogen. However, carbapenemase-
producing Enterobacteriaceae (CPE) have emerged as a challenge, but
the epidemiology of this pathogen is poorly understood. Conclusion ESBL producers were the most frequently isolated Gram-
negative bacterial isolates in this tertiary-care hospital in north India. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S28 pp
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P76). Prevalence of drug-resistant Gram-negative
isolates. Figure 2 (abstract P76). Prevalence of Gram-negative isolates. P <0.05 from the previous model and those mentioned at recent
medical literature as signifi cantly related with A. baum septicemia and
its mortality. Figure 1 (abstract P76). Prevalence of drug-resistant Gram-negative
isolates. y
Results A total of 101 patients (10.78%) developed bloodstream
infection caused by A. baum and the mortality rate due to A. baum
septicemia was estimated as 49.5% (50/101). Whitman TJ: Infection control challenges related to war wound infections
in the ICU setting. J Trauma 2007, 62:S53. Prevalence of multidrug resistance, extensive drug resistance
and pandrug resistance among multiple Gram-negative isolates:
experience in a tertiary-care hospital ICU in North India
S Dewan, T Sahoo, N Chandra, A Varma
Fortis Escorts Heart Institute, New Delhi, India
Critical Care 2013, 17(Suppl 2):P76 (doi: 10.1186/cc12014) Multiple regression
analysis revealed adjusted mortality >55% (Exp(B) = 2.01, P = 0.013),
MIS (Exp(B) = 1.97, P = 0.017), ABL (Exp(B) = 2.34, P = 0.009), LS >14
days (Exp(B) = 1.34, P = 0.034), MV (Exp(B) = 2.67, P = 0.005) and INV.PR
(Exp(B) = 3.27, P = 0.001) as independent risk factors associated with
A. baum septicemia. p
Conclusion The fact that the probability of A. baum bacteremia
increases in immunocompromised patients refl ects the opportunistic
characteristic of these infections. MV accelerates respiratory A. baum
colonization, which is a risk factor for AB bacteremia. Recent INV. PR increased the incidence of A. baum bacteremia and this result is
probably related to the severe status in patients with central venous
catheter. The administration of carbapenems inhibits the growth of
other more susceptible bacteria, allowing the growth of multidrug-
resistant A. baum. Figure 1 (abstract P76). Prevalence of drug-resistant Gram-negative
isolates. Figure 2 (abstract P76). Prevalence of Gram-negative isolates. P78 Prevalence of colonization by multiresistant bacteria on admission
to the ICU in the French military hospital in Kabul, Afghanistan
JV Schaal, P Pasquier, H Delacour, A Salvadori, A Jarrassier, JR Renner,
SM Mérat
Military Teaching Hospital Bégin, Saint-Mandé, France
Critical Care 2013, 17(Suppl 2):P78 (doi: 10.1186/cc12016) Prevalence of colonization by multiresistant bacteria on admission
to the ICU in the French military hospital in Kabul, Afghanistan Prevalence of colonization by multiresistant bacteria on admission
to the ICU in the French military hospital in Kabul, Afghanistan
JV Schaal, P Pasquier, H Delacour, A Salvadori, A Jarrassier, JR Renner,
SM Mérat
Military Teaching Hospital Bégin, Saint-Mandé, France
Critical Care 2013, 17(Suppl 2):P78 (doi: 10.1186/cc12016) Military Teaching Hospital Bégin, Saint-Mandé, France Military Teaching Hospital Bégin, Saint-Mandé, France
Critical Care 2013, 17(Suppl 2):P78 (doi: 10.1186/cc12016) Figure 2 (abstract P76). Prevalence of Gram-negative isolates. Introduction The French military hospital at the Kaboul International
Airport (KaIA) base provides surgical care for International Force
and Afghan National Army soldiers, and also local patients. The
development of multiresistant bacteria (MRB) nosocomial infections
has raised a major problem complicating the care of combat casualties
[1]. The aim of this study is to assess the prevalence of MRB carriage on
admission to the ICU in this combat support hospital. Introduction The French military hospital at the Kaboul International
Airport (KaIA) base provides surgical care for International Force
and Afghan National Army soldiers, and also local patients. The
development of multiresistant bacteria (MRB) nosocomial infections
has raised a major problem complicating the care of combat casualties
[1]. The aim of this study is to assess the prevalence of MRB carriage on
admission to the ICU in this combat support hospital. Among ESBL producers, XDR organisms were most frequent, followed
by MDR and PDR organisms. Few of the XDR isolates were NDM
producers, which have propensity to spread to other bacteria. In view of
signifi cant prevalence of multidrug resistance amongst Gram-negative
organisms in the ICU, regular surveillance of antibiotic susceptibility
patterns plays a crucial role for setting orders to guide the clinician in
choosing empirical or directed therapy of infected patients. References Methods We used a prospective observation study on patients
admitted to the French military ICU in KaIA over 3 months (July to
September 2012). All hospitalized patients were assessed for the
presence of colonization with MRB: nasal and rectal swabs were
performed to identify, respectively, methicillin-resistant Staphylococcus
aureus (MRSA) and extended-spectrum β-lactamases bacteria (ESBLB). The following data were recorded for each patient on admission:
demographic characteristics, bacteriological results, length of stay,
type of previous hospitalization. References
1. Radji M, et al.:Asian Pac J Trop Biomed 2011, 1:39-42. 2. Falagas ME, et al.: Clin Infect Dis 2008, 46:1121-1122. 3. Souli M, et al.: Euro Surveill 2008, 13:30-40. 1.
Radji M, et al.:Asian Pac J Trop Biomed 2011, 1:39-42.
2.
Falagas ME, et al.: Clin Infect Dis 2008, 46:1121-1122.
3.
Souli M, et al.: Euro Surveill 2008, 13:30-40. References 1. Wu HS, et al.: Clin Microbiol Infect 2012, 18:E373-E376
2. Huang ST, et al.: J Microbiol Immunol Infect 2012, 45:356-362. Risk factors associated with Acinetobacter baumannii septicemia
and its mortality rates in critically ill patients Data collected included
underlying diseases, malignancies and immune suppression (MIS),
prognostic factors (APACHE score, adjusted mortality), age, sex, length
of ICU stay (LS), recent administration of broad-spectrum β-lactam
antibiotics (especially carbapenems; ABL), mechanical ventila-
tion (MV), implementation of invasive procedures (central venous
catheter and urine catheter; INV.PR) and outcome. At fi rst a univariate
statistical model was used with signifi cance level set at P = 0.05. For
the multivariate statistical analysis model we used all variables with Conclusion In this study, prevalence of colonization with ESBLB
at admission is very high, suggesting a high prevalence of MDR
colonization in the local population in Afghanistan. It remains important
to intensify the prevention policy against MRB cross-transmission in
the deployed ICU. p y
Reference p y
Reference Whitman TJ: Infection control challenges related to war wound infections
in the ICU setting. J Trauma 2007, 62:S53. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S29 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (63 patients: 30 received SD and 33 HD), the clinical cure rate and
microbiological eradication percentage were higher when TGC was
used at higher doses (57.6% vs. 33.3%; P = 0.08 and 57.1% vs. 30.4%;
P = 0.1). Table 1 shows multivariate analysis of clinical cure predictors
in the VAP subgroup. (63 patients: 30 received SD and 33 HD), the clinical cure rate and
microbiological eradication percentage were higher when TGC was
used at higher doses (57.6% vs. 33.3%; P = 0.08 and 57.1% vs. 30.4%;
P = 0.1). Table 1 shows multivariate analysis of clinical cure predictors
in the VAP subgroup. P79
Extended-spectrum β-lactamase Providencia stuartii in a general
ICU
P Myrianthefs1, E Evodia2, G Fildissis1, G Baltopoulos1
1Athens University, Athens, Greece; 2Iatriko Kentro Athinon, Athens, Greece
Critical Care 2013, 17(Suppl 2):P79 (doi: 10.1186/cc12017) Table 1 (abstract P80). Predictors of clinical cure in patients with VAP
(multivariate analysis)
OR (95% CI)
P value
HD TGC
5.93 (1.38 to 25.6)
0.02
IIAT
0.23 (0.06 to 0.85)
0.03
SOFA score
0.62 (0.45 to 0.86)
<0.01 Table 1 (abstract P80). Predictors of clinical cure in patients with VAP
(multivariate analysis) Introduction Providencia stuartii, a member of the family of Entero-
bacteriacea, is a Gram-negative pathogen causing colonization and
opportunistic infections in ICU patients. pp
p
Methods We retrospectively recorded the characteristics of P. stuartii
infections in our ICU in a total period of 1 year (six-bed general ICU). Methods We retrospectively recorded the characteristics of P. stuartii
infections in our ICU in a total period of 1 year (six-bed general ICU). Results A total of 116 patients (80 males, 68.9%) were hospitalized in
our ICU of mean age 58.5 ± 1.8, mean ICU stay: 23.2 ± 3.3 days, APACHE
II: 19.3 ± 0.7, SAPS II: 45.7 ± 1.6, SOFA: 7.9 ± 0.4 and mortality: 18.9%. Admission diagnosis was multiple trauma (29.3%), emergency surgery
(37.1%), and medical (33.6%). Of them 21 (18.1%) developed P. Reference 1. Tasina E, et al.: Effi cacy and safety of tigecycline for the treatment of
infectious diseases: a meta-analysis. Lancet Infect Dis 2011, 11:834-844. P81 p y
Reference stuartii
infection that was related to multiple trauma (P = 0.0289), length of ICU
stay 69.8 ± 12.6 (median 51.0) versus 12.6 ± 1.7 (median 6.0) (P <0.0001)
and illness severity APACHE II (21.7 ± 1.3 vs. 17.5 ± 0.8; P = 0.0056), SAPS
II (54.9 ± 2.9 vs. 43.6 ± 1.8; P = 0.0296) and SOFA (10.6 ± 0.5 vs. 7.2 ± 0.4;
P <0.0001). There was no statistically signifi cant diff erence regarding
sex, age or mortality (P = 0.3789). Mean day of fi rst isolation was
31.1 ± 2.1 (median 28.0). The number of isolations per site were – blood:
11 (median day: 25.0), tracheal aspirates: 9 (median: 28.5), catheter tip:
15 (median: 31.0), urine: 6 (median: 32.0), wound: 12 (median: 25.0),
biological fl uids: 4 (median: 29.5), other catheters: 1 (median: 28.0),
middle ear: 2 (median: 32.5), and nose: 1 (median: 25.0). Six patients
had only one site isolation and the remaining 15 had multiple sites
of P. stuartii isolation. We totally recorded 63 isolates of which 82.5%
were second-generation and third-generation cephalosporin-resistant,
80.3% aztreonam-resistant and 81% carbapenem-resistant strains
expressing an extended-spectrum β-lactamase phenotype. All patients
had previously received colistin or meropenem or tygecycline for a
median period of 18.5, 15 and 10.5 days, respectively. Conclusion In critically ill patients, HD TGC use seems to be safe and,
combined with other active antibiotics, may increase the rate of MDR
germ VAP clinical success. IIAT and the severity degree of patients’
clinical condition still remain major determinants of VAP treatment
failure. In vitro effi ciency of Amikacin Inhale, a novel drug-device delivery
system y
N Kadrichu1, S Boc1, K Corkery1, P Challoner2
1Novartis Pharmaceuticals, San Carlos, CA, USA; 2Nektar Therapeutics, San
Francisco, CA, USA
Critical Care 2013, 17(Suppl 2):P81 (doi: 10.1186/cc12019) Introduction Amikacin Inhale (NKTR-061, BAY41-6551) is a drug-
device combination in clinical development for adjunctive treatment
of intubated and mechanically ventilated patients with Gram-negative
pneumonia. The product uses a proprietary vibrating mesh nebulizer
system (PDDS Clinical) with amikacin sulfate formulated for inhalation
(3.2 ml of 125 mg/ml amikacin solution) for a 10-day twice-daily course
of therapy. It is designed for use with two delivery systems: one system
for intubated patients (On-vent; Figure 1), and a second Handheld (HH)
system for patients who are extubated before completing the course
of therapy (Figure 2). We investigated in vitro the amikacin lung dose
delivered by PDDS Clinical. Conclusion P. stuartii infection/colonization may develop in critically
ill patients, especially those with multiple trauma and prolonged
ICU stay, and maybe isolated in blood after the 25th day of ICU stay. Also, infections due to extended-spectrum β-lactamase-producing
multidrug-resistant P. stuartii are an emerging problem. P80
High-dose tigecycline use in severe infections
G De Pascale1, L Montini1, T Spanu2, V Bernini1, A Occhionero1, DL Grieco1,
M Biancone1, P De Santis1, ES Tanzarella1, SL Cutuli1, MA Pennisi1,
MA Antonelli1
1Sacro Cuore Catholic University, Rome, Italy; 2Institute of Microbiology, Sacro
Cuore Catholic University, Rome, Italy
Critical Care 2013, 17(Suppl 2):P80 (doi: 10.1186/cc12018) P80
High-dose tigecycline use in severe infections
G D P
l
1 L M
i i1 T S
2 V B
i i1 A O Methods An estimated lung dose (ELD) for On-vent setting was
measured in vitro after collecting aerosolized amikacin from a fi lter at
the end of an endotracheal tube during ventilation. The ELD for the HH
device was calculated from the fi ne particle fraction (FPF <5 μm) post-
mouthpiece, multiplied by the in vitro delivered dose post-mouthpiece. FPF <5 μm refl ects lung deposition observed during phase 2 clinical
trials [1]. Eighty-one nebulizers with volume median diameter (VMD) Introduction The aim of this study is to describe the clinical and
epidemiological profi le of ICU patients receiving tigecycline (TGC) and
to evaluate the potential benefi ts of TGC higher doses. Figure 1 (abstract P81). On-vent system with brown blinder shell for
clinical trials. i
Methods All patients admitted to our ICU between 1 June 2009 and
31 May 2012 who received TGC were evaluated. Cases were excluded
when infections were not microbiologically confi rmed.i i
Results Over the study period, 100 patients fulfi lled the inclusion
criteria: 54 in the SD group (50 mg every 12 hours) and 46 in the HD
group (100 mg every 12 hours). The SD group and the HD group were
not signifi cantly diff erent in terms of age, severity of disease, duration
of TGC therapy, rate of concomitant other active antibiotic use and
of inadequate empirical antimicrobial therapy (IIAT) (P = NS). MDR
A. baumannii and K. pneumoniae were the main pathogens isolated. The percentage of germs other than A. baumannii and K. pneumoniae
was higher in the SD TGC group (P <0.01). Otherwise infections due to
less susceptible germs (TGC MIC value ≥1 μg/ml) were mainly treated
with TGC higher doses (P <0.01). No signifi cant diff erences were found
in terms of ICU mortality (P = 0.8). The rate of abnormal laboratory
measures during TGC treatment was similar between the two groups
(P = NS). No patients required TGC discontinuation or dose reduction
because of suspected adverse events. In the VAP subpopulation Figure 1 (abstract P81). On-vent system with brown blinder shell for
clinical trials. Critical Care 2013, Volume 17 Suppl 2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S30 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 parameters infl uencing the relation between amikacin dose and peak
concentration. Figure 2 (abstract P81). Handheld system with brown blinder shell for
clinical trials. Cost implication of antibiotic-associated diarrhoea and the fi nancial
impact of probiotic use in its prevention p
p
p
C Ebm, M Cecconi, C Moran, A Rhodes, T Rahman C Ebm, M Cecconi, C Moran, A Rhodes,
St Georges Healthcare Trust, London, UK St Georges Healthcare Trust, London, UK Reference g
Critical Care 2013, 17(Suppl 2):P83 (doi: 10.1186/cc12021) 1. Fink J, et al.: High in vivo amikacin lung deposition after NKTR-061 dosing
correlates with in vitro aerosol characterization. Am J Respir Crit Care Med
2008, 177:A530. Introduction Antibiotic-associated diarrhoea (AAD) occurs in as many
as 30% of patients receiving antibiotics, often leading to increased
morbidity, prolonged in-hospital stay and additional healthcare
resource utilisation. Age, antibiotics and prolonged postoperative ward
and ICU stay have been suggested to be independent risk factors. In
such patient populations, probiotics may be used to prevent antibiotic-
associated diarrhoea, yet they are not routinely recommended as a
component of perioperative care. The aim of this study was to model
the long-term costs associated with AAD and to assess the eff ectiveness
of probiotics as a preventive strategy. P80
High-dose tigecycline use in severe infections
G D P
l
1 L M
i i1 T S
2 V B
i i1 A O Methods In this retrospective study we looked at 47 patients (128 peak
levels) between 2003 and 2012. Multivariate linear regression analysis
was done for several parameters: administered dose calculated with
total body weight, ideal body weight, adjusted body weight, type of
intensive care patient, BMI, daily fl uid balance, SOFA score and APACHE
score, and patient characteristics were analyzed. Results A linear correlation between dose and amikacin peak level was
confi rmed (Figure 1). A total 54.69% of all amikacin administrations did
not result in a therapeutic peak level. The multivariate linear regression
analysis showed the best linear correlation with adjusted body weight
and SOFA score. The comparison of variables between four patient
groups, based on the deviation between measured peak level and
predicted peak level (according the linear correlation), showed new
variables that may infl uence peak level. yl
Conclusion This confi rms that low doses (<18 mg/kg) of amikacin in
intensive care patients seldom result in a therapeutic peak level. The
proposed loading dose of 25 mg/kg is good for reaching a therapeutic
level, although 29.6% remains subtherapeutic. Due to the linear
correlation, more therapeutic levels may be reached with higher doses
(25 to 30 mg/kg). New variables need further investigation to explain
the high variability in achieved peak level. Figure 2 (abstract P81). Handheld system with brown blinder shell for
clinical trials. of 4.4 ± 0.5 μm and output rates of 0.23 ± 0.10 ml/minute were tested
for each system. Delivered dose data were fi t to the independent
variables (that is, VMD and output rates) using a least-squares fi t with
95% confi dence limits. P83 y y
Conclusion The results support the use of the PDDS Clinical with either
system to administer aerosolized amikacin with high effi ciency and no
dose adjustment is required when switching from the On-vent to the
HH system for extubated patients. References References
1. Delattre IK, et al.: Ther Drug Monit 2010, 32:749-756. 2. Taccone FS, et al.: Crit Care 2010, 14:R53. 3. Gálvez R, et al.: Int J Antimicrobial Agents 2011, 38:146-151. 4. Avent ML, et al.: Internal Med J 2011, 41:441-449. 5. Fernández de Gatta MM, et al.: J Clin Pharm Ther 1996, 21:417-421. 1. Delattre IK, et al.: Ther Drug Monit 2010, 32:749-756. 2. Taccone FS, et al.: Crit Care 2010, 14:R53. 3. Gálvez R, et al.: Int J Antimicrobial Agents 2011, 38:146-151. 4. Avent ML, et al.: Internal Med J 2011, 41:441-449. 5. Fernández de Gatta MM, et al.: J Clin Pharm Ther 1996, 21:417-421. 1. Delattre IK, et al.: Ther Drug Monit 2010, 32:749-756. 2. Taccone FS, et al.: Crit Care 2010, 14:R53. 2. Taccone FS, et al.: Crit Care 2010, 14:R53. 3. Gálvez R, et al.: Int J Antimicrobial Agents 2011, 38:146-151. i
Results Total percentage recoveries for On-vent and HH test runs were
between 85% and 115% of the nominal dose. The mean ELDs were
50 ± 9% (On-vent) and 49 ± 11% (HH) of the nominal dose. Nebulizers
with longer dosing times and lower VMDs had higher ELD values for
both delivery systems. 4. Avent ML, et al.: Internal Med J 2011, 41:441-449. 4. Avent ML, et al.: Internal Med J 2011, 41:441-449. 5
Fernández de Gatta MM et al : J Clin Pharm Ther 1996 21:417 42 5. Fernández de Gatta MM, et al.: J Clin Pharm Ther 1996, 21:417-42 Empirical versus preemptive antimycotic therapy in terms of
outcome benefi t
l
d
l Figure 1 (abstract P85). Cologne antifungal strategy. Figure 1 (abstract P85). Cologne antifungal strategy. Introduction Novel treatment strategies for invasive candidiasis (IC)
are constantly emerging. Nevertheless, diffi culties in diagnosis pose
a challenge on their reliability, effi cacy and safety. We have previously
developed and approbated in our ICU an algorithm for empirical
antimycotic therapy, combining the most signifi cant risk factors for
IC with three major clinical criteria for persistent nonbacterial sepsis
[1]. On the other hand, preemptive therapy, based on identifi cation of
mycotic antigens and/or anti-mycotic antibodies in serum, is regarded
as more reliable, even though it is known for its low sensitivity. The
aim of the current study was to compare and evaluate the possible
outcome benefi t of our protocol implementation versus detection of
galactomanan in patient’s serum as a trigger for antimycotic treatment
initiation. Results Mean age was 67.4 years with 64% male patients. Most
patients had combined CABG and valve procedure (n = 20), other
groups were HTX and LTX (n = 4), assist therapy (n = 4), TAVI (n = 3)
and other procedures. Mean predicted mortality using the logarithmic
CASUS score at the onset of IFI was 59%. C. albicans was isolated in 73%,
C. glabrata in 21%. Length of antifungal treatment using micafungin in
30 cases was 14 ± 5 days. Eradication of yeast was successful in 79%
but mortality of all patients remains high at 36.8% but was lower than
predicted in the CASUS score. Mortality was not yeast related. Methods A randomized prospective controlled trial was carried out
from September 2010 to September 2012. After the implication
of the inclusion and exclusion criteria, patients were submitted to
block randomization and stratifi ed on the basis of their initial SAPS
II exp score. Antimycotic therapy was started on the day of inclusion
in the control group and only with positive galactomanan serum
test in the preemptive therapy group. Initial data were gathered on
demographics, proven risk factors for IC-related mortality, severity of
infl ammatory response and organ dysfunction. Dynamics of SIRS and
SOFA values, Candida colonization index, ventilator-free days, length of
ICU stay and outcome were followed for each patient.i p
y
y
Conclusion Our described treatment approach shows encouraging
results for the treatment of IFI especially in high-risk cardiothoracic
patients. Results A total of 106 patients were enrolled. Empirical versus preemptive antimycotic therapy in terms of
outcome benefi t
l
d
l No statistically signifi cant
diff erences in their basal characteristics were found. The subsequent
SIRS and SOFA scores showed fi rm dynamics in the control group,
although the new organ dysfunction severity was insignifi cantly lower. The length of ICU stay and the number of ventilator-free days were
comparable. The in-hospital mortality was 47.1% in the preemptive
therapy group versus 31.3% in the control group (P = 0.94). A total of
seven adverse reactions were observed among treated patients, yet
not associated with higher mortality risk. Analysis of parameters in critically ill patients infl uencing amikacin
peak levels p
M Hollevoet1, SJ Toye2, J De Waele1 p
IM Hollevoet1, SJ Toye2, J De Waele1 Introduction Recent studies demonstrate that a loading dose
of 25 mg/kg (total body weight) of amikacin in septic patients is
required to reach a suffi cient peak concentration. This study examines Methods We developed a simulation model to determine clinical costs
and outcomes attributable to AAD. To assess the cost-eff ectiveness of
probiotics, as part of a perioperative regime, we constructed a decision Figure 1 (abstract P82). Correlation between dose of amikacin and peak concentration. Figure 1 (abstract P82). Correlation between dose of amikacin and peak concentration. S31 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 tree. The model observes long-term costs and outcomes of probiotics
as compared with conventional therapy, from a societal perspective. Input parameters, extracted from meta-analysis, clinical trials and
national databases, include incidence numbers, costs and quality-
adjusted health states for the remaining life (QALYs). Outcomes assessed
were overall costs attributable to ADD and the cost-eff ectiveness of
probiotics, described as costs/QALY. References . Eggiman et al.: Invasive Candida infections in the ICU. Mycoses 20 1. Eggiman et al.: Invasive Candida infections in the ICU. Mycoses 2012,
55(Suppl):65-72. 55(Suppl):65-72. 2. Andes et al.: Impact of treatment strategy on outcomes in patients with
candidemia and other forms of invasive candidiasis: a patient-level
quantitative review of randomized trials. Clin Infect Dis 2012, 54:1110-1122. P86 Conclusion The choice of empirical versus preemptive therapy led to
earlier and more stable reduction in the degree of organ dysfunction
severity. It showed to be at least not inferior if not equal; in terms of
survival benefi t and expediency of treatment. Moreover, galactomanan
detection fails to guide the choice of the individual antimycotic, based
on the expected Candida spp. Impact of echinocandins as fi rst-choice therapy in cardiothoracic
high-risk patients with candidemia Impact of echinocandins as fi rst-choice therapy in cardiothoracic
high-risk patients with candidemia g
p
G Langebartels, Y Choi, J Catteleans, T Wahlers
University of Cologne, Germany
Critical Care 2013, 17(Suppl 2):P85 (doi: 10.1186/cc12023) g
p
G Langebartels, Y Choi, J Catteleans, T Wahlers
University of Cologne, Germany
Critical Care 2013, 17(Suppl 2):P85 (doi: 10.1186/cc12023) Introduction Invasive candidemia is a major cause of increased
mortality among ICU patients. Antifungal agents like liposomale
amphotericin B and azoles could not accomplish the claim to be fi rst
choice in the treatment of invasive fungal infection (IFI) because of side
eff ects and eff ectiveness. Especially, cardiothoracic surgery patients
as a group of high-risk patients are in a focus for new strategies
and agents. A new class of antimycotic agents, the echinocandins,
with a low profi le of side eff ects, low interactive potential and high
eff ectiveness in the treatment of candidemia, is a powerful option in
the treatment of IFI. We report our single-center experience with a
modifi ed clinical treatment approach based on clinical score of Leon
and using echinocandins as fi rst-line therapy for proven and suspected
fungal infection. Results Our results indicate an estimated incremental lifetime cost of
£13,272.53 per ADD patient, largely driven by increased ICU length of
stay and readmission rates. The addition of probiotics to the standard
perioperative regime is associated with a small survival benefi t of
1.2 months, yet a cost reduction of £917.3/ADD patient. The main cost
was increased duration of ICU stay and readmissions, which contribute
to 85% of total expenses. p
Conclusion AAD is associated with a signifi cant increase in costs from a
societal perspective. The provision of probiotics can achieve substantial
cost savings and can be recommended as a cost-eff ective regime in the
perioperative setting. Preventing ADD off ers a potentially signifi cant
reduction of in-hospital costs and resource expenditures. g
Methods From May 2011 to October 2012, 2,844 patients were
treated on our cardiothoracic ICU. We evaluated 37 cardiothoracic
postoperative patients with proven or suspected IFI or prophylaxis
(Figure 1). The records were evaluated for cardiothoracic procedures,
microbiological and yeast date, cardiothoracic surgery score (CASUS),
ICU and clinical data. P84
Empirical versus preemptive antimycotic therapy in terms of
outcome benefi t
S Milanov, VT Todorova, G Georgiev, M Milanov
Emergency Hospital ‘Pirogov’, Sofi a, Bulgaria
Critical Care 2013, 17(Suppl 2):P84 (doi: 10.1186/cc12022) Cost-eff ectiveness analysis of anidulafungin in the treatment of
candidaemia g
Results A total of 2,426 patients were admitted to Heart of England
NHS Foundation Trusts ICUs during the study period. Of these, 31
patients were identifi ed as having an IFI (1.3%). Baseline demographic
data were similar between groups. Patients with IFI had signifi cantly
longer ICU length of stay (19 days vs. 5 days, P <0.0001) and required
more days of advanced organ support (12 days vs. 3 days, P <0.0001). A trend towards higher hospital mortality (41% vs. 27%, P = 0.08) was
observed. Candida albicans was the predominant organism cultured
(64%), followed by Candida glabrata (23%), with other Candida species
accounting for the remaining 13% of IFIs. Sites of IFIs were blood (45%),
intra-abdominal (39%), and pleural (16%). Most patients (52%) who
acquired an IFI had had intra-abdominal surgery prior to ICU admission. The majority of patients (71%) were treated with echinocandins, whilst
of the nine patients who were initially treated with fl ucanazole, six
(67%) required therapy escalation to an echinocandin. G Auzinger1, G Playford2, C Graham3, H Narula3, C Charbonneau4,
D Weinstein4, M Kantecki4, H Schlamm5, M Ruhnke6
1King’s College Hospital, London, UK; 2Princess Alexandra Hospital, Brisbane,
Australia; 3RTI Health Solutions, Durham, NC, USA; 4Pfi zer International
Operations, Paris, France; 5Pfi zer Inc., New York, NY, USA; 6Charité University
Medicine, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P87 (doi: 10.1186/cc12025) G Auzinger1, G Playford2, C Graham3, H Narula3, C Charbonnea
D Weinstein4, M Kantecki4, H Schlamm5, M Ruhnke6 1King’s College Hospital, London, UK; 2Princess Alexandra Hospital, Brisbane,
Australia; 3RTI Health Solutions, Durham, NC, USA; 4Pfi zer International
Operations, Paris, France; 5Pfi zer Inc., New York, NY, USA; 6Charité University
Medicine, Berlin, Germany y
Critical Care 2013, 17(Suppl 2):P87 (doi: 10.1186/cc12025) Critical Care 2013, 17(Suppl 2):P87 (doi: 10.1186/cc12025) Introduction Echinocandins are recommended fi rst-line treatment
for candidaemia [1]. A cost-eff ectiveness model developed from a UK
perspective examined costs and outcomes of antifungal treatment
for candidaemia and other forms of invasive candidiasis based on
European clinical guidelines [1]. Methods Costs and treatment outcomes with the echinocandin
anidula fungin were compared with caspofungin, micafungin,
fl uconazole, voriconazole and amphotericin B. The model included
non-neutropenic patients aged ≥16 years with confi rmed candidaemia/
another form of invasive candidiasis receiving intravenous fi rst-line
treatment [2]. Fungal ventricular assist device infections occur in colonized
patients and are associated with high mortality rate Fungal colonization (FC), infection and outcome
during VAD support
n
FC
Mortality
No IFI
24
12
9
IFI
10
9
8
P value
0.029
0.024
Conclusion In our center, we observed a high incidence of IFI in IC
patients with VAD that was associated with a mortality rate of 80%
Screening of fungal colonization appears to be very important durin
the ICU stay for VAD patients. Trials are needed for investigating th
use, the drug choice and the timing of antifungal prophylaxis for suc
high-risk patients. Reference
1. Aslam S, et al.: Clin Infect Dis 2010, 50:664-671. P87 were all-cause 6-week mortality, cost of treatment-related adverse
events (AEs) and other medical resource use costs. Life-years were
calculated using a published model [4]. Antifungal agent-related AEs
were taken from the product label/literature. Resource use was derived
from the literature and discussion with clinical experts. Drug acquisition/
administration costs were taken from standard UK costing sources. with fungi [1]. The relationship between colonization and invasive
fungal infection (IFI) in severely ill ICU patients with a VAD support is
not described. This study analyzes the incidence and outcome of fungal
infection and colonization in VAD patients in bridge to transplantation
or in destination therapy. py
Methods We conducted a retrospective review of all VAD implantations
in our surgical ICU between 2007 and 2012. The incidence of fungal
colonization, antifungal prophylaxis, bacterial sepsis and the mortality
of IFI versus no IFI patients were compared. Results First-line anidulafungin for treatment of candidaemia was
cost-eff ective per life-year gained versus fl uconazole (incremental
cost-eff ectiveness ratio £813). Anidulafungin was cost saving versus
caspofungin and micafungin in terms of life-years gained due to
lower ICU costs and a higher rate of survival combined with a higher
probability of clinical success.f Results In the study period, 34 patients with severe heart failure
or cardiogenic shock were selected for a VAD implantation (nine
in destination therapy). The overall mortality rate was 50% during
mechanical assistance. Confi rmed (n = 8) and highly suspected (n = 2)
IFI occurred during the ICU stay in 29% of patients who were treated
with echinocandins, voriconazole and/or liposomal amphotericin B. The
isolated fungi were: six Candida albicans, two parapsilosis, one glabrata
and one invasive pulmonary aspergillosis. Antifungal prophylaxis with
fl uconazole was administered to 18% of patients at mean for 5 days
mainly in the more recent implantations. Allou N, et al.: Curr Infect Dis Rep 2011, 13:426-432.
Zaragosa R, et al.: Adv Sepsis 2008, 6:90-98. Fungal ventricular assist device infections occur in colonized
patients and are associated with high mortality rate In the no IFI population,
54% (n = 13) had a systemic or VAD bacterial sepsis with a mortality
rate about 54%. The mortality without any sepsis was reduced to 18%. Fungal colonization was signifi cantly more present (90% vs. 50%)
before IFI in VAD patients. The mortality rate was dramatically higher
with IFI (80% vs. 38%) in accordance with the literature [1]. See Table 1. y
Conclusion
Anidulafungin
was
cost-eff ective
compared
with
fl uconazole for treatment of candidaemia and was cost saving versus
other echinocandins in the UK. European guidelines recommend
echinocandins as fi rst-line treatments for candidaemia [1]; this
model indicates that anidulafungin marries clinical eff ectiveness and
cost-eff ectiveness. f
References 1. Cornely OA: Clin Microbiol Infect 2012, 18:19-37. 2. Reboli AC, et al.: N Engl J Med 2007, 356:2472-2482. 3. Mills EJ, et al.: Ann Clin Microbiol Antimicrob 2009, 8:23. 4. Sidhu MK, et al.: Curr Med Res Opin 2009, 25:2049-2059 2. Reboli AC, et al.: N Engl J Med 2007, 356:2472 2482. 3. Mills EJ, et al.: Ann Clin Microbiol Antimicrob 2009, 8:23. 4. Sidhu MK, et al.: Curr Med Res Opin 2009, 25:2049-2059 Table 1 (abstract P86). Fungal colonization (FC), infection and outcome
during VAD support Table 1 (abstract P86). Fungal colonization (FC), infection and outcome
during VAD support
n
FC
Mortality
No IFI
24
12
9
IFI
10
9
8
P value
0.029
0.024 Reference 1. Aslam S, et al.: Clin Infect Dis 2010, 50:664-671. Cost-eff ectiveness analysis of anidulafungin in the treatment of
candidaemia Patients were categorised as a clinical success or failure
(patients with persistent/breakthrough infection); frequency data for
each outcome were taken from a mixed-treatment comparison [3]. Successfully treated patients switched to oral therapy. Clinical failures
switched to a diff erent antifungal class. It was assumed that second-line
treatment duration was equivalent to that of fi rst-line treatment and
only two lines of therapy were required to treat infection. Other inputs Conclusion The results of our study are consistent with other published
data, in that whilst IFI prevalence is low, they are associated with
increased morbidity in critically ill patients. This study has led to a
change in hospital policy regarding antifungal use in the ICU, with
echinocandins being fi rst-line in the pre-emptive treatment of IFI. We
keenly await the results of the FIRE study, which will provide important
insights to identifi cation of patients at risk of IFIs and optimal drug
therapy. Prevalence and impact of invasive fungal infections in intensive care
JM Patel K Couper T Melody R O’Brien D Parekh Introduction Invasive fungal infections (IFI) aff ect 1% of ICU patients
and are increasing in incidence. IFIs are associated with a poor
prognosis, which is further complicated by diffi culties in identifi cation
of fungal organisms by traditional culture methods and the emergence
of Candida species resistant to triazole therapy [1,2]. This study aimed to
assess the prevalence of IFIs, the organisms responsible and outcomes
of patients aff ected. Conclusion In our center, we observed a high incidence of IFI in ICU
patients with VAD that was associated with a mortality rate of 80%. Screening of fungal colonization appears to be very important during
the ICU stay for VAD patients. Trials are needed for investigating the
use, the drug choice and the timing of antifungal prophylaxis for such
high-risk patients. R f f
Methods Patients admitted to the Heart of England NHS Foundation
Trust ICUs who acquired an IFI were identifi ed through the hospital
Fungal Infection Risk Evaluation (FIRE) study database. All ICU patients
admitted between October 2009 and March 2011 were used as a
comparative cohort. Data collected included: baseline demographics,
length of stay, ICU and hospital mortality, and nature of IFI. Data were
analysed using Student’s t test for continuous data and Fischer’s exact
test for categorical data. Fungal ventricular assist device infections occur in colonized
patients and are associated with high mortality rate P Gaudard, J Eliet, N Zeroual, G Culas, P Colson CHRU Montpellier, France Critical Care 2013, 17(Suppl 2):P86 (doi: 10.1186/cc12024) Referencefi Introduction Infection is a common complication of the ventricular
assist device (VAD) and is associated with poor outcome especially 1. S Milanov, et al.: Effi cacy and utility of a protocol for pre-emptive
antimycotic therapy. Crit Care 2010, 14(Suppl 1):P70. 1. S Milanov, et al.: Effi cacy and utility of a protocol for pre-emptive
antimycotic therapy. Crit Care 2010, 14(Suppl 1):P70. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S32 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 with fungi [1]. The relationship between colonization and invasiv
fungal infection (IFI) in severely ill ICU patients with a VAD support
not described. This study analyzes the incidence and outcome of fung
infection and colonization in VAD patients in bridge to transplantatio
or in destination therapy. Methods We conducted a retrospective review of all VAD implantation
in our surgical ICU between 2007 and 2012. The incidence of fung
colonization, antifungal prophylaxis, bacterial sepsis and the mortali
of IFI versus no IFI patients were compared. Results In the study period, 34 patients with severe heart failu
or cardiogenic shock were selected for a VAD implantation (nin
in destination therapy). The overall mortality rate was 50% durin
mechanical assistance. Confi rmed (n = 8) and highly suspected (n =
IFI occurred during the ICU stay in 29% of patients who were treate
with echinocandins, voriconazole and/or liposomal amphotericin B. Th
isolated fungi were: six Candida albicans, two parapsilosis, one glabra
and one invasive pulmonary aspergillosis. Antifungal prophylaxis wi
fl uconazole was administered to 18% of patients at mean for 5 da
mainly in the more recent implantations. In the no IFI populatio
54% (n = 13) had a systemic or VAD bacterial sepsis with a mortali
rate about 54%. The mortality without any sepsis was reduced to 18%
Fungal colonization was signifi cantly more present (90% vs. 50%
before IFI in VAD patients. The mortality rate was dramatically high
with IFI (80% vs. 38%) in accordance with the literature [1]. See Table
Table 1 (abstract P86). References Allou N, et al.: Curr Infect Dis Rep 2011, 13:426-432. Zaragosa R, et al.: Adv Sepsis 2008, 6:90-98. S33 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P89
Beliefs and actual practice of oxygen therapy in the ICU
HJ Helmerhorst1, MJ Schultz2, PH Van der Voort3, E De Jonge1,
DJ Van Westerloo1
1Leiden University Medical Center, Leiden, the Netherlands; 2Academic
Medical Center, Amsterdam, the Netherlands; 3Onze Lieve Vrouwe Gasthuis,
Amsterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P89 (doi: 10.1186/cc12027) P90
FiO2/PEEP index: a simple tool for opitimizing ventilator settings
D Trasy, M Nemeth, K Kiss, Z Till, Z Molnar
University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P90 (doi: 10.1186/cc12028) Introduction During mechanical ventilation, oxygenation can be
infl uenced by adjusting FiO2 and positive end-expiratory pressure
(PEEP). There have been recommendations for how the FiO2 and
PEEP should be set [1]. However, in a recent audit we found that the
compliance of doctors of these recommendations is very low [2]. Therefore we invented a simple parameter called the FiO2/PEEP index
(FPi) of which the physiologic value is ≤7 (that is, FiO2 = 21%/PEEP =
3 cmH2O), which corresponds to the ARDSNet trial’s minimum FiO2/
PEEP settings: 35%/5 cmH2O [2]. The aim of this case–control study
was to investigate the impact of an FPi ≤7 targeted protocol on clinical
practice. Introduction The aim of this study was to compare self-reported beliefs
with actual clinical practice of oxygen therapy in the ICU. Hyperoxia is
frequently encountered in ventilated patients and prolonged exposure
has repeatedly been shown to induce lung injury and (systemic)
toxicity. y
Methods An online questionnaire for ICU clinicians was conducted
to investigate beliefs and motives regarding oxygen therapy for
critically ill patients. Furthermore, arterial blood gas (ABG) samples
and corresponding ventilator settings were retrieved to retrospectively
assess objective oxygenation between 1 April 2011 and 31 March 2012
in the ICUs of three teaching hospitals in the Netherlands. p
Methods A prospective observational study in 2010 and 2012. Every
mechanically ventilated patient was recruited. Demographics, outcome
data and Lung Injury Score (LIS) were recorded after admission. Respiratory settings, oxygenation and ventilation parameters were
recorded after mechanical ventilation was commenced and the fi rst
arterial blood gas sample was taken (T0). Measurements were repeated
in 24 hours (T1). Data are presented as mean ± SD, paired-sample and
independent-sample t test and chi-square tests were used for statistics. References Results In 2010, 75 patients, and in 2012, 130 patients were included. There was no diff erence in demographics, disease severity, but LIS
was higher in 2012: 1.34 ± 1.13 versus 0.84 ± 0.98, P = 0.001. There
was no signifi cant diff erence in FPi between the two groups at T0:
10.91 ± 4.25 versus 10.26 ± 5.01 (2012 vs. 2010, respectively). At T24 the
FPi was signifi cantly lower in 2012 as compared with 2010: 7.28 ± 2.58
versus 8.17 ± 3.3, P = 0.001; which was due to the higher PEEP applied:
7.08 ± 2.87 versus 6.63 ± 2.91, P = 0.014. Although in 2012 signifi cantly
more patients, 112 (86%), were ventilated with FiO2 ≥50% at T0 as
compared with 2010, 44 (58%) (P = 0.001), by T24 signifi cantly less
patients received FiO2 ≥50%, 46 (35%) vs. 34 (45%) (P = 0.011). There
was no signifi cant diff erence between the two groups regarding FiO2,
PaO2 and PaCO2 at T0 and T24.i Results Analyzable questionnaire responses were received from 200
ICU physicians and nurses. The majority of respondents believed
that oxygen-induced lung injury is a concern, although barotrauma
and volutrauma are generally considered to impose a greater risk in
mechanical ventilation. Frequently allowed minimal saturation ranges
in the questionnaire were 85 to 95% and 7 to 10 kPa (Figure 1). Self-
reported FiO2 adjustment in hypothetical patient cases with variable
saturation levels was moderately impacted by the underlying clinical
condition. To study actual clinical practice, a total of 107,888 ABG
samples with corresponding ventilator settings, covering 5,565 patient
admissions, were retrieved. Analysis showed a median (IQR) PaO2 of
11.7 kPa (9.9 to 14.3), median FiO2 was 0.4 (0.4 to 0.5), median PEEP was
5 (5 to 8). A total 63.5% of all PaO2 registries were higher than previously
suggested oxygenation goals (7.3 to 10.7 kPa) [1]. In 56.8% of cases
with PaO2 higher than the target range, neither FiO2 nor PEEP levels
had been lowered when the next ABG sample was taken. Figure 1 (abstract P89). Conclusion Most clinicians acknowledge the detrimental eff ects of
prolonged exposure to hyperoxia in the ICU and report a low tolerance
for high saturation levels. 1.
Brower RG, et al.: N Engl J Med 2004, 351:327-336. References However, the self-reported intention for
conservative oxygen therapy is not consistently expressed in our
objective data of actual clinical practice and a large proportion of
patients was exposed to high and potentially toxic oxygen levels. Reference
1. Brower RG, et al.: N Engl J Med 2004, 351:327-336. Figure 1 (abstract P89). 2
2
Conclusion Implementing an FPi ≤7-based algorithm signifi cantly
reduced the FiO2 and increased the PEEP applied in mechanically
ventilated within the fi rst 24 hours. Whether this has any impact on
earlier weaning due to reaching the weaning criteria of FiO2 sooner,
and as a result shortening the duration of mechanical ventilation, has
to be investigated in the future. g
References References
1. ARDSNet: N Engl J Med 2000, 342:1301. 2. Kiss K, et al.: Intensive Care Med 2011, 37(Suppl 1):S195. 1. ARDSNet: N Engl J Med 2000, 342:1301. 1. ARDSNet: N Engl J Med 2000, 342:1301. 2. Kiss K, et al.: Intensive Care Med 2011, 37(Suppl 1):S195. g
,
2. Kiss K, et al.: Intensive Care Med 2011, 37(Suppl 1):S195. 2. Kiss K, et al.: Intensive Care Med 2011, 37(Suppl 1):S195. P92 9
Relation between PaO2/FiO2 ratio, SpO2/FiO2 ratio, oxygenation
index and ventilation ratio in critically ill patients
T Aslanidis, A Myrou, E Chytas, E Anastasiou, E Geka, E Efthimiou,
V Ourailoglou, I Soultati, S Primikiri, M Giannakou-Peftoulidou
A.H.E.P.A. University Hospital, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P92 (doi: 10.1186/cc12030) Relation between PaO2/FiO2 ratio, SpO2/FiO2 ratio, oxygenation
index and ventilation ratio in critically ill patients y
A Batchinsky1, C Necsoiu1, T Langer2, V Vecchi2, W Baker1, J Salinas1,
L Cancio1
1US Army Institute of Surgical Research, Fort Sam Houston, TX, USA;
2University of Milan, Italy
Critical Care 2013, 17(Suppl 2):P93 (doi: 10.1186/cc12031) A Batchinsky1, C Necsoiu1, T Langer2, V Vecchi2, W Baker1, J Salinas1,
L Cancio1
1US Army Institute of Surgical Research, Fort Sam Houston, TX, USA;
2University of Milan, Italy
Critical Care 2013, 17(Suppl 2):P93 (doi: 10.1186/cc12031) Introduction Respiratory complexity (RC) as assessed by sample
entropy (SampEn) is lower in patients failing spontaneous breathing
trials. We evaluated the role of RC in monitoring subjects with severe
acute respiratory distress syndrome (ARDS) treated with the Cardiohelp
(Maquet, Rastatt, Germany) extracorporeal life-support system (ECLS). We hypothesized that RC is reduced during apnea. Introduction Many authors have proposed less invasive ways to
measure oxygenation in patients with ALI. The aim of the present study
is to compare the already popular oxygenation index (OI) and PaO2/
FiO2 ratio (PFr) with other recently proposed indices such as SpO2/FiO2
ratio (SFr) and ventilation ratio (VR) in a mixed ICU population. Methods During a 6-month prospective observational study carried
out in a polyvalent 10-bed adult ICU, ABGs were obtained from 145
patients. Two independent measurements were taken from each
patient under the same mode of ventilation (SIMVPSV). PFr, SFr, VR
and OI were calculated. Demographic data (APACHE II score, age, sex, yp
g p
Methods Six sheep sedated with midazolam were connected to the
Cardiohelp via a 23F Avalon catheter. Blood fl ow was ~2 l/minute
and FiO2 was 0.5. Sheep were on CPAP of 8 cmH2O with FiO2 of 1 via
tracheostomy. P91
Use of high-fl ow nasal canulae: eff ect on alveolar pressure and its
limitation limitation
H Hayami, K Mizutani, M Shioda, S Takaki, H Maejima, K Ueno,
Y Yamaguchi, T Kariya, T Gotoh
Yokohama City University Hospital, Yokohama, Japan
Critical Care 2013, 17(Suppl 2):P91 (doi: 10.1186/cc12029) Figure 1 (abstract P89). Introduction High-fl ow nasal canulae (HFNC) deliver high-fl ow
humidifi ed gas at up to 60 l/minute. There are two types of respiratory
circuit to generate mix gas fl ow, Blender type (typeB) and Venturi type
(typeV). The therapy is well established in the pediatric population
and HFNC use has been described in the adult population. It has been
reported that HFNC provide higher FIO2 compared with low-fl ow
canulae, and also create mild positive pharyngeal airway pressure, but
the eff ect on alveolar pressure is unknown. We aimed to investigate
the eff ect of HFNC on alveolar pressure, by measuring intratracheal
pressure in patients with a cricothyrotomy catheter (CTC). At the same
time, we measured the actual gas fl ow rate (AGFR) by fl owmeter and
compared it with assumed fl ow. Conclusion Most clinicians acknowledge the detrimental eff ects of
prolonged exposure to hyperoxia in the ICU and report a low tolerance
for high saturation levels. However, the self-reported intention for
conservative oxygen therapy is not consistently expressed in our
objective data of actual clinical practice and a large proportion of
patients was exposed to high and potentially toxic oxygen levels. Reference l
Methods Seven patients with a CTC were participated. A tube was
connected to the CTC and the tube was then connected to a pressure
transducer to measure intratracheal pressure. The HFNC (Optifl ow S34 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 diagnosis, body weight) were also recorded. Kolmogorov–Smirnov
test for normality was calculated followed by bivariate nonparametric
analysis (statistical signifi cance: P <0.05). system) were applied with the humidifi er to optimize humidication. TypeB was used in three patients and typeV in four patients. The fl ow
was started at 10 l/minute. This fl ow rate was titrated upwards to a
maximum of 60 l/minute (10, 25, 30, 40, 50, 60 l/minute) and the AGFR
was measured. Intratracheal pressure tracing was done over 1 minute. Airway pressure measurement was repeated and the maximal
expiratory pressure was measured in mmHg. y
gi
Results A total of 290 measurements were included for further analysis. References References
1. Rice TW, et al.: Chest 2007, 132:410-417. 2. Sinha P, et al.: Br J Anaesth 2009, 102:692-697. 1. Rice TW, et al.: Chest 2007, 132:410-417. 1. Rice TW, et al.: Chest 2007, 132:410-417. 1. Rice TW, et al.: Chest 2007, 132:410-417. 2. Sinha P, et al.: Br J Anaesth 2009, 102:692-697. Conclusion HFNC are eff ective in providing higher expiratory pressure. It is important to know the fl ow rate is lower than expected when the
Venturi type is used. 1. Rice TW, et al.: Chest 2007, 132:410-417. 2. Sinha P, et al.: Br J Anaesth 2009, 102:692-697. *Signifi cant diff erence versus Baseline. #Signifi cant diff erence between Preinjury and Postinjury apnea. P91
Use of high-fl ow nasal canulae: eff ect on alveolar pressure and its
limitation Mean ± SD values for age and APACHE II score were 61.2 ± 16 years and
15.4 ± 1.8, respectively, while median ± SD values for PFr are 257 ± 135,
for SFr 211 ± 58, for OI 4.8 ± 4.46 and for VR 1.19 ± 0.31. Relations
between PFr, SFr and OI are displayed in Table 1. p
y p
g
Results The AGFR in the respiratory circuit was almost same in typeB,
but there was obvious decrease in the AGFR in typeV (7.1 ± 1.0,
17.7 ± 0.8, 21.9 ± 0.9, 29.9 ± 3.6, 36.9 ± 2.7, 45.0 ± 5.5 l/minute at assumed
fl ow, 10, 25, 30, 40, 50, 60 l/minute, respectively). HFNC signifi cantly
increased maximal expiratory pressure in both groups, 1.5 ± 2.1,
2.0 ± 1.0, 3.0 ± 2.8, 4.5 ± 3.5 mmHg for typeV and 2.5 ± 0.7, 5.8 ± 2.4,
6.0 ± 2.8, 8.0 ± 2.8 mmHg (maximum 10 mmHg) for typeB, when AGFR
was set at 30, 40, 50, 60 l/minute. Higher AGFRs were found to result
in larger increase in maximum expiratory pressure. The data indicate
that HFNC are associated with an increase in intratracheal expiratory
pressure. Because it was diffi cult to determine end-expiratory pressure,
we chose maximal expiratory pressure for a substitute. The reason why
AGFR in typeV was lower than assumed fl ow may be the resistance
generated by NC. The larger increase in expiratory pressure in our study
than previously reported may be due to the eff ect of high respiratory
resistance of Japanese who have relatively small airway structure
compared with western people.f Table 1 (abstract P92). Correlations (Spearman ρ) between VR, PFr, SFr and OI Table 1 (abstract P92). Correlations (Spearman ρ) between VR, PFr, SFr and OI
PFr
SFr
OI Table 1 (abstract P92). Correlations (Spearman ρ) between VR, PFr, SFr and OI
PFr
SFr
OI
VR
–0.326*
–0.325*
0.373*
PFr
–
0.759*
–0.939*
SFr
–
–
–0.773*
*P <0.01. Conclusion Our study identifi ed a strong relation between PFr, SFr and
OI but not VR. Thus, these markers may be used interchangeably as
bedside indices of oxygenation in critically ill patients. Yet larger studies
are needed to come to a safer conclusion. R f Acute respiratory distress syndrome – the Berlin defi nition: impact
on an ICU of a university hospital Acute respiratory distress syndrome – the Berlin defi nition: impact
on an ICU of a university hospital
A Agrifoglio, J Lopez, J Figueira, M Hernández, L Fernández, M Irazabal,
S Yus, M Jiménez
Hospital Universitario La Paz, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P94 (doi: 10.1186/cc12032) y
A Agrifoglio, J Lopez, J Figueira, M Hernández, L Fernández, M Irazabal,
S Yus, M Jiménez Hospital Universitario La Paz, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P94 (doi: 10.1186/cc12032) g
Results A weaning-induced pulmonary edema was diagnosed in 12
instances (PAOP signifi cantly increased from 15.6 ± 0.6 to 25.8 ± 0.9
in these cases). EVLWI, BNP, plasma protein and hemoglobin
concentrations signifi cantly increased in these instances (28.3 ± 5.7%,
20.2 ± 7.8%, 9.6 ± 0.8% and 9.3 ± 1.3%, respectively) while they did not
signifi cantly changed in cases without weaning-induced pulmonary
edema. The increase of EVLWI ≥8.5% (+1.5 ml/kg), an increase in
BNP ≥6.7% (+23 pg/ml), an increase in plasma protein concentration
≥5% and in hemoglobin concentration ≥5% exhibited good areas
under the ROC curves to predict weaning-induced pulmonary edema
(0.97 ± 0.03, 0.80 ± 0.11, 1.0 ± 0.00 and 0.92 ± 0.05, respectively). These
areas under the ROC curves were not statistically diff erent. The baseline
values of EVLWI, BNP, plasma protein and hemoglobin concentrations
did not predict weaning-induced pulmonary edema. Introduction The current defi nition of ARDS was established in 1994 by
the AECC. In 2011 emerged the Berlin defi nition. The aim of this study
was to fi nd a new approach from the impact of ARDS in an ICU and to
analyze APACHE II score, days of mechanical ventilation (MV), duration
of ARDS and mortality. y
Methods We analyzed the database of our ICU collected prospectively
from October 2008 to January 2011 and regarded all patients
undergoing MV for 48 hours or more and who met the AECC criteria. All
patients were ventilated according to the ARDSNet protocol. p
g
p
Results During this period 1,900 patients were admitted, 607 were
under MV for at least 48 hours and 104 had ARDS criteria. Moderate
ARDS 60.6%: 60% male, median age 53 years, APACHE II mean 22.49
(95% CI = 20.71 to 24.27), mean duration MV 23.84 (95% CI = 18.08 to
29.60), ARDS duration 12.87 (95% CI = 9.83 to 15.90), average stay 29.22,
mortality 52.4%. P92 Reference ICU admission have a longer duration of ARDS, which means more days
on MV and therefore increased ICU stay. Reference of 0.1 ml/kg oleic acid. Apneas (defi ned as >60% reduction in minute
ventilation) developed both before (Preinjury) and after ARDS
(Postinjury). Heart rate (HR), mean arterial pressure (MAP), respiratory
rate (RR) and tidal volume (TV) were recorded, and the PaO2:FiO2 ratio
(PFR) calculated corresponding to sections of respiratory pressure
waveforms containing 200 breaths during Baseline, during Preinjury
and Postinjury apnea periods. Similar periods were selected before
and after apneas. Inter-breath interval (IBI) and its deviation, SampEn,
percentage of forbidden words (FW), bit per-word entropy (BPWEn),
symbol distribution entropy (DisNEn), and stationarity (StatAv)
were calculated using software. Statistics by one-way ANOVA with
adjustment for multiple comparisons. 1. Ranieri M, et al.: JAMA 2012, 307:2526-2533. P95
Extravascular lung water, B-type natriuretic peptide and blood
volume contraction for diagnosing weaning-induced pulmonary
edema
M Dres, JL Teboul, N Anguel, C Richard, X Monnet
Service de réanimation médicale, Hôpital de Bicêtre, Le Kremlin Bicêtre, France
Critical Care 2013, 17(Suppl 2):P95 (doi: 10.1186/cc12033) Extravascular lung water, B-type natriuretic peptide and blood
volume contraction for diagnosing weaning-induced pulmonary
edema M Dres, JL Teboul, N Anguel, C Richard, X Monnet
Service de réanimation médicale, Hôpital de Bicêtre, Le Kremlin Bicêtre, France
Critical Care 2013, 17(Suppl 2):P95 (doi: 10.1186/cc12033) M Dres, JL Teboul, N Anguel, C Richard, X Monnet
Service de réanimation médicale, Hôpital de Bicêtre, Le Kremlin Bicêtre, France
Critical Care 2013, 17(Suppl 2):P95 (doi: 10.1186/cc12033) Results See Table 1. SampEn was associated with PFR (r2 = 0.31,
P <0.001). Introduction We compared three diff erent methods for diagnosing
pulmonary edema induced by weaning from mechanical ventilation:
the increase in extravascular lung water index (EVLWI), the increase
in B-type natriuretic peptide (BNP) and blood volume contraction,
refl ected by increases in plasma protein and in hemoglobin concen-
trations, all observed during a spontaneous breathing trial (SBT).fi Conclusion Respiratory complexity during apnea and ECLS remains
unchanged in a healthy state but decreases after ARDS. It has a
signifi cant association with PFR and may serve as an index of injury
severity. P92 After ~6 hours of ECLS, ARDS was induced by injection Table 1 (abstract P93)
Preinjury
Preinjury
Preinjury
Postinjury
Postinjury
Postinjury
BL
pre-apnea
apnea
post-apnea
pre-apnea
apnea
post-apnea
RR (breaths/minute)
22 ± 2
22 ± 3
17 ± 2
23 ± 4
24 ± 7
5 ± 1*
25 ± 7
TV (ml)
388 ± 31
217 ± 32
145 ± 14
205 ± 21
211 ± 51
116 ± 30*
229 ± 40
HR (beats/minute)
156 ± 15
153 ± 21
152 ± 21
166 ± 21
162 ± 22
165 ± 21
151 ± 16
MAP (mmHg)
102 ± 4.12
102 ± 6.13
103 ± 6.91
104 ± 6.08
106 ± 3.76
108 ± 3.86
111 ± 5.69
PFR (unitless)
435 ± 24.80
417 ± 48.06
354 ± 49.52
429 ± 37.87
139 ± 50.90*
92 ± 30.43*#
91 ± 27.25*
IBI (ms)
2,765.1 ± 230.5
3,149.8 ± 378.0
5,151.7 ± 467.7
3,404.7 ± 663.0
2,253.2 ± 427.0
10,334.0 ± 1,739.2*#
2,162.0 ± 387.6
IBI Dev (ms)
771.8 ± 117.49
1,630.0 ± 230.64
4,505.7 ± 785.34
1,598.3 ± 292.45
1,404.8 ± 445.04 25,065.0 ± 11,480.37*#
844.6 ± 236.65
SampEn (unitless)
1.7 ± 0.12
1.6 ± 0.16
1.3 ± 0.13
1.6 ± 0.11
1.2 ± 0.14
0.5 ± 0.25*#
1.3 ± 0.20
FW (%)
29.9 ± 4.60
41.5 ± 7.43
41.8 ± 10.15
34.5 ± 6.10
44.0 ± 8.80
62.2 ± 4.16*
33.6 ± 5.26
BPwEn (unitless)
4.9 ± 0.18
4.3 ± 0.27
4.1 ± 0.37
4.6 ± 0.22
4.1 ± 0.42
2.7 ± 0.38*#
4.4 ± 0.24
DisNEn (unitless)
0.8 ± 0.03
0.7 ± 0.04
0.7 ± 0.06
0.8 ± 0.05
0.7 ± 0.07
0.4 ± 0.06*#
0.7 ± 0.04
StatAv (unitless)
0.42 ± 0.06
0.37 ± 0.09
0.37 ± 0.04
0.41 ± 0.10
0.31 ± 0.05
0.51 ± 0.07
0.42 ± 0.08
*Signifi cant diff erence versus Baseline. #Signifi cant diff erence between Preinjury and Postinjury apnea. S35 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 ICU admission have a longer duration of ARDS, which means more days
on MV and therefore increased ICU stay. Reference
1. Ranieri M, et al.: JAMA 2012, 307:2526-2533. ICU admission have a longer duration of ARDS, which means more days
on MV and therefore increased ICU stay. P94 Methods We included 12 diffi cult-to-wean patients (22 recordings). Before and at the end of a SBT (T tube), we measured pulmonary
occlusion arterial pressure (PAOP), EVLWI (PiCCO device), BNP, plasma
protein and hemoglobin concentrations. Weaning-induced pulmonary
edema was confi rmed if a clinical intolerance to SBT was associated
with an increase of PAOP >18 mmHg at the end of SBT. Acute respiratory distress syndrome – the Berlin defi nition: impact
on an ICU of a university hospital Severe ARDS 39.4%: 63% male, median age 45 years,
APACHE II mean score 23.19 (95% CI = 20.62 to 25.75), mean duration
MV 18.52 (95% CI = 13.01 to 24.03), ARDS duration 12.15 (95% CI =
8.72 to 15.58), average stay 22.65, mortality 48.8% with no statistically
signifi cant diff erence with moderate ARDS (P = 0.72). Leading cause of
death in both groups: multiorgan dysfunction (60%). See Tables 1 and 2. Conclusion In this new approach of our ARDS patients we found no
statistically signifi cant diff erences in relation to the variables of interest
to analyze in the two groups. Patients with severe ARDS who survive Conclusion The increases in EVLWI, in plasma protein and hemoglobin
concentration and in BNP are valuable alternatives to the pulmonary
artery catheter for diagnosing weaning-induced pulmonary edema. P96 P96 Airway pressures and S36 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 the other hand, auto open lung management off ers a distinctive result
of ventilation. A signifi cant improvement of oxygenation and lung
compliance was observed within a few breaths after the recruitment
maneuvers. Both subjects were ventilated at the same tidal volume of
6 ml/kg and the comparative results of automatic ventilation settings
and BGA are provided in Table 1 for every 2 hours. Conclusion The auto open lung management concept gave much
better gas exchange than the auto ARDSNet protocol. These
preliminary results showed a necessity to evaluate the two diff erent
ventilation strategies. Therefore, further experiments with pig models
will be implemented in the near future to obtain results with statistical
signifi cance and to ensure the safety of automation in a mechanical
ventilation system. the other hand, auto open lung management off ers a distinctive result
of ventilation. A signifi cant improvement of oxygenation and lung
compliance was observed within a few breaths after the recruitment
maneuvers. Both subjects were ventilated at the same tidal volume of
6 ml/kg and the comparative results of automatic ventilation settings
and BGA are provided in Table 1 for every 2 hours. ventilatory mechanics were measured twice daily. Data are presented
as mean ± 95% CI. *P > 0.05. the other hand, auto open lung management off ers a distinctive result
of ventilation. A signifi cant improvement of oxygenation and lung
compliance was observed within a few breaths after the recruitment
maneuvers. Both subjects were ventilated at the same tidal volume of
6 ml/kg and the comparative results of automatic ventilation settings
and BGA are provided in Table 1 for every 2 hours. Results A total of 166 patients with mean age 55 (52 to 58) years, LIS
of 2.27 (2.16 to 2.37) and MODS of 3.13 (2.9 to 3.3) were enrolled and
stratifi ed by LISL (65.8%) and LISH (34.2%). Mortality, and incidence
of pneumothorax and atelectasis were 21% (14.9 to 28.2), 1.6% (0.8
to 3.1) and 6.4 (3.2 to 11.2). Duration of MV was 6 days (4.83 to 7.16). Pneumothorax in LISL and LISH was 1% (0 to 5.2) versus 2.4% (0.7
to 6.1)*, and mortality 11.4% (3.8 to 24.6) versus 25% (9.8 to 46.7)*. P97 Comparison of a fully automatic ARDSNet protocol and a
feedback-controlled open lung management concept
A Pomprapa1, D Schwaiberger2, B Lachmann2, S Leonhardt1
1RWTH Aachen University, Aachen, Germany; 2Charite Berlin, Germany
Critical Care 2013, 17(Suppl 2):P97 (doi: 10.1186/cc12035) Introduction Intellivent-ASV has been developed to provide fully
closed loop mechanical ventilation using a ventilation controller keep-
ing EtCO2 and SpO2 within expert-based ranges. Ventilation of ARDS
patients focuses on delivering adequate oxygenation and allowing
elimination of CO2 while protecting the lung. The objectives were to
compare Intellivent-ASV with conventional ventilation on safety and
effi cacy, and to compare the number of manual adjustments between
the two ventilatory modalities. Introduction The aim of this study is to compare two ventilation
strategies, the ARDSNet protocol and open lung management, using
computer control for 6 hours. The standard therapy for patients with
ARDS does typically apply a mechanical ventilator to support breathing. The cost of therapy is high and it requires much attention from
physicians to adjust the proper ventilation settings in a timely manner. A closed-loop ventilation concept has therefore been developed and
tested with two induced ARDS pigs. y
Methods A randomized, controlled study including all consecutive
patients receiving mechanical ventilation for at least 48 hours. Patients
were randomly ventilated either with Intellivent-ASV or conventional
ventilation, with a S1 (Hamilton, Bonaduz, Switzerland). Parameters
were adjusted by the clinician in charge of the patient. Ventilatory and
oxygenation parameters were recorded cycle by cycle during 48 hours
and blood gases were performed every 6 hours. g
Methods The hardware system is composed of a ventilator (Servo 300),
a spectrophotometry (CEVOX), a capnography device (CO2SMO+), an
electrical impedance tomography device (GOE MF II) and a patient
monitor (Sirecust). The software is developed with Labview 7.1. With
approval from the ethical committee, two 27 kg pigs were exposed to
surfactant depletion with a warm saline washout to induce ARDS (PaO2/
FiO2 <200 mmHg). One pig model was ventilated with an automatic
ARDSNet protocol and another was automatically ventilated with open
lung management. Blood gas analysis (BGA) was carried out every half
an hour.i Results Twenty-four patients with ARDS were included, 10 female, 14
male, median age 58 (46 to 63) years, APACHE II score 22 (17 to 29),
PaO2/FiO2 at inclusion 136 (107 to 154). Eleven were ventilated in the
conventional group and 13 in the Intellivent-ASV group. P96 Pressure support ventilation with minimal sedation as the main
ventilatory mode in critically ill patients with lung injury: eff ect on
mortality and incidence of complications
A Gomez1, A Leon1, G Fernandez1, G Montenegro1, H Gomez2
1Clinica Palermo, Bogota, Colombia; 2University of Pittsburgh, PA, USA
Critical Care 2013, 17(Suppl 2):P96 (doi: 10.1186/cc12034) Pressure support ventilation with minimal sedation as the main
ventilatory mode in critically ill patients with lung injury: eff ect on
mortality and incidence of complications A Gomez1, A Leon1, G Fernandez1, G Montenegro1, H Gomez2
1Clinica Palermo, Bogota, Colombia; 2University of Pittsburgh, PA, USA
Critical Care 2013, 17(Suppl 2):P96 (doi: 10.1186/cc12034) Table 1 (abstract P94). Subgroup analysis: moderate ARDS
Variable
Survivors
Nonsurvivors
P value
APACHE II
20.06 ± 6.96
24.83 ± 6.17
0.006
MV (days)
25.63 ± 29.13
22.21 ± 15.43
0.557
ARDS (days)
10.29 ± 9.07
15.20 ± 12.92
0.107
ICU stay
34.33 ± 33.26
24.57 ± 16.91
0.142
Table 2 (abstract P94). Subgroup analysis: severe ARDS
Variable
Survivors
Nonsurvivors
P value
APACHE II
20.82 ± 8.89
25.31 ± 5.63
0.761
MV (days)
26.95 ± 18.03
10.10 ± 11.64
0.001
ARDS (days)
15.8 ± 11.60
8.5 ± 8.6
0.031
ICU stay
33.52 ± 18.25
11.25 ± 11.80
0.001 Table 1 (abstract P94). Subgroup analysis: moderate ARDS Introduction The primary aim of this study is to assess the impact of
pressure support ventilation (PSV) on the rate of pneumothorax and
mortality in critically ill patients with lung injury. The secondary aim is
to evaluate pressure–volume (P–V) relationships. Spontaneous modes
of ventilation have been associated with lower rates of atelectasis,
less muscle atrophy, better airfl ow distribution and importantly lower
sedation requirements, which relates to lower mortality. Accordingly,
we hypothesized that the use of PSV in patients with moderate/severe
lung injury would have rates of pneumothorax and mortality within the
standard of care. We further hypothesized that given its spontaneous
nature, set pressures (PEEP and PS) but not tidal volume (Vt) would be
related to airway pressures. Methods All adult patients admitted to two surgical/medical ICUs
subjected to invasive mechanical ventilation (MV) were enrolled. Patients were stratifi ed by Lung Injury Score (LIS) in two groups:
<2.5 (LISL); ≥2.5 (LISH). Exclusion criteria included pneumothorax on
admission, use of other ventilatory strategies, and inability to trigger
ventilation. Patients were ventilated with PSV, and treated only with
pro re nata haldol, morphine and clozapine. P96 Mean parameters were: Vt 10.3 ml/kg (10.1 to 10.6); PEEP 10.6 (9.9 to
11); PS 16.6 (15.9 to 17.1); plateau pressure (Ppl) 25.7 (25.1 to 26.2). Ppl was >26.2 only in 2.5%. PEEP and PS (P <0.0001), and MODS were
associated with Pplt, but not Vt* or LIS. Only lower Vt was associated
with barotrauma (OR = 0.996, P = 0.02). p
y
Conclusion The auto open lung management concept gave much
better gas exchange than the auto ARDSNet protocol. These
preliminary results showed a necessity to evaluate the two diff erent
ventilation strategies. Therefore, further experiments with pig models
will be implemented in the near future to obtain results with statistical
signifi cance and to ensure the safety of automation in a mechanical
ventilation system. Conclusion We demonstrate that PSV in minimally sedated patients
with severe lung injury is safe as it is associated with low incidence
of barotrauma, atelectasis and mortality, and with Ppl and duration
of MV within standard of care. We also demonstrate in PSV that P–V
relationships may diff er and that in this setting higher Vt may not be
deleterious. P98 Comparison of an entirely automated ventilation mode,
Intellivent-ASV, with conventional ventilation in ARDS patients:
a 48-hour study
E Bialais1, L Vignaux2, X Wittebole1, D Novotni2, J Meyer2, M Wysocki2,
T Sottiaux3, G Reychler1, J Roeseler1, P Laterre1, P Hantson1
1Cliniques Universitaires Saint Luc, Brusells, Belgium; 2Hamilton Medical AG,
Bonaduz, Switzerland; 3Clinique Notre-Dame de Grâce, Gosselies, Belgium
Critical Care 2013, 17(Suppl 2):P98 (doi: 10.1186/cc120236) P97 The study was
stopped for one patient from the Intellivent-ASV group because of a
pneumothorax not caused by ventilation. The delivered Vt was slightly
higher during Intellivent-ASV (7.9 (7.5 to 8.5) vs. 7.2 (6.8 to 7.8) ml/kg,
P = 0.013). The time spent by the various parameters in the suboptimal
zone (safety) is the same for the two ventilation modes. The time spent
in the optimal zone (effi cacy) is the same for the two ventilation modes, Results Artifi cial ventilation using the auto ARDSNet protocol success-
fully stabilized oxygenation, minimized plateau pressure (<30 cmH2O),
and controlled the pH value for acidosis and alkalosis management. On Table 1 (abstract P97). Comparative results between auto ARDSNet protocol and auto open lung management
Before auto ventilation
After 2 hours
After 4 hours
After 6 hours
Auto ARDSNet
Auto OLM
Auto ARDSNet
Auto OLM
Auto ARDSNet
Auto OLM
Auto ARDSNet
Auto OLM
FiO2
1.0
1.0
0.40
0.25
0.40
0.25
0.50
0.25
RR (bpm)
22
40
23
38
23
39
23
35
I:E ratio
1:2
1:1
1:2
1:1
1:2
1:1
1:2
1:1
VT (ml/kg)
6
6
6
6
6
6
6
6
Pplat /PIP (mmHg)
22
22
18
20.6
15.8
17.7
14.6
15.4
PEEP (cmH2O)
5
10
8
15
2
13
10
11
pH
7.34
7.30
7.32
7.52
7.37
7.60
7.38
7.51
pO2/FiO2 (mmHg)
59
59
180
380.8
155
387.2
122
404
pCO2 (mmHg)
52
64
56
39.6
58
31.5
54
41
OLM, open lung management. Table 1 (abstract P97). Comparative results between auto ARDSNet protocol and auto open lung management . Comparative results between auto ARDSNet protocol and auto open lung management S37 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 expect for SpO2 (78% (75 to 89) with Intellivent-ASV vs. 44% (41 to 58),
P = 0.001). Intellivent-ASV required less manual adjustments during
48 hours (2 (0 to 7) vs. 17 (10 to 27), P <0.001). The total number of
adjustments during 48 hours, including automatic regulation, was
higher in the Intellivent-ASV group (379 (274 to 493) vs. 17 (10 to 27),
P <0.001). Methods This observational study in ICU mechanically ventilated septic
patients with intra-abdominal surgical infections was carried out at the
V.A. Negovsky Research Institute of General Reanimatology in 2010
to 2012. ARDS was diagnosed and staged according to the research
institute criteria [1] and the American–European criteria. Diff erence in pulmonary permeability between indirect and
direct acute respiratory distress syndrome assessed by the
transpulmonary thermodilution technique K Morisawa1, Y Taira1, M Yanai1, Y Takamatu1, S Kushimoto2, S Fujitani3
1St Marianna University, Kawasaki-Shi, Japan; 2Tohoku University Graduate
School of Medicine, Sendai-shi, Japan; 3Tokyo Bay Urayasu/Ichikawa Medical
Center, Urayasu-shi, Japan
Critical Care 2013, 17(Suppl 2):P99 (doi: 10.1186/cc12037) Introduction ARDS is characterized by increased pulmonary capillary
permeability secondary to diff use alveolar infl ammation and injury. Common risk factors can be classifi ed into two groups: extrapulmonary
causes (indirect etiologies: ARDSexp) or pulmonary causes (direct
etiologies: ARDSp). There are few quantitative methods to distinguish
between the diff erences in these two ARDS categories. Conclusion Plasma SPD level ≤253.0 ng/ml is a sensitive and specifi c
biomarker of the early stage of ARDS in septic patients. Reference f
g
Methods A subanalysis of the trial by the PiCCO Pulmonary Edema
Study (prospective, observational, multi-institutional study) in 23 ICUs
of academic tertiary referral hospitals in Japan. All consecutive adult
patients requiring mechanical ventilation with the diagnosis of ARDS
were monitored by the transpulmonary thermodilution technique
system (PiCCO; Pulsion Medical Systems, Munich, Germany) for 3 days:
day 0, day 1 and day 2. The pulmonary vascular permeability index
(PVPI), extravascular lung water index and intrathoracic blood volume
index were measured concurrently. Three experts retrospectively
determined the pathophysiological mechanism causing ARDS by
patient history, clinical presentation, chest computed tomography and
radiography. Patients were classifi ed into two groups: patients with
ARDS triggered by ARDSexp and ARDSp. 1. Kuzovlev A, et al.: Semin Cardiothorac Vasc Anesth 2010, 14:231-24 P101 Intravenous perfl uorocarbons for prevention of
ventilator-associated ARDS A Scultetus1, A Haque1, F Arnaud1, G McNamee2, C Auker1, R McCarron1,
P McKay2, R Mahon1
1Naval Medical Research Center, Silver Spring, MD, USA; 2Uniformed Services
University of the Health Sciences, Bethesda, MD, USA
Critical Care 2013, 17(Suppl 2):P101 (doi: 10.1186/cc12039) A Scultetus1, A Haque1, F Arnaud1, G McNamee2, C Auker1, R McCarron1,
P McKay2, R Mahon1 Results During the study period from March 2009 to August 2011, a
total of 173 patients were assessed including 56 ARDSexp patients
and 117 ARDSp patients, with the most common cause of ARDSexp
secondary to sepsis (71%) and of ARDSp pneumonia (80%). The
measure ment of PVPI was signifi cantly elevated in the ARDSp group
on all days. There were no signifi cant diff erences in mortality at 28 days,
mechanical ventilation days, and hospital length-of-stay between the
two groups, while the ARDSexp group seemed to be associated with
prolonged mechanical ventilation days and hospital length-of-stay.f Introduction Emulsifi ed perfl uorocarbons (PFC) are synthetic hydro-
carbons that can carry 50 times more oxygen than human plasma. Their
properties may be advantageous in applications requiring preservation
of tissue viability in oxygen-deprived states [1,2], making them a
potential candidate for combat and civilian prehospital resuscitation. Our hypothesis is that an intravenous dose of PFC increases vital organ
tissue oxygenation, improves survival after hemorrhagic shock (HS)
and may reduce or prevent the development of ventilator-associated
ARDS. Here we report data from the fi rst part (HS only) of a multiphase
swine study to study the benefi ts of PFC in treating HS and preventing
ARDS. This initial study was designed to ensure safe use of PFC in
traumatized animals. Conclusion This study suggests the existence of several diff erences
in pathogenetic pathways and the degree of pulmonary permeability
between patients with ARDSexp and ARDSp. We therefore believe that
using the PVPI may provide us with timely quantitative information to
make clinical decisions. Methods Anesthetized Yorkshire swine were hemorrhaged 55% of their
estimated blood volume (Time 0 (T0)) over 15 minutes. At T15 minutes,
pigs received a bolus of the PFC Oxycyte or 10% hydroxyethyl starch
(HES). At T60 animals underwent continuous hemorrhage (0.5 cm3/kg/
minute) until death. Time to death and physiological parameters were
primary endpoints. P97 Plasma
concentrations of SPD and CCP were measured on ARDS diagnosis
(day 0) and days 3, 5, and 7 by immunoenzyme essay (Bio Vendor, USA). Data were statistically analyzed by STATISTICA 7.0, ANOVA method,
and presented as mean ± σ, ng/ml. P <0.05 was considered statistically
signifi cant. Areas under the receiver operating curves (ROC) were
calculated. Conclusion Over 48 hours, Intellivent-ASV allowed safe ventilation for
ARDS patients, with better oxygenation and an identical ventilation
effi cacy when compared with conventional ventilation, with less
manual adjustments. Results Fifty-fi ve patients (out of 250 screened) were enrolled according
to the inclusion/exclusion criteria. Patients were assigned into groups:
ARDS (n = 30; subgroups – ARDS stage 1 (n = 15), stage 2 (n = 15)) and
noARDS (n = 25). In the ARDS group SPD was higher at all points than
in the noARDS group (day 0 – 352.5 ± 287.8 vs. 141.0 ± 103.4; day 3 –
278.6 ± 204.8 vs. 151.8 ± 125.9; day 5 – 339.9 ± 331.4 vs. 162.3 ± 138.8;
day 7 – 339.9 ± 300.1 vs. 169.8 ± 154.5; P <0.05). SPD levels were lower
at ARDS stage 1 in comparison with stage 2 (day 0 – 154.6 ± 125.2 vs. 451.8 ± 299.2; day 3 – 129.3 ± 74.9 vs. 362.9 ± 202.0; day 5 – 167.8 ± 120.4
vs. 428.5 ± 372.4; day 7 – 186.2 ± 127.1 vs. 415.5 ± 337.0; P <0.05). Plasma SPD had a good diagnostic capacity for stage 1 ARDS: SPD on
day 0 ≤253.0 ng/ml yielded a sensitivity of 90% and specifi city of 74%
(AUC = 0.83; 95% CI = 0.631 to 0.951; P = 0.0001). No such diff erences
for CCP were detected.i P103
End-expiratory esophageal pressure versus lower infl ection point in
acute lung injury Results In the multivariable model, age, lung compliance, serum lactate
and serum ACE levels were signifi cantly associated with both 28-day
and 90-day mortality. No signifi cant correlation was found between
serum and BALF ACE levels (Spearman’s ρ = 0.054; P = 0.66). Serum
ACE concentrations were signifi cantly higher (P = 0.046) in patients
with D/D genotype versus the two other groups combined (I/D and
I/I genotypes). A meta-analysis of six studies (including ours) provided
evidence that the D allele is signifi cantly associated with increased
mortality in ALI/ARDS patients, yielding a per-allele odds ratio of 1.76
(95% CI: 1.19 to 2.59). See Figure 1 and Table 1.f Introduction No recommendations available concerning protocols of
static PV loop and esophageal pressure measurements use set positive
end-expiratory pressure (PEEP). The aim of the study was evaluation
of the signifi cance of the lower infl ection point (LIP) and esophageal
pressure monitoring for PEEP adjustment in ALI and ARDS. Conclusion Serum ACE levels appear to be aff ected by the I/D poly-
morphism and are correlated with prognosis in patients with ALI/ARDS,
indicating that further investigation of the clinical signifi cance of the
ACE in ARDS might be of value. Methods A prospective study performed in one general ICU. We include
72 patients who received mechanical ventilation before evaluation
and met ARDS criteria by AECC (1994) – acute onset, PaO2/FiO2 lower
than 250 Torr, bilateral infi ltrates on chest X-ray. Exclusion criteria were
age <15 years and pregnancy. We drew a static pressure–volume loop
with sustained infl ation 40×30 (PV loop) for all patients using a low-
fl ow technique (Hamilton G5) and measured the esophageal pressure
(Avea) in 36 of 72 patients. After that PEEP was set according to zero
end-expiratory transpulmonary pressure. We compare PV loop data
with esophageal pressure measurements.l Figure 1 (abstract P102). Figure 1 (abstract P102). Results The low infl ection point median was 8 (95% CI = 5 to 10.5)
mbar, which does not correspond to the empirically set optimal PEEP
of 13 (95% CI = 12 to 15) mbar (P <0.001, Wilcoxon test). End-expiratory
esophageal pressure (EEEP) median was 14 (95% CI = 12 to 18) mbar,
the correlation between LIP and EEEP was poor (ρ = 0.279, P = 0.049). We fi nd signifi cant correlation between static compliance and EEEP
(ρ = –0.421, P = 0.005). P100 P102
Eff ect of angiotensin-converting enzyme gene I/D polymorphism
and its expression on clinical outcome in acute respiratory distress
syndrome
I Tsangaris, A Tsantes, P Kopterides, G Tsaknis, S Kokkori, I Karampela,
D Konstantonis, S Karabi, E Vrigkou, A Pappas, S Orfanos, A Armaganidis
University Hospital ‘Attikon’, University of Athens Medical School, Athens,
Greece
Critical Care 2013, 17(Suppl 2):P102 (doi: 10.1186/cc12040)
Introduction The role of the D allele of the angiotensin-converting
enzyme (ACE) gene I/D polymorphism in the clinical outcomes of
patients with acute lung injury and acute respiratory distress syndrome
(ALI/ARDS) remains controversial. We assessed simultaneously the
eff ect of the ACE I/D polymorphisms as well as the serum and BALF ACE
levels on prognosis of ARDS patients. Methods We recruited 69 mechanically ventilated ALI/ARDS patients. ACE activity levels in serum and BALF were assessed by chemical
th d
P ti
t
t
d f
ACE I/D
l
hi
Ti
Table 1 (abstract P102). Characteristics of the study population (n = 69)
Age (years)
64.4 ± 17.9; 69 (28 to 89)
Sex (males, %)
43 (62.3%)
APACHE II score
22.1 ± 6.2; 21 (14 to 35)
SOFA score
8.4 ± 3.2; 8 (4 to 16)
Lung compliance (ml/cmH2O)
29.4 ± 7.2; 30 (17 to 40)
PaO2/FiO2
139, 1 ± 47.1; 140 (72 to 223)
PEEP
8.1 ± 2.9; 8 (5 to 15)
Lung Injury Score
2.5 ± 0.6; 2.5 (1.75 to 3.50)
Blood lactate (mmol/l)
1.7 ± 1.7; 1.3 (0.5 to 5.2)
Septic status
39 (56.5%)
Serum ACE (U/l)
16.5 ± 10.8; 13.6 (4.8 to 39.8)
BALF (U/l)
2.3 ± 1.4; 2.0 (0.3 to 4.7)
ACE I/D polymorphism:
D/D
27 (39.1%)
I/D
28 (40.6%)
I/I
14 (20.3%)
28-day mortality
34/69 (49.3%)
90-day mortality
43/69 (62.3%)
Ventilator-free days
5.1 ± 7.8; (0 to 23)
Days w/o cardiovascular failure
14.0 ± 10.2; 16 (0 to 27)
Days w/o renal failure
15.4 ± 10.6; 16 (0 to 28) Table 1 (abstract P102). 0
Eff ect of angiotensin-converting enzyme gene I/D polymorphism
and its expression on clinical outcome in acute respiratory distress
syndrome Eff ect of angiotensin-converting enzyme gene I/D polymorphism
and its expression on clinical outcome in acute respiratory distress
syndrome y
I Tsangaris, A Tsantes, P Kopterides, G Tsaknis, S Kokkori, I Karampela,
D Konstantonis, S Karabi, E Vrigkou, A Pappas, S Orfanos, A Armaganidis
University Hospital ‘Attikon’, University of Athens Medical School, Athens,
Greece Critical Care 2013, 17(Suppl 2):P102 (doi: 10.1186/cc12040) Introduction The role of the D allele of the angiotensin-converting
enzyme (ACE) gene I/D polymorphism in the clinical outcomes of
patients with acute lung injury and acute respiratory distress syndrome
(ALI/ARDS) remains controversial. We assessed simultaneously the
eff ect of the ACE I/D polymorphisms as well as the serum and BALF ACE
levels on prognosis of ARDS patients. p
g
p
Methods We recruited 69 mechanically ventilated ALI/ARDS patients. ACE activity levels in serum and BALF were assessed by chemical
methods. Patients were genotyped for ACE I/D polymorphisms. Time-
to-event analysis evaluated the variables associated with the 28-day
and 90-day mortality. P100 Characteristics of the study population (n = 69)
Age (years)
64.4 ± 17.9; 69 (28 to 89)
Sex (males, %)
43 (62.3%)
APACHE II score
22.1 ± 6.2; 21 (14 to 35)
SOFA score
8.4 ± 3.2; 8 (4 to 16)
Lung compliance (ml/cmH2O)
29.4 ± 7.2; 30 (17 to 40)
PaO2/FiO2
139, 1 ± 47.1; 140 (72 to 223)
PEEP
8.1 ± 2.9; 8 (5 to 15)
Lung Injury Score
2.5 ± 0.6; 2.5 (1.75 to 3.50)
Blood lactate (mmol/l)
1.7 ± 1.7; 1.3 (0.5 to 5.2)
Septic status
39 (56.5%)
Serum ACE (U/l)
16.5 ± 10.8; 13.6 (4.8 to 39.8)
BALF (U/l)
2.3 ± 1.4; 2.0 (0.3 to 4.7)
ACE I/D polymorphism:
D/D
27 (39.1%)
I/D
28 (40.6%)
I/I
14 (20.3%)
28-day mortality
34/69 (49.3%)
90-day mortality
43/69 (62.3%)
Ventilator-free days
5.1 ± 7.8; (0 to 23)
Days w/o cardiovascular failure
14.0 ± 10.2; 16 (0 to 27)
Days w/o renal failure
15.4 ± 10.6; 16 (0 to 28) Table 1 (abstract P102). Characteristics of the study population (n = 69) on these data we will continue to the next phase of this project and
test PFC in the prevention of ARDS alone, and in combination with HS. References on these data we will continue to the next phase of this project and
test PFC in the prevention of ARDS alone, and in combination with HS. References 1. Keipert PE, et al.: Enhanced oxygen delivery by perfl ubron emulsion during
acute hemodilution. Artif Cells Blood Substit Immobil Biotechnol 1994,
22:1161-1167.l 2. Habler O, et al.: IV perfl ubron emulsion versus autologous transfusion in
severe normovolemic anemia: eff ects on left ventricular perfusion and
function. Res Exp Med (Berl) 1998, 197:301-318. 1.
Keipert PE, et al.: Enhanced oxygen delivery by perfl ubron emulsion during
acute hemodilution. Artif Cells Blood Substit Immobil Biotechnol 1994,
22:1161-1167. P100 Biomarkers for early stage of acute respiratory distress syndrome in
septic patients: surfactant protein D and Clara cell protein
A Kuzovlev1, V Moroz1, A Goloubev1, S Polovnikov2
1V.A. Negovsky Scientifi c Research Insitute of General Reanimatology RAMS,
Moscow, Russia; 2N.N. Burdenko Main Military Hospital, Moscow, Russia
Critical Care 2013, 17(Suppl 2):P100 (doi: 10.1186/cc12038) Results Average survival time after onset of second hemorrhage
was 60 minutes in PFC-treated animals versus 65 minutes in the HES
group. Mean arterial pressure (MAP) was similar between T0 and T60,
thereafter PFC-treated animals had lower MAP, mean pulmonary artery
pressure (MPAP), heart rate and cardiac output (P >0.05).if Conclusion There was no signifi cant diff erence in survival time, MAP
and MPAP in the PFC group compared with HES control. These data
suggest that it is safe to administer in this HS model. Regulatory
approval of hemoglobin-based oxygen carriers has been halted due to
possible side eff ects related to vasoconstriction. In this model, and with
this class of oxygen-carrying drugs (PFCs), we did not observe evidence
of vasoconstriction. Using PFCs did not result in a survival advantage
here; however, there was also no observation of adverse events. Based Introduction The injury of alveolar epithelium and endothelium is the
basis of pathogenesis for the early stage of acute respiratory distress
syndrome (ARDS). A prompt detection of this injury will provide us
with a possibility for early ARDS diagnosis and treatment. Currently
no biomarkers of alveolar epithelial injury are clinically available. The
purpose of our study was to investigate the role of surfactant protein D
(SPD) and Clara cell protein (CCP) as biomarkers of early ARDS. S38 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 on these data we will continue to the next phase of this project and
test PFC in the prevention of ARDS alone, and in combination with HS. References
1. Keipert PE, et al.: Enhanced oxygen delivery by perfl ubron emulsion during
acute hemodilution. Artif Cells Blood Substit Immobil Biotechnol 1994,
22:1161-1167. 2. Habler O, et al.: IV perfl ubron emulsion versus autologous transfusion in
severe normovolemic anemia: eff ects on left ventricular perfusion and
function. Res Exp Med (Berl) 1998, 197:301-318. P103 P103
End-expiratory esophageal pressure versus lower infl ection point in
acute lung injury
A Yaroshetskiy1, D Protsenko1, E Larin2, O Ignatenko1, B Gelfand1
1Russian National Research Medical University, Moscow, Russia; 2City
Hospital #7, Moscow, Russia
Critical Care 2013, 17(Suppl 2):P103 (doi: 10.1186/cc12041) 2.
Habler O, et al.: IV perfl ubron emulsion versus autologous transfusion in
severe normovolemic anemia: eff ects on left ventricular perfusion and
function. Res Exp Med (Berl) 1998, 197:301-318. Quantifi cation of stress raisers in ARDS Quantifi cation of stress raisers in ARDS
M Cressoni1, M Amini1, P Cadringher2, C Chiurazzi1, D Febres1, E Gallazzi1,
A Marino1, M Brioni1, F Menga1, I Cigada1, E Carlesso1, D Chiumello2,
L Gattinoni2
1Università degli Studi di Milano, Milan, Italy; 2Fondazione IRCCS Ca’ Granda-
Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P105 (doi: 10.1186/cc12043) Results We included in our study the CT scans of 89 patients (24 mild,
55 moderate and 10 severe ARDS, age 58 ± 17 years, BMI 25.6 ± 5.4,
PaO2/FiO2 165 ± 66, PaCO2 42 ± 9, 60 discharged alive from ICU
(67%)). At PEEP 5 cmH2O 39 ± 16% of the lung parenchyma was not
infl ated, 33 ± 13% poorly infl ated, 28 ± 15% well infl ated and 0 ± 1%
overinfl ated. The median lung recruitability between 5 and 15 cmH2O
PEEP was 6% (3 to 10%) of lung parenchyma (71 g (26 to 182 g)). Mean
EELV at PEEP 5 cmH2O was 1,233 ± 709 while it increased to 1,784 ± 848
at PEEP 15 cmH2O (ΔEELV = 551 ± 407 ml). More than one-half (52% (31
to 75%)) of the increase in end-expiratory lung volume (EELV) was due
to an increase in infl ation of the already well-aerated tissue; 38% (23 to
43%) was due to the infl ation of poorly aerated tissue, while only 4%
(1 to 8%) to the recruitment of lung tissue. The CLsp of the well, poorly
and recruited tissue was 0.07 (0.031 to 0.13), 0.03 (0.01 to 0.06) and 0.02
(0.01 to 0.04) ml/cmH2O/g, respectively (P <0.0001). Introduction Ventilator-induced lung injury (VILI) is a well-known side
eff ect of mechanical ventilation. The pressures and volumes needed
to induce VILI in healthy animals are far greater than pressure and
volumes applied in clinical practice [1]. A possible explanation may be
the presence of local pressure multipliers (stress raisers). Methods We retrospectively analyzed CT scans of 147 patients with
ARDS and CT scans of 100 healthy subjects. A homogeneous lung
would have the same gas/tissue ratio in all its regions. If a lung region
expands less than the neighbour regions these will be more strained
to vicariate the non/less expanding region. Recruited lung tissue does not resume normal mechanical
properties g
Results Patients were 65 ± 13 years old, ventilation days were 9 ± 10
and PaO2/FiO2 was 194 ± 49 mmHg at PEEP 8 ± 2 cmH2O. Introduction
of Sigh improved PaO2/FiO2 (P <0.001) and ΔEELVgl (P <0.001). The increase in EELV was diff use across all lung regions as both
ΔEELVnondep and ΔEELVdep changed (P <0.01 and P = 0.06). minute
ventilation, arterial blood pH and hemodynamics were not signifi cantly
aff ected by introduction of Sigh, while mean airway pressure increased
(P <0.001), albeit only at the highest Sigh rate and by a limited extent
(~2 cmH2O). Patients with mean ΔEELVgl over the three Sigh phases
>200 ml (that is, Sigh-responders) had signifi cantly lower PaO2/
FiO2 values at baseline (that is, at Sigh rate = 0/minute) than Sigh-
nonresponders (163 ± 25 mmHg vs. 222 ± 39 mmHg, P <0.01). p
p
M Cressoni1, C Chiurazzi1, M Amini1, D Febres1, E Gallazzi1, E Carlesso1,
P Cadringher1, T Langer1, D Chiumello1, L Gattinoni2
1Università degli Studi di Milano, Milan, Italy; 2Fondazione IRCCS Ca’ Granda –
Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P106 (doi: 10.1186/cc12044) Introduction Positive end-expiratory pressure (PEEP) is commonly
used in patients with ARDS to prevent end-expiratory collapse. The
increase in gas volume and/or change in mechanical properties of
the lung are used at the bedside to estimate lung recruitment and
individualize PEEP selection. Methods Retrospective analysis of patients who underwent whole-
lung CT scan at 5 and 15 cmH2O PEEP. Every chest CT was divided both
into 10 apex-base levels of equal height and into 10 sterno-vertebral
levels, yielding 100 units per scanned lung (200 units per patient);
each unit was classifi ed according to its average CT number (<–100
not infl ated, –100 to –500 poorly infl ated, –500 to –900 well infl ated
and –900 overinfl ated). Lung regions defi ned as not infl ated at PEEP
5 cmH2O and infl ated (poorly, well or over) at PEEP 15 cmH2O were
classifi ed as recruited. Moreover, a surrogate specifi c lung compliance
(CLsp) was determined on the expiratory limb of the pressure–volume
curve (15 to 5 cmH2O PEEP at CT) and was defi ned as Δgas/cmH2O
PEEP/gram of tissue. Gattinoni L, et al.: Int Care Med 2005. 31:776-784. Lung recruitment induced by Sigh in hypoxemic intubated critically
ill patients p
TM Mauri1, G Bellani1, A Coppadoro1, P Tagliabue2, A Barletta1, V Meroni2,
M Teggia Droghi1, N Patroniti1, A Pesenti1
1University of Milan-Bicocca, Monza, Italy; 2San Gerardo Hospital, Monza, Italy
Critical Care 2013, 17(Suppl 2):P104 (doi: 10.1186/cc12042) of the stress raisers the fraction of lung volume above this threshold. Results The extent of stress raisers increased with the severity of
ARDS (14 ± 5, 18 ± 8, 23 ± 1% of lung parenchyma in mild, moderate
and severe ARDS, P <0.0001). The extent of stress raisers correlated
with the dead space fraction (r2 = 0.34, P <0.001), with the fraction
of poorly aerated tissue (r2 = 0.36, P <0.0001) and also has a negative
correlation with the fraction of well infl ated tissue (r2 = 0.47, P <0.0001). The response to PEEP, passing from 5 to 45 cmH2O is minimal (average
decrease of stress raiser extent 6 ± 5%) and inter-individual variability
is great (in 11 patients, stress raisers increased passing from PEEP 5 to
PEEP 45). Stress raisers turn out to be greater in nonsurvivor patients
than in survivor patients (17 ± 7 vs. 20 ± 9% of lung volume, P = 0.03). Introduction In intubated critically ill patients, cyclic short recruitment
manoeuvres (Sigh) introduced during pressure support ventilation
(PSV) improve oxygenation, probably by increasing end-expiratory
lung volume (EELV). We assessed Sigh eff ects on regional distribution
of EELV by electrical impedance tomography (EIT): a bedside, radiation-
free lung imaging technique. Moreover, we investigated baseline
characteristics correlated with response to Sigh. Methods We enrolled 20 intubated critically ill patients undergoing
PSV with PaO2/FiO2 ≤300 mmHg and PEEP ≥5 cmH2O. We applied on
each patient’s thorax a 16-electrode belt connected to an EIT monitor
(PulmoVista 500®; Dräger Medical GmbH, Lübeck, Germany). Sigh was
introduced as 35 cmH2O continuous positive airway pressure phase
lasting 3 to 4 seconds at diff erent rates (0, 0.5, 1, 2 per minute, random
order) for 20 minutes. From raw EIT data, we calculated global changes
in EELV (ΔEELVgl), calculated as changes in end-expiratory lung
impedance calibrated versus tidal volume to estimate changes in ΔEELV
and considering 0 Sigh/minute as baseline; and ΔEELV of nondependent
and dependent lung regions (ΔEELVnondep and ΔEELVdep). Together,
we collected ventilation parameters, hemodynamics and arterial blood
gases. Reference 1. Protti A: Am J Respir Crit Care Med 2011, 183:1354-1362. 1. Protti A: Am J Respir Crit Care Med 2011, 183:1354-1362. Recruited lung tissue does not resume normal mechanical
properties p
g
g
Conclusion Introduction of Sigh in intubated critically ill patients
undergoing PSV improves gas exchange by inducing alveolar
recruitment across all lung regions with minimal increase of airway
pressure. Patients with more severe gas exchange impairment present
the highest degree of potentially recruitable lung. Quantifi cation of stress raisers in ARDS We measured the stress
raisers by computing the ratio between the gas fraction of the region
of interest and the neighbouring regions: if the infl ation would be the
same (homogeneity), the ratio will be equal to one; if the infl ation of the
surrounding regions would be greater than the region of interest (that
is, more strained), the ratio between the two will be greater than one 2
Conclusion Most of the PEEP-related EELV increase is due to an
increased infl ation of already well aerated lung tissue. In agreement
with the baby lung theory [1], the well-aerated tissue showed near to
normal mechanical properties. Lung recruitment induced by Sigh in hypoxemic intubated critically
ill patients P106 Recruited lung tissue does not resume normal mechanical
properties
M Cressoni1, C Chiurazzi1, M Amini1, D Febres1, E Gallazzi1, E Carlesso1,
P Cadringher1, T Langer1, D Chiumello1, L Gattinoni2
1Università degli Studi di Milano, Milan, Italy; 2Fondazione IRCCS Ca’ Granda –
Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P106 (doi: 10.1186/cc12044) P103
End-expiratory esophageal pressure versus lower infl ection point in
acute lung injury Sustained infl ation did not lead to improved
oxygenation (P >0.05). PEEP adjustment by EEEP led to an increase in
PaO2/FiO2 – median 107 mmHg (95% CI = 18 to 147, P <0.001). EEEP was
similar to empirically set PEEP (P >0.05). Conclusion LIP has poor correlation with EEEP. PEEP adjustment by
esophageal pressure was close to empirically set PEEP and can improve
oxygenation. S39 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P104 and was taken as a measure of stress raiser. We considered pathological
stress raisers as the regions showing infl ation ratio greater than the
95th percentile of the control group (1.61) and defi ned as the extent P104
Lung recruitment induced by Sigh in hypoxemic intubated critically
ill patients
TM Mauri1, G Bellani1, A Coppadoro1, P Tagliabue2, A Barletta1, V Meroni2,
M Teggia Droghi1, N Patroniti1, A Pesenti1
1University of Milan-Bicocca, Monza, Italy; 2San Gerardo Hospital, Monza, Italy
Critical Care 2013, 17(Suppl 2):P104 (doi: 10.1186/cc12042) Reference Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S40 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P107
Intraoperative management of hypoxemia with recruitment
maneuvers: are the benefi ts worth the costs? V Grosomanidis, B Fyntanidou, K Karakoulas, K Kotzampasi, E Oloktsidou,
C Nouris, C Skourtis
Aristotle Medical School, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P107 (doi: 10.1186/cc12045) P107 and without inotropic support were enrolled. A transthoracic echo-
cardiography was performed to assess diastolic function: E/A, isovolumic
relaxation time (IVRT), deceleration time (DT), E/e’ were assessed. Eight
patients (fi ve with poor relaxation pattern, three with decreased left
ventricle compliance) were identifi ed in diastolic dysfunction group
(Group 1) and eight were included in the normal diastolic function
group (Group 2). A recruitment manoeuvre was performed by applying
a PEEP of 40 cmH2O for 30 seconds. Haemodynamic parameters of
cardiac output (CO), stroke volume (SV), heart rate (HR), pulse pressure
variation (PPV), and cardiac cycle effi ciency (CCE) were continuously
recorded during the manoeuvre using the MostCare pulse contour
method.ii and without inotropic support were enrolled. A transthoracic echo-
cardiography was performed to assess diastolic function: E/A, isovolumic
relaxation time (IVRT), deceleration time (DT), E/e’ were assessed. Eight
patients (fi ve with poor relaxation pattern, three with decreased left
ventricle compliance) were identifi ed in diastolic dysfunction group
(Group 1) and eight were included in the normal diastolic function
group (Group 2). A recruitment manoeuvre was performed by applying
a PEEP of 40 cmH2O for 30 seconds. Haemodynamic parameters of
cardiac output (CO), stroke volume (SV), heart rate (HR), pulse pressure
variation (PPV), and cardiac cycle effi ciency (CCE) were continuously
recorded during the manoeuvre using the MostCare pulse contour
method. Introduction Several strategies can be applied for the prevention and
management of anesthesia-related hypoxemia. The aim of our study
was to investigate the cardiovascular eff ects of a specifi c recruitment
maneuver (RM) used for the management of intraoperative hypoxemia. Methods Forty-nine patients (29 male/20 female) of mean age
57.4 ± 9.9 years, mean weight 80.2 ± 13.6 kg and ASA-PS classifi cation I
to IV (I: 2, II: 27, III: 16, IV: 6), undergoing general surgery procedures, who
developed intraoperative hypoxemia (PaO2/FiO2 <200), were enrolled
in our study. For the management of hypoxemia a total of 67 RMs
have been applied, which consisted of a manual increase of the airway
pressure to 40 cmH2O. Reference This rise has been maintained for 10 seconds and
was followed by an increase of positive end-expiratory pressure from 5
to 10 cmH2O. Before RM application an oesophageal Doppler monitor
probe was inserted into the patients for measuring stroke volume (SV),
cardiac output (CO), peak velocity (PV) and fl ow corrected time (FTc). Standard monitoring also included ECG, IBP, ETCO2 and SpO2. Heart rate
(HR), systemic arterial pressure (SAP), SV, CO, PV and FTc were recorded
directly before, during, right after and 5 minutes after RM application
(T1 to T4). Kolmogorov–Smirnoff was used to test normal distribution
of data and ANOVA was used for the statistical analysis. P <0.05 was
considered statistically signifi cant.i Results No signifi cant haemodynamic changes happened in the fi rst
part of the recruitment (t0 to t15 seconds). A signifi cant decrease in
CO (P <0.05), SV (P <0.01), systolic pressure (P <0.04) and PPV (P <0.04)
occurred in Group 1 during the remaining part of the manoeuvre. No
signifi cant decrease in the same haemodynamic parameters were
noted when only E/A was used to discriminate the diastolic dysfunction. See Table 1. Table 1 (abstract P108). CO diff erence associated with diastolic dysfunction
parameters
E/A <1
E/A >1
P value
CO 15 to 30
3.9
4.8
0.20
E/e’ <1
E/e’ >1
CO 15 to 30
3.92
5.6
0.02 Table 1 (abstract P108). CO diff erence associated with diastolic dysfunction
parameters y
gi
Results HR, SAP, SV, CO, PV and FTc showed a statistically signifi cant
decrease during and right after RM compared with the baseline
values but they were gradually restored to control values after RM
discontinuation (Table 1). Conclusion The recruitment manoeuvre can compromise left diastolic
fi lling by increasing the transpulmonary pressure. In patients with
diastolic dysfunction a signifi cant decrease in SV can be observed
during the recruitment manoeuvre with no compensatory mechanisms
evoked determining a signifi cant decrease in CO. The E/A ratio is not
able to discriminate between the two groups, confi rming the power of
tissue Doppler imaging to recognize the correct diastolic pattern. Reference Table 1 (abstract P107)
T1
T2
T3
T3
HR (beats/minute)
69.8±7.1
60.2±10.6*
67.1±7.4*^
68.4±6.9^
SAP mean (mmHg)
89.8±7.7
53.4±5.7*
81.5±7.7*^
87.6±6.7^
SV (ml)
82.2±21.2
22.5±15.5*
75.4±21.7*^
79.6±6.4^
CO (l/minute)
5.75±1.7
1.4±0.9*
5.1±1.7*^
5.5±1.5^
PV (cm/second)
72.6±17.1
26±15.9*
68±16.8*^
70.7±15.9^
FTc (seconds)
0.4±0.06
0.19±0.08*
0.36±0.06*^
0.36±0.06^
*P <0.05 versus T1. ^P <0.05 versus T2. 1. Reference Gernoth C, et al.: Respiratory and haemodynamic changes during
decremental open lung positive end-expiratory pressure titration in
patients with acute respiratory distress syndrome. Crit Care 2009, 13:R59. P109 Alveolar Recruitment for ARDS Trial: preliminary results
AB Cavalcanti1, EA Suzumura1, M Abreu1, GF Ribeiro1, A Kodama1,
F Moreira1, HP Guimarães1, E Romano1, MB Amato2, O Berwanger1,
CR Carvalho2, ART Investigators1
1Hospital do Coração – HCor, São Paulo, Brazil; 2Universidade de São Paulo,
Brazil
Critical Care 2013, 17(Suppl 2):P109 (doi: 10.1186/cc12047) Conclusion According to our results, RM application causes a profound
impairment of the cardiovascular system. This can be attributed to the
increase of the airway pressures, which results in preload decrease
and afterload increase of the right ventricle and cardiac contractility
attenuation. Nevertheless, these eff ects are transient and reversible
and RM application is a safe technique for the management of
intraoperative hypoxemia, provided that adequate cardiac preload is
ensured and the anesthesiologist is alert to discontinue the RM if it
exceeds cardiovascular reserves. Critical Care 2013, 17(Suppl 2):P109 (doi: 10.1186/cc12047 Introduction The aim of the ongoing Alveolar Recruitment for ARDS
Trial (ART) is to evaluate whether a maximum stepwise alveolar recruit-
ment maneuver (MSARM) followed by ventilation at optimal PEEP may
decrease 28-day mortality in patients with moderate to severe ARDS
compared with ventilation with the ARDSNet strategy (NCT01374022). Here we report the results of a planned a priori preliminary analysis
involving 100 patients to assess feasibility, physiologic variables and
safety outcomes. P108 P108
Haemodynamic changes during alveolar recruitment manoeuvre in
patients with diastolic dysfunction
A Rubino, E Marini, B Ferro, M Collareta, F Forfori, F Guarracino, F Giunta
Azienda Ospedaliero Universitaria Pisana, Pisa, Italy
Critical Care 2013, 17(Suppl 2):P108 (doi: 10.1186/cc12046) y
Methods ART is an event-driven multicenter randomized controlled
trial planned to last until 520 deaths within 28 days are observed. Patients assigned to the experimental group receive a MSARM
achieving PEEP of 45 cmH2O and plateau pressure of 60 cmH2O plus
PEEP titrated according to the static compliance of the respiratory
system (ART strategy). The target tidal volume is 4 to 6 ml/kg predicted
body weight (PBW) and plateau pressure ≤30 cmH2O in both groups. Results We randomized 101 patients between November 2011 and
October 2012 in 51 centers. Considering 124 active sites in July 2013
and the current recruitment rate, we will fi nish enrollment by November
2014. MSARM was complete in 34/47 (72%) of patients allocated to the P112f Eff ect of protective ventilation on organ-specifi c cytokine
production in an experimental postoperative sepsis model
J Sperber1, M Lipcsey2, A Larsson3, A Larsson2, J Sjölin3, M Castegren1
1Centre for Clinical Research Sörmland, Uppsala University, Uppsala, Sweden;
2Surgical Sciences, Uppsala University, Uppsala, Sweden; 3Medical Sciences,
Uppsala University, Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P112 (doi: 10.1186/cc12050) Methods We prospectively evaluated 270 patients immediately after
cardiac surgery, presenting hypoxemia and PaO2/FiO2 <250. Patients
were randomized to an intensive alveolar recruitment maneuver (ARM)
or a moderate ARM strategy. Intensive ARM group: recruitment with
an inspiratory pressure amplitude of 15 cmH2O and PEEP of 30 cmH2O,
followed by ventilation with PEEP = 13 cmH2O, during 4 hours of
protective mechanical ventilation with VT = 6 ml/kg/pbw. Moderate
ARM group: recruitment with opening pressures of 20 cmH2O in the
airways, followed by ventilation with PEEP = 8 cmH2O, during 4 hours of
protective mechanical ventilation with VT = 6 ml/kg/pbw. The primary
outcome was a composite endpoint of severe pulmonary complications
in the postoperative period defi ned as intra-hospital death, need for
mechanical ventilation for more than 48 hours after surgery, pulmonary
infection or after reintubation within 28 days after randomization. The
secondary outcome was the incidence of nonpulmonary complications
as postoperative myocardial ischemia, acute renal failure (RIFLE-R),
respiratory mechanics and blood gas analysis after ARM, ICU length of
stay, hospital length of stay and 30-day mortality. Introduction Low tidal volume (VT) ventilation in intensive care
patients without lung injury attenuates the systemic infl ammatory
response [1]. The contribution of the specifi c organ infl ammatory
responses to the systemic picture remains to be elucidated. We
investigated the eff ect of low VT ventilation compared with medium
high VT on hepatic, splanchnic and cerebral cytokine responses in an
experimental large animal postoperative sepsis model. Methods Twenty pigs, group Protective Ventilation (PV), were ventilated
with low VT (6 ml/kg) and PEEP 10 cmH2O while 10 pigs, group Control
(C), were ventilated with a VT of 10 ml/kg and PEEP 5 cmH2O. Catheters
were introduced into an artery, the jugular bulb, the hepatic vein and
the portal vein. Laparotomy for 2 hours simulated a surgical procedure
after which baseline ensued and a continuous endotoxin infusion was
started at 0.25 μg/kg/hour for 5 hours. Diff erences were analyzed with
ANOVA for repeated measures. 1.
Amato MB, et al.: N Engl J Med 1998, 338:347-354. P110 0
Intensive alveolar recruitment protocol reduces pulmonary
complications and intensive care permanence after cardiac surgery
A Leme1, L Hajjar1, E Nozawa1, C Hashizume1, J Almeida1, J Fukushima1,
J Auler Jr1, R Santiago1, R Ianotti1, M Amato2, E Osawa1, M Feltrim1, F Galas1
1Heart Institute, São Paulo, Brazil; 2Faculdade de Medicina da Universidade de
São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P110 (doi: 10.1186/cc12048) Conclusion The reliability of pressure measurements and also of
compliance estimation via the tested catheters is high. Only in
two catheters was the fi lling volume a critical point for a precise
measurement of pressure or for estimation of compliance. Immediately
after unpacking, adhesion of the balloon material might prevent
reliable pressure measurement, therefore before the fi rst measurement
overfi lling of the balloon and retention of the excess gas seems strongly
recommended. Introduction Cardiac surgical procedures are associated with a high
incidence of postoperative complications, increasing costs and
mortality. The purpose of this study is to evaluate prospectively the
impact of two protective mechanical ventilation strategies, both using
low-tidal volume ventilation (6 ml/kg/ibw) after cardiac surgery. Haemodynamic changes during alveolar recruitment manoeuvre in
patients with diastolic dysfunction Introduction To investigate the haemodynamic eff ects of an alveolar
recruitment manoeuvre in intubated patients with diastolic dysfunction
compared with patients with normal diastolic function in a mixed ICU. Methods Between October 2011 and February 2012, 16 mechanically
ventilated patients admitted to the ICU with normal systolic function 2
Results We randomized 101 patients between November 2011 and
October 2012 in 51 centers. Considering 124 active sites in July 2013
and the current recruitment rate, we will fi nish enrollment by November
2014. MSARM was complete in 34/47 (72%) of patients allocated to the S41 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 ART group. The main reason for MSARM interruption was hypotension
(8/9 (90%)). Mean titrated PEEP in the ART group was 16.1 ± 3.5 versus
12.9 ± 3.4 cmH2O in ARDSNet (P <0.001). One hour after randomization,
tidal volume was similar between the ART and ARDSNet groups
(5.1 ± 0.8 vs. 5.3 ± 0.7 ml/kg PBW, respectively; P = 0.08). Mean values
remained below 6 ml/kg PBW up to day 3. Few patients had plateau
pressure >30 cmH2O 1 hour after randomization in the ART and
ARDSNet groups (3/43 (7.0%) vs. 4/47 (8.5%), respectively; P = 1.00) and
on subsequent days. PaO2/FiO2 was signifi cant higher in the ART group
(179.5 ± 84.5 vs. 143.3 ± 46.8, P = 0.01) and increased over time up to
7 days after randomization (272.3 ± 136.5 vs. 192.6 ± 72.3, P = 0.003). The ART strategy did not increase the risk of barotrauma (relative risk
(RR) = 0.78, 95% CI = 0.19 to 3.30) in the fi rst 7 days after randomization
or the need to initiate or increase vasopressors or mean arterial
pressure <65 mmHg (RR = 1.14, 95% CI = 0.65 to 2.02, P = 0.67) 1 hour
after randomization. However, the ART strategy increased the risk for
severe acidosis (pH <7.10) 1 hour after randomization (RR = 3.20, 95%
CI = 1.12 to 9.20, P = 0.03). Reliability of pressure measurements via balloon catheters is high:
an evaluation of six esophageal catheters Reliability of pressure measurements via balloon catheters is high:
an evaluation of six esophageal catheters p
g
S Walterspacher, L Isaak, HJ Kabitz, J Guttmann, S Schumann
University Medical Center Freiburg, Germany
Critical Care 2013, 17(Suppl 2):P111 (doi: 10.1186/cc12049) y
g
y
Critical Care 2013, 17(Suppl 2):P111 (doi: 10.1186/cc12049) Introduction Reliable measurement of esophageal pressure is a
prerequisite for analysis of respiratory system mechanics in spontaneous
breathing patients. For that purpose, various types of balloon catheters
exist that diff er in material, size and shape. In physical models we
studied the quality of pressure measurement via six diff erent balloon
catheters, three of them containing a second balloon for measurement
of gastric pressure. g
p
Methods Nine balloons of six esophageal catheters were investigated
in three conditions: measurement of balloon pressure during initial
infl ations immediately after unpacking; measurement of static pressures
at diff erent fi lling volumes; and compliance estimation in physical
models (27, 54, 90 ml/cmH2O) at diff erent levels of superimposed
pressure. Conclusion ART is feasible. The incidence of adverse events was similar
between groups except for severe acidosis 1 hour after randomization. Hence we adjusted the study protocol, increasing the respiratory rate
(from 10 to 15/minute) during MSARM. Results During the initial infl ation most catheters showed pressure
artifacts resulting from material adhesion. Those artifacts disappeared
during following infl ations. Static pressure measurements could
be performed with an error below 1 cmH2O if the balloon was fi lled
appropriately. Overfi lling of the balloon resulted in larger errors only in
two catheters. Compliance estimations resulted in errors below 1 ml/
cmH2O. Superimposed pressure had no relevant eff ect on compliance
estimation. P112f y
y
y
y
Results The intensive ARM group compared with the moderate ARM
group had lower incidence of the primary outcome, mainly due to
the reduced rate of pulmonary infection (2.3% vs. 10.1%, P = 0.009). Moreover, the intensive ARM group presented higher lung compliance
(68 ± 19 vs. 51 ± 17 ml/cmH2O, P <0.001) and PaO2/FiO2 ratio (360 ± 68
vs. 240 ± 74, P <0.001) after intervention when compared with the
moderate group. Also, the intensive ARM presented a lower length of
ICU stay (3 days vs. 4 days, P = 0.027) than the moderate ARM. There are
no diff erences regarding severe nonpulmonary complications and 30-
day mortality between groups. Results TNFα levels were higher in the hepatic vein than in the artery,
the jugular bulb and the portal vein. IL-6 levels were higher in the
artery and the jugular bulb compared with the portal and hepatic
veins. IL-10 levels were higher in the portal vein compared with the
jugular bulb and hepatic vein. The organ-specifi c IL-10 concentrations
were all higher than the arterial concentration. Comparison between
the ventilation groups showed that TNFα, IL-6 and IL-10 in the hepatic
vein were higher in group C compared with group PV at the end of
the experiment. Peak concentrations of TNFα and IL-6 in the portal vein
were higher in group C compared with group PV. Conclusion An intensive ARM strategy reduces postoperative pulmo-
nary complications, reduces hypoxemia, increases lung compliances
and decreases the length of ICU stay after cardiac surgery. Reference S42 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods Mechanically ventilated patients (PEEP 10 cmH2O and TV
7 ml/kg – Baseline) were randomized to two groups: in the PEEP 15
group, PEEP was increased from 10 to 15 cmH2O; while in the PEEP 5
group, PEEP was decreased from 10 to 5 cmH2O. Arterial gas analyses
were performed in both groups after 5, 15, 30 and 60 minutes from the
change of PEEP. Conclusion In this experiment TNFα was mainly generated in the
liver while the results point to signifi cant nonhepatic IL-6 and IL-10
production. Ventilation with low VT and medium-high PEEP attenuated
hepatic and splanchnic cytokine production compared with medium-
high VT and lower PEEP. Reference 1. N Engl J Med 2006, 354:1775-1786. 1. N Engl J Med 2006, 354:1775-1786. 1. N Engl J Med 2006, 354:1775-1786. P112f Reference g
Results We enrolled 44 ARDS patients: 23 in the PEEP 15 group and 21
in the PEEP 5 group. At Baseline, PaO2/FiO2 (P/F) and PaCO2 were similar
in both groups (P/F 169.5 ± 78.8 vs. 165.4 ± 80.6; PaCO2 45.6 ± 8.5 vs. 41.7 ± 5.0 mmHg, PEEP 15 vs. PEEP 5, respectively). In the PEEP 15
group, P/F signifi cantly continuously increased over time. In PEEP 5,
P/F signifi cantly decreased after 5 minutes and remained stable over
time. In the PEEP 15 group, PaCO2 did not change within 60 minutes
after PEEP increase. When PEEP was reduced (PEEP 5) PaCO2 remained
stable for the fi rst two steps, while at 30 and 60 minutes PaCO2 was
signifi cantly higher than at Baseline (Figures 1 and 2). 1. Determann et al.: Crit Care 2010, 14:R1. doi:10.1186/cc8230 P114 Comparison between the standard and low-dose chest CT scans on
the lung quantitative analysis in critically ill patients. S Froio1, D Chiumello1, I Cigada2, M Brioni2, S Coppola1, F Menga2,
L Gattinoni2
1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P114 (doi: 10.1186/cc12052) Comparison between the standard and low-dose chest CT scans on
the lung quantitative analysis in critically ill patients. S Froio1, D Chiumello1, I Cigada2, M Brioni2, S Coppola1, F Menga2,
L Gattinoni2 Comparison between the standard and low-dose chest CT scans on
the lung quantitative analysis in critically ill patients. S Froio1, D Chiumello1, I Cigada2, M Brioni2, S Coppola1, F Menga2,
L Gattinoni2 Figure 2 (abstract P113). Two-way ANOVA RM. P <0.05, #versus
5 minutes. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Comparison between the standard and low do
the lung quantitative analysis in critically ill pat
S Froio1, D Chiumello1, I Cigada2, M Brioni2, S Coppo
L Gattinoni2
1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore P
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P114 (doi: 10.1186/cc
Introduction Quantitative analysis of a lung C
the reference method to study lung recruitab
ventilatory strategy in ARDS patients. However
radiation exposure, especially when CT analysis is
the evolution of ARDS. The aim of this study was t
of a lower radiation dose on lung CT quantitati
with analysis computed by standard CT. Methods Sedated and paralyzed patients und
scans: a Standard CT (120 kV, 110 mAs, pitch 1.2
Care Dose Technology) and a CT performed by a
(Lowdose: 120 kV, 60 mAs). CTs were perform
inspiratory or expiratory hold, for diff erent valu
Each CT image was manually delineated exclud
and mediastinal structures. We analyzed Stand
images with dedicated software to quantify
lung regions with diff erent degrees of infl atio
data computed by Standard and by Lowdose s
according to Bland–Altmann analysis. Figure 2 (abstract P113). Two-way ANOVA RM. P <0.05, #versus
5 minutes. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Figure 1 (abstract P114). Total tissue weight. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Time required for gas exchange equilibration after a change of
positive end-expiratory pressure in acute respiratory distress
syndrome y
S Coppola1, D Chiumello1, F Menga2, M Brioni2, I Cigada2, S Froio1
1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P113 (doi: 10.1186/cc12051) i
y
Conclusion Our data indicate that it is important in critically ill
patients to allow suffi cient time for the full eff ect of PEEP increase on
oxygenation and to prevent excessive delay when P/F decrease occurs
following the application of a lower level of PEEP. Introduction Positive end-expiratory pressure (PEEP) is fundamental
to prevent lung collapse in ARDS patients. A common method to
titrate PEEP is to perform a PEEP test, recording the variation of cardio-
respiratory parameters after a PEEP change [1]. The aim of this study
was to evaluate the time-course changes of gas exchanges following
a PEEP test. P116 P116
Feasibility and eff ectiveness of prone position in morbidly obese
ARDS patients: a case–control clinical study
A De Jong, N Molinari, M Sebbane, A Prades, S Jaber
Montpellier University Hospital, Montpellier, France
Critical Care 2013, 17(Suppl 2):P116 (doi: 10.1186/cc12054) Introduction Obese patients are at risk of developing atelectasis and
acute respiratory distress syndrome (ARDS) [1]. The prone position
(PP) may reduce atelectasis, and improves oxygenation and outcome
in severe hypoxemic patients in ARDS [2], but little is known about its
eff ect in obese ARDS patients. Results We enrolled 13 patients admitted to our ICU. In the Bland–
Altman analysis of the lung total tissue and not infl ated tissue, the
bias and agreement bands for Standard CT scan and Lowdose CT
quantitative analysis were –16.97 g (–77.43 to +43.49 g) and –9.65 g
(–116.81 to 77.51 g), respectively (Figures 1 and 2). Results We enrolled 13 patients admitted to our ICU. In the Bland–
Altman analysis of the lung total tissue and not infl ated tissue, the
bias and agreement bands for Standard CT scan and Lowdose CT
quantitative analysis were –16.97 g (–77.43 to +43.49 g) and –9.65 g
(–116.81 to 77.51 g), respectively (Figures 1 and 2). f
Methods Morbidly obese patients (body mass index (BMI) ≥35 kg/m2) in
PP with ARDS (PaO2/FiO2 ratio ≤200 mmHg) were matched to nonobese
(BMI <30 kg/m2) ARDS patients in a case–control clinical study. The
primary endpoints were safety and complications of PP; the second
endpoints were the eff ect on oxygenation (PaO2/FiO2 ratio at the end
of PP), length of mechanical ventilation and ICU stay, nosocomial
infections and mortality. g
p
y
g
Conclusion Lung CT quantitative analysis computed by Lowdose
CT scans could be a useful tool for monitoring and ventilatory
management of ARDS patients. y
Results Between January 2005 and December 2009, 149 patients were
admitted for ARDS. Thirty-three obese patients were matched with
33 nonobese patients. Median PP duration was 9 (6 to 11) hours in
obese patients and 8 (7 to 12) hours in nonobese patients (P = 0.28). We collected 51 complications, of which 25 in obese patients and
26 in nonobese patients. The number of patients with at least one
complication was similar across groups (n = 10, 30%). P114 1Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P114 (doi: 10.1186/cc12052) Introduction Quantitative analysis of a lung CT scan is considered
the reference method to study lung recruitability to optimize the
ventilatory strategy in ARDS patients. However, CT implies a risk of
radiation exposure, especially when CT analysis is necessary to monitor
the evolution of ARDS. The aim of this study was to evaluate the impact
of a lower radiation dose on lung CT quantitative analysis compared
with analysis computed by standard CT. y
p
y
Methods Sedated and paralyzed patients underwent two chest CT
scans: a Standard CT (120 kV, 110 mAs, pitch 1.2, collimation 0.6 mm;
Care Dose Technology) and a CT performed by a lower radiation dose
(Lowdose: 120 kV, 60 mAs). CTs were performed during the same
inspiratory or expiratory hold, for diff erent values of airway pressure. Each CT image was manually delineated excluding pleural eff usions
and mediastinal structures. We analyzed Standard and Lowdose CT
images with dedicated software to quantify the tissue weight of
lung regions with diff erent degrees of infl ation. Lung quantitative
data computed by Standard and by Lowdose scans were compared
according to Bland–Altmann analysis. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Figure 1 (abstract P113). Two-way ANOVA RM. P <0.05, *versus Time 0,
#versus 5 minutes, °versus 15 minutes. Figure 2 (abstract P113). Two-way ANOVA RM. P <0.05, #versus
5 minutes according to Bland Altmann analysis. Figure 2 (abstract P113). Two-way ANOVA RM. P <0.05, #versus
5 minutes. Figure 1 (abstract P114). Total tissue weight. Figure 1 (abstract P114). Total tissue weight. Critical Care 2013, Volume 17 Suppl 2
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pp
http://ccforum.com/supplements/17/S2 therapy was higher in APRV than the CMV group (72% vs. 49%,
respectively, P <0.0001). The mortality rate was signifi cantly higher in
CMV (n = 46, 51%) as compared with APRV (n = 11, 29%) (P = 0.022). Conclusion The use of APRV in septic shock patients restores
hemodynamic stability earlier than the CMV mode. There was a
signifi cant improvement in ICU survival using APRV over CMV. P114 Early
initiation of APRV in ventilated septic shock patients was associated
with a decrease in ICU mortality. Figure 2 (abstract P114). Not infl ated tissue weight. Figure 2 (abstract P114). Not infl ated tissue weight. P116 The PaO2/FiO2 ratio
(Figure 1) increased signifi cantly more in obese patients (from 118 ± 43
to 222 ± 84 mmHg) than in nonobese patients (from 113 ± 43 mmHg
to 174 ± 80 mmHg, P = 0.03). Length of mechanical ventilation, ICU stay
and nosocomial infections did not diff er signifi cantly, but mortality at
90 days was signifi cantly lower in obese patients (27 vs. 48%, P <0.05). 1. N Engl J Med 2006, 354:1775-1786. Benefi cial eff ects of intrapulmonary percussive ventilation in
patients with respiratory insuffi ciency in the ICU I Blum, R Janssen-Dean, A Overdijk van, B Speelberg
St Anna Hospital Geldrop, Geldrop, the Netherlands
Critical Care 2013, 17(Suppl 2):P119 (doi: 10.1186/cc12057) I Blum, R Janssen-Dean, A Overdijk van, B Speelberg
St Anna Hospital Geldrop, Geldrop, the Netherlands
Critical Care 2013, 17(Suppl 2):P119 (doi: 10.1186/cc12057) l
Methods A case–control study was conducted following ethics
approval. A case was defi ned as a ventilated pregnant/postpartum
woman reported to the Australian and New Zealand INFINITE H1N1 09
study from 1 June 2009 to 31 August 2009. Controls were ventilated
nonpregnant women (15 to 49 years) reported to the INFINITE H1N1
09 study during the same time frame. Data were entered into SPSS
and analysed using nonparametric statistics; two-tailed P <0.05 was
considered signifi cant. Introduction Intrapulmonary percussive ventilation (IPV) is a therapy
that is used to clear endobronchial secretions. The IPV ventilator was
designed and developed by FM Bird in 1979. The ventilator consists
of a phasitron that delivers rapid, high-fl ow, mini bursts of oxygen,
mixed with air. The potential aims of this mechanism are pulmonary
recruitment, improved mucus clearance with a direct high-frequency
oscillatory eff ect. We investigated whether IPV has a positive eff ect on
ventilatory values in adult patients on the ICU. i
Results We studied 36 index cases and 38 controls. Index cases were
more likely to have a single arterial blood gas (ABG) taken prior to
intubation (P <0.05). Similar reasons were given for the trigger to
intubate (high respiratory rate, low PaO2, increased work of breathing)
apart from a high PaCO2, which was a trigger in the control group only
(P <0.05). There were no diff erences in the pre-intubation and post-
intubation ABGs apart from a lower PaCO2 (P <0.05) and lower HCO3
(P <0.05) in cases, and cases presented with a lower haemoglobin
(P <0.05). There were six diffi cult intubations documented with no
diff erences between groups. Initial ventilator settings including mode,
tidal volume, minute volume and respiratory rate demonstrated no
diff erences. Both groups showed increases in PaO2 and PaCO2, and
a decrease in pH from the pre-intubation to post-intubation ABGs
(P <0.05).f y
p
Methods All patients presenting during a 4-month period in 2011 with
respiratory insuffi ciency on our mixed adult ICU were included in this
study. Patients were monitored before, directly after and 15 minutes
after therapy with IPV using a Bird Intrapulmonary Ventilator Model
IPV-2C. Preliminary experience with airway pressure release ventilation on
hemodynamics in patients with septic shock in a medical/surgical
ICU A Hussain, H Lababidi, A Mir, A AlHamoud, A Al Oheli, A Al Enezi
King Fahd Medical City, Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P115 (doi: 10.1186/cc12053) Introduction Airway pressure release ventilation (APRV) allows
spontaneous breathing throughout the ventilation cycle. It increases
venous return and cardiac index, which will signifi cantly improve
organ perfusion. This is important in septic shock patients to prevent
extrathoracic organ system failure secondary to poor perfusion. Benefi ts of APRV with cardiovascular changes are noticed in patients
with acute lung injury and acute respiratory distress syndrome. It is
not well established whether applying APRV will improve the survival
outcome for septic shock patients. The primary outcome is whether the
use of APRV in septic shock patients restores hemodynamic stability
earlier than the CMV mode. The secondary hypothesis is whether the
use of APRV in septic shock patients improves their survival in the ICU. Methods After Institutional Review Board approval, we retrospectively
analyzed the clinical data of 129 septic shock patients who received
ventilator support between January and December 2011 at a tertiary
care hospital. The Cox proportional hazards model was used in
adjusting potential confounding factors. The nonparametric Wilcoxon
rank sum test was used to assess signifi cant outcome diff erences
between groups. Time to event/survival data will be analyzed using
Kaplan–Meier methods. These analyses were accomplished using SAS,
version 9.3. Figure 1 (abstract P116). Individual variations of PaO2/FiO2 ratio between
supine and prone positions in obese and nonobese patients. Results Among the 187 patients, 58 were excluded as per the exclusion
criteria: incomplete data (n = 28), do not resuscitate (n = 16), ICU
readmission (n = 12) and head injury (n = 4). Finally, 129 patients were
included, from these 91 received CMV and 38 received APRV. At the
beginning of the study, there were no diff erences between the groups
in relation to hemodynamic parameters. Reversal of shock achieved
in less than 72 hours was statistically signifi cant between the groups
(APRV, n = 16 (42%) and CMV, n = 8 (9%), P = 0.0101). The proportion
of patients recovering from septic shock after initiation of ventilator Figure 1 (abstract P116). Individual variations of PaO2/FiO2 ratio between
supine and prone positions in obese and nonobese patients. S44 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 years. Provision of mechanical ventilation to pregnant/postpartum
women with H1N1 infl uenza: a case–control study l
WE Pollock1, R Bellomo2, S Webb3, I Seppelt4, A Davies5, E Sullivan6,
S Morrison7, B Howe7 1Mercy Hospital for Women, Heidelburg, Australia; 2Austin Health, Heidelberg,
Australia; 3Royal Perth Hospital, Perth, Australia; 4Nepean Hospital, Penrith,
Australia; 5Alfred Hospital, Prahran, Australia; 6University of New South Wales,
Randwick, Australia; 7Monash University, Prahran, Australia
Critical Care 2013, 17(Suppl 2):P117 (doi: 10.1186/cc12055) Conclusion In this study, inhalation injury is common in burn patients. They had higher severity scores (APACHE II, ABSI) and higher mortality. These patients had a higher need for mechanical ventilation and
lower PaO2/FiO2 ratio, but there was no signifi cant increase in ARDS or
respiratory sepsis. Introduction During the infl uenza pandemic of 2009, clinicians
delivered mechanical ventilation to pregnant women with little
evidence to guide practice. The objective of this study was to compare
the provision of mechanical ventilation to pregnant/postpartum
women and a nonpregnant matched control group admitted to the
ICU with H1N1 infl uenza. P120 P120
High-frequency ventilation for acute traumatic and nontraumatic
lung injury
R Varutti1, R Bigai1, M Fiorillo1, D Tomasello1, W Mercante1, G Trillò2
1Azienda Ospedaliera Santa Maria degli Angeli, Pordenone, Italy; 2Udine
University Hospital, Udine, Italy
Critical Care 2013, 17(Suppl 2):P120 (doi: 10.1186/cc12058) Preliminary experience with airway pressure release ventilation on
hemodynamics in patients with septic shock in a medical/surgical
ICU The mean total burn surface area (TBSA) was 28% (± 25). Forty-
fi ve patients (53.5%) had TBSA >20%. These patients had higher severity
scores: ABSI 8 (± 2.8) versus 6 (± 2.2) (P = 0.0001), APACHE II 13 (± 6.9)
versus 7 (± 6.7) (P = 0.0001). They also had a higher need for mechanical
ventilation (88.1% vs. 34.9%, P = 0.001) and shock (59.5% vs. 27.0%, P =
0.001). The PaO2/FiO2 ratio at admission with and without inhalation
was 245 versus 359 (P = 0.003), and at 72 hours was 207 versus 306
(P = 0.002). There were no signifi cant diff erences in the appearance
of sepsis, ARDS and renal failure. The length of stay with and without
inhalation was 25.2 (1 to 95) days versus 22.5 (1 to 92) days (P = 0.4). The
inhalation injury group showed a signifi cantly higher mortality (9.0%
vs. 28.9%, P = 0.001). Conclusion PP seems safe in obese patients and may improve
oxygenation more than in nonobese patients. Obese patients could be
a subgroup of ARDS patients who may benefi t most from PP. References
1. Gong MN, et al.: Thorax 2010, 65:44-50. 2. Charron C, et al.: Intensive Care Med 2011, 37:785-790. References
1. Gong MN, et al.: Thorax 2010, 65:44-50. 2. Charron C, et al.: Intensive Care Med 2011, 37:785-790. 1. Gong MN, et al.: Thorax 2010, 65:44-50. 2. Charron C, et al.: Intensive Care Med 2011, 37:785-790. Benefi cial eff ects of intrapulmonary percussive ventilation in
patients with respiratory insuffi ciency in the ICU All patients received IPV for a period of 20 minutes consisting
of two cycles of 10 minutes. Peripheral oxygen saturation (SpO2), tidal
volume (Vt), respiratory rate, end-tidal CO2 (ET-CO2), dynamic lung
compliance (C-dyn) and work of breathing (WOB) were monitored
at the diff erent time points. Paired Student t tests were performed in
order to compare the values immediately before IPV, with directly after
therapy and 15 minutes later. P <0.05 was considered signifi cant.i yi
Results Eighty-three patients were examined. SpO2 improved signifi -
cantly from 93.7 ± 3.7 before IPV to 95.7 ± 2.8 after IPV (P <0.001) and
15 minutes later to 95.2 ± 2.8 (P <0.001). Vt improved from 418 ± 111
before IPV to 476 ± 102 directly after (P <0.001) and to 480 ± 131
15 minutes later (P <0.01). Respiratory rate improved from 24 ± 6 to
23 ± 6 only after 15 minutes signifi cantly (P <0.01). WOB and C-dyn did
not change. ET-CO2 decreased from 34.9 ± 14.8 to 33.3 ± 13.3 (P <0.05)
directly after IPV and to 32.1 ± 12.8 (P <0.01) 15 minutes later.if Conclusion There were physiological diff erences between the two
groups with pregnant/postpartum women showing lower PaCO2 and
HCO3. However, initial ventilator support was not signifi cantly diff erent
for pregnant/postpartum women compared with controls. Reference p
p
Critical Care 2013, 17(Suppl 2):P118 (doi: 10.1186/cc12056) 1. Salim A, Martin M: High-frequency percussive ventilation. Crit Care Med
2005, 33:S241-S245. Introduction The objective is to analyze the epidemiology and
mortality of critical burn patients with inhalation injury. Introduction The objective is to analyze the epidemiology and
mortality of critical burn patients with inhalation injury. Methods A prospective, observational and descriptive study was
carried out in all patients admitted to an ICU from October 2008 to June
2011. Inhalation injury was defi ned with two or more of the following
criteria: history of injury in an enclosed space, facial burns with singed
nasal hair, carbonaceus sputum or stridor. If they were intubated it
was diagnosed by bronchoscopy. Demographic data, length of stay,
ABSI, APACHE II, duration of mechanical ventilation, hospital course
and mortality data were collected. Data are presented as number and
percentage or as median and interquartile range and were analyzed
with the Fisher exact test and Mann–Whitney test. P119i Benefi cial eff ects of intrapulmonary percussive ventilation in
patients with respiratory insuffi ciency in the ICU
I Blum, R Janssen-Dean, A Overdijk van, B Speelberg
St Anna Hospital Geldrop, Geldrop, the Netherlands
Critical Care 2013, 17(Suppl 2):P119 (doi: 10.1186/cc12057) Inhalation injury in critical burn patients
L C
h f
M S
h
E H
L F
á d Conclusion In this study we demonstrated a benefi cial eff ect of IPV on
oxygen saturation, tidal volume and end-tidal CO2. IPV has this eff ect in
addition to the mobilization of bronchial secretions. Reference P120
High-frequency ventilation for acute traumatic and nontraumatic
lung injury high-frequency oscillatory ventilation (HFOV) was considered mostly
a rescue therapy. py
Methods Fifteen adult patients admitted to our ICU for acute
traumatic and nontraumatic lung injury were submitted to HFOV when
conventional mechanical ventilation failed. Acknowledgement This research has been cofi nanced by the European
Union and Greek national funds through the operating programme
‘Education and Lifelong Learning’ – Research Funding Programme
Heracleitus II. Results Clinical and demographic data are shown in Table 1. Figure 1
shows the trend of gas parameters during the recovery. At baseline
PaO2 was 94 ± 28 mmHg; after 6 hours of HFOV: 135 ± 41 mmHg,
P <0.01. At baseline PaO2/FiO2 was 182 ± 97 mmHg; after 6 hours of
HFOV: 264 ± 101 mmHg, P <0.01. The benefi ts are maintained when
returned to conventional ventilation. Reference 1. Zeravik J, et al.: Acta Anaesthesiol Scand 1989, 33(Suppl 90):149-152. Figure 1 (abstract P121). Scatter plot and fi t line of ELWI versus
dPaO2/FiO2. Conclusion HFOV may therefore be anticipated to improve end-
organ perfusion and gas exchange; it should be considered in severe
traumatic and nontraumatic respiratory failure [1]. 1. BMJ 2010, 340:c2327. 1. BMJ 2010, 340:c2327. P120
High-frequency ventilation for acute traumatic and nontraumatic
lung injury R Varutti1, R Bigai1, M Fiorillo1, D Tomasello1, W Mercante1, G Trillò2
1Azienda Ospedaliera Santa Maria degli Angeli, Pordenone, Italy; 2Udine
University Hospital, Udine, Italy
C i i
l C
2013 17(S
l 2) P120 (d i 10 1186/
12058) y
p
y
Critical Care 2013, 17(Suppl 2):P120 (doi: 10.1186/cc12058) Introduction ARDS is commonly observed in trauma patients. In
some instances the severity of the clinical presentation is such
that all conventional ventilatory support mode fails. In this setting, Results Of the 362 patients admitted, 84 (23.2%) had inhalation injury. Seventy-six percent were male and the average age was 52 (± 17.5) S45 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P120). Mean PaO2, PaCO2, PaO2/FiO2 at baseline and during HFOV (P, percussionator; C, conventional). *P <0.05 versus baseline, †P <0.01
versus baseline. Figure 1 (abstract P120). Mean PaO2, PaCO2, PaO2/FiO2 at baseline and during HFOV (P, percussionator; C, conventional). *P <0.05 versus baseline, †P <0.01
versus baseline. Figure 1 (abstract P120). Mean PaO2, PaCO2, PaO2/FiO2 at baseline and during HFOV (P, percussionator; C, conventional). *P <0.05 v
versus baseline. Table 1 (abstract P120). Clinical and demographic data
Mean
Age (years)
64
Male/female
11/4
Dead
4
Trauma
3
SAPS II
45/40
APACHE
23/8 Table 1 (abstract P120). Clinical and demographic data fl ow were 3.5 Hz, 85 to 95 cmH2O, and 40 l/minute, respectively; a 2.5 to
3.5 cmH2O tracheal tube cuff leak was used. HFO mean airway pressure
(mPaw) exceeded preceding conventional ventilation (CV)-mPaw by
7 to 13 cmH2O. PaO2/FiO2, lung mechanics, and hemodynamics were
documented during lung-protective CV (baseline) and 1 hour following
the initiation of HFO-TGI ventilation. PULSION PICCOplus v7.0 was used
for hemodynamic measurements including the ELWI.i y
g
Results Oxygenation (PaO2/FiO2) improved signifi cantly with HFO-
TGI compared with CMV (125.5 ± 54.7 vs. 195.6 ± 108.7, P <0.001). Changes in PaO2/FiO2 were positively correlated with ELWI at baseline
(Spearman’s ρ = 0.56, P = 0.016). See Figure 1. There were no signifi cant
changes in patients’ fl uid balance and hemodynamics including the
ELWI. Conclusion Estimation of the ELWI can help to predict the oxygenation
response of ARDS patients considered for HFO-TGI ventilation. The
possibility that HFO-TGI exerts an eff ect on pulmonary oedema needs
further investigation.i high-frequency oscillatory ventilation (HFOV) was considered mostly
a rescue therapy. P121 Improvement in oxygenation with high-frequency oscillatory
ventilation combined with tracheal gas insuffl ation is correlated to
the extravascular lung water index
CS Vrettou, S Malachias, SG Zakynthinos, SD Mentzelopoulos
Evaggelismos General Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P121 (doi: 10.1186/cc12059) Physiologic eff ects and regional ventilation of high-frequency
positive-pressure ventilation using a conventional ventilator in a
severe ARDS animal model associated with an inspiratory pause or
recruitment maneuvers RL Cordioli1, M Park2, M Amato3, S Gomes3, E Leite2, L Azevedo2
1Hopitaux Universitaire de Geneve, Geneva, Switzerland; 2Hospital Sirio
Libanes IEP, São Paulo, Brazil; 3Hospital das Clinicas LIM 09, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P122 (doi: 10.1186/cc12060) RL Cordioli1, M Park2, M Amato3, S Gomes3, E Leite2, L Azevedo2
1Hopitaux Universitaire de Geneve, Geneva, Switzerland; 2Hospital Sirio
Libanes IEP, São Paulo, Brazil; 3Hospital das Clinicas LIM 09, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P122 (doi: 10.1186/cc12060) i
g
Results Data displayed by VDR4® overestimated pulmonary pressures
by more than 10%, but were reliable for other parameters. During
ascent, an off set appeared for all respiratory parameters: Vt increased
by 59% and PmEI by 53% between 0 and 8,000 ft. During descent,
the off set was reversely directed with a 39% decrease in Vt and a 28%
decrease in PmEE between 8,000 and 0 ft. Modifying working pressure
adequately corrected PmEI and PmEE, but not Vt. In all cases, manually
correcting VDR4® parameters to their 0 ft level also corrected these
off sets. Multivariate analysis further established that, adjusting for
other parameters, Vt, PmEI and PmEE did practically not depend on
altitude. Oxygen consumption of the respirator was high, 25 l/minute
at 0 ft, and stable with altitude. It was reduced with percussive rate and
with FiO2. Introduction Protective mechanical ventilation (MV) in ARDS is based
on reduced stretch of pulmonary tissue, sometimes resulting in severe
hypoventilation that can be avoided when using high respiratory rate. High-frequency positive-pressure ventilation (HFPPV) has not been
fully explored, especially when associated with other strategies aiming
to avoid hypercapnia. y
Methods We induced ARDS in eight pigs by lung lavage with saline
plus 3 hours of injurious MV with low PEEP and high driving pressure
(DP). We then performed a recruitment maneuver (RM) followed
by PEEP titration using the amount of alveolar collapse in electrical
impedance tomography (EIT). Then stabilization during 1 hours with
tidal volume (VT) at 6 ml/kg, respiratory rate (RR) 35 breaths/minute
and PEEP selected with the PEEP-FiO2 table (ARMA study), which was
kept constant during two steps of HFPPV with a RR 60: one without
an inspiratory pause (HFPPV-60), and one with a pause of 30% of
inspiratory time (HFPPV-60 w/P30%). In another HFPPV step, we used
PEEP titrated with EIT after RM (HFPPV-60 w/RM). Recruitable volume is comparable in acute respiratory distress
syndrome and in healthy lungs CA Stahl1, K Moeller2, D Steinmann1, D Henzler3, S Lundin4, O Stenqvist4
1University Medical Center Freiburg, Germany; 2Biomedical Engineering,
Villingen-Schwenningen, Germany; 3Klinikum Herford, Germany;
4Sahlgrenska University Hospital, Gothenburg, Sweden
Critical Care 2013, 17(Suppl 2):P124 (doi: 10.1186/cc12062) Table 1 (abstract P122). Physiological variables
Variable
VT = 6, stabilization
HFPPV-60
HFPPV-60 w/RM
DP (cmH2O)
16 ± 2
16 ± 3
12 ± 2*
P/F ratio
95 ± 13
149 ± 60
246 ± 99*
*Tukey’s post-hoc analysis, P < 0.05 versus others. Table 1 (abstract P122). Physiological variables Introduction The application of PEEP is commonly used in acute
respiratory distress syndrome (ARDS) and has been shown to improve
oxygenation. To identify patients that most benefi t from the application
of PEEP, the discrimination of recruiters and nonrecruiters has been
postulated by Gattinoni and colleagues [1]. Recently, Dellamonica and
colleagues [2] presented a method to predict alveolar recruitment. We hypothesised that the amount of recruitable volume allows the
discrimination between ARDS patients and patients with healthy lungs
(HL). Conclusion HFPPV with a conventional mechanical ventilator is able
to maintain stable PaCO2 in clinically acceptable values, allowing
reductions in VT. HFPPV-60 w/RM and PEEP titration using EIT allowed
further physiologic benefi ts in a severe ARDS model. (
)
Methods We recalculated the recruited volume (RV) in 25 patients
with ARDS [3] according to the method proposed by Dellamonica and
colleagues during an incremental PEEP manoeuvre (PEEP increased
until the plateau pressure reached 45 cmH2O). RV was calculated as the
change in end-expiratory lung volume minus total respiratory system
compliance times the PEEP change (RV = ΔEELV – CTOT×ΔPEEP). For comparison, 12 patients with HL undergoing elective surgery in
general anaesthesia were measured using the same protocol. Results Both ARDS and HL patients exhibited typical P–V curves and
stepwise recruitment (Figure 1). By raising PEEP from 0 to 12 cmH2O,
ARDS patients recruited 331 ± 195 ml (mean ± SD) and HL patients
435 ± 43 ml. There was a strong correlation (R2 = 0.88) of the total RV
with the end-inspiratory volume at a plateau pressure of 45 cmH2O in
both groups; that is, recruitment was found to the same extent in both
groups (Figure 2). Methods We recalculated the recruited volume (RV) in 25 patients
with ARDS [3] according to the method proposed by Dellamonica and
colleagues during an incremental PEEP manoeuvre (PEEP increased
until the plateau pressure reached 45 cmH2O). g
References g
References
1. Velmahos GC, et al.: Chest 1999, 116:440-446. 2. Chung KK, et al.: Crit Care Med 2010, 38:1970-1977. References
1. Velmahos GC, et al.: Chest 1999, 116:440-446. 2. Chung KK, et al.: Crit Care Med 2010, 38:1970-1977. 2
Results HFPPV allowed reduction in PaCO2 levels: 81 (77, 94) versus
60 (58, 61), 59 (58, 60), 60 (58, 61) mmHg, besides using lower VT:
5.2 (5.0, 5.9), 5.1 (4.5, 6.0), 4.7 (4.2, 5.7) and 4.8 (4.5, 5.6) ml/kg during
stabilization, HFPPV-60, HFPPV-60 w/P30% and HFPPV-60 w/RM,
respectively. It had no signifi cant diff erent results comparing HFPPV-60
with and without an inspiratory pause. HFPPV-60 w/RM allowed a
better alveolar homogenization and improvement in oxygenation,
shunt, dead space and DP compared with the other steps. See Table 1. P124 Recruitable volume is comparable in acute respiratory distress
syndrome and in healthy lungs
CA Stahl1, K Moeller2, D Steinmann1, D Henzler3, S Lundin4, O Stenqvist4
1University Medical Center Freiburg, Germany; 2Biomedical Engineering,
Villingen-Schwenningen, Germany; 3Klinikum Herford, Germany;
4Sahlgrenska University Hospital, Gothenburg, Sweden
Critical Care 2013, 17(Suppl 2):P124 (doi: 10.1186/cc12062) Recruitable volume is comparable in acute respiratory distress
syndrome and in healthy lungs RV was calculated as the
change in end-expiratory lung volume minus total respiratory system
compliance times the PEEP change (RV = ΔEELV – CTOT×ΔPEEP). For comparison, 12 patients with HL undergoing elective surgery in
general anaesthesia were measured using the same protocol. Physiologic eff ects and regional ventilation of high-frequency
positive-pressure ventilation using a conventional ventilator in a
severe ARDS animal model associated with an inspiratory pause or
recruitment maneuvers During each HFPPV
step, VT was set to reach a PaCO2 of 60 ± 3 mmHg. Distribution of
regional ventilation was analyzed using EIT. Equilibrium was considered
if PaCO2 was stable (<5% of variation) for >30 minutes. 2
Conclusion HFPV can be safely used at altitude, provided that VDR4®-
displayed parameters are used to manually adjust settings in order to
avoid exposing patients to volutrauma or barotrauma during ascent,
and to major hypoventilation and alveolar collapse during descent. The
high oxygen consumption is currently the main limit to its use for long-
range aeromedical evacuations. Improvement in oxygenation with high-frequency oscillatory
ventilation combined with tracheal gas insuffl ation is correlated to
the extravascular lung water index HFPPV
better alveolar homogenization and improvem
shunt, dead space and DP compared with the ot
Table 1 (abstract P122). Physiological variables
Variable
VT = 6, stabilization
HFPPV-6
DP (cmH2O)
16 ± 2
16 ± 3
P/F ratio
95 ± 13
149 ± 60
*Tukey’s post-hoc analysis, P < 0.05 versus others. Conclusion HFPPV with a conventional mechan
to maintain stable PaCO2 in clinically accept
reductions in VT. HFPPV-60 w/RM and PEEP titra
further physiologic benefi ts in a severe ARDS mo
P123
High-frequency percussive ventilation at altitu
hypobaric chamber with a mechanical test lung
AC Cirodde1, S Montmerle2, ND Donat1, CB Bourhill
L Bargues1, T Leclerc1
1Military Hospital Percy, Clamart, France; 2IRBA, Brétig
Critical Care 2013, 17(Suppl 2):P123 (doi: 10.1186/c
Introduction High-frequency percussive ventilat
technique for most severe acute lung injury/acu
syndrome (ARDS) patients [1], especially with
respiratory burns [2]. This study aimed at ch
delivered by Percussionnaire VDR4® and at evalua
interferes with HFPV, in order to assess its usabili
Methods Using a mechanical test lung mimicki
17 ml/cmH2O) with two resistance levels (5 and
and ventilated with VDR4® in a hypobaric cham
between 0 and 5,000 and then 0 and 8,000 ft were P122 VDR4® parameters were modifi ed one at a time at each altitude. Besides
these parameters (cross-measured with standalone hardware), oxygen
consumption of the respirator and three calculated parameters were
studied: low-frequency tidal volume (Vt, integrated from instantaneous
fl ows measured with a Fleisch pneumotachograph), end-inspiratory
(PmEI) and end-expiratory (PmEE) mean pressures. PmEI and PmEE in
HFPV refl ect plateau pressure and positive end-expiratory pressure in
conventional ventilation. The correction of altitude-induced off set with
the modifi cation of working pressure was also tested. Physiologic eff ects and regional ventilation of high-frequency
positive-pressure ventilation using a conventional ventilator in a
severe ARDS animal model associated with an inspiratory pause or
recruitment maneuvers
RL Cordioli1, M Park2, M Amato3, S Gomes3, E Leite2, L Azevedo2
1Hopitaux Universitaire de Geneve, Geneva, Switzerland; 2Hospital Sirio
Libanes IEP, São Paulo, Brazil; 3Hospital das Clinicas LIM 09, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P122 (doi: 10.1186/cc12060) Physiologic eff ects and regional ventilation of high-frequency
positive-pressure ventilation using a conventional ventilator in a
severe ARDS animal model associated with an inspiratory pause or
recruitment maneuvers Improvement in oxygenation with high-frequency oscillatory
ventilation combined with tracheal gas insuffl ation is correlated to
the extravascular lung water index Introduction
High-frequency
oscillatory
ventilation
combined
with tracheal gas insuffl ation (HFO-TGI) can signifi cantly improve
oxygenation in patients with ARDS. It has been demonstrated that
oxygenation in patients with ARDS has a better response to HFO when
extravascular lung water is >15 ml/kg body weight (BW). Our aim is to
examine whether the extravascular lung water index (ELWI) correlates
with changes in PaO2/FiO2 in patients ventilated with HFO-TGI. Figure 1 (abstract P121). Scatter plot and fi t line of ELWI versus
dPaO2/FiO2. 2
2
Methods Data from 18 sessions of HFO-TGI in six patients were included
in the analysis. HFO frequency, oscillatory pressure amplitude, and bias Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S46 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P122
Physiologic eff ects and regional ventilation of h
positive-pressure ventilation using a conventio
severe ARDS animal model associated with an
recruitment maneuvers
RL Cordioli1, M Park2, M Amato3, S Gomes3, E Leite2,
1Hopitaux Universitaire de Geneve, Geneva, Switzerlan
Libanes IEP, São Paulo, Brazil; 3Hospital das Clinicas LIM
Critical Care 2013, 17(Suppl 2):P122 (doi: 10.1186/c
Introduction Protective mechanical ventilation
on reduced stretch of pulmonary tissue, sometim
hypoventilation that can be avoided when using
High-frequency positive-pressure ventilation (H
fully explored, especially when associated with o
to avoid hypercapnia. Methods We induced ARDS in eight pigs by lu
plus 3 hours of injurious MV with low PEEP and
(DP). We then performed a recruitment man
by PEEP titration using the amount of alveolar
impedance tomography (EIT). Then stabilization
tidal volume (VT) at 6 ml/kg, respiratory rate (R
and PEEP selected with the PEEP-FiO2 table (AR
kept constant during two steps of HFPPV with
an inspiratory pause (HFPPV-60), and one wit
inspiratory time (HFPPV-60 w/P30%). In another
PEEP titrated with EIT after RM (HFPPV-60 w/RM
step, VT was set to reach a PaCO2 of 60 ± 3 m
regional ventilation was analyzed using EIT. Equil
if PaCO2 was stable (<5% of variation) for >30 mi
Results HFPPV allowed reduction in PaCO2 lev
60 (58, 61), 59 (58, 60), 60 (58, 61) mmHg, be
5.2 (5.0, 5.9), 5.1 (4.5, 6.0), 4.7 (4.2, 5.7) and 4.8 (
stabilization, HFPPV-60, HFPPV-60 w/P30% a
respectively. It had no signifi cant diff erent results
with and without an inspiratory pause. P123 We compared incidence
rates of nosocomial bacteremia using the Hospitals in Europe Link for
Infection Control through Surveillance (HELICS) between general ICU
and VV-ECMO patients and used multiple logistic regression analysis to
control for possible confounders. Figure 2 (abstract P124). p
Results During the study period 1,146 patients were admitted and 16
received VV-ECMO. The incidence of bloodstream infection in patients
with ECMO was 19/1,000 exposure-days versus 4,9/1,000 exposure-
days in general ICU patients (incidence rate ratio of 3.82; 95% CI: 2.0
to 7.3; P <0.001). Bacteremia was mostly due to Gram-negative agents
(65%). The patients with bloodstream infections under ECMO (n =
10) had a nonsignifi cant younger age (P = 0.08) and a lower SAPS II
score (P = 0.03) compared with non-ECMO patients (n = 25). VV-ECMO
patients had a signifi cantly higher risk of primary bacteremia than non-
ECMO patients (P = 0.04). Patients with bloodstream infections in the
VV-ECMO group had a lower crude mortality rate (OR: 0.1, P = 0.04), not
confi rmed in the adjusted analysis. There were no crude or adjusted
diff erences in the time to bacteremia or infections due to multiple
drug-resistant microorganisms (OR: 0.26; P = 0.085) between groups. g
g
g
p
Conclusion This study suggests that VV-ECMO patients have a
signifi cantly higher risk for primary nosocomial bloodstream infection. A larger study is needed to confi rm such fi ndings and to assess the
need for specifi c intervention, namely routine antibiotic prophylaxis. Referencesfi Figure 2 (abstract P124). 1. Peek GJ, et al.: Effi cacy and economic assessment of conventional
ventilatory support versus extracorporeal membrane oxygenation for
severe adult respiratory failure (CESAR): a multicentre randomised
controlled trial. Lancet 2009, 374:1351-1363. 1. Peek GJ, et al.: Effi cacy and economic assessment of conventional
ventilatory support versus extracorporeal membrane oxygenation for
severe adult respiratory failure (CESAR): a multicentre randomised
controlled trial. Lancet 2009, 374:1351-1363. P123 P123
High-frequency percussive ventilation at altitude: study in a
hypobaric chamber with a mechanical test lung
AC Cirodde1, S Montmerle2, ND Donat1, CB Bourhillon2, P Jault1,
L Bargues1, T Leclerc1
1Military Hospital Percy, Clamart, France; 2IRBA, Brétigny, France
Critical Care 2013, 17(Suppl 2):P123 (doi: 10.1186/cc12061) Results Both ARDS and HL patients exhibited typical P–V curves and
stepwise recruitment (Figure 1). By raising PEEP from 0 to 12 cmH2O,
ARDS patients recruited 331 ± 195 ml (mean ± SD) and HL patients
435 ± 43 ml. There was a strong correlation (R2 = 0.88) of the total RV
with the end-inspiratory volume at a plateau pressure of 45 cmH2O in
both groups; that is, recruitment was found to the same extent in both
groups (Figure 2). Introduction High-frequency percussive ventilation (HFPV) is a rescue
technique for most severe acute lung injury/acute respiratory distress
syndrome (ARDS) patients [1], especially with smoke inhalation or
respiratory burns [2]. This study aimed at characterizing HFPV as
delivered by Percussionnaire VDR4® and at evaluating how hypobarism
interferes with HFPV, in order to assess its usability at altitude. Conclusion The relative contribution of RV to lung volume gain is similar
in ARDS and in patients with healthy lungs. Our results question the
relevance of recruitability as defi ned by Dellamonica and colleagues as
a typical phenomenon of ARDS, but support the baby lung concept, as
the recruited volume was closely related to the size of the lung. Methods Using a mechanical test lung mimicking ARDS (compliance
17 ml/cmH2O) with two resistance levels (5 and 15 cmH2O/l/second)
and ventilated with VDR4® in a hypobaric chamber, ascents/descents
between 0 and 5,000 and then 0 and 8,000 ft were performed. Adjustable Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 S47 http://ccforum.com/supplements/17/S2 References
1. Gattinoni et al.: N Engl J Med 2006, 354:1775. 2. Dellamonica et al.: Intensive Care Med 2011, 37:1595. 3. Stahl et al.: Crit Care Med 2006, 34:2090. P125
Nosocomial bloodstream infection and venovenous extracorporeal
membrane oxygenation: a retrospective cohort study
B Serra de Oliveira, S Mendes Fernandes, C França
Hospital de Santa Maria CHLN, Lisboa, Portugal
Critical Care 2013, 17(Suppl 2):P125 (doi: 10.1186/cc12063)
Methods A retrospective cohort study from patients admitted to
our ICU between April 2009 and June 2012. P123 We compared incidence
rates of nosocomial bacteremia using the Hospitals in Europe Link for
Infection Control through Surveillance (HELICS) between general ICU
and VV-ECMO patients and used multiple logistic regression analysis to
control for possible confounders. Results During the study period 1,146 patients were admitted and 16
received VV-ECMO. The incidence of bloodstream infection in patients
with ECMO was 19/1,000 exposure-days versus 4,9/1,000 exposure-
days in general ICU patients (incidence rate ratio of 3.82; 95% CI: 2.0
to 7.3; P <0.001). Bacteremia was mostly due to Gram-negative agents
(65%). The patients with bloodstream infections under ECMO (n =
10) had a nonsignifi cant younger age (P = 0.08) and a lower SAPS II
score (P = 0.03) compared with non-ECMO patients (n = 25). VV-ECMO
patients had a signifi cantly higher risk of primary bacteremia than non-
ECMO patients (P = 0.04). Patients with bloodstream infections in the
VV-ECMO group had a lower crude mortality rate (OR: 0.1, P = 0.04), not
confi rmed in the adjusted analysis. There were no crude or adjusted
diff erences in the time to bacteremia or infections due to multiple
drug-resistant microorganisms (OR: 0.26; P = 0.085) between groups. Conclusion This study suggests that VV-ECMO patients have a
signifi cantly higher risk for primary nosocomial bloodstream infection. A larger study is needed to confi rm such fi ndings and to assess the
need for specifi c intervention, namely routine antibiotic prophylaxis. References
1. Peek GJ, et al.: Effi cacy and economic assessment of conventional
ventilatory support versus extracorporeal membrane oxygenation for
severe adult respiratory failure (CESAR): a multicentre randomised
controlled trial. Lancet 2009, 374:1351-1363. 2. Conrick-Martin I, et al.: Nosocomial infections in a cohort of extracorporeal
life support patients. Crit Care Resusc 2012, 14:198-201. P126
Monitoring anticoagulation during extracorporeal membrane
oxygenation in patients with acute respiratory failure
M Panigada1, C Mietto2, F Pagan2, L Bogno2, V Berto2, L Gattinoni2
1F
d
i
IRCCS C ’ G
d O
d l M
i
P li li i
Mil
It l
Figure 1 (abstract P124). Figure 2 (abstract P124). Figure 1 (abstract P124). Figure 1 (abstract P124). Figure 2 (abstract P124). Methods A retrospective cohort study from patients admitted to
our ICU between April 2009 and June 2012. References 1. Gattinoni et al.: N Engl J Med 2006, 354:1775. 2. Dellamonica et al.: Intensive Care Med 2011, 37:1595. 3. Stahl et al.: Crit Care Med 2006, 34:2090. 2. Conrick-Martin I, et al.: Nosocomial infections in a cohort of extracorporeal
life support patients. Crit Care Resusc 2012, 14:198-201. 2. Conrick-Martin I, et al.: Nosocomial infections in a cohort of extracorporeal
life support patients. Crit Care Resusc 2012, 14:198-201. P125 Nosocomial bloodstream infection and venovenous extracorporeal
membrane oxygenation: a retrospective cohort study
B Serra de Oliveira, S Mendes Fernandes, C França
Hospital de Santa Maria CHLN, Lisboa, Portugal
Critical Care 2013, 17(Suppl 2):P125 (doi: 10.1186/cc12063) P127
Extracorporeal membrane oxygenation: the MOTOR of cytokine
production? p
Y Hara1, O Nishida1, T Nakamura1, S Uchiyama1, J Shibata1, C Yamashita1,
M Yumoto1, Y Shimomura1, N Kuriyama1, N Yasuoka1, M Ito1, K Kawata1,
S Hayakawa1, S Yamada2, T Miyasho3, K Moriyama1
1Fujita Health University School of Medicine, Toyoake, Japan; 2Shino
Corporation, Tokyo, Japan; 3Rakuno Gakuen University, Ebetsu, Japan
Critical Care 2013, 17(Suppl 2):P127 (doi: 10.1186/cc12065) g
Results In all patients RR decreased with the increase of extracorporeal
CO2 removal and a negative correlation was found between RR and
ECCO2 Cl/total VCO2 (r2 = 0.42, P <0.01). In all patients we were able to
obtain a reduction of RR below 15 (28 ± 4 vs. 8 ± 4, RR at low gas fl ow vs. RR at maximal gas fl ow, P <0.001). The selected maximal gas fl ow was
variable between diff erent patients (6.7 ± 2 l/minute), corresponding to
diff erent levels of ECCO2 Cl/total VCO2 (83 ± 17%, range 53 to 100%) and
RR response (8 ± 4, range 5 to 14). Introduction The usefulness of extracorporeal membrane oxygenation
(ECMO) is being rediscovered in the wake of the pandemic of H1N1
infl uenza. However, it has been reported that patients who received
ECMO often developed virus-associated hemophagocytic syndrome
(VAHS), compared with those without ECMO support. Although
there is ample evidence that extensive cytokine activation is a key
factor in VAHS, ECMO itself could be a potential trigger to exacerbate
the pathology by amplifying cytokine activation. In this study, we
investigated whether mediators such as cytokines may be produced
by ECMO. Conclusion In patients with COPD exacerbation, who failed non-
invasive ventilation, VV-ECMO allows one to maintain spontaneous
breathing. Titration of extracorporeal CO2 removal leads to control
RR. This approach could interrupt the vicious circle of dynamic
hyperinfl ation and allow the defl ation of lung parenchyma. Figure 1 (abstract P128). y
Methods Patients with severe respiratory failure who were placed
on ECMO were enrolled between June and July 2012. This study was
approved by the ethics committee. Blood specimens were drawn from
the blood circuit at the inlet of the centrifugal pump (before) and
outlet of the hollow fi ber oxygenator (after) at a frequency of three
to four times per day. Blood IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10,
IL-12(p70), IL-13, IL-17, G-CSF, GM-CSF, IFNγ, MCP-1, MIP-1β, and TNFα
were measured globally using a multiplex cytokine bead array system
(Bio-Plex; Bio-Rad, Tokyo, Japan). Eff ect of extracorporeal CO2 removal on respiratory rate in
spontaneously breathing patients with chronic obstructive
pulmonary disease exacerbation Table 1 (abstract P126). Comparison between aPTT and K-TEG R Table 1 (abstract P126). Comparison between aPTT and K-TEG R
aPTT range (ratio)
K-TEG R range
Low
Therapeutic
High
(minutes)
(<1.5)
(1.5 to 2)
(>2)
Low (<20)
13 (9%)
8 (5%)
0
21 (14%)
Therapeutic (20 to 90) 10 (7%)
25 (17%)
11 (7%)
46 (30%)
High (>90)
15 (10%)
55 (36%)
15 (10%)
85 (56%)
38 (25%)
88 (58%)
26 (17%)
152 total
samples
Conclusion Anticoagulation was excessive in more than one-half of the
samples according to TEG monitoring, while negligible based on aPTT. Reference
1. Oliver WC: Semin Cardiothorac Vasc Anesth 2009, 13:154-175. P127
Extracorporeal membrane oxygenation: the MOTOR of cytokine
production? Y Hara1, O Nishida1, T Nakamura1, S Uchiyama1, J Shibata1, C Yamashita1,
M Yumoto1, Y Shimomura1, N Kuriyama1, N Yasuoka1, M Ito1, K Kawata1,
S Hayakawa1, S Yamada2, T Miyasho3, K Moriyama1
1Fujita Health University School of Medicine, Toyoake, Japan; 2Shino
Corporation, Tokyo, Japan; 3Rakuno Gakuen University, Ebetsu, Japan
Critical Care 2013 17(Suppl 2):P127 (doi: 10 1186/cc12065) Table 1 (abstract P126). Comparison between aPTT and K-TEG R
aPTT range (ratio) Table 1 (abstract P126). Comparison between aPTT and K-TEG R
PTT
(
ti ) Table 1 (abstract P126). Comparison between aPTT and K-TEG R
aPTT range (ratio)
K-TEG R range
Low
Therapeutic
High
(minutes)
(<1.5)
(1.5 to 2)
(>2)
Low (<20)
13 (9%)
8 (5%)
0
21 (14%)
Therapeutic (20 to 90) 10 (7%)
25 (17%)
11 (7%)
46 (30%)
High (>90)
15 (10%)
55 (36%)
15 (10%)
85 (56%)
38 (25%)
88 (58%)
26 (17%)
152 total
samples Introduction During severe exacerbation of chronic obstructive
pulmonary disease (COPD) tachypnea, as a consequence of respiratory
acidosis, and airfl ow limitation, due to small airway obstruction, lead to
lung hyperinfl ation, respiratory distress and gas exchange impairment. Invasive mechanical ventilation could worsen lung hyperinfl ation
and produce a vicious circle. We investigated whether increasing
extracorporeal carbon dioxide removal (ECCO2 Cl) could reduce
the respiratory rate (RR), so prolonging time for lung emptying and
allowing resolution of hyperinfl ation. g
yl
Methods Six patients with COPD exacerbation with respiratory acidosis
(PaCO2 83 ± 27 mmHg, pH 7.19 ± 0.1) and tachypnea (RR 39 ± 5)
despite maximal non-invasive ventilation underwent venovenous
extracorporeal membrane oxygenation (VV-ECMO). All patients were
awake and spontaneously breathing an adequate air–oxygen mixture
to correct hypoxemia (PaO2 72 ± 27 mmHg). Monitoring anticoagulation during extracorporeal membrane
oxygenation in patients with acute respiratory failure yg
p
B Serra de Oliveira, S Mendes Fernandes, C França Hospital de Santa Maria CHLN, Lisboa, Portugal p
,
,
g
Critical Care 2013, 17(Suppl 2):P125 (doi: 10.1186/cc12063) M Panigada1, C Mietto2, F Pagan2, L Bogno2, V Berto2, L Gattinoni2
1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy;
2IRCCS Ca’ Granda Maggiore Policlinico Hospital Foundation, Milan, Italy
Critical Care 2013, 17(Suppl 2):P126 (doi: 10.1186/cc12064) Introduction Venovenous extracorporeal membrane oxygenation
(VV-ECMO) for respiratory failure in the ICU is used in a variety of
clinical situations and has been demonstrated to signifi cantly improve
survival without disability in adult respiratory distress syndrome [1]. ECMO has been presented as a risk factor for bloodstream infection
although recently published data do not support this view or the use
of antibiotic prophylaxis [2]. We aimed to examine VV-ECMO as a risk
factor for nosocomial bloodstream infection. Introduction aPTT is a common tool for anticoagulation monitor ing
during extracorporeal membrane oxygenation (ECMO). Thrombo-
elasto graphy (TEG) is another available option in this setting. Methods A prospective observational study on 12 consecutive
patients during venovenous ECMO. Anticoagulation was provided g p y (
)
p
g
Methods A prospective observational study on 12 consecutive
patients during venovenous ECMO. Anticoagulation was provided S48 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 0.22; IL-8, P = 0.43; IL-10, P = 0.84; MCP-1, P = 0.10; and MIP-1β, P = 0.65;
Wilcoxon signed-rank test). with unfractioned heparin titrated to an aPTT ratio target of 1.5 to 2. Kaolin-activated TEG (K-TEG) was contemporarily measured but did not
guide heparin infusion. Baseline K-TEG reaction time (R) >20 minutes
is accepted for anticoagulation but when it exceeds 90 minutes
anticoagulation may be too great [1]. with unfractioned heparin titrated to an aPTT ratio target of 1.5 to 2. Kaolin-activated TEG (K-TEG) was contemporarily measured but did not
guide heparin infusion. Baseline K-TEG reaction time (R) >20 minutes
is accepted for anticoagulation but when it exceeds 90 minutes
anticoagulation may be too great [1]. g
Conclusion The use of ECMO in patients with severe respiratory failure
did not induce systemic infl ammatory changes. These observations are
preliminary, but may nevertheless have important implications for the
future management of patients with severe infections. g
y
g
Results Mean ECMO duration was 9 ± 4 days. Eff ect of extracorporeal CO2 removal on respiratory rate in
spontaneously breathing patients with chronic obstructive
pulmonary disease exacerbation While keeping the blood
fl ow stable (2.9 ± 0.5 l/minute), we changed the gas fl ow of the artifi cial
lung to modify the extracorporeal CO2 clearance as a percentage of
total patient CO2 production (% ECCO2 Cl/total VCO2) and we observed
the variations of RR. We recorded RR at three levels of gas fl ow in each
patient (Figure 1). Conclusion Anticoagulation was excessive in more than one-half of the
samples according to TEG monitoring, while negligible based on aPTT. Reference Reference
1. Oliver WC: Semin Cardiothorac Vasc Anesth 2009, 13:154-175. Monitoring anticoagulation during extracorporeal membrane
oxygenation in patients with acute respiratory failure A total of 152 K-TEGs were
collected. Comparison between aPTT and K-TEG R is reported in Table 1. Four patients (33%) had hemorrhagic complications. Neither aPTT
nor K-TEG R were signifi cantly diff erent in patients with hemorrhagic
events compared with patients without hemorrhagic events but the
latter received a signifi cantly lower total heparin dose (P = 0.0097). P128f P128f Eff ect of extracorporeal CO2 removal on respiratory rate in
spontaneously breathing patients with chronic obstructive
pulmonary disease exacerbation
E Spinelli, S Crotti, L Zacchetti, N Bottino, V Berto, R Russo, M Chierichetti,
A Protti, L Gattinoni
Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P128 (doi: 10.1186/cc12066) P131 Feasibility of minimally invasive stimulation of the phrenic nerves
for supporting ventilation in fully anesthetized swine
J Bijwadia1, M Karamanoglu2
1Regions Hospital, St Paul, MN, USA; 2Respithera, Bloomington, MN, USA
Critical Care 2013, 17(Suppl 2):P131 (doi: 10.1186/cc12069) ECCO2 removal with a phosphorylcholine-coated membrane
oxygenator in diffi cult respiratory weaning patients Table 1 (abstract P129) p
g
Results There was a close correlation between the measured build-up
of end-expiratory lung volume during a PEEP-step manoeuvre and
ΔPEEP/EL where EL was conventionally determined using oesophageal
pressure measurements (see Figure 1).fi Conclusion Esophageal pressure measurements are diffi cult to perform
[3] and rarely used in routine clinical practice. Our fi ndings indicate that
a change in PEEP together with measurements of the resulting change
in end-expiratory volume by spirometry in the ventilator could be used
to determine lung elastance separately, the relation between lung and
chest wall elastance as well as the transpulmonary pressure. References Conclusion ECCO2 removal with a phosphorylcholine-coated mem-
brane oxygenator is clinically safe, avoids clotting of the oxygenator
circuit and allows adequate CO2 removal. Reference 1. Stenqvist O, et al.: Acta Anaesthesiol Scand 2012, 56:738-747. 2. Baydur A: Chest 2002, 121:324-326. 3. Hedenstierna G: Minerva Anestesiol 2012, 78:959-966. 1. Stenqvist O, et al.: Acta Anaesthesiol Scand 2012, 56:738-747. 2. Baydur A: Chest 2002, 121:324-326. 2. Baydur A: Chest 2002, 121:324 326. 3. Hedenstierna G: Minerva Anestesiol 2012, 78:959-966. 1. Livigni S, et al.: Crit Care 2006, 10:R151. P127
Extracorporeal membrane oxygenation: the MOTOR of cytokine
production? One-way
ANOVA was used to compare the changes of parameters. P <0.05 was
considered signifi cant. Results Table 1 presents the main results. The CO2 removal by mem-
brane oxygenator ranged from 56 to 37 ml/minute. All patients
survived to the treatment and 7/10 were weaned from the ventilator
at the end of ECCO2 removal. Only one oxygenator was used for every
patient without clotting of the circuit or any major bleeding problem. Table 1 (abstract P129)
Day 0
Day 2
Day 4
pH
7.24 ± 0.06
7.38 ± 0.1
7.41 ± 0.07*
PaCO2 (mmHg)
70 ± 5
57 ± 8
52 ± 3*
PP (cmH2O)
48 ± 4
28 ± 4*
25 ± 4
VCO2 (ml/minute)
220 ± 15
201 ± 14
180 ± 13*
*P <0.05 versus T0. Conclusion ECCO2 removal with a phosphorylcholine-coated mem-
brane oxygenator is clinically safe, avoids clotting of the oxygenator
circuit and allows adequate CO2 removal. Reference
1. Livigni S, et al.: Crit Care 2006, 10:R151. P130
Lung elastance can be determined without esophageal pressure
measurements
C Grivans, S Lundin, O Stenqvist
Sahlgrenska University Hospital, Gothenburg, Sweden
Critical Care 2013, 17(Suppl 2):P130 (doi: 10.1186/cc12068)
Introduction We have previously shown, in an ex vivo porcine model,
that lung elastance calculated as the PEEP change divided by lung
volume increase (ΔPEEP/ΔEELV) is closely correlated to conventionally
measured lung elastance using oesophageal pressure [1]. In this study
we hypothesize that the successive change in lung volume during
a PEEP-step manoeuvre could be predicted from ΔPEEP and lung
l
h
b
f h
d
l d
h P129
ECCO2 removal with a phosphorylcholine-coated membrane
oxygenator in diffi cult respiratory weaning patients
F Turani, S Martini, A Marinelli, M Falco, R Barchetta, F Candidi, F Gargano
1Aurelia Hospital/European Hospital, Rome, Italy
Critical Care 2013, 17(Suppl 2):P129 (doi: 10.1186/cc12067) Figure 1 (abstract P130). Correlation between spirometrically measured
and calculated increase in EELV. ECCO2 removal with a phosphorylcholine-coated membrane
oxygenator in diffi cult respiratory weaning patients Introduction ECCO2 removal may be a useful support in patients
with diffi cult respiratory weaning. The aim of this study is to evaluate
a new phosphorylcholine-coated EECO2 removal system with no
thrombogenic activity to assess the clinical safety of the system, the
changes of main cardiorespiratory indices and CO2 removal by the
system. y
Methods Ten patients were enrolled. Before starting with ECCO2
removal all patients were ventilated with TV <6 ml/kg, peak pressure
>35 cmH2O and pH <7.25. ECCO2 removal was initiated using a modifi ed
continuous venovenous hemofi ltration system with a membrane
oxygenator (ABYLCAP; Bellco, Mirandola, Italy; membrane surface
area: 0.67 m2, blood fl ow 280 to 350 ml/minute, phosphorylcholine
coated). Femoral vein cannulation with a double-lumen catheter was
used to connect the patients to the extracorporeal system. Heparin was
infused to maintain ACT <190 to 240 seconds. All patients had ECCO2
for 4 days. Every 12 hours the pH, PaCO2, peak pressure and PaO2 were
evaluated. VCO2 was determined by indirect calorimetry using a gas
analyser equipped on an Engstrom Carestation Ventilator. During the
ECCO2 removal the patients were ventilated with TV ≤6 ml/kg and peak
pressure <30 cmH2O. All data are expressed as mean ± SD. One-way
ANOVA was used to compare the changes of parameters. P <0.05 was
considered signifi cant. Figure 1 (abstract P130). Correlation between spirometrically measured
and calculated increase in EELV. i
Results Table 1 presents the main results. The CO2 removal by mem-
brane oxygenator ranged from 56 to 37 ml/minute. All patients
survived to the treatment and 7/10 were weaned from the ventilator
at the end of ECCO2 removal. Only one oxygenator was used for every
patient without clotting of the circuit or any major bleeding problem. pressure minus tidal variation in oesophageal pressure divided by tidal
volume. Position of the oesophageal catheter was verifi ed according
to Baydur [2]. The measured change in end-expiratory lung volume
during the PEEP-step manoeuvre using spirometry was compared
with the end-expiratory lung volume change calculated from EL and
stepwise changes in PEEP as ΔPEEP/EL. Table 1 (abstract P129)
Day 0
Day 2
Day 4
pH
7.24 ± 0.06
7.38 ± 0.1
7.41 ± 0.07*
PaCO2 (mmHg)
70 ± 5
57 ± 8
52 ± 3*
PP (cmH2O)
48 ± 4
28 ± 4*
25 ± 4
VCO2 (ml/minute)
220 ± 15
201 ± 14
180 ± 13*
*P <0.05 versus T0. P127
Extracorporeal membrane oxygenation: the MOTOR of cytokine
production? HMGB1 was measured using an ELISA
kit (Shino-Test, Tokyo, Japan). Results Two patients with interstitial pneumonia were studied. The
ECMO system consisted of a Rotafl ow Centrifugal Pump (Maquet Japan,
Tokyo, Japan), a Biocube TNC coating 6000 (NIPRO, Osaka, Japan), and
a percutaneous cardiopulmonary support system (Capiox EBS; Terumo,
Tokyo, Japan). The blood fl ow rate was 2.0 ± 4.0 l/minute. A total of 34
blood sets were collected. In most cases, blood levels of IL-1β, IL-2, IL-
4, IL-5, IL-12(p70), IL-13, IL-17, GM-CSF, IFNγ, and TNFα were below the
detection limit and did not increase during ECMO. The other mediators
were detected at the inlet (before), but no signifi cant increase was
observed at the outlet (after) (HMGB1, P = 0.33; IL-6, P = 0.12; IL-7, P = Figure 1 (abstract P128). S49 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P129
ECCO2 removal with a phosphorylcholine-coated membrane
oxygenator in diffi cult respiratory weaning patients
F Turani, S Martini, A Marinelli, M Falco, R Barchetta, F Candidi, F Gargano
1Aurelia Hospital/European Hospital, Rome, Italy
Critical Care 2013, 17(Suppl 2):P129 (doi: 10.1186/cc12067)
Introduction ECCO2 removal may be a useful support in patients
with diffi cult respiratory weaning. The aim of this study is to evaluate
a new phosphorylcholine-coated EECO2 removal system with no
thrombogenic activity to assess the clinical safety of the system, the
changes of main cardiorespiratory indices and CO2 removal by the
system. Methods Ten patients were enrolled. Before starting with ECCO2
removal all patients were ventilated with TV <6 ml/kg, peak pressure
>35 cmH2O and pH <7.25. ECCO2 removal was initiated using a modifi ed
continuous venovenous hemofi ltration system with a membrane
oxygenator (ABYLCAP; Bellco, Mirandola, Italy; membrane surface
area: 0.67 m2, blood fl ow 280 to 350 ml/minute, phosphorylcholine
coated). Femoral vein cannulation with a double-lumen catheter was
used to connect the patients to the extracorporeal system. Heparin was
infused to maintain ACT <190 to 240 seconds. All patients had ECCO2
for 4 days. Every 12 hours the pH, PaCO2, peak pressure and PaO2 were
evaluated. VCO2 was determined by indirect calorimetry using a gas
analyser equipped on an Engstrom Carestation Ventilator. During the
ECCO2 removal the patients were ventilated with TV ≤6 ml/kg and peak
pressure <30 cmH2O. All data are expressed as mean ± SD. P134 Novel biomarkers for prediction of mortality after acute
exacerbation of chronic obstructive pulmonary disease
H Michalopoulou, H Michalopoulou, P Stamatis, F Kattis, D Stamatis
Metaxa Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P134 (doi: 10.1186/cc12072) Results Ninety patients were included (77% male, 23% female). Mean
age 69 years (± 11.88). Fifty-fi ve percent were smokers, 52% had
arterial hypertension, 39% received long-term oxygenotherapy, 18%
received antibiotics in the 7 days before, 18% corticosteroids, and
14% were on long-term NIV support at home. An emergency medical
ambulance was immediately sent for 86% of patients. Ninety-two
percent had normal consciousness (Glasgow Coma Scale 15), 78%
had bronchospasm, 71% had signs of respiratory struggle, and 12%
were unable to speak. The mean respiratory rate was 31.4 cycles/
minute (± 8.18), the average cardiac pulse was 103.6 beats/minute
(± 23.14). Nasal EtCO2 44.92 mmHg (± 16.38), pulse oximetry with air
was 83.48% (± 12.09), and the average fl ow rate of oxygen delivered
was 5.69 l/minute (± 2.93). None of the patients had fever. Eighty-fi ve
percent were supported on spontaneous ventilation, 22% received
prehospital non-invasive ventilation, they all showed signs of severity
and 3% need tracheal intubation. Seventy-fi ve percent of patients
received β2-agonist and anticholinergic nebulization, 45% intravenous
corticosteroids. Seventy-one percent were admitted to the emergency
room, 29% to the ICU. Introduction Retrospective studies suggest that cardiac troponin
levels are often elevated in patients with acute exacerbation of
chronic obstructive pulmonary disease (AECOPD) indicating a poor
survival. Novel high-sensitivity cardiac troponin (hs-cTnT) assays have
better analytical precision than standard troponin (cTnT) assays. We
elaborated a prospective cohort study to investigate the prognostic
value of this novel biomarker in patients with AECOPD. Methods Fifty-six patients (mean age 64 years, 68% male) with the
fi nal diagnosis of AECOPD were enrolled. Those who were diagnosed
with acute coronary syndromes were excluded. We measured cardiac
troponin T with a standard fourth-generation assay and a high-
sensitivity assay. Clinical, electrocardiographic and echocardiographic
data were collected at admission and the primary prognostic endpoint
was death during 30 days of follow-up. Results Mean hs-cTnT levels at admission were 34 ng/l. During the
follow-up period seven patients (12%) died. Thirty-eight percent of
patients had hs-cTnT above the range of 14 ng/l. Lung elastance can be determined without esophageal pressure
measurements C Grivans, S Lundin, O Stenqvist Sahlgrenska University Hospital, Gothenburg, Sweden Sahlgrenska University Hospital, Gothenburg, Sweden g
y
g
Critical Care 2013, 17(Suppl 2):P130 (doi: 10.1186/cc12068) Critical Care 2013, 17(Suppl 2):P130 (doi: 10.1186/cc12068) Introduction Long-term use of mechanical ventilators may lead to
ventilator-induced diaphragmatic dysfunction (VIDD) and increase the
duration of weaning from MV [1]. It was hypothesized that stimulating
the diaphragm during MV may prevent VIDD and may lead to early
weaning [2]. In this study, the feasibility of generating coordinated
contraction of both diaphragms was investigated using a novel
transvenous diaphragmatic pacing system. Introduction We have previously shown, in an ex vivo porcine model,
that lung elastance calculated as the PEEP change divided by lung
volume increase (ΔPEEP/ΔEELV) is closely correlated to conventionally
measured lung elastance using oesophageal pressure [1]. In this study
we hypothesize that the successive change in lung volume during
a PEEP-step manoeuvre could be predicted from ΔPEEP and lung
elastance as ΔPEEP/EL. The objective of the study was to validate this
hypothesis in patients with acute respiratory failure (ARF). Methods Two juvenile pigs were anesthetized with propofol (150 to
250 μg/kg/minute) and ventilated (VENT) with an assist control mode
MV (Nellcor Puritan Bennett 840). Using fl uoroscopy, a novel multipolar
neurostimulation catheter (Inspirx RL PICC53; Respithera, Bloomington,
MN, USA) was threaded into the left internal jugular vein and advanced
to the junction of right atrium and the superior vena cava using a Methods Thirteen patients with ARF were studied during an incremental
PEEP trial, 0–4–8–12–16 cmH2O. ΔEELV was determined as the change
in expiratory tidal volume following each PEEP step. Conventional
calculation of lung elastance was obtained from tidal variation in airway S50 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 modifi ed Seldinger technique. The successful capture of the right
and left phrenic nerves was confi rmed by fl uoroscopic visualization. Peak airway pressures (PAWP) and blood gases were determined after
10 minutes MV (MV), MV and stimulation applied together (MV+STIM)
and stimulation only (STIM). modifi ed Seldinger technique. The successful capture of the right
and left phrenic nerves was confi rmed by fl uoroscopic visualization. Peak airway pressures (PAWP) and blood gases were determined after
10 minutes MV (MV), MV and stimulation applied together (MV+STIM)
and stimulation only (STIM). P133
N-terminal pro-brain natriuretic peptide as a prognostic marker
in patients with exacerbation of chronic obstructive pulmonary
disease Table 1 (abstract P131). Ventilation parameters
MV
MV+STIM
STIM
PAWP (cmH2O)
20
16
9
PO2 (mmHg)
89
101
95
PCO2 (mmHg)
32
26
26 Table 1 (abstract P131). Ventilation parameters Table 1 (abstract P131). Ventilation parameters P Stamatis, H Michalopoulou, D Stamatis
Metaxa Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P133 (doi: 10.1186/cc12071) P Stamatis, H Michalopoulou, D Stamatis
Metaxa Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P133 (doi: 10.1186/cc12071) Introduction Plasma N-terminal pro-brain natriuretic peptide (NT-pro-
BNP) levels are elevated in patients with pulmonary disease especially
in those with concomitant right ventricular dysfunction. The aim of the
present study was to investigate the use of plasma NT-pro-BNP levels as
a prognostic tool in patients with exacerbation of chronic obstructive
pulmonary disease (COPD). Conclusion It was possible to capture and stimulate both phrenic
nerves using a minimally invasive approach to support respiration
and sustain blood gases at physiological levels. This development
could help wean MV-dependent ICU patients earlier. Further long-term
studies are needed to assess the full potential of this novel system. References Methods A retrospective medical records analysis of all patients
hospitalized between June 2009 and June 2012 with the fi nal diagnosis
of acute exacerbation of COPD who had undergone NT-pro-BNP
measurements at admission followed by echocardiogram. 1. Vassilakopoulos T, Petrof BJ: Am J Respir Crit Care Med 2004, 169:336-341. 2. Sassoon CS, Zhu E, Caiozzo VJ: Am J Respir Crit Care Med 2004, 170:626-632. 1. Vassilakopoulos T, Petrof BJ: Am J Respir Crit Care Med 2004, 169:336-341. 2. Sassoon CS, Zhu E, Caiozzo VJ: Am J Respir Crit Care Med 2004, 170:626-632. 1. Vassilakopoulos T, Petrof BJ: Am J Respir Crit Care Med 2004, 169:336-341. y
Results Seventy-two patients (mean age 69 years, 58% male) with
COPD exacerbation but without clinical or echocardiography-oriented
evidence of acute cardiac disease were enrolled. The mean forced
expiratory volume in 1 second (FEV 1)/forced vital capacity ratio was
44.2% and the mean FEV 1 was 0.82 l. Median NT-pro-BNP levels
at admission were 424.4 pg/ml and the tertile limits were 262.1 and
930 pg/ml. NT-pro-BNP levels signifi cantly predicted 30-day mortality
(OR: 7.2, 95% CI: 3.9 to 15.2; P <0.001), (OR: 25, 95% CI: 16.3 to 37.3;
P <0.001) and (OR: 44, 95% CI: 44.5 to 76.2) for each tertile. These
associations persisted after adjusting for arterial CO2 pressure, body
mass index, age, gender and systolic pulmonary artery pressure. References References
1. Antoine G: The epidemiology and outcome of medical emergency team
call patients treated with non invasive ventilation. Resuscitation 2011,
82:1218-1223. 2. Schmidbauer W: Early prehospital use of non invasive ventilation improves
acute respiratory failure in acute exacerbation of chronic obstructive
pulmonary disease. Emerg Med J 2011, 28:626-627. 1. Antoine G: The epidemiology and outcome of medical emergency team
call patients treated with non invasive ventilation. Resuscitation 2011,
82:1218-1223. 2. Schmidbauer W: Early prehospital use of non invasive ventilation improves
acute respiratory failure in acute exacerbation of chronic obstructive
pulmonary disease. Emerg Med J 2011, 28:626-627. y
Results No animal–ventilator dyssynchrony during stimulation
(MV+STIM) was noted while peak airway pressures were reduced. During STIM there was no discernible paradoxical movement of
the diaphragm. In addition, PCO2 and PO2 confi rmed that adequate
ventilation and oxygenation can be provided by the system, while
PAWP could be reduced (Table 1). Prehospital management of COPD patients in respiratory failure
and short-term outcome G Campagne, J Cuny, P Gosselin, P Goldstein, N Assez, E Wiel
Lille University Hospital Center, Lille Cedex, France y
p
Critical Care 2013, 17(Suppl 2):P132 (doi: 10.1186/cc12070) Introduction Respiratory failure in COPD patients is a frequent call
in French emergency dispatching centers. We have evaluated the
prehospital management of COPD patients and severity signs, and
analyse outcome in the emergency department or ICU. Conclusion Among patients with acute exacerbation of chronic
pulmonary disease, NT-pro-BNP could be a useful marker for severity
and poor prognosis. Methods We conducted an observational, descriptive, retrospective,
single-center study during a 4-month period. All COPD patients with
respiratory failure and prehospital care were included. Diff erent data
were recorded. P134 Prognostic accuracy
of hs-cTnT for death was signifi cantly higher, with area under the
ROC curve (AUC) of 0.83 (95% CI: 0.72 to 0.86), than that of cTnT (AUC:
0.63, 95% CI: 0.56 to 0.74; P <0.001). After adjustment for age, gender,
creatinine levels, heart rate, left ventricular ejection fraction, arterial
O2 pressure and systolic pulmonary artery pressure, hs-cTnT above the Conclusion Most of the patients had signs of severity and
bronchospasm. The absence of fever and antibiotics allows us to think
that the cause of decompensation is not pneumonia. Although most of
them were hypoxic and hypercapnic, they seem to be good candidates
for NIV support in prehospital care. Very few studies report the use of
NIV in cases of COPD respiratory failure in the fi rst care delivered at
home. Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 equal groups: group I (cases, 25 patients) were subjected to additional
intervention, early FOB during the fi rst 6 to 12 hours from admission;
while group II (control, 25 patients) received the conventional treatment
and NIPPV only. Outcome parameters measured were changes in ABG
data, duration of NIPPV, rate of its success, ICU stay and mortality as
well as the safety of FOB and possible complications.if 99th percentile was associated with a hazard ratio for death of 3.2 (95%
CI: 1.5 to 6.7). Conclusion In AECOPD, novel biomarkers such as hs-cTnT appear to be
positively associated with COPD severity and could be a determinant
of mortality. 99th percentile was associated with a hazard ratio for death of 3.2 (95%
CI: 1.5 to 6.7). Conclusion In AECOPD, novel biomarkers such as hs-cTnT appear to be
positively associated with COPD severity and could be a determinant
of mortality. y
Results No signifi cant diff erence was detected between the two
groups regarding the baseline characteristics. No serious complications
happened from FOB, oxygen desaturation occurred in 4/25 patients
(16%), tachycardia in 2/25 patients (8%). In group I, 23 patients (92%)
were successfully weaned from NIPPV versus 16 patients (64%) in
group II (P = 0.037). Total duration of NIPPV was 28.52 hours in group I
versus 56.25 hours in group II (P = 0.001). Length of ICU stay was 4.84
days in group I versus 8.68 days in group II (P = 0.001). SMART-COP score for patients admitted with community-acquired
pneumonia (CAP) to an ICU in a district general hospital: a smarter
way of identifying patients with severe CAP? SMART-COP score for patients admitted with community-acquired
pneumonia (CAP) to an ICU in a district general hospital: a smarter
way of identifying patients with severe CAP? y
y
g p
M Pachucki1, F Kelly2, A Padkin2 M Pachucki1, F Kelly2, A Padkin2 y
1Frenchay Hospital, Bristol, UK; 2RUH, Bath, UK Results Thirty-four patients were enrolled. Median age was 66 (54 to
71) years; 65% of the patients were male; median APACHE IV score on
admission was 76 (61 to 96). Median ICU stay was 8 (5 to 17) days, with
an overall hospital mortality of 12%. Thirteen patients (38%) tested
positive for a respiratory virus. The most frequently found virus was
infl uenza (37%), followed by RSV (15%), rhinovirus (15%), MPV (15%),
corona virus (9%) and parainfl uenza virus (9%). Two of the patients
(6%) had a bacterial–viral co-infection (blood culture and PCR positive
for Streptococcus pneumonia and rhinovirus in one patient and for
Haemophilus infl uenza and infl uenza virus in the second patient). Length of stay in the ICU was signifi cantly longer in PCR-negative
patients, in comparison with PCR-positive patients (11 (6 to 20) vs. 7 (2
to 8) days, respectively, P <0.05).l y
p
Critical Care 2013, 17(Suppl 2):P137 (doi: 10.1186/cc12075) Introduction British Thoracic Society guidelines on community-
acquired pneumonia (CAP) advocate ICU referral for patients with
CURB65 score of 4 and 5. A recently developed scoring system, SMART-
COP, designed to identify patients at need of intensive respiratory or
vasopressor support (IRVS), has been validated in a variety of settings. It predicts the need for ICU admission (defi ned as need for IRVS) with
greater accuracy than CURB65, but is not used routinely in our UK
institution. Methods We retrospectively analysed critical care admissions of
patients with a diagnosis of CAP in a UK district general hospital –
ICNARC-coded diagnoses of pneumonia (bacterial, viral, no organisms
isolated) over a 7-month period (August 2011 to January 2012). We
ascertained the CURB65 and SMART-COP scores on referral to the ICU
and matched them in relation to the need for IRVS, length of inotropic
and ventilatory support and ICU length of stay. Conclusion Respiratory viruses, and particularly infl uenza virus, are
frequently found in adult patients with respiratory failure admitted to
the ICU. P136 Role of bronchoscopy during non-invasive ventilation in
hypercapnic respiratory failure
W Rady, A Abouelela, A Aly, W Youssef
Alexandria University, Alexandria, Egypt
Critical Care 2013, 17(Suppl 2):P136 (doi: 10.1186/cc12074) References 1. Carrat F, et al.: Intensive Care Med 2006, 2:156-159. 2. Choi SH, et al.: Am J Respir Crit Care Med 2012, 186:325-332. 3. Miggins M, et al.: PLoS One 2011, 6:18890. 1. Carrat F, et al.: Intensive Care Med 2006, 2:156-159. 2. Choi SH, et al.: Am J Respir Crit Care Med 2012, 186:325-332. 3. Miggins M, et al.: PLoS One 2011, 6:18890. Results Our search revealed 28 potential matches. Five patients were
excluded (not CAP) and the notes for seven patients were not available
for analysis. We analysed the notes of 16 patients matching our criteria. In this small sample, there was a strong association between increasing
SMART-COP score and the need for IRVS (correlation coeffi cient r =
0.96). There was also a strong correlation with longer inotropic support
(r = 0.85) and longer ventilatory support (r = 0.96) with increasing
SMART-COP scores but a weaker correlation with length of ICU stay (r =
0.49). Moreover, none of the patients admitted to the ICU had CURB65
score higher than 3 at the time of ICU referral. P134 Only one patient
died in group I versus three patients in group II (P = 0.609). High incidence of respiratory viruses in critically ill adult patients
with respiratory failure Introduction Laboratory testing for viral infections is not routinely
performed in adult patients admitted to the ICU. However, reports
indicate that viruses may contribute to signifi cant morbidity and
mortality and that viral–bacterial co-infection is associated with poor
outcome in this particular patient population. The aim of the study was
to document the incidence of respiratory virus infections in critically ill
adult patients admitted to the ICU for acute respiratory failure. g
p
p
g
p
Conclusion The early application of FOB during NIPPV in patients
with ARF due to COPD exacerbation was shown to be safe. Signifi cant
improvement in the outcome of patients who underwent FOB was
noticed in terms of improved ABG data, shorter duration of NIPPV,
higher percentage of success and shorter ICU stay while no signifi cant
diff erence was detected in mortality. References 1. Scala R, et al.: Crit Care 2010, 14:R80. y
Methods The prospective, observational study took place during
two consecutive winter seasons in a mixed 16-bed ICU at the Medical
Center Leeuwarden, the Netherlands. Adult patients admitted to
the ICU, suspected of respiratory failure due to community-acquired
pneumonia, were included. After informed consent brushed
nasopharyngeal swab (Copan®) samples were taken within 24 hours
after admission and analyzed by PCR assays for the presence of
respiratory syncytial virus (RSV), infl uenza virus, metapneumovirus
(MPV), parainfl uenza virus, adenovirus, rhinovirus and coronavirus. Data are presented as median (IQR). 1. Scala R, et al.: Crit Care 2010, 14:R80. 2. Ambrosino N, et al.: Eur Respir J 2008, 31:874-886. 2. Ambrosino N, et al.: Eur Respir J 2008, 31:874-886. 2. Ambrosino N, et al.: Eur Respir J 2008, 31:874-886. 3. Heunks LM, et al.: Intensive Care Med 2010, 36:143-147. p
3. Heunks LM, et al.: Intensive Care Med 2010, 36:143-147. 3. Heunks LM, et al.: Intensive Care Med 2010, 36:143-147. Role of bronchoscopy during non-invasive ventilation in
hypercapnic respiratory failure W Rady, A Abouelela, A Aly, W Youssef W Rady, A Abouelela, A Aly, W Youssef Alexandria University, Alexandria, Egypt
Critical Care 2013, 17(Suppl 2):P136 (doi: 10.1186/cc12074) Conclusion In our small sample, higher SMART-COP score was
associated with increased likelihood of IRVS. This suggests that a further
study with a larger sample size should be performed to investigate
whether SMART-COP is an improvement on CURB65 in predicting the
need for IRVS in UK intensive care patients. Introduction Non-invasive positive-pressure ventilation (NIPPV) is
the fi rst-line treatment for hypercapnic acute respiratory failure (ARF)
secondary to COPD exacerbation in selected patients. Limited data
exist supporting the use of fi beroptic bronchoscopy (FOB) during this
clinical setting. The aim of this study is to assess the role of FOB during
NIPPV in patients with decompensated COPD acute exacerbation. P135 High incidence of respiratory viruses in critically ill adult patients
with respiratory failure
M Sietses1, TE Faber1, L Bont2, H Buter1, EC Boerma1
1Medical Center Leeuwarden, the Netherlands; 2University Medical Center
Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P135 (doi: 10.1186/cc12073) Ventilation distribution measured with electrical impedance
tomography at varying levels of assist during pressure support
versus neurally adjusted ventilatory assist in ICU patients
MS Van Mourik1, P Blankman1, D Hasan2, D Gommers1
1Erasmus Medical Center, Rotterdam, the Netherlands; 2Maasstad, Rotterdam,
the Netherlands
Critical Care 2013, 17(Suppl 2):P140 (doi: 10.1186/cc12078) 1Erasmus Medical Center, Rotterdam, the Netherlands; 2Maasstad, Rotterdam,
the Netherlands y
y
Results The APACHE scores in the S group and NS group were 27 ± 10
and 23 ± 4, Sequential Organ Failure Assessment scores were 8 ± 4
and 7 ± 3, respectively. These scores showed no signifi cant diff erence
between the groups. Procalcitonin (PCT) in the S and NS groups was
20.7 ± 21.7 and 45.0 ± 47.7 ng/ml, respectively, and there was no
signifi cant diff erence between the groups. PCT declined signifi cantly
in both groups. PaO2/FiO2 of the NS group was signifi cantly higher
than the S group on ICU admission and 4 days after admission, but
no signifi cant diff erence on 7 days after ICU admission. IL-6 of the NS
group declined signifi cantly after ICU admission, and the S group also
tended to decline. Introduction Electrical impedance tomography (EIT) is a non-invasive
and nonradiating imaging technique, which can be used to visualize
ventilation distribution of the lungs and could distinguish between the
dependent (dorsal) and nondependent (ventral) parts. Introduction Electrical impedance tomography (EIT) is a non-invasive
and nonradiating imaging technique, which can be used to visualize
ventilation distribution of the lungs and could distinguish between the
dependent (dorsal) and nondependent (ventral) parts. Methods The aim of this study was to observe ventilation distribution
between dependent and nondependent lung regions, for the individual
patient, during three diff erent levels of support during pressure
support (PS) and neurally adjusted ventilatory assist (NAVA) ventilation. Ten mechanically ventilated patients in the ICU were included. y
p
Results The ratio for dependent/nondependent distribution of
ventilation is signifi cantly higher at lower support levels compared
with higher support levels in both PS and NAVA. However, during NAVA
there was signifi cantly less impedance loss between the diff erent levels
of assist compared with PS. Tidal volumes decreased when decreasing
assist levels during PS whereas not during NAVA ventilation. The
electrical activity of the diaphragm decreased in both PS and NAVA with
higher levels of assist. Three patients showed an increase in dependent
tidal impedance variation (TIV) after lowering the assist level from 15 to
10 cmH2O. This increase in TIV did not occur during NAVA ventilation. 3.
Robins-Browne K, et al.: The SMART-COP score performs well for
pneumonia risk stratifi cation in Australia’s Tropical Northern Territory: a
prospective cohort study. Trop Med Int Health 2012, 17:914-919.
4.
Fang et al.: Application and comparison of scoring indices to predict
outcomes in patients with healthcare associated pneumonia. Crit Care
2011, 15:R32. P138f P138
Eff ects of steroid therapies to severe streptococcal pneumoniae
that required mechanical ventilation
T Nagura, T Ikeda, K Ikeda, T Ueno, S Suda
Tokyo Medical University, Hachioji Medical Center, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P138 (doi: 10.1186/cc12076) Introduction Streptococcal pneumonia remains the most common
cause of community-acquired pneumonia (CAP), bacterial meningitis
and bacteremia. Severe pneumonia caused by streptococcal pneu-
monia frequently exists in the emergency room or ICU. We performed
this study to evaluate the eff ect of steroid therapy for severe strepto-
coccal pneumonia patients with mechanical ventilation retrospectively. Methods We enrolled 13 adults of streptococcal pneumonia patients
who required mechanical ventilation. Seven of 13 patients (S group)
were administered with steroid (hydrocortisone 200 to 300 mg/day),
and the remaining six patients received no steroid therapy (NS group). As the conventional therapies, mechanical ventilation was commenced
when a patient’s PaO2/FiO2 showed less than 200 or they clinically
complained of being short of breath. All patients received appropriate
fl uid therapies, vasoactive agents and blood transfusion according
to the protocol of early goal-directed therapy in the Surviving Sepsis
Campaign Guidelines 2008, and also were treated with antibiotics,
immunoglobulins (5 g/day for 3 days) and sivelestat sodium hydrate
(4.8 mg/kg/day for 7 days). Conclusion The presence of diabetes mellitus and immunosuppression,
higher APACHE II scores, longer mechanical ventilation duration, and
presence of open wounds were predictors of VAP. Reference 1. Guidelines for the management of adults with hospital-acquired,
ventilator-associated, and healthcare-associated pneumonia. Am J Respir
Crit Care Med 2005, 171:388-416. Ventilation distribution measured with electrical impedance
tomography at varying levels of assist during pressure support
versus neurally adjusted ventilatory assist in ICU patients
MS Van Mourik1, P Blankman1, D Hasan2, D Gommers1
1Erasmus Medical Center, Rotterdam, the Netherlands; 2Maasstad, Rotterdam,
the Netherlands
Critical Care 2013, 17(Suppl 2):P140 (doi: 10.1186/cc12078) Conclusion Steroid therapy for severe streptococcal pneumonia
patients with mechanical ventilation may have a potential to maintain
oxygenation of the lung, but no signifi cant eff ects on changes of
infl ammatory markers (IL-6, CRP). P139 Risk factors for ventilator-associated pneumonia in a surgical ICU
A Kundakci, O Ozkalaycı, P Zeyneloglu, H Arslan, A Pirat
Baskent University Hospital, Ankara, Turkey
Critical Care 2013, 17(Suppl 2):P139 (doi: 10.1186/cc12077) 2
Conclusion There is more ventilation in the dependent part of the
lung, compared with the nondependent part, at lower levels of assist. This could indicate that at higher support levels the contribution
of the diaphragm is reduced. During NAVA ventilation, there is an
autoregulation in which the patient is adjusting his tidal ventilation to
maintain homogeneous ventilation distribution. Introduction Ventilator-associated pneumonia (VAP) is the most
common nosocomial infection in ICU patients who require mechanical
ventilation support. The aim of this study was to determine predictors
for the development of VAP in surgical ICU patients admitted to Baskent
University Hospital. y
Methods Following Institutional Review Board approval we performed
this retrospective study, including 876 patients admitted to the surgical
ICU between January 2009 and July 2012. After completing a review of
patient data, 45 patients diagnosed with VAP were compared with 26
appropriate matches who were not. Patients’ demographical features
(age, sex, body weight), underlying diseases, etiology for ICU admission,
APACHE II and Sequential Organ Failure Assessment scores, duration of
hospitalization, organ dysfunctions, fl uid balances, laboratory values,
use of vasopressors, mechanical ventilation, nutrition, antibiotics,
transfusions, features related to central venous catheterization, urinary
catheterization, and intubation were the recorded parameters. Patients P140 Ventilation distribution measured with electrical impedance
tomography at varying levels of assist during pressure support
versus neurally adjusted ventilatory assist in ICU patients
MS Van Mourik1, P Blankman1, D Hasan2, D Gommers1
1Erasmus Medical Center, Rotterdam, the Netherlands; 2Maasstad, Rotterdam,
the Netherlands
Critical Care 2013, 17(Suppl 2):P140 (doi: 10.1186/cc12078) References 1. Lim WS, et al.: BTS guidelines for the management of community acquired
pneumonia in adults: update 2009. Thorax 2009, 64(Suppl 3):iii1-iii55. doi:10.1136/thx.2009.121434 Methods This study is a randomized prospective case–control pilot
study carried out on 50 patients admitted to critical care units at
Alexandria University Hospital, Egypt suff ering from hypercapnic ARF
secondary to COPD exacerbation with Kelly Matthay Score from 2 to 4. All patients received NIPPV. Patients were divided randomly into two doi:10.1136/thx.2009.121434 2. Charles PG, et al.: SMART-COP: a tool for predicting the need for intensive
respiratory or vasopressor support in community-acquired pneumonia. J
Clin Infect Dis 2008, 47:375-384. 2. Charles PG, et al.: SMART-COP: a tool for predicting the need for intensive
respiratory or vasopressor support in community-acquired pneumonia. J
Clin Infect Dis 2008, 47:375-384. Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 S52 who were not intubated and were discharged or died within 2 days of
ICU admission were excluded. Results Out of 71 patients who were included in the fi nal analysis, 45
patients (63%) had VAP. Comparing with the control group, patients
who developed VAP were more likely to have diabetes mellitus and
immunosuppression (P = 0.020 and P = 0.014, respectively). These
patients also had higher APACHE II scores (P = 0.020) and a longer
duration of mechanical ventilation (P <0.001), and more frequently had
an open wound (P = 0.001). Following regression analysis, presence
of diabetes mellitus (OR: 12.048; 95% CI: 1.157 to 125.000; P = 0.037),
immunosuppression (OR: 16.949, 95% CI: 2.463 to 111.111; P = 0.004),
and open wound (OR: 5.714; 95% CI: 1.017 to 37.258; P = 0.048), higher
APACHE II scores (OR: 1.132; 95% CI: 1.022 to 1.254; P = 0.018), and
prolonged duration of mechanical ventilation (OR: 1.084; 95% CI: 1.002
to 1.171; P = 0.043) were determined as risk factors for VAP. Fourteen-
day and 28-day mortality rates for VAP were 19% and 29%, respectively
(P = 0.760 and P = 1.000, respectively). References 1. Marik PE, et al.: J Intensive Care Med 2012. [Epub ahead of print]
2. Piper AJ, et al.: Am J Respir Crit Care Med 2011, 183:292-298. 1. Marik PE, et al.: J Intensive Care Med 2012. [Epub ahead of print]
2. Piper AJ, et al.: Am J Respir Crit Care Med 2011, 183:292-298. 1. Marik PE, et al.: J Intensive Care Med 2012. [Epub ahead of print]
2. Piper AJ, et al.: Am J Respir Crit Care Med 2011, 183:292-298. Non-invasive ventilation in patients with a clinical diagnosis of
pneumonia: a clinical audit Results Three hundred and fi fty-nine subjects were analyzed. Table 1
shows the time on BiPAP per visit. Children whose weight was >97
percentile revealed trends towards longer treatment times on BiPAP
compared with the other two groups. The moderate subjects who
weighed >97 percentile had statistically signifi cant longer treatment
periods (P <0.006) when compared with the <90 percentile moderate
group. Initial BiPAP settings are listed in Table 2. When controlling for
age, higher BiPAP settings correlated with increasing weight. There
were no weight-based trends for admissions to the PICU or overall
hospital lengths of stay. No serious complications were noted.if Introduction The benefi ts of non-invasive ventilation (NIV) in patients
with type 2 respiratory failure secondary to exacerbations of COPD
are widely acknowledged [1], but its effi cacy in respiratory failure of
other aetiologies is less clear. We describe use of NIV (bilevel and/
or continuous positive airway pressure modes) in the subgroup of
patients with a clinical diagnosis of pneumonia in a 35-bed adult
critical care unit (ACCU). Methods A retrospective review of data recorded prospectively on
an electronic clinical information management system at University
College Hospital (UCH), London, UK. Patients requiring NIV acutely
at UCH are managed on the ACCU. Electronic records reviewed for
patients admitted from 1 November 2010 to 2 December 2011 with
clinical diagnosis of pneumonia, and in whom NIV was recorded on ≥6
occasions on the electronic chart. We concede some patients on NIV
<6 hours may have been missed. We excluded patients in whom NIV was
the ceiling of therapy and those using domiciliary NIV. The following
data were collected: baseline patient demographics, comorbidities,
therapy received in the ACCU, respiratory physiological variables,
length of stay/unit outcome. Descriptive statistics of the cohort and
variables associated on univariate analysis with requirement for
intubation were generated using MedCalc Version 12.3.0.0 (MedCalc
Software, Mariakerke, Belgium).i Conclusion This is the fi rst study to analyze the weight eff ect on BiPAP
application in children with status asthmaticus. Subjects who weighed
more trended greater mean time on BiPAP and initial BiPAP settings. Weight did aff ect PICU admissions or overall length of hospital stay. Analysis of bilevel positive airway pressure therapy in children
based on weight Subjects were divided into moderate and severe exacerbations and
then further subdivided into the following growth curve-based weight
subgroups: <90 percentile, 90 to 97 percentile and >97 percentile. Subjects received standard asthma therapies in addition to BiPAP. Data
were obtained at the bedside by the respiratory therapist or collected
retrospectively by study investigators. Data were stored and analyzed
using a RedCap database.i in status asthmaticus. Our purpose was to analyze BiPAP use and
outcomes for children with status asthmaticus and obesity in our PED. Methods Patients placed on BiPAP in the PED for status asthmaticus
from 1 January 2010 to 31 August 2012 were included in the analysis. Subjects were divided into moderate and severe exacerbations and
then further subdivided into the following growth curve-based weight
subgroups: <90 percentile, 90 to 97 percentile and >97 percentile. Subjects received standard asthma therapies in addition to BiPAP. Data
were obtained at the bedside by the respiratory therapist or collected
retrospectively by study investigators. Data were stored and analyzed
using a RedCap database.i Analysis of bilevel positive airway pressure therapy in children
based on weight Analysis of bilevel positive airway pressure therapy in children
based on weight
A Williams, T Abramo
Vanderbilt University Medical Center, Nashville, TN, USA
Critical Care 2013, 17(Suppl 2):P141 (doi: 10.1186/cc12079) A Williams, T Abramo
Vanderbilt University Medical Center, Nashville, TN, USA
Critical Care 2013, 17(Suppl 2):P141 (doi: 10.1186/cc12079) Introduction Obesity as a comorbidity adds a challenge when treating
children presenting to the PED in status asthmaticus. Bilevel positive
airway pressure (BiPAP) is an accepted treatment modality for children S53 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Table 1 (abstract P141). Time on BiPAP by weight percentile and severity of asthma exacerbation (hours)
Moderate exacerbation
Severe exacerbation Table 1 (abstract P141). Time on BiPAP by weight percentile and severity of asthma exacerbation (hours) Methods A monocentric prospective observational study including 48
patients with MOHS and treated by NIV for ARF over a 2-year period. Results NIV failed to reverse ARF in only seven patients. NIV failure
was associated with a sixfold increase in in-hospital mortality (85.7%
vs. 14.6%; P <0.001). Factors associated with NIV failure and need for
endotracheal intubation included pneumonia (n = 5, 71.4% vs. n = 8,
19.5%; P = 0.01), higher SOFA (11 vs. 5; P <0.001) and SAPS 2 (69 vs. 39; P = 0.001) score at admission. The only factor associated with
successful response to NIV was idiopathic decompensated MOHS
(n = 19, 100% vs. n = 0, 0%; P = 0.03). In these patients, pH (7.23 vs. 7.28
vs. 7.44, respectively, at H0, H2, H24; P <0.05) and PCO2 (74.5 vs. 68.5 vs. 57 mmHg, respectively, at H0, H2, H24; P <0.05) dramatically improved
with NIV.fi Conclusion NIV provides a very effi cient therapy to reverse hypercapnic
ARF in morbidly obese patients. Multiple organ failure and pneumonia
are the main factors associated with NIV failure in patients with MOHS. Conclusion NIV provides a very effi cient therapy to reverse hypercapnic
ARF in morbidly obese patients. Multiple organ failure and pneumonia
are the main factors associated with NIV failure in patients with MOHS. References in status asthmaticus. Our purpose was to analyze BiPAP use and
outcomes for children with status asthmaticus and obesity in our PED. Methods Patients placed on BiPAP in the PED for status asthmaticus
from 1 January 2010 to 31 August 2012 were included in the analysis. Non-invasive ventilation in patients with a clinical diagnosis of
pneumonia: a clinical audit
CJ Wright1, J Hornsby2, DA Barr3, SR Moonesinghe2
1Glasgow Royal Infi rmary, Glasgow, UK; 2University College Hospital, London,
UK; 3Brownlee Institute, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P143 (doi: 10.1186/cc12081) Non-invasive ventilation in patients with a clinical diagnosis of
pneumonia: a clinical audit
CJ Wright1, J Hornsby2, DA Barr3, SR Moonesinghe2
1Glasgow Royal Infi rmary, Glasgow, UK; 2University College Hospital, London,
UK; 3Brownlee Institute, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P143 (doi: 10.1186/cc12081) P142 Determinants of non-invasive ventilation success or failure in
morbidly obese patients in acute respiratory failure
M Lemyze, P Taufour, J Mallat, O Nigeon, D Thevenin
Schaff ner Hospital, Lens, France
Critical Care 2013, 17(Suppl 2):P142 (doi: 10.1186/cc12080) Introduction Malignant obesity hypoventilation syndrome (MOHS)
refers to the association between morbid obesity (body mass index
>40 kg/m2), daytime hypercapnia (PCO2 >45 mmHg) after other
respiratory or neuromuscular causes of alveolar hypoventilation have
been excluded, and multiple organ disorders (diabetes mellitus, arterial
hypertension, metabolic syndrome, obstructive sleep apnea syndrome)
[1]. Acute respiratory failure (ARF) is a common life-threatening
complication of MHOS. Non-invasive ventilation (NIV) provides the
cornerstone of the therapeutic management of decompensated MOHS
[2]. We aimed to identify the determinants of NIV success or failure in
this indication. g
Results Fifty-two patients fulfi lled the inclusion criteria, 29 (55.8%)
were male, median age was 62.4 (IQR 48.8 to 71.6), median unit stay
was 8 days (IQR 4.5 to 16.5), mean duration of NIV use was 2 days (IQR 1
to 3), 27 (51.9%) required intubation of whom 48% died. Total mortality
was 13 (25%) – all intubated. Patients with recorded comorbidities were
more likely to be intubated than those without (OR = 4.3, P = 0.0362);
pH was signifi cantly lower in those requiring intubation at all recorded
time points; and mean FiO2 at 4 to 6 hours was signifi cantly higher in
those requiring intubation(0.72 vs. 0.56, P = 0.032). There was a trend S54 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 toward patients with COPD requiring intubation more often (OR = 8.4,
P = 0.0711). are specialized and relatively invasive. Visualizing diaphragm thickening
during inspiration by ultrasound may permit non-invasive monitoring. We explored the feasibility, reliability and physiological signifi cance of
diaphragm thickening on ultrasound. Conclusion NIV was successful in 48.1% of patients with pneumonia,
the remainder requiring intubation. Given the high mortality in those
patients who failed NIV we believe its use in a ward setting is hazardous. We conclude that NIV may be useful in some individuals with
pneumonia, but they should be managed in the ACCU setting. Further
work is required to identify those patients at risk of deterioration on NIV
given the possible excess mortality. Methods Five healthy subjects participated. Analysis of management of non-invasive ventilation support in
prehospital care for COPD patients and short-term outcome
J Cuny, G Campagne, P Gosselin, P Goldstein, N Assez, E Wiel
Lille University Hospital Center, Lille Cedex, France
Critical Care 2013, 17(Suppl 2):P144 (doi: 10.1186/cc12082) Analysis of management of non-invasive ventilation support in
prehospital care for COPD patients and short-term outcome
J Cuny, G Campagne, P Gosselin, P Goldstein, N Assez, E Wiel
Lille University Hospital Center, Lille Cedex, France
Critical Care 2013, 17(Suppl 2):P144 (doi: 10.1186/cc12082) y
Results Diaphragm thickening in the zone of apposition was readily
visualized by ultrasound in all fi ve subjects. Mean end-expiratory
diaphragm thickness was 2.1 mm (SD = 0.3 mm). During tidal
breathing, the diaphragm thickened by a mean of 35% (SD = 31%). The
Bland–Altman coeffi cient of reproducibility was 0.5 mm; approximately
50% of measurement variability arose from caliper positioning on the
ultrasound machine; diaphragm thickness measurements changed as
the probe was placed in diff erent intercostal interspaces. Diaphragm
inspiratory
thickening
increased
signifi cantly
with
increasing
inspiratory eff ort but also varied with lung volume independent of
eff ort. At inspiratory volumes below 40% of inspiratory capacity, lung
volume change contributed minimally to diaphragm thickening. Introduction Benefi ts of the use of NIV in emergency departments are
well established. Training and available staff , and choice of respiratory
machines are essential criteria for success. Methods We conducted an observational, descriptive, retrospective,
single-center study in a 4-month period. COPD patients with respiratory
failure who received prehospital NIV were included. We compared two
groups: COPD patients with NIV, and COPD patients without NIV. g
p
p
p
Results Forty-two patients were included, mean age 68.86 years
(± 11.98), 57.14% smokers, 64.28% arterial hypertension, 100% long-
term oxygenotherapy, 23.80% antibiotics in the 7 days before, 28.57%
corticosteroids. A total of 88.09% had bronchospasm, 78.26% had
struggle signs, 28.57% were unable to speak, 14.28% of patients were
sweating. The mean respiratory rate was 30.5 cycles/minute (± 7.17),
mean pulse rate was 105.76 (± 25.34). Nasal EtCO2 was 47.75 mmHg
(± 16.53), pulse oxymetry in air was 85% (± 10.94), oxygen fl ow
rate was 5.45 l/minute (± 2.42), temperature was 37.14°C (± 8.15). Twenty patients received NIV. A total of 61.90% were admitted to
the emergency department, 35.71% to the ICU, and one patient
was left at home. One patient received tracheal intubation in the
hosting service. Mortality in the fi rst month was 13.04%. Analysis of management of non-invasive ventilation support in
prehospital care for COPD patients and short-term outcome
J Cuny, G Campagne, P Gosselin, P Goldstein, N Assez, E Wiel
Lille University Hospital Center, Lille Cedex, France
Critical Care 2013, 17(Suppl 2):P144 (doi: 10.1186/cc12082) A signifi cant
diff erence (P <0.05) was found for: sweats (30%/0), respiratory rate
(34 ± 8.23/27 ± 6.11), nasal EtCO2 (55.0 ± 24.4/40.50 ± 9.03), pulse
oxymetry in air (80% ± 8.63/90% ± 13.25), pulse oxymetry with
oxygen (89.4% ± 4.24/87.90% ± 2.55), β2-mimetic and anti-cholinergic
nebulization (60% ± 0.5/90% ± 0.29), emergency room admission
(35% ± 0.35/86% ± 0.48), ICU admission (60% ± 0.5/13% ± 0.35),
arterial blood gases on arrival in the host service (PaCO2
76.6 ± 18.66/43.93 ± 11.78). No diff erence in mortality at 1 month (2/3). Conclusion Non-invasive ventilation has improved the management
and prognosis of COPD patients admitted to the emergency room. Very few studies concern the prehospital management. NIV seems to
show an eff ect on prehospital care, especially in patients with signs of
severity, hypercapnia, and without fever. Oxygenation and hypercapnia
seem to be improved. Also fewer patients are admitted to the ICU. Bronchospasm does not seem to be an obstacle. R f y
Conclusion Visualizing diaphragm thickening in the zone of appo-
sition by ultrasound provides a feasible non-invasive technique for
monitoring diaphragm activation in healthy subjects. Diaphragm
thickening primarily refl ects muscular eff ort rather than altered muscle
conformation induced by changes in lung volume, especially at lower
inspiratory volumes. Reference 1. Brochard L, et al.: Noninvasive ventilation for acute exacerbations of
chronic obstructive pulmonary disease. N Engl J Med 1995, 333:817-822. Retrospective review of utilisation and outcomes of diaphragmatic
EMG monitoring and neurally adjusted ventilatory assist in a central
London teaching hospital over a 3-year periodi Retrospective review of utilisation and outcomes of diaphragmatic
EMG monitoring and neurally adjusted ventilatory assist in a central
London teaching hospital over a 3-year period
A Skorko, D Hadfi eld, A Vercueil, C Bell, A Feehan, K Peters, P Hopkins
King’s Health Partners AHSC, London, UK
Critical Care 2013, 17(Suppl 2):P146 (doi: 10.1186/cc12084) Introduction The theoretical advantages of monitoring the electrical
activity of the diaphragm (EAdi) and neural triggering of support
breaths (NAVA-Maquet) have not yet been shown to translate into
signifi cant clinical benefi t [1]. Here we assess the eff ect of EAdi
monitoring, in patients at risk of prolonged weaning, on outcomes. Figure 1 (abstract P146). Figure 1 (abstract P146). 1. Schmidbauer W, et al.: Early pre hospital use of NIV. Emerg Med J 2011,
28:626-627. P145 P145
A window into diaphragm kinetics: feasibility, precision, and
physiological meaning of ultrasound measurements of diaphragm
thickness
EC Goligher1, F Laghi2, ND Ferguson1
1University of Toronto, Canada; 2Loyola University, Chicago, IL, USA
Critical Care 2013, 17(Suppl 2):P145 (doi: 10.1186/cc12083) P142 We monitored inspiratory
fl ow, volume, esophageal and gastric pressures, and diaphragm
electrical activity (by esophageal and surface electromyography) while
subjects performed a series of inspiratory maneuvers: tidal breathing,
threshold-loaded breathing, a Muller maneuver, and inspiration to
various lung volumes above functional residual capacity. At the end
of each inspiratory eff ort, subjects were instructed to close the glottis
and relax the respiratory muscles (so as to maintain lung volume while
eliminating diaphragm activation). Sonographic images of diaphragm
thickening during these maneuvers were obtained using M-mode with
a 13 MHz linear array probe placed in the right ninth, 10th, or 11th
intercostal space between the middle and anterior axillary lines. Risk of weaning failure with PAOP ≥15 versus PAOP ≥18
Y Nassar, A Abdelbary Risk of weaning failure with PAOP ≥15 versus PAOP ≥18
Y Nassar, A Abdelbary
Cairo University, Giza, Egypt
Critical Care 2013, 17(Suppl 2):P148 (doi: 10.1186/cc12086) Cairo University, Giza, Egypt
Critical Care 2013, 17(Suppl 2):P148 (doi: 10.1186/cc12086) Introduction Despite 18 mmHg being recognized as a classical cutoff
value, there is no defi nite value of PAOP above which cardiogenic
oedema develops. We aimed to assess the risks of weaning failure with
PAOP ≥15 and with PAOP ≥18. Results In total, 493/2,684 (18.3%) patients had heart–lung risk factors
for prolonged weaning. One hundred and four patients received EAdi
monitoring. Ventilated days were signifi cantly reduced (Figure 1) in
the EAdi monitored group (median 9) versus the nonmonitored group
(median 12); P = 0.024 (Mann–Whitney U test). ICU mortality was not
signifi cantly diff erent and there was no correlation between time spent
in NAVA and days ventilated.i Methods A prospective enrollment of 30 adult medical ICU patients
on invasive mechanical ventilation who were fulfi lling the criteria for
initiation of weaning from mechanical ventilation according to the
Sixth International Conference Statement of Intensive Care Medicine. We followed the two-step weaning strategy, which involves assessment
regarding readiness for weaning followed by a spontaneous breathing
trial (SBT) as a diagnostic test to determine the likelihood of successful
extubation. Invasive right heart catheterization to measure PAOP, and
non-invasive transthoraxic echo and tissue Doppler parameters E/E’,
E/A, DT, IVRT were measured before initiation and after termination of
the SBT. Conclusion EAdi catheter insertion was associated with a signifi cant
reduction in time spent on MV. A prospective RCT will be needed to
confi rm benefi t and explore mechanisms. Reference 1. Brander L, et al.: Chest 2009, 135:695-703. Results E/E’ was signifi cantly correlated with PAOP (r = 503, P <0.001). A
cutoff value of septal E/E’ >11.3 predicted PAOP elevation ≥18 mmHg
with a sensitivity of 90.9% and specifi city of 87.8%, while a value of
septal E/E’ >10.9 predicted PAOP elevation ≥15 mmHg with a sensitivity
of 59.1% and specifi city of 87.8%. The incidence of weaning failure
was 23%, and 72% of patients who failed to be weaned exhibited
signifi cantly higher PAOP during the trial. Patients with PAOP ≥18 at
the end of the SBT mostly (72.4%) failed weaning while a minority
(27.6%) successfully weaned. 9
Impact of continuous positive airway pressure on the treatment of
acute exacerbation of asthma: a randomised controlled trial
Y Sutherasan, S Kiatboonsri, P Theerawit, C Kiatboonsri, S Trakulsrichai
Ramathibothi Hospital, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P149 (doi: 10.1186/cc12087) Introduction Despite the extensive use of continuous positive airway
pressure (CPAP) in various respiratory failure conditions, its role in acute
asthmatic attack is uncertain [1-3]. This study aims at exploring the
effi cacy of CPAP when use in addition to the conventional treatment of
acute asthma exacerbation in the emergency department (ED). Results Forty-two patients were evaluated (42% male), mean age was
73 years (SD ± 15) and APACHE II score was 18.77 (median 20). Based
upon evaluation of the fi ndings of BLUE protocol, a positive correlation
was found in 83.78% of the sample. For the diagnosis of pulmonary
edema (n = 15), sensitivity was 85% and specifi city was 86%, while for
the diagnosis of pneumonia (n = 17), sensitivity was 88% and specifi city
was 90%. In this sample there were only four cases of reversible airway
obstruction and one case of pulmonary embolism. Five patients were
excluded of the analysis because they had rare diagnoses. g
y
Methods During May to December 2009, acute asthma patients
attending ED were randomly assigned to CPAP and control groups. In
addition to the conventional treatment, CPAP ventilation of 8 cmH2O
was applied to patients in the CPAP group. Data collections in both
groups included peak expiratory fl ow rates (PEFR, %predicted) at
each 15-minute interval (T0, T15, …), the number of short-acting
bronchodilators (SABA) used, ED length of stay, relapse rate and
admission rate. The primary outcome is the improvement in peak
expiratory fl ow rate during a short period stay in the ED. Results There were 86 patients enrolled, with 43 patients in each
study group. PEFR in the CPAP group showed no diff erence from the
control group at T0 (52.7 ± 20.33 vs. 47.2 ± 16.64, P = NS) but was
signifi cantly better at T15 (65.58 ± 21.64 vs. 56.49 ± 18.53%, P <0.05). Such improvement was consistently observed for all subsequent PEFR
measurement, along with the fewer SABA used and relapse rate, but
the fi gures did not meet statistical diff erence. Methods During May to December 2009, acute asthma patients
attending ED were randomly assigned to CPAP and control groups. In
addition to the conventional treatment, CPAP ventilation of 8 cmH2O
was applied to patients in the CPAP group. 1.
Lichtenstein DA, Mezière GA: Relevance of lung ultrasound in the diagnosis
of acute respiratory failure: the BLUE protocol. Chest 2008, 134:117-125. 1.
Brander L, et al.: Chest 2009, 135:695-703. 1.
Lichtenstein DA, Mezière GA: Relevance of lung ultrasound in the diagnosis
of acute respiratory failure: the BLUE protocol Chest 2008 134:117 125 Risk of weaning failure with PAOP ≥15 versus PAOP ≥18
Y Nassar, A Abdelbary The majority (91.3%) of the patients
with PAOP <18 had a successful weaning outcome and 8.7% failed
weaning outcome. PAOP ≥18 signifi cantly negatively correlated with
weaning success (P = 0.001, r = –0.627). On the other hand, PAOP ≥15
did not show a signifi cant correlation (P >0.05, r = –0.274). The PAOP
≥18 weaning failure risk estimate was 8.2, with 95% CI 0.004 to 0.3. The
PAOP ≥15 weaning failure risk estimate was 1.83, with a signifi cant 95%
CI of 0.913 to 3.648.i P147
Validation of a lung ultrasound protocol in acute respiratory failure:
preliminary results
RV Cremonese1, AC Tabajara Raupp1, JM Stormovski de Andrade1,
RS Townsend1, FS Neres1, AS Machado1, MC Prestes1, JG Maccari2,
FL Dexheimer Neto1
1Ernesto Dornelles, Porto Alegre, Brazil; 2Hospital Moinhos de Vento, Porto
Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P147 (doi: 10.1186/cc12085) P147
Validation of a lung ultrasound protocol in acute respiratory failure:
preliminary results
RV Cremonese1, AC Tabajara Raupp1, JM Stormovski de Andrade1,
RS Townsend1, FS Neres1, AS Machado1, MC Prestes1, JG Maccari2,
FL Dexheimer Neto1
1Ernesto Dornelles, Porto Alegre, Brazil; 2Hospital Moinhos de Vento, Porto
Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P147 (doi: 10.1186/cc12085) Critical Care 2013, 17(Suppl 2):P147 (doi: 10.1186/cc12085) Introduction Patients with acute respiratory failure demand dynamic
evaluation and interventions. Lung ultrasound is a bedside technique,
very promising in this setting. Previously, Lichtenstein and colleagues
proposed an algorithm approach with a diagnostic accuracy of 90.5%,
named Bedside Lung Ultrasound in Emergency (BLUE protocol). However, this approach has never been validated in other populations
or ultrasound operators. The aim of this study was to evaluate the
diagnostic accuracy of the lung ultrasound algorithm (BLUE protocol)
in ICU patients admitted with acute respiratory failure. Conclusion E/E’ was signifi cantly correlated with PAOP (r = 503,
P <0.001). E/E’ > 11.3 predicted PAOP elevation ≥18 mmHg (sensitivity
90.9% and specifi city 87.8%), while E/E’ >10.9 predicted PAOP elevation
≥15 mmHg (sensitivity 59.1% and specifi city 87.8%). Weaning failure
risk is greater (8.2 vs. 1.8) with PAOP ≥18 versus PAOP ≥15, respectively. P149 y
Methods A prospective study, single-center consecutive case series
evaluating critically ill adult patients in acute respiratory failure,
admitted to a 23-bed general ICU between October 2011 and
November 2012. The ultrasound examination was performed by an ICU
physician until 20 minutes after admission, without interfering with
usual care, and the ultrasound operator was blinded to the medical
history of the patient. Three items were assessed: artifacts (horizontal
A lines or vertical B lines indicating interstitial syndrome), lung sliding,
alveolar consolidation and/or pleural eff usion. BLUE protocol results
were compared with clinical diagnosis by the medical assistant team
at the end of the ICU stay. P149
Impact of continuous positive airway pressure on the treatment of
acute exacerbation of asthma: a randomised controlled trial
Y Sutherasan, S Kiatboonsri, P Theerawit, C Kiatboonsri, S Trakulsrichai
Ramathibothi Hospital, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P149 (doi: 10.1186/cc12087) A window into diaphragm kinetics: feasibility, precision, and
physiological meaning of ultrasound measurements of diaphragm
thickness EC Goligher1, F Laghi2, ND Ferguson1 g
,
g
,
g
1University of Toronto, Canada; 2Loyola University, Chicago, IL, USA
Critical Care 2013, 17(Suppl 2):P145 (doi: 10.1186/cc12083) Introduction Inadequate respiratory muscle activity has been linked
to ventilator-induced diaphragm dysfunction. Current techniques for
monitoring respiratory muscle activity during mechanical ventilation S55 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P148 Methods Institutional ethics approval was obtained. The Medtrack
clinical information system was searched to identify patients who
received MV for >48 hours and who had signifi cant chronic pulmonary
disease or left/right ventricular impairment between April 2009 and
March 2012. Age, APACHE II score, ventilated days, time in NAVA and
outcome were compared between groups who had or had not received
EAdi monitoring. P150
Novel method to confi rm tracheal intubation based on airway
pressures: validation in patients with lung disease
A Stemerdink1, KG Monsieurs2, F Luijmes1, W Dieperink1, JG Zijlstra1,
AF Kalmar1
1University Medical Center Groningen, the Netherlands; 2Antwerp University
Hospital, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P150 (doi: 10.1186/cc12088) P150
Novel method to confi rm tracheal intubation based on airway
pressures: validation in patients with lung disease
A Stemerdink1, KG Monsieurs2, F Luijmes1, W Dieperink1, JG Zijlstra1,
AF Kalmar1
1University Medical Center Groningen, the Netherlands; 2Antwerp University
Hospital, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P150 (doi: 10.1186/cc12088) P150
Novel method to confi rm tracheal intubation based on airway
pressures: validation in patients with lung disease
A Stemerdink1, KG Monsieurs2, F Luijmes1, W Dieperink1, JG Zijlstra1,
AF Kalmar1
1University Medical Center Groningen, the Netherlands; 2Antwerp University
Hospital, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P150 (doi: 10.1186/cc12088) Results A total of 470 BC were carried out in 240 patients admitted to
the ICU (68% were male and 32% were female). All of the procedures
were performed in mechanically ventilated patients. The mean patient
age was 61 ± 14 years, with an APACHE II score at admission of 20 ± 7. The
main cause for ICU admission was respiratory failure (27%). The most
common comorbidities were smoking, chronic obstructive pulmonary
disease, cardiovascular risk factors and atrial fi brillation. The main
indications for performing the BC were percutaneous tracheostomy
endoscopically guided (28%), resolution of atelectasis (27%), aspiration
of secretions (24%) and hemoptysis (10%). Bronchial brushing was
performed in 435 cases (93%) obtaining microbiological growth in 64%
of them, mainly Pseudomonas aeruginosa and Acinetobacter baumanii. Bronchoalveolar lavage was performed in 62 cases (13%) obtaining
microbiological growth in 61% of them, mainly cytomegalovirus and
Stenotrophomona maltophilia. Citology was performed in 60 cases
(13%); 7% showed the presence of microorganism and 13% revealed
precancerous/cancerous tissue changes. Introduction Emergency endotracheal intubation results in accidental
oesophageal intubation in up to 17% of patients often with disastrous
consequences. We have previously published a highly specifi c and
sensitive novel method to detect endotracheal intubation based on
diff erences in ventilation pressure waveforms in the oesophagus and
in the trachea in patients with healthy lungs [1]. A detection algorithm,
based on diff erences in compliance/elasticity between the lung and
the oesophagus, generated a D-value indicating tracheal intubation if
D >0.5 and oesophageal intubation if D <0.5. The aim of the current
study was to validate the algorithm in patients with lung disease. p
g
Conclusion BC can be performed at the bedside, avoiding transfers
outside the ICU that may cause clinical worsening. Most frequent
indications were bronchoscopy-guided percutaneous tracheostomy,
resolution of atelectasis, aspiration of secretions and hemoptysis. P150
Novel method to confi rm tracheal intubation based on airway
pressures: validation in patients with lung disease
A Stemerdink1, KG Monsieurs2, F Luijmes1, W Dieperink1, JG Zijlstra1,
AF Kalmar1
1University Medical Center Groningen, the Netherlands; 2Antwerp University
Hospital, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P150 (doi: 10.1186/cc12088) Microbiological results obtained revealed the presence of aggressive
pathogens. BC in ICU provides valuable diagnostic information and has
therapeutic utility. y
g
p
g
Methods After obtaining institutional approval, 20 intubated and
ventilated ICU patients were included. Inclusion criteria were controlled
mechanical ventilation and at least mild to moderate lung injury
according to a Murray lung injury score >0.1. A connecting piece was
placed between the endotracheal tube and the ventilation bag. This
piece comprised a thin air-fi lled catheter inserted through the tube
lumen at 1 cm from the distal end, and a second catheter located at
the proximal end of the tube. We performed three consecutive manual
bag ventilations while recording the pressure curves through both
catheters. For each ventilation, a D-value was calculated. y
References References
1. Shivaram U, et al.: Eff ects of continuous positive airway pressure in acute
asthma. Respiration 1987, 52:157-162. 2. Meduri GU, et al.: Noninvasive positive pressure ventilation in status
asthmaticus. Chest 1996, 110:767-774. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. 1. Shivaram U, et al.: Eff ects of continuous positive airway pressure in acute
asthma. Respiration 1987, 52:157-162. 2. Meduri GU, et al.: Noninvasive positive pressure ventilation in status
asthmaticus. Chest 1996, 110:767-774. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. 1. Shivaram U, et al.: Eff ects of continuous positive airway pressure in acute
asthma. Respiration 1987, 52:157-162. 2. Meduri GU, et al.: Noninvasive positive pressure ventilation in status
asthmaticus. Chest 1996, 110:767-774. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. 1. Shivaram U, et al.: Eff ects of continuous positive airway pressure in acute
asthma. Respiration 1987, 52:157-162. Introduction Bronchoscopy (BC) is a frequently used diagnostic and
therapeutic tool in our ICU. We planned to review the patients and
conditions in which the procedure was performed. The objective was
to determine indications, diagnostic and therapeutic yield. 2. Meduri GU, et al.: Noninvasive positive pressure ventilation in status
asthmaticus. Chest 1996, 110:767-774. 2. Meduri GU, et al.: Noninvasive positive pressure ventilation in status
asthmaticus. Chest 1996, 110:767-774. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. 3. Lin HC, et al.: Eff ect of nasal continuous positive airway pressure on
methacholine-induced bronchoconstriction. Respir Med 1995, 89:121-128. Methods A retrospective, single-center observational study was carried
out. All BC performed in a 20-bed polyvalent ICU of a tertiary university
hospital over a period of 30 months (from January 2010 to June 2012)
were included. The main variables collected were demographic data
and clinical data including comorbidities, endobronchial techniques
and results. References 1. Lucena CM, et al.: Med Intensiva 2012, 36:389-395. 2. Estella A, et al.: Med Intensiva 2012, 36:396-401. 3. Shennib H, et al.: Chest Surg Clin N Am 1996, 6:349-361. 4. Turner JS, et al.: Crit Care Med 1994, 22:259-264. 1. Lucena CM, et al.: Med Intensiva 2012, 36:389-395. Results Mean age (SD) of the patients was 60 (16) years, 60% were
male. The mean (SD) Murray score was 1.4 (0.6). Pathologies included
pulmonary oedema, pneumonia, atelectasis and traumatic lung
injury. All 60 D-values are represented in Figure 1. The median
(IQR, range) D-value was 38 (16 to 74, 0.8 to 1,272). Our algorithm
therefore confi rmed a high sensitivity to detect correct endotracheal
intubation also in patients with lung disease. Under the hypothesis that
oesophageal compliance does not increase signifi cantly in patients
with lung disease, the specifi city of our algorithm will not be aff ected. 9
Impact of continuous positive airway pressure on the treatment of
acute exacerbation of asthma: a randomised controlled trial
Y Sutherasan, S Kiatboonsri, P Theerawit, C Kiatboonsri, S Trakulsrichai
Ramathibothi Hospital, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P149 (doi: 10.1186/cc12087) Data collections in both
groups included peak expiratory fl ow rates (PEFR, %predicted) at
each 15-minute interval (T0, T15, …), the number of short-acting
bronchodilators (SABA) used, ED length of stay, relapse rate and
admission rate. The primary outcome is the improvement in peak
expiratory fl ow rate during a short period stay in the ED. Conclusion Preliminary results demonstrate diagnostic accuracy of
lung ultrasound examination performed on the admission of patients
with acute respiratory failure similar to the original publication. Lung
ultrasound, as proposed in the BLUE protocol, has good accuracy, and
seems reproducible and useful in this group of patients. f p
yl
g
p
y
Results There were 86 patients enrolled, with 43 patients in each
study group. PEFR in the CPAP group showed no diff erence from the
control group at T0 (52.7 ± 20.33 vs. 47.2 ± 16.64, P = NS) but was
signifi cantly better at T15 (65.58 ± 21.64 vs. 56.49 ± 18.53%, P <0.05). Such improvement was consistently observed for all subsequent PEFR
measurement, along with the fewer SABA used and relapse rate, but
the fi gures did not meet statistical diff erence. 1. Lichtenstein DA, Mezière GA: Relevance of lung ultrasound in the diagnosis
of acute respiratory failure: the BLUE protocol. Chest 2008, 134:117-125. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S56 http://ccforum.com/supplements/17/S2 Conclusion The addition of CPAP to conventional acute asthma
treatment in the ED could accelerate PEFR improvement with a trend
showing fewer SABA needed, lower relapse rate and shorter ED length
of stay. P151
Main indications, diagnostic and therapeutic yield of bronchoscopy
in the ICU J Iglesias, B Gonzalez, O Rajas
La Princesa Hospital, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P151 (doi: 10.1186/cc12089) P152 P152
Benchtop study comparing leakages across cuff s of three
endotracheal tubes
SM Lam, CW Lau, WW Yan
Pamela Youde Nethersole Eastern Hospital, Hong Kong
Critical Care 2013, 17(Suppl 2):P152 (doi: 10.1186/cc12090) Conclusion The algorithm to detect correct endotracheal intubation
performed excellent in patients with lung disease. Reference Introduction The aim was to compare two novel endotracheal tubes
(ETT), Mallinckrodt TaperGuard (TG, tapered polyvinyl chloride (PVC)
cuff ) and KimVent Microcuff (MC, cylindrical polyurethrane cuff ), 1. Kalmar AF, et al.: Resuscitation 2012, 83:177-182. Figure 1 (abstract P150). D-value of individual test ventilations. Figure 1 (abstract P150). D-value of individual test ventilations. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2 S57 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P152). Downward leak in PEEP 0 cmH2O without
suction. Each point represents the mean and standard deviation of eight
repeated measures. with SSD. However, the shape most suitable for preventing dripping
onto the subglottis has not been determined. The purpose of this study
was to determine whether an ET with tapered-type cuff can reduce the
incidence of VAP. Methods After approval from the appropriate ethics committee, we
conducted a single-institutional prospective randomized clinical trial
on the eff ectiveness of using an ET with a diff erent cuff type. Adult
patients (n = 399; ≥18 years old) were screened between April 2010
and March 2012, and 289 patients expected to require mechanical
ventilation (MV) for at least 48 hours were randomized. Patients were
assigned to intubation with one of the following two HVLP ETs with
SSD on the experimental tube: tapered type (Taperguard™ Evac®;
Covidien, Dublin, Ireland) or spindle type (Hi-Lo Evac®; Covidien). Cuff
pressure measurement was carried out every 6 hours, and cuff pressure
was maintained between 20 and 30 cmH2O. The primary outcome was
VAP incidence based on semiquantitative bronchoalveolar lavage fl uid
culture of 3+ or phagocytosis on Gram staining in patients intubated
for at least 48 hours. The other outcome was the rate of achieving an
appropriate cuff pressure (20 to 30 cmH2O), time to VAP onset, duration
of MV, duration of ICU stay, mortality, and adverse events. Figure 1 (abstract P152). Downward leak in PEEP 0 cmH2O without
suction. Each point represents the mean and standard deviation of eight
repeated measures. Eff ect of body position changes on endotracheal tube cuff pressure
in ventilated patients Fernandez JF, et al.: Chest 2012, 142:231-238. 1.
Fernandez JF, et al.: Chest 2012, 142:231-238. P152 y
y
Results The rate of microbiologically confi rmed VAP was 21.7%
(23/106) for the tapered-type ET patients and 21.7% (23/106) for the
spindle-type ET patients (P = 1.00). The rate of achieving appropriate
cuff pressure was 83.2% (332/1,974) for the tapered type and 82.4%
(328/1,867) for the spindle type (P = 0.549). No signifi cant diff erences
between groups were observed for time to VAP onset, duration of MV,
ICU stay. The incidence of reintubation due to laryngeal edema after
extubation was slightly higher in patients with the tapered-type ET
(11.5%, 6/52) than in patients with the spindle-type ET (2.0%, 1/49), but
the diff erence was not signifi cant (P = 0.113).ff with conventional Portex (PT, globular PVC cuff ) in leakages across
cuff s (microaspiration) under simulated clinical situations. It has been
shown that globular PVC cuff s protect poorly against leakages due
to microchannels formed from infolding of redundant cuff material
[1]. We hypothesized that TG and MC better prevent microaspiration,
which is a major mechanism of ventilator-associated pneumonia (VAP). Methods Each ETT was inserted into a silicone cylinder of 2 cm
wide inclined at 35°. Then 20 ml water was added above the cuff
and leakage measured every minute under fi ve diff erent simulated
clinical conditions: mechanical ventilation for acute severe asthma
(positive end-expiratory pressure (PEEP) 0 cmH2O), normal lungs (PEEP
5 cmH2O) and acute respiratory distress syndrome (PEEP 10 cmH2O),
and disconnection from the ventilator with and without spontaneous
breathing eff ort. Spontaneous breathing was simulated with a
respiratory gas exchange simulator. Suction was applied at 200 cmH2O
sustained for 3 minutes at the Murphy eye. Each scenario was repeated
with cuff pressures (Pcuff ) 10, 20 and 30 cmH2O maintained by a Pcuff
maintenance device. Conclusion Diff erences in cuff type and shape under identical
conditions of cuff pressure control have no infl uence on the incidence
of VAP. Acknowledgement UMIN-CTR UMIN000003371. Acknowledgement UMIN-CTR UMIN000003371. P154f Eff ect of body position changes on endotracheal tube cuff pressure
in ventilated patients Eff ect of body position changes on endotracheal tube cuff pressu
in ventilated patients
C Lizy1, W Swinnen2, S Labeau3, S Blot1
1Gent University, Ghent, Belgium; 2General Hospital Sint Blasius,
Dendermonde, Belgium; 3University College Ghent, Belgium
Critical Care 2013, 17(Suppl 2):P154 (doi: 10.1186/cc12092) C Lizy1, W Swinnen2, S Labeau3, S Blot1
1Gent University, Ghent, Belgium; 2General Hospital Sint Blasius,
Dendermonde, Belgium; 3University College Ghent, Belgium
Critical Care 2013, 17(Suppl 2):P154 (doi: 10.1186/cc12092) Results PT leaked grossly in all scenarios without PEEP and at PEEP
5 cmH2O in the presence of suction irrespective of Pcuff (Pcuff
30 cmH2O: PT 19.7 ± 0.4, TG 0.0 ± 0.1, MC 0.0 ± 0.0, P <0.001; Pcuff
20 cmH2O: PT 19.9 ± 0.4, TG 7.4 ± 6.2, MC 0.0 ± 0.0, P <0.001; Pcuff
10 cmH2O: PT 20.0 ± 0.0, TG 12.7 ± 5.1, MC 0.9 ± 0.8, P <0.001). Leakage
under these scenarios can be reduced in TG and prevented in MC by
Pcuff ≥20 cmH2O (Figure 1).f Introduction In order to avoid either microaspiration (<20 cmH2O) or
tracheal injury (>30 cmH2O), the target for endotracheal tube (ETT) cuff
pressure (CP) is between 20 and 30 cmH2O. Our objective was to assess
the eff ect of patient positioning changes on CP in adult patients. f
2
Conclusion Microcuff outperformed the others in preventing
microaspiration, while Portex leaked grossly even at a recommended
Pcuff of 20 to 30 cmH2O whenever PEEP was lost. The eff ect of ETT type
on the incidence of VAP warrants further investigation. Reference Methods A sample of 12 orally intubated and sedated patients was
selected. Patients received neuromuscular blockade and were positioned
in a neutral start position (backrest, head of bed elevation (HoB)
30°, head in neutral position) and CP set at 25 cmH2O. Subsequently
16 diff erent position changes were performed: antefl exion head,
hyperextension head, left and right lateral fl exion of head, left and right
rotation of the head, semirecumbent position (HoB 45°), recumbent
position (HoB 10°), horizontal position, trendelenburg (–10°), left
and right lateral positioning over 30°, 45° and 90°. Once the patient
was correctly positioned, CP was recorded during an end-expiratory
ventilatory hold. CP observed was compared with CP at start position
(25 cmH2O). Also the number of values outside the target range (20 to
30 cmH2O) was reported as considered clinically relevant. 1. Fernandez JF, et al.: Chest 2012, 142:231-238. 1. P153
Endotracheal tube with tapered-type cuff for preventing
ventilator-associated pneumonia: a randomized clinical trial
N Saito, T Yagi, Y Hara, H Matsumoto, K Mashiko
Nippon Medical School Chiba Hokusoh Hospital, Inzai, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P153 (doi: 10.1186/cc12091) P153
Endotracheal tube with tapered-type cuff for preventing
ventilator-associated pneumonia: a randomized clinical trial
N Saito, T Yagi, Y Hara, H Matsumoto, K Mashiko
Nippon Medical School Chiba Hokusoh Hospital, Inzai, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P153 (doi: 10.1186/cc12091) 2
y
Results A total of 192 measurements were performed (12 subjects×16
positions). Results are shown in Table 1 (CP reported as median, IQR). In every position a signifi cant deviation in CP was observed. In total,
40.6% of values exceeded the upper limit of 30 cmH2O. No values
beneath the lower target limit of 20 cmH2O were observed. In each
position the upper target limit was exceeded at least once. Conclusion Simple changes in patient positioning may result in
potentially harmful CP (>30 cmH2O). These observations call for strict
CP monitoring. 2
Results A total of 192 measurements were performed (12 subjects×16
positions). Results are shown in Table 1 (CP reported as median, IQR). In every position a signifi cant deviation in CP was observed. In total,
40.6% of values exceeded the upper limit of 30 cmH2O. No values
beneath the lower target limit of 20 cmH2O were observed. In each
position the upper target limit was exceeded at least once. Introduction The most common cause of ventilator-associated
pneumonia (VAP) is aspiration of oral secretion through the
endotracheal tube (ET). Subglottic suction drainage (SSD) has been
recommended as a safety measure against aspiration due to its
high eff ectiveness. Currently, two types of cuff shape – spindle and
tapered – are predominant in high-volume, low-pressure (HVLP) ETs Conclusion Simple changes in patient positioning may result in
potentially harmful CP (>30 cmH2O). These observations call for strict
CP monitoring. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S58 Table 1 (abstract P154). P155
Towards safer airway management in the critically ill: lessons from
National Audit Project 4 Introduction National Audit Project 4 (NAP4) highlighted the need to
improve airway management in ICUs and key recommendations were
the continuous use of end-tidal carbon dioxide (ETCO2) monitoring,
pre-intubation checklists and diffi cult airway trolleys [1]. This complete
cycle audit aimed to quantify the current state of airway management
on our ICU and the eff ectiveness of implementing the NAP4
recommendations. Results The studied ETTs remained in place for a median of 7 days
(IQR range 4 to 15). The amount of secretions assessed by CT scan was
0.293 ± 0.290 ml (range 0.032 to 0.777 ml). Secretion volumes were
not related to patient severity at admission (SAPS 2, P/F ratio) or days
of intubation; an inverse correlation with patient’s age was present
(P = 0.032, R2 = 0.46). Bacterial growth was present in 9/11 (82%)
ETT fl uids cultures and Candida spp. showed an elevated prevalence
(6/11, 55%). In the bench tests, the cleaning device reduced resistance
to airfl ow (diff erence before and after cleaning 5.5 (95% CI = 8.9 to
1.6) cmH2O/l/second, P = 0.006). After cleaning, resistance resulted
higher than intact ETTs, although with a clinically negligible diff erence
(diff erence 0.3 (95% CI = 0.2 to 0.6 cmH2O/l/second), P = 0.032). Methods Data collection was carried out prospectively for both
phases and included documentation of intubation, use of ETCO2 and
the incidence of serious adverse events (SAEs). The contents of the
intubation boxes were compared against the Diffi cult Airway Society
(DAS) guidelines [2]. The re-audit was carried out 6 months following
the introduction of a pre-intubation checklist, a documentation sticker,
a diffi cult airway trolley and standardization of the basic bedside
airway boxes with a checklist of contents. A training program in airway
management for all ICU staff was also introduced. f
2
Conclusion Micro-CT scan is a feasible and promising technique to
assess secretions volume in ETTs after extubation. The use of an ETT
cleaning device decreases resistance to airfl ow in bench tests; the
eff ectiveness of such a device in the clinical setting could be properly
assessed by post-extubation CT scan. gf
Results The baseline characteristics of both groups were similar. The
initial audit included 45 patients and the re-audit 58 patients. In the initial
audit 19% of patients had accurate documentation of intubation with
the use of ETCO2. The continuous use of ETCO2 was 80%. P153
Endotracheal tube with tapered-type cuff for preventing
ventilator-associated pneumonia: a randomized clinical trial
N Saito, T Yagi, Y Hara, H Matsumoto, K Mashiko
Nippon Medical School Chiba Hokusoh Hospital, Inzai, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P153 (doi: 10.1186/cc12091) Endotracheal tube cuff pressure values following
patient position changes
Cuff
P for Δ from
Measurements
pressure
start position
>30 cmH2O
(cmH2O)a
(25 cmH2O)
(n (%))
Antefl exion head
33.5 (27.5 to 42.0)
0.005
9 (75)
Hyperextension head
30.5 (27.5 to 42.0)
0.004
6 (50)
Lateral fl exion head, left
26.5 (25.3 to 28.8)
0.025
1 (8)
Lateral fl exion head, right
27.0 (25.3 to 29.8)
0.049
2 (17)
Rotation head, left
27.0 (26.3 to 33.8)
0.003
3 (25)
Rotation head, right
28.0 (26.3 to 38.8)
0.020
4 (33)
Semirecumbent, 45° HoB
28.0 (25.3 to 31.5)
0.009
2 (17)
Recumbent, 10° HoB
28.0 (25.3 to 31.5)
0.016
3 (25)
Horizontal back rest
28.5 (25.3 to 31.5)
0.011
3 (25)
Trendelenburg, 10°
29.0 (25.3 to 32.0)
0.008
5 (42)
Left lateral position, 30°
30.5 (25.3 to 32.0)
0.010
6 (50)
Left lateral position, 45°
30.5 (27.3 to 32.8)
0.005
6 (50)
Left lateral position, 90°
34.0 (27.0 to 36.8)
0.006
7 (58)
Right lateral position, 30°
31.0 (27.0 to 34.5)
0.003
6 (50)
Right lateral position, 45°
32.5 (28.3 to 35.8)
0.002
8 (50)
Right lateral position, 90°
31.0 (29.3 to 35.0)
0.003
7 (58)
Total (192 measurements)
–
–
78 (40.6)
HoB, head of bed elevation. aCuff pressure values reported as median (fi rst to
third quartile). Table 1 (abstract P154). Endotracheal tube cuff pressure values following
patient position changes This audit demonstrated that simple measures in conjunction with
a structured training program can help to improve the safety of
airway management in critically ill patients. Further work is required
to improve compliance with the pre-intubation checklist alongside a
continued education program. References 1. The Royal College of Anaesthetists [www.rcoa.ac.uk/nap4]
2. Diffi cult Airway Society Equipment List
[http://www.das.uk.com/equipmentlistjuly2005.htm] P156
Quantitative assessment of endotracheal tube (ETT) biofi lm by
micro-CT scan: evaluation of the eff ectiveness of ETT cleaning
devices
A Coppadoro1, G Bellani1, T Mauri1, R Borsa1, V Meroni1, A Barletta1,
M Teggia Droghi1, A Lucchini2, R Marcolin2, N Patroniti1, S Bramati2,
A Pesenti1
1University of Milan-Bicocca, Monza, Italy; 2San Gerardo Hospital, Monza, Italy
Critical Care 2013, 17(Suppl 2):P156 (doi: 10.1186/cc12094) Introduction The goal of this study was to assess the amount of
biofi lm within the endotracheal tube (ETT) by CT scan and to evaluate
in a bench test a device aimed at ETT biofi lm removal. P157 P157
Early identifi cation of patients at risk of diffi cult intubation in
the ICU: development and validation of the MACOCHA score in a
multicenter cohort study
A De Jong1, N Molinari1, N Terzi2, N Mongardon3, B Jung1, S Jaber1
1Montpellier University Hospital, Montpellier, France; 2Caen University
Hospital, Caen, France; 3Cochin University Hospital, Paris, France
Critical Care 2013, 17(Suppl 2):P157 (doi: 10.1186/cc12095) P153
Endotracheal tube with tapered-type cuff for preventing
ventilator-associated pneumonia: a randomized clinical trial
N Saito, T Yagi, Y Hara, H Matsumoto, K Mashiko
Nippon Medical School Chiba Hokusoh Hospital, Inzai, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P153 (doi: 10.1186/cc12091) Biofi lm grows
within the ETT soon after intubation, resulting in increased resistance
to airfl ow. Biofi lm detachment from the ETT can result in bacterial
colonization of the distal airways. The use of an ETT cleaning device
could eff ectively reduce the patient’s airfl ow resistance burden. The
quantifi cation and distribution of biofi lm within the ETT might help to
evaluate ETT cleaning device eff ectiveness. gf
Methods We collected ETTs of 11 critically ill patients after extubation. Micro-CT scan (SkyScan 1176; Bruker, Belgium) was performed using
a resolution of 35 μm. Axial sections of the 20 cm above the cuff were
reconstructed, and the volume of secretions was assessed by a density
criterion. Microbiological cultures of the ETT lavage fl uid were then
obtained. Patient’s demographics and clinical data were recorded. In
a diff erent set of bench experiments, we injected 1 ml water-based
polymer into new ETTs of diff erent sizes. We measured resistance to
airfl ow before and after using an ETT cleaning device (Airway Medix
Closed Suction System; Biovo Technologies, Tel Aviv, Israel). We also
obtained resistance values of intact ETTs as controls. HoB, head of bed elevation. aCuff pressure values reported as median (fi rst to
third quartile). P155
Towards safer airway management in the critically ill: lessons from
National Audit Project 4 Bedside airway
boxes did not contain a checklist of standardized equipment. A diffi cult
airway trolley was not located on the ICU. Following introduction of the
changes described, a signifi cant improvement in the use of ETCO2 to
confi rm intubation (46% vs. 19%, P <0.01) and continuous use of ETCO2
monitoring (100% vs. 80% P ≤0.0004) was observed. The use of the pre-
intubation checklist (7%) and stickers (11%) was poor. All bedside boxes
had a checklist and the required equipment. A diffi cult airway trolley is
located on the ICU with an equipment list and DAS algorithm attached. No SAEs were recorded in either phase. Early identifi cation of patients at risk of diffi cult intubation in
the ICU: development and validation of the MACOCHA score in a
multicenter cohort study A De Jong1, N Molinari1, N Terzi2, N Mongardon3, B Jung1, S Jaber1
1Montpellier University Hospital, Montpellier, France; 2Caen University
Hospital, Caen, France; 3Cochin University Hospital, Paris, France
Critical Care 2013, 17(Suppl 2):P157 (doi: 10.1186/cc12095) A De Jong1, N Molinari1, N Terzi2, N Mongardon3, B Jung1, S Jaber1
1Montpellier University Hospital, Montpellier, France; 2Caen University
Hospital, Caen, France; 3Cochin University Hospital, Paris, France
Critical Care 2013, 17(Suppl 2):P157 (doi: 10.1186/cc12095) Introduction Diffi cult intubation (DI) in the ICU is a challenging
issue, associated with severe life-threatening complications [1,2]. The
objective was to develop and validate a simplifi ed score for identifying
patients with DI in the ICU and to report related complications. Introduction Diffi cult intubation (DI) in the ICU is a challenging
issue, associated with severe life-threatening complications [1,2]. The
objective was to develop and validate a simplifi ed score for identifying
patients with DI in the ICU and to report related complications. Conclusion Serious airway complications are rare. Safer airway
manage ment is essential in the ICU to prevent morbidity and mortality. S59 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods Data collected in a prospective multicenter-study from 1,000
consecutive intubations from 42 ICUs were used to develop a simplifi ed
score of DI, which was then validated externally in 400 consecutive
intubation procedures from 18 other ICUs and internally by bootstrap
on 1,000 iterations. common indications of intubation were acute respiratory failure
(61.0%) and coma (17.1%). The primary outcome occurred in 138 of
246 intubations (56.1%). Within the fi rst hour, the incidence of severe
hypotension was 47.2%, of severe hypoxemia was 20.3%, of cardiac
arrest was 5.7% and of death was 4.5%. Results In multivariate analysis, the main predictors of DI (incidence =
11.3%) were related to the patient (Mallampati score III or IV,
obstructive sleep apnea syndrome, reduced mobility of cervical spine,
limited mouth opening), to pathology (severe hypoxia, coma) and to
the operator (non-anesthesiologist). From the β-parameter, a seven-
item simplifi ed score (MACOCHA score; Table 1) was built, with an area
under the curve (AUC) of 0.89 (95% CI = 0.85 to 0.94). Tracheal intubation for a diffi cult airway using Airway scope®,
KingVision® and McGRATH®: a comparative manikin study of
inexperienced personnel J Itai1, Y Tanabe2, T Nishida1, T Inagawa1, Y Torikoshi1, Y Kida1, T Tamura1,
K Ota1, T Otani1, T Sadamori1, K Une1, R Tsumura1, Y Iwasaki1, N Hirohashi1,
K Tanigawa1 J Itai1, Y Tanabe2, T Nishida1, T Inagawa1, Y Torikoshi1, Y Kida1, T Tamura1,
K Ota1, T Otani1, T Sadamori1, K Une1, R Tsumura1, Y Iwasaki1, N Hirohashi1,
K Tanigawa1 g
1Hiroshima University Hospital, Hiroshima-shi, Japan; 2Hiroshima City
Hospital, Hiroshima-shi, Japan
Critical Care 2013, 17(Suppl 2):P159 (doi: 10.1186/cc12097) g
1Hiroshima University Hospital, Hiroshima-shi, Japan; 2Hiroshima City
Hospital, Hiroshima-shi, Japan
Critical Care 2013, 17(Suppl 2):P159 (doi: 10.1186/cc12097) g
1Hiroshima University Hospital, Hiroshima-shi, Japan; 2Hiroshima City
Hospital, Hiroshima-shi, Japan
Critical Care 2013, 17(Suppl 2):P159 (doi: 10.1186/cc12097) Table 1 (abstract P157). MACOCHA score calculation worksheet
Points
Factors related to patient
Mallampati score III or IV
5
Obstructive apnea syndrome
2
Reduced mobility of cervical spine
1
Limited mouth opening <3 cm
1
Factors related to pathology
Coma
1
Severe hypoxemia (<80%)
1
Factor related to operator
Non-anaesthesiologist
1
Total
12
Conclusion DI in ICU is strongly associated with severe life-threatening
complications. A simple score including seven clinical items
discriminates diffi cult and non-DI in ICU. References
1. Jaber et al.: Crit Care Med 2006, 34:2355-2361. 2. Jaber et al.: Intensive Care Med 2010, 36:248-255. P158
Incidence of major complications related to endotracheal
intubation in ICUs of nonacademic Brazilian public hospitals
AB Cavalcanti1, K Normilio-da-Silva1, JC Acarine Mouro1, F Moreira1,
AA Kodama1, R Del-Manto2, O Berwanger1
1Hospital do Coração – HCor, São Paulo, Brazil; 2Hospital Militar de Área de
São Paulo – HMASP, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P158 (doi: 10.1186/cc12096)
d
b
d
h
d
f Table 1 (abstract P157). MACOCHA score calculation worksheet
Points
Factors related to patient
Mallampati score III or IV
5
Obstructive apnea syndrome
2
Reduced mobility of cervical spine
1
Limited mouth opening <3 cm
1
Factors related to pathology
Coma
1
Severe hypoxemia (<80%)
1
Factor related to operator
Non-anaesthesiologist
1
Total
12 Table 1 (abstract P157). MACOCHA score calculation worksheet Table 1 (abstract P157). MACOCHA score calculation worksheet Introduction The Airway scope® (AWS), the KingVision® (KV) and the
McGRATH® (MG) are new indirect video laryngoscopes designed
to facilitate eff ective and safe tracheal intubation under various
conditions. However, there are few comparative studies as for
performance in tracheal intubation attempted by inexperienced
personnel. Early identifi cation of patients at risk of diffi cult intubation in
the ICU: development and validation of the MACOCHA score in a
multicenter cohort study In the validation
cohort (prevalence of DI = 8%), the AUC was of 0.86 (95% CI = 0.76
to 0.96), with a sensitivity of 73%, a specifi city of 89%, a negative
predictive value of 98% and a positive predictive value of 36%. After
internal validation by bootstrap, the AUC was 0.89 (95% CI = 0.86 to
0.93). Severe life-threatening events (severe hypoxia, collapse, cardiac
arrest or death) occurred in 38% of the 1,000 cases. Patients with DI
(n = 113) had signifi cantly higher severe life-threatening complications
than those who had a non-DI (51% vs. 36%, P <0.0001). Conclusion Incidence of major complications associated with
endotracheal intubation is unacceptably high in Brazilian public ICUs. There is an urgent need for eff ective measures to increase safety of this
common procedure in our environment. Tracheal intubation for a diffi cult airway using Airway scope®,
KingVision® and McGRATH®: a comparative manikin study of
inexperienced personnel The purpose of this study was to evaluate success rates, time
to intubation with use of these devices by inexperienced personnel in a
simulated manikin diffi cult airway.i Methods Twenty-nine fi fth-year medical students with no previous
experience in tracheal intubation participated in this study. We used
an advanced patient simulator (SimMan®; Laerdal Medical, Stavanger,
Norway) to simulate normal and diffi cult airway scenarios including
cervical spine rigidity, swollen tongue, and pharyngeal edema. The
sequences in selecting devices and scenarios were randomized. Success
rate for tracheal intubation, and the time required for visualization
of the glottis (T1), tracheal intubation (T2), and infl ation of the lungs
(T3) were analyzed. Also, numbers of audible dental click during the
intubation attempt were recorded. The three diff erent intubation
devices were tested in four diff erent scenarios by 29 students. Results All three devices had very high success rates of tracheal
intubation (AWS 100%; KV 100%; MG 99%). In the normal airway, T1,
T2 and T3 were AWS: 4.4 ± 3.3 seconds, 7.5 ± 3.8 seconds, 11.1 ± 3.8
seconds; KV: 6.9 ± 6.9 seconds, 10.3 ± 7.9 seconds, 14.1 ± 8.3 seconds;
MG: 4.6 ± 1.3 seconds, 11.4 ± 5.2 seconds, 16.1 ± 5.4 seconds,
respectively (NS). In the three diffi cult airway scenarios, T1, T2 and T3
were AWS: 5.3 ± 4.7* seconds, 10.4 ± 6.9* seconds, 14.2 ± 6.9* seconds;
KV: 10.9 ± 12.8 seconds, 21.5 ± 18.1 seconds, 25.4 ± 18.3 seconds;
MG: 13.0 ± 15.3 seconds, 26.2 ± 24.6 seconds, 31.0 ± 24.3 seconds,
respectively (*P <0.05 AWS vs. KV and MG). The number of audible
dental click sounds with the MG was greater than with the AWS and KV
(AWS 5%*; KV 8%*; MG 28%*; *P <0.05 AWS and KV vs. MG). 1. Jaber et al.: Crit Care Med 2006, 34:2355-2361. 2. Jaber et al.: Intensive Care Med 2010, 36:248-255. P159 P159
Tracheal intubation for a diffi cult airway using Airway scope®,
KingVision® and McGRATH®: a comparative manikin study of
inexperienced personnel
J Itai1, Y Tanabe2, T Nishida1, T Inagawa1, Y Torikoshi1, Y Kida1, T Tamura1,
K Ota1, T Otani1, T Sadamori1, K Une1, R Tsumura1, Y Iwasaki1, N Hirohashi1,
K Tanigawa1
1Hiroshima University Hospital, Hiroshima-shi, Japan; 2Hiroshima City
Hospital, Hiroshima-shi, Japan
Critical Care 2013, 17(Suppl 2):P159 (doi: 10.1186/cc12097) Incidence of major complications related to endotracheal
intubation in ICUs of nonacademic Brazilian public hospitals Conclusion The AWS, KV and MG had very high success rates of tracheal
intubation and are suitable as intubation devices for inexperienced
personnel. In the diffi cult airway, however, the intubation time with
AWS was signifi cantly shorter than with KV and MG. These fi ndings
suggested that AWS may be most useful device particularly in diffi cult
situations such as emergency settings. Further studies in a clinical
setting are needed to confi rm these fi ndings. Introduction Our objective is to determine the incidence of major
complications related to endotracheal intubations (death, cardiac
arrest, severe hemodynamic instability or severe hypoxemia) per-
formed in the ICUs of nonacademic Brazilian public hospitals. Methods This is a report of the baseline phase of a cluster randomized
trial with two parallel arms. In this baseline phase, we collected data
from sequential patients needing endotracheal intubation in 17 ICUs
from Brazil. Patients needing endotracheal intubation after cardiac
arrest were excluded. In a second ongoing phase, not reported
here, we have randomized ICUs to a multifaceted intervention to
prevent complications of endotracheal intubation. Primary outcome
was defi ned as the occurrence of death, cardiac arrest, severe
cardiovascular instability (systolic blood pressure <60 mmHg at least
one time, <90 mmHg and persisting >30 minutes or need to initiate
vasopressors) or severe hypoxemia (SpO2 <80%) within 1 hour after
intubation. P160 NT-proBNP and cardiac cycle effi ciency changes during extubation
process in critically ill patients i
Results One hundred patients were surveyed. The ET and LT groups
included 49 and 51 patients, respectively. Tracheostomy was performed
using a percutaneous procedure in 48 patients (ET: 25, LT: 23) and a
surgical procedure in 52 patients (ET: 24, LT: 28). Sixty-two patients (ET:
34, LT: 28) survived to discharge and 16 patients died in the ICU (ET:
7, LT: 9). Fifty-six patients (ET: 31, LT: 25) were weaned from ventilator
support and tracheostomy cannula was removed in 20 patients (ET:
11, LT: 9). There were no signifi cant diff erences in type of tracheostomy
procedure, period from tracheostomy until ICU and hospital discharge,
rate of patients who could be weaned from ventilator and removed
tracheostomy cannula, and ICU and hospital mortality between the
groups. The length of mechanical ventilation and the time to removal
of tracheostomy cannula were signifi cantly shorter in the ET group
(5 ± 7 vs. 26 ± 41 and 29 ± 24 vs. 94 ± 83 days, respectively). Introduction Several factors can lead to weaning failure from
mechanical ventilation (MV). Among these, cardiac dysfunction is one
of the main causes. NT-proBNP has been proposed as a biomarker of
cardiovascular function during weaning from MV. Unfortunately it does
not provide for a continuous monitoring of cardiac function. Pulse
wave analysis may serve as a continuous bedside monitoring tool of
cardiovascular performance. Cardiac cycle effi ciency (CCE) is an indirect
index of left ventricular performance obtained by the pulse contour
method MostCare (Vygon, Padova, Italy). The aim of this study was to
evaluate the correlation between NT-proBNP and CCE and the potential
usefulness of such variables during the weaning process from MV. Conclusion In this retrospective study, early tracheostomy reduced
the length of weaning after tracheostomy and the time to removal of
tracheostomy cannula, while there were no diff erences in the length of
ICU stay and patient outcome. In critically ill adult patients who require
mechanical ventilation, a tracheostomy performed at an earlier stage
may shorten the duration of artifi cial ventilation. A further randomized
clinical trial is essential to determine the eff ectiveness and safety of
early tracheostomy. Methods Twenty-two long-term (>48 hours) mechanically ventilated
patients capable of performing a weaning trial of spontaneous
breathing (SBT) were enrolled in the study. Inclusion criteria were:
age >18 years and equipment with a standard arterial catheter line. Resistance of neonatal endotracheal tubes: a comparison of four
commercially available types Introduction In mechanically ventilated neonates the fl ow-dependent
resistance of the endotracheal tube (ETT) causes a noticeable pressure
diff erence between airway and tracheal pressure [1]. This may
potentially lead to retardation of the passive driven expiration and
dynamic lung infl ation consecutively but more importantly increases Introduction In mechanically ventilated neonates the fl ow-dependent
resistance of the endotracheal tube (ETT) causes a noticeable pressure
diff erence between airway and tracheal pressure [1]. This may
potentially lead to retardation of the passive driven expiration and
dynamic lung infl ation consecutively but more importantly increases Results We evaluated 246 intubations. The mean age was
49 ± 21.9 years; the SAPS 3 admission score was 57.4 ± 15.4. The most S60 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 after extubation (T2, T3) compared with T1; conversely, CCE showed
an inverse trend. Overall, a negative correlation was found between
NT-proBNP and CCE values (R = –0.81, P <0.001). Signifi cant inverse
correlations were found between NT-proBNP and CCE at T1, T2, and
T3 (R = –0.91, –0.75 and –0.73 respectively; P < 0.001). The overall
correlation between NT-proBNP and CCE was –0.74 in the SB group and
–0.86 in the CPAP group. Standard hemodynamic and ventilatory data
did not show signifi cant changes during the study. the work of breathing. The aim of this study was to compare the
resistive pressure drop of four commercially available neonatal ETTs
with internal diameter (ID) of 2.0 mm. the work of breathing. The aim of this study was to compare the
resistive pressure drop of four commercially available neonatal ETTs
with internal diameter (ID) of 2.0 mm. Methods The pressure–fl ow relationship of neonatal ETTs (internal
diameter 2.0 mm) of four diff erent manufacturers (Mallinckrodt, Hi-
Contour, Covidien, Dublin, Ireland; Portex, Cole’s Neonatal Tube, Smith
Medical, St Paul, MN, USA; Rüsch, Silko Clear, Telefl ex Medical, Kernen,
Germany; and Vygon, Pediatric Endotracheal Tube, Ecouen, France)
was determined in a physical model consisting of a tube connector,
an anatomically curved ETT and an artifi cial trachea. The model was
ventilated with a sinusoidal gas fl ow with an amplitude of ±100 ml/
second and a ventilation rate ranging from 40 to 60 cycles/minute. The
coeffi cients of an approximation equation considering ETT resistance
were fi tted separately to the measured pressure–fl ow curves for
inspiration and expiration. Comparison of outcomes between early and late tracheostomy for
critically ill patients Guttmann J, et al.: Crit Care Med 2000, 28:1018-1026. Comparison of outcomes between early and late tracheostomy for
critically ill patients Results The pressure drop profi les of all ETTs were nonlinearly fl ow
dependent. The expiratory pressure drop (Pexp) slightly exceeded
the inspiratory one (Pinsp). The ETT of Portex Cole’s Neonatal Tube
had a signifi cant lower pressure drop (Pinsp 8.8 ± 0.3 cmH2O vs. Pexp
–11.2 ± 0.2 cmH2O at an air fl ow of 100 ml/second) compared with all
other ETTs (Mallinckrodt: Pinsp 19.2 ± 0.4 cmH2O vs. Pexp –24.3 ± 0.7 cmH2O;
Rüsch: Pinsp 28.3 ± 0.8 cmH2O vs. Pexp –33.0 ± 0.8 cmH2O; Vygon: Pinsp
19.2 ± 0.4 cmH2O vs. Pexp –22.6 ± 0.6 cmH2O, all P <0.05) for all fl ow and
ventilation rates, respectively. K Suzuki1, S Kusunoki1, T Yamanoue1, K Tanigawa2
1Hiroshima Prefectural Hospital, Hiroshima, Japan; 2Hiroshima University
Hospital, Hiroshima, Japan
Critical Care 2013, 17(Suppl 2):P162 (doi: 10.1186/cc12100) Introduction Tracheostomy is one of the more commonly performed
procedures in critically ill patients requiring long-term mechanical
ventilation. However, the optimal timing or method of performing
tracheostomies in this population remains to be established. In the
present study, we compared outcomes of early and late tracheostomy
in critically adult patients with diff erent clinical conditions. y
Conclusion The ETT resistance highly contributes to the total airway
resistance in neonatal ventilation. The fl ow-dependent pressure drop
of shouldered Cole’s Neonatal Tube (Portex) was up to 70% less in
inspiration (resp. 67% in expiration) compared with straight tubes
with an internal diameter corresponding to the narrow part of Cole’s
tube. We conclude that neonatal intubation with a Cole’s tube can
clearly reduce the resistive load due to the endotracheal tube and thus
potentially prevent additionally work of breathing. Reference yf
Methods All patients needing tracheostomy in the Critical Care
Medical Center of Hiroshima Prefectural Hospital from January 2009
to December 2011 were surveyed. Patients with tracheostomy who
were not indicated for mechanical ventilation were excluded from the
subjects. Early tracheostomy (ET) was defi ned as <10 days after tracheal
intubation and late tracheostomy (LT) was defi ned as ≥10 days after
intubation. We compared patient characteristics, type of tracheostomy
procedure, length of weaning from ventilator and outcomes between
the groups. Data are shown as the mean ± SD, with unpaired t test
and Mann–Whitney U test used for statistical analyses. Statistical
signifi cance was accepted at P <0.05. 1. Guttmann J, et al.: Crit Care Med 2000, 28:1018-1026. 1. NT-proBNP and cardiac cycle effi ciency changes during extubation
process in critically ill patients Exclusion criteria were: neuromuscular disease, tracheotomy, renal
failure, and traumatic brain injury. During the weaning process, NT-
proBNP plasma levels, CCE, and standard hemodynamic and ventilatory
data were collected 30 minutes before extubation (T1), 2 hours (T2)
and 12 hours later (T3). After removal of tracheal tube, patients with
a history of heart failure received continuous positive airway pressure
(CPAP group). Patients with normal cardiac function were maintained
with spontaneous breathing (SB group). Reference . Arabi Y, et al.: Crit Care 2004, 8:R347-R352. Arabi Y, et al.: Crit Care 2004, 8:R347-R352. Arabi Y, et al.: Crit Care 2004, 8:R347-R352. Resistance of neonatal endotracheal tubes: a comparison of four
commercially available types gi
g
g
y
Conclusion NT-proBNP correlated well with CCE. The latter seems to be
an additional attractive index of cardiovascular state that, in association
with NT-proBNP changes, may provide information about cardiac
function on a beat-by-beat basis during weaning process from MV. P162 Ultrasound scanning for percutaneous dilatational tracheostomy:
a systematic review y
R Pugh, A Slater y
R Pugh, A Slater y
R Pugh, A Slater R Pugh, A Slater
Glan Clwyd Hospital, Bodelwyddan, UK
Critical Care 2013, 17(Suppl 2):P163 (doi: 10.1186/cc12101) Results Sixty-six paired NT-proBNP and CCE values were obtained. Patients in the SB group and in the CPAP group were 10 and 12,
respectively. In both groups there was a trend towards an increase in
NT-proBNP values after extubation, an opposite trend was observed
regarding CCE values (P <0.05). NT-proBNP levels showed an increase Glan Clwyd Hospital, Bodelwyddan, UK
Critical Care 2013, 17(Suppl 2):P163 (doi: 10.1186/cc12101) Introduction Percutaneous dilatational tracheostomy (PDT) remains
a frequently performed procedure in the ICU. However, there is great S61 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P164). Oxygenation time to oxygen alveolar pressure
>100 mmHg. Figure 2 (abstract P164). Mean carbon dioxide alveolar pressure at the
start and at the end of the test. variability in the course of blood vessels in the pre-tracheal area. A 5%
risk of clinically relevant bleeding was recently reported for patients
undergoing PDT [1]. We conducted a systematic review of reports
evaluating clinical outcomes following use of ultrasound scanning (US)
for PDT. variability in the course of blood vessels in the pre-tracheal area. A 5%
risk of clinically relevant bleeding was recently reported for patients
undergoing PDT [1]. We conducted a systematic review of reports
evaluating clinical outcomes following use of ultrasound scanning (US)
for PDT. Figure 1 (abstract P164). Oxygenation time to oxygen alveolar pressure
>100 mmHg. Methods Two investigators performed a search of the literature using
the following databases: CENTRAL, Embase, MEDLINE and SCOPUS. The following eligibility criteria were used: population including adults
>16 years managed in the ICU; use of ultrasound to guide decision-
making pre-PDT or guide PDT performance; report of clinically relevant
outcome measures. Nonrandomised controlled trials were classifi ed
according to Cochrane Non-Randomised Study Methods Group criteria
[2] and evaluated for risk of bias. [ ]
Results An initial search identifi ed 2,043 reports, of which 10 studies
met eligibility criteria: eight case series, one randomised controlled trial
(RCT) and one prospective cohort study, incorporating 488 patients. Two studies specifi cally reported data on patients with obesity (n = 29
patients) and one study reported data for a group of patients with
spinal cord fi xation (n = 6). Reference 1. Cook TM, Nolan JP, Magee PT, Cranshaw JH: Needle cricothyroidotomy. Anaesthesia 2007, 62:289-291. P Persona, P Diana, A Ballin, F Baratto, M Micaglio, C Ori g
Clinica di Anestesia e Medicina Intensiva, Padova, Italy y
Critical Care 2013, 17(Suppl 2):P164 (doi: 10.1186/cc12102) Introduction Failed airway situations are potentially catastrophic
events and require a correct approach with appropriate tools. Recently,
Ventrain has been presented as a manual device for emergency
ventilation through a small-bore cannula, which can provide expiratory
assistance by applying the Venturi eff ect. y
R Pugh, A Slater US was used to guide decision to perform
PDT or surgical tracheostomy in fi ve studies, with decision to perform
surgical tracheostomy ranging from 0 to 27% of cases. US was used
to guide insertion point in seven studies, and used real-time in four
studies. Times to perform US-guided PDT were reported in four studies
(ranging from 8 to 12 minutes). No studies compared time taken with
or without US. Data on complications of procedure were reported
in nine studies. Minor bleeding was reported for eight cases (1.6%
overall). Prolonged bleeding was reported in two cases (0.4%). There
were no episodes of catastrophic bleeding among 488 cases. High risk
of bias was identifi ed in fi ve studies in terms of patient selection. An
intervention protocol was not defi ned in three reports. No attempt was
made at blinding any aspect of the 10 studies. Figure 2 (abstract P164). Mean carbon dioxide alveolar pressure at the
start and at the end of the test. g
y
Conclusion Use of US guidance could theoretically help minimise risk
of haemorrhagic complications during PDT and perhaps reduce time
taken to perform PDT. However, there is currently inadequate evidence
from controlled cohort studies or RCTs to suggest that routine use
for PDT in selected or unselected groups improves clinically relevant
outcome measure. References 1. Delaney et al.: Crit Care 2006, 10:R55. 2. Higgins JPT, Green S (Eds): Cochrane Handbook for Systematic Reviews of
Interventions. Version 5.1.0. The Cochrane Collaboration; 2011. [www.cochrane-handbook.org] 2. Higgins JPT, Green S (Eds): Cochrane Handbook for Systematic Reviews of
Interventions. Version 5.1.0. The Cochrane Collaboration; 2011. [www.cochrane-handbook.org] vs. Melker –0.3; vs. Quicktrach –5.9) (Figure 2) and moreover the users
judged it more favorably. Conclusion In this manikin study, Ventrain seemed to be able to
appropriately oxygenate and ventilate a patient in a CICV situation. When compared with the best available choices, it has shown not to
be inferior. R f Evaluation of a new device for emergency transcricoid ventilation in
a manikin model Reference g
CW Carspecken, D Talmor This may represent a paucity of evidence
on the subject, suggesting that further clinical trials on the topic are
needed to contribute to the evidence base. This also highlights the
need for international consensus on the topic, to reduce duplication of
eff orts, standardise practice, and improve outcomes. Reference
1. Appraisal of Guidelines for Research and Evaluation: Instrument. AGREE II
[http://www.agreetrust.org/]
ble 1 (abstract P166). Summary of results returned following systematic review
Total
Results with
Full texts
Guidelines
AGREE instrument
atabase
results
limits
retrieved
found
compliant
bMed
4,685
1,596
33
EDLINE
3,859
1,337
17
26
2
NAHL
711
172
6
edical societies/guideline websites/clearinghouses
N/A
N/A
N/A
54
3
oogle
17,600,000
497
N/A Introduction Protocol-based care of the tracheostomised patient is
important, as adverse events confer a high rate of mortality. Little is
known regarding the existence of formal evidence-based guidelines on
tracheostomy care. The aim of this study was to perform a systematic
review for evidence-based guidelines on adult tracheostomy care. (<$11) electronics were designed such that no power supply would be
needed apart from a computer or mobile device. Collection of airfl ow
signal and calibration was performed with a standard 3 l syringe with
fl ow volume measurement on one adult subject. Results Calibration of the assembled device over range of 0.5 to 3 l/
second enabled conversion of pressure diff erence to air fl ow with a
mean measured tube resistance of 0.258 (kPa*second)/l. Robust signal
responses to an adult subject’s continuous respiratory maneuvers on
the tube itself were demonstrated. Subject performance of the pMDI
technique with subsequent tidal volume breathing was recorded and
analyzed (Figure 1). Methods A systematic search of PubMed, MEDLINE, guideline
clearinghouses, centres of evidence-based practice, and professional
societies’ guidelines relating to care of adult patients with a
tracheostomy was performed by two reviewers. In addition, a
Google search of publicly available tracheostomy care guidelines
was performed. Search terms: (tracheostom* OR tracheotom*) AND
(protocol* OR guideline* OR standard* OR management OR consensus
OR algorithm*). Filters: English language, human, from 1 January
1990 to date, adult patients. Guideline appraisal criteria: the quality
of guidelines retrieved was assessed using the Appraisal of Guidelines
Research and Evaluation II (AGREE II) instrument [1]. y
Conclusion Design and calibration of a novel low-cost monitoring
device for bronchodilator delivery monitoring enabled detection
and recording of characteristic fl ow volume respiratory patterns for
point-of-care diagnostics in resource-limited settings. g
CW Carspecken, D Talmor Future work will
require clinical testing and automated detection of the correct pMDI
patient technique. Results The search results are summarised in Table 1. A total of 80
guidelines were identifi ed. Five were found to satisfy the AGREE II
criteria and only three related to the entire spectrum of tracheostomy
management. The majority was informal and was not published or
evidence based. p
References References
1. Ari A, et al.: Respir Care 2012, 57:613-626. 2. Riviello E, et al.: Crit Care Med 2011, 39:860. 2. Riviello E, et al.: Crit Care Med 2011, 39:860. Conclusion Five evidence-based guidelines on adult tracheostomy
management were identifi ed. This may represent a paucity of evidence
on the subject, suggesting that further clinical trials on the topic are
needed to contribute to the evidence base. This also highlights the
need for international consensus on the topic, to reduce duplication of
eff orts, standardise practice, and improve outcomes. g
CW Carspecken, D Talmor Harvard Medical School and Beth Israel Deaconess Medical Center, Boston,
MA, USA y
yf
Methods We used the SimulARTI Human Patient Simulator to evaluate
Ventrain. Initially, we studied the eff ectiveness and security in
ventilating and oxygenating the patient. In a second phase, the Ventrain
performance was compared with what is considered to be the present
gold standard (Quicktrach II, Portex Mini-Trach II Seldinger Kit, Melker
Emergency Cricothyrotomy Catheter Set). Seven anesthesiologists
performed an emergency transcricoid ventilation with each device in
the same setting. Critical Care 2013, 17(Suppl 2):P165 (doi: 10.1186/cc12103) Critical Care 2013, 17(Suppl 2):P165 (doi: 10.1186/cc12103) Introduction Eff ective delivery of aerosolized bronchodilators for
patients with asthma is crucial for adequate therapy in critical care
and emergent settings. Often administered with pressure-metered
dose inhalers (pMDIs), bronchodilator delivery depends on the correct
patient technique during administration [1] and the ability to measure
treatment response, which are diffi cult to monitor at the point of
care and particularly so in resource-poor settings where standard in-
hospital monitoring is unavailable [2].l Results Ventrain provided an average tidal volume of 334 ml and an
average minute volume of 2.4 l in the considered situation, with a
modifi cation of PAO2 from 32 to 702 mmHg and of PACO2 from 54.5
to 38.8 mmHg. In the second phase, the time needed to obtain an
eff ective oxygenation with Ventrain was found to be shorter than
other devices (median diff erence; vs. Minitrach –60 seconds; vs. Melker
–35 seconds; vs. Quicktrach –25 seconds) (Figure 1); the ability to
remove CO2 resulted bigger (average diff erence: vs. Minitrach –11.9; Methods A point-of-care device for airfl ow measurement during
bronchodilator delivery was designed and tested for use in resource-
limited settings. The handheld device was constructed from a clinical
aerosol delivery tube with a bidirectional sensor for pressure diff erential
detection about the aerosol element (Figure 1). The custom low-cost S62 Critical Care 2013, Volume 17 Suppl 2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P165). Figure 1 (abstract P165). Figure 1 (abstract P165). Figure 1 (abstract P165). $11) electronics were designed such that no power supply would be
eeded apart from a computer or mobile device. Collection of airfl ow
gnal and calibration was performed with a standard 3 l syringe with l
ow volume measurement on one adult subject. g
CW Carspecken, D Talmor esults Calibration of the assembled device over range of 0.5 to 3 l/
econd enabled conversion of pressure diff erence to air fl ow with a
ean measured tube resistance of 0.258 (kPa*second)/l. Robust signal
sponses to an adult subject’s continuous respiratory maneuvers on
e tube itself were demonstrated. Subject performance of the pMDI
chnique with subsequent tidal volume breathing was recorded and
nalyzed (Figure 1). onclusion Design and calibration of a novel low-cost monitoring
evice for bronchodilator delivery monitoring enabled detection
nd recording of characteristic fl ow volume respiratory patterns for
oint-of-care diagnostics in resource-limited settings. Future work will
quire clinical testing and automated detection of the correct pMDI
atient technique. eferences
Ari A, et al.: Respir Care 2012, 57:613-626. Riviello E, et al.: Crit Care Med 2011, 39:860. 166
vidence-based guidelines and protocols for the management of
dult patients with a tracheostomy: a systematic review
Siddiqui1, PB Sherren2, MA Birchall3
North West London Hospitals NHS Trust, London, UK; 2Barts and The London
HS Trust, London, UK; 3The Royal National Throat Nose and Ear Hospital,
ndon, UK
itical Care 2013, 17(Suppl 2):P166 (doi: 10.1186/cc12104)
Introduction Protocol-based care of the tracheostomised patient is
important, as adverse events confer a high rate of mortality. Little is
known regarding the existence of formal evidence-based guidelines on
tracheostomy care. The aim of this study was to perform a systematic
review for evidence-based guidelines on adult tracheostomy care. Methods A systematic search of PubMed, MEDLINE, guideline
clearinghouses, centres of evidence-based practice, and professional
societies’ guidelines relating to care of adult patients with a
tracheostomy was performed by two reviewers. In addition, a
Google search of publicly available tracheostomy care guidelines
was performed. Search terms: (tracheostom* OR tracheotom*) AND
(protocol* OR guideline* OR standard* OR management OR consensus
OR algorithm*). Filters: English language, human, from 1 January
1990 to date, adult patients. Guideline appraisal criteria: the quality
of guidelines retrieved was assessed using the Appraisal of Guidelines
Research and Evaluation II (AGREE II) instrument [1]. Results The search results are summarised in Table 1. A total of 80
guidelines were identifi ed. Five were found to satisfy the AGREE II
criteria and only three related to the entire spectrum of tracheostomy
management. The majority was informal and was not published or
evidence based. Conclusion Five evidence-based guidelines on adult tracheostomy
management were identifi ed. Can we keep neck breathers safe? A survey on training adequacy of
medical staff caring for tracheostomy/laryngectomy patients Results PDTs with LMA were successful in 99.3% of the patients (n = 149). The ratings were 1 or 2 in 96% of cases with regards to ventilation and
to blood gas analysis, in 96.6% for identifi cation of relevant structures
and tracheal puncture site, and in 93.3% for the view inside the trachea
during PDT. A rating of 5 was assigned to one patient requiring tracheal
reintubation for inadequate ventilation. There were no damages to the
bronchoscope or reports of gastric aspiration. Can we keep neck breathers safe? A survey on training adequacy of
medical staff caring for tracheostomy/laryngectomy patients
C Tjen, G Rajendran, S Hutchinson
Norfolk and Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P168 (doi: 10.1186/cc12106) C Tjen, G Rajendran, S Hutchinson j
j
Norfolk and Norwich University Hospital, Norwich, UK y
Critical Care 2013, 17(Suppl 2):P168 (doi: 10.1186/cc12106) Introduction A fatal incident related to a blocked tracheostomy
tube prompted a review in our Trust. To provide safe tracheostomy
care, changes in staffi ng, education and operational policies were
recommended. Training of potential fi rst responders to tracheostomy
or laryngectomy emergencies remains outstanding. We aim to quantify
the training defi cit. Tracheostomies are common in critical care but
these patients require ongoing management of an artifi cial airway on
discharge to the ward and even the community. In 2010 our critical care
unit cared for 108 tracheostomy patients, of which 30 were transferred
to the wards. The 4th National Audit Project highlighted complications
including hypoxic brain injury and death [1] and the National Patient
Safety Agency recognised a number of avoidable aspects [2]. Existing
guidelines for management of these patients including emergencies
are not widely known. p
p
g
p
Conclusion The Blue Dolphin PDT using a LMA showed defi nite
advantages regarding inspection of dilation process. This method
improves visualization of the trachea and larynx during a video-assisted
procedure and prevents the diffi culties associated with the use of an
ET such as cuff puncture, tube transection by the needle, accidental
extubation, and bronchoscope lesions. The LMA results as an eff ective
and successful ventilatory device during PDT. This may be especially
relevant in cases of diffi cult patient anatomy where improved structural
visualization optimizes operating conditions. The intensivist performing
PDT should be scrupulous when deciding which method to use. References 1. Cook T, et al.: Br J Anaesth 2011, 106:632-642. 1. Cook T, et al.: Br J Anaesth 2011, 106:632-642. 2. McGrath B, et al.: Postgrad Med J 2010, 86:522-525. Results All 11 ICUs questioned perform and manage tracheostomies. Of
22 respondents, 10 knew of guidelines covering all of the emergencies
described above and their location. Four respondents thought that
these guidelines were accessible in an emergency setting, one-half
of which were on computer systems requiring a login and search
function. With regards to emergency management, 19 respondents felt
competent in a tracheostomy emergency; almost exclusively through
experience and in-house teaching. No respondents were aware of any
formal emergency tracheostomy management courses. 2. McGrath B, et al.: Postgrad Med J 2010, 86:522-525. Laryngeal mask as a safe and eff ective ventilatory device during
Blue Dolphin tracheostomy in the ICU The insertion of the LMA, the quality
of ventilation, the blood gas values, the view of the tracheal puncture
site, and the view of the balloon dilatation were rated as follows: very
good (1), good (2), barely acceptable (3), poor (4), and very poor (5). Results PDTs with LMA were successful in 99.3% of the patients (n = 149). The ratings were 1 or 2 in 96% of cases with regards to ventilation and
to blood gas analysis, in 96.6% for identifi cation of relevant structures
and tracheal puncture site, and in 93.3% for the view inside the trachea
during PDT. A rating of 5 was assigned to one patient requiring tracheal
reintubation for inadequate ventilation. There were no damages to the
bronchoscope or reports of gastric aspiration. 1. Cook TM, et al.: Results of the 4th National Audit Project of the Royal
College of Anaesthetists. Br J Anaesth 2011, 106:632-642. College of Anaesthetists. Br J Anaesth 2011, 106:632-642. 2. McGrath BA, et al.: Multidisciplinary guidelines for the management of
tracheostomy and laryngectomy airway emergencies. Anaesthesia 2012,
67:1025-1041. p
y g
g
tracheostomy and laryngectomy airway emergencies. Anaesthesia 2012,
67:1025-1041. tracheostomy and laryngectomy airway emergencies. Anaesthesia 2012,
67:1025-1041. Can we keep neck breathers safe? A survey on training adequacy of
medical staff caring for tracheostomy/laryngectomy patients In our
ICU the Blue Dolphin PDT with LMA has become the procedure of choice. References Can we keep neck breathers safe? A survey on training adequacy of
medical staff caring for tracheostomy/laryngectomy patients
C Tjen, G Rajendran, S Hutchinson
Norfolk and Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P168 (doi: 10.1186/cc12106) Can we keep neck breathers safe? A survey on training adequacy of
medical staff caring for tracheostomy/laryngectomy patients
C Tjen, G Rajendran, S Hutchinson
Norfolk and Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P168 (doi: 10.1186/cc12106) P167
Survey of emergency tracheostomy management in the East of
England region
N Lawrence, L Oakley, C Swanavelder, M Palmer
West Suff olk Hospital, Bury St Edmunds, UK
Critical Care 2013, 17(Suppl 2):P167 (doi: 10.1186/cc12105) Results We achieved a response rate of 39% (65/168). Respondents
comprised: 33% anaesthesia/critical care, 47% medicine and 14%
surgery. Over one-half (36/65) had managed tracheostomy/laryngec-
tomy emergencies, with 42% (15/36) of these incidents occurring
on the wards and one in an outpatient clinic. Only 20% (13/65) had
received any formal training on management of a blocked/misplaced
tracheostomy tube and only 18% (12/65) were aware of any guidelines. One-third of responders lacked confi dence in management of these
emergencies and 88% felt they would benefi t from formal training
including simulation. Introduction The 4th National Audit Project of the Royal College
of Anaesthetists [1] concluded that the majority of airway-related
signifi cant complications in ICUs resulted from displaced or blocked
tracheostomies and recommended together with the Intensive Care
Society and the National Tracheostomy Safety Project that each ICU
in the UK should have an emergency airway management plan and
guidelines [2]. The aim of this survey was to establish whether such
guidelines exist and are familiar to those working within the ICUs of the
East of England (EoE), their ease of availability in an emergency and the
degree of emergency tracheostomy training within the region. Conclusion The population of patients with exteriorised tracheas is
increasing and represents a high-risk group. Management of airway
emergencies in these patients is not part of standard life-support
courses. According to our trainees, these scenarios are relatively
common and a signifi cant proportion of fi rst responders are poorly
equipped to deal with them. Our Trust will be including specifi c training
on the emergency management of neck breathers as part of in-house
resuscitation training. We would contend that national resuscitation
courses should consider doing the same. f Methods Data collection was via a telephone survey of 11 ICUs in the
EoE training region during July 2012 with one senior ICU nurse and one
ICU trainee questioned per hospital. Questions related to the existence
and accessibility of guidelines for tracheostomy emergencies, and to
the respondent’s degree of emergency tracheostomy training and their
perceived availability of formal training. P166 P166
Evidence-based guidelines and protocols for the management of
adult patients with a tracheostomy: a systematic review 1. Appraisal of Guidelines for Research and Evaluation: Instrument. AGREE II
[http://www.agreetrust.org/] 1. Appraisal of Guidelines for Research and Evaluation: Instrument. AGREE II
[http://www.agreetrust.org/] Table 1 (abstract P166). Summary of results returned following systematic review
Total
Results with
Full texts
Guidelines
AGREE instrument
Database
results
limits
retrieved
found
compliant
PubMed
4,685
1,596
33
MEDLINE
3,859
1,337
17
26
2
CINAHL
711
172
6
Medical societies/guideline websites/clearinghouses
N/A
N/A
N/A
54
3
Google
17,600,000
497
N/A Table 1 (abstract P166). Summary of results returned following systematic review Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S63 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P167 ENT, thoracics and A&E were approached. All completed forms were
included. P167
Survey of emergency tracheostomy management in the East of
England region
N Lawrence, L Oakley, C Swanavelder, M Palmer
West Suff olk Hospital, Bury St Edmunds, UK
Critical Care 2013, 17(Suppl 2):P167 (doi: 10.1186/cc12105) 2.
De Nicola A, Sucre MJ: Abstract 829. Proceedings Book of 40th Congress of the
Society of Critical Care Medicine 2011: January 15–19 2011; San Diego, CA.
Mount Prospect, IL: Society of Critical Care Medicine. Laryngeal mask as a safe and eff ective ventilatory device during
Blue Dolphin tracheostomy in the ICU A De Nicola1, MJ Sucre1, G Donnarumma1, A Corcione2
1San Leonardo Hospital, Castellammare di Stabia, Italy; 2Monaldi Hospital,
Naples, Italy A De Nicola1, MJ Sucre1, G Donnarumma1, A Corcione2
1San Leonardo Hospital, Castellammare di Stabia, Italy; 2Monaldi Hospital,
Naples, Italy Critical Care 2013, 17(Suppl 2):P169 (doi: 10.1186/cc12107) g
y
y
g
Conclusion Despite national guidance within the UK this survey
highlights that implementation and awareness of emergency
tracheostomy guidelines in ICUs in the EoE region is poor, and when
present they are not readily accessible in an emergency. Emergency
training has largely been informal and the availability of formal training
courses has not been recognised. In order to improve patient safety
there is a need to ensure that emergency tracheostomy management
including guidelines, equipment and formalised tracheostomy
emergency training are adopted and embraced universally. References Introduction Usually percutaneous tracheostomy is accomplished via
the tracheal tube. Some severe complications during percutaneous
dilatational tracheostomy (PDT) may be related to poor visualization
of tracheal structures. The alternative implies extubation and re-
insertion of a laryngeal mask (LMA). An accidental extubation as well
as an injuring of the vocal cords (because of the infl ated cuff during
dislocation) appears impossible in this method. Subjectively, the
bronchoscopic view obtained via a LMA seems to be better than that
obtained with an endotracheal tube (ET) [1,2]. Cook TM, et al.: Results of the 4th National Audit Project of the Royal 1. Cook TM, et al.: Results of the 4th National Audit Project of the Royal
College of Anaesthetists. Br J Anaesth 2011, 106:632-642. Methods In this prospective observational study, the bedside PDT
was performed using the Ciaglia Blue Dolphin method in 150 critically
ill patients. The patient’s tracheal tube was exchanged for a LMA
Fastrach™ before undertaking PDT. The insertion of the LMA, the quality
of ventilation, the blood gas values, the view of the tracheal puncture
site, and the view of the balloon dilatation were rated as follows: very
good (1), good (2), barely acceptable (3), poor (4), and very poor (5). Methods In this prospective observational study, the bedside PDT
was performed using the Ciaglia Blue Dolphin method in 150 critically
ill patients. The patient’s tracheal tube was exchanged for a LMA
Fastrach™ before undertaking PDT. References 1. Linstedt U, et al.: Anesth Analg 2010, 110:1076-1082. Methods An anonymous online survey was sent to all trainees who
may respond to a tracheostomy emergency in our organisation. Trainees in anaesthesia/critical care, general medicine, general surgery, 2. De Nicola A, Sucre MJ: Abstract 829. Proceedings Book of 40th Congress of the
Society of Critical Care Medicine 2011: January 15–19 2011; San Diego, CA. Mount Prospect, IL: Society of Critical Care Medicine. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S64 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P170 Figure 1 (abstract P171). P170
Real-time ultrasound-guided balloon dilatational percutaneous
tracheostomy is a safe procedure in critically ill patients:
an evaluation study
A Taha, A Shafi e, M Mostafa, P Wallen, H Hon, R Marktanner
Sheikh Khalifa Medical City, Abu Dhabi, United Arab Emirates
Critical Care 2013, 17(Suppl 2):P170 (doi: 10.1186/cc12108) Introduction Balloon dilatational percutaneous tracheostomy with
radial outward dilation minimizes bleeding and injury to tracheal
rings [1]. Using the bronchoscope during the puncture of the trachea
holds a risk of inaccurate placement, of hypoventilation and of needle
puncturing the bronchoscope. This study was conducted in order to
evaluate the safety features and feasibility of combining real-time
ultrasound-guided puncture of the trachea with visualization of
the needle path during tracheal puncture [2]. We provide precise
localization of tracheal cartilage and proper site puncture for the stoma
by ultrasound, with subsequent performance of balloon dilatational
percutaneous tracheostomy using the Blue dolphin technique.i p
y
g
p
q
Methods Twenty-fi ve patients including 15 males and 10 females,
mean age 61 years, with age range from 23 to 102 years, underwent
bedside percutaneous tracheostomy combining real-time ultrasound
and the Blue Dolphin technique. Results The median time for the procedure was 15 (12 to 20) minutes. Targeted placement for the tracheostoma between the second and
third or the third and fourth tracheal ring was achieved in 100%. No
signifi cant complications (for example, tracheal bleeding, puncture
posterior tracheal wall, misplacement of the tracheal cannula) occurred. One fractured tracheal ring was identifi ed using bronchoscopy after
the procedure. No conversion into a bronchoscopically guided or into a
surgical open technique was necessary. found, with no signifi cant diff erence between RV systolic function,
pulmonary acceleration time and pulmonary velocity time integral
between survivors and nonsurvivors. Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome Critical Care 2013, 17(Suppl 2):P172 (doi: 10.1186/cc12110) G Tavazzi1, M Bojan2, S Canestrini2, M White2, S Price2
1University of Pavia Foundation Policlinico San Matteo IRCCS, Pavia, Italy;
2Royal Brompton Hospital, London, UK Introduction Global left ventricular electromechanical dyssynchrony
(GLVD) is uncoordinated LV contraction that reduces the extent of
intrinsic energy transfer from the myocardium to the circulation leading
to a reduction in peak LV pressure rise, prolonged total isovolumic
time (t-IVT) and fall in stroke volume [1]. This potentially important
parameter is not routinely assessed in critically ill cardiothoracic
patients. Introduction Acute cor pulmonale (ACP) is associated with increased
mortality in patients ventilated for acute respiratory distress syndrome
(ARDS). Interventional lung assist (iLA) allows a lung-protective
ventilatory strategy, whilst allowing CO2 removal, but requires adequate
right ventricular (RV) function. RV restriction (including presystolic
pulmonary A wave) [1] is not routinely assessed in ARDS. p
Methods A prospective analysis of retrospectively collected data
in cardiothoracic ICU patients who underwent echocardiography
was performed. In addition to epidemiological factors, echo data
included comprehensive assessment of LV/RV systolic and diastolic
function including Doppler analysis of isovolumic contraction/
relaxation, ejection time (ET) and fi lling time (FT). t-IVT was calculated
as (60 – (total ET + total FT)) and the Tei Index as (ICT + IRT) / ET. t-IVT
>14 second/minute and Tei index >0.48 were used to defi ne GLVD [2]. Data are shown as mean ± SD/median (interquartile range). p
y
y
Methods A prospective analysis of retrospectively collected data
in patients with echo during iLA was performed. Data included
epidemiologic and ventilatory factors, LV/RV function, evidence of
RV restriction and pulmonary hemodynamics. Data are shown as
mean ± SD/median (interquartile range). Results Thirty-two patients (45 ± 17 years), 22 male (68%), SOFA score
11.15 ± 2.38 were included. Pulmonary hypertension (PHT) was 53%,
and hospital mortality 43%. Mortality was not associated with age,
days on iLA, length of ICU stay, inotropic support, nitric oxide or level
of ventilatory support, but was associated with pressor requirement
(P = 0.005), a worse PaO2:FiO2 ratio (9.4 (7.8 to 12.6) vs. 15.2 (10.7 to
23.9), P = 0.009) and higher pulmonary artery pressures (56.5 mmHg
(50 to 60) vs. 44.5 (40.5 to 51.2), P = 0.02). No echo features of ACP were g
Results A total of 103 patients (63.5 ± 18.4 years), 65 male (63%), APACHE
II score (14.6 ± 7.4) were included. References References
1. Gromann TW, et al.: Anesth Analg 2009, 108:1862-1866. 2. Venkatakrishna Rajajee et al.: Crit Care 2011, 15:R54. 3. Guinot et al.: Crit Care 2012, 16:R40. P172 Left ventricular electromechanical dyssynchrony and mortality in
cardiothoracic intensive care G Tavazzi , M Bojan , A Duncan , A Vazir , S Price
1University of Pavia Foundation Policlinico San Matteo IRCCS, Pavia, Italy;
2Necker-Enfants Malades University Hospital, Paris, France; 3Royal Brompton
Hospital, London, UK References The incidence of RV restriction
was high (43%), and independent of PHT, RV systolic function and level
of respiratory support, but correlated with CO2 levels (restrictive 7.1 kPa
(7.4 to 8.0) vs. 6.1 (5.8 to 6.8), P = 0.03). See Figure 1. Conclusion This study demonstrates the feasibility and safeness
combining real-time ultrasound guidance and balloon dilatational
percutaneous tracheostomy (Blue Dolphin technique) in critically
ill patients. In our division, this technique has become the standard
bedside tracheostomy procedure because it combines excellent
patient safety features with avoidance of intraprocedural tube
misplacement, hypoventilation and accidental bronchoscopy damage
even in technically diffi cult cases [3]. g
Conclusion Typical echo features of ACP were not seen in this study,
possibly because of the protective ventilatory strategies allowed by
use of iLA. The incidence of RV restriction may refl ect more subtle
abnormalities of RV function. Further studies are required to elucidate
RV pathophysiology in critically ill adult patients with ARDS. Reference 1. Cullen S, et al.: Circulation 1995, 91:1782-1789. P171 P171
Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome
G Tavazzi1, M Bojan2, S Canestrini2, M White2, S Price2
1University of Pavia Foundation Policlinico San Matteo IRCCS, Pavia, Italy;
2Royal Brompton Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P171 (doi: 10.1186/cc12109) P171
Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome
G Tavazzi1, M Bojan2, S Canestrini2, M White2, S Price2
1University of Pavia Foundation Policlinico San Matteo IRCCS, Pavia, Italy;
2Royal Brompton Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P171 (doi: 10.1186/cc12109) Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome
G Tavazzi1, M Bojan2, S Canestrini2, M White2, S Price2
1University of Pavia Foundation Policlinico San Matteo IRCCS, Pavia, Italy;
2Royal Brompton Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P171 (doi: 10.1186/cc12109) Intracardiac ECG for confi rmation of correct positioning of central
venous catheters is safe and cost-eff ective M Eriksson, R Dörenberg Surgical Sciences, Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P175 (doi: 10.1186/cc12113) Methods Data for all ultrasound-guided interventions, including
complications, are prospectively collected in our department for audit
purposes; in this study we involved only CVC insertions in the ICU
between February 2011 and November 2012. Electronic medical and
laboratory records and paper-based nursing charts were retrospectively
studied for all interventions, specifi cally looking for blood results,
coagulation abnormalities and intervention-related complications. Introduction About 10 years ago the use of chest radiographs as
the golden standard to ensure correct positioning of central venous
catheters (CVC) was questioned. The frequent use of CVCs was also
challenged. We decided to retrospectively evaluate our routines in a
large surgical unit in a Swedish university hospital. Methods All X-rays were centrally registered. Chest X-ray performed in
our unit is almost entirely used to confi rm CVC positioning. The Certofi x
CVC set for the Seldinger technique in combination with Certodyn –
Universaladapter (B Braun, Germany) is now used as the routine
equipment and the right jugular vein is our standard approach. Results In the study period, ultrasound guidance was employed for a
total of 291 central line insertions in 220 ICU patients. Coagulopathy
was detected in 127 cases at the time of CVC placement (43.6%). On the
day of CVC insertion, coagulation abnormalities were corrected in 20
cases (15.7%); 33 out of 50 patients with severe coagulopathy (66.0%)
and 74 out of 77 patients with coagulopathy of moderate severity
(96.1%) had no correction at all. Correction was started only after CVC
insertion for reasons unrelated to CVC placement in a further eight
and two patients with severe and less severe coagulopathy (16.0% and
2.6%), respectively. No bleeding complications were observed. Results In 2002 the total number of X-rays performed in patients at
our unit was 2,306, which corresponds to the approximate number
of inserted CVCs at that time, since a confi rmatory X-ray was routine. X-rays were rarely performed on other indications in our unit. X-ray
costs were at that time approximately €300,000 (~€130/each). The year
after, 1,726 chest X-rays were performed, refl ecting both the use of
intracardiac confi rmation of correct CVC position and also a reduced
use of CVCs. This trend has continued over time. In 2011 approximately
600 CVCs were inserted at our unit. X-rays were performed in about
20% of these cases. Cyanoacrylate glue prevents early bleeding of the exit site after CVC
or PICC placement p
G Scoppettuolo, MG Annetta, C Marano, E Tanzarella, M Pittiruti
Catholic University, Rome, Italy
Critical Care 2013, 17(Suppl 2):P174 (doi: 10.1186/cc12112) p
G Scoppettuolo, MG Annetta, C Marano, E Tanzarella, M Pittiruti
Catholic University, Rome, Italy
C i i
l C
2013 17(S
l 2) P174 (d i 10 1186/
12112) y
y
itical Care 2013, 17(Suppl 2):P174 (doi: 10.1186/cc12112) Intracardiac ECG for confi rmation of correct positioning of central
venous catheters is safe and cost-eff ective The cost for a chest X-ray is today ~€200, meaning
that X-ray costs were approximately €24,000. We have not experienced
any medical problems when intracardiac ECG was used for positioning
confi rmation. On the contrary, aspiration of venous blood without
apparent p-waves in a patient with sinus rhythm may suggest improper
placement of the CVC; for example, the right brachial vein. p
y
g
p
Conclusion In patients undergoing CVC insertion in our ICU,
coagulopathy is common. We observed uncomplicated CVC placement
in all 41 patients with severe uncorrected coagulopathy and in a further
76 patients with coagulopathy of moderate severity. When combined
with other studies, our data suggest that ultrasound-guided CVC
placement without routine correction of coagulation abnormalities
may be safe in the ICU. Safety of ultrasound-guided central venous access in critically ill
patients with uncorrected coagulopathy Safety of ultrasound-guided central venous access in critically ill
patients with uncorrected coagulopathy
G Reusz1, C Langer1, G Egervari1, P Sarkany1, A Csomos2
1Markhot Ferenc Hospital, Eger, Hungary; 2Semmelweis University, Budapest,
Hungary
Critical Care 2013, 17(Suppl 2):P173 (doi: 10.1186/cc12111) Conclusion Glue is an inexpensive and highly eff ective tool for avoiding
the risk of early bleeding of the exit site after catheter placement. We
also suggest that in the next future the glue might prove to have
benefi cial collateral eff ects on the risk of extraluminal contamination
(by reducing the entrance of bacteria in the space between the catheter
and the skin), as well as on the risk of dislocation (by increasing the
stability of the catheter inside the skin breach). y
G Reusz1, C Langer1, G Egervari1, P Sarkany1, A Csomos2 g
g
y
1Markhot Ferenc Hospital, Eger, Hungary; 2Semmelweis University, Budapest,
Hungary g
g
y
1Markhot Ferenc Hospital, Eger, Hungary; 2Semmelweis University, Budapest,
Hungary y
Critical Care 2013, 17(Suppl 2):P173 (doi: 10.1186/cc12111) Introduction Correction of coagulopathy before central venous
catheter (CVC) insertion is a common practice; however, when
ultrasound guidance is used this is controversial as mechanical
complications are rare. Studies in oncology patients suggest that CVC
placement without prior correction of coagulopathy is safe but no
studies are available for critically ill patients and guidelines do not give
recommendations [1,2]. We do not routinely correct coagulopathy,
even if severe, when ultrasound guidance is used and the purpose of
this retrospective study was to evaluate the safety of this practice. P175 Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome Results In 65 consecutive patients (45 PICCs, 11 dialysis catheters
and nine CVCs), there was no signifi cant local bleeding at 1 hour or at
24 hours after catheter placement. No local adverse reaction occurred. No damage to the polyurethane of the catheters was detected. Conclusion Glue is an inexpensive and highly eff ective tool for avoiding
the risk of early bleeding of the exit site after catheter placement. We
also suggest that in the next future the glue might prove to have
benefi cial collateral eff ects on the risk of extraluminal contamination
(by reducing the entrance of bacteria in the space between the catheter
and the skin), as well as on the risk of dislocation (by increasing the
stability of the catheter inside the skin breach). Methods The aim of this pilot study was to verify the effi cacy of a
cyanoacrylate glue in reducing the risk of early bleeding at the exit
site after CVC or PICC placement. We studied a group of adult patients
consecutively undergoing placement of polyurethane CVCs or PICCs
without reverse tapering in a non-intensive ward of our hospital. All
lines were inserted according to the same protocol, which included 2%
chlorhexidine antisepsis, maximal sterile barriers, ultrasound guidance,
EKG guidance and securement with sutureless device. Two minutes
after placement of the glue, the exit site was covered with a temporary
gauze dressing, which was replaced by transparent membrane at
24 hours. All patients were assessed at 1 hour and at 24 hours. fi
Conclusion GLVD that limits cardiac output is common in the
cardiothoracic ICU, and signifi cantly related to mortality. When
diagnosed, the underlying cause should be sought and treatment
instigated to minimize the t-iVT (pacing optimization/revascularization/
inotrope titration/volaemia optimization). References References
1. Cavanna L, et al.: World J Surg Oncol 2010, 8:91. 2. Lamperti M, et al.: Intensive Care Med 2012; 38:1105-1117. 1. Cavanna L, et al.: World J Surg Oncol 2010, 8:91. 2. Lamperti M, et al.: Intensive Care Med 2012; 38:1105-1117. Conclusion If we had continued to use CVCs at the same frequency
as we did 10 years ago, and used X-ray confi rmation in practically all
cases, we would have paid approximately €460,000 annually. Reduced
use of CVCs, in combination with intracardiac confi rmation of CVC
positioning, has not only allowed us to reduce costs associated with
CVC insertion by more than €400,000, corresponding to a reduction
rate of more than 90%, but also decreased the patient’s exposure to
X-ray irradiation. Stas M, et al.: Eur J Surg Oncol 2001, 27:316-320.
Joshi A, et al.: Indian Crit Care Med 2008, 12:10-14. p
References 1. Duncan A, et al.: J Am Coll Cardiol 2003, 41:121-128. 2. Tei C, et al.: J Am Coll Cardiol 1996, 28:658-664. 1. Duncan A, et al.: J Am Coll Cardiol 2003, 41:121-128. 2. Tei C, et al.: J Am Coll Cardiol 1996, 28:658-664. p
Results In 65 consecutive patients (45 PICCs, 11 dialysis catheters
and nine CVCs), there was no signifi cant local bleeding at 1 hour or at
24 hours after catheter placement. No local adverse reaction occurred. No damage to the polyurethane of the catheters was detected.f Right ventricular restriction in interventional lung assist for acute
respiratory distress syndrome The prevalence of GLVD was high
(24/103, 22%) and associated with signifi cantly increased mortality,
7.5% vs. 25% (P = 0.02). There was no diff erence in requirement for
cardiorespiratory support between the two populations, but there
were signifi cant diff erences (no GLVD vs. GLVD) in requirement for Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 patient’s care and logistical problems. In our experience, the rate of
signifi cant local bleeding after placement of PICCs without reverse
tapering may be as high as 40% at 1 hour and 15% at 24 hours, while
the rate of bleeding after placement of a large-bore dialysis catheter is
above 50% at 1 hour. pacing (35% vs. 62%, P = 0.02), atrial fi brillation (20% vs. 41%, P = 0.03),
QRS duration (92.0 (80.0 to 120.0) vs. 116.5 (95.0 to 154.0), P = 0.01) and
QTc (460.0 (416.0 to 498.5) vs. 477.5 (451.2 to 541.0), P = 0.02). There
was no signifi cant diff erence in ejection fraction (no GLVD 43.0 (35.0
to 49.5) vs. GLVD 39.6 (29.5 to 49.7), P = 0.43), mitral regurgitation
(40.5% vs. 62.5%, P = 0.06), or any other measures of LV systolic or
diastolic function between the two groups. There was good correlation
between the two methods used to assess dyssynchrony (LV t-IVT:LV Tei
index correlation coeffi cient = 0.80, P <0.001). Methods The aim of this pilot study was to verify the effi cacy of a
cyanoacrylate glue in reducing the risk of early bleeding at the exit
site after CVC or PICC placement. We studied a group of adult patients
consecutively undergoing placement of polyurethane CVCs or PICCs
without reverse tapering in a non-intensive ward of our hospital. All
lines were inserted according to the same protocol, which included 2%
chlorhexidine antisepsis, maximal sterile barriers, ultrasound guidance,
EKG guidance and securement with sutureless device. Two minutes
after placement of the glue, the exit site was covered with a temporary
gauze dressing, which was replaced by transparent membrane at
24 hours. All patients were assessed at 1 hour and at 24 hours. Is automated brachial cuff measurement of arterial pressure less
accurate in cases of arrhythmia? y
K Lakhal1, S Faiz2, M Martin1, AS Crouzet1, F Reminiac2, S Ehrmann2,
R Cinotti1, X Capdevila3, K Asehnoune4, Y Blanloeil1, B Rozec1, T Boulain2
1Centre Hospitalier Universitaire Laennec, Nantes, France; 2Centre Hospitalier
Universitaire Bretonneau, Tours, France; 3Centre Hospitalier Universitaire
Lapeyronie, Montpellier, France; 4Centre Hospitalier Universitaire Hotel-Dieu,
Nantes, France Introduction In cases of arrhythmia, the beat-to-beat variation of
arterial pressure (AP) may impair the accuracy of automated cuff
measurements. Indeed, this oscillometric device relies on the detection
of arterial wall oscillations. Our aim was to determine, in ICU patients,
whether brachial cuff measurements are really less reliable during
arrhythmia than during regular rhythm. y
g
g
y
Methods Patients with arrhythmia and carrying an intra-arterial
catheter were prospectively and consecutively included in this multi-
center study. After each arrhythmic inclusion, a regular rhythm patient
was included. A second inclusion was possible in case of change in
the cardiac rhythm. Three pairs of invasive and brachial cuff (Philips®
MP70 monitor) measurements of mean arterial pressure (MAP) were
respectively averaged. Some patients underwent a second set of
measurements, after a cardiovascular intervention (passive leg raising,
volume expansion, initiation of/increase in catecholamine infusion)
allowing the assessment of MAP changes. Figure 2 (abstract P177). Use of CO data. Results In the 111 analyzed inclusions (in 103 patients) there was
only one failure in displaying a brachial cuff measurement of MAP. Arrhythmic patients (atrial fi brillation 88%, frequent extrasystoles
7%, fl utter 5%) were similar (P >0.3) to patients in regular rhythm
for MAP (median 74 (IQR 67 to 80) vs. 75 (69 to 84) mmHg), SAPS II
score, BMI, arm circumference, norepinephrine administration (36%
vs. 35%), mechanical ventilation (80% vs. 81%), and site of the intra-
arterial catheter (radial artery: 80% vs. 85%). Between arrhythmic
and regular rhythm patients: the agreement (Bland–Altman analysis)
between invasive and brachial cuff measurements of MAP was similar
(mean bias –0.4 ± 7.2 (limits of agreement –14/14) mmHg vs. 3.0 ± 8.2
(–13/19) mmHg, respectively); the detection of hypotension (invasive
MAP <65 mmHg) by the brachial cuff was of similar reliability (area
under the ROC curve (AUC) = 0.91 (95% CI = 0.80 to 0.97) vs. AUC = 0.98
(0.89 to 1), P = 0.2); and the detection of a response (>10% increase in
MAP) to therapy was of similar reliability (AUC = 1 (0.85 to 1) (n = 22) vs. Is automated brachial cuff measurement of arterial pressure less
accurate in cases of arrhythmia? AUC = 0.99 (0.78 to 1) (n = 17), P = 0.5). Figure 2 (abstract P177). Use of CO data. Results Fifty-three patients (18.5%) had evidence of CO monitoring. LiDCO was the most popular method (Figure 1). A total of 264 (94%)
patients received treatment with vasopressors and/or inotropes. CO
data were utilised in a variety of ways (Figure 2). Conclusion The majority of potential donors require vasopressors
and/or inotropes post BSD, but it seems only a minority currently have
their CO monitored. There is variation in how CO data are utilised to
direct haemodynamic management. We welcome the development of
standardised bundle-driven donor management. Conclusion These preliminary results suggest that arrhythmia does not
impair the reliability of automated cuff measurements of MAP. y
References Introduction Early bleeding from the exit site after CVC or PICC
placement is a very common event that causes diffi culties in the Stas M, et al.: Eur J Surg Oncol 2001, 27:316-320. Joshi A, et al.: Indian Crit Care Med 2008, 12:10-14. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S66 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P177). CO device. Figure 2 (abstract P177). Use of CO data. P176 Figure 1 (abstract P177). CO device. P176
Is automated brachial cuff measurement of arterial pressure less
accurate in cases of arrhythmia? K Lakhal1, S Faiz2, M Martin1, AS Crouzet1, F Reminiac2, S Ehrmann2,
R Cinotti1, X Capdevila3, K Asehnoune4, Y Blanloeil1, B Rozec1, T Boulain2
1Centre Hospitalier Universitaire Laennec, Nantes, France; 2Centre Hospitalier
Universitaire Bretonneau, Tours, France; 3Centre Hospitalier Universitaire
Lapeyronie, Montpellier, France; 4Centre Hospitalier Universitaire Hotel-Dieu,
Nantes, France
Critical Care 2013, 17(Suppl 2):P176 (doi: 10.1186/cc12114) Reference 1. Murugan R, et al.; HIDonOR Study Investigators: Preload responsiveness is
associated with increased interleukin-6 and lower organ yield from brain-
dead donors. Crit Care Med 2009, 37:2387-2393. Cardiac output monitoring in brain-stem-dead potential organ
donors: an audit of current UK practice
CJ Wright1, A Broderick2, G Mandersloot3
1Glasgow Royal Infi rmary, Glasgow, UK; 2National Health Service Blood and
Transplant, London, UK; 3The Royal London Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P177 (doi: 10.1186/cc12115) P179 Prediction of 28-day mortality by indocyanine green disappearance
rate, other markers of hepatic function and transpulmonary
thermodilution parameters: a prospective study in 154 patients
W Huber, D Ertekin, T Langer, B Saugel, T Lahmer, M Messer, C Spinner,
C Schultheiss, RM Schmid
Klinikum rechts der Isar, Technical University of Munich, Germany
Critical Care 2013, 17(Suppl 2):P179 (doi: 10.1186/cc12117) Methods A prospective observational study was designed and
conducted. Inclusion criteria were: patients who had underwent
elective or urgent/emergent cardiac surgery, with cardiac index (CI)
<2.5 l/minute/m2 estimated by means of a PAC, left ventricle ejection
fraction (LVEF) <40%, lactate >2 mmol/l, age >18 years. A central venous
catheter (CVC) and a PAC were inserted for each patient before surgery
in the same right internal jugular vein in accordance with standard
procedure. Proper position of the PAC was confi rmed with pressure
tracings and chest X-ray. Mixed and central venous blood samples
were collected from the distal ports of the PAC and CVC respectively
30 minutes after ICU admission, and every 6 hours for a total of three
samples in a 24-hour period for each patient. All blood samples were
analyzed by a co-oximeter (Radiometer ABL800 fl ex; Radiometer,
Copenhagen, Denmark). Statistical analysis was performed by Stats
Direct (Ver.2.5.8, Cheshire, UK) and GraphPad (Vers. Prism 4.0; San
Diego, CA, USA). All data were tested for normal distribution with the
Kolmogorov–Smirnov test. Statistical analysis was performed by linear
regression analysis. The agreement between absolute values of ScvO2
and SvO2 were assessed by the mean bias and 95% limits of agreement
(LOA) ((mean bias ± 1.96)×standard deviation) according to the method
described by Bland and Altman [5]. Introduction Hepatic dysfunction has been associated with outcome of
ICU patients. However, most scoring systems including APACHE II only
marginally refl ect acute liver dysfunction on admission. Indocyanine
green (ICG) is eliminated by hepatobiliary excretion. Therefore, the
ICG plasma disappearance rate (ICG-PDR) is used as a dynamic liver
function test. ICG-PDR has been associated with mortality in several
studies. Methods A prospective study to compare prediction of 28-day
mortality by ICG-PDR, other markers of liver function and scoring
systems (primary endpoint). In the subgroup of patients with trans-
pulmonary thermodilution (TPTD) monitoring (PiCCO device; Pulsion,
Munich, Germany), predictive capabilities of ICG-PDR were compared
with cardiac index (CI), extravascular lung water index (EVLWI),
global end-diastolic volume index (GEDVI) and pulmonary vascular
permeability index (PVPI). ICG-PDR (i.v. Indocyanine green plasma disappearance rate for assessment
of liver function: re-evaluation of normal ranges and impact of
biometric data Introduction Signifi cant changes in haemodynamics occur after brain
stem death (BSD) and there is evidence that yield of transplantable
organs may be decreased in donors who remain preload responsive
prior to donation [1], suggesting that optimisation of the cardiac
output (CO) may be benefi cial in potential organ donors. We describe
current UK practice with regard to CO monitoring in this group. W Huber, M Kranzmayr, C Schultheiss, W Reindl, A Krug, B Saugel, U Mayr,
RM Schmid W Huber, M Kranzmayr, C Schultheiss, W Reindl, A Krug, B Saugel, U Mayr,
RM Schmid Klinikum rechts der Isar, Technical University of Munich, Germany
Critical Care 2013, 17(Suppl 2):P178 (doi: 10.1186/cc12116) Introduction The indocyanine green plasma disappearance rate
(ICG-PDR) is a dynamic liver function test that can be non-invasively
measured by pulse densitometry. ICG-PDR is associated with mortality
and other markers of outcome. Due to predominant use of ICG-PDR
in the ICU setting, the normal range is based on scarce data available
outside the ICU and given with 18 to 25%/minute. Methods We reviewed a database of 287 brain-stem-dead potential
organ donors collected by specialist nurses in organ donation
(SN-OD) over a 6-month period (30 April 2011 to 31 October 2011)
across multiple UK centres. The database contained data on donor
management in the period from initial SN-OD review to immediately
prior to transfer to the operating theatre. We analysed data on CO
monitoring and vasopressor/inotrope use. Where information was
missing/not recorded in the dataset, the treatment referred to was
interpreted as not given/not done. Methods To prospectively re-evaluate the normal range and to analyze
the potential impact of biometric data on ICG-PDR, we measured
ICG-PDR (i.v. injection of 0.25 mg/kg ICG; LiMON, Pulsion, Munich, S67 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 independently associated with age (P = 0.025), but not with any other
biometric parameter (gender, weight and height). Germany) in 95 outpatients with ulcerative colitis (UC). Due to a
prevalence of primary sclerosing cholangitis (PSC) of 5% in UC, patients
were asked regarding previous diagnosis of PSC. Additionally, serum
bilirubin, AST, ALT, INR, AP, γGT and cholinesterase were determined. Association of biometric data (age, gender, height, weight) and
diagnosis of PSC on ICG-PDR were evaluated using Spearman corre-
lation, ROC and multivariate analysis (statistics: IBM SPSS 20). Indocyanine green plasma disappearance rate for assessment
of liver function: re-evaluation of normal ranges and impact of
biometric data Conclusion ICG-PDR on admission is an independent predictor of
28-day mortality. Predictive capabilities particularly of APACHE II can
be improved by combination with ICG-PDR. Among TPTD-derived
parameters, only PVPI provides signifi cant prediction of mortality. y
Results A total of 95 patients (54 male, 41 female), age 43.2 ± 15.5 (21 to
76) years, weight 74 ± 16 kg, height 175 ± 10 cm, BMI 23.7 ± 4, bilirubin
0.48 ± 0.27 mg/dl, AST 26.8 ± 11.4 U/l, ALT 26.8 ± 14.9 U/l, previous
diagnosis of PSC 8/95 (8.4%). ICG-PDR ranged from 13.8 to 44.0 with a
mean of 28.2 ± 6.8 and a median of 27.0%/minute. In univariate analysis
ICG-PDR was signifi cantly associated with age (r = –0.480; P <0.001),
weight (r = –0.262; P = 0.011) and female gender (r = 0.221; P = 0.032),
but not to height (P = 0.681). In multivariate analysis (R = 0.459)
including the variables age, gender, height, weight and diagnosis of
PSC, only age was independently (P <0.001) associated with ICG-PDR. With each year in age, ICG-PDR decreased by 0.206%/minute. In ROC
analysis, ICG-PDR above the upper normal range (>25%/minute) was
signifi cantly associated with young age (AUC 0.746; P <0.001). P180 P180 Mixed and central venous oxygen saturation are not
interchangeable in patients with cardiogenic shock after cardiac
surgery
S Romagnoli, P Balsorano, F Landucci, A De Gaudio
AOUC Careggi, Florence, Italy
Critical Care 2013, 17(Suppl 2):P180 (doi: 10.1186/cc12118) g
y
S Romagnoli, P Balsorano, F Landucci, A De Gaudio
AOUC Careggi, Florence, Italy Introduction Mixed venous oxygen saturation (SVO2) represents a
well-recognized parameter of oxygen delivery (DO2)–consumption
(VO2) mismatch and its use has been advocated in critically ill patients
in order to guide hemodynamic resuscitation [1] and oxygen delivery
optimization. Nevertheless, the pulmonary artery catheter (PAC) is not
readily available and its use is not devoid of risks. Furthermore, its use
has been decreasing in recent years in surgical and cardiac surgical
patients as the benefi t of guiding therapy with this device is unclear
[2-4]. Central venous oxygen saturation (ScVO2) has been suggested
as an alternative to SVO2 monitoring due to its feasibility in several
settings. Unfortunately concerns arise from its capability to correlate
with SVO2, the relationship being infl uenced by several factors, such
as hemodynamic impairment and pathological process. Hemodynamic
instability and shock often complicate cardiac surgery, and the SVO2–
ScVO2 relationship has not been specifi cally investigated in this setting. The aim of this study is to compare SVO2 and ScVO2 values in patients
with cardiogenic shock after cardiac surgery. i
Conclusion ICG-PDR (normal) values should be corrected for age. With
a decrease in ICG-PDR of 0.206% per year, the range between the upper
and lower normal level (25 to 18%/minute) is passed through within
about 35 years of life. These fi ndings are in accordance with functional
loss of other organ functions (for example, cardiac output, glomerular
fi ltration rate) with increasing age. Normal ranges of innovative
markers of organ function mainly derived from ICU populations should
be re-evaluated outside the ICU. Utility of transesophageal echocardiography in the ICU:
a preliminary US perspective References
1. Rivers E, et al.: Early goal-directed therapy in the treatment of severe sepsis
and septic shock. N Engl J Med 2001, 345:1368-1377. 2. Edwards JD: Oxygen transport in cardiogenic and septic shock. Crit Care
Med 1991, 19:658-663. 3. Hadian M, et al.: Evidence based of the use of the pulmonary artery
catheter: impact data and complications. Crit Care 2006, 10(Suppl
3):S11-S18. 4. Connors AF, et al.: The eff ectiveness of right heart catheterization in the
initial care of critically ill patients. JAMA 1996, 276:889-897. 5. Bland JM, Altman DG: Statistical methods for assessing agreement
between two methods of clinical measurement. Lancet 1986, 1:307-310. 6. Walley KR: Use of central venous oxygen saturation to guide therapy. Am J
Respir Crit Care Med 2011, 184:514-520. Introduction While TEE is providing a direct assessment of the cardiac
function and volume status as a diagnostic tool, until recently it has
been impractical to be continuously available for monitoring. A new
disposable, monoplanar TEE probe (ImaCor) can remain in the patient
for up to 72 hours, allowing repeated measures of ventricular function
and volume status, parameters needed to monitor response to therapy. Methods We assessed the benefi t these TEE data provided in the
assessment of fi ve domains: hypovolemia, right ventricular dysfunction,
left ventricular dysfunction, sepsis, and valvular abnormality. Bedside
practitioners listed their diagnoses before and after seeing primary
TEE images perform by trained physicians. We used a 0 to 5 Likert scale
to assess diff erential diagnosis before and after the TEE, comparing
changes using a paired t test. 1. Rivers E, et al.: Early goal-directed therapy in the treatment of severe sepsis
and septic shock. N Engl J Med 2001, 345:1368-1377. 1. Rivers E, et al.: Early goal-directed therapy in the treatment of severe sepsis
and septic shock. N Engl J Med 2001, 345:1368-1377. 2. Edwards JD: Oxygen transport in cardiogenic and septic shock. Crit Care
Med 1991, 19:658-663. 3. Hadian M, et al.: Evidence based of the use of the pulmonary artery
catheter: impact data and complications. Crit Care 2006, 10(Suppl
3):S11-S18.f 3. Hadian M, et al.: Evidence based of the use of the pulmonary artery
catheter: impact data and complications. Crit Care 2006, 10(Suppl
3):S11-S18.f g
g
p
Results All requests for TEE were to access hemodynamic instability. A
total of 18 patients were screened and nine were eligible, in which 16
total TEE studies were performed. Utility of transesophageal echocardiography in the ICU:
a preliminary US perspective There were no complications with
TEE and all patients tolerated the long-term placement of the probe
well. Of the fi ve diagnostic domains studied, right ventricular failure was
the most commonly underdiagnosed contributor to the hemodynamic
instability among patients prior to TEE (P = 0.054) (Figures 1 and 2). 7. Reinhart K, et al.: Comparison of central-venous to mixed-venous oxygen
saturation during changes in oxygen supply/demand. Chest 1989,
95:1216-1221. 7. Reinhart K, et al.: Comparison of central-venous to mixed-venous oxygen
saturation during changes in oxygen supply/demand. Chest 1989,
95:1216-1221. P179 During circulatory shock,
not homogeneous oxygen extraction and regional blood fl ow Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S68 Figure 1 (abstract P180). Table 1 (abstract P181). Hemodynamic parameters before and after
treatment Group 2 compared with Group 1 (P <0.001). The increase in plasma NO
level was signifi cantly higher in Group 2 (Table 1). Conclusion L-Arginine i.v. infusion 5 ml/kg during 24 hours increased
NO blood level and decreased MPAP in pediatric sepsis, but did not
deteriorate hemodynamic system values. Table 1 (abstract P181). Hemodynamic parameters before and after
treatment
Group 1
Group 1
Group 2
Group 2
Value
before
after
before
after
MAP (mmHg)
66.9 ± 3.4
68.8 ± 3.0
72.8 ± 2.0
71.8 ± 1.3
MPAP (mmHg)
43.5 ± 2.6
44.3 ± 3.4
50.8 ± 3.0
28.8 ± 2.1
CO (l/minute/m2)
4.2 ± 0.2
5.0 ± 0.4
4.8 ± 0.2
4.8 ± 0.3
SpO2 (%)
93.7 ± 1.2
93.4 ± 1.5
94.6 ± 0.7
98.1 ± 0.3 redistribution make SVO2 a more reliable parameter suggesting the
global adequacy of cardiac output rather than ScVO2. In this study
we aimed at evaluating SvO2–ScVO2 diff erences in patients with
cardiogenic shock, as defi ned by hyperlactatemia, low CI, and LVEF
<40%, after cardiac surgery. Our results highlighted a great variability
for these two parameters, with a clinically unacceptable mean bias and
LOA. As expected, ScVO2 values were consistently higher. References P179 bolus of 0.25 mg/kg ICG; LiMON
device, Pulsion) and all other predictors were determined within
48 hours after admission. Statistics: IBM SPSS 20. Results A total of 20 patients were enrolled. In 18 out of 20 cases all
three blood samples were collected. In two patients only two blood
samples were drawn as they exited the inclusion criteria. Linear
regression analysis between the two variables resulted in an r2 of 0.708. Bland–Altman analysis (Figure 1) for the pooled measurements of SvO2
and ScvO2 showed a mean bias and LOA of 6.82% (SD of bias 5.3) and
–3.71 to +17.3% respectively. Results A total of 154 patients (46 female, 108 male), age 59 ± 13 years,
APACHE II score 16.0 ± 5.7, SOFA score 7.6 ± 4.2. Etiology: sepsis 14.4%,
cirrhosis 28.8%, GI bleeding 8.9%, ARDS 17.8%, cardiogenic shock
4.1%, acute renal failure 3.4%, various 22.6%. The 28-day mortality was
signifi cantly predicted by APACHE II (ROC-AUC: 0.762; P <0.001) and
SOFA (AUC: 0.784; P <0.001). Among markers of hepatic function on
admission, ICG-PDR provided the largest AUC (0.742; P <0.001), which
was larger than for GOT (AUC: 0.646; P = 0.019), bilirubin (AUC: 0.641;
P = 0.023) and INR (AUC: 0.614; P = 0.067). Among TPTD parameters, only
PVPI signifi cantly predicted 28-day mortality (AUC: 0.643; P = 0.043),
whereas CI, GEDVI and EVLWI were not predictive. Prediction of 28-day
mortality by SOFA could not be improved by models including any
hepatic parameter. By contrast, ICG-PDR was independently (P = 0.036)
associated with mortality when included in a model (R = 0.58) with
APACHE II. This model based on APACHE II and ICG-PDR provided
the largest of all ROC-AUCs (AUC: 0.804; P <0.001). ICG-PDR itself was y
Conclusion ScVO2 has been advocated as an attractive and simple
indicator of DO2–VO2 mismatch [6]. Its role as a surrogate of the well-
established SVO2 has been investigated in several settings, and it
has been purposed in the hemodynamic resuscitation of critically
ill septic patients [1]. Nevertheless, the SVO2–ScVO2 relationship
can be infl uenced by several factors due to ScVO2 dependency from
global blood fl ow redistribution that can occur during hemodynamic
impairments. It has been shown previously that in healthy people
ScVO2 values tend to underestimate SVO2 values, due to the higher
oxygen content from inferior vena cava [7]. P181 Pulmonary hemodynamic disorder in pediatric sepsis and their
correction with L-arginine infusion
M Georgiyants, V Korsunov
Kharkov Medical Academy Post-Graduate Education, Kharkov, Ukraine
Critical Care 2013, 17(Suppl 2):P181 (doi: 10.1186/cc12119) g
M Georgiyants, V Korsunov Figure 1 (abstract P182). Ventricular function according to the Likert
scale. Kharkov Medical Academy Post-Graduate Education, Kharkov, Ukraine
Critical Care 2013, 17(Suppl 2):P181 (doi: 10.1186/cc12119) Introduction The acute respiratory distress syndrome and pulmonary
hypertension (PH) is one of the factors of septic mortality. Some data
demonstrate arginine defi ciency as an important pathogenic factor of
septic PH. We suppose that intravenous L-arginine infusion improves
NO production and reduce PH. Methods We examined 46 patients with sepsis, severe sepsis and septic
shock in accordance with San Antonio Criteria. Organ dysfunction
severity was defi ned according to SOFA. The mean pulmonary artery
pressure (MPAP) was estimated by Doppler method, cardiac output
(CO), and stroke volume (SV) – by ultrasound M-mode. SpO2, mean
arterial pressure (MAP), blood gas, and plasma NO level was evaluated. The patients of Group 1 (n = 21, age 22.1 ± 8.5 months) had respiratory
and hemodynamic support, and antibiotics. Group 2 (n = 25, age
27.0 ± 11.2 months) had the same treatment with continuous i.v. infusion of 5 ml/kg body weight 4.2% L-arginine solution during
24 hours. Results The patients of Groups 1 and 2 did not have statistical
diff erence of SOFA (4.1 ± 1.0 vs. 4.4 ± 0.7), need for ventilator support
(57.0 ± 11.0% vs. 44.0 ± 10.0%), and dose of inotropes (P >0.05). The
reduction of MPAP and increase of SaO2 was signifi cantly higher in Figure 1 (abstract P182). Ventricular function according to the Likert
scale. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S69 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 2 (abstract P182). Volume status, valve, sepsis according to the
Likert scale. ICU in Thailand that was capable of measuring extravascular lung
water. None of the Thai ICUs used transcutaneous PCO2, near-infrared
spectroscopy, gut mucosal tonometry and sublingual sidestream dark-
fi eld for tissue perfusion monitoring. Only four ICUs had transcutaneous
PO2. However, measuring the level of lactate as one of the tissue
perfusion markers was routinely performed in about 50% of the ICUs. Conclusion There were variations in monitoring performance in Thai
ICUs. These vary by type of hospital. Academic ICUs had a tendency for
advance monitoring in overall aspects. Establishing a critical care echocardiography laboratory
K Yastrebov1, A McLean2 Establishing a critical care echocardiography laboratory
K Yastrebov1, A McLean2
1The St George Hospital, Sydney, Australia; 2Nepean Hospital, Sydney, Australia
Critical Care 2013, 17(Suppl 2):P184 (doi: 10.1186/cc12122) Introduction Echocardiography is increasingly utilized by inten sive care
physicians in everyday practice. Standardization of echocardiographic
studies and reporting, quality assurance and medicolegal requirements
necessitate establishment of a dedicated system within the critical
care setting. We describe the process of setting up a critical care
echocardiography (CCE) laboratory based on our experience from
three separate ICUs. Figure 2 (abstract P182). Volume status, valve, sepsis according to the
Likert scale. Methods A retrospective review and analysis of the process involved in
establishment of echocardiography laboratories within ICUs. Methods A retrospective review and analysis of the process involved in
establishment of echocardiography laboratories within ICUs. Results Creating a CCE service involves a number of stages and
takes several years to achieve. Major components include staffi ng,
equipment, quality control, study archiving and networking capability. For staffi ng the objective is to identify and recruit staff with adequate
training and expertise in CCE, providing 24/7 specialist cover in
addition to supporting and training junior medical and nursing staff . There is further a need to acquire funding for high-quality ultrasound
machines and related hardware as well as long-term DICOM-based
archiving and reporting systems. This should be based on projections
of annual volumes of echo studies and corresponding digital storage. Networking connectivity is highly desirable, including obligatory
back-up solutions and site allocations. A business case incorporating
all the above should precede any development as identifi able funding
sources and administrative approval are essential. The implementation
stage requires the presence of a project leader who can organize the
trialing of scanners, archiving, reporting and research systems, ensure
compatibility with existing hospital and cardiology networks, and who
can assist in individualizing archiving and reporting software refl ecting
institutional and ICU specifi cs. Coordination with the IT department
is very important. Clear contractual vendor obligations for service,
maintenance and future upgrades of hardware and software need to
be specifi ed. Training and credentialing of staff is best achieved within
a systematic framework that includes ongoing competency review,
education and QA programs. Partnership with cardiology may benefi t
both groups. Major pitfalls are associated with poor initial training, lack
of expertise and leadership, and bad vendor contracts. Cardiopulmonary monitoring in Thai ICUs: results of ICU-RESOURCE
I surveys K Chittawatanarat1, A Wattanatham2, D Sathaworn2, C Permpikul3,
TSCCM Study Group4
1Chiang Mai University, Chiang Mai, Thailand; 2Pramongkudklao Hospital,
Bangkok, Thailand; 3Siriraj Hospital, Mahidol University, Bangkok, Thailand;
4Thai Society of Critical Care Medicine, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P183 (doi: 10.1186/cc12121) Introduction Although rapid progress in ICU monitoring with advanced
equipments has been developed, there were limited data on ICU
monitoring in Thailand. The objective of this study was to determine
the current utilization of monitoring in Thai ICUs. g
Methods A self-administered questionnaire was developed by the
TSCCM research subcommittee. Data verifi cation was processed by an
online medical research tools program (OMERET). p
g
Results A total of 350 questionnaires were sent to ICUs throughout
Thailand. In total, 256 questionnaires were confi rmed after being
received at the end of June 2012. Of these, 140 fi lled forms (56.9%)
were returned for fi nal analysis. More than 70% of the ICUs had basic
hemodynamic monitoring. Less than 10% of general and regional
hospitals could perform cardiac output monitoring by thermodilution
technique compared with 60% of academic teaching hospitals. New and advanced hemodynamic monitoring techniques such as
pulse pressure variation, systolic pressure variation, stroke volume
variation, PiCCO, Vigileo-Flo Tract, Pleth variability index device and
echocardiography were available only in ICUs of academic teaching
hospitals except ultrasound-based techniques including transthoracic
and transesophageal echocardiography and USCOM. For respiratory
monitoring, all ICUs had a SpO2 monitoring device but only one-half of
them had end-tidal CO2 monitoring. Nearly 80% of ventilator support
in participating ICUs was capable of displaying graphic waveform
monitoring. Only 43.6% of participating ICUs had a ventilator machine
that could calculate lung mechanics data. Advanced respiratory
monitoring such as EIT and esophageal pressure monitoring are
available only in ICUs of academic teaching hospitals. There was no Conclusion Establishment of a CCE laboratory requires careful
planning, and allocation of adequate human and fi nancial resources. Many potential problems can be identifi ed and prevented in advance. Strong expert leadership plays an important role. P185 Establishing a critical care echocardiography laboratory
K Yastrebov1, A McLean2 Conclusion Our results suggest that having continuously available
TEE for monitoring and management of hemodynamically unstable
patients increases awareness of right ventricular dysfunction in the ICU. References Conclusion Our results suggest that having continuously available
TEE for monitoring and management of hemodynamically unstable
patients increases awareness of right ventricular dysfunction in the ICU. References 1. Vieillard-Baron A, et al.: Intensive Care Med 2004, 30:1734-1739. 2. Monnet X, et al.: Intensive Care Med 2005, 31:1195-1201. 1. Vieillard-Baron A, et al.: Intensive Care Med 2004, 30:1734-1739. 2. Monnet X, et al.: Intensive Care Med 2005, 31:1195-1201. P181 Some advance monitoring used
in developed countries is also unavailable in Thailand. P187l P187
Infl uence of positive end-expiratory pressure on four-chamber
longitudinal strain analysis by speckle tracking echocardiography
F Franchi, A Faltoni, M Cameli, S Cecchini, M Lisi, M Contorni, S Mondillo,
S Scolletta
University of Siena, Italy
Critical Care 2013, 17(Suppl 2):P187 (doi: 10.1186/cc12125) Introduction Speckle-tracking echocardiography (STE) has emerged as
an ultrasound technique for accurately evaluating myocardial function
also in critically ill patients. By tracking the displacement of the speckles
during the cardiac cycle, the strain rate can be measured offl ine after
adequate image acquisition. The aim of the study was to evaluate the
eff ects of the positive end-expiratory pressure (PEEP) on four-chamber
longitudinal strain (LS) analysis in critically ill patients. g
y
y
p
Methods We enrolled 20 consecutive patients (mean age 64 ± 18) who
needed mechanical ventilation and were admitted to the ICU due to
heterogeneous causes. Inclusion criteria were: hypoxia requiring PEEP
titration, invasive arterial pressure monitoring, age >18. Exclusion
criteria were: myocardial dysfunction, cardiac arrhythmias and
valvular pathologies. The same operator performed three standard
echocardiography measurements (MyLab 70 Xvision; Esaote), each
of them after having increased PEEP at 5, 10, and 15 cmH2O (T1, T2,
T3, respectively). Blood gas analysis, respiratory, and hemodynamic
parameters provided by a pulse contour method were also recorded. STE analysis was performed offl ine (XStrain™MyLab 70 Xvision; Esaote). Results Left peak atrial LS (LA-PALS) was signifi cantly reduced from
T1 to T2, and from T2 to T3 (40.2 ± 12%, 35.9 ± 9%, 28.4 ± 8%, T1,
T2, T3, respectively; P <0.05). Right peak atrial LS (RA-PALS) and
right ventricular (RV)-LS showed a signifi cant reduction only at T3
(RA-PALS: 44.7 ± 48.5% at T1, 35.9 ± 11% at T3; RV-LS: –20.2 ± 2% at
T1, –16.3 ± 1.1% at T3; P <0.05). Left ventricular (LV)-LS did not change
signifi cantly during titration of PEEP. Cardiac chambers’ volumes and
cardiac output (CO) showed a signifi cant reduction at higher levels of
PEEP. Pulse pressure variation was signifi cantly aff ected by higher levels
of PEEP (P <0.05). Conclusion Contrast-enhanced ultrasonography clearly improves
visualization of the perfusion in various tissues. It is very likely to be
superior to standard Doppler ultrasound, and is safe and well tolerated
in critically ill patients. P185
Contrast-enhanced ultrasonography in the ICU: promising tool or
exciting toy?i Contrast-enhanced ultrasonography in the ICU: promising tool or
exciting toy?i Contrast-enhanced ultrasonography in the ICU: promising tool or
exciting toy? I Göcze, R Herzog, BM Graf, A Agha, HJ Schlitt, K Pfi ster, E Jung, T Bein
University Medical Centre Regensburg, Germany
Critical Care 2013, 17(Suppl 2):P185 (doi: 10.1186/cc12123) I Göcze, R Herzog, BM Graf, A Agha, HJ Schlitt, K Pfi ster, E Jung, T Bein
University Medical Centre Regensburg, Germany
Critical Care 2013, 17(Suppl 2):P185 (doi: 10.1186/cc12123) Introduction Contrast-enhanced ultrasonography (CEUS) is a
dynamic digital ultrasound-based imaging technique, which allows
quantifi cation of the microvascularisation up to the capillary vessels. As
a novel method for assessment of tissue perfusion it is ideally designed
for use in the ICU. CEUS is cost-eff ective and safe and can be repeatedly
performed at the bedside without radiation and nephrotoxicity. S70 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods The frequency of CEUS use in the multidisciplinary surgical
ICU was retrospectively evaluated for the period from 1 September
2011 to 1 September 2012. Furthermore, contributions of this novel
method to the management of critically ill ICU patients as well as its
accuracy were assessed. References Krishnagopalan S, Kumar A, Parrillo JE, Kumar A: Myocardial dysfunction in
the patient with sepsis. Curr Opin Crit Care 2002, 8:376-388. 2. Parker MM, McCarthy KE, Ognibene FP, et al.: Right ventricular dysfunction
and dilatation, similar to left ventricular changes,characterize the cardiac
depression of septic shock in humans. Chest 1999, 97:126-131. y
Results In total, 33 CEUS studies were performed in critically ill ICU
patients. The most frequent indications included: assessment of the
liver perfusion, assessment of the pancreas and kidney perfusion after
pancreas and kidney transplantation, assessment of the renal perfusion
in acute kidney injury (AKI), assessment of active bleeding and
assessment of the bowel perfusion. In all studies, the correct diagnosis
was achieved and the transport of critically ill patients to the radiology
department for further diagnostic procedures as well as application
of iodinated contrast agents was avoided. In 16 cases signifi cant new
fi ndings were detected. Twelve of them were missed by conventional
standard Doppler ultrasound prior to CEUS. In assessment of seven
cases with AKI, impaired or delayed perfusion and microcirculation
of the kidney was observed in six patients. In three patients urgent
surgical intervention was performed because of CEUS results. In three
cases active bleeding was excluded at the bedside due to absence of
contrast agent extravasation into hematoma (thigh and perihepatic) or
into abdominal cavity, without need for complementary CT imaging or
angiography. In one case the regular perfusion of intestinal anastomosis
was confi rmed with no need for surgical exploration. None of patients
undergoing CEUS manifested any adverse reactions or developed any
complications associated with the imaging technique. P186 Hemodynamic disclosure of septic shock patients by intensive care
ultrasound
P Theerawit, Y Sutherasan, T Hongpanat
Ramathibodi Hospital, Mahidol University, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P186 (doi: 10.1186/cc12124) Hemodynamic disclosure of septic shock patients by intensive care
ultrasound P Theerawit, Y Sutherasan, T Hongpanat P Theerawit, Y Sutherasan, T Hongpanat
Ramathibodi Hospital, Mahidol University, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P186 (doi: 10.1186/cc12124) Introduction Even though invasive hemodynamic devices are usually
used for assessment of septic shock victims, they cannot evaluate the
heart function. LV dysfunction as well as right heart syndrome are not
uncommon in sepsis and critical patients. Intensive care ultrasound
discloses these data and leads to appropriate treatment. Conclusion In hypoxic patients with normal cardiac function, PEEP
titration determined a reduction of LA-LS, RA-LS and RV-LS values. LV-LS values were not infl uenced by PEEP changes. The fall in CO,
observed with higher values of PEEP, seemed to be related to the
impairment of preload and not of myocardial contractility. Whenever
interpreting data on cardiac function obtained with longitudinal strain
analysis, attention of the clinician should be drawn to diff erent levels
of PEEP. The higher the PEEP, the more the probability of misleading
interpretation of STE data. Methods The study was a prospective cross-sectional study. The
measurement was performed within 24 hours of ICU admission. We
excluded patients with history of COPD and pulmonary hypertension
from any diseases. Only good-quality images acquired from subjects
were included for analysis. The primary objective was to disclose how
the hemodynamic changed in septic patients by ICU-US. y
g
p
p
y
Results A total of 133 septic patients were measured by ICU-US. Good
image quality was acquired in 115 cases (86.47%). The mean ages were
57.48 ± 17.87 years. The three major causes of sepsis were pneumonia,
unknown source, abdominal infection and bacteremia. Heart failure at
admission was found only in 1.79%. Previous history of hypertension,
DM, and coronary artery disease was found in 12.17%, 11.30%, and
1.74% of patients. The mean LV ejection fraction (LVEF) was 54 ± 15.43%. The percentages of patients with LVEF <35%, 35 to 40%, 41 to 45%
and >45% were 13.2%, 7.5%, 8.5%, and 70.8% respectively. Diastolic
dysfunction defi ned by E/A ratio <1 was observed in 47.5% of patients. A total 44.6% of cases had cardiac output under 4 l/minute whereas
CO over 6 l/minute was found in 18.1% of cases. The average mean
pulmonary artery pressure was 34.75 ± 15.13 mmHg. The proportion of
patients with meanPAP over 25 mmHg was 76.2%. RV to LV ratio >1 was
found in 42.4% of septic patients. P187l Promising indications for the use of CEUS in the
ICU may be the assessment of kidney microcirculation and assessment
of liver perfusion in liver transplant and liver trauma patients. P186 P189 Validation of less-invasive hemodynamic monitoring with Pulsiofl ex
in critically ill patients: interim results of a multicentre study
K Van de Vijver1, C Pigozzi1, L Vervliet1, V Vanbiervliet1, V Brabers1, I Vos1,
H Maes1, N Van Regenmortel1, I De laet1, K Schoonheydt1, H Dits1, J Belda2,
Z Molnar3, M Malbrain1
1Ziekenhuis Netwerk Antwerpen, ZNA Stuivenberg, Antwerp, Belgium;
2Hospital Clinico Universitario, Valencia, Spain; 3University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P189 (doi: 10.1186/cc12127) Results A total of 103 measurements in 77 patients were performed. In seven patients measurement with Nexfi n was not possible. For CO
(55 paired measurements), values were 6 ± 2.1 l/minute (range 2.6
to 12). Pearson’s correlation coeffi cient comparing Nexfi n-CO with
reference CO showed a good correlation (R2 = 0.52). Bland–Altman
analysis comparing both CO techniques revealed a mean bias ± 2SD
(LA) of 0.3 ± 3.6 l/minute (58% error). The MAP was 84.2 ± 15.6 mmHg
(53 to 131.5) and values obtained with the Nexfi n correlated well
with the reference method with an R2 of 0.72. Bland–Altman analysis
comparing both MAP techniques revealed a mean bias ± 2SD (LA) of
–0.3 ± 18 mmHg (20.9% error). However, Nexfi n-MAP did not correlate
well with NIBP (R2 = 0.36). Hemoglobin values obtained with Nexfi n
Massimo technique did not correlate well with laboratory values
(R2 = 0.26, 33% error). The 26 patients that died in the ICU had higher
APACHE II (P = 0.017), SAPS II (P <0.0001), SOFA (P <0.0001) and SOS
(P = 0.004) scores and signifi cantly lower MAP (P <0.0001), hemoglobin
(P = 0.01) and lower dp/dtmax (P = 0.003), a marker for contractility. There were no outcome diff erences with regard to subgroup analysis in
patients with either low or high CO or SVR. Introduction Thermodilution (TD) is considered a gold standard for
measurement of cardiac index (CI) in critically ill patients. The aim of
this study is to compare intermittent bolus TD CI with intermittent
automatic calibration CI (AutoCI) and two continuous CIs obtained by
pulse contour analysis with PiCCO2 (PiCCI) and Pulsiofl ex (PuCCI). 2l
Methods Interim results of an ongoing prospective multicentre study
in 53 patients. Age 58.7 ± 15.4, SAPS II score 51.4 ± 14.7 and SOFA score
10 ± 3.2. P188 Evaluation of a new calibration index suggesting recalibration of
the pulse contour cardiac index by transpulmonary thermodilution:
a prospective study
W Huber, K Waldleitner, S Mair, B Saugel, RM Schmid
Klinikum rechts der Isar, Technical University of Munich, Germany
Critical Care 2013, 17(Suppl 2):P188 (doi: 10.1186/cc12126) Introduction After calibration by thermodilution (TD), the PiCCO
device is able to assess cardiac index (CI) using pulse contour (PC)
analysis. The manufacturer suggests recalibration by TD after 8 hours. Recently, we suggested a calibration index indicating a certain
probability of a relevant bias and triggering the next calibration. With
changes of CIpc compared with the previous CItd being a key predictor
of the bias of CIpc compared with the following CItd, the manufacturer
implemented a soft alarm indicating changes in CIpc compared with
the last CItd (thresholds adjustable to 15%, 25% and 35%). The aim of Conclusion Cardiac dysfunction, namely left ventricle and probably
right ventricle, was not uncommon in septic shock patients. Without
intensive care ultrasound, all crucial information was delayed until
patient deterioration and initial treatment may be harmful. Thus
cardiac ultrasound should be used initially to disclose hemodynamic
features before routine resuscitation is initiated. S71 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Results In total, 940 CCI and 382 TDCI values were obtained: 940 paired
PiCCI and PuCCI; 358 paired AutoCI-TDCI measurements. TDCI values
ranged from 1.5 to 6.9 l/minute/m2 (mean 3.6 ± 1.1), AutoCI from 1.8
to 7.2 (3.6 ± 0.9), PiCCI from 1.0 to 7.1 (3.5 ± 1.1) and PuCCI from 1.3 to
7.6 (3.6 ± 1). Pearson’s correlation coeffi cient comparing mean PuCCI
and PiCCI values per patient had an R2 of 0.79. Comparison between
AutoCI and TDCI had an R2 of 0.51. Changes in AutoCI correlated well
with changes in TDCI (R2 = 0.44, concordance coeffi cient = 95.7), as
did changes in PuCCI versus changes in PiCCI (R2 = 0.99, CC = 93.4%). Changes in PiCCI and PuCCI induced by an intervention correlated
well with each other (R2 = 0.86, CC = 100%). The percentage error
(PE) obtained by Bland and Altman analysis and R2 for the diff erent
comparisons are presented in Table 1. the study was prospective evaluation of predictive capabilities of the
15% alarm regarding a bias of CIpc compared with CItd exceeding
critical thresholds of 15%, 20% and 25%. P190 P190 P190
Interim results of an ongoing study on the use of non-invasive
hemodynamic monitoring with Nexfi n in critically ill patients
K Van de Vijver, V Brabers, C Pigozzi, L Vervliet, V Vanbiervliet, H Maes,
I Vos, M Peetermans, N Van Regenmortel, I De laet, K Schoonheydt, H Dits,
M Malbrain
Ziekenhuis Netwerk Antwerpen, ZNA Stuivenberg, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P190 (doi: 10.1186/cc12128) Interim results of an ongoing study on the use of non-invasive
hemodynamic monitoring with Nexfi n in critically ill patients
K Van de Vijver, V Brabers, C Pigozzi, L Vervliet, V Vanbiervliet, H Maes,
I Vos, M Peetermans, N Van Regenmortel, I De laet, K Schoonheydt, H Dits,
M Malbrain Ziekenhuis Netwerk Antwerpen, ZNA Stuivenberg, Antwerp, Belgium
Critical Care 2013, 17(Suppl 2):P190 (doi: 10.1186/cc12128) Conclusion Bias of CIpc compared with the next CItd does not depend
on the time since last CItd. Changes in CIpc itself compared with the
last CItd are associated with the bias. A 15% change in CIpc trend alarm
provided by the new PiCCO algorithm is signifi cantly associated with
bias values exceeding 15%, 20% or 25% of CItd, whereas time to last
calibration >8 hours is not associated with bias exceeding these critical
thresholds. Introduction The Nexfi n monitor (BMEYE, the Netherlands) enables
continuous non-invasive analysis of blood pressure (MAP) as well as
cardiac output (CO) measurements. The aim of the present study was
to validate the Nexfi n in a mixed population of medical ICU patients. Introduction The Nexfi n monitor (BMEYE, the Netherlands) enables
continuous non-invasive analysis of blood pressure (MAP) as well as
cardiac output (CO) measurements. The aim of the present study was
to validate the Nexfi n in a mixed population of medical ICU patients. Introduction The Nexfi n monitor (BMEYE, the Netherlands) enables
continuous non-invasive analysis of blood pressure (MAP) as well as
cardiac output (CO) measurements. The aim of the present study was
to validate the Nexfi n in a mixed population of medical ICU patients. Methods Interim results of a prospective ongoing study in 77 patients
admitted to the medical ICU (46 patients mechanically ventilated, M/F
ratio 1/1). Age 65.6 ± 15.9, BMI 25.6 ± 4.8, APACHE II score 22.9 ± 10.9,
SAPS II 48 ± 20.1, SOFA score 7.5 ± 4.5. A modifi ed outreach score (SOS)
was calculated on admission. P190 For all patients, simultaneous recording
of arterial pressure by radial line (n = 78), PiCCO (n = 44) or by NIBP with
arm cuff (n = 47) was compared with the Nexfi n monitor. Statistical
analysis was performed with Student’s t test, Pearson correlation and
Bland–Altman analysis. i
Methods Interim results of a prospective ongoing study in 77 patients
admitted to the medical ICU (46 patients mechanically ventilated, M/F
ratio 1/1). Age 65.6 ± 15.9, BMI 25.6 ± 4.8, APACHE II score 22.9 ± 10.9,
SAPS II 48 ± 20.1, SOFA score 7.5 ± 4.5. A modifi ed outreach score (SOS)
was calculated on admission. For all patients, simultaneous recording
of arterial pressure by radial line (n = 78), PiCCO (n = 44) or by NIBP with
arm cuff (n = 47) was compared with the Nexfi n monitor. Statistical
analysis was performed with Student’s t test, Pearson correlation and
Bland–Altman analysis. P188 the study was prospective evaluation of predictive capabilities of the
15% alarm regarding a bias of CIpc compared with CItd exceeding
critical thresholds of 15%, 20% and 25%. Methods A prospective analysis of 329 routine TPTD measurements
in 70 patients. The CI-trend alarm was set to 15%, and CIpc, trend
alarm (yes/no), relative and absolute changes in CIpc were recorded
immediately before TD providing exact measurement of CItd. Predictive
capabilities of the 15% trend alarm regarding the bias were evaluated
using Spearman correlation, chi-square test, ROC analysis and Wilcoxon
test (IBM SPSS 20). Results A total of 70 patients (24 female, 46 male), age 62 ± 14 years,
APACHE II score 19.5 ± 7.5, SOFA score 8.9 ± 4.4. At 937 ± 904 (50 to 5,795)
minutes after the last CItd, CIpc provided a bias of –0.0566 ± 0.761 l/
minute*m2 compared with the next CItd. Percentage error was 34.6%. Absolute and relative bias of CIpc compared with the new CItd did not
correlate to time to last TD, but correlated (all P <0.001) to CIpc itself
(r = 0.333; r = 0.385) and relative (r = 0.531; r = 0.537) and absolute
(r = 0.533; r = 0.529) changes in CIpc compared with last CItd. The 15%
CIpc trend alarm was indicated before 83/329 measurements (25.2%). The amount of bias exceeded 15% and 20% in 101 (30.7%) and 79
(24%) of TDs. In TDs with indicated trend alarm (≥15% deviation of CIpc
to the last CItd), the amount of bias more frequently exceeded 15%
(P = 0.019), 20% (P <0.001) and 25% (P <0.001). Time to last calibration
≥8 hours was not associated with bias exceeding 15% (P = 0.735), 20%
(P = 0.888) or 25% (P = 281). Conclusion The preliminary results indicate that in unstable critically
ill patients, CI can be reliably monitored with Pulsiofl ex technology via
a femoral line. Pulsiofl ex was also able to keep track of changes in CI. P191
Cost-eff ectiveness analysis of stroke volume variation guided
perioperative hemodynamic optimization
J Benes, J Zatloukal, A Simanova, I Chytra, E Kasal
The Medical Faculty and Hospital in Plzen – Charles University Prague, Plzen,
Czech Republic
Critical Care 2013, 17(Suppl 2):P191 (doi: 10.1186/cc12129) P191
Cost-eff ectiveness analysis of stroke volume variation guided
perioperative hemodynamic optimization
J Benes, J Zatloukal, A Simanova, I Chytra, E Kasal
The Medical Faculty and Hospital in Plzen – Charles University Prague, Plzen,
Czech Republic
Critical Care 2013, 17(Suppl 2):P191 (doi: 10.1186/cc12129) Introduction Perioperative goal-directed therapy (pGDT) can sub-
stantially improve the outcome of high-risk surgical patients [1]. But the
approach needs an initial investment and increases the staff workload. Economic factors might participate in the weak adherence to the
pGDT concept. Some model studies support pGDT cost-eff ectiveness,
but real economic data based on a recent clinical trial are lacking. We
performed an economic analysis of hemodynamic optimization using
the stroke volume variation trial [2] in order to elucidate this issue. Figure 1 (abstract P192). Correlation between PaO2/FiO2 and EVLWIc. Methods The hospital care invoices of all 120 patients included in the
trial were retrospectively extracted. Due to the nature of the data we
have adopted the healthcare payer’s perspective. We performed a
comparison of induced costs between the Vigileo (n = 60) and Control
(n = 60) groups and constructed a cost tree using the study group and
complications occurrence as distributive parameters. The incremental
cost-eff ectiveness ratio per complication avoided was calculated and,
fi nally, diff erent reimbursing categories were assessed as potential cost
drivers. (EVLWI) refl ects pulmonary edema [2]. The new EV1000/VolumeView
(Edwards Lifesciences) can accurately measure EVLWI corrected for the
actual volume of lung parenchyma (EVLWIc). The aim of our study is to
prove a stronger correlation between EVLWIc and PaO2/FiO2 compared
with EVLWI in patients undergoing pulmonary resection. Methods A prospective observational study. Seven patients with
lung cancer undergoing pulmonary resection were monitored using
the EV1000 plathform. EVLWI was assessed by thermodilution at
the following time points: after intubation (t1); during single-lung
ventilation (t2); after lung resection (t3); after ICU admission (t4);
12 hours (t5) and 18 hours after ICU admission (t6). EVLWIc values were
also collected at t3 and t4. PaO2/FiO2 was measured at the same time
points. Results A decreased rate (18 vs. 35 patients) and number of
complications (34 vs. 78) were observed in the original trials Vigileo
group. The mean cost of intervened patient was lower (€2,877 ± 2,336
vs. €3,371 ± 3,238; P = 0.38). According to the cost-tree analysis,
patients with complications (n = 53; 44%) consumed signifi cantly more
resources (€235,623; 63%). References 1. Pinsky MR: Clin Chest Med 2003, 24:549-560. 2. Sakka SG, et al.: Chest 2002, 122:2080-2086. 1. Pinsky MR: Clin Chest Med 2003, 24:549-560. 2. Sakka SG, et al.: Chest 2002, 122:2080-2086. Acknowledgements The study was supported by the MSM0021620819
and the Charles University Research Fund (project number P36). References P193 1. Hamilton MA, et al.: Systematic review and meta-analysis on the use of
preemptive hemodynamic intervention to improve postoperative
outcomes in moderate and high-risk surgical patients. Anesth Analg 2011,
112:1392-1402.l Goal-directed resuscitation therapy in high-risk patients
undergoing cardiac surgery (GRICS study): a randomized controlled
trial – preliminary results
E Osawa1, A Rhodes2, J Vincent3, J Almeida1, J Fukushima1, B Pileggi1,
C Park1, L Camara1, J Auler Jr1, R Chan1, M Piccioni1, M Lima1, F Galas1,
L Hajjar1
1Heart Institute, São Paulo, Brazil; 2St George’s Healthcare NHS Trust and St
George’s University of London, UK; 3Universite Libre de Bruxelles, Brussels,
Belgium
Critical Care 2013, 17(Suppl 2):P193 (doi: 10.1186/cc12131) Goal-directed resuscitation therapy in high-risk patients
undergoing cardiac surgery (GRICS study): a randomized controlled
trial – preliminary results P191
Cost-eff ectiveness analysis of stroke volume variation guided
perioperative hemodynamic optimization
J Benes, J Zatloukal, A Simanova, I Chytra, E Kasal
The Medical Faculty and Hospital in Plzen – Charles University Prague, Plzen,
Czech Republic
Critical Care 2013, 17(Suppl 2):P191 (doi: 10.1186/cc12129) A gain of €634 per avoided complications
confi rms that the lower complications rate was the most important cost
driver. Both the clinical care for patients costs (€505 vs. 912; P = 0.04)
and ward stay costs (€244 vs. 402; P = 0.03) were decreased by the
intervention. On the contrary, the intervention increased anaesthesia
costs (€880 vs. 688; P = 0.001). Results No signifi cant correlation was found between EVLWI and PaO2/
FiO2 (r = –0.3124, P >0.05), while a signifi cant correlation was seen
between EVLWIc and PaO2/FiO2 (r = –0.528, P =0.009; Figure 1). Conclusion Despite the small sample size, this study shows that in
patients undergoing pulmonary resection the EVLWIc is more strongly
correlated to PaO2/FiO2 than EVLWI. Therefore, the EV1000 may be
a valuable tool for more reliable hemodynamic monitoring in this
subgroup of patients. R f Results No signifi cant correlation was found between EVLWI and PaO2/
FiO2 (r = –0.3124, P >0.05), while a signifi cant correlation was seen
between EVLWIc and PaO2/FiO2 (r = –0.528, P =0.009; Figure 1). 2
2
g
Conclusion Despite the small sample size, this study shows that in
patients undergoing pulmonary resection the EVLWIc is more strongly
correlated to PaO2/FiO2 than EVLWI. Therefore, the EV1000 may be
a valuable tool for more reliable hemodynamic monitoring in this
subgroup of patients. Conclusion Intraoperative fl uid optimization with the use of stroke
volume variation and the Vigileo/FloTrac system showed not only a
substantial improvement of morbidity, but was also associated with an
economic benefi t. This observed benefi t highly exceed the increased
monitoring costs in our trial. P192 P192
EV1000/VolumeView: a new device for a more reliable
measurement of extravascular lung water index in patients with
lung resections
A Donati, C Melia, V Monaldi, R Domizi, E Damiani, A Carsetti, C Scorcella,
R Castagnani, P Pelaia
Università Politecnica delle Marche, Ancona, Italy
Critical Care 2013, 17(Suppl 2):P192 (doi: 10.1186/cc12130) P192
EV1000/VolumeView: a new device for a more reliable
measurement of extravascular lung water index in patients with
lung resections
A Donati, C Melia, V Monaldi, R Domizi, E Damiani, A Carsetti, C Scorcella,
R Castagnani, P Pelaia
Università Politecnica delle Marche, Ancona, Italy
Critical Care 2013, 17(Suppl 2):P192 (doi: 10.1186/cc12130) Goal-directed resuscitation therapy in high-risk patients
undergoing cardiac surgery (GRICS study): a randomized controlled
trial – preliminary results p
y
E Osawa1, A Rhodes2, J Vincent3, J Almeida1, J Fukushima1, B Pileggi1,
C Park1, L Camara1, J Auler Jr1, R Chan1, M Piccioni1, M Lima1, F Galas1,
L Hajjar1 2. Benes J, et al.: Intraoperative fl uid optimization using stroke volume
variation in high risk surgical patients: results of prospective randomized
study. Crit Care 2010, 14:R118. 2. Benes J, et al.: Intraoperative fl uid optimization using stroke volume
variation in high risk surgical patients: results of prospective randomized
study. Crit Care 2010, 14:R118. jj
1Heart Institute, São Paulo, Brazil; 2St George’s Healthcare NHS Trust and St
George’s University of London, UK; 3Universite Libre de Bruxelles, Brussels,
Belgium
l
l
(d
) P189 All patients underwent PiCCO monitoring via a femoral line
whilst a radial line was kept in place during four 8-hour time periods
(in the fi rst two periods, the Pulsiofl ex was connected to a radial line;
in the last two it was connected to a femoral line). In the fi rst and third
periods, the Pulsiofl ex was calibrated with TDCI, for the second and
fourth periods Pulsiofl ex was calibrated with AutoCI. Simultaneous
PiCCI and PuCCI measurements were obtained every 2 hours while
simultaneous TDCI and AutoCI were obtained every 8 hours. We also
looked at the eff ects of 40 interventions. p
g
Conclusion In unstable critically ill patients, MAP (and CO) can be
monitored with the Nexfi n. The exact patient population for this Table 1 (abstract P189). Results of Bland and Altman analysis
Pulsiofl ex
AutoCal
PE (CCl) (%)
n
R2
PE (TD-Pi) (%)
n
R2
PE (TD-Pu) (%)
n
R2
All
All
37.9
940
0.73
22.8
382
0.88
38.5
382
0.66
All
Yes
43.4
510
0.50
20.4
210
0.88
42.3
210
0.47
All
No
27.8
430
0.83
25.6
172
0.85
32.5
172
0.74
Fem
All
30.6
464
0.73
20.2
192
0.88
33.0
192
0.66
Rad
All
44.2
476
0.58
25.2
190
0.85
43.7
190
0.56 Table 1 (abstract P189). Results of Bland and Altman analysis S72 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Figure 1 (abstract P192). Correlation between PaO2/FiO2 and EVLWIc. technology has yet to be defi ned and more patients are probably
needed for pattern recognition, although the results indicate that low
MAP and dp/dtmax are associated with poor outcome. In the future,
Nexfi n data could theoretically be incorporated in a new outreach
score. P191 Treatment-related cardiac complications associated with
goal-directed therapy in high-risk surgical patients: a meta-analysis
N Arulkumaran, C Corredor, M Hamilton, M Grounds, J Ball, A Rhodes,
M Cecconi Treatment-related cardiac complications associated with
goal-directed therapy in high-risk surgical patients: a meta-analysis
N Arulkumaran, C Corredor, M Hamilton, M Grounds, J Ball, A Rhodes,
M Cecconi
St George’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P195 (doi: 10.1186/cc12133) Treatment-related cardiac complications associated with
goal-directed therapy in high-risk surgical patients: a meta-analysis
N Arulkumaran, C Corredor, M Hamilton, M Grounds, J Ball, A Rhodes,
M Cecconi St George’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P195 (doi: 10.1186/cc12133) Introduction We hypothesized that goal-directed therapy (GDT) is not
associated with an increased risk of cardiac complications in high-risk,
noncardiac surgical patients. Patients with limited cardiopulmonary
reserve are at risk of mortality and morbidity after major surgery [1]. Augmentation of the oxygen delivery index (DO2I) with a combination
of intravenous fl uids and inotropes (GDT) has been shown to reduce
the postoperative mortality and morbidity in high-risk patients [2]. However, concerns regarding cardiac complications associated with
fl uid challenges and inotropes used to augment cardiac output may
deter clinicians from instituting early GDT in the very patients who are
more likely to benefi t. Results Twenty patients from the GDT group were compared with 20
control patients. Both groups were comparable concerning baseline
characteristics and severity scores, except for a higher prevalence of
hypertension and heart failure in the GDT group. Intraoperative data
showed no diff erence regarding length of extracorporeal circulation,
fl uid balance, transfusion or inotropic requirement. Patients from the
GDT group were given more fl uids within the fi rst 8 hours as compared
with the conventional group (1,250 ml vs. 500 ml, P <0.001). GDT
patients showed a median ICU stay of 3 days (95% CI: 3 to 4) compared
with 5 days in control patients (95% CI: 4 to 7). Moreover, hospital stay
was less prolonged in GDT patients (10 days vs. 14 days, P = 0.043). Inotropic dependence was lower in the GDT group (29 hours vs. 55 hours, P = 0.003) as well as the cumulative dobutamine dosage (8 vs. 19 μg/kg/day, P = 0.025). Also, GDT group presented a lower incidence
of infections, tachyarrithmias and acute renal failure (RIFLE R) when
compared with the control group. yi
Methods Systematic search of MEDLINE, Embase and CENTRAL
databases for randomized controlled trials of GDT in high-risk surgical
patients. Studies including cardiac surgery, trauma, and pediatric
surgery were excluded to minimize heterogeneity. References 1. Pearse RM, et al.: Lancet 2012, 380:1059-1065. 2. Hamilton MA, et al.: Anesth Analg 2011, 112:1392-1402. 2. Hamilton MA, et al.: Anesth Analg 2011, 112:1392-1402. Results In the group treated with a restrictive volume approach,
patients received fl uids at the rate of 7.0 ± 1.0 ml/kg/hour. PaO2/FiO2
was 288 ± 14 after intubation and 270 ± 22 before extubation. In the
group treated with a liberal volume approach, fl uids were replaced
at 11.0 ± 2.0 ml/kg/hour. PaO2/FiO2 was 259 ± 24 after intubation and
223 ± 43 before extubation. Surgery combined with OLV was found to
signifi cantly aff ect the PaO2/FiO2 value (ANOVA, F1,14 = 15.85a, P = 0.001,
partial η2 = 0.531). The average PaO2/FiO2 level was signifi cantly higher
in the restrictive-replacement group than in the liberal-replacement
group (ANOVA, F1,14 = 9.66, P = 0.008, partial η2 = 0.408). There was
no interaction between the groups (ANOVA, F1,14 = 1.7a, P = 0.215,
partial η2 =0.108). Mean length of stay in the ICU was similar between
the restrictive-replacement group (5.2 ± 2.3 days) and the liberal-
replacement group (6.3 ± 1.6 days) (ANOVA, F1,14 = 0.814a, P = 0.382,
partial η2 = 0.055). Perioperative volume management for esophageal cancer surgery
M Karaman Ilic, G Madžarac, J Kogler, D Stančić Rokotov, N Hodoba
University Hospital Centre Zagreb, Croatia
Critical Care 2013, 17(Suppl 2):P194 (doi: 10.1186/cc12132) Perioperative volume management for esophageal cancer surgery
M Karaman Ilic, G Madžarac, J Kogler, D Stančić Rokotov, N Hodoba
University Hospital Centre Zagreb, Croatia
Critical Care 2013, 17(Suppl 2):P194 (doi: 10.1186/cc12132) Introduction Pulmonary complications are the primary reason for
extending a patient’s stay in the ICU. The aim of this study was to
investigate whether perioperative volume management can infl uence
the PaO2/FiO2 value and total length of stay in the ICU. Conclusion Perioperative, physiologically guided, GDT in high-risk
surgical patients is not associated and actually reduces postoperative
cardiovascular complications. 2
2
Methods Sixteen patients were divided into two groups: one group
was treated with a restrictive approach (≤8 ml/kg/hour), and the other
with a liberal approach (> 8 ml/kg/hour). Patients were randomly
allocated using sealed envelopes. During the thoracic part of the
surgical procedure, all patients received one-lung ventilation (OLV). P195 or extracardiac arteriopathy) were allocated to GDT or conventional
hemodynamic therapy. We excluded patients with endocarditis,
previous use of dobutamine, need for IABP, high dose of vasopressors
and emergency surgery. The GDT protocol involved hemodynamic
resuscitation aimed at a target of a cardiac index >3 l/minute/m2
through a three-step approach: fl uid therapy of 250 ml lactated Ringer’s
solution, dobutamine infusion up to a dose of 20 μg/kg/minute, and
red blood cell transfusion to reach a hematocrit level above 28%. EV1000/VolumeView: a new device for a more reliable
measurement of extravascular lung water index in patients with
lung resections g
Critical Care 2013, 17(Suppl 2):P193 (doi: 10.1186/cc12131) g
Critical Care 2013, 17(Suppl 2):P193 (doi: 10.1186/cc12131) Introduction A goal-directed resuscitation therapy (GDT) through
optimization of cardiac output reduces complications in noncardiac
surgeries. We investigated whether the implementation of a GDT
protocol in high-risk cardiac surgery with the use of LiDCO Rapid
reduces postoperative complications as compared with the standard
of care. Introduction A goal-directed resuscitation therapy (GDT) through
optimization of cardiac output reduces complications in noncardiac
surgeries. We investigated whether the implementation of a GDT
protocol in high-risk cardiac surgery with the use of LiDCO Rapid
reduces postoperative complications as compared with the standard
of care. A Donati, C Melia, V Monaldi, R Domizi, E Damiani, A Carsetti, C Scorcella,
R Castagnani, P Pelaia
Università Politecnica delle Marche, Ancona, Italy
Critical Care 2013, 17(Suppl 2):P192 (doi: 10.1186/cc12130) Introduction Hemodynamic monitoring is important in high-risk
surgical patients in order to detect and correct circulatory instability,
thereby improving outcome [1]. The extravascular lung water index Methods We performed a prospective and randomized study whereby
consecutive patients fulfi lling one high-risk criteria (EuroSCORE >5,
ejection fraction <50%, recent myocardial infarction, unstable angina S73 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 1.
Nisanevich V, Felsenstein I, Almogy G, Weissman C, Einav S, Matot I: Eff ect of
intraoperative fl uid management on outcome after intraabdominal
surgery. Anesthesiology 2005, 103:25-32.l 1.
Nisanevich V, Felsenstein I, Almogy G, Weissman C, Einav S, Matot I: Eff ect of
intraoperative fl uid management on outcome after intraabdominal
surgery. Anesthesiology 2005, 103:25-32.
2.
Wei S, Tian J, Song X, Chen Y: Association of perioperative fl uid balance and
adverse surgical outcomes in esophageal cancer and esophagogastric
junction cancer. Ann Thorac Surg 2008, 86:266-272. Treatment-related cardiac complications associated with
goal-directed therapy in high-risk surgical patients: a meta-analysis
N Arulkumaran, C Corredor, M Hamilton, M Grounds, J Ball, A Rhodes,
M Cecconi We reviewed the
rates of all cardiac complications, arrhythmias, acute myocardial
ischemia, and acute pulmonary edema. Meta-analyses were performed
and forest plots drawn using RevMan software. Data are presented as
odd ratios (ORs) (95% CIs), and P values. Conclusion A hemodynamic therapy tailored to an optimized cardiac
output reduced the length of ICU stay, vasoactive drug requirement
and postoperative complications. Results We identifi ed 23 randomized controlled trials including 2,219
patients, who reported cardiac complications. GDT was associated
with a reduction in total cardiovascular complications (OR = 0.55 (0.39
to 0.78), P = 0.0007), and with a signifi cantly reduced incidence of
arrhythmias (OR = 0.59 (0.38 to 0.91), P = 0.02). GDT was not associated
with an increase in acute pulmonary edema (OR = 0.68 (0.42 to 1.10),
P = 0.11) or acute myocardial ischemia (OR = 0.70 (0.38 to 1.27), P = 0.23). Subgroup analysis of overall cardiovascular complications revealed that
the benefi t is most pronounced in patients receiving fl uid and inotrope
therapy (OR = 0.55 (0.34 to 0.89), P = 0.01) to achieve a supranormal
oxygen delivery target (OR = 0.50 (0.32 to 0.79), P = 0.003), guided by
the use of minimally invasive cardiac outmonitoring (OR = 0.50 (0.33 to
0.77), P = 0.002). p
p
p
Acknowledgement Grants received from FAPESP 2011/19987-5. Acknowledgement Grants received from FAPESP 2011/19987-5. Estimation of potential cost-savings related to the implementation
of perioperative hemodynamic goal-directed therapy COVIG (T1)
0.31 (CI 0.24 to 1.63)
–0.49
–3.41 to 2.43
COECHO vs. COMC (T1)
0.71 (CI 0.56 to 1.08)
–0.055
–1.19 to 1.08
COECHO vs. COVIG (T2)
0.27 (CI 0.16 to 1.53)
–0.49
–3.24 to 2.34
COECHO vs. COMC (T2)
0.79 (CI 0.66 to 1.107)
–0.023
–0.94 to 0.9
CI, confi dence interval; LoA, limits of agreement. between 0.35 and 0.55. Importantly, these odd ratios were not
related to the morbidity rates. In the US publication [2], extra costs for
treating patients with 1+ complication were $17,949. In the Swiss (CH)
publication [3], they were $34,446. See Table 1. y
Conclusion Depending on the pre-implementation morbidity rate,
the degree of pGDT-induced morbidity reduction and the country,
cost-savings ranged between $808 and $13,434 per patient. This large
variability suggests that local/hospital estimations are desirable before
starting pGDT implementation. These tailored evaluations would also
allow more precise cost-saving estimations by taking into account the
actual and expected pGDT compliance rates. and compared with those calculated with the echocardiographic
standard formulation (stroke volume = cross-sectional area×velocity
time integral; COECHO = SV×heart rate). In every patient CO was
measured twice: at baseline (T1) and after volume loading (500 ml
lactate Ringer solution) (T2). Agreements between COVIG, COMC, and
COECHO were evaluated by means of simple linear regression (r2) and
Bland–Altman analysis. and compared with those calculated with the echocardiographic
standard formulation (stroke volume = cross-sectional area×velocity
time integral; COECHO = SV×heart rate). In every patient CO was
measured twice: at baseline (T1) and after volume loading (500 ml
lactate Ringer solution) (T2). Agreements between COVIG, COMC, and
COECHO were evaluated by means of simple linear regression (r2) and
Bland–Altman analysis. 1. Hamilton MA, et al.: A systematic review and meta-analysis on the use of
preemptive hemodynamic intervention to improve postoperative
outcomes in moderate and high-risk surgical patients. Anesth Analg 2011,
112:1392-1402. 2. Boltz et al.: Synergistic implications of multiple postoperative outcomes. Am J Med Qual 2012, in press. 3. Vonlanthen R, et al.: The impact of complications on costs of major surgical
procedures. Ann Surg 2011, 254:907-913. 3. Vonlanthen R, et al.: The impact of complications on costs of major surgical
procedures. Ann Surg 2011, 254:907-913. y
Results Twenty patients were enrolled in the study. Values of r2, bias
and limit of agreement at T1 and T2 are summarized in Table 1. Estimation of potential cost-savings related to the implementation
of perioperative hemodynamic goal-directed therapy p
p
T Simon, G Marx RWTH University Hospital Aachen, Germany y
y
Critical Care 2013, 17(Suppl 2):P196 (doi: 10.1186/cc12134) Introduction Many studies have demonstrated the ability of peri-
operative hemodynamic goal-directed therapy (pGDT) to decrease
post operative morbidity in patients undergoing medium-to-high-risk
surgery [1]. As a result, pGDT may be a cost-saving strategy. Our goal
was to provide an estimation of potential cost-savings based on recent
literature. Methods The largest and most recent meta-analysis [1] on pGDT
was used to estimate what could be the reduction of postoperative
morbidity if pGDT was to be adopted. Costs related to the treatment
of patients developing at least one (1+) postoperative complication
were obtained from two recent US [2] and Swiss [3] publications. Potential cost-savings related to the adoption of pGDT were calculated
according to the actual morbidity rate, assuming 0% pGDT use so far,
and 100% compliance rate after implementation. Conclusion Results from this small sample indicate that esophageal
carcinoma surgery by itself had a detrimental eff ect on the PaO2/FiO2
value, which restriction of perioperative volume did not signifi cantly
aff ect. Volume restriction also did not aff ect length of stay in the ICU. References 1. Nisanevich V, Felsenstein I, Almogy G, Weissman C, Einav S, Matot I: Eff ect of
intraoperative fl uid management on outcome after intraabdominal
surgery. Anesthesiology 2005, 103:25-32.l Results The 2011 meta-analysis [1] of 29 RCTs (4,805 patients) showed
that pGDT is associated with a reduction in the rate of patients
developing 1+ postoperative complications with odd ratios ranging 2. Wei S, Tian J, Song X, Chen Y: Association of perioperative fl uid balance and
adverse surgical outcomes in esophageal cancer and esophagogastric
junction cancer. Ann Thorac Surg 2008, 86:266-272. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S74 Actual morbidity rate no pGDT (%)
10
20
30
40
50
60
Expected morbidity rate pGDT (%)
3.5 to 5.5
7 to 11
11 to 17
14 to 22
18 to 28
21 to 33
Expected cost reduction/US patient ($)
808 to 1,167
1,615 to 2,333
2,423 to 3,500
3,231 to 4,667
4,039 to 5,833
4,846 to 7,000
Expected cost reduction/CH patient ($)
1,550 to 2,239
3,100 to 4,478
4,478 to 6,545
6,200 to 8,956
7,578 to 11,023
9,300 to 13,434 Table 1 (abstract P197)
Pearson r2
Bias
LoA
COECHO vs. Cardiac output estimation: Vigileo and Mostcare versus
echocardiography S Romagnoli, D Quattrone, AR De Gaudio S Romagnoli, D Quattrone, AR De Gaudio S Romagnoli, D Quattrone, AR De Gaudio
Azienda Ospedaliero Universitaria Careggi, Florence, Italy
Critical Care 2013, 17(Suppl 2):P197 (doi: 10.1186/cc12135) g
Azienda Ospedaliero Universitaria Careggi, Florence, Italy y
Critical Care 2013, 17(Suppl 2):P197 (doi: 10.1186/cc12135) Conclusion Vigileo did not prove to be a substitute to the reference
system; pre-loaded data, necessary for vascular impedance estimation,
may be one of the main limitations that made Vigileo measurements
less accurate than the MostCare ones. On the contrary, MostCare, an
uncalibrated totally independent system, was shown to properly
estimate the vascular impedance in these hemodynamically stable
patients. Further comparisons in unstable conditions are needed to
confi rm our observations. Introduction In the present study we analyzed the reliability for
cardiac output (CO) measurement of Vigileo (Edwards Lifescience,
Irvine, CA, USA) and MostCare (pressure recording analytical method;
Vygon-Vytech, Padova, Italy) in comparison with transthoracic Doppler
echocardiography (as the reference method) in patients undergoing
vascular surgery. Estimation of potential cost-savings related to the implementation
of perioperative hemodynamic goal-directed therapy CO
values ranged from 3.9 and 8.6 l/minute (echo), from 3.4 to 9.9 (Vigileo)
and from 4 to 8.3 (MostCare); the Pearson’s and Bland–Altman methods
showed poor agreement between COECHO and COVIG, demonstrating
a tendency to overestimation (see Figure 1). The percentage of error (PE)
was 51.7% at T1 and 49.3% at T2. On the contrary, MostCare measures
showed good agreement with echocardiography (see Table 1) with a
PE of 22.4% at T1 and of 17% at T2. Computerized decision support system improves consistency of
haemodynamic assessment amongst ICU team members
AE Aneman, R Ranganatha
Liverpool Hospital, Liverpool, Australia
Critical Care 2013, 17(Suppl 2):P199 (doi: 10.1186/cc12137) Computerized decision support system improves consistency of
haemodynamic assessment amongst ICU team members
AE Aneman, R Ranganatha
Liverpool Hospital, Liverpool, Australia
Critical Care 2013, 17(Suppl 2):P199 (doi: 10.1186/cc12137) Methods The in vitro set-up consisted of a centrifugal pump, a network
of tubes with variable volumes, an electromagnetic fl owmeter to
measure and adjust the generated fl ow, heating devices to maintain
constant temperature (37°C), two thermistors for thermodilution
measurement, an ultrasound transducer and a pressure stabilizer. A
small bolus of UCA diluted in cold saline (1 mg SonoVue® in 20 ml saline
at 4°C) was injected into the system. The cold UCA passage through a
fi rst and a second region of interest (ROI) was measured simultaneously
with the ultrasound transducer and the thermistors. The measurements
were performed at diff erent fl ows and volumes. BVs were estimated
using the two diff erent approaches, namely CEUS and thermodilution. The IDCs were processed and fi tted separately with a dedicated model
to estimate the ΔMTT of the cold UCA bolus between the two ROIs and
the two thermistors. All the measurements were repeated three times. Results A linear relation between BVs estimated by the two techniques
was observed with a correlation coeffi cient of 0.94. The bias of
CEUS with respect to the true volumes was –40.1 ml; the bias of
thermodilution was 84.3 ml. The most prominent diff erences between
the two techniques were observed in case of high volume and low fl ow,
possibly due to diff erent transport kinetics between UCAs and heat. AE Aneman, R Ranganatha Liverpool Hospital, Liverpool, Australia p
p
p
Critical Care 2013, 17(Suppl 2):P199 (doi: 10.1186/cc12137) Critical Care 2013, 17(Suppl 2):P199 (doi: 10.1186/cc12137) Introduction This study evaluated the potential of the NAVIGATOR
(Applied Physiology) haemodynamic monitor to improve consistency
of cardiovascular assessments amongst ICU members with diff erent
levels of expertise and experience. Methods Patients (n = 20) post cardiac surgery were monitored by
NAVIGATOR to display heart effi ciency (Eh), mean systemic fi lling
pressure (Pms) and vascular resistance (SVR) against targets for
mean arterial pressure and cardiac output set by the clinical team. i
References y
Methods Both Vigileo and MostCare were connected to the FloTrac
transducer (Edwards Lifescience) for CO calculation. The data from
Vigileo and MostCare were registered (COVIG and COMC respectively) . Romano SM, et al.: Crit Care 2002, 30:1834-1841. . Romano SM, et al.: Crit Care 2002, 30:1834-1841. 2. Geisen M, et al.: Curr Opin Crit Care 2012, 18:377-384. Figure 1 (abstract P197). Bland–Altman analysis (T1). Figure 1 (abstract P197). Bland–Altman analysis (T1). Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S75 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P198 Agreement (maximum two steps deviation for each assessment
between staff ; number of patients), disagreement (median and
interquartile range of steps) and therapeutic agreement (intervention/s
to change Eh, Pms and SVR in similar direction/s, number of patients)
were recorded and analysed for statistical diff erence BLIND versus
OPEN (Fisher’s exact test, Mann–Whitney test, P <0.05). P198
Brachial pulse waveform characteristics predict development of
organ failure in septic patients
S Kazune, I Jagmane
Hospital of Traumatology and Orthopaedics, Riga, Latvia
Critical Care 2013, 17(Suppl 2):P198 (doi: 10.1186/cc12136) y
Results Eh was commonly overestimated, Pms commonly under-
estimated with no clear trend for SVR. Agreement amongst all
categories of staff increased and disagreement score decreased for
Eh, Pms and SVR (Table 1) comparing BLIND versus OPEN assessments. Agreement for therapeutic interventions also increased signifi cantly
from 4/20 (BLIND) to 18/20 patients (OPEN). Introduction Pulse waveform characteristics (pulse wave transit
time and augmentation index) are measures of arterial stiff ness. Previous studies have found an association between severity of
acute infl ammatory states and increased arterial stiff ness but it is
not known whether non-invasive pulse waveform analysis could
predict development of multiple organ failure in septic patients. The
purpose of this study was to evaluate the photoplethysmographic
brachial artery pulse wave transit time and augmentation index and
their changes in response to induced forearm ischemia in septic ICU
patients and correlate these indices to the development of subsequent
end organ damage. Table 1 (abstract P199)
BLIND
OPEN
Eh
6/20, 2 (1 to 3)
17/20*, 1 (0 to 1)*
Pms
6/20, 2.5 (2 to 4)
17/20*, 2 (1 to 2)*
SVR
8/20, 2 (1 to 2)
16/20*, 1 (1 to 2)*
Patients (n/20) and score (median and IQR). *P <0.05. Methods A prospective observational study in patients with sepsis
within 24 hours of admission. i
References Severity of sepsis was assessed with
APACHE II score (median 18.5) and SOFA score (median 7.5). Three-
minute signal recording was done concurrently from the brachial
artery at the elbow and the radial artery at the wrist with an originally
designed photoplethysmograph at rest and after 5 minutes of induced
forearm ischemia. Recordings were analyzed to obtain the pulse wave
transit time and augmentation index at rest and 60 seconds after
induced ischemia. The SOFA score was recalculated at 48 hours post
recording. Patients (n/20) and score (median and IQR). *P <0.05. Conclusion The assessment of heart function, intravascular fi lling
and resistance state in postoperative cardiac patients was made
signifi cantly more consistent amongst diff erent ICU staff members
using the NAVIGATOR haemodynamic monitor. Such enhanced
consistency could potentially make the haemodynamic management
more eff ective with improved clinical outcomes. Results We studied 14 consecutive general ICU patients. There was
a negative linear relationship between the pulse wave transit time
(median 22.6 ms) at rest and increase in SOFA score in 48 hours
(P = 0.02, r = 0.96). The postischemic pulse wave transit time increased
in all patients (median 25.7 ms) but no association was found
between the proportion of increase and subsequent change in SOFA. Correlation between rest (median 7.6) and postischemic (median 7.2)
augmentation index and 48-hour SOFA scores was not statistically
signifi cant (r = 0.57, P = 0.46). Reference Reference 1. Vlachopoulos C, Dima I, Aznaouridis K, Vasiliadou C, Ioakeimidis N, Aggeli C,
Toutouza M, Stefanadis C: Acute systemic infl ammation increases arterial
stiff ness and decreases wave refl ections in healthy individuals. Circulation
2005, 112:2193-2200. Volume quantifi cation by contrast-enhanced ultrasound and
thermodilution: an in vitro comparison gi
Conclusion This study indicates that in early sepsis pulse waveform
characteristics could predict the risk of developing end organ failure. The pulse wave transit time is more robust than the augmentation
index and could be easier to use in patients with poor perfusion. Vascular reactivity indices do not seem to have predictive value in this
context. Introduction In clinical practice, blood volumes (BV) are typically
measured by thermodilution. Recently, contrast-enhanced ultrasound
(CEUS) has been proposed as an alternative minimally invasive
approach for BV assessment [1]. This method measures BV using a
single peripheral injection of a small bolus of ultrasound contrast
agent (UCA) detected by an ultrasound scanner. By measuring the
acoustic backscatter, two indicator dilution curves (IDCs) can be
derived from two diff erent sites in the circulatory system. IDC analysis
permits deriving the mean transit time (MTT) the bolus takes to travel
between the injection site and two measurement sites. Assessment of
the BV between these sites is obtained by multiplying the diff erence
in MTT (ΔMTT) by the blood fl ow. In this study, we compare diff erent
volumes in an in vitro set-up by CEUS with true set-up volumes and
thermodilution acquired volumes. Introduction In clinical practice, blood volumes (BV) are typically
measured by thermodilution. Recently, contrast-enhanced ultrasound
(CEUS) has been proposed as an alternative minimally invasive
approach for BV assessment [1]. This method measures BV using a
single peripheral injection of a small bolus of ultrasound contrast
agent (UCA) detected by an ultrasound scanner. By measuring the
acoustic backscatter, two indicator dilution curves (IDCs) can be
derived from two diff erent sites in the circulatory system. IDC analysis
permits deriving the mean transit time (MTT) the bolus takes to travel
between the injection site and two measurement sites. Assessment of
the BV between these sites is obtained by multiplying the diff erence
in MTT (ΔMTT) by the blood fl ow. In this study, we compare diff erent
volumes in an in vitro set-up by CEUS with true set-up volumes and
thermodilution acquired volumes. Fluid responsiveness in pressure support ventilation: role of asynchrony
A Messina, D Colombo, G Cammarota, M De Lucia, F Della Corte,
P Navalesi
Università Maggiore della Carità A. Avogadro, Novara, Italy
Critical Care 2013, 17(Suppl 2):P202 (doi: 10.1186/cc12140) Introduction Pulse pressure variation (PPV) is a dynamic indicator
of fl uid responsiveness, which is known to have a low sensibility and
specifi city in patients ventilated in pressure support (PS) [1]. We aim
to investigate patient–ventilator asynchrony as a potential source of
hemodynamic interference in PS. Methods Institutional ethical approval was obtained. Intubated major
trauma patients were sequentially enrolled. Exclusions included major
thoracic burns and children. Bioreactance fl ow monitoring (NICOM;
Cheetah) was applied at the same time as ECG leads and the calibration
step performed during handover from the prehospital team. Time to
availability of oxygen delivery data was recorded and trauma team
members surveyed regarding for perceived benefi ts and concerns from
this monitoring. The infl uence of fl ow monitoring on fl uid resuscitation,
time to CT and defi nitive disposal (to OR/ICU) was measured and
compared with a control population matched for injury severity score,
age and sex. y
Methods We performed a prospective study including PS ventilated
patients who met inclusion criteria for fl uid depletion [1]. Patients who
showed an asynchrony index (AI) exceeding 10% were included in the
asynchrony group (AG). The remaining patients were included in the
synchrony group (SG) [2]. Beat-to-beat hemodynamic variables were
recorded through PRAM (Mostcare; Vytech Health srl, Padova, Italy). PPV cutoff of 13% was used to identify fl uid responders/nonresponders. A fl uid challenge of 500 ml normal saline was given in 5 minutes. An increase of 15% of cardiac index after 10 minutes indicated fl uid
responsiveness. Results Cardiac index was available at mean 10.6 minutes (median
9 minutes; SD 3.9), fl uid responsiveness at mean 35.9 minutes (median
35; SD 11.3) and oxygen delivery calculation at mean 25.3 minutes
(median 25; SD 7.7). Passive leg raise was not performed in 63% of
patients due to concerns about pelvic or brain injury. Volume of fl uid
infused (mean 738 vs. 925 ml; P = 0.124), time to CT (mean 57.4 vs. 68.8 minutes; P = 0.08), and time to defi nitive disposal (mean 124.9 vs. 146.1 minute; P = 0.069) were all reduced in the fl ow monitored group,
although not signifi cantly diff erent when compared with a matched Results So far, eights patients showed an AI >10% while 16 did not. Overall sensitivity was 28.6% versus 50% in SG; overall specifi city was
76.5% versus 91.7% in AG. Computerized decision support system improves consistency of
haemodynamic assessment amongst ICU team members
AE Aneman, R Ranganatha
Liverpool Hospital, Liverpool, Australia
Critical Care 2013, 17(Suppl 2):P199 (doi: 10.1186/cc12137) Four categories of staff participated: nine consultants (C), eight
senior registrars (SR), nine registrars (R) and 11 nurses (N) (median
ICU experience 15, 7, 2 and 10 years, respectively) and were asked
to score Eh, Pms, and SVR ranging (discrete steps of one) from –5
(grossly subnormal) to 0 (normal) to 5 (grossly supranormal) fi rst
without (BLIND) and then given (OPEN) access to the NAVIGATOR
display. Recommendations for therapeutic interventions were noted. S76 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 control group (Mann–Whitney U rank sum). Eighty-four percent of
trauma team members surveyed felt the fl ow monitoring data to be
useful, and only 11% felt it may impair clinical management.l Conclusion Given the good correlation between BVs estimated with
CEUS and thermodilution, CEUS is not inferior to thermodilution with
the advantage of being minimally invasive. Reference Conclusion Cardiac index, fl uid responsiveness and oxygen delivery
data can be obtained inform a primary survey. Rather than introducing
delays, the use of fl ow monitoring was associated with a trend towards
decreased time to imaging; less fl uid use pre-damage control point
and reduced time to defi nitive disposal. Further research is required to
confi rm benefi ts and mechanism. 1. Mischi M, et al.: IEEE Trans UFFC 2004, 51:1137-1141. 1. Mischi M, et al.: IEEE Trans UFFC 2004, 51:1137-1141. P202 Fluid responsiveness in pressure support ventilation: role of asynchrony
A Messina, D Colombo, G Cammarota, M De Lucia, F Della Corte,
P Navalesi and disposal y
,
,
3. Dunham M, et al.: J Trauma Acute Care Surg 2012, 73:479-485. y
3. Dunham M, et al.: J Trauma Acute Care Surg 2012, 73:479-485. King’s Health Partners AHSC, London, UK Critical Care 2013, 17(Suppl 2):P201 (doi: 10.1186/cc12139) Fluid responsiveness in pressure support ventilation: role of asynchrony
A Messina, D Colombo, G Cammarota, M De Lucia, F Della Corte,
P Navalesi Fluid responsiveness in pressure support ventilation: role of asynchrony
A Messina, D Colombo, G Cammarota, M De Lucia, F Della Corte,
P Navalesi
Università Maggiore della Carità A. Avogadro, Novara, Italy
Critical Care 2013, 17(Suppl 2):P202 (doi: 10.1186/cc12140) Introduction The use of cardiac output monitoring has been shown to
be benefi cial in the setting of perioperative medicine and critical illness
[1,2]. More recently, its application in the setting of major trauma has
been described [3]. Here, we describe our preliminary experience of
embedding bioreactance fl ow monitoring within the major trauma
primary survey of severely injured patients and the subsequent eff ect
on patient management. P203
Comparison of pulse pressure variation with radial arterial systolic,
diastolic and pulse transit time interval variation in pediatric
patients undergoing liver transplantation
YJ Moon, IG Jun, WJ Shin, BH Sang, GS Hwang
Asan Medical Center, Seoul, South Korea
Critical Care 2013, 17(Suppl 2):P203 (doi: 10.1186/cc12141) P203
Comparison of pulse pressure variation with radial arterial systolic,
diastolic and pulse transit time interval variation in pediatric
patients undergoing liver transplantation
YJ Moon, IG Jun, WJ Shin, BH Sang, GS Hwang
Asan Medical Center, Seoul, South Korea
Critical Care 2013, 17(Suppl 2):P203 (doi: 10.1186/cc12141) l
g
p
Methods We measured the eff ects after the administration of 50, 100
and 500 ml bolus colloid infusions on CO (Modelfl ow (COm) and LiDCO
(COli)), CVP and MAP in 21 patients on mechanical ventilation after
elective cardiothoracic surgery. From the data we analysed the smallest
volume that was predictive for the eff ects of 500 ml on cardiac output. Results COli and COm increased after 50, 100 and 500 ml fl uid loading. Best results are observed for changes in COm after 100 ml fl uid loading
(area under the ROC 0.86, 95% CI between 0.65 and 1.00). A change in
Modelfl ow CO of at least 4.3% has a sensitivity of 67% and a specifi city
of 100% after 100 ml fl uid loading. Sensitivity is 60% and specifi city
83% for a similar cutoff in CO measured with the LiDCO device after
100 ml fl uid loading. In our patient population, MAP and COli did not
predict responsiveness with more accuracy than mathematical chance. See Figure 1.l Introduction In pediatric patients, dynamic preload indices to predict
fl uid responsiveness have confl icting results in comparison with adults. A recent study demonstrated that pulse pressure variation (PPV) ≥16%
has provided an accurate method for predicting fl uid responsiveness
in pediatric congenital heart surgery. We aimed to compare PPV and
respiratory systolic, diastolic and pulse transit time interval variation
(STV, DTV and PTTV, respectively) as predictors of fl uid responsiveness
during pediatric liver transplantation. g p
p
Methods A total of 61 data from 16 pediatric patients, median age 5.4 years
(range 0.1 to 9 years), were retrospectively evaluated from electrically
recorded radial arterial and central venous pressure (CVP) waveform. The
time from the onset of systolic upstroke to the dicrotic notch was defi ned
as the systolic time interval (STI), and the time from dicrotic notch to the
beginning of systolic upstroke was defi ned as the diastolic time interval
(DTI). The time from peak R wave on electrocardiography to the onset of
systolic upstroke was defi ned as the pulse transit time (PTT) interval. P203
Comparison of pulse pressure variation with radial arterial systolic,
diastolic and pulse transit time interval variation in pediatric
patients undergoing liver transplantation
YJ Moon, IG Jun, WJ Shin, BH Sang, GS Hwang
Asan Medical Center, Seoul, South Korea
Critical Care 2013, 17(Suppl 2):P203 (doi: 10.1186/cc12141) STV
was calculated by averaging of three consecutive respiratory cycles with
the following: (STImaximum – STIminimum) / STImean. The same method was used
for calculating DTV, PTTV and PPV. STV, DTV and PTTV were corrected
by cardiac period. Averaged CVP was used as a static preload index. PPV
threshold ≥16% was used to discriminate fl uid responsiveness. Receiver
operating characteristic (ROC) curves and Pearson’s correlation analysis
were used for the comparison. Conclusion Changes in pulse contour CO can be used in a mini-fl uid
challenge to assess fl uid responsiveness in our postcardiac surgery
patients. Effi cacy of intraoperatory optimisation of fl uids guided with
transoesophageal Doppler monitorisation: a multicentre
randomised controlled trial
S Maeso1, R Villalba2, J Ripollés3, S Asuero2, J Blasco1, J Calvo3
1Agencia Lain Entralgo, Madrid, Spain; 2Hospital Universitario Ramón y Cajal,
Madrid, Spain; 3Hospital Infanta Leonor, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P205 (doi: 10.1186/cc1243) Fluid responsiveness in pressure support ventilation: role of asynchrony
A Messina, D Colombo, G Cammarota, M De Lucia, F Della Corte,
P Navalesi
Università Maggiore della Carità A. Avogadro, Novara, Italy
Critical Care 2013, 17(Suppl 2):P202 (doi: 10.1186/cc12140) Overall Cohen’s k was 33.3% versus 61.2% in
AG (see Figure 1). However, because none of the responders in the AG
group was detected by PPV, statistical analysis was not feasible within
this subgroup. Figure 1 (abstract P202). Figure 1 (abstract P202). S77 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Conclusion The consistency of PPV in predicting fl uid responsiveness
during PS seems to be more reliable in the patients with better patient–
ventilator synchrony. References References
1. Heenen S, et al.: Crit Care 2006, 10:R102. 2. Vitacca M, et al.: Chest 2004, 126:851-859. Introduction The mini-fl uid challenge is a widely used strategy to
manage fl uid loading in the ICU and OR. Although it might be a rational
strategy, data on the mini-fl uid challenge and its reliability are very
limited. We investigated the value of changes in pulse contour cardiac
output as a result of a mini-fl uid challenge of 50 and 100 ml to predict
fl uid loading responsiveness. Mini-fl uid challenge: how much fl uid and what parameter to use?
B Geerts, RB De Wilde, JJ Maas, LP Aarts, JR Jansen
Leiden University Medical Centre, Leiden, the Netherlands
Critical Care 2013, 17(Suppl 2):P204 (doi: 10.1186/cc12142) Mini-fl uid challenge: how much fl uid and what parameter to use? B Geerts, RB De Wilde, JJ Maas, LP Aarts, JR Jansen
Leiden University Medical Centre, Leiden, the Netherlands
Critical Care 2013, 17(Suppl 2):P204 (doi: 10.1186/cc12142) y
y
References
1. Heenen S, et al.: Crit Care 2006, 10:R102. 2. Vitacca M, et al.: Chest 2004, 126:851-859. Effi cacy of intraoperatory optimisation of fl uids guided with
transoesophageal Doppler monitorisation: a multicentre
randomised controlled trial S Maeso1, R Villalba2, J Ripollés3, S Asuero2, J Blasco1, J Calvo3
1Agencia Lain Entralgo, Madrid, Spain; 2Hospital Universitario Ramón y Cajal,
Madrid, Spain; 3Hospital Infanta Leonor, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P205 (doi: 10.1186/cc1243) Results PPV showed correlations with STV, DTV and PTTV (r = 0.65,
0.57 and 0.60, respectively), but less with CVP (r = –0.30). Area under
ROC curves (AUC) of STV, DTV, PTTV and CVP were 0.834, 0.872, 0.832
and 0.613, respectively. Cutoff values of STV, DTV, PTTV and CVP were
7.7% (sensitivity/specifi city, 0.80/0.83), 7.7% (sensitivity/specifi city,
0.70/0.88), 8.7% (sensitivity/specifi city 0.67/1.0) and 3.1% (sensitivity/
specifi city 0.50/0.85), respectively. Introduction The objective is to compare stay in surgery monitored
with oesophageal Doppler with unmonitored. Introduction The objective is to compare stay in surgery monitored
with oesophageal Doppler with unmonitored. g
Methods A randomized trial. We present preliminary results obtained
in the fi rst 55 cases. The surgeries were general and urological. ISRCTN93543537. Results There were no diff erences in any of the baseline variables. A
total of 69.1% were men. The mean age was 65.80 years. There were
63.6% general surgery and 36.4% urologic surgery. There were 81.5%
open surgeries and 18.5% laparoscopic. The results were favorable to
the intervention group for most outcomes; these diff erences did not i
y
y
Conclusion This study shows that STV, DTV and PTTV can be used as
a surrogate for PPV ≥16%, suggesting that this novel method can be
used to predict hemodynamic response during pediatric surgery. Reference 1. Renner J, et al.: Br J Anaesth 2012, 108:108-115. Figure 1 (abstract P204). Cardiac function curve: a fl uid challenge of 50 ml in a (non)responder. gure 1 (abstract P204). Cardiac function curve: a fl uid challenge of 50 ml in a (non)responder. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S78 Table 1 (abstract P205). Analysis of the outcome variables
ODM
Control
Outcome
Units
(n = 32)
(n = 23)
P value
Surgery time
Minutes
206.20
225.59
0.532
Total stay
Days
13.92
18.19
0.292
Postsurgery
Days
10.93
14.36
0.280
Time to enteral
Days
2.73
3.13
0.567
Time to deambulation
Days
2.52
3.00
0.359
Complications
%
34.4
21.7
0.377 Table 1 (abstract P205). Effi cacy of intraoperatory optimisation of fl uids guided with
transoesophageal Doppler monitorisation: a multicentre
randomised controlled trial Analysis of the outcome variables P207
Prediction of stroke volume response to fl uid bolus in 100 children
R Saxena, A Durward, I Murdoch, S Tibby
Guy’s & St Thomas’ NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P207 (doi: 10.1186/cc12145)
Figure 1 (abstract P206). Figure 1 (abstract P206). reach statistical signifi cance (Table 1). We emphasize the diff erence in
postoperative stay of 3.4 days. Only complications results were against
the intervention. Conclusion The preliminary results obtained for the postsurgical length
of stay, as for most of the outcomes, were favorable to monitoring by
oesophageal Doppler. P206 P206
Looking for the infl exion point of the Frank–Starling curve
HD Aya, M Geisen, C Ebm, N Fletcher, M Grounds, A Rhodes, M Cecconi
St George’s Healthcare NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P206 (doi: 10.1186/cc12144) Introduction Fluid overload is associated with poor outcome in the
critically ill. Thus, an accurate predictor of a positive haemodynamic
response (increase in stroke volume) to fl uid challenge is vital.i l
Methods We studied the predictive value (positive response defi ned
as change in stroke volume >15% after 10 ml/kg fl uid bolus) of a range
of haemodynamic variables: static (CVP, active circulating volume,
central blood volume, total end diastolic volume), dynamic (systolic
pressure variation, stroke volume variation) and contactility (dp/dt),
in a group of 100 ventilated children (median weight 10 kg). Variables
were measured using transpulmonary ultrasound dilution and PRAM
(an arterial pulse contour method). Introduction Fluid responsiveness is defi ned based on an arbitrary
increase of cardiac output (CO) or stroke volume (SV) of 10 to 15%. We
hypothesise that the variation of heart effi ciency (Eh) and the slope
(S) defi ned by the relative increase of CO over the relative increase of
mean fi lling pressure (Pmsa) can be used as alternative defi nitions of
fl uid responsiveness. l
p
Methods Patients admitted to the ICU were monitored with a calibrated
LiDCOplus (LiDCO, UK) and Navigator (Applied Physiology, Australia) to
estimate Pmsa and Eh (Pmsa – central venous pressure/Pmsa). A 250 ml
fl uid challenge was performed over 5 minutes. Categorical data were
compared by Pearson chi-square test. Correlation was assessed by
Kappa test. The infl exion point of S to defi ne responders was obtained
by ROC curve analysis. p
Results We performed 168 paired measurements (pre-fl uid and post-
fl uid challenge), with a SV response rate of 45%. Overall predictive
values were poor, but slightly better for static versus dynamic variables
(Table 1). When SV response was analysed as a continuous variable,
the two predictive multivariable variables were change in TEDVI and
baseline dp/dt (r2 = 0.30, both P <0.001). Table 1 (abstract P207). ROC areas for haemodynamic variables
AUC
95% CI
CVP
0.50
0.41
0.59
ACVI
0.62
0.54
0.71
CBVI
0.59
0.50
0.68
TEDVI
0.65
0.56
0.73
SPV
0.59
0.50
0.69
SVV
0.48
0.39
0.58
dp/dt
0.59
0.50
0.68 Table 1 (abstract P207). ROC areas for haemodynamic variables Table 1 (abstract P207). P207
d P207
Prediction of stroke volume response to fl uid bolus in 100 children
R Saxena, A Durward, I Murdoch, S Tibby
Guy’s & St Thomas’ NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P207 (doi: 10.1186/cc12145) P206 ROC areas for haemodynamic variables
AUC
CI y
y
Results A total of 104 fl uid challenges were observed in 40 patients. ROC
curve analysis reveals an area under the curve of 0.93 (95% CI = 0.85
to 1, P <0.001). The best cutoff for the slope was 0.76 (sensitivity 0.92,
specifi city 0.93). The proportions of responders identifi ed by the ΔEh
(Table 1) and by the slope method (Table 2) are smaller compared with
the relative increase of SV method. Signifi cant correlation was found
between both methods and the ΔSV (ΔEh κ = 0.54, P <0.001; S κ = 0.55,
P <0.001). See Figure 1. g
Conclusion Moderate agreement is observed between new and
current defi nitions of fl uid responsiveness. 1. Michard F, et al.: Chest 2002, 121:2000. Table 1 (abstract P206). Distribution of events according to ΔSV and ΔEh
Response by ΔSV ≥10%
Response by ΔEh ≥0
P value
Nonresponder
62 (59.6%)
76 (73.1%)
<0.001
Responder
42 (40.4%)
28 (26.9%)
<0.001
Table 2 (abstract P206). Distribution of events according to ΔSV and the
slope (S)
Response by ΔSV ≥10%
Response by S ≥0.76
P value
Nonresponder
62 (59.6%)
75 (72.1%)
<0.001
Responder
42 (40.4%)
29 (27.9%)
<0.001 Table 1 (abstract P206). Distribution of events according to ΔSV and ΔEh Conclusion The predictive ability for typical static and dynamic
haemodynamic variables, when taken in isolation, is poor. However,
improved prediction is seen when baseline contractility is taken into
account. SP De Wolf, J Stens, RJ Van der Zwan, N Koning, C Boer
VU University Medical Centre, Amsterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P208 (doi: 10.1186/cc12146) Introduction This study compared the eff ect of pulse pressure variation
(PPV) and cardiac index (CI)-guided fl uid therapy versus mean arterial S79 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Conclusion SDF imaging is a feasible, non-invasive bedside method
to study the postoperative intestinal microcirculation. The established
reference ranges are useful for early detection of postoperative local
complications and studies of microcirculatory changes induced by
systemic pathologies; for example, in sepsis. pressure (MAP)-guided fl uid therapy on microcirculatory perfusion in
patients undergoing abdominal surgery. g
g
g
y
Methods Patients undergoing elective abdominal surgery were
randomized into a PPV/CI-guided group (n = 11) or a MAP-guided
(n = 12) group. P206 PPV, CI and MAP were measured using the non-invasive
fi nger arterial blood pressure measurement device ccNexfi n (Edwards
Lifesciences BMEYE, Amsterdam, the Netherlands). Tidal volumes were
≥8 ml/kg with PEEP ≥8 mmHg. In both groups, MAP of 70 mmHg
was maintained. In the PPV/CI group, an intraoperative algorithm
was used keeping the PPV under 12% and CI above 2.5 l/minute/
m2 using fl uid therapy and dobutamine and noradrenaline infusion,
respectively. Sublingual microvascular perfusion was measured after
anesthesia induction, and every subsequent hour using sidestream
dark-fi eld imaging (Microscan; Microvision Medical, Amsterdam, the
Netherlands). The perfused small vessel density (PVD) values were
offl ine quantifi ed.i P210 P210
Monitoring microcirculatory blood fl ow with a new sublingual
tonometer in a porcine model of haemorrhagic shock
P Palagyi, J Kaszaki, Z Molnar
University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P210 (doi: 10.1186/cc12148) Introduction Tissue capnometry has been used to assess organ
perfusion but it is not generally available at the bedside [1,2]. Our aim
was to test a new sublingual capillary tonometer in haemorrhagic
shock. fl
qi
Results The fi rst hour during surgery, the PPV/CI-guided group tended
to receive more fl uids than the MAP-guided group (1,014 ± 501ml vs. 629 ± 463 ml; P = 0.07). At this time point, the PVD was slightly lower in
the PPV/CI-guided group (16.7 ± 3.1 mm/mm2) when compared with
the MAP-guided group (17.9 ± 3.9 mm/mm2; P = 0.41). In both groups
the PVD remained stable during the fi rst 2 hours of surgery. However,
2 hours after the start of surgery, the PVD in the PPV/CI group restored
and tended to be higher than in the MAP-guided group (21.1 ± 1.9 vs. 18.1 ± 3.4 mm/mm2; P = 0.09). After 1 hour of surgery, the administered
fl uid volume correlated inversely with PVD (r = –0.59, P = 0.011).l Methods Thirty-six mini-pigs were anaesthetised, ventilated and divided
into sham operated and shock groups. Instrumentation included:
intestinal and sublingual tonometry, haemodynamic monitoring
(PiCCO; Pulsion, Germany) and orthogonal polarization spectroscopy
(OPS). After baseline measurements (T0) haemorrhagic shock was
induced and maintained by reducing mean arterial pressure (MAP)
to ~40 mmHg for 60 minutes. Measurements were repeated every
30 minutes (T1 to T6). Fluid resuscitation started after T2 aiming to
increase MAP to 75% of the baseline value. OPS imaging was performed
at T0, T2 and T6. P211 Methods For this observational prospective cohort study 77 patients
were screened. In total, 165 SDF measurements could be obtained. All
patients included had bowel surgery for chronic infl ammatory bowel
disease or intestinal malignancies and received an ileostomy. Patients
were excluded if they had signs of local infection or bleeding, or if they
were admitted to the ICU for sepsis or perioperative complications. The
SDF device was gently inserted into the stoma at a depth of 3 to 4 cm, and
fi ve real-time images were recorded. All videos recorded were analyzed
offl ine using AVA® software (Microvision Medical, Amsterdam, the
Netherlands). The following parameters were quantifi ed: microvascular
fl ow index (MFI), total vessel density (TVD), perfused vessel density
(PVD), and proportion of perfused vessels (PPV). Patients were followed
for 3 days post surgery with fi ve images captured every day. References 1. Creteur et al.: Curr Opin Crit Care 2006, 12:272-277. 2. Cammarata et al.: Shock 2009, 31:207-211. 1. Creteur et al.: Curr Opin Crit Care 2006, 12:272-277. 2. Cammarata et al.: Shock 2009, 31:207-211. Postoperative imaging of the intestinal microcirculation
I Abd
1 R H ll1 D H
l
1 V C
2 CH L h
1 Abdo1, R Hall1, D Henzler1, V Cerny2, CH Lehmann1 1Dalhousie University, Halifax, Canada; 2Charles University, Medical Faculty in
Hradec Kralove, Czech Republic Conclusion In this model of haemorrhagic shock, sublingual PCO2
gap showed good correlation with RBCV, suggesting that this new
sublingual capillary tonometer may be an appropriate tool for
monitoring microcirculation at the bedside. f Introduction Sidestream dark-fi eld (SDF) imaging was introduced
recently to study the sublingual microcirculation in humans. Patients
with ileostomies off er a unique access to the intestinal microcirculation. To date, no reference ranges for standard microcirculatory parameters
of the gut are available. Therefore, the aim of our study was to
establish a database for postoperative microcirculatory parameters for
ileostomies. P209 Postoperative imaging of the intestinal microcirculation
I Abdo1, R Hall1, D Henzler1, V Cerny2, CH Lehmann1
1Dalhousie University, Halifax, Canada; 2Charles University, Medical Faculty in
Hradec Kralove, Czech Republic
Critical Care 2013, 17(Suppl 2):P209 (doi: 10.1186/cc12147) P206 Data are presented as the median (interquartile
range), for statistical analysis Friedmann ANOVA, Mann–Whitney and
Spearman tests were used as appropriate. l
y
Conclusion Goal-directed fl uid management resulted in a higher
administered fl uid volume in the beginning of surgery, and this was
associated with a slightly reduced microcirculatory perfusion when
compared with MAP-guided fl uid management. Microcirculatory
perfusion tended to improve as surgery progressed in the goal-directed
fl uid therapy group. Our fi ndings suggest that goal-directed and MAP-
guided fl uid management are associated with distinct patterns in
fl uid resuscitation, which may be of consequence for microvascular
perfusion. p
pp
p
Results Bleeding resulted in a signifi cant decrease in MAP and cardiac
index, and an increase in heart rate. Macrohaemodynamic changes
were accompanied by signifi cant changes in red blood cell velocity
(RBCV) and a signifi cant increase in the intestinal and sublingual
mucosal-to-arterial carbon dioxide partial pressure diff erence (PCO2
gap): from 4 (2 to 11) to 30 (23 to 37) mmHg in the sublingual, and from
25 (17 to 31) to 50 (33 to 64) mmHg in the ileum. RBCV decreased from
1,075 (945 to 1,139) to 520 (449 to 621) μm/second in the sublingual
area, and from 646 (596 to 712) to 419 (350 to 451) μm/second in the
ileum. There was signifi cant correlation between RBCV and PCO2 gap in
sublingual and intestinal regions alike: r = –0.58; r = –0.71, P <0.0001,
respectively. Sublingual microcirculatory changes during transient
intra-abdominal hypertension: a study in laparoscopic surgery
patients Introduction Microvascular alterations play an important role in
development of organ failure [1]. It is not known whether increased
intra-abdominal pressure (IAP) is associated with microcirculatory
perfusion derangements. Our hypothesis was that transiently
increased IAP is related to microcirculatory alterations in laparoscopic
cholecystectomy patients. Introduction Microvascular alterations play an important role in
development of organ failure [1]. It is not known whether increased
intra-abdominal pressure (IAP) is associated with microcirculatory
perfusion derangements. Our hypothesis was that transiently
increased IAP is related to microcirculatory alterations in laparoscopic
cholecystectomy patients. Results We were able to capture clear images of the small intestine
microvasculature. Distinct villi were visible with a dense network
of microvessels (diameter: 6 to 17 μm). Mean TVD (± 2SD) was 19.3
(± 1.0) mm/mm2, PVD: 18.5 (± 1.1) mm/mm2, PPV: 94.5 (± 5)% and
MFI: 2.8 (± 0.1). Patients’ age, sex and comorbidity had no signifi cant
impact on postoperative microvascular parameters. No signifi cant
changes of the microvascular parameters were observed during the
fi rst 3 postoperative days. Methods Sixteen patients (14 female, two male) who underwent
laparoscopic cholecystectomy were studied. Sublingual orthogonal
polarization spectral (OPS) imaging was used to detect microcirculatory
function. OPS was done before surgery, at least 15 minutes after
initiation of pneumoperitoneum and 1 hour after the end of pneumo-
peritoneum. The microcirculation cutoff value for vessels was 20 μm. Data are presented as medians with interquartile ranges. S80 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Results Patient median age was 54 (39 to 63) years, ASA score 2 (2 to 3),
BMI 29.7 (24.9 to 34.7), haemoglobin concentration 138 (133 to 142) g/l,
and hematocrit 42 (39 to 43). IAP was held at 12.5 (12 to 13) mmHg,
median duration of pneumoperitoneum was 45 (24 to 55) minutes. Median MAP was 86 (69 to 93), abdominal perfusion pressure (APP)
73 (57 to 81) mmHg during the pneumoperitoneum. Median fl uid
administration during anesthesia was 1,050 (1,000 to 1,400) ml. Altogether 448 microcirculation videos were taken. Interobserver
variability was 24%. The following microcirculatory parameter values
describe before, during and after pneumoperitoneum periods. Total
vascular density was 19.4 (17.0 to 21.1); 18.5 (17.0 to 20.9); 19.3 (16.9 to
20.9) n/mm2. Perfused vessels density was 13.3 (10.9 to 15.2); 13.8 (8.9
to 18.0); 13.1 (11.0 to 16.0) n/mm2. Microcirculatory response to experimentally-induced whole body
heat stress in healthy humans A reduction in glycocalyx thickness,
represented by an increased PBR, correlates with impairment of
perfused microvascular vessel density. Our data support the use
of intraoperative PBR monitoring as a novel clinical indicator of
microcirculatory perfusion. g y
y
Results The perfused vessel density remained stable during off -
pump surgery, while the PBR decreased from 2.6 ± 0.1 μm (baseline)
to 2.3 ± 0.1 μm (ICU). Nonpulsatile CPB was associated with loss
of microcirculatory perfusion during bypass (15.1 ± 2.7 mm/mm2)
and ICU admission (15.3 ± 2.6 mm/mm2) compared with baseline
(19.8 ± 2.8 mm/mm2). Pulsatile CPB reduced the perfused vessel
density from 20.9 ± 2.4 to 16.7 ± 2.6 mm/mm2, but this was restored
towards baseline levels upon ICU admission (20.3 ± 2.3 mm/mm2;
P = 0.02 between groups). The PBR increased upon initiation of CPB
in both groups, but remained elevated in the nonpulsatile fl ow group
only. In the pulsatile group, the PBR started at 2.4 ± 0.1 μm, increased
to 2.7 ± 0.1 μm during bypass but restored already during CPB towards
a PBR value of 2.3 ± 0.1 μm (ICU). The PBR and perfused vessel density
showed a good correlation (r = –0.65; P = 0.002), demonstrating loss of
perfused vascular density when the glycocalyx is damaged. y
y
y
y
Methods Eight healthy men with no history of cold and/or heat injury
were recruited to this study. Passive body heating was performed by
continuous immersion up to the waist in the water bath at 44°C and
continued until rectal temperature reached 39.5°C. Before, at the end
of whole body heating and 1 hour after heating was ended, systemic
hemodynamics and direct in vivo observation of the sublingual
microcirculation were obtained with sidestream dark-fi eld imaging. Assessment of microcirculatory parameters of convective oxygen
transport (microvascular fl ow index (MFI), proportion of perfused
vessels (PPV)), and diff usion distance (perfused vessel density (PVD)
and total vessel density (TVD)) was done using a semiquantitative
method. Vessels were separated into large (mostly venules) and small
(mostly capillaries) using a diameter cutoff value of 20 μm.i p
y
g y
y
g
Conclusion The glycocalyx is damaged after initiation of CPB, and
restoration is impaired after exposure to nonpulsatile fl ow during the
use of the heart–lung machine. A reduction in glycocalyx thickness,
represented by an increased PBR, correlates with impairment of
perfused microvascular vessel density. Changes in microcirculatory perfusion during cardiac surgery are
paralleled by alterations in glycocalyx integrityf Conclusion Defined cold exposure had no effect on the micro-
circulation. Introduction Previous studies demonstrate that loss of glycocalyx
integrity is associated with impaired microvascular function. We
investigated whether glycocalyx dimensions are reduced in patients
undergoing cardiac surgery with or without cardiopulmonary bypass
(CPB), and are paralleled by loss of microcirculatory perfusion using in
vivo microcirculation measurements. Microvascular response to cold stress in healthy humans
A Pranskunas1, R Rasimaviciute1, J Paltanaviciute1, L Daniuseviciute2,
V Pilvinis1, M Brazaitis2 Microvascular response to cold stress in healthy humans
A Pranskunas1, R Rasimaviciute1, J Paltanaviciute1, L Daniuseviciute2,
V Pilvinis1, M Brazaitis2
1Lithuanian University of Health Sciences, Kaunas, Lithuania; 2Lithuanian
Sports University, Kaunas, Lithuania
Critical Care 2013, 17(Suppl 2):P213 (doi: 10.1186/cc12151) Introduction Cold exposure can be adapted for exercise or therapeutic
purposes, but its impact on microcirculation in healthy humans has not
been well defi ned. We hypothesize that whole body cold stress may
impair microcirculation. Introduction Cold exposure can be adapted for exercise or therapeutic
purposes, but its impact on microcirculation in healthy humans has not
been well defi ned. We hypothesize that whole body cold stress may
impair microcirculation. Methods Seven volunteers were recruited for the water immersion
procedure. During the cooling protocol the volunteers every
20 minutes of immersion were asked to step out from the bath and
rest for 10 minutes in a room environment and then return to the
water bath for the next 20 minutes of immersion. This head-out
immersion procedure in bath water at 14°C continued until the rectal
temperature was dropped to 35.5°C or the time of 180 minutes was
terminated. Maximum cold water immersion time was 120 minutes. Before, at the end of whole body cooling and 1 hour after cooling was
ended, systemic hemodynamics and direct in vivo observation of the
sublingual microcirculation were obtained with sidestream dark-fi eld
imaging. Assessment of microcirculatory parameters of convective
oxygen transport (microvascular fl ow index (MFI), proportion of
perfused vessels (PPV)), and diff usion distance (perfused vessel density
(PVD) and total vessel density (TVD)) was done using a semiquantitative
method.i Conclusion Microcirculatory alterations are mild during transient
increase of intra-abdominal pressure in laparoscopic surgery patients. References 1. De Backer D, et al.: Crit Care Med 2006, 34:1918-1924. 2. Vellinga NAR, et al.: Crit Care Res Pract 2012. doi:10.1155/2012/121752 1. De Backer D, et al.: Crit Care Med 2006, 34:1918-1924. 2. Vellinga NAR, et al.: Crit Care Res Pract 2012. doi:10.1155/2012/121752 1. De Backer D, et al.: Crit Care Med 2006, 34:1918-1924. 1. De Backer D, et al.: Crit Care Med 2006, 34:1918-1924. 2. Vellinga NAR, et al.: Crit Care Res Pract 2012. doi:10.1155/2012/121752 P212 Changes in microcirculatory perfusion during cardiac surgery are
paralleled by alterations in glycocalyx integrity
C Boer1, NJ Koning1, J Van Teeff elen2, AB Vonk1, H Vink2
1VU University Medical Center, Amsterdam, the Netherlands; 2Maastricht
University, Maastricht, the Netherlands
Critical Care 2013, 17(Suppl 2):P212 (doi: 10.1186/cc12150) Sublingual microcirculatory changes during transient
intra-abdominal hypertension: a study in laparoscopic surgery
patients Proportion of perfused vessels (PPV)
was 61 (50 to 69); 64 (45 to 76); 60 (54 to 67)%. Microvascular fl ow index
was 2.4 (2.0 to 2.5); 2.5 (2.0 to 3.0); 2.3 (2.0 to 2.9) and heterogeneity
index was 0.8 (0 to 0.9); 0.6 (0 to 1.0); 0.6 (0 to 0.8). No signifi cant
diff erences in microcirculatory parameters were observed between
time points. PPV was somewhat less than that of described in healthy
volunteers (61% vs. 90%) [2]. Microcirculatory response to experimentally-induced whole body
heat stress in healthy humans A Pranskunas1, E Milieskaite1, I Maraulaite1, L Daniuseviciute2,
Z Pranskuniene1, V Pilvinis1, M Brazaitis2
1Lithuanian University of Health Sciences, Kaunas, Lithuania; 2Lithuanian
Sports University, Kaunas, Lithuania
Critical Care 2013, 17(Suppl 2):P214 (doi: 10.1186/cc12152) Methods Patients undergoing on-pump surgery with nonpulsatile
(n = 11) or pulsatile (n = 13) CPB or off -pump surgery (n = 13) underwent
sublingual sidestream dark-fi eld imaging at baseline, during coronary
grafting and upon ICU admission to assess perfused microvascular
vessel density. Glycocalyx integrity was evaluated using the GlycoCheck
Measurement Software, and expressed as the perfused boundary
region (PBR). An increase in PBR represents deeper penetration of
erythrocytes into the glycocalyx, and is indicative for compromised
glycocalyx thickness. Introduction Vasodilation and increased skin blood fl ow (also
sweating) are infl uential in heat dissipation during heat exposure and
exercise. It is unclear how heat stress infl uences microcirculation. Side
dark-fi eld imaging visualizes the blood fl ow at the capillary level and
helps to assess perfusion heterogeneity. Clinical and experimental
data show that the sublingual region is clinically relevant for
detecting microcirculatory alterations and more represents central
microcirculation than cutaneous perfusion. We hypothesize that whole
body heat stress may increase capillary density. g y
y
Results The perfused vessel density remained stable during off -
pump surgery, while the PBR decreased from 2.6 ± 0.1 μm (baseline)
to 2.3 ± 0.1 μm (ICU). Nonpulsatile CPB was associated with loss
of microcirculatory perfusion during bypass (15.1 ± 2.7 mm/mm2)
and ICU admission (15.3 ± 2.6 mm/mm2) compared with baseline
(19.8 ± 2.8 mm/mm2). Pulsatile CPB reduced the perfused vessel
density from 20.9 ± 2.4 to 16.7 ± 2.6 mm/mm2, but this was restored
towards baseline levels upon ICU admission (20.3 ± 2.3 mm/mm2;
P = 0.02 between groups). The PBR increased upon initiation of CPB
in both groups, but remained elevated in the nonpulsatile fl ow group
only. In the pulsatile group, the PBR started at 2.4 ± 0.1 μm, increased
to 2.7 ± 0.1 μm during bypass but restored already during CPB towards
a PBR value of 2.3 ± 0.1 μm (ICU). The PBR and perfused vessel density
showed a good correlation (r = –0.65; P = 0.002), demonstrating loss of
perfused vascular density when the glycocalyx is damaged. Conclusion The glycocalyx is damaged after initiation of CPB, and
restoration is impaired after exposure to nonpulsatile fl ow during the
use of the heart–lung machine. Changes in microcirculatory perfusion during cardiac surgery are
paralleled by alterations in glycocalyx integrity
C Boer1, NJ Koning1, J Van Teeff elen2, AB Vonk1, H Vink2
1VU University Medical Center, Amsterdam, the Netherlands; 2Maastricht
University, Maastricht, the Netherlands
Critical Care 2013, 17(Suppl 2):P212 (doi: 10.1186/cc12150) Results During cooling and 1 hour after cooling was ended, a signifi cant
increase in cardiac output (P = 0.028 and P = 0.043) was observed,
but there were no changes in heart rate or mean arterial pressure in
comparison with baseline variables. There were no signifi cant changes
in PPV, MFI, PVD and TVD of small vessels in comparison with baseline
variables during all observational time. Changes in microcirculatory perfusion during cardiac surgery are
paralleled by alterations in glycocalyx integrity
C Boer1, NJ Koning1, J Van Teeff elen2, AB Vonk1, H Vink2
1VU University Medical Center, Amsterdam, the Netherlands; 2Maastricht
University, Maastricht, the Netherlands
Critical Care 2013, 17(Suppl 2):P212 (doi: 10.1186/cc12150) P215 Observational study of the eff ects of age, diabetes mellitus,
cirrhosis and chronic kidney disease on sublingual microvascular
fl ow
T Reynolds1, S Jhanji2, A Vivian-Smith1, RM Pearse1
1Royal London Hospital, London, UK; 2Royal Marsden Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P215 (doi: 10.1186/cc12153) i
Methods In a prospective cohort during 1 year, we followed consecu-
tively enrolled patients admitted immediately postoperative to the
surgical ICU of Hospital Santa Luzia, Brasília, Brazil. Patients were
assigned to two groups: LC >10% and LC ≤10%. The primary outcome
measure was mortality at 7 and 28 days. The secondary outcome
included hospital and ICU length of stay (LOS). Introduction Sidestream dark-fi eld (SDF) imaging is an important
new technology that has been used to demonstrate microcirculatory
abnormalities in a variety of critical illnesses [1]. The microcirculation
is also aff ected by age and chronic comorbidities. However, the eff ect
of these conditions on SDF microcirculatory parameters has not been
well described. p
g
y (
)
Results A total of 417 patients were followed. In total, 50.4% were
male and 83% underwent elective surgery. The mean age was 59 ± 16,
APACHE II score 8 ± 5, SAPS 2 26 ± 11. The mortality at 7 days was 0.95%
(n = 4) and the mortality at 28 days was 2.15% (n = 9), respectively. Hospital mortality was 4.79% (n = 20). Sixty-one percent (n = 255) of
the patients had LC >10% versus 39% (n = 162) with LC ≤10%. Those
who had LC ≤10% were older (62 ± 16 vs. 57 ± 17, P = 0.00) and had
greater APACHE II score (9 ± 6 vs. 7 ± 4, P = 0.00) and SAPS 2 (28 ± 12
vs. 25 ± 10, P = 0.02). There was no diff erence in ICU LOS (5 ± 12 vs. 4 ± 9 days, P = 0.54) and hospital LOS (10 ± 15 vs. 9 ± 11 days, P = 0.48). Initial lactate levels were lower in the group with LC ≤10% (1.1 ± 0.9
vs. 1.9 ± 1.6, P = 0.00); however, mean lactate was higher in 24 hours
(2.0 ± 1.8 vs. 1.0 ± 0.7, P = 0.00). P215 All of the patients who died in the
fi rst 7 days had LC ≤10% (2.46%, n = 4, P = 0.02); this group also had a
higher mortality at 28 days (4.32%, n = 7 vs. 0.78%, n = 2; P = 0.03). The
relative risk for mortality LC ≤10% in 7 and 28 days was 1.02 (95% CI:
1.00 to 1.05) and 5.07 (95% CI: 1.17 to 27.09), respectively. Signifi cant
diff erence was observed in the Kaplan–Meier survival curves for 7 and
28 days (P = 0.01 and 0.02, respectively). The sensibility of LC ≤10% was
100% (95% CI: 51 to 100%) for 7-day mortality and 78% (95% CI: 45 to
94%) for 28-day mortality. The specifi city was 62% (95% CI: 57 to 66%)
for 7-day mortality and 62% (95% CI: 57 to 66%) for 28-day mortality. C
l
i
D
i
i i i l l
l
l l
l
d Methods Sublingual SDF images were obtained from fi ve groups of
20 participants: healthy volunteers under 25 years, healthy volunteers
over 55 years, and stable patients over 55 years with one of diabetes
mellitus (DM), cirrhosis and stage 5 chronic kidney disease (CKD). Microcirculatory parameters [1] between the groups were then com-
pared for signifi cance using ANOVA for parametric data and the
Kruskal–Wallis test for nonparametric data. This was approved by the
local ethics committee. Results All DM patients were type 2, with mean glycated haemoglobin
(HbA1c) of 8.8% (SD 1.7%). Seventeen cirrhotic patients were Child–
Pugh–Turcotte score A and one was score B. For CKD, the mean
estimated glomerular fi ltration rate was 11.5 ml/minute (SD 2.9). Median microvascular fl ow index (MFI) was 2.85 (IQR 2.75 to 3.0) for
participants aged <25, 2.81 (2.66 to 2.97) for those aged >55, 2.88 (2.75
to 3.0) for those with DM, 3.0 (2.83 to 3.0) for those with cirrhosis and
3.0 (2.78 to 3.0) for those with CKD (P for diff erence = 0.14). There were
no signifi cant diff erences in the proportion of perfused vessels and
perfused vessel density between the groups. See Figure 1. Conclusion Despite initial lactate levels, lactate clearance ≤10% proved
to be a good predictor of mortality in 7 and 28 days in surgical patients
admitted in the postoperative period to the ICU. References e e e ces
1. Nguyen HB, et al.: Crit Care Med 2004, 32:1637-1642. 2. Jansen TC, et al.: Am J Respir Crit Care Med 2010, 182:752-756. y p
y
Reference 1. De Backer D, et al.: How to evaluate the microcirculation: report of a round
table conference. Crit Care 2007, 11:R101. Figure 1 (abstract P215). Microcirculatory response to experimentally-induced whole body
heat stress in healthy humans Our data support the use
of intraoperative PBR monitoring as a novel clinical indicator of
microcirculatory perfusion. f
Results Whole body heating resulted in signifi cantly increased heart
rate (P = 0.012) and cardiac output (P = 0.046) in comparison with
baseline variables. One hour after heating was ended, the heart rate S81 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 remained increased (P = 0.012), but cardiac output returned to baseline
values. During all observational time the mean arterial pressure remain
unaltered. There was no signifi cant diff erence in MFI and PPV of small
vessels at the end of heating and 1 hour after heating in comparison
with baseline variables. One hour after heating we observed signifi cant
increase in PVD (P = 0.046) and TVD (P = 0.028) of small vessels
regardless of lack of diff erence at the end of heating (P = 0.753 and
P = 0.075, respectively) in comparison with baseline variables. C
l
i
Wh l b d h
i
i d
d i
d
d
h
i remained increased (P = 0.012), but cardiac output returned to baseline
values. During all observational time the mean arterial pressure remain
unaltered. There was no signifi cant diff erence in MFI and PPV of small
vessels at the end of heating and 1 hour after heating in comparison
with baseline variables. One hour after heating we observed signifi cant
increase in PVD (P = 0.046) and TVD (P = 0.028) of small vessels
regardless of lack of diff erence at the end of heating (P = 0.753 and
P = 0.075, respectively) in comparison with baseline variables. P217 Assessment of the mottling score as a mortality predictor in
critically ill patients
E Bastos de Moura, F Ferreira Amorim, C Darwin Silveira, M Oliveira Maia
Hospital Santa Luzia, Brasília, Brazil
Critical Care 2013, 17(Suppl 2):P217 (doi: 10.1186/cc12155) P215 y
Conclusion Older age, diabetes, and chronic kidney and liver disease
need not be considered confounding factors for comparison of SDF
microcirculatory parameters in the critically ill. Microcirculatory response to experimentally-induced whole body
heat stress in healthy humans Lactate clearance as a determinant of mortality in surgical patients
FF Amorim1, EB Moura2, AR Santana1, FB Soares1, LG Godoy1,
TA Rodrigues1, LJ Almeida1, GM FIlho1, TA Silva1, AP Amorim3, JA Neto2,
MO Maia2
1Escola Superior de Ciências da Saúde, Brasília, Brazil; 2Unidade de Terapia
Intensiva Adulto do Hospital Santa Luzia, Brasília, Brazil; 3Liga Academica de
Medicina Intensiva do Distrito Federal (LIGAMI), Brasilia, Brazil
Critical Care 2013, 17(Suppl 2):P216 (doi: 10.1186/cc12154) Lactate clearance as a determinant of mortality in surgical patients
FF Amorim1, EB Moura2, AR Santana1, FB Soares1, LG Godoy1,
TA Rodrigues1, LJ Almeida1, GM FIlho1, TA Silva1, AP Amorim3, JA Neto2,
MO Maia2 1Escola Superior de Ciências da Saúde, Brasília, Brazil; 2Unidade de Terapia
Intensiva Adulto do Hospital Santa Luzia, Brasília, Brazil; 3Liga Academica de
Medicina Intensiva do Distrito Federal (LIGAMI), Brasilia, Brazil
Critical Care 2013, 17(Suppl 2):P216 (doi: 10.1186/cc12154) Conclusion Whole body heating induced time-dependent changes in
capillary density. Introduction Serial measurements of lactate over time may be a better
prognosticator than a single lactate concentration [1]. Early lactate-
guided therapy also reduces ICU length of stay and ICU and hospital
mortality [2]. This study aims to assess the prognostic value of the
lactate clearance (LC) in the fi rst 24 hours in surgical patients. Introduction Serial measurements of lactate over time may be a better
prognosticator than a single lactate concentration [1]. Early lactate-
guided therapy also reduces ICU length of stay and ICU and hospital
mortality [2]. This study aims to assess the prognostic value of the
lactate clearance (LC) in the fi rst 24 hours in surgical patients. Impact of diff erent haemodynamic resuscitation strategies on
brain perfusion and tissue oedema markers in a model of severe
haemorrhagic shock pi
yf
p
g
y
y
Results Of the 770 patients included in the analysis, 52 died within
30 days. The standard SI at a cutoff point of 0.7 had the highest
predictability and sensitivity of 30-day mortality (area under the curve
(AUC) = 0.7, sensitivity = 0.79, specifi city = 0.56). In comparison, one
of the modifi ed SIs (pulse/diastolic blood pressure) had 0.65 sensitivity
and 0.71 specifi city (AUC = 0.73). A heart rate >100 bpm predicted 30-
day mortality with 0.40 sensitivity and 0.82 specifi city (AUC = 0.63). Conclusion To our knowledge, this is the fi rst study to examine
the relationship between the SI and mortality in patients with a
GI haemorrhage. It appears that the standard SI, when compared
with various modifi ed SIs and conventional vital signs, had the
highest combined predictability and sensitivity of 30-day mortality
in a population of patients suff ering from GI haemorrhage. Further
prospective studies are needed to confi rm these fi ndings. Faculdade de Medicina, Universidade de São Paulo, Brazil i
yf
g
y
y
Results Of the 770 patients included in the analysis, 52 died within
30 days. The standard SI at a cutoff point of 0.7 had the highest
predictability and sensitivity of 30-day mortality (area under the curve
(AUC) = 0.7, sensitivity = 0.79, specifi city = 0.56). In comparison, one
of the modifi ed SIs (pulse/diastolic blood pressure) had 0.65 sensitivity
and 0.71 specifi city (AUC = 0.73). A heart rate >100 bpm predicted 30-
day mortality with 0.40 sensitivity and 0.82 specifi city (AUC = 0.63).i Faculdade de Medicina, Universidade de São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P218 (doi: 10.1186/cc12156) Critical Care 2013, 17(Suppl 2):P218 (doi: 10.1186/cc12156) Introduction This study aimed to compare the cerebral eff ects of
terlipressin (TERLI) with conventional prehospital fl uid resuscitation
with lactated Ringer’s solution (LR) in a model of haemorrhagic shock
(HS). y
y
y
pi
y
Conclusion To our knowledge, this is the fi rst study to examine
the relationship between the SI and mortality in patients with a
GI haemorrhage. It appears that the standard SI, when compared
with various modifi ed SIs and conventional vital signs, had the
highest combined predictability and sensitivity of 30-day mortality
in a population of patients suff ering from GI haemorrhage. Further
prospective studies are needed to confi rm these fi ndings. Impact of diff erent haemodynamic resuscitation strategies on
brain perfusion and tissue oedema markers in a model of severe
haemorrhagic shock Methods Pigs (20 to 30 kg) were randomized into one of the groups:
Sham (n = 2), HS (n = 9), LR (3× volume bled; n = 9) or TERLI (2 mg bolus;
n = 9). HS induced to target MAP of 40 mmHg was maintained for
30 minutes. Brain tissue oxygen pressure (PbtO2), intracranial pressure
(ICP), cerebral perfusion pressure (CPP), haemodynamics and blood gas
analyses were assessed prior to HS (baseline) up to 120 minutes after
treatment. Tissue markers of brain oedema (aquaporin-4 (AQP4) and
Na–K–Cl cotransporter-1 (NKCC1)), apoptosis (pre-apoptotic protein
(Bax)) and oxidative stress (thiobarbituric acid reactive substances
(TBARS)) were also measured. Results Sham animals had no signifi cant changes in the variables
assessed. HS resulted in a signifi cant decrease in CPP (mean varied
from 36 to 39 mmHg), PbtO2 (from 23.6 to 26.6 mmHg), ICP (from 1
to 2 mmHg) and haemodynamics (MAP from 38 to 40 mmHg; CI from
1.8 to 2.1 l/minute/m2), and a signifi cant increase in blood lactate (from
6.7 to 8.9 mmol/l) and cerebral AQP4 (mean ± SE; 167 ± 54% of sham),
NKCC1 (237 ± 47% of sham), Bax (167 ± 44% of sham) and TBARS. Fluid
resuscitation was followed by an increase in ICP (from 7 to 9 mmHg) and
a decrease in CPP (from 41 to 52 mmHg), with an increased expression
of cerebral AQP4 (210 ± 56% of sham), NKCC1 (163 ± 32% of sham) and
Bax (137 ± 24% of sham). Only TERLI restored baseline values of CPP
(from 54 to 61 mmHg) and did not change the cerebral expression of
AQP4 (100 ± 6% of sham), NKCC1 (100 ± 1% of sham), Bax (102 ± 6% of
sham) and TBARS. Both TERLI and LR recovered baseline levels of PbtO2
(TERLI from 30.0 to 34.2 mmHg; LR from 29.4 to 40.7 mmHg) and MAP
(TERLI from 53 to 64 mmHg; LR: 48 to 61 mmHg). Blood lactate levels
were not recovered in any group (TERLI from 5.7 to 8.1 mmol/l; LR from
4.5 to 7.7 mmol/l). Inhaled carbon monoxide or nebulized sodium nitrite protect
against hemorrhagic shock-induced mitochondrial dysfunction
H Gomez, D Escobar, B Ataya, L Gordon, O Ogundele, M Pinsky, S Shiva,
B Zuckerbraun
University of Pittsburgh, PA, USA
Critical Care 2013, 17(Suppl 2):P220 (doi: 10.1186/cc12158) Introduction Shock induces mitochondrial damage, which can lead
to tissue injury and infl ammation. Accuracy of the shock index and various modifi ed shock indexes to
predict early mortality in patients suff ering from gastrointestinal
haemorrhage Introduction The shock index (SI) is an easy-to-use clinical tool that
rapidly identifi es patients at risk of haemodynamic decompensation. Previous studies focused primarily on patients suff ering from
pneumonia, pulmonary embolism, ruptured ectopic pregnancy and
traumatic haemorrhagic shock. Modifi ed SIs have also been studied. Could the SI be accurate in patients suff ering from gastrointestinal (GI)
haemorrhage? The aim of this study was to compare the performance
of the SI with various modifi ed SIs and conventional vital signs in
predicting 30-day mortality in a population of patients with a GI
haemorrhage. Conclusion The mottling score was an objective reproducible system
to bedside use and a good predictor of septic shock mortality. References 1. Hernandez G, et al.: Evolution of peripheral vs metabolic perfusion 1. Hernandez G, et al.: Evolution of peripheral vs metabolic perfusion
parameters during septic shock resuscitation. A clinical-physiologic stu 1. Hernandez G, et al.: Evolution of peripheral vs metabolic perfusion
parameters during septic shock resuscitation. A clinical-physiologic study. J Crit Care 2012, 27:283-288. parameters during septic shock resuscitation. A clinical-physiologic study. J Crit Care 2012, 27:283-288. 2. Ait-Oufella H, et al.: Mottling score predicts survival in septic shock. Intensive
Care Med 2011, 37:801-807. g
Methods A single-center post-hoc analysis was conducted of
prospectively collected data from patients diagnosed with a GI
haemorrhage episode in an academic emergency department (ED)
from March 2008 to December 2011. Data were extracted from two
databases used at our ED. The SI (pulse/systolic blood pressure) and
nine modifi ed SIs were calculated from the available fi rst documented
vital signs. ROC curves were used to determine sensitivity and
specifi city of the diff erent SIs in predicting 30-day mortality. References 1. Cavus E, et al.: Resuscitation 2009, 80:567-572. 2. Bayram B, et al.: Am J Emerg Med 2011, 30:1176-1182. y
Results One hundred and seventeen patients were analyzed; 20 were
excluded (18 terminal illness, two with incomplete data). Ninety-seven
patients were included; the mean age was 72.8 years, mean SAPS II
score was 46.8 (SD ± 15.7), mean APACHE II score was 19.2 (SD ± 8.1);
mean norepinephrine dose was 1.25 μg/kg/minute; mean length of
stay in ICU was 19.2 days (1 to 176); mottling score distribution was:
score 0: 41 patients; score 1: 33 patients; score 2: 14 patients; score 3:
two patients; score 4: three patients; score 5: four patients. The sepsis
origin was as follows: 65, pulmonary (67%); 18, abdominal (18.5%);
nine, urinary (9.5%); two, osseous (2%); one, mediastinal; one, skin and
soft tissue; and one, central nervous system (1% each). Comparing
the mortality in Groups 1, 2 and 3, we found a signifi cant diff erence
(P = 0.042), even greater when considering 28-day mortality (P = 0.004). The Kaplan–Meier survival method showed P = 0.000. P219 P219
Accuracy of the shock index and various modifi ed shock indexes to
predict early mortality in patients suff ering from gastrointestinal
haemorrhage
J St-Cyr Bourque, J Cliche, J Chauny, R Daoust, J Paquet, É Piette
Hôpital du Sacré-Coeur de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P219 (doi: 10.1186/cc12157) Assessment of the mottling score as a mortality predictor in
critically ill patients y
E Bastos de Moura, F Ferreira Amorim, C Darwin Silveira, M Oliveira Maia
Hospital Santa Luzia, Brasília, Brazil
Critical Care 2013, 17(Suppl 2):P217 (doi: 10.1186/cc12155) Introduction The use of peripheral perfusion objective parameters to
anticipate successful resuscitation in septic shock has been recently
investigated [1]. The mottling score, a perfusion parameter used for
decades, has been proposed to correlate with septic shock survival [2],
and was tested in this study as a clinical tool in predicting mortality. Methods A prospective observational study was conducted, with
patients consecutively admitted to a tertiary hospital ICU in Brasília,
Brazil. From July 2011 to May 2012, all patients diagnosed with septic
shock were enrolled. Demographic data, diagnoses, shock origin and
severity scores were recorded. After initial resuscitation, the score
was registered in the fi rst 3 days by the same observer, considering
the score on the lower limb without an arterial catheter, or the worst
between the lower limbs, and the worst in the 3 days. Exclusion criteria
were terminal illness with no intervention decision and incomplete S82 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 contributed to the cerebral apoptosis. None of the treatments caused
cerebral oxidative stress. data. The scores are pooled in Group 1 (scores 0 and 1), Group 2 (scores
2 and 3) and Group 3 (scores 4 and 5) to compare mortality. Statistical
analysis was made using the chi-square test. contributed to the cerebral apoptosis. None of the treatments caused
cerebral oxidative stress. data. The scores are pooled in Group 1 (scores 0 and 1), Group 2 (scores
2 and 3) and Group 3 (scores 4 and 5) to compare mortality. Statistical
analysis was made using the chi-square test. P221 Is norepinephrine more eff ective than other vasopressors for septic
shock? A systematic review and meta-analysis
F Zhou1, Z Peng2, W Zhang3, J Bishop2, Q Song1
1Chinese People’s Liberation Army General Hospital, Beijing, China; 2University
of Pittsburgh, PA, USA; 3University of Pittsburgh School of Medicine, PA, USA
Critical Care 2013, 17(Suppl 2):P221 (doi: 10.1186/cc12159) Is norepinephrine more eff ective than other vasopressors for septic
shock? A systematic review and meta-analysis y
y
Results A total of 300 patients underwent randomization, were
infused with the study drug (148 patients received vasopressin, and
152 norepinephrine), and were included in the analysis. Patients who
received vasopressin had a lower rate of morbidity (23.5% vs. 34%,
P = 0.001) as compared with the norepinephrine group. The 30-day
mortality rate was 6.1% in the norepinephrine group and 4.6% in the
vasopressin group (P = 0.570). There were no signifi cant diff erences in
the overall rates of serious adverse events (7.4% and 6.6%, respectively;
P = 0.772). Introduction Norepinephrine has been widely used in septic
shock. However, its eff ect remains controversial. We conduct a
systematic review and meta-analysis to compare the eff ect between
norepinephrine and other vasopressors. Methods The PubMed, Embase, and Cochrane Library databases
from database inception until October 2012 were searched. We
selected randomized controlled trials in adults with septic shock and
compared norepinephrine with other vasopressors. The quality of
each study included was assessed with Jadad score. After assessing for
heterogeneity of treatment eff ect across trials using the I2 statistic, we
used a fi xed eff ect model (P ≥0.1) or random-eff ects model (P <0.1) and
expressed results as the risk ratio (RR) for dichotomous outcomes or the f Conclusion Vasopressin reduces major morbidity after cardiac surgery
as compared with norepinephrine among patients with cardiac surgery
with vasoplegic shock. p g
Acknowledgement Clinical Trials number NCT01505231. p g
Acknowledgement Clinical Trials number NCT01505231. P223 standardized mean diff erence (SMD) for continuous data with 95% CI. Results Eighteen trials (n = 2,715) met inclusion criteria, which
compared norepinephrine with fi ve diff erent vasopressors (dopamine,
vasopressin, epinephrine, terlipressin and phenylephrine). The mean
Jadad score was 3.11. Overall, there was no diff erence in mortality
in the comparisons between norepinephrine and vasopressin,
epinephrine, terlipressin and phenylephrine (P >0.05, respectively). However, norepinephrine had a trend in decreasing mortality
compared with dopamine (RR, 0.84; 95% CI, 0.68 to 1.02; P = 0.08). P222 11 mg in PBS×30 minutes, n = 6), and sham (n = 3). CO or NaNO2 were
initiated ~30 minutes before resuscitation. Primary endpoints were
changes in muscle and platelet mitochondrial respiration between
BL and EndObs, quantifi ed by muscle respiratory control ratio (RCR,
traditional respirometry), and by the change in proton-leak respiration
(PLR) and mitochondrial reserve capacity in platelets. Secondary
endpoint was mortality at EndObs. P222
Vasopressin Versus Norepinephrine for the Management of Shock
After Cardiac Surgery (VaNCS study): a randomized controlled trial
L Hajjar1, JL Vincent2, A Rhodes3, D Annane4, F Galas1, J Almeida1,
S Zeferino1, L Camara1, V Santos1, J Pereira1, E Osawa1, E Maciel1,
A Rodrigues1, J Jardim1, D Blini1, E Araujo1, F Bergamin5, R Kalil Filho1,
J Auler Jr1
1Heart Institute, São Paulo, Brazil; 2Universite Libre de Bruxelles, Brussels,
Belgium; 3University of London, UK; 4Faculte Medicine Paris, France; 5Cancer
Institute, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P222 (doi: 10.1186/cc12160) Vasopressin Versus Norepinephrine for the Management of Shock
After Cardiac Surgery (VaNCS study): a randomized controlled trial
L Hajjar1, JL Vincent2, A Rhodes3, D Annane4, F Galas1, J Almeida1,
S Zeferino1, L Camara1, V Santos1, J Pereira1, E Osawa1, E Maciel1,
A Rodrigues1, J Jardim1, D Blini1, E Araujo1, F Bergamin5, R Kalil Filho1,
J Auler Jr1
1Heart Institute, São Paulo, Brazil; 2Universite Libre de Bruxelles, Brussels,
Belgium; 3University of London, UK; 4Faculte Medicine Paris, France; 5Cancer
Institute, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P222 (doi: 10.1186/cc12160) y
Results Skeletal muscle RCR decreased in the HSR group (P = 0.04) but
not in sham. Decrease in RCR was primarily due to decreased ADP-
dependent respiration, without change in state 4 respiration. HSR
also resulted in platelet mitochondrial dysfunction as demonstrated
by increased PLR and decreased reserve capacity. This correlated
with increased platelet activation (%CD62P+ by fl ow cytometry) in
HSR. CO or NaNO2 treatment prevented these deleterious changes in
both muscle and platelet mitochondrial respiration, as well as limited
HSR-induced platelet activation. CO treatment also improved reserve
capacity compared with baseline. Mortality was higher in HSR than in
CO or NaNO2 (80 vs. 33 and 33%, respectively). Introduction Vasoplegic syndrome is a common complication after
cardiac surgery, with negative impact on patient outcomes and
hospital costs. Pathogenesis of vasodilatory phenomenon after cardiac
surgery remains a matter of controversy. Loss of vascular tone can
be partly explained by the depletion of neurohypophyseal arginine
vasopressin stores. 1.
Jadad AR, et al.: Control Clin Trials 1996, 17:1-12.
2.
Higgins JP, et al.: BMJ 2003, 327:557-560. Intraaortic counterpulsation in a second-level institution:
indications, management and outcome
R Gómez López, P Fernández Ugidos, P Vidal Cortés, M Lorenzo Lage,
A Tizón Varela, E Rodríguez Álvarez
Complexo Hospitalario Universitario de Ourense, Spain
Critical Care 2013, 17(Suppl 2):P223 (doi: 10.1186/cc12161) Introduction Counterpulsation is an important support for patients
with cardiac diseases. The use of these devices has been limited
typically to hospitals with a cardiac surgery service. The aim of this
study is to describe the management of patients with an intraaortic
balloon pump (IABP) in our second-level institution. Methods An observational study that includes all patients with IABP
in our hospital from January 2010 to September 2012, followed to
November 2012. Epidemiological and clinical variables as an indication
of IABP, therapeutic management and outcome were collected. Because of the small size sample, statistical analysis was limited to
descriptive parameters. Conclusion There is not suffi cient evidence to prove that
norepinephrine is superior to vasopressin, epinephrine, terlipressin
and phenylephrine in terms of mortality. However, norepinephrine is
associated with a decreased HR, cardiac index and an increased SVRI,
and appears to have a greater eff ect on decreasing mortality compared
with dopamine. Results Seventeen patients (76.5% men, age 66.6 ± 11.6 (43 to 85) years,
APACHE score 21.5 ± 14.9 (3 to 46) points) were treated with IABP. The
reason for ICU admission was STEMI (n = 6 patients, 35.3%), no-STEMI
(n = 5, 29.4%), acute heart failure (n = 4, 23.5%) and out-of-hospital Impact of diff erent haemodynamic resuscitation strategies on
brain perfusion and tissue oedema markers in a model of severe
haemorrhagic shock Resuscitative adjuncts to limit
mitochondrial injury may be eff ective to reduce tissue injury and
protect against the sequelae of hemorrhagic shock (HS). Others and we
have demonstrated the protective eff ects of inhaled carbon monoxide
(CO) or nebulized sodium nitrite (NaNO2) in models of HS. Our aim was
to test the hypothesis that CO and NaNO2 protect against hemorrhagic
shock-induced tissue injury/infl ammation by limiting mitochondrial
damage and preventing bioenergetic failure. Methods Twenty anesthetized female Yorkshire pigs were subjected
to severe hemorrhage until unable to compensate or 90 minutes, and
were then resuscitated with volume/pressors. Muscle and platelet
samples were obtained at baseline (BL) and 2 hours after resuscitation
(EndObs). Animals were randomized to: standard of care (HSR, n = 5);
HSR+CO (CO; 250 ppm×30 minutes, n = 6); or HSR+NaNO2 (NaNO2; Conclusion TERLI recovered cerebral perfusion and oxygenation
with no signifi cant changes in ICP and cerebral markers of oedema,
apoptosis and oxidative stress. LR did not recover CPP probably due to
a signifi cant increase in ICP caused by brain oedema, which may have S83 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P222 Vasopressin is commonly used as an adjunct to
catecholamines to support blood pressure in refractory septic shock,
but its eff ect on vasoplegic shock is unknown. We hypothesized that
the use of vasopressin would be more eff ective on treatment of shock
after cardiac surgery than norepinephrine, decreasing the composite
endpoint of mortality and severe morbidity. 2
y
Conclusion In severe HS, mitochondrial injury in platelets and muscle
was limited by CO or NaNO2. Although not powered for a secondary
endpoint, mortality was double in HSR versus adjunctive therapies. This suggests that CO and NaNO2 may protect mitochondrial function
by maintaining ATP-coupled respiration and reserve capacity, and that
this may confer a survival advantage. However, further investigations
are required. Methods In this prospective and randomized, double-blind trial,
we assigned patients who had vasoplegic shock to receive either
vasopressin (0.01 to 0.06 U/minute) or norepinephrine (0.01 to 1 μg/
kg/minute) in addition to open-label vasopressors. All vasopressor
infusions were titrated and tapered according to protocols to maintain
a target blood pressure. The primary endpoint was major morbidity
according to STS (30-day mortality, mechanical ventilation >48 hours,
mediastinitis, surgical re-exploration, stroke, acute renal failure). Secondary outcomes were time on mechanical ventilation, ICU and
hospital stay, new infection, the time to attainment of hemodynamic
stability, occurrence of adverse events and safety. P221 There
were a decreased heart rate (HR) (SMD, –2.10; 95% CI, –3.95 to –0.25;
P = 0.03), cardiac index (SMD, –0.73; 95% CI, –1.14 to –0.03; P = 0.004)
and an increased systemic vascular resistance index (SVRI) (SMD, 1.03;
95% CI, 0.61 to 1.45; P <0.0001) with the treatment of norepinephrine
compared with dopamine.fi Intraaortic counterpulsation in a second-level institution:
indications, management and outcome
R Gómez López, P Fernández Ugidos, P Vidal Cortés, M Lorenzo Lage,
A Tizón Varela, E Rodríguez Álvarez
Complexo Hospitalario Universitario de Ourense, Spain
Critical Care 2013, 17(Suppl 2):P223 (doi: 10.1186/cc12161) 3.
Mayo Clin Proc 2012, 87:620-628. 1.
J Cardiothorac Vasc Anesth 2011, 25:526-535. P224 Results Our urosepsis population contained 364 patients, of which 129
(35.4%) were either using a BB, a CCB or both before their admission. Mean age was 74.9 years and 48.1% of the patients were women. A total
of 36 patients died (9.9%) in a 30-day period. The group taking either BB
or CCB had a signifi cantly lower mean SI (0.76, 95% CI: 0.72 to 0.81 vs. 0.93, 95% CI: 0.89 to 0.98, P <0.0001). In our urosepsis population, a SI
of 0.7 had a sensitivity of 0.76 and a specifi city of 0.27 (area under the
curve (AUC): 0.596) for patients taking neither BB nor CCB. In the group
taking either or both medications, a SI of 0.7 had a sensitivity of 0.57
and a specifi city of 0.42 (AUC: 0.578). In both groups, lowering the SI to
0.5 increased the sensitivity to more than 0.95 but lowered specifi city
signifi cantly.f Impact of left ventricular ejection fraction and elevated fi lling
pressures on mortality in mechanically ventilated patients in severe
sepsis or septic shock P225 Eff ect of β-blockers and calcium channel blockers on shock index
predictability in patients suff ering from urosepsis
J Cliche, J St-Cyr Bourque, R Daoust, J Chauny, J Paquet, E Piette
Sacré-Coeur Hospital, Montreal, Canada
Critical Care 2013, 17(Suppl 2):P225 (doi: 10.1186/cc12163) Eff ect of β-blockers and calcium channel blockers on shock index
predictability in patients suff ering from urosepsis Eff ect of β-blockers and calcium channel blockers on shock index
predictability in patients suff ering from urosepsis f
J Cliche, J St-Cyr Bourque, R Daoust, J Chauny, J Paquet, E Piette
Sacré-Coeur Hospital, Montreal, Canada
Critical Care 2013, 17(Suppl 2):P225 (doi: 10.1186/cc12163) Introduction In our busy emergency departments the identifi cation of
patients at risk of rapid hemodynamic decompensation is crucial. The
shock index (SI) (pulse/systolic blood pressure) is a non-invasive clinical
sign associated with more complications if higher than 0.7. The eff ect
of β-blockers (BB) and calcium channel blockers (CCB) on the SI has
not yet been described. Most studies excluded these patients, owing
to the medications’ eff ect on the cardiac pulse. Considering that BB
and CCB are commonly prescribed, we studied their eff ect on the SI’s
predictability on 30-day mortality in a urosepsis population. Methods This single-center post-hoc analysis of prospectively collected
data was conducted in an academic Canadian emergency department
(ED) between March 2008 and February 2011. Data were extracted
from two institutional databases. We included patients with a fi nal
diagnosis of urosepsis, sepsis, pyelonephritis and urinary tract infection. Selected patients also had a documented positive urine culture. The SI
predictability on 30-day mortality was calculated for patients taking
BB and/or CCB as well as patients taking none of these medications. Sensitivity and specifi city were determined using ROC curves. t tests
were used to compare mean SI between both groups. g
pp
y
Conclusion In our hospital the IABP was mainly used in refractory
cardiogenic shock because of myocardial ischemia, with an all-cause
mortality of 41.2%. Low rates of complications were observed. Transfer
of patients with IABP was performed safely. 2.
Crit Care Med 2009, 37:441-447. P226 Levosimendan in critically ill adults: a utilisation review
J Aron1, S Harrison2, A Milne2, S Patel2, R Maharaj2
1Lewisham NHS Trust, London, UK; 2King’s College Hospital NHS Trust, London,
UK ii
Results All cause 30-day mortality was 50% (n = 29). Forty-six patients
(79%) had normal LV function. Forty-four (76%) patients had normal
LV fi lling pressures. Patients who survived had lower E/e’ ratio, (median
E/e’ 10.6 ± 4 vs. 13.6 ± 7, P = 0.1) but this was not statistically signifi cant. However, when defi ned as E/e’ >15, the mortality was 71%. On the
contrary, patients with low LVEF had a mortality of 41% and LVEF was
no diff erent between survivors and nonsurvivors (55 ± 16 vs. 58 ± 14,
P = 0.34). Introduction Levosimendan improves haemodynamic performance
and may have cardioprotective eff ects. Small trials have demonstrated
effi cacy in well-defi ned populations [1]. Given the uncertain benefi ts
we describe contemporaneous patient selection, utilisation of
levosimedan, haemodynamic eff ects and outcomes. f
Methods The study was performed at a single centre’s three ICUs. Data from 38 consecutive admissions, from 2010 to 2012, treated
with levosimendan were collected retrospectively. Demographics,
illness severity, comorbidity, haemodynamic, metabolic, biochemical,
resource utilisation, organ support and hospital outcomes were
analysed. Conclusion Myocardial dysfunction is a well-known entity in patients
with septic shock. The clinical spectrum of this entity is broad, including
LV, RV and diastolic dysfunction. Although the E/e’ ratio has been a
known prognostic indicator in other cardiac conditions, its role in these
patients is less clear. This study demonstrated that when septal mitral
annulus E/e’ was >15, it was a better marker for mortality than LVEF
in mechanically ventilated patients with severe sepsis or septic shock. Larger studies incorporating diastolic evaluation and TDI should be
performed to further clarify this fi nding. Results Our cohort had a mean age of 58 (95% CI = 53.4 to 64.4). Only
18% underwent cardiothoracic surgery: sepsis, pulmonary emboli and
myocardial/pericardial diseases were also treated with levosimendan. Admission characteristics included mean PaO2/FiO2 ratio (PFR) of
186.7 mmHg (95% CI = 160 to 212) and mean cardiac index (CI) of 2.3
(95% CI 1.9 to 2.8). The median APACHE II score was 24 (interquartile
range 18.5 to 30). Levosimendan was initiated at a variable point
during ICU treatment (days 0 to 14) and was usually the third inotrope References S84 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 cardiac arrest (n = 2, 11.8%). The indications of IABP were refractory
cardiogenic shock (n = 12, 70.6%), high-risk percutaneous coronary
intervention (n = 2, 11.8%), refractory angina (n = 1, 5.9%), refractory
pulmonary edema (n = 1, 5.9%) and electrical storm (n = 1, 5.9%). Five IABPs (27.4%) were inserted in the ICU and the rest in the
catheterization laboratory. Six patients (35.3%) suff ered a cardiac
arrest prior to hemodynamic stabilization. Three patients (17.6%, the
electrical storm and the out-of-hospital cardiac arrest) died before
coronary catheterization was performed and the other three were
treated with mild therapeutic hypothermia. Thirteen patients (76.5%)
needed invasive mechanical ventilation. In six patients (35.3%) invasive
hemodynamic monitoring was performed (one pulmonary artery
catheter, fi ve PICCO). Transthoracic echocardiography was performed
in all patients and transesophageal in six (35.3%). Six of the patients
(35.3%) were transferred to the reference centre for immediate coronary
artery bypass grafting (CABG). No complications were reported during
the transfer. During the ICU stay, femoral artery pseudoaneurysm was
reported in one of the patients and inguinal hematoma after IABP
withdrawal in two (no transfusion required). Four patients (27.4%) died
because of refractory cardiogenic shock despite revascularization. The
length of mechanical support was 1.67 ± 1.5 days (0 to 6). Impact of left ventricular ejection fraction and elevated fi lling
pressures on mortality in mechanically ventilated patients in severe
sepsis or septic shock p
p
S Gillespie, J Pulido
Mayo Clinic, Rochester, MN, USA
Critical Care 2013, 17(Suppl 2):P224 (doi: 10.1186/cc12162) S Gillespie, J Pulido Mayo Clinic, Rochester, MN, USA Introduction Myocardial dysfunction in septic shock is common and
the presentation is broad. There are confl icting data regarding the
prognostic implications of low left ventricular (LV) ejection fraction and
elevated E/e’ ratio on mortality in this patient population. We sought
to assess the impact of LV dysfunction and elevated E/e’ ratio on 30-
day mortality in mechanically ventilated patients with severe sepsis or
septic shock. Conclusion To our knowledge this is the only study analysing the eff ect
of BB and CCB on the SI predictability of 30-day mortality. Our results
indicate that SI cannot be used to accurately predict mortality with
patients suff ering from urosepsis. In both our groups, SI performance
was poor, as shown by the ROC curves. BB or CCB did not infl uence
these results. Methods Fifty-eight mechanically ventilated patients with severe
sepsis or septic shock admitted from 1 August 2007 to 31 January
2009 were prospectively evaluated with transthoracic echocardiogram
within 24 hours of admission. Left ventricular ejection fraction was
assessed using the modifi ed Simpson method as recommended by
the American Society of Echocardiography. Normal LV function was
defi ned as LVEF 40%. Mitral infl ow pulsed wave Doppler of peak E
waves and tissue Doppler imaging (TDI) of the septal mitral annulus
peak velocities were measured, the E/e’ ratio was obtained. Elevated LV
fi lling pressures was defi ned as E/e’ >15. P227 P227
Levosimendan versus dobutamine in Tako-tsubo cardiomyopathy
FR Righetti, MP Parolini, GC Castellano
St. Boniface Hospital Verona, Italy
Critical Care 2013, 17(Suppl 2):P227 (doi: 10.1186/cc12165) and their interaction. Data are reported as mean ± standard deviation. Results Ivabradine decreased the HR from 223 ± 18 to 196 ± 15 bpm
(P <0.05). Contractility and relaxation remained unchanged. Ivabradine
reduced the positive chronotropic eff ect at all doses of dobutamine
(10 nM: 232 ± 37 vs. 187 ± 19, 100 nM: 265 ± 37 vs. 211 ± 40, 1 μM:
316 ± 35 vs. 250 ± 39, 10 μM: 320 ± 33 vs. 235 ± 40 bpm; P <0.05). It
shifted the maximum positive inotropic action of dobutamine to lower
dose ranges (100 nM: 2,924 ± 841 vs. 2,978 ± 955, 300 nM: 3,743 ± 925
vs. 4,795 ± 1,298, 1 μM: 4,138 ± 935 vs. 4,896 ± 1,861 mmHg/second;
P <0.05). A comparable shift was seen for relaxation (100 nM:
–2,178 ± 686 vs. –2,520 ± 742, 300 nM: –2,615 ± 726 vs. –3,150 ± 888,
1 μM: –2,903 ± 752 vs. –2,972 ± 967 mmHg/second; P <0.05). Levosimendan increased the HR only at high doses. With ivabradine, no
positive chronotropic eff ect of levosimendan was observed (100 nM:
185 ± 30 vs. 162 ± 22, 1 μM: 208 ± 28 vs. 166 ± 29, 10 μM: 242 ± 27 vs. 168 ± 36 bpm; P <0.05). Ivabradine attenuated the positive inotropic
eff ect of levosimendan (100 nM: 2,303 ± 303 vs. 1,737 ± 262, 1 μM:
2,977 ± 481 vs. 1,940 ± 449, 10 μM: 3,480 ± 941 vs. 2,189 ± 542 mmHg/
second; P <0.05) but did not signifi cantly alter its lusitropic eff ect. Introduction Tako-tsubo cardiomyopathy is a clinical entity charac-
terized by a dysfunction of the left ventricle, usually transient, which
manifests itself with symptoms that can mimic an acute coronary
syndrome. It is characterized by transient ballooning modifi cation of
the left ventricular apex, probably due to neurogenic stimuli resulting in
low cardiac output syndrome. The aim of this prospective randomized
study was to evaluate, by serial transesophageal echocardiography
(TEE), what is the best drug treatment between levosimendan and
dobutamine to restore a satisfactory cardiac function in the case of low
cardiac output. Methods Twelve adult patients, aged 18 years, were admitted to
the ICU with Tako-tsubo cardiomyopathy at entrance. P228
Combined eff ects of ivabradine with dobutamine or levosimendan
in isolated perfused hearts
D Konrad, R Vicenzi-Moser, M Vicenzi, U Aldenhoff , E Schwarzl, W Toller
Medical University Graz, Austria
Critical Care 2013, 17(Suppl 2):P228 (doi: 10.1186/cc12166) (range 0 to 5) commenced. Treatment with levosimendan (Table 1)
resulted in improved CI and PFR with reductions in EVLW, BE, lactate
and creatinine. The ICU length of stay was 15.4 days (95% CI = 13.2 to
17.6) and the hospital mortality was 54%. No signifi cant adverse eff ects
were reported. (range 0 to 5) commenced. Treatment with levosimendan (Table 1)
resulted in improved CI and PFR with reductions in EVLW, BE, lactate
and creatinine. The ICU length of stay was 15.4 days (95% CI = 13.2 to
17.6) and the hospital mortality was 54%. No signifi cant adverse eff ects
were reported. Introduction Dobutamine and levosimendan improve cardiac
contractility in patients with heart failure, septic cardiomyopathy or
cardiac surgery. Tachycardia in these cases is undesired because of
aggravating ischemia. Ivabradine, a selective sinus node inhibitor,
does not aff ect contractility. A combination of ivabradine with positive
inotropic drugs might be favorable. Thus, we compared the cardiac
eff ects of dobutamine or levosimendan alone, and combined with
ivabradine using the Langendorff method of isolated perfused hearts. Methods Isolated guinea pig hearts (n = 37) were perfused with
incremental doses of dobutamine (10 nM to 10 μM) or levosimendan
(30 nM to 10 μM) either alone or combined with 3 μM ivabradine. Heart rate (HR), left ventricular pressures, contractility (+dLVP/dt) and
relaxation (–dLVP/dt) were recorded. Data for each drug (dobutamine
or levosimendan) were analyzed by two-way ANOVA for repeated
measures including the two main eff ects of ivabradine and drug dose
and their interaction. Data are reported as mean ± standard deviation. Results Ivabradine decreased the HR from 223 ± 18 to 196 ± 15 bpm
(P <0.05). Contractility and relaxation remained unchanged. Ivabradine
reduced the positive chronotropic eff ect at all doses of dobutamine
(10 nM: 232 ± 37 vs. 187 ± 19, 100 nM: 265 ± 37 vs. 211 ± 40, 1 μM:
316 ± 35 vs. 250 ± 39, 10 μM: 320 ± 33 vs. 235 ± 40 bpm; P <0.05). It
shifted the maximum positive inotropic action of dobutamine to lower
dose ranges (100 nM: 2,924 ± 841 vs. 2,978 ± 955, 300 nM: 3,743 ± 925
vs. 4,795 ± 1,298, 1 μM: 4,138 ± 935 vs. 4,896 ± 1,861 mmHg/second;
P <0.05). A comparable shift was seen for relaxation (100 nM:
–2,178 ± 686 vs. –2,520 ± 742, 300 nM: –2,615 ± 726 vs. P227 The patients
were divided randomly into two groups: levosimendan (six patients)
treated with levosimendan and standard treatment, and the control
group (six patients) with dobutamine and standard treatment. In all
patients, serial TEE was performed studying the systolic function, by
ejection fraction of the left ventricle with Simpson’s method. The TEEs
were performed at the entrance of the patient, after 12 and 24 hours
of treatment. The results were expressed as mean with standard
deviation. For the comparison between the two groups we used the
Fisher test, considered signifi cant with P <0.05. Conclusion Addition of ivabradine to dobutamine attenuates its
chronotropic actions without diminishing its inotropic eff ects. A
combination of levosimendan with ivabradine does not seem to
provide benefi t. Clinical studies are necessary to confi rm these
experimental results. P228 P228
Combined eff ects of ivabradine with dobutamine or levosimendan
in isolated perfused hearts
D Konrad, R Vicenzi-Moser, M Vicenzi, U Aldenhoff , E Schwarzl, W Toller
Medical University Graz, Austria
Critical Care 2013, 17(Suppl 2):P228 (doi: 10.1186/cc12166) References Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S85 Table 1 (abstract P226). Treatment outcomes
Variable
Mean
95% CI
CI Δd5 – d0
+1.34
0.4 to 2.2
PFR Δd5 – d0
+284
223 to 345
BE Δd5 – d0
+2.9
0.85 to 5
Lac Δd5 – d0
–1
–2 to 0.2
d0, treatment commenced. Table 1 (abstract P226). Treatment outcomes control group there remains a dysfunction in systolic function. We
have shown the drug therapy based on levosimendan contributes to
improving the systolic function of the left ventricle compared with
treatment with dobutamine despite the initial cardiac stunning. Reference 1. Landoni G, et al.: Eff ects of levosimendan on mortality and hospitalization. A metanalysis of randomized controlled studies. Crit Care Med 2012,
40:634-646. 1. Landoni G, et al.: Eff ects of levosimendan on mortality and hospitalization. A metanalysis of randomized controlled studies. Crit Care Med 2012,
40:634-646. P228
Combined eff ects of ivabradine with dobutamine or levosimendan
in isolated perfused hearts
D Konrad, R Vicenzi-Moser, M Vicenzi, U Aldenhoff , E Schwarzl, W Toller
Medical University Graz, Austria
Critical Care 2013, 17(Suppl 2):P228 (doi: 10.1186/cc12166) –3,150 ± 888,
1 μM: –2,903 ± 752 vs. –2,972 ± 967 mmHg/second; P <0.05). Levosimendan increased the HR only at high doses. With ivabradine, no
positive chronotropic eff ect of levosimendan was observed (100 nM:
185 ± 30 vs. 162 ± 22, 1 μM: 208 ± 28 vs. 166 ± 29, 10 μM: 242 ± 27 vs. 168 ± 36 bpm; P <0.05). Ivabradine attenuated the positive inotropic
eff ect of levosimendan (100 nM: 2,303 ± 303 vs. 1,737 ± 262, 1 μM:
2,977 ± 481 vs. 1,940 ± 449, 10 μM: 3,480 ± 941 vs. 2,189 ± 542 mmHg/
second; P <0.05) but did not signifi cantly alter its lusitropic eff ect. Conclusion Levosimendan is usually used in patients with cardiogenic
shock unresponsive to conventional inotropes or mechanical
augmentation. The mortality of this group is high but represents
patients with shock refractory to conventional treatment. Its use in
sepsis, myocarditis and pulmonary embolism is not well established. In this group levosimendan appears to have a favourable eff ect on
gas exchange, renal function and tissue perfusion. Limitations include
retrospective analysis and missing data. p
y
g
Reference
1. Landoni G, et al.: Eff ects of levosimendan on mortality and hospitilisation. A meta-analysis of RCT. Crit Care Med 2012, 40:634-646. Intraabdominal hypertension and abdominal compartment
syndrome in patients admitted to a medical and surgical ICU at a
third referral university hospital Intraabdominal hypertension and abdominal compartment
syndrome in patients admitted to a medical and surgical ICU at a
third referral university hospital y
p
N Chindavech1, S Yenarkart2, S Kongsayreepong2
1Siriraj Hospital, Mahidol University, Bangkok, Thailand; 2Division of Critical
Care Medicine, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P231 (doi: 10.1186/cc12169) Introduction Intraabdominal hypertension (IAH), especially abdominal
compartment syndrome (ACS), can aff ect organ function leading
to multiple organ failure. Appropriate and prompt management
could improve survival. Less recognition of this problem in critically
ill patients has been reported. The aim of this study was to study the
prevalence, predictive factors and clinical outcome of IAH and ACS in
a mixed population of critically ill patients by intermittent measuring
bladder pressure during the ICU stay. Results The positive inotropic action of dobutamine was least at 37°C,
more pronounced at 40°C and best at 34°C (37°C vs. 40°C vs. 34°C: 300
nM 280 ± 71 vs. 301 ± 94 vs. 345 ± 99%, 1 μM 310 ± 74 vs. 327 ± 92
vs. 389 ± 144%, 10 μM 297 ± 69 vs. 339 ± 96 vs. 359 ± 175%; P <0.05). Dobutamine’s positive lusitropic eff ect was not signifi cantly altered by
temperature. The positive inotropic action of levosimendan was best at
37°C, in hyperthermia and hypothermia only the three highest doses
of levosimendan increased contractility (37°C vs. 40°C vs. 34°C: 100 nM
121 ± 21 vs. 134 ± 15 vs. 87 ± 12%, 300 nM 135 ± 22 vs. 137 ± 17 vs. 94 ± 15%, 1 μM 153 ± 22 vs. 147 ± 21 vs. 108 ± 21%, 3 μM 172 ± 19
vs. 150 ± 24 vs. 115 ± 23%, 10 μM 173 ± 31 vs. 156 ± 28 vs. 117 ± 28%;
P <0.05). The positive lusitropic eff ect of levosimendan at 37°C was
almost absent in hypothermia and hyperthermia (P <0.05). g
y
Methods This prospective observational study was done in 130 adult
patients (age >18 years) admitted to a medical and general surgical
ICU at a third referral university hospital during June to November
2011. Variables about the patient’s profi le laboratory data and clinical
outcome as ICU and hospital length of stay, and ICU, in-hospital and
28-day mortality were recorded. y
y
Results There were 33 (25.4%) medical and 93 (74.6%) surgical ICU
patients in this study. Intraabdominal hypertension and abdominal compartment
syndrome in patients admitted to a medical and surgical ICU at a
third referral university hospital Surgical patients had higher prevalence of IAH
than medical patients (57.8% vs. 33.3%, P = 0.015). Medical patients
were admitted with severe sepsis/septic shock, AKI, pneumonia and
ARDS. Surgical patients were more acutely ill with high ASA (III to IV),
severity score, underwent emergency abdominal surgery and received
more transfusion but were no diff erent in type of fl uid replacement. Signifi cant risk factors of IAH were coagulopathy (OR = 2.09, 95%
CI = 1.62 to 2.69), intraabdominal infection (OR = 1.87, 95% CI = 1.40
to 2.48), retroperitonium hematoma (OR = 1.82, 95% CI = 1.36 to 2.44),
marked ascites (OR = 1.76, 95% CI = 1.32 to 2.36), acidosis (pH <7.2)
(OR = 1.82, 95% CI = 1.37 to 2.43), severe sepsis/septic shock (OR = 1.63,
95% CI = 1.14 to 2.33), and massive transfusion (OR = 1.51, 95%
CI = 1.10 to 2.08). Patients with IAH had more reopened surgeries and
had higher ICU, hospital and 28-day mortality. Sixteen (12.3%) patients
had ACS, 15 patients underwent emergency surgery, two patients
had temporary abdominal closured and one-half of the patients had
severe abdominal sepsis and massive transfusion. Fourteen patients
died despite temporary abdominal closure. Delayed release abdominal
tamponade were most causes of death. Conclusion In isolated perfused hearts, dobutamine has its best
positive inotropic eff ect in hypothermia whereas levosimendan
increases contractility best at normothermic conditions. Clinical studies
are necessary to confi rm these experimental results. Confl icting results between V/Q SPECT and angioscan in pulmonary
embolism: what to do? Confl icting results between V/Q SPECT and angioscan in pulmonary
embolism: what to do? JM Martel, MN Nadeau, RD Daoust, JP Paquet, JC Chauny
Hôpital du Sacré-Coeur de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P230 (doi: 10.1186/cc12168) Confl icting results between V/Q SPECT and angioscan in pulmonary
embolism: what to do? JM Martel, MN Nadeau, RD Daoust, JP Paquet, JC Chauny
Hôpital du Sacré-Coeur de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P230 (doi: 10.1186/cc12168) Introduction The objective of this study is to determine the fi nal
clinical diagnosis of patients who underwent the double investigation
for pulmonary embolism with confl icting results. Pulmonary embolism
is a prevalent pathology in the emergency department (ED). A certain
proportion of patients undergo a double radiological investigation
(V/Q SPECT and angioscan), which incurs higher costs and X-ray doses. However, no study to date has addressed this issue. p
Conclusion The prevalence and morbidity/mortality of IAH and ACS
were high in this institute. Early appropriate and prompt management,
especially fl uid and releasing a tamponade eff ect, were important. P232 P231
Intraabdominal hypertension and abdominal compartment
syndrome in patients admitted to a medical and surgical ICU at a
third referral university hospital
N Chindavech1, S Yenarkart2, S Kongsayreepong2
1Siriraj Hospital, Mahidol University, Bangkok, Thailand; 2Division of Critical
Care Medicine, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P231 (doi: 10.1186/cc12169) Intraabdominal hypertension and abdominal compartment
syndrome in patients admitted to a medical and surgical ICU at a
third referral university hospital
N Chindavech1, S Yenarkart2, S Kongsayreepong2
1Siriraj Hospital, Mahidol University, Bangkok, Thailand; 2Division of Critical
Care Medicine, Bangkok, Thailand
Critical Care 2013, 17(Suppl 2):P231 (doi: 10.1186/cc12169) P229l i
Results Patients in the two groups were statistically comparable with
respect to sex (P = 0.31) and age (P = 0.53). The causes of the syndrome
of Tako-tsubo were: subarachnoid hemorrhage (six patients) after
coronary artery bypass graft (four patients), and polytrauma (two
patients). All patients had low cardiac output. In the levosimendan
group the ejection fraction at entrance was 25 ± 6%, after 12 hours
36 ± 10%, and 47 ± 5% after 24 hours. In the control group the ejection
fraction at entrance was 24 ± 7%, after 12 hours 28 ± 6% and after
24 hours 33 ± 4%. Comparing the two groups we reached statistical
signifi cance, P = 0.026. Infl uence of temperature on the cardiac action of dobutamine and
levosimendan in isolated perfused hearts
U Aldenhoff , R Vicenzi-Moser, M Vicenzi, D Konrad, E Schwarzl, W Toller
Medical University Graz, Austria
Critical Care 2013, 17(Suppl 2):P229 (doi: 10.1186/cc12167) Introduction Inotropic agents (catecholamines or calcium-sensitizers)
are frequently used in hypothermic as well as hyperthermic patient
conditions. Divergent results from animal experiments raise doubt as
to whether they act to the same extent at diff erent body temperatures. Thus, we studied the infl uence of clinically relevant temperatures on
the hemodynamic eff ects of dobutamine and levosimendan using the
Langendorff model of isolated perfused hearts. Conclusion Comparing the two groups, we noticed that both started
from a low cardiac output. However, in the group who used the drug
therapy based on levosimendan we saw a return of systolic function
of the left ventricle to near-normal levels within 24 hours, while in the S86 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods Isolated guinea pig hearts (n = 60) were perfused with
incremental doses (10 nM to 10 μM) of dobutamine or levosimendan
either at normothermic (37°C), hyperthermic (40°C) or hypothermic
(34°C) perfusion conditions. Contractility (+dLVP/dt), relaxation
(–dLVP/dt), left ventricular pressures and heart rate were recorded. Data
with increasing drug dosage were calculated in percent from baseline
for each temperature tested. Data for each drug were analysed by two-
way ANOVA for repeated measures including the two main eff ects of
temperature and drug dose and their interaction. Data are reported as
mean ± standard deviation. P232 Methods This retrospective study included patients who underwent
a double investigation in the ED of Hôpital du Sacré-Coeur de
Montréal from April 2008 to October 2012. Patients were selected
from a computerized database of medical prescriptions (MediaMed
Technologie). Data were then extracted from the patient’s fi les: patient
characteristics, radiology report, diagnosis and treatment. Descriptive
statistics were conducted. 3
Red cell distribution width predicts cardiovascular complications
after high-risk surgery
M Geisen, HD Aya, C Ebm, MA Hamilton, J Ball, M Grounds, A Rhodes,
M Cecconi
St George’s Healthcare NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P232 (doi: 10.1186/cc12170) Red cell distribution width predicts cardiovascular complications
after high-risk surgery
M Geisen, HD Aya, C Ebm, MA Hamilton, J Ball, M Grounds, A Rhodes,
M Cecconi
St George’s Healthcare NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P232 (doi: 10.1186/cc12170) Myocardial injury in critically ill patients admitted with noncardiac
diagnosesfi Hunziker H, et al.: Crit Care 2012, 16:R89. 1. Bazick HS, et al.: Crit Care Med 2011, 39:1913-1921. 2. Hunziker H, et al.: Crit Care 2012, 16:R89. 1. Bazick HS, et al.: Crit Care Med 2011, 39:1913-1921. 2. Hunziker H, et al.: Crit Care 2012, 16:R89. References 1. Jin R, et al.: Circulation 2005, 111:3359-3365. 2. Stamous SC: Ann Thorac Surg 2011, 91:42-48. 1. Jin R, et al.: Circulation 2005, 111:3359-3365. 2. Stamous SC: Ann Thorac Surg 2011, 91:42-48. Myocardial injury in critically ill patients admitted with noncardiac
diagnosesfi g
J Lo1, K Lei2, I Webb2, J Coutts2, J Chambers2, A Griffi ths2, J Smith2,
E Connell3, P Collinson3, J Peacock1, D Treacher2, M Ostermann2
1King’s College London, London, UK; 2Guys & St Thomas Foundation Hospital,
London, UK; 3St George’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P234 (doi: 10.1186/cc12172) Introduction In the critically ill, the incidence of raised cardiac troponin
T (cTnT) levels is high. Although the mechanisms of myocardial injury
are not well understood, raised cTnT levels are associated with increased
mortality. The aim of our study was to determine the incidence,
prevalence and outcome of silent myocardial injury as determined by
raised cTnT levels and concomitant ECG changes in critically ill patients
admitted for noncardiac reasons. Results A total of 119 patients were included. Seventy-six (63%)
patients developed complications postoperatively. Thirty patients
(25.2%) developed cardiovascular complications. These patients had
a higher median RDW than patients without cardiovascular compli-
cations (14.7 vs. 13.8%, P <0.05). RDW on ICU admission was associated
with serum lactate concentration on ICU admission: receiver operating
characteristic analysis showed an area under the curve of 0.68 (SE = 0.06,
P = 0.005; see Figure 1). RDW >14.35% predicted hyperlactataemia with
a sensitivity of 76.0% and a specifi city of 71.1%. ad
tted o
o ca d ac easo s. Methods ECGs were taken and cTnT was measured daily during the fi rst
week and on alternate days during the second week until discharge
from the ICU or death. After completion of the study, all cTnT levels and
ECGs were analysed independently and patients were classifi ed into
four groups: defi nite MI (cTnT ≥15 ng/l and defi nite ECG changes of MI),
possible MI (cTnT ≥15 ng/l and ischaemic changes on ECG), troponin
rise alone (cTnT ≥15 ng/l with no ischaemic ECG changes), or normal. All medical notes were reviewed independently by two ICU clinicians. Results A total of 144 patients were included in the analysis (42%
female; mean age 61.9 (SD 16.9); mean APACHE II score 19.4). In total,
121 patients (84%) had at least one cTnT level ≥15 ng/l during their
stay in the ICU. Twenty patients (14%) fulfi lled criteria for a defi nite MI,
of whom 65% were septic and 50% were on noradrenaline at the time
(ICU and hospital mortality: 25% and 30%, respectively). P234 Myocardial injury in critically ill patients admitted with noncardiac
diagnoses
J Lo1, K Lei2, I Webb2, J Coutts2, J Chambers2, A Griffi ths2, J Smith2,
E Connell3, P Collinson3, J Peacock1, D Treacher2, M Ostermann2
1King’s College London, London, UK; 2Guys & St Thomas Foundation Hospital,
London, UK; 3St George’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P234 (doi: 10.1186/cc12172) Consequences of obesity in outcomes after cardiac surgery:
analysis of the ARIAM registry EC Curiel-Balsera1, J Muñoz-Bono1, MJ Delgado-Amaya1, ,
,
g
y ,
R Hinojosa-Pérez2, A Reina-Toral3, A Gordillo-Brenes4, R Rivera-Fernández1
1Hospital Regional Carlos Haya, Málaga, Spain; 2Virgen del Rocio Hospital,
Seville, Spain; 3Virgen de las Nieves Hospital, Granada, Spain; 4Puerta del mar
Hospital, Cádiz, Spain Conclusion Eighty-four per cent of critically ill patients had a raised
cTnT level at some stage during their stay in the ICU. More than 40% of
patients fulfi lled criteria for a possible or defi nite MI, of whom only 20%
were recognised clinically. ICU and hospital outcome were signifi cantly
worse in patients with a cTnT rise. The proportion of patients with sepsis
was similar between the patients with a defi nite, possible or no MI. Critical Care 2013, 17(Suppl 2):P233 (doi: 10.1186/cc12171) Critical Care 2013, 17(Suppl 2):P233 (doi: 10.1186/cc12171) Introduction Obesity is a disease that aff ects a large part of the
population and has been associated with worse outcomes after cardiac
surgery. The aim of our study is to evaluate the consequences of
obesity related to postoperative complications, hospital length of stay
and mortality. Red cell distribution width predicts cardiovascular complications
after high-risk surgery Results In all, 125 patients underwent the double investigation. Our
sample had a mean age of 63.1 years (SD ±18.6) and was composed of
82 (65.6%) women. One hundred and fi fteen patients (92%) underwent
the V/Q SPECT fi rst. The result of 66 (52.8%) SPECT was intermediate
or indeterminate. The fi nal diagnosis was pulmonary embolism for
23 patients (18.4%). A signifi cant proportion of patients (19, 38.0%)
had confl icting results with the two tests. In this subpopulation, four
(21.1%) had a fi nal diagnosis of pulmonary embolism. In the 16 patients
with a result of high probability V/Q SPECT and negative angioscan,
one (6.3%) had a fi nal diagnosis of pulmonary embolism, but three
(100%) with low probability SPECT and positive angioscan were given
this fi nal diagnosis. Introduction The red-cell distribution width (RDW) is associated
with cardiovascular morbidity and mortality and is a predictor of ICU
survival. It was the aim of this study to investigate the potential of RDW
to predict postoperative cardiovascular complications (new onset of
treated arrhythmias, myocardial ischaemia or initiation of vasopressor
support) and its association with markers of tissue perfusion (serum
lactate >2.5 mmol/l). Methods Analysis of prospectively recorded data for a register of
patients admitted to an 18-bed ICU in a large teaching hospital
after undergoing high-risk surgery. Haemodynamic and laboratory
parameters during the fi rst 12 hours of ICU stay were recorded as
well as demographic characteristics. Assessment for postoperative
complications was performed using the postoperative morbidity
survey and the Clavien-Dindo classifi cation. In addition, clinical
outcome data (hospital mortality, length of ICU stay, length of hospital
stay, readmission to the ICU) were recorded. i
Conclusion Most patients underwent the double investigation
because of intermediate or indeterminate V/Q SPECT results. In case
of confl icting results, clinicians based their decision on the angioscan. In future studies, it would be useful to identify contributing factors for
this discordance. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2 S87 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P232). Prediction of hyperlactataemia according to
RDW. in the development of overall postoperative complications (33% in
obese and 35.8% in nonobese, P = 0.07), although less appreciated
were reoperation rate or stroke, as well as further development of
postoperative renal failure. Red cell distribution width predicts cardiovascular complications
after high-risk surgery After adjusting for severity and length
of cardio bypass time, obese patients had lower mortality without
being statistically signifi cant, OR = 0.94 (0.79 to 1.04). There were no
diff erences in ICU length of stay, but obese patients had greater ward
length of stay, 9.04 (10.43) versus 1.18 (9.2) days, P = 0.01. in the development of overall postoperative complications (33% in
obese and 35.8% in nonobese, P = 0.07), although less appreciated
were reoperation rate or stroke, as well as further development of
postoperative renal failure. After adjusting for severity and length
of cardio bypass time, obese patients had lower mortality without
being statistically signifi cant, OR = 0.94 (0.79 to 1.04). There were no
diff erences in ICU length of stay, but obese patients had greater ward
length of stay, 9.04 (10.43) versus 1.18 (9.2) days, P = 0.01. Conclusion Obese patients undergoing cardiac surgery have mortality,
rate of complications and length of stay similar to nonobese patients. Obese patients required less reoperation but developed more frequent
postoperative renal failure. R f Myocardial injury in critically ill patients admitted with noncardiac
diagnosesfi Thirty-nine
patients (27%) had a possible MI, of whom 69% were septic and on
noradrenaline (ICU and hospital mortality: 31% and 41%, respectively). Sixty-two patients (43%) had a raised troponin without ECG, of whom
69% were septic and 50.7% were on noradrenaline (ICU and hospital
mortality: 23% and 31%, respectively). Twenty-three patients had
normal cTnT results and serial ECGs, of whom 61% had sepsis. ICU
and hospital mortality was 4%. Only 25% of defi nite MIs and 18% of
possible MIs were recognised by the clinical teams at the time. Methods ECGs were taken and cTnT was measured daily during the fi rst
week and on alternate days during the second week until discharge
from the ICU or death. After completion of the study, all cTnT levels and
ECGs were analysed independently and patients were classifi ed into
four groups: defi nite MI (cTnT ≥15 ng/l and defi nite ECG changes of MI),
possible MI (cTnT ≥15 ng/l and ischaemic changes on ECG), troponin
rise alone (cTnT ≥15 ng/l with no ischaemic ECG changes), or normal. All medical notes were reviewed independently by two ICU clinicians. y
pi
y
Conclusion RDW is a potential parameter for perioperative risk
stratifi cation. Further studies will have to elucidate the ability of acute
elevations in RDW to refl ect impaired tissue perfusion. References p
y y
Results A total of 144 patients were included in the analysis (42%
female; mean age 61.9 (SD 16.9); mean APACHE II score 19.4). In total,
121 patients (84%) had at least one cTnT level ≥15 ng/l during their
stay in the ICU. Twenty patients (14%) fulfi lled criteria for a defi nite MI,
of whom 65% were septic and 50% were on noradrenaline at the time
(ICU and hospital mortality: 25% and 30%, respectively). Thirty-nine
patients (27%) had a possible MI, of whom 69% were septic and on
noradrenaline (ICU and hospital mortality: 31% and 41%, respectively). Sixty-two patients (43%) had a raised troponin without ECG, of whom
69% were septic and 50.7% were on noradrenaline (ICU and hospital
mortality: 23% and 31%, respectively). Twenty-three patients had
normal cTnT results and serial ECGs, of whom 61% had sepsis. ICU
and hospital mortality was 4%. Only 25% of defi nite MIs and 18% of
possible MIs were recognised by the clinical teams at the time. 1. Bazick HS, et al.: Crit Care Med 2011, 39:1913-1921. 2. P235 Methods An observational, prospective, multicenter study of patients
included in the ARIAM registry of adult cardiac surgery between March
2008 and March 2011. We analyzed clinical variables, the surgical
procedure, postoperative complications and mortality, comparing
the group of patients with body mass index (BMI) greater or less than
30 kg/m2. P235
Validation of the GRACE score risk in our population
L Sayagues1, J Sieira1, E Abbu2, J Chico3, C Pena3, J Gonzalez Juantey2
1HULA, Lugo, Spain; 2CHUS, Santiago de Compostela, Spain; 3CHUVI, Vigo,
Spain
Critical Care 2013, 17(Suppl 2):P235 (doi: 10.1186/cc12173) P235
Validation of the GRACE score risk in our population
L Sayagues1, J Sieira1, E Abbu2, J Chico3, C Pena3, J Gonzalez Juantey2
1HULA, Lugo, Spain; 2CHUS, Santiago de Compostela, Spain; 3CHUVI, Vigo,
Spain
Critical Care 2013, 17(Suppl 2):P235 (doi: 10.1186/cc12173) 35
Validation of the GRACE score risk in our population Validation of the GRACE score risk in our population
L Sayagues1, J Sieira1, E Abbu2, J Chico3, C Pena3, J Gonzalez Juantey2
1HULA, Lugo, Spain; 2CHUS, Santiago de Compostela, Spain; 3CHUVI, Vigo,
Spain
C iti
l C
2013 17(S
l 2) P235 (d i 10 1186/
12173) p p
L Sayagues1, J Sieira1, E Abbu2, J Chico3, C Pena3, J Gonzalez Juantey2
1HULA, Lugo, Spain; 2CHUS, Santiago de Compostela, Spain; 3CHUVI, Vigo,
Spain Critical Care 2013, 17(Suppl 2):P235 (doi: 10.1186/cc12173 Results The study included 4,712 patients with a mean age of 64.03 (SD
±12.08) years, BMI 28.53 (SD ±4.7) and EuroSCORE 5.58 (SD ±2.91). In
1,940 patients (35.7%) BMI was >30 kg/m2. There were no diff erences Introduction The Global Registry of Acute Coronary Events (GRACE)
risk score provides an estimation of the probability of death within S88 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 6 months of hospital discharge in patients with acute coronary
syndrome (ACS). Our aim was to assess the validity of this risk score
in our contemporary cohort of patients admitted to our third-level
hospital. Figure 2 (abstract P236). Incomplete and complete revascularization. p
Methods The study involved 1,185 consecutive patients with ACS
evaluated between February 2004 and February 2009. Their virtual
status was determined 6 months after hospital discharge and the
validity of the GRACE risk score was evaluated. y
Results In total, 450 (38.8%) patients were admitted for STEMI and 725
(61.2%) for NSTEMI. Percutaneous revascularisation was performed
in 846 (71.5%). p
Critical Care 2013, 17(Suppl 2):P236 (doi: 10.1186/cc12174) Introduction This is a prospective study in which all ACS cases
attending a level 3 hospital were collected consecutively from February
2004 to 2010, and were clinically followed-up until 2012. Introduction In patients with severe and inoperable aortic stenosis,
some trials have proven that TAVI excels standard procedures and has
also been proven less cost-eff ective. We aim at reviewing short-term
results after 2 years since the implementation of this technique in our
premises. y
Methods Of a total 4,589 cases, only 2,515 were revascularized with ICP
(1,742 complete and 907 incomplete; 160 failed). We split the cases into
two main groups: STEMI and NSTEMI. p
Methods A study of a retrospective cohorts of patients who underwent
isolated aortic valve replacement (AVR) by either conventional
surgery or TAVI (CoreValve device) from June 2010 to December 2011
at the University Hospital Carlos de Haya (Málaga, Spain). Clinical
epidemiologic, complication and short-term outcome variables were
registered. Qualitative variables are expressed as percentages, while
quantitative variables are expressed as means and SD. Fisher’s exact
test and Mann–Whitney’s U-test were used where necessary (5%
maximum error ratio). g
p
Results We observed a 0.6 higher accumulative survival rate in patients
with complete ICP over patients who underwent surgery, incomplete
ICP or mixed treatment. Those with conservative measures solely have,
obviously, worst prognosis. See Figures 1 and 2. Figure 1 (abstract P236). FEVI and prognosis. Results A total number of 27 TAVI and 154 isolated AVR procedures
were completed. Intervention typology was chosen according to the
recommendations of scientifi c societies, apart from patients’ fulfi llment
of the anatomic criteria required for percutaneous implant. Mean age
was 67 ± 11 years (54% males) in AVR and 80 ± 6 years (44% males)
in TAVI (P <0.05). The additive EuroSCORE in AVR was 7 ± 2 and 9 ± 2
in TAVI (P <0.05). However, 55.6% of the percutaneous-valve patients
presented previous coronary-tree alterations with stent implantation,
while only 7% of AVR patients showed these alterations (P <0.001). ICU
mortality in TAVI and AVR patients was 3.7% and 8.2%, respectively
(P = NS). Regarding complications, 48.1% of TAVI patients showed
altered heart rhythm and 33% required a permanent pacemaker. Electrical disorders were observed in 4% of AVR patients, while 1.9%
of these patients required a permanent pacemaker (P <0.001 for both). Reoperation was necessary in 14.8 and 1.9% of TAVI and AVR patients,
respectively (P <0.001). P235 The median GRACE risk score was 121 (interquartile
range 96/144). Mortality after discharge was 4.4%. The calibration of
the GRACE score was Hosmer–Lemeshow P >0.2 and its discriminatory
capacity was excellent. Area under the ROC curve was 0.86, 95% CI
0.807 to 0.916, in all patients. See Table 1. Table 1 (abstract P235)
GRACE risk
Total
STEMI
NSTEMI
Low
270 (22.7)
177 (38.3)
93 (12.8)
Medium
334 (28.2)
141 (30.7)
193 (26.7)
High
580 (49)
142 (30.9)
438 (60.5) Figure 2 (abstract P236). Incomplete and complete revascularization. Figure 2 (abstract P236). Incomplete and complete revascularization. Conclusion In our study patients undergoing ICP have higher survival
rates in comparison with cardiac surgery except those >65 years old
and diabetic groups in which cardiac surgery has higher ratios than ICP. In the rest of the groups, no matter how many coronary arteries were
aff ected, only those with complete ICP present higher survival rates. It
would be important to repeat this study in a multicentric cohort. Conclusion The GRACE risk score for predicting death within 6 months
of hospital discharge was validated and can be used in patients with
ACS. It would be perfect in the future to include the GRACE risk score
in the medical records of this type of patients. Also it would be very
interesting to validate this in a multicentric study. Non-invasive valve implantation with TAVI versus conventional
aortic valve replacement Non-invasive valve implantation with TAVI versus conventional
aortic valve replacement
E Trujillo-García, C Joya-Montosa, MJ Delgado-Amaya, E Curiel-Balsera
Hospital Regional Carlos Haya, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P237 (doi: 10.1186/cc12175) p
p
L Sayagues1, J Sieira1, E Abbu Assi2, J Chico3, C Pena3, J Gonzalez Juanatey2
1HULA, Lugo, Spain; 2CHUS, Santiago de Compostela, Spain; 3CHUVI, Vigo,
Spain aortic valve replacement
E Trujillo-García, C Joya-Montosa, MJ Delgado-Amaya, E Curiel-Balsera
Hospital Regional Carlos Haya, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P237 (doi: 10.1186/cc12175) P238
Time to recover from shock is determinant of a positive fl uid
balance in septic shock
S Lobo, AL Cunha
Faculdade de Medicina de São José do Rio Preto, Brazil
Critical Care 2013, 17(Suppl 2):P238 (doi: 10.1186/cc12176) Introduction Excess fl uids may be harmful in critically ill patients. We aimed to evaluate the cumulative fl uid balance during 7 days in
patients with septic shock after recovery from shock. Methods A prospective and observational study in septic shock
patients. Patients with MAP >65 mmHg and lactate <2.0 mEq/l were
included <12 hours after weaning from vasopressors. Daily fl uid
balance was registered during 7 days after the enrollment. Patients
were divided into two groups according to the full cohort’s median
cumulative fl uid balance administered during the period of shock (use
of vasopressors) calculated on study day 1: Group 1 ≤4.4 l (n = 20) and
Group 2 >4.4 l (n = 20). i
Results Seventy-nine patients were enrolled. Fifty-one patients
were with moderate UGIB, and 28 patients were with severe UGIB. As indicated in Tables 1 and 2, there was a signifi cant relationship
between CLac and sustained bleeding in moderate UGIB (P = 0.02). On
the other hand, there was no signifi cant relationship between CLac and
sustained bleeding in severe UGIB (P = 0.58). p
Results Cumulative fl uid balance was 1.6 ± 1.8 l in Group 1 and
10.2 ± 4.1 l in Group 2 and 8.5 ± 5.3 l in Group 1 and 18.5 ± 7.9 l in
Group 2 on study day 8 (P <0.001 for both). Time for recovery from
shock was predictive of receiving larger volume of fl uids (OR: 1.38, 95%
CI: 1.08 to 1.75, P = 0.009). After zeroing fl uid balance on study day 2,
7 days cumulative fl uid balance continues to increase in both groups
(Figure 1). Patients in Group 2 had more prolonged length of stay in the
ICU and in hospital than patients in Group 1. Table 1 (abstract P239). Relationship between lactate clearance and
sustained bleeding in moderate UGIB (n = 51) Table 1 (abstract P239). Relationship between lactate clearance and
sustained bleeding in moderate UGIB (n = 51) Sustained bleeding
Nonsustained bleeding
(n = 19)
(n = 32)
P value
CLac <0.5
18 (95%)
21 (65%)
0.02
CLac ≥0.5
1 (5%)
11 (34%) Conclusion After recovery from septic shock we notice a huge accu-
mulated fl uid balance. A more positive fl uid balance was associated
with a more prolonged length of stay in the ICU and in the hospital. P239
Lactate clearance is a predictor of sustained bleeding in emergency
room patients with moderate upper gastrointestinal bleeding
T Wada1, A Hagiwara2, N Yahagi1, A Kimura2
1University of Tokyo, Japan; 2National Center for Global Health and Medicine,
Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P239 (doi: 10.1186/cc12177)
Figure 1 (abstract P238). Figure 1 (abstract P238). Table 2 (abstract P239). Relationship between lactate clearance and
sustained bleeding in severe UGIB (n = 28)
Sustained bleeding
Nonsustained bleeding
(n = 6)
bleeding (n = 22)
P value
CLac <0.5
4 (67%)
18 (82%)
0.58
CLac ≥0.5
2 (33%)
4 (18%)
Conclusion If an initial fl uid therapy for moderate UGIB results in high
CL
h bl
di
i h
l
d h
d F
h
i Table 2 (abstract P239). Relationship between lactate clearance and
sustained bleeding in severe UGIB (n = 28) Conclusion If an initial fl uid therapy for moderate UGIB results in high
CLac, the bleeding might already have stopped. For such a patient, we
may save urgent endoscopy. p
Critical Care 2013, 17(Suppl 2):P236 (doi: 10.1186/cc12174) y
Conclusion Even with our limited experience, TAVI patients are
observed to be older, to involve higher surgical risk, and to have
undergone previous coronary-tree interventions. Although no Figure 1 (abstract P236). FEVI and prognosis. S89 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 signifi cant diff erences were found regarding mortality, a higher rate
of complications was observed in TAVI patients. With no short-term
diff erences, a signifi cant rate of postsurgical complications, and a cost-
effi ciency handicapped technique, analysis of long-term outcomes
seems necessary to assess TAVI’s advantages over conventional AVR. UGIB patient needs an intervention or not. However, the intervention
which the GBS mentions includes not only endoscopy but also blood
transfusion. Therefore, we cannot determine whether a UGIB patient
needs urgent endoscopy or just blood transfusion by GBS alone. We
hypothesized that high lactate clearance (CLac) would decrease the
likelihood of sustained UGIB. Methods This is a retrospective study. UGIB patients, who visited the
emergency department (ED) of the National Center for Global Health
and Medicine from April 2011 to March 2012 and received urgent
endoscopy in the ED, were enrolled. We collected for each patient
the GBS, the blood lactate value on arrival in the ED, the blood lactate
value after bolus administration of 20 to 40 ml/kg Ringer’s acetate
(initial fl uid therapy) and the report of urgent endoscopy. We classifi ed
the severity of UGIB according to GBS. A score ≤12 was classifi ed as
moderate, and a score ≥13 was classifi ed as severe. CLac was defi ned
as the percentage decrease in blood lactate from the time of arrival
in the ED to the time when an initial fl uid therapy was fi nished. CLac
<50% was defi ned as low, and CLac ≥50% was defi ned as high. Whether
a patient had sustained bleeding or not was determined based on
the report of urgent endoscopy. The relationship between CLac and
sustained bleeding was examined by Fisher’s exact test, and P <0.05
was considered statistically signifi cant. P238 P238
Time to recover from shock is determinant of a positive fl uid
balance in septic shock
S Lobo, AL Cunha
Faculdade de Medicina de São José do Rio Preto, Brazil
Critical Care 2013, 17(Suppl 2):P238 (doi: 10.1186/cc12176) P240 P240
Muscular glucose assessed by microdialysis and blood glucose can
predict mortality in septic shock
A Massoudi, M Belhadj Amor, C Romdhani, A Ben Gabsia, I Labbene,
M Ferjani
Military Hospital of Tunis, Tunisia
Critical Care 2013, 17(Suppl 2):P240 (doi: 10.1186/cc12178) predict mortality in septic shock
A Massoudi, M Belhadj Amor, C Romdhani, A Ben Gabsia, I Labbene,
M Ferjani
Military Hospital of Tunis, Tunisia
Critical Care 2013, 17(Suppl 2):P240 (doi: 10.1186/cc12178) Conclusion After recovery from septic shock we notice a huge accu-
mulated fl uid balance. A more positive fl uid balance was associated
with a more prolonged length of stay in the ICU and in the hospital. Introduction The aim of our study was to assess the muscular glucose
by microdialysis and its association with mortality in septic shock
patients. Introduction The aim of our study was to assess the muscular glucose
by microdialysis and its association with mortality in septic shock
patients. P239
Lactate clearance is a predictor of sustained bleeding in emergency
room patients with moderate upper gastrointestinal bleeding
T Wada1, A Hagiwara2, N Yahagi1, A Kimura2
1University of Tokyo, Japan; 2National Center for Global Health and Medicine,
Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P239 (doi: 10.1186/cc12177) Methods We conducted a preliminary prospective study. We included
septic shock patients hemodynamically optimized according to
international recommendations. A microdialysis catheter was inserted
in the femoral quadriceps. Interstitial fl uid samples were collected
every 6 hours for 5 days. The determination of muscular glucose was
performed by the CMA 600 analyzer (CMA/Microdialysis AB, Sweden). We also performed a dosage of concomitant blood glucose. The
study population was divided into two groups according to hospital
mortality. Statistic analysis: Mann–Whitney test and chi-squared test: Introduction There is no useful predictor of sustained upper gastro-
intestinal bleeding (UGIB). Glasgow–Blatchford scoring (GBS) is based
on simple clinical and laboratory variables and can predict whether a S90 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 comparisons between groups. Quantitative variables were expressed
as mean ± standard deviation or median (interquartile range) as
appropriate. Nasogastric tube dislodgement: a problem on our ICU
B Morton, R Hall, T Ridgway, O Al-Rawi
Liverpool Heart & Chest, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P242 (doi: 10.1186/cc12180) Results We included 12 patients with septic shock. The mortality rate
was 50%. P240 Demographics were comparable between groups except
for age (66 ± 9 vs. 41 ± 12, dead patients vs. survivors, respectively;
P = 0.002). Pneumonia was the major cause of septic shock (10 patients). We analysed 167 blood samples and 166 muscular glucose samples. We found a positive association between muscular glucose, blood
glucose and mortality. Tissue glucose was signifi cantly higher among
dead patients compared with survivors at the 54th hour. Comparing all
data, muscular glucose (P = 0.02) and blood glucose (P = 0.007) were
signifi cantly higher in dead patients (Table 1). Introduction It was noted on our unit that dislodgement of nasogastric
tubes occurred commonly. This can lead to an increased risk of
aspiration, interruptions in nutritional support, skin breakdown and
radiographic exposure [1]. It is recommended that the position of
nasogastric tubes should be confi rmed by aspiration and pH testing,
with radiographic confi rmation used only when this is not possible [2]. Methods We performed a retrospective review of chest X-ray (CXR)
requests for the 3-month period June to August 2012 using the trust
radiology information system. The proportion of CXR requests for
confi rmation of position and patient demographics were measured
with an estimation of the fi nancial cost performed. Table 1 (abstract P240). Association between muscular glucose, blood
glucose and mortality
Dead patients
Survivors
(n = 6)
(n = 6)
P value
Muscular glucose (mmol/l)
9.4 (5.66; 13.71)
7.87 (5.62; 10.41)
0.02
Blood glucose (mmol/l)
10.9 (8.3; 15.5)
8.6 (6.9; 11.4)
0.007 Table 1 (abstract P240). Association between muscular glucose, blood
glucose and mortality i
Results There were 541 patients admitted to the critical care area in
the study period. In total, 207 out of 2,340 (8.8%) CXRs performed were
for confi rmation of position. Repeated X-rays were required in some
patients (see Table 1); these patients were older and tended to have
a longer length of stay. A mobile CXR costs £25 in our trust, if one CXR
is accepted per patient with a nasogastric tube; there was an excess of
160 images with a cost of £4,000 in the 3-month period. Conclusion Our data suggest that muscular glucose assessed by
microdialysis and blood glucose are associated with mortality in septic
shock patients. Therefore, muscular glucose may refl ect the metabolic
alterations and microcirculatory dysfunction induced by septic shock. y
References References 1. Lorente L, et al.: Accidental catheter removal in critically ill patients: a 1. Lorente L, et al.: Accidental catheter removal in critically ill patients: a prospective and observational study. Crit Care 2004, 8:R229-R233. 2. National Patient Safety Agency: Reducing Harm Caused by the Misplacement of
Nasogastric Feeding Tubes. NPSA/2011/PSA002. NHS; 2011. [http://www.nrls.npsa.nhs.uk/alerts/?entryid45=129640] Conclusion There was a high prevalence of constipation among critical
care patients with poor adherence to the bowel protocol, which requires
improvements in staff awareness and new recommendations (drug chart
amendment, and so forth). Duration of mechanical ventilation and failure
to feed were greater in constipated than nonconstipated patients. This
may indicate a common correlation and further studies are warranted. Due to the possible signifi cant impact of constipation on patients’
recovery, each critical care unit should introduce a bowel protocol or
ensure compliance with the existing one before the evidence is clearly
established. f Handling of dietary protein in critically ill patients Handling of dietary protein in critically ill patients Handling of dietary protein in critically ill patients
F Liebau1, O Rooyackers1, L Van Loon2, J Wernerman1
1Karolinska University Hospital Huddinge, Stockholm, Sweden; 2NUtrIm Handling of dietary protein in critically ill patients
F Liebau1, O Rooyackers1, L Van Loon2, J Wernerman1
1Karolinska University Hospital Huddinge, Stockholm, Sweden; 2NUtrIm,
Maastricht University, Maastricht, the Netherlands
Critical Care 2013, 17(Suppl 2):P243 (doi: 10.1186/cc12181) g
y
y
Liebau1, O Rooyackers1, L Van Loon2, J Wernerman1 ,
y
,
,
1Karolinska University Hospital Huddinge, Stockholm, Sweden; 2NUtrIm
M
i h U i
i
M
i h
h N h l
d Maastricht University, Maastricht, the Netherlands
Critical Care 2013, 17(Suppl 2):P243 (doi: 10.1186/cc12181) y
Critical Care 2013, 17(Suppl 2):P243 (doi: 10.1186/cc12181) Introduction Protein turnover measurements by stable isotope
techniques can be applied to assess the nutritional/metabolic status in
critically ill patients and their response to feeding. Because of uncertain
gastrointestinal transport and uptake of nutrients, their contribution
needs to be measured separately. We studied whole-body protein
kinetics, with special emphasis on the contribution of dietary protein,
in ICU patients and healthy controls. Introduction Protein turnover measurements by stable isotope
techniques can be applied to assess the nutritional/metabolic status in
critically ill patients and their response to feeding. Because of uncertain
gastrointestinal transport and uptake of nutrients, their contribution
needs to be measured separately. We studied whole-body protein
kinetics, with special emphasis on the contribution of dietary protein,
in ICU patients and healthy controls. 2.
Palácio de Azevedo R: Intestinal constipation in intensive care unit. Rev Bras
Ter Intensiva 2009, 21:324-331. P241
Constipation in critically ill patients and its relationship to feeding
and weaning from respiratory support g
p
E Spodniewska, A Guha p
Royal Liverpool University Hospital, Liverpool, UK y
p
y
p
p
Critical Care 2013, 17(Suppl 2):P241 (doi: 10.1186/cc12179) Critical Care 2013, 17(Suppl 2):P241 (doi: 10.1186/cc12179) Introduction Constipation can be defi ned as a failure of the bowels to
open for 3 consecutive days. It is a common problem in intensive care
settings but not many studies have so far raised the subject. Methods The audit had the Trust Audit Committee’s approval. The
existing protocol was used as the benchmark. Patients were studied
prospectively to assess compliance with the local bowel protocol,
incidence of constipation and relationship to weaning from respiratory
support and feeding. All HDU and all mechanically ventilated
ICU patients who stayed on the ward for more than 3 days were
included, except for patients after bowel surgery and patients with
encephalopathy. Conclusion An excess of CXRs were performed for confi rmation
of nasogastric tube in our patient population. The recommended
methods for position confi rmation were reinforced amongst medical
staff . The high number of repeated imaging for some patients indicates
that dislodgement of tubes was also a problem. We propose that
nasogastric tubes should be bridled after fi rst dislodgement or at
tracheostomy insertion to minimise dislodgement in the future. Results Among the 24 HDU and 21 ICU patients audited in the
Royal Liverpool University Hospital, 67% and 57% respectively were
constipated. Laxatives were prescribed when patients had not opened
their bowels for 3 days in 25% HDU and 75% ICU cases. Taking into
consideration that the median age, APACHE II score and length
of stay for constipated and nonconstipated patients were similar,
the relationship to feeding and respiratory support were assessed. Constipated patients required mechanical ventilation for an average of
6.8 days and nonconstipated for 4.3 days. Failure to feed was observed
at least once in 58% constipated and 44% nonconstipated ICU patients
and 19% constipated and 12.5% nonconstipated HDU patients. 1.
Mostafa SM: Constipation and its implications in the critically ill patient. Br
J Anaesth 2003, 91:815-819. P240 Table 1 (abstract P242) Table 1 (abstract P242)
Mean days
CXR for
Total NG
Total
from fi rst to Mean
NG
Patients
CXR
CXR
Proportion
last CXR
age
1
47
47
321
0.15
8.45
67.55
2
16
32
181
0.18
15.94
69.63
3
7
21
87
0.24
17.86
70.00
4
10
40
149
0.27
15.90
69.20
5
3
15
52
0.29
23.33
66.00
6
3
18
61
0.30
40.00
76.67
8
2
16
52
0.31
39.50
80.00
9
2
18
46
0.39
29.50
83.00 Confi rmation of nasogastric tube placement in critical care
M Moore, R Thomson Confi rmation of nasogastric tube placement in critical care
M Moore, R Thomson
St Georges Healthcare NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P244 (doi: 10.1186/cc12182) St Georges Healthcare NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P244 (doi: 10.1186/cc12182) Conclusion In this population of ICU patients, routine/daily checks of
NG pH aspirate appear to be limited. This is almost certainly due to the
use of continuous NG feed together with PPIs. The usefulness of pH
testing in newly placed NG tubes, however, appears more reliable. Introduction Placement of nasogastric tubes (NGTs) is commonplace
in critical care. Misplacement of NGTs is rare and considered a never
event [1]. Strategies to avoid never events (by confi rming NGT position)
include pH analysis of gastric secretions or chest X-ray confi rmation
of tube position. For this reason the authors set out to establish the
effi ciencies surrounding safe placement of NGTs in a 17-bed adult
cardiothoracic critical care unit in a large teaching hospital. Figure 2 (abstract P245). pH values – new NG tubes. Figure 1 (abstract P245). pH values – routine checks. Figure 1 (abstract P245). pH values – routine checks. Methods This small-scale study of 25 NGT placements during a 5-week
period collated data supplied by questionnaire by healthcare workers
responsible for NGT placements.i Results Analysis of Adverse Incident Reports identifi ed no never
events of misplaced NGTs within the previous 10 years. This audit
revealed that the commonest type of NGT was a radio-opaque tube
with stylet (corfl o) (92% of placements), with occasional use of the
electromagnetic placement system (cortrak) (8% of placements). Sizes 10 (40%) and 12 (56%) were most common. Tube placement was
confi rmed by: X-ray (72%); pH of aspirates (35%); electromagnetic tube
placement (one patient). The time taken from decision to place NGT to
use varied (range 15 to 510 minutes). Little distinction was seen in the
time taken to use and NGT confi rmed by aspirate alone (205 minutes)
or by X-ray (220 minutes), although the shortest interval was seen
in electromagnetic NGT placement (15 minutes). The cost of NGTs
confi rmed by aspirate alone was low (approximately £10.00), higher
with X-ray confi rmation/electromagnetic placement (approximately
£45.00). Figure 1 (abstract P245). pH values – routine checks. Figure 2 (abstract P245). pH values – new NG tubes. pH testing to confi rm nasogastric tube position on the ICU: are we
wasting our time? p
Results Protein net balance was lower in patients than in the reference
group at baseline (–1.8 ± 1.7 vs. 0.6 ± 0.6 mg/kg BW/hour, P = 0.003),
and after enteral feeding (–1.1 ± 1.5 vs. 0.6 ± 0.6 mg/kg BW/hour,
P = 0.049). Recovery of labelled Phe from enteral feeding into the
systemic circulation was higher in the reference group as compared
with patients (20.3 + 11.2% vs. 7.0 + 4.8%, P = 0.005). Enteral feeding
did not aff ect protein metabolism in the reference group. In patients,
protein breakdown became slightly lower during enteral feeding
(10.6 ± 3.3 vs. 11.2 ± 3.3 mg/kg BW/hour, P = 0.086) and protein net
balance became slightly higher (–1.1 ± 1.5 vs. –1.8 ± 1.7 mg/kg BW/
hour, P = 0.086). P Temblett, S George Introduction For such a simple procedure, the insertion and position
checking of nasogastric (NG) tubes can be very problematic. The UK’s
National Patient Safety Agency declared that ‘Placement of NG tubes
together with confi rmation of correct placement can carry signifi cant
risks’ and recommends that measuring the pH of NG aspirate should
be the fi rst-line method of determining correct NG position (safe range
≤5.5) [1]. Methods In order to assess the usefulness of pH testing of NG aspirates
in critical care patients, a prospective survey of pH testing of NG tube
aspirate was carried out. This was undertaken both in patients who
had a newly placed NG tube and in patients who were having regular/
routine checks of their existing NG tube. Conclusion Intrinsically isotope-labelled casein can be used to
quantify dietary contribution to protein metabolism in critically ill
patients. Hypocaloric enteral feeding marginally improved protein
balance in these patients. The low recovery of enterally administered
labelled amino acid underlines the need to quantify uptake from
the gastrointestinal tract when protein turnover measurements are
performed in critically ill patients on enteral nutrition. Results A total of 168 separate pH readings in 41 ICU patients receiving
continuous enteral nutrition were recorded. In total, 137/168 patients
were receiving proton pump inhibitors (PPIs). Eighteen readings were
from newly placed NG tubes and 150 readings from old NG tubes. Fifty-
three per cent of routine pH readings were falsely high; that is, pH 6 or
above despite the NG tube being in the stomach (Figure 1). References 1. Mostafa SM: Constipation and its implications in the critically ill patient. Br
J Anaesth 2003, 91:815-819. 2. Palácio de Azevedo R: Intestinal constipation in intensive care unit. Rev Bras
Ter Intensiva 2009, 21:324-331. S91 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods Mechanically ventilated, not enterally fed ICU patients (n = 9)
were recruited from an interdisciplinary ICU. Healthy, overnight-fasted
volunteers (n = 6) served as reference. A primed constant i.v. infusion
of 2H-labeled phenylalanine (Phe) and tyrosine was used to quantify
whole-body protein metabolism. Patients remained on parenteral
nutrition (PN) as clinically indicated; controls received PN starting
2.5 hours before starting enteral feeding. Intrinsically 13C-Phe-labeled
casein was infused for 6 hours by nasogastric tube at 0.75 g protein/
hour, together with maltodextrin at 2.73 g/hour. Protein breakdown,
synthesis, net balance, and Phe splanchnic extraction were calculated
before and at the end of the enteral feeding period, using equations
for steady-state whole-body protein kinetics. Comparisons were made
by Wilcoxon matched pairs and Mann–Whitney U tests; values are
reported as mean ± SD. confi rmations. Even if the frequency and volume of gastric aspiration
were greater, there is a belief that pH testing may not be suffi ciently
accurate (since many factors alter patients’ gastric pH). It is possible
that new technologies such as electromagnetic NGT placement may
allow faster/equally safe practices. Further study including cost/benefi t
analysis will be needed to confi rm this. Reference 1. National Patient Safety Agency: Patient Safety Alert (5): Reducing the Harm
Caused By Misplaced Nasogastric Feeding Tubes. London: NPSA; 2005. [http://www.nrls.npsa.nhs.uk/resources/?EntryId45=59794] pH testing to confi rm nasogastric tube position on the ICU: are we
wasting our time? Twenty-
eight per cent of newly placed NG tubes had falsely high pH readings
(Figure 2). Reference Reference
1. National Patient Safety Agency: Reducing the Harm Caused by Misplaced
Nasogastric Feeding Tubes. NPSA/PSA001./PSA002. NPSA; 2005, 2011. [www.nrls.npsa.nhs.uk] P246
Eff ect of probiotic containing lactobacillus, bifi dobacterium and
streptococcus thermophilus in critically ill patients
M Ebrahimi Mamaghani, S Sanaie, A Mahmoudpour, H Hamishehkar
Tabriz University of Medical Sciences, Tabriz, Iran
Critical Care 2013, 17(Suppl 2):P246 (doi: 10.1186/cc12184) Introduction Sepsis is the most common cause of death in ICUs [1]. Destruction of intestinal barrier function and increased translocation
of bacteria to systemic blood fl ow can lead to sepsis [2]. Probiotics
may have benefi cial eff ects in improvement of critically ill patients
by modulating intestinal barrier and reduction of infl ammation
[3]. The aim of this trial was to determine the eff ect of probiotic on
infl ammatory biomarkers and mortality rate of sepsis in critically ill
patients in the ICU. p
Methods This double-blind, randomized controlled trial was
conducted on 40 critically ill patients admitted to the ICU. They were
randomly assigned to receive placebo or probiotic for 7 days. The
APACHE score, Sequential Organ Failure Assessment (SOFA) and
systemic concentrations of IL-6, procalcitonin (PCT) and protein C were
measured before initiation of the study and on days 4 and 7. Also, 28-
day mortality was evaluated for each patient. y
y
p
Results IL-6 and PCT levels decreased and protein C levels increased
signifi cantly in probiotic group over the treatment period (P <0.001). There was a signifi cant diff erence in IL-6, PCT and protein C levels of the
7th day between two groups (P = 0.001, 0.006 and <0.001, respectively). Compared with controls, probiotic was eff ective in improving APACHE
and SOFA scores in 7 days (P <0.001). There was signifi cant diff erence
between the probiotic and control group in the 28-day mortality rate
(20% vs. 55% respectively, P = 0.048).l y
Conclusion Probiotics reduce infl ammation and mortality rate in
critically ill patients and might be considered as an adjunctive therapy
to sepsis. Figure 1 (abstract P247). Selenium levels before and after 10 days of
admission. p
References Confi rmation of nasogastric tube placement in critical care
M Moore, R Thomson Conclusion Despite the small dataset the results demonstrate a
concerning delay in the application of enteral feeding and/or drug
administration. Whilst reassuring in the steps taken to avoid never
events, this study demonstrates that there may be delays in time-critical
administration of enteral medicine or optimal nutritional practices. This study reveals a signifi cant problem with aspirating gastric
contents for pH testing, necessitating a large number of X-ray position Figure 2 (abstract P245). pH values – new NG tubes. S92 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Table 1 (abstract P247). Mean selenium levels in diff erent sources of sepsis
Source
Quantity
Level (mg/dl)
Abdominal
12
0.43
Chest
12
0.71
Skin
3
0.55
CNS
2
0.53
GU
2
0.58
Figure 1 (abstract P247). Selenium levels before and after 10 days of
admission. Table 1 (abstract P247). Mean selenium levels in diff erent sources of sepsis Selenium levels in patients with diff erent sources of sepsis
L French, P Temblett
Morriston Hospital, Swansea, UK
Critical Care 2013, 17(Suppl 2):P247 (doi: 10.1186/cc12185) Selenium levels in patients with diff erent sources of sepsis
L French, P Temblett
Morriston Hospital, Swansea, UK
Critical Care 2013, 17(Suppl 2):P247 (doi: 10.1186/cc12185) p
References 1. Angus DC, Linde-Zwirble WT, Lidicker J, Clermont G, Carcillo J, Pinsky MR:
Epidemiology of severe sepsis in the USA: analysis of incidence, outcome,
and associated costs of care. Crit Care Med 2001, 29:1303-1310. 1. Angus DC, Linde-Zwirble WT, Lidicker J, Clermont G, Carcillo J, Pinsky MR:
Epidemiology of severe sepsis in the USA: analysis of incidence, outcome,
and associated costs of care. Crit Care Med 2001, 29:1303-1310. Conclusion All patients admitted with early septic shock had subnormal
selenium levels. Patients with an abdominal source of septic shock had
the lowest levels. This survey supports previous studies indicating early
supplementation may be benefi cial in septic shock patients. References 2. Hassoun HT, Kone BC, Mercer DW: Post-injury multiple organ failure: the
role of gut. Shock 2001, 15:1-10. 2. Hassoun HT, Kone BC, Mercer DW: Post-injury multiple organ failure: the
role of gut. Shock 2001, 15:1-10. 3. Morrow LE: Probiotics in the intensive care unit. Curr Opin Crit Care 2009,
15:144-148. 3. Morrow LE: Probiotics in the intensive care unit. Curr Opin Crit Care 2009,
15:144-148. References
1. Berger MM: Nutr Clin Pract 2006, 21:438-449. 2. Angstwurm MW, et al.: Crit Care Med 2007, 35:118-126. 3. Andrews PJ, et al.: BMJ 2011, 342:d1542. References
1. Berger MM: Nutr Clin Pract 2006, 21:438-449. 2. Angstwurm MW, et al.: Crit Care Med 2007, 35:118-126. 3. Andrews PJ, et al.: BMJ 2011, 342:d1542. 1. Berger MM: Nutr Clin Pract 2006, 21:438-449. 1. Berger MM: Nutr Clin Pract 2006, 21:438-449. 2. Angstwurm MW, et al.: Crit Care Med 2007, 35:118-126. 3. Andrews PJ, et al.: BMJ 2011, 342:d1542. P248 Selenium levels in patients with diff erent sources of sepsis
L French, P Temblett
Morriston Hospital, Swansea, UK
Critical Care 2013, 17(Suppl 2):P247 (doi: 10.1186/cc12185) P248
Is a combined i.v. and enteral glutamine regimen superior to a
single i.v. glutamine regimen in severe thoracic trauma?
D Pavelescu, I Grintescu, I Luca-Vasiliu, L Mirea
Emergency Hospital Floreasca, Bucharest, Romania
Critical Care 2013, 17(Suppl 2):P248 (doi: 10.1186/cc12186) Morriston Hospital, Swansea, UK Morriston Hospital, Swansea, UK D Pavelescu, I Grintescu, I Luca-Vasiliu, L Mirea D Pavelescu, I Grintescu, I Luca Vasiliu, L Mirea
Emergency Hospital Floreasca, Bucharest, Romania g
y
p
Critical Care 2013, 17(Suppl 2):P248 (doi: 10.1186/cc12186) Introduction The aim of this study is to establish whether diff erent
types of sepsis have an impact on selenium levels. Selenium is an
essential trace element involved in antioxidant and immunological
reactions. Selenium levels have been shown to be low in patients
with systemic infl ammatory response syndrome and sepsis. Selenium
replacement has been recommended in patients with sepsis [1,2]. Greater than 5 days of supplementation may also help to prevent the
development of new infections on ICUs [3]. Introduction Glutamine regulates many biological functions in
preserving the cell, acts as a key respiratory fuel and nitrogen donor
for rapidly dividing cells, and modulates the expression of many genes
associated with metabolism, cell defences and repair, and cytokine
production. In severe thoracic trauma, glutamine supplementation
is essential because the body consumes more than it produces and
glutamine eff ects become dependent on its route of delivery. Methods This is a prospective survey where selenium levels were
collected from patients admitted with septic shock to a tertiary ICU, for
6 months from October 2010 to March 2011. f
Methods Fifty-two patients 19 to 78 years old with surgery for severe
thoracic trauma were assessed in two groups: Group A received 0.3
to 0.5 g/kg/day i.v. glutamine + 20 g enteral glutamine for 7 days,
supplementation to enteral nutrition; Group B receive only i.v. glutamine supplementation to enteral nutrition 0.3 to 0.5 g/kg/day for
7 days. Weaning time, the duration of p.o. ileus, incidence and time to
resolution of VAP, glycemic level and the percentage decrease of CRP at
96 hours were assessed in both groups. Results Selenium levels were measured in 31 patients with septic shock. Abdominal and chest sepsis were the main sources of infection. Those
with an abdominal source of sepsis had the lowest levels, as shown in
Table 1. All septic shock patients who had selenium levels taken within
the fi rst 10 days of admission had subnormal levels (<0.8 mg/dt), and
after 10 days had levels within the normal range, as shown in Figure 1. P248
Is a combined i.v. and enteral glutamine regimen superior to a
single i.v. glutamine regimen in severe thoracic trauma?
D Pavelescu, I Grintescu, I Luca-Vasiliu, L Mirea
Emergency Hospital Floreasca, Bucharest, Romania
Critical Care 2013, 17(Suppl 2):P248 (doi: 10.1186/cc12186) Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S93 Critical Care 2013, Volume 17 Suppl 2 pp
http://ccforum.com/supplements/17/S2 Results Weaning time and the duration of p.o. ileus were signifi cantly
lower in Group A; although the incidence of VAP is similar in both
groups, the time of VAP resolution is lower, the glycemic control is
better in Group A. The percentage of CRP decrease is higher in Group
A. See Figure 1. Conclusion Glutamine becomes an essential amino acid in severe
thoracic trauma and when the patients are fed other than TPN (enteral
P249
Clinical eff ect of altered lipid emulsion composition containing
fi sh oil in postoperative patients following abdominal surgery:
a prospective, randomized, open-label, comparative, multicenter
phase 3 clinical study
M Keum1, S Hong1, H Han2, D Yoon3, J Seo4, I Yun5
1Asan Medical Center Seoul South Korea; 2Seoul National University
Figure 1 (abstract P248). Results. tract P248). Results Results Weaning time and the duration of p.o. ileus were signifi cantly
lower in Group A; although the incidence of VAP is similar in both
groups, the time of VAP resolution is lower, the glycemic control is
better in Group A. The percentage of CRP decrease is higher in Group
A. See Figure 1. Conclusion Glutamine becomes an essential amino acid in severe
thoracic trauma and when the patients are fed other than TPN (enteral,
oral); although hard evidence is lacking, both administration routes
may be effi cient as soon as possible. Results Weaning time and the duration of p.o. ileus were signifi cantly
lower in Group A; although the incidence of VAP is similar in both
groups, the time of VAP resolution is lower, the glycemic control is
better in Group A. The percentage of CRP decrease is higher in Group
A. See Figure 1. P249 P249
Clinical eff ect of altered lipid emulsion composition containing
fi sh oil in postoperative patients following abdominal surgery:
a prospective, randomized, open-label, comparative, multicenter
phase 3 clinical study
M Keum1, S Hong1, H Han2, D Yoon3, J Seo4, I Yun5
1Asan Medical Center, Seoul, South Korea; 2Seoul National University
Bundang Hospital, Seongnam, South Korea; 3Gangnam Severance Hospital,
Seoul, South Korea; 4Samsung Medical Center, Seoul, South Korea; 5Konkuk
University Medical Center, Seoul, South Korea
Critical Care 2013, 17(Suppl 2):P249 (doi: 10.1186/cc12187) P249
Clinical eff ect of altered lipid emulsion composition containing
fi sh oil in postoperative patients following abdominal surgery:
a prospective, randomized, open-label, comparative, multicenter
phase 3 clinical study g
Conclusion Glutamine becomes an essential amino acid in severe
thoracic trauma and when the patients are fed other than TPN (enteral,
oral); although hard evidence is lacking, both administration routes
may be effi cient as soon as possible. Introduction Omega-3 fatty acids have been shown to decrease
infl ammatory responses after trauma and surgery resulting in S94 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 vs. 16.2 ± 19.4 IE, P = 0.155; EN: 52.9 ± 25.6 vs. 61.7 ± 25%, P = 0.304;
day 3: gluc: 129 ± 17 vs. 133 ± 21 mg/dl, P = 0.578; ins: 35.1 ± 30.9 vs. 32.3 ± 30.5 IE, P = 0.802; EN: 78.3 ± 28.9 vs. 83.4 ± 29%, P = 0.617; day 5:
gluc: 124 ± 16 vs. 132 ± 17 mg/dl, P = 0.276; ins: 25.7 ± 37 vs. 33 ± 26.4
IE, P = 0.562; EN: 73.9 ± 41.4 vs. 87.1 ± 23%, P = 0.354). Interestingly,
the calorie achievement was not associated with insulin demand (day
1: R = 0.241, P = 0.156) or average blood glucose (day 1: R = 0.248,
P = 0.14) throughout the study. reduction of morbidity and mortality in postoperative patients. The
n-6:n-3 polyunsaturated fatty acid (PUFA) ratio in parenteral nutrition
(PN) also infl uences the immune modulation. The aim of this study
was to evaluate the optimal ratio of PUFA in postoperative patients
following abdominal surgery. Methods In a prospective, randomized, open-label, comparative,
multicenter, phase 3 clinical study, we compared the safety and
effi cacy of a 2.1:1 ratio of n-6:n-3 fatty acid compared with 2.5:1 in
postoperative patients requiring PN. Eff ect of hypocaloric versus normocaloric parenteral nutrition on
whole body protein kinetics in critically ill neurosurgical patients
O Rooyackers, A Berg, J Wernerman y
O Rooyackers, A Berg, J Wernerman y
,
g,
Karolinska Institutet and University Hospital, Huddinge, Sweden
Critical Care 2013, 17(Suppl 2):P251 (doi: 10.1186/cc12189) y
,
g,
arolinska Institutet and University Hospital, Huddinge, Sweden y
g
Karolinska Institutet and University Hospital, Huddinge, Sweden
Critical Care 2013, 17(Suppl 2):P251 (doi: 10.1186/cc12189) y
ritical Care 2013, 17(Suppl 2):P251 (doi: 10.1186/cc12189) Table 1 (abstract P249). Lipid emulsion composition of CFO and KAB Table 1 (abstract P249). Lipid emulsion composition of CFO and KAB
CFO
KAB Table 1 (abstract P249). Lipid emulsion composition of CFO and KAB
CFO
KAB Table 1 (abstract P249). Lipid emulsion composition of CFO and KAB
CFO
KAB
LCT (g/l)
60
60
MCT (g/l)
50
60
Olive oil (g/l)
50
50
Fish oil (g/l)
40
30
n-6:n-3
2.1:1
2.5:1 Introduction The optimal feeding of critically ill patients treated in the
ICU is controversial. Present guidelines for protein feeding are based
on weak evidence obtained with suboptimal methods. Whole body
protein kinetics is an attractive technique to assess optimal protein
intake by measuring the eff ect of protein feeding strategies on protein
synthesis rates, protein degradation rates and protein balance. Here
protein kinetics were measured in critically ill neurosurgical patients
during hypocaloric and normocaloric parenteral nutrition. g y
Methods Neurosurgical patients on mechanical ventilation (n = 16)
were studied. Energy expenditure was measured with indirect
calorimetry. After that, the patients were randomized to receiving
24 hours of 50% of measured energy expenditure followed by 24 hours
of 100% or 100% before 50%. Whole body protein kinetics were
measured during the last half hour of the feeding periods using stable
isotope-labeled phenylalanine as a tracer. During a continuous infusion
of labeled phenylalanine and tyrosine, plasma samples were obtained
and later analyzed for the content of the labeled amino acids using
mass spectrometry. Protein kinetics were calculated using standard
steady-state kinetics. In addition, amino acid concentrations were
analyzed by HPLC. Student’s t test was used for statistical analyses. Conclusion CFO containing low n-6:n-3 PUFA ratio was comparable
with KAB in safety and effi cacy in postoperative patients requiring PN
and also had advantages with regard to lipid metabolism aspect. References References
1. Simopoulos AP: Biomed Pharmacother 2002, 56:365-379. 2. Heller AR, et al.: Crit Care Med 2006, 34:972-979. References
1. Simopoulos AP: Biomed Pharmacother 2002, 56:365-379. 2. Heller AR, et al.: Crit Care Med 2006, 34:972-979. P249 Fifty-four patients were assigned
to receive PN with Combifl ex Omega peripheral® (CFO, low ratio group,
n = 28) or SMOFKabiven peripheral® (KAB, high ratio group, n = 26) for
3 days after abdominal surgery. Safety and effi cacy were monitored
daily with laboratory parameters, vital signs, and adverse events from
the operation day (day 0) until the end of the study (day 4). Conclusion Medical critically ill patients with fat-based or glucose-
based EN achieve similar glucose control. EN was not associated with
glucose concentrations or insulin demand. P250
Infl uence of fat-based versus glucose-based enteral nutrition
formulas on glucose homeostasis y
y
y
Results The patients received 0.5 ± 0.1 and 1.1 ± 0.2 g amino acids/
kg/day (P <0.001) on the days with 50 and 100% of measured energy
expenditure respectively. Energy expenditures were 23.4 ± 2.7 and
24.5 ± 2.3 kcal/kg/day (P = 0.05) on the 50 and 100% days respectively. Plasma amino acids concentrations were 2.8 ± 0.5 and 2.9 ± 0.4 mM
(P = 0.085) on the 2 days respectively. Whole body protein synthesis
was 12% lower when 50% of energy expenditure was given, 11.7 ± 3.0
versus 13.3 ± 2.2 mg/kg/hour (P = 0.025), whilst protein degradation
was unaltered 13.6 ± 3.5 versus 14.0 ± 2.6 mg/kg/hour (P = 0.56). Also
protein oxidation was unaltered 3.0 ± 2.1 versus 2.9 ± 1.4 mg/kg/hour
(P = 0.85). This resulted in a 60% higher whole body protein balance
with the normocaloric nutrition, –1.9 ± 2.1 versus –0.7 ± 1.3 mg/kg/
hour (P = 0.014). M Wewalka, A Drolz, C Zauner Introduction Trauma patients who receive fat-based parenteral
nutrition (PEN) achieve better glucose control compared with those fed
with glucose-based PEN [1,2]. Therefore, we determined whether fat-
based enteral nutrition (EN) has the same benefi t on glucose control
and exogenous insulin demand in medical intensive care patients
compared with glucose-based EN. Here we present preliminary data
for this ongoing randomized controlled cohort study. Methods Medical critically ill patients with need for mechanical
ventilation and without contraindications for EN are included in the
study. Patients are randomly assigned to receive either fat-based
(n = 30) or glucose-based (n = 30) EN. To evaluate the individual calorie
demand, indirect calorimetry is performed after an overnight fast. The
determined amount of EN is administered continuously for 5 days. Glucose concentrations are measured at least three times per day and
averaged. Furthermore, exogenous insulin demand per 24 hours and
calorie achievement per 24 hours are evaluated daily. Conclusion The protein kinetics measurements and the protocol used
were useful to assess the effi cacy of nutritional support in critically ill
patients. In the critically ill neurosurgical patients treated in the ICU,
hypocaloric feeding was associated with a more negative protein
balance, while the amino acid oxidation was not diff erent. References 1. Tappy L, et al.: Crit Care Med 1998, 26:860-867. 1. Tappy L, et al.: Crit Care Med 1998, 26:860-867. 2. Huschak G, et al.: Intensive Care Med 2005, 31:1202-1208. 2. Huschak G, et al.: Intensive Care Med 2005, 31:1202-1208. Results Total cholesterol (TC) and low-density lipoprotein-cholesterol
(LDL-C) levels were less changed signifi cantly in the low ratio group
(3 ± 18 vs. 16 ± 23 mg/dl, P = 0.027 for TC, 4 ± 12 vs. 12 ± 18 mg/dl,
P = 0.026 for LDL-C) compared with the high ratio group in postoperative
patients. Other laboratory parameters and adverse events did not show
statistically signifi cant diff erences between the groups. See Table 1. P252
Withholding parenteral nutrition for 1 week reduces ICU-acquired
weakness Results So far, 37 patients, 16 with fat-based, 21 with glucose-based EN
have been included in the study. Both groups had similar age (62 ± 10
vs. 58 ± 16 years, P = 0.44), body mass index (26.7 ± 5.8 vs. 28.4 ± 4.4 kg/
m2, P = 0.302), SAPS II score (62.4 ± 12.7 vs. 64 ± 12.3, P = 0.697),
and fasting plasma glucose (132 ± 34 vs. 121 ± 26 mg/dl, P = 0.269). Furthermore, resting energy expenditure was similar in both groups
(1,522 ± 365 vs. 1,573 ± 313 kcal/day, P = 0.647). Throughout the entire
study period, average blood glucose, exogenous insulin demand,
and calorie achievements per day were similar between the groups
(day 1: gluc: 139 ± 30 vs. 127 ± 20 mg/dl, P = 0.143; ins: 27.8 ± 28.4 G Hermans, B Clerckx, T Vanhullebusch, F Bruyninckx, M Casaer,
P Meersseman, D Mesotten, S Vancromphaut, P Wouters, R Gosselink,
A Wilmer, G Van den Berghe
UZ Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P252 (doi: 10.1186/cc12190) G Hermans, B Clerckx, T Vanhullebusch, F Bruyninckx, M Casaer,
P Meersseman, D Mesotten, S Vancromphaut, P Wouters, R Gosselin
A Wilmer, G Van den Berghe Critical Care 2013, 17(Suppl 2):P252 (doi: 10.1186/cc12190) Critical Care 2013, 17(Suppl 2):P252 (doi: 10.1186/cc12190 Introduction ICU-acquired weakness (ICUAW) is a frequent and
important complication of critical illness [1]. A large randomized S95 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 intake. In the abdominal muscle compartments, changes were similar,
albeit smaller. Femoral and abdominal subcutaneous adipose tissue
compartments were unaff ected by disease and nutritional strategy. Conclusion Early PN did not prevent the pronounced wasting of
skeletal muscle observed over the fi rst week of critical illness. Moreover,
early PN increased the amount of adipose tissue within the muscle
compartments. R f controlled trial (EPaNIC: clinicaltrials.gov: NCT00512122) [2] showed
that withholding parenteral nutrition during the fi rst week of ICU stay
whereby tolerating substantial caloric defi cit (late PN) accelerated
recovery and shortened weaning time as compared with early
parenteral substitution for defi cient enteral feeding (early PN). We
examined the impact of late PN, as compared with early PN, on
incidence and recovery of ICUAW. y
Methods A preplanned subanalysis of adult patients included in the
EPaNIC trial. P254 P254
Withholding parenteral nutrition during the fi rst week of critical
illness increases plasma bilirubin but lowers the incidence of
cholestasis and gallbladder sludge
YM Vanwijngaerden1, L Langouche1, M Gielen1, Y Debaveye1,
M Casaer1, C Liddle2, S Coulter2, R Brunner1, P Wouters1, A Wilmer1,
G Van den Berghe1, D Mesotten1
1University Hospitals KU Leuven, Belgium; 2University of Sydney, Australia
Critical Care 2013, 17(Suppl 2):P254 (doi: 10.1186/cc12192) 5
Withholding parenteral nutrition during the fi rst week of critical
illness increases plasma bilirubin but lowers the incidence of
cholestasis and gallbladder sludge
YM Vanwijngaerden1, L Langouche1, M Gielen1, Y Debaveye1,
M Casaer1, C Liddle2, S Coulter2, R Brunner1, P Wouters1, A Wilmer1,
G Van den Berghe1, D Mesotten1
1University Hospitals KU Leuven, Belgium; 2University of Sydney, Australia
Critical Care 2013, 17(Suppl 2):P254 (doi: 10.1186/cc12192) Introduction Cholestatic liver dysfunction (CLD) during critical illness,
defi ned by hyperbilirubinemia, often occurs and is associated with
poor outcome. Parenteral nutrition (PN) is assumed to aggravate CLD. However, hyperbilirubinemia more frequently occurred when the start
of PN was delayed until day 8 in the ICU (late PN) [1]. Late PN accelerated
recovery as compared with early initiation of PN. Results Clinical ICUAW evaluation was performed in 600 patients
(matched n = 558), electrophysiological testing in 730 (matched
n = 684). Late PN reduced the incidence of ICUAW at fi rst evaluation
from 43.1% to 34.4%, P = 0.03 (matched: early PN 41.6%, late PN 33.3%,
P = 0.04). Signifi cantly fewer patients in the late PN group developed
weakness at any time during ICU stay (late PN 37.0%, early PN 46.4%,
P = 0.02; matched: late PN 36.2%, early PN 45.2%, P = 0.03). ICUAW
may have recovered faster with late PN than with early PN (P = 0.05,
matched P = 0.06). Other outcomes were not diff erent. y
y
Methods This was a preplanned subanalysis of a large randomized
controlled trial on early versus late initiation of PN (n = 4,640) [1]. Plasma
total bilirubin was quantifi ed in all patients daily while in the ICU. Liver
enzymes ALT, AST, GGT and ALP were quantifi ed twice weekly in all
patients while in the ICU. In a random predefi ned subset of patients,
circulating bile salts were also quantifi ed with MS-HPLC at baseline
and on day 3, day 5 and the last day in the ICU (n = 280). Impact of early parenteral nutrition on muscle and adipose tissue
compartments during critical illness L Langouche1, MP Casaer2, W Coudyzer2, D Vanbeckevoort2,
B De Dobbelaer2, FG Güiza1, PJ Wouters2, D Mesotten2, G Van den Berghe2
1KU Leuven, Belgium; 2University Hospitals Leuven – KU Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P253 (doi: 10.1186/cc12191) g
y
Conclusion Tolerating substantial caloric defi cit by withholding PN
until day 8 of critical illness increased circulating levels of bilirubin but
reduced the occurrence of gallbladder sludge and lowered GGT, ALP
and ALT levels. These results suggest that hyperbilirubinemia during
critical illness dies not necessarily refl ect cholestasis and instead may
be an adaptive response. Additional analyses on a propensity score-
matched patient population are ongoing. Introduction The goal of enhanced nutrition in critically ill patients is
to improve outcome by reducing lean tissue wasting. However, such
eff ect has not been proven. This study aimed to assess the eff ect of
early administration of parenteral nutrition (PN) on muscle volume and
composition by repeated quantitative computer tomography (qCT). Introduction The goal of enhanced nutrition in critically ill patients is
to improve outcome by reducing lean tissue wasting. However, such
eff ect has not been proven. This study aimed to assess the eff ect of
early administration of parenteral nutrition (PN) on muscle volume and
composition by repeated quantitative computer tomography (qCT). p
y
p
q
p
g
p y (q
)
Methods We performed a preplanned substudy of a randomized
controlled trial (EPaNIC) that compared early initiation of PN
when enteral nutrition was insuffi cient (early PN) with tolerating a
pronounced nutritional defi cit for 1 week in the ICU (late PN) [1]. Late
PN prevented infections and accelerated recovery. We studied 15
EPaNIC study neurosurgical patients requiring prescheduled repeated
follow-up CT scans and six healthy volunteers matched for age, gender
and BMI. Repeated abdominal and femoral qCT images were obtained
in a standardized manner on median ICU day 2 (IQR 2 to 3) and day 9
(8 to 10). Intramuscular, subcutaneous and visceral fat compartments
were delineated manually. Muscle and adipose tissue volume and
composition were quantifi ed using standard Hounsfi eld Unit ranges. Results Critical illness evoked substantial loss of femoral muscle volume
in 1 week, irrespective of the nutritional regimen. Early PN reduced the
quality of the muscle tissue, as refl ected by the attenuation, revealing
increased intramuscular water/lipid content. Early PN also increased the
volume of adipose tissue islets within the femoral muscle compartment. P252
Withholding parenteral nutrition for 1 week reduces ICU-acquired
weakness The study was performed between October 2008 and
November 2010 and included those patients who required intensive
care for ≥8 days as well as a computer-generated, admission category-
matched, random sample of short-stay ICU patients, the latter to
correct for possible bias evoked by earlier ICU discharge in one of the
two study groups. Assessors blinded for treatment allocation evaluated
muscle strength clinically three times weekly from awakening onward
and performed nerve conduction studies and electromyography (NCS
and EMG) weekly. The primary outcome was the incidence of ICUAW,
diagnosed clinically by the Medical Research Council (MRC) sum score
(<48/60) [3] at fi rst evaluation. Secondary outcomes included ICUAW
at worst and last MRC evaluation, recovery from ICUAW and incidence
of abnormal fi ndings on NCS and EMG. All analyses were performed
on the total dataset and on a for-baseline characteristics propensity
score-matched sample to correct for possible imbalances between the
groups. Reference 1. Casaer MP, et al.: N Engl J Med 2011, 365:506-517. 1. Casaer MP, et al.: N Engl J Med 2011, 365:506-517. 1. Casaer MP, et al.: N Engl J Med 2011, 365:506-517. Impact of early parenteral nutrition on muscle and adipose tissue
compartments during critical illness These changes in skeletal muscle integrity correlated with caloric p
y
p
q
p
g
p y q
Methods We performed a preplanned substudy of a randomized
controlled trial (EPaNIC) that compared early initiation of PN
when enteral nutrition was insuffi cient (early PN) with tolerating a
pronounced nutritional defi cit for 1 week in the ICU (late PN) [1]. Late
PN prevented infections and accelerated recovery. We studied 15
EPaNIC study neurosurgical patients requiring prescheduled repeated
follow-up CT scans and six healthy volunteers matched for age, gender
and BMI. Repeated abdominal and femoral qCT images were obtained
in a standardized manner on median ICU day 2 (IQR 2 to 3) and day 9
(8 to 10). Intramuscular, subcutaneous and visceral fat compartments
were delineated manually. Muscle and adipose tissue volume and
composition were quantifi ed using standard Hounsfi eld Unit ranges. Reference 1. Casaer et al.: N Engl J Med 2011, 365:506-516. 1. Casaer et al.: N Engl J Med 2011, 365:506-516. P254 Gallbladder
sludge was evaluated by ultrasound on ICU day 5 by blinded assessors
(n = 776). f
Conclusion As compared with early PN, late PN reduced the incidence
of ICUAW and may have accelerated recovery thereof. References Results From day 1 after randomization until the end of the 7-day
intervention window, plasma bilirubin was higher in the late PN than
in the early PN group (all P <0.001). In the late PN group, as soon as
PN was started on day 8, plasma bilirubin also fell and the two groups
became comparable. Maximum levels of GGT, ALP and ALT during the
ICU stay were higher in the early PN group (all P <0.01). Compared with
baseline, the circulating glycine and taurine conjugated primary bile
salts were elevated on day 3, day 5 and last day of the ICU stay (P <0.01
for all). However, there was no diff erence between the two groups. More patients in the early PN than in the late PN group had gallbladder
sludge on day 5 (45% vs. 37%; P = 0.04).i 1. Stevens et al.: Int Care Med 2007, 33:1876-1891. 2. Casaer et al.: N Engl J Med 2011, 365:506-516. 3. De Jonghe et al.:, JAMA 2002, 288:2859-2867. 1. Stevens et al.: Int Care Med 2007, 33:1876-1891. 2. Casaer et al.: N Engl J Med 2011, 365:506-516. 3. De Jonghe et al.:, JAMA 2002, 288:2859-2867. References the duration of renal replacement therapy (RRT) [1]. The impact of the
intervention on early markers of catabolism has not been investigated. Methods We studied the impact of early versus late PN on daily
markers of catabolism in the ICU in the total study population and in
propensity score-matched subgroups of long-stay patients. In addition,
we calculated the net incorporation rate of the extra amino acids
supplied by early PN. 1. Debaveye, Van den Berghe: Annu Rev Nutr 2006, 26:513-538. 2. Masiero et al.: Cell Metab 2009, 10:507-515. 3. Vanhorebeek et al.: J Clin Endocrinol Metab 2011, 96:E633-E645. 4. Derde et al.: Endocrinology 2012, 153:2267-2276. 5. Casaer et al.: N Engl J Med 2011, 365:506-517. Results Plasma urea, the urea/creatinine ratio and nitrogen excretion
increased over time in the ICU. Early PN further increased these markers
of catabolism, from the fi rst day of amino acid infusion onward, and
only marginally improved the nitrogen balance. Also in the group
that received PN only after the fi rst week in the ICU, ureagenesis was
increased by infusing amino acids. Over the fi rst 2 weeks, approximately
two-thirds of the extra amino acids supplied by early PN were net
wasted in urea. The above fi ndings were confi rmed in propensity score-
matched subgroups of long-stay patients. The higher urea levels with
early PN, rather than the kidney function as such, may have driven the
observed longer duration of RRT, as supported by multiple regression
analysis. P255 Impact of early parenteral nutrition on catabolism
J Gunst1, I Vanhorebeek1, MP Casaer1, G Hermans1, PJ Wouters1, J Dubois2,
K Claes1, M Schetz1, G Van den Berghe1
1KU Leuven, Belgium; 2Jessa Hospital, Hasselt, Belgium
Critical Care 2013, 17(Suppl 2):P255 (doi: 10.1186/cc12193) Introduction Prolonged critically ill patients enter a state of
hypercatabolism and muscle weakness, which has been associated
with increased morbidity and mortality. Early, full feeding of ICU
patients has been advocated to counteract catabolism. However, a
large, multicenter study found that early parenteral nutrition (PN) had
no signifi cant impact on mortality and even increased dependency
on intensive care with, among others, a signifi cant prolongation of Introduction Prolonged critically ill patients enter a state of
hypercatabolism and muscle weakness, which has been associated
with increased morbidity and mortality. Early, full feeding of ICU
patients has been advocated to counteract catabolism. However, a
large, multicenter study found that early parenteral nutrition (PN) had
no signifi cant impact on mortality and even increased dependency
on intensive care with, among others, a signifi cant prolongation of S96 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Overcrowding estimation in the emergency department: is the
simplest score the best? Overcrowding estimation in the emergency department: is the
simplest score the best? V Winkin, V Pinckaers, V D’Orio, A Ghuysen
CHU Liège, Belgium
Critical Care 2013, 17(Suppl 2):P258 (doi: 10.1186/cc12196) p
V Winkin, V Pinckaers, V D’Orio, A Ghuysen
CHU Liège, Belgium
Critical Care 2013, 17(Suppl 2):P258 (doi: 10.1186/cc12196) V Winkin, V Pinckaers, V D’Orio, A Ghuysen
CHU Liège, Belgium
Critical Care 2013, 17(Suppl 2):P258 (doi: 10.1186/cc12196) Results As compared with controls, muscle from critically ill patients
showed reduced myofi ber density, a shift to smaller (especially type I)
myofi bers, lower myosin and actin mRNA, upregulated mRNA of the
ubiquitin ligases muscle-ring-fi nger-1 and atrogin-1, a small increase
in the autophagosome formation marker LC3-II/LC3-I, and increases
in the autophagic substrates ubiquitin and p62 (all P ≤0.006). Late-PN,
resulting in a larger caloric defi cit than early-PN, had no substantial
impact on atrophy markers. In contrast, late-PN increased LC3-II/LC3-I
(P = 0.02), which coincided with less accumulation of ubiquitinated
proteins/aggregates (P = 0.05). Fewer patients on late-PN developed
muscle weakness as compared with early-PN (42% vs. 68%, P = 0.05). In multivariable analysis, a lower LC3-II/LC3-I ratio (P = 0.05) and higher
myofi ber density (P = 0.04) were independently associated with muscle
weakness. Introduction Emergency department (ED) overcrowding is a major
international problem with a negative impact on both patient care and
providers. Among validated methods of measurement, emergency
physicians have to choose between simple and complex scores [1,2]. The aim of the present study was to compare the complex National
Emergency Department Overcrowding Scale (NEDOCS) with the
simple occupancy rate (OR) determination. We further evaluated the
correlation between these scores and a qualitative assessment of
crowding. Methods The study was conducted in two academic hospitals and one
county hospital in Liège, Chênée and Verviers; each with an ED census
of over 40,000 patient visits per year. Samplings occurred over a 2-week
period in January 2011, with fi ve sampling times each day.f Methods The study was conducted in two academic hospitals and one
county hospital in Liège, Chênée and Verviers; each with an ED census
of over 40,000 patient visits per year. Samplings occurred over a 2-week
period in January 2011, with fi ve sampling times each day. Results ED staff considered overcrowding as a major concern in the
three EDs. Median OR ranged from 68 to 100, while the NEDOCS
ranged from 64.5 to 76.3. P256 f
Results In the period studied, the gross attendance fi gure increased
by 20,480 (33.72%), whilst the admission rate rose from 22 to 27%. Following closure of the neighbouring ED, the proportion of high-
acuity patients attending our institution increased dramatically,
with the proportion of category one and two patients (Manchester
Triage Scale) increasing by 8.33% (P = 0.076) and 18.80% (P <0.001),
respectively. Likewise, ambulance arrivals increased out of proportion
to the total increase in attendances (P = 0.016). Admissions from the ED
to the ICU increased by 63.04%. Consultants workloads now include
50% more category 1 and 2 patients (P = 0.001).i g
p
Reference g
p
Reference 1. Casaer et al.: N Engl J Med 2011, 365:506-517. P257
Impact of closing an emergency department on a neighbouring
teaching hospital: the concentrate eff ect
J Millar, R Wilson, P O’Connor, R McLaughlin
Royal Victoria Hospital, Belfast, UK
Critical Care 2013, 17(Suppl 2):P257 (doi: 10.1186/cc12195) P257
Impact of closing an emergency department on a neighbouring
teaching hospital: the concentrate eff ect
J Millar, R Wilson, P O’Connor, R McLaughlin
Royal Victoria Hospital, Belfast, UK
Critical Care 2013, 17(Suppl 2):P257 (doi: 10.1186/cc12195) Introduction Closure of an acute hospitals emergency department
(ED) has important ramifi cations for those centres expected to take up
the resultant workload. The continued reconfi guration of emergency
care is likely to produce an increasing number of these scenarios. Little
evidence is available to support planning of such initiatives and thus the
implications are diffi cult to anticipate. This study aims to demonstrate
one hospital’s experience of the rationalisation of emergency care and
its eff ect on workload. y
Conclusion The extra amino acids supplied by early PN appeared
ineffi cient to reverse the negative nitrogen balance, not because of
insuffi cient amino acid delivery, but rather because of insuffi cient
incorporation with, instead, increased degradation into urea. The
substantial catabolism of the extra amino acids, leading to pronounced
urea generation, may have prolonged the duration of RRT in the early
PN group. Methods This retrospective study was conducted in a large teaching
hospital. Activity data were analysed for a 12-month period following
the closure of a neighbouring ED. The results were subsequently
compared against the year prior to closure. Attendance, triage data,
admission rates and waiting times were compared across the two
periods, as were workload data for all grades of physician. The chi-
squared test was used to examine diff erences between groups.i Impact of early versus late parenteral nutrition on morphological
and molecular markers of atrophy and autophagy in skeletal muscle
of critically ill patients Impact of early versus late parenteral nutrition on morphological
and molecular markers of atrophy and autophagy in skeletal muscle
of critically ill patients
I Vanhorebeek, MP Casaer, F Güiza, S Derde, I Derese, PJ Wouters,
Y Debaveye, J Gunst, G Hermans, G Van den Berghe
KU Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P256 (doi: 10.1186/cc12194) Introduction Muscle weakness of critical illness is associated
with prolonged dependency on ventilatory support and delayed
rehabilitation. Muscle wasting related to poor nutrition has long been
considered a major determinant, whereas the importance of myofi ber
integrity only recently emerged [1-4]. We hypothesized that nutrient
restriction early during illness aggravates atrophy while preserving
myofi ber integrity by activating the crucial cellular quality control
pathway autophagy. The latter could be important to preserve muscle
function. Conclusion Reconfi guration of emergency care can have dramatic
implications for existing services; these may not always be anticipated. Rationalisation of ED’s may result in a concentration of high-acuity
patients accompanied by a downturn in the numbers of patients
whose presentations are amenable to care delivered in other settings. This abrupt change in case mix requires a re-examination of existing
workforces and their seniority. Methods Critically ill patients (n = 122) were randomized to early (early-
PN) or late (late-PN) initiation of parenteral nutrition to complete failing
enteral nutrition, while maintaining normoglycemia (80 to 110 mg/
dl) with insulin, in the EPaNIC study [5]. Vastus lateralis biopsies were
harvested after 1 week and compared with matched controls (n = 20). P259
GP-led walk-in centre in the UK: another way for urgent healthcare
provision M Arain, J Nicholl, M Campbell
The University of Sheffi eld, UK
Critical Care 2013, 17(Suppl 2):P259 (doi: 10.1186/cc12197) M Arain, J Nicholl, M Campbell
The University of Sheffi eld, UK
Critical Care 2013, 17(Suppl 2):P259 (doi: 10.1186/cc12197) Introduction It is evident that accident and emergency departments
are overloaded with patients, which results in delays in healthcare
provision [1]. A large proportion of patients consist of patients with
minor illness that can be seen by a healthcare provider in a primary care
setting. The aim of the study was to determine the characteristics of
patients using GP walk-in services, patients’ satisfaction and the eff ect
on emergency department (ED) services. Introduction It is evident that accident and emergency departments
are overloaded with patients, which results in delays in healthcare
provision [1]. A large proportion of patients consist of patients with
minor illness that can be seen by a healthcare provider in a primary care
setting. The aim of the study was to determine the characteristics of
patients using GP walk-in services, patients’ satisfaction and the eff ect
on emergency department (ED) services. Figure 1 (abstract P260). GP patient visits before and after the reform of
emergency services. has a negative impact on patient safety [1]. The aim of this study was
to analyse whether a reform of emergency care can reduce patient fl ow
into the ED. y
Methods The survey was conducted in Sheffi eld and Rotherham
walk-in centres over 3 weeks during September and October 2011. A self-reported, validated questionnaire was used to conduct survey
on the patients presenting at these centres. We estimated that a
sample size of around 400 patients from each centre was required to
achieve statistically robust results. A post-visit, short questionnaire
was also sent to those who agreed for the second questionnaire and
provided contact details. ED data were also obtained from April 2008
to March 2010, 1 year before and 1 year after the opening of the GP
walk-in centre. Data were entered and analysed in PASW Statistics 18. Ethical approval of the study was obtained from the NHS ethical review
committee. Methods A substantial reform of emergency care took place in the
province of Kanta-Häme in Southern Finland. Three separate out-of-
hours services in primary healthcare (PHC) and one ED in the hospital
were combined into one large ED in April 2007. P259
GP-led walk-in centre in the UK: another way for urgent healthcare
provision Basic principles of
the new ED were: the ED is only for those patients who are seriously
ill or injured, and need immediate care; PHC (healthcare centres) take
care of acute ordinary illnesses and nonserious injuries during offi ce
hours. To achieve these principles a regional fi ve-scale triage system
was planned and implemented. The information plan was established. Citizens were systematically informed about the principles of the new
ED by mail, articles in the newspapers and interviews in the radio and
television. The ED’s Internet pages were planned and established. The
number of patient visits (Hämeenlinna region) was analyzed 2 years
before and after establishing the new ED. Results A total of 1,030 patients participated in the survey (Rotherham
501; Sheffi eld 529). The mean age of the participants was 32.1 years at
Sheffi eld and 30.88 years at Rotherham. A higher proportion of users
were female, around 59% at both centres. Most of the patients rated
high for convenience of the centre opening hours and location (above
85%, apart from the location of Sheffi eld centre, which was rated high
by around 72% of the research participants). Overall 93% patients
were satisfi ed with the service at Rotherham centre and around 86%
at the Sheffi eld centre. Based on the estimation of the monthly counts
of patients attending ED and the GP walk-in centre, around 14%
monthly reduction in minor attendances at ED was expected. However,
ED routine data did not show any signifi cant reduction in minor
attendances as a result of the opening of the GP walk-in centre. g
Results During the 2-year period before the establishment of the
new ED the mean number of GP patient visits was 1,845 ± 43/month. During the 2-year period after the reform the number was diminished
to 1,364 ± 21/month. This change was not associated with the increase
of the patient visits taken care of by specialists and hospital residents. See Figure 1. Conclusion An extensive reform of the emergency services can notably
reduce patient fl ow into the ED. Reference 1. Boyle A, et al.: Emerg Med Int 2012:838610. [Epub ahead of print] 1. Boyle A, et al.: Emerg Med Int 2012:838610. [Epub ahead of print] p
g
Conclusion These walk-in centres have been shown to increase
accessibility to healthcare service through longer opening hours and
walk-in facility. P259 P259
GP-led walk-in centre in the UK: another way for urgent healthcare
provision
M Arain, J Nicholl, M Campbell
The University of Sheffi eld, UK
Critical Care 2013, 17(Suppl 2):P259 (doi: 10.1186/cc12197) Reference 1. Gerard K, Lattimer V: Preferences of patients for emergency services
available during usual GP surgery hours: a discrete choice experiment. Fam Pract 2005, 22:28-36. Introduction Rio de Janeiro’s Fire Squad is responsible for EMS in
the city. During 2010 we implemented 10 ambulances with EKG
transmission capability in our city. Our intention was to access the
prevalence of acute myocardial infarction in the prehospital setting. Methods We used the Aerotel HeartView EKG system to acquire patient
examination and a blackberry phone to transmit and receive the PDF
EKG trace. The PDF comes with the cardiologist’s interpretation from a
remote hospital, the HCOR São Paulo. Introduction Rio de Janeiro’s Fire Squad is responsible for EMS in
the city. During 2010 we implemented 10 ambulances with EKG
transmission capability in our city. Our intention was to access the
prevalence of acute myocardial infarction in the prehospital setting. P259
GP-led walk-in centre in the UK: another way for urgent healthcare
provision Although the eff ect on the reduction of patients’ load at
the ED is not visible as these centres cover a fraction of the population,
the centre has a potential to divert patients from the ED. P261 Prehospital EKG evaluation in Rio de Janeiro ambulances
RV Vasconcellos, FE Erthal, RV Vargas
Instituto Nacional de Cardiologia, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P261 (doi: 10.1186/cc12199) P260 P260
Establishing a new emergency department: eff ects on patient fl ow
V Rautava1, T Valpas1, M Nurmikari2, A Palomäki1
1Kanta-Häme Central Hospital, Hämeenlinna, Finland; 2City of Hämeenlinna,
Finland
Critical Care 2013, 17(Suppl 2):P260 (doi: 10.1186/cc12198) P260
Establishing a new emergency department: eff ects on patient fl ow
V Rautava1, T Valpas1, M Nurmikari2, A Palomäki1
1Kanta-Häme Central Hospital, Hämeenlinna, Finland; 2City of Hämeenlinna,
Finland
Critical Care 2013, 17(Suppl 2):P260 (doi: 10.1186/cc12198) Overcrowding estimation in the emergency department: is the
simplest score the best? We found a signifi cant correlation between Conclusion Early-PN did not counteract muscle atrophy whereas
it suppressed autophagy and aggravated weakness. Statistically,
muscle weakness was not explained by atrophy or wasting but rather
by impaired autophagy and preservation of muscle density. Thus,
tolerating nutrient restriction early during critical illness may preserve
myofi ber integrity by activating autophagy. i
Results ED staff considered overcrowding as a major concern in the
three EDs. Median OR ranged from 68 to 100, while the NEDOCS
ranged from 64.5 to 76.3. We found a signifi cant correlation between Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S97 the OR and the NEDOCS (Pearson R = 0.973, 0.974 and 0.972), as well as
between the OR, the NEDOCS and the subjective evaluation (P = 0.001). Conclusion Crowding evaluation in the ED requires validated, easy-to-
use, scores. Our study indicates that the simple OR is as accurate as the
complex NEDOCS, allowing continuous crowding assessment. References Figure 1 (abstract P260). GP patient visits before and after the reform of
emergency services. the OR and the NEDOCS (Pearson R = 0.973, 0.974 and 0.972), as well as
between the OR, the NEDOCS and the subjective evaluation (P = 0.001). Conclusion Crowding evaluation in the ED requires validated, easy-to-
use, scores. Our study indicates that the simple OR is as accurate as the
complex NEDOCS, allowing continuous crowding assessment. References 1. Hwang U, McCarthy ML, Bernstein SL, et al.: Measures of crowding in the
emergency department: a systematic review. Acad Emerg Med 2011,
18:527-538. 1. Hwang U, McCarthy ML, Bernstein SL, et al.: Measures of crowding in the
emergency department: a systematic review. Acad Emerg Med 2011,
18:527-538. 2. Hoot NR, Zhou C, Jones I, Aronsky D: Measuring and forecasting emergency
department crowding in real time. Ann Emerg Med 2007, 49:747-755. 2. Hoot NR, Zhou C, Jones I, Aronsky D: Measuring and forecasting emergency
department crowding in real time. Ann Emerg Med 2007, 49:747-755. Establishing a new emergency department: eff ects on patient fl ow
V Rautava1, T Valpas1, M Nurmikari2, A Palomäki1 y
Methods We used the Aerotel HeartView EKG system to acquire patient
examination and a blackberry phone to transmit and receive the PDF
EKG trace. The PDF comes with the cardiologist’s interpretation from a
remote hospital, the HCOR São Paulo. Establishing a new emergency department: eff ects on patient fl ow
V Rautava1, T Valpas1, M Nurmikari2, A Palomäki1
1Kanta-Häme Central Hospital, Hämeenlinna, Finland; 2City of Hämeenlinna,
Finland Critical Care 2013, 17(Suppl 2):P260 (doi: 10.1186/cc12198) p
Results We realized 503 EKG examinations in total. Of these, 248
(49%) had as the chief complaint chest pain, 101 (20%) shortness of
breath, 47 (9%) syncope, 36 (7%) palpitation; other complaints were
15%. We detected 32 examinations (6.36%) with ST elevation MI and Critical Care 2013, 17(Suppl 2):P260 (doi: 10.1186/cc12198) Introduction Overcrowding in emergency departments (EDs) is a
widely known problem. It causes problems and delays in the ED and S98 Critical Care 2013, Volume 17 Suppl 2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P261). Acute inferior MI with complete heart block. Figure 2 (abstract P261). Paroxysmal supraventricular tachycardia. Figure 1 (abstract P261). Acute inferior MI with complete heart block. Figure 1 (abstract P261). Acute inferior MI with complete heart block. Figure 1 (abstract P261). Acute inferior MI with complete heart block. Figure 2 (abstract P261). Paroxysmal supraventricular tachycardia. Figure 2 (abstract P261). Paroxysmal supraventricular tachycardia. Figure 2 (abstract P261). Paroxysmal supraventricular tachycardia. S99 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 leg and patient satisfaction were used to assess the outcome. The
assessments were made before the FIB, at 15 minutes, 30 minutes and
45 minutes. 44 examinations (8.75%) with ST depression. Atrial fi brillation was
detected in 43 examinations (8.5%). See Figures 1 and 2. 44 examinations (8.75%) with ST depression. Atrial fi brillation was
detected in 43 examinations (8.5%). See Figures 1 and 2. Conclusion This experience gave us an idea of the prevalence for
acute ST elevation MI in the prehospital setting, so that we can better
develop our prehospital thrombolysis protocol and focus our training
for cardiology care. Results A convenience sample of 19 patients was enrolled in this study. The compartment block was placed by three mergence ultrasound
trainers. The mean age was 58.5 years. There were 13 females and
six males. P264
Abdominal pain in adolescent females: a single-centre audit and
review of management Abdominal pain in adolescent females: a single-centre audit and
review of management Introduction Abdominal pain in adolescent females has undergone
recent changes with regards to its management under various
specialities. The authors report a single-centre audit looking at the
correct investigation and management of 12-year-old to 16-year-old
girls with abdominal pain in the emergency department setting. Introduction Abdominal pain in adolescent females has undergone
recent changes with regards to its management under various
specialities. The authors report a single-centre audit looking at the
correct investigation and management of 12-year-old to 16-year-old
girls with abdominal pain in the emergency department setting. Methods A single-centre audit and retrospective analysis of patients
took place using case notes and computerised records. Documentation
was analysed using statistical analysis and minimum standards were
set and reviewed. Results In 1 year, 76 long peripheral catheters were inserted in
emergency conditions, using ultrasound guidance. The success rate
was 100%; most lines lasted >1 week and were used for diff erent
purposes, including contrast medium injection. Results After exclusion criteria 62 females between the ages of 12 and
16 presented to the paediatric emergency department in Leicester
with abdominal pain as the predominant admission symptom during
a 12-month period. Documentation of the gynaecological history
was poor (menstrual history 47%, sexual history 14%, contraception
8%), as was the performance of basic investigations (urine dipsticks
65%, pregnancy test 42%). Documentation was analysed with regard
to discharge diagnosis. Ultrasound investigation was performed on
seven of the patients but only once admitted to various specialities. No
ultrasound was undertaken upon admission. p
p
g
j
Conclusion The direct Seldinger technique allows a rapid and safe
placement of the catheter in a vein of the arm or of the forearm,
even when the vessel cannot be palpated or seen, as long as it can
be visualized by ultrasound and it is not deeper than 2 cm. The long
life of this type of peripheral line (up to 2 weeks) is guaranteed by the
material (polyurethane being more biocompatible than Tefl on) and
by the length of the catheter (which reduces the risk of dislodgment). Also, these catheters are particularly cost-eff ective if compared with a
central line or with a midline catheter, since a complete kit including
catheter, 20 G needle and 20 cm guidewire costs between €15 and €20. Conclusion Improvement in documentation of minimum standards
for these patients is needed. P265 Introduction The lumbar plexus block provides excellent analgesia
after hip and knee surgery. One approach to the three major nerves
(femoral, lateral cutaneous and obturator) of the lubar plexus is
the fascia iliaca compartment block, fi rst described by Dalens and
colleagues. Using this blind approach, complete nerve block in the
distribution of the lumbar plexus may be achieved only in 38% of cases. Ultrasound-guided regional techniques off er a number of advantages
including real-time needle guidance and direct observation of local
anesthetic spread within tissue planes. Establishing a new emergency department: eff ects on patient fl ow
V Rautava1, T Valpas1, M Nurmikari2, A Palomäki1 The mean pain score was 9.5 at time 0. The mean pain
score had improved to 5.5 at 15 minutes, 4.5 at 30 minutes and 3 at
45 minutes, respectively. The patient satisfaction was scored 4.5 on a
scale of 1 to 5 where 5 was the most satisfi ed. No patient required any
further analgesia up to 90 minutes and no issues were raised about
the pain or discomfort upon patient transfer. We have no documented
complications or side eff ects. Technique of ultrasound-guided peripheral venous access in the
emergency room f
Conclusion FIB is now widely used by non-anaesthetic trainees to
combat pain in preoperative care due to its safe landmarks. Our
results have shown that it can be used safely and eff ectively for pain
management in hip fracture by emergency physicians, who are trained
in the technique, on the shop fl oor. MG Annetta, G Scoppettuolo, M Biancone, F Toni, M Pittiruti
Catholic University, Rome, Italy MG Annetta, G Scoppettuolo, M Biancone, F Toni, M Pittiruti
Catholic University, Rome, Italy
Critical Care 2013, 17(Suppl 2):P262 (doi: 10.1186/cc12200) Catholic University, Rome, Italy
Critical Care 2013, 17(Suppl 2):P262 (doi: 10.1186/cc12200) Critical Care 2013, 17(Suppl 2):P262 (doi: 10.1186/cc12200) Introduction In emergency situations, patients may need a fast and
reliable peripheral venous access, which sometimes may be diffi cult
to obtain, because of poor visualization of the superfi cial veins due
to edema, obesity, hypovolemia or local abnormalities. In such cases,
insertion of a central line is potentially time consuming and possibly
associated with complications. Furthermore, central lines inserted
in emergency are known to be at high risk of infection, so guidelines
recommend that they should removed within 24 to 48 hours. In this
setting, ultrasound-guided placement of a peripheral venous access
might be more rapid, safer and more cost-eff ective than a central line. Methods We have reviewed retrospectively our experience with the
emergency use of 18 G or 20 G polyurethane catheters, 8 to 10 cm long,
inserted by direct Seldinger technique. P264 Reference 1. Ioannidis JP, Salem D, Chew PW, Lau J: Accuracy and clinical eff ect of out-of-
hospital electrocardiography in the diagnosis of acute cardiac ischemia: a
meta-analysis. Ann Emerg Med 2001, 37:461-470. P264
Abdominal pain in adolescent females: a single-centre audit and
review of management A multidisciplinary care pathway could
improve outcome. Consideration should be given to whether early
ultrasound investigation is appropriate and there is a further need for
investigation as to whether this would improve longer term outcomes. P263 Multidisciplinary approach to improving documentation of visua
acuity in patients presenting with ocular trauma
L Low, M Johnston
NHS Tayside, Dundee, UK
Critical Care 2013, 17(Suppl 2):P265 (doi: 10.1186/cc12203) Critical Care 2013, 17(Suppl 2):P265 (doi: 10.1186/cc12203) Introduction This study aimed to implement a multidisciplinary
quality improvement project in Perth Royal Infi rmary A&E department
to improve documentation of visual acuity (VA) in patients presenting
with ocular injury. Methods We hypothesized that real-time UFIB could be successfully
performed in the ED and would provide an excellent adjunct or
alternative to repeat doses of IVMS for pain control in patients with HFx. The study was conducted at University Hospital, Birmingham, where
we see about 90,000 patients every year. All patients with confi rmed
femoral (neck/shaft) fractures and pain score >7 were included in
the study. Patients with local wounds, or suspected signifi cant pelvic
injury were excluded. A combination of 15 ml lignocaine 1% and 15 ml
bupvivcaine 0.25% was used. VAS (0 to 10), pain on movement of the Methods The improvement project involves a three-pronged multi-
disciplinary approach: ensure that equipment required for VA testing
(Snellen chart and pinhole mask) was readily available; encourage VA
testing at fi rst point of contact with A&E staff , both nursing/medical staff ;
and refresher online course on how to test for VA quickly and accurately,
in the 6/X format. We compared the pre-intervention (2 September to
2 October 2012) and post-intervention (11 October to 19 November
2012) rates of VA documentation using the chi-square test. S100 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P265). Table 1 (abstract P266). Comparison of children receiving BiPAP for status
asthmaticus by severity of illness Table 1 (abstract P266). Multidisciplinary approach to improving documentation of visua
acuity in patients presenting with ocular trauma
L Low, M Johnston
NHS Tayside, Dundee, UK
Critical Care 2013, 17(Suppl 2):P265 (doi: 10.1186/cc12203) Comparison of children receiving BiPAP for status
asthmaticus by severity of illness
Moderate
Severe
(n = 206)
(n = 197)
P value
Age (years)
6.8 ± SD 2.8
7.1 ± SD 4.2
X
Gender
62% male
61% male
X
History of asthma
92%
87%
X
Prior PICU admission
21%
32%
X
Prior history of BiPAP use
19%
24%
X
Prior history of intubation
6%
7%
X
Mean triage for BiPAP start (hours)
2.8 ± SD 2.1
1.2 ± SD 1.25
<0.0001
Mean time on BiPAP (hours)
3.9 ± SD 3.6
5.6 ± SD 5.2
<0.0001
Median time on BiPAP (hours)
2.7
3.8
X
Admission to PICU
58%
75%
X
Total length of hospital stay
42 ± SD 26
50 ± SD 33
<0.06
Serious complications
None
None
X
72-hour returns
n = 1 (0.5%) –
n = 4 (2%) –
X
no admits
no admits Results During the pre-intervention period, of a total of 44 patients
who presented to A&E with eye injury, only 36 patients (95%) had their
VA tested. Following intervention, there was signifi cant improvement
in VA testing, where all 43 patients presenting to A&E with eye injuries
had their VA tested (100%, P = 0.02). Documentation of VA in the correct
(6/X) format increased from 82 to 84% following intervention. There
was a 15% improvement in documentation of best-corrected VA, from
48 to 63% post intervention. See Figure 1. Results During the pre-intervention period, of a total of 44 patients
who presented to A&E with eye injury, only 36 patients (95%) had their
VA tested. Following intervention, there was signifi cant improvement
in VA testing, where all 43 patients presenting to A&E with eye injuries
had their VA tested (100%, P = 0.02). Documentation of VA in the correct
(6/X) format increased from 82 to 84% following intervention. There
was a 15% improvement in documentation of best-corrected VA, from
48 to 63% post intervention. See Figure 1. g
Conclusion Through a multidisciplinary approach, we were successful
in achieving our target of 100% VA documentation rate in all patients
presenting with eye injury to PRI A&E. P267
Report of an outbreak of toxicity from a novel drug of abuse in the
UK: Eric-3 C Morden, S Haig, C Kelly Conclusion All patients recovered without any sequelae. Three patients’
hair started to grow 3 months from ingestion of thallium, and after half
a year their hair was restored to their former state. We had diffi culty
ordering iron(III)hexacyanoferrate(II) because this is also known as an
antidote for cesium. On 11 March 2011 a megathrust earthquake and
tsunami hit Japan and the giant tsunami gave rise to an accident at
a nuclear power generation plant. Because the rumor of radioactive
substances including cesium might be spread was the talk in the city
near the nuclear power plant, the authorities put the antidote under
heavy supervision. We could also collect the data for the course of
thallium concentration. Thallium concentration of the patients who
had an antidote was reduced more rapidly but these patients had a
loose stool, thought to be a side eff ect of this antidote. Great Western Hospital, Swindon, UK Critical Care 2013, 17(Suppl 2):P267 (doi: 10.1186/cc12205) Introduction We present a case series of toxicity due to a novel sub-
stance in the UK: Eric-3. Novel drugs of abuse are becoming more
common throughout the world, and they represent particular
diffi culties in their acute management. A recent report from the
European Monitoring Centre for Drugs and Drug Addiction and
Europol has reported 49 new psychoactive substances reported via its
early warning system. Methods This was a retrospective case-note review over a 6-month
period. Patients were included if their presentation was due to recent
ingestion of Eric-3. Physiological data, symptoms, outcome and
destination of the patient from the emergency department were
collected. Postmortem toxicological analysis was obtained for one of
the two patients who died.i Reference 1. Centers for Disease Control and Prevention: Thallium poisoning from eating
contaminated cake – Iraq, 2008. MMWR Morb Mortal Wkly Rep 2008,
57:1015-1018. 1. Centers for Disease Control and Prevention: Thallium poisoning from eating
contaminated cake – Iraq, 2008. MMWR Morb Mortal Wkly Rep 2008,
57:1015-1018. Results Forty-one attendances were identifi ed from 18 patients. Two patients died and fi ve were admitted to the ICU. Heart rate and
temperature on arrival tended to be above normal (mean heart rate
was 112 bpm, with an SD of 18; mean temperature was 37.45°C with an
SD of 0.95). In total, 63.4% of attendances included agitation and 34.1%
choreiform movements. α-Methyltryptamine and 3-/4-fl uoroephedrine
were found in the blood of one of the patients who died. P270 Heart rate variability in children with tricyclic antidepressant
intoxication BiPAP for treating moderate and severe asthma exacerbations in a
PED A Williams, T Abramo A Williams, T Abramo Introduction BiPAP utilization for the treatment of severe refractory
status asthmaticus patients has become an accepted therapy but is
not well described for moderate exacerbations. We sought to analyze
outcomes from our BiPAP quality database for children presenting in
status asthmaticus at varying levels of severity. Methods PED status asthmaticus patients requiring BiPAP from
1 January 2010 to 31 August 2012 had a bedside interview and
documentation of information at the time therapies were given. Incomplete data were collected retrospectively. All data were stored
and analyzed using a RedCap database. Subjects were stratifi ed into
severity groups based on asthma score at the time of BiPAP placement. Results There were 206 subjects in the moderate severity group
and 197 in the severe group. Table 1 shows the groups were well matched and compares other pertinent data. Children with severe
presentations were placed on BiPAP sooner (P <0.001) and remained
on BiPAP longer (P <0.001). The moderate group had a longer wait
until BiPAP placement. Tables 2 and 3 demonstrate higher initial BiPAP
(IPAP/EPAP) settings with increasing age and severity. Figure 1 trends
initiation and termination asthma scores stratifi ed by severity at BiPAP matched and compares other pertinent data. Children with severe
presentations were placed on BiPAP sooner (P <0.001) and remained
on BiPAP longer (P <0.001). The moderate group had a longer wait
until BiPAP placement. Tables 2 and 3 demonstrate higher initial BiPAP
(IPAP/EPAP) settings with increasing age and severity. Figure 1 trends
initiation and termination asthma scores stratifi ed by severity at BiPAP Figure 1 (abstract P266). Initiation and termination asthma scores grouped by severity. Figure 1 (abstract P266). Initiation and termination asthma scores grouped by severity. S101 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 placement. More of the severe group was admitted to the PICU and
had overall longer hospitalizations (P <0.06). None experienced severe
complications. Conclusion BiPAP is a benefi cial therapy for children presenting to
the PED with severe asthma exacerbations. It may have utility for less
severe asthma exacerbations. Table 2 (abstract P266). BiPAP for treating moderate and severe asthma exacerbations in a
PED Three patients consented to oral
administration of an antidote. Two patients rejected administration
because their symptoms were mild and getting better. All symptoms of
all patients gradually disappeared by August. We also followed up the
course of blood concentration of thallium. The concentration in three
patients who took the antidote was reduced more rapidly than the two
patients who did not take it. p
Conclusion BiPAP is a benefi cial therapy for children presenting to
the PED with severe asthma exacerbations. It may have utility for less
severe asthma exacerbations. P267 BiPAP for treating moderate and severe asthma exacerbations in a
PED Mean settings by age: moderate group
Age (years)
IPAP
EPAP
<3 (n = 22)
12.1 ± SD 2.1
6.4 ± SD 1.3
3 to 4 (n = 48)
12.1 ± SD 1.6
6.4 ± SD 1.2
5 to 6 (n = 51)
13.0 ± SD 2.3
6.6 ± SD 1.3
7 to 8 (n = 24)
13.9 ± SD 2.1
7.2 ± SD 1.2
9 to 10 (n = 28)
13.9 ± SD3.2
7.4 ± SD 2.0
11 to 12 (n = 25)
13.8 ± SD 1.8
7.0 ± SD 1.3
13 to 14 (n = 10)
13.8 ± SD 3.2
7.4 ± SD 1.4
15 to 17 (n = 8)
17.0 ± SD 2.4
9.5 ± SD 0.9
Table 3 (abstract P266). Mean settings by age: severe group
Age (years)
IPAP
EPAP
<3 (n = 21)
12.7 ± SD 2.0
6.8 ± SD 1.3
3 to 4 (n = 50)
13.3 ± SD 2.8
6.8 ± SD 1.4
5 to 6 (n = 26)
13.9 ± SD 2.2
7.3 ± SD 1.3
7 to 8 (n = 29)
15.0 ± SD 2.4
7.7 ± SD 1.5
9 to 10 (n = 26)
15.7 ± SD 3.5
8.0 ± SD 1.8
11 to 12 (n = 22)
14.5 ± SD 3.2
7.6 ± SD 1.6
13 to 14 (n = 9)
15.3 ± SD 3.2
8.0 ± SD 1.7
15 to 17 (n = 14)
17.5 ± SD 3.0
9.5 ± SD 1.8 References
1. EMCDDA: Europol Annual Report on the Implementation of Council Decision
2005/387/JHA. Lisbon: EMCDDA/Europol; 2011. 2. Brandt SD, et al.: Drug Test Anal 2010, 2:377-382. 3. Sikk K, et al.: Acta Neurol Scand 2007, 115:385-389. 4. Stepens A, et al.: N Engl J Med 2008, 358:1009-1017. 5. Murphree HB, et al.: Clin Pharmacol Ther 1961, 2:722-726. P268
Abstract withdrawn
P269
Thallium group poisoning incident in Japan 2011
Y Namba, R Suzuki, J Sasaki, M Takayasu, K Watanabe, D Kenji, M Hayashi,
Y Kitamura, M Kawamo, H Masaki, E Kyuuno, M Hayashi, M Yamaguchi,
A Maeda
Showa University Fujigaoka Hospital, Yokohama, Japan
Critical Care 2013, 17(Suppl 2):P269 (doi: 10.1186/cc12207) Introduction Thallium is an odorless, tasteless, heavy metal that has
been often used for intentional poisonings. In severe patients, thallium
poisoning produces neuromuscular symptoms such as extreme pain
and muscle weakness. Introduction Thallium is an odorless, tasteless, heavy metal that has
been often used for intentional poisonings. BiPAP for treating moderate and severe asthma exacerbations in a
PED In severe patients, thallium
poisoning produces neuromuscular symptoms such as extreme pain
and muscle weakness. Methods Five case reports. p
Results All patients worked at a pharmaceutical factory. They joined
a tea party held at their workplace at the end of April 2011. The fi ve
patients drank tea from a teapot someone had put thallium in. A few
days later, they complained of femoral numbness and pain caused by
pressure. About a week later, three of fi ve patients had profound hair
loss. Three weeks after the party, they came to our ER. We thought that
their symptoms might be caused by some chemicals. We searched the
keywords: ‘lower extremity pain’, ‘hair loss’ and ‘poison’ in the Internet. As a result, thallium, mercury, lead, and so forth, were suspicious
metals. In those metals, thallium was most likely because it was used
in their factory. We immediately examined the blood concentration of
several metals and ordered iron(III)hexacyanoferrate(II) that is known
as the antidote for thallium poisoning. Only thallium was positive in
the blood metal concentration test. Three patients consented to oral
administration of an antidote. Two patients rejected administration
because their symptoms were mild and getting better. All symptoms of
all patients gradually disappeared by August. We also followed up the
course of blood concentration of thallium. The concentration in three
patients who took the antidote was reduced more rapidly than the two
patients who did not take it. Results All patients worked at a pharmaceutical factory. They joined
a tea party held at their workplace at the end of April 2011. The fi ve
patients drank tea from a teapot someone had put thallium in. A few
days later, they complained of femoral numbness and pain caused by
pressure. About a week later, three of fi ve patients had profound hair
loss. Three weeks after the party, they came to our ER. We thought that
their symptoms might be caused by some chemicals. We searched the
keywords: ‘lower extremity pain’, ‘hair loss’ and ‘poison’ in the Internet. y
y p
p
As a result, thallium, mercury, lead, and so forth, were suspicious
metals. In those metals, thallium was most likely because it was used
in their factory. We immediately examined the blood concentration of
several metals and ordered iron(III)hexacyanoferrate(II) that is known
as the antidote for thallium poisoning. Only thallium was positive in
the blood metal concentration test. Heart rate variability in children with tricyclic antidepressant
intoxication Heart rate variability in children with tricyclic antidepressant
intoxication
EC Dinleyici, Z Kilic, S Sahin, R Tutuncu-Toker, M Eren, ZA Yargic, P Kosger
B Ucar
Eskisehir Osmangazi University Faculty of Medicine, Eskisehir, Turkey
Critical Care 2013, 17(Suppl 2):P270 (doi: 10.1186/cc12208) EC Dinleyici, Z Kilic, S Sahin, R Tutuncu-Toker, M Eren, ZA Yargic, P Kosger,
B Ucar Conclusion In this outbreak in the UK, Eric-3 gave symptoms similar
to other stimulants known as legal highs, including death. It may have
been a novel substance, 3-/4-fl uoroephedrine. This underlines the need
for prospective data collection and early national and international
information sharing. Eskisehir Osmangazi University Faculty of Medicine, Eskisehir, Turkey
Critical Care 2013, 17(Suppl 2):P270 (doi: 10.1186/cc12208) Eskisehir Osmangazi University Faculty of Medicine, Eskisehir, Turkey
Critical Care 2013, 17(Suppl 2):P270 (doi: 10.1186/cc12208) Introduction Tricyclic antidepressant (TCA) intoxication is among the
most common causes of poisonings in our country. The TCA group of Introduction Tricyclic antidepressant (TCA) intoxication is among the
most common causes of poisonings in our country. The TCA group of S102 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 demonstrated edematous wall thickening of gastric mucosa, and
ascites was observed in 44.4% (12/27) of these patients. The patients
with small intestinal anisakiasis demonstrated limited edematous wall
thickening of the intestinal tract, and ascites was observed in 90.9%
(40/44) of these patients. Furthermore, phlegmon of mesentery fat was
observed in 72.7% (32/44) of the small intestinal anisakiasis patients. Conclusion When the cause of abdominal pain cannot be determined
by initial assessment, anisakiasis should be considered, especially if the
patient has a history of raw or undercooked seafood ingestion. In the
ED, certain methods of diagnosis are evaluation of the time to develop
symptoms and CT imaging, and a history of raw or undercooked
seafood ingestion should be considered in the diagnosis. References drugs aff ects the central nervous and cardiovascular systems, resulting
in severe arrhythmia and death. Heart rate variability (HRV) analysis is a
non-invasive assessment method that allows evaluation of the cardiac
autonomic (sympathetic and parasympathetic) activity. The aim of this
study was to evaluate HRV in children requiring ICU stay due to TCA
poisoning. Methods Twenty children with isolated TCA poisoning aged between 3
and 16 years who were hospitalized in the pediatric ICU, between March
2009 and July 2010, and 20 healthy children as a control group were
enrolled. Diagnosing anisakiasis in the emergency department St. Luke’s International Hospital, Tokyo, Japan p
y
p
Critical Care 2013, 17(Suppl 2):P271 (doi: 10.1186/cc12209) Introduction Humans can be incidentally parasitized by third-stage
Anisakis larvae after ingestion of raw or undercooked seafood. Although the clinical symptom of anisakiasis is abdominal pain, the
clinical fi nding is nonspecifi c and may be misdiagnosed as appendicitis,
ileus, and so forth. Every year, >40,000 patients visit our ED and our
hospital near the Tsukiji market (famous seafood market in Tokyo). We
often observed cases of anisakiasis in patients who visited the hospital
with abdominal pain as the chief complaint. Thus, we researched to
determine factors useful to diagnose anisakiasis. Methods We retrospectively reviewed data of 83 patients (58 men,
25 women) diagnosed with anisakiasis in our ED (22 July 2003 to 22
July 2012) and examined the usefulness of clinical history, blood test,
diagnostic imaging, and so forth, for anisakiasis diagnosis. Diagnosis
was made after (A) endoscopically proven Anisakis polypide and (B) a
hematologically positive Anisakis antibody (IgG, IgA) and CT diagnostic
imaging. y
Conclusion The implementation of the TM Project at a community
hospital has a major impact in the eICU and eED Sepsis Management
Program, and improved compliance with recommended care bundles. f Heart rate variability in children with tricyclic antidepressant
intoxication Clinical and electrocardiographic (ECG) fi ndings were noted
in the TCA poisoning group. In both groups, 24-hour time domain
HRV analysis (SDNN, SDANN, SDNNi, RMSDD, NN50, and pNN50) was
performed. We also recorded frequency domain analysis results at the
fi rst 5 minutes and the last 5 minutes of the 24-hour record (VLF, nLF,
nHF, LF/HF ratio). 1. Valle et al.: J Med Case Rep 2012, 6:114. 2. David Hwang et al.: Chonnam Med J 2012, 48:73-75. 1. Valle et al.: J Med Case Rep 2012, 6:114. Results The average level of TCA in the study group was 1,116 ± 635
and TCA levels were positively correlated with the duration of QRS
interval (P <0.01). In time-domain nonspectral evaluation, SDNN
(P <0.001), SDNN (P <0.05), RMSDD (P <0.01), and pNN50 (P <0.01)
were found signifi cantly lower in the TCA intoxication group compared
with the control group, while NN50 (P <0.01) was signifi cantly higher
in value. The spectral analysis (frequency domain) of data recorded at
fi rst 5 minutes after intensive care admission showed that the values of
the nLF (P <0.05) and LF/HF ratio (P = 0.001) were signifi cantly higher
in the TCA intoxication group than the controls, while nHF (P = 0.001)
values were signifi cantly lower. The frequency domain spectral analysis
of data recorded at the last 5 minutes showed a lower nHF (P = 0.001)
in the TCA intoxication group than the controls, and the LF/HF ratio
was signifi cantly higher (P <0.05) in the intoxication group. SDNN
(P <0.001), RMSDD (P <0.01), SDNNi (P <0.01), and pNN50 (P <0.01)
levels were higher in patients with positive ECG fi ndings than those
without positive ECG fi ndings. The LF/HF ratio was higher in seven
children with seizures (P <0.001).i P272 Implementation of a sepsis protocol in a community hospital using
a Telemedicine Program
M Steinman1, CA Abreu Filho1, A Andrade2, R Cal1, N Akamine1, J Teixeira2,
E Silva1, A Kanamura1, M Cenderoglo1, C Lottenberg1
1Hospital Israelita Albert Einstein, São Paulo, Brazil; 2Hospital Municipal Dr. Moysés Deutsch, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P272 (doi: 10.1186/cc12210) Implementation of a sepsis protocol in a community hospital using
a Telemedicine Program Introduction Sepsis has high incidence around the world, approximately
400,000 new cases occur annually in Brazil. In developing countries
it is still an important cause of death due to poor adherence to best
practice medicine protocols, mainly in community hospitals. The
Brazilian mortality rate of septic shock is around 60%. The aim of this
study is to describe the fi rst Brazilian initiative of implementation of a
Telemedicine (TM) Project for therapeutic support of septic patients in
a community hospital in São Paulo, Brazil. Conclusion Existing fi ndings give us an idea about HRV’s value to
determine arrhythmia and predict convulsion risk in TCA poisonings. HRV can be used as a non-invasive method in determining the
treatment and prognosis of TCA poisoning. y
p
,
Methods Since May 2012, a TM Program has been implemented at
two hospitals in São Paulo – Hospital Municipal Dr. Moysés Deutsch
(HMMD), a public, secondary hospital, and Hospital Israelita Albert
Einstein (HIAE), a tertiary private philanthropic entity – due to a
partnership with Brazilian Health Ministry. A TM Central Command
was located at HIAE with Endpoint 97 MXP Cisco® Solution and a
mobile Intern MXP ISDN/IP Cisco® for the remote hospital (HMMD) via
dedicated GB/sec connection. Imaging examinations were evaluated
using PACS technology. At HMMD, the Sepsis Protocol, based on the
Surviving Sepsis Campaign, was started for every recruited patient
admitted to the emergency department (ED) or the ICU, and assessed
by the Central Command through TM with an experienced consultant. Results Over a 6-month period, 33 patients with diagnosis of sepsis
(including severe sepsis and septic shock) admitted to HMMD were
evaluated by skilled doctors of HIAE via TM, and the Surviving Sepsis
Campaign protocol was instituted. In total, 87.8% of the consultations
originated from the ICU and 12.2% from the ED; 16 patients were
male (48.4%), mean age was 44.2 years old. TM improved diagnosis in
12.1% and infl uenced clinical management in 87.9% of the cases; in
11 patients (33.3%) TM consultation led to antibiotic change. Hospital
mortality rate was 33.3%. P271 Diagnosing anisakiasis in the emergency department
T Takabayashi, S Ishimatsu, N Otani, T Mochizuki, R Miyamichi
St. Luke’s International Hospital, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P271 (doi: 10.1186/cc12209) Reference 1. Beale R, et al.: Promoting Global Research Excellence in Severe Sepsis
(PROGRESS): lessons from an international sepsis registry. Infection 2009,
37:222-232. Results Of the 83 patients, 39 had gastric anisakiasis and 44 had small
intestinal anisakiasis. All gastric and small intestinal anisakiasis patients
were diagnosed by methods A and B, respectively. A blood test was
unable to show the specifi c infl ammatory reaction. A history of raw
or undercooked seafood ingestion was noted in 95.2% (79/83) of the
patients. This was observed in 100% (39/39) of the gastric anisakiasis
patients and 91% (40/44) of the small intestinal anisakiasis patients. With regard to the development of symptoms, symptoms for gastric
anisakiasis developed within 48 hours and reached a peak in less than
6 hours, whereas the symptoms for small intestinal anisakiasis reached
a peak in 48 hours and persisted for a maximum of 5 days. Diagnostic
CT imaging revealed that all the patients with gastric anisakiasis P273 Reform of emergency services: immediate eff ects on cardiac care
unit and ICU patient intake AO Alaspää, T Valpas, V Rautava, A Palomäki
Kanta-Häme Central Hospital, Hämeenlinna, Finland
Critical Care 2013, 17(Suppl 2):P275 (doi: 10.1186/cc12213) Introduction Early onset eff ective care in the emergency department
(ED) has been reported to have a great infl uence on the intensive care
patients’ morbidity and mortality [1]. Little is known about the infl uence
of the reorganisation of the ED on patient intake to the ICUs. The aim
of this study was to analyse monthly intake of patients from the ED
to the cardiac care unit (CCU) and ICU before and after the reform of
emergency services. Results HMMD receives an average of 30 cases of stroke monthly,
and thrombolysis did not occur before the implementation of the TM
Project, because of the lack of neurologists available to conduce the
cases. After implementation of the TM Program, six cases of ischemic
stroke were thrombolyzed with alteplase; only one case (16%)
progressed to death from septic shock, and one case (16%) presented
symptomatic intracranial hemorrhage. g
y
Methods In Kanta-Häme Central Hospital, a new ED started on 1 April
2007. Four older emergency rooms were combined into one bigger
emergency department and an observation ward was introduced
with continuous follow-up of vital signs. This study is a retrospective
analysis of the patient intake to the CCU and ICU 1 year before and after
the reorganisation. Using as data the Finnish Intensive Care Quality
Consortium (Intensium, Finland) database and the cardiac database
of the hospital, patient transfer from ED to the ICU and CCU was
collected and analysed. Monthly pre/post comparisons were carried
out statistically by a nonparametric Wilcoxon signed-rank test. Conclusion Thrombolysis in ischemic stroke reduces 30% the risk
of disability and 18% the mortality rate. This procedure has been
only feasible to be done in the community setting because of the
implementation of the TM Project, which permits the presence of a real
time consultation with a specialized neurological team from a tertiary
center. P274 P274
Analysis of emergency calls achieved in a French emergency
dispatching centre: what resources for which patients
O Tilak, J Cuny, N Assez, P Goldstein, E Wiel
Lille University Hospital Center, Lille Cedex, France
Critical Care 2013, 17(Suppl 2):P274 (doi: 10.1186/cc12212) Figure 1 (abstract P275). Combined patient fl ow to the ICU and CCU
before and after the reformation of the ED. Initial Brazilian experience of Telestroke for thrombolysis in a
community hospital CA Abreu Filho1, A Caluza1, M Steinman1, G Silva1, R Cal1, N Akamine1,
J Teixeira2, A Andrade2, E Silva1, A Kanamura1, M Cenderoglo1,
C Lottenberg1 CA Abreu Filho1, A Caluza1, M Steinman1, G Silva1, R Cal1, N Akamine1,
J Teixeira2, A Andrade2, E Silva1, A Kanamura1, M Cenderoglo1,
C Lottenberg1 1Hospital Israelita Albert Einstein, São Paulo, Brazil; 2Hospital Municipal Dr. Moysés Deutsch, São Paulo, Brazil
C
l C
(S
l )
(d
/
) S103 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 analyses, and then returned home. In total, 25.50% of patients were
hospitalized in a medical or surgical department, and 12.42% in the
short-term hospitalised unit of the emergency department (stay
duration <24 hours). Some 5.10% of patients worsened and were
oriented in the ICU. A total 3.77% of patients in a cardiac ICU. In total,
73.84% of patients had stay duration less than 6 hours in the ED, 24.45%
<24 hours. Forty percent of patients supported by fi remen and 54%
supported by private ambulance left the hospital after a single medical
consultation. Introduction Stroke is the leading cause of death in Brazil, the number
of stroke patients receiving thrombolysis therapy in the developing
world is extremely low. Prehospital delay, fi nancial constraints, and
lack of infrastructure are the main barriers for thrombolysis therapy in
developing countries. In this way, telemedicine (TM) allows knowledge
transfer of medical expertise to remote community hospitals, where
there is a shortage of specialized doctors. The aim of this study is
to describe the fi rst Brazilian initiative of a Telestroke Program for
therapeutic support and monitoring cases of acute stroke, and venous
thrombolysis of eligible cases in a community hospital in São Paulo,
Brazil. analyses, and then returned home. In total, 25.50% of patients were
hospitalized in a medical or surgical department, and 12.42% in the
short-term hospitalised unit of the emergency department (stay
duration <24 hours). Some 5.10% of patients worsened and were
oriented in the ICU. A total 3.77% of patients in a cardiac ICU. In total,
73.84% of patients had stay duration less than 6 hours in the ED, 24.45%
<24 hours. Forty percent of patients supported by fi remen and 54%
supported by private ambulance left the hospital after a single medical
consultation. Conclusion Nearly 70% of patients calling the French emergency
medical dispatching centre are sent to hospital. Initial Brazilian experience of Telestroke for thrombolysis in a
community hospital Those transportations
are supported for two-thirds of cases by a private ambulance or
fi remen ambulance. One out of two patients only receive a simple
medical consultation in the ED, and go back home. This may concur
to the defi ciency of using general medicine in town. They prefer
using emergency services for free. Only one patient out of four was
hospitalized more than 24 hours. Methods Since May 2012 a TM Program has been implemented at
two hospitals in São Paulo – Hospital Municipal Dr. Moysés Deutsch
(HMMD), a public, secondary hospital, distant about 60 minutes
from the nearest tertiary center, and Hospital Israelita Albert Einstein
(HIAE), a tertiary private philanthropic entity – due to a partnership
with Brazilian Health Ministry. Patients admitted to the community
hospital’s emergency department with acute ischemic stroke as
possible diagnosis are submitted to brain imaging examinations,
and considered for remote consultation. A TM Central Command was
located at HIAE with Endpoint 97 MXP Cisco® Solution and a mobile
Intern MXP ISDN/IP Cisco® for the remote hospital (HMMD). Imaging
examinations were evaluated using PACS technology. At HIAE the
TM Center has skilled doctors, including neurologists, available for
consultation 24 hours, 7 days a week. They discussed the neurological
cases via TM, and selected stroke patients for intravenous thrombolysis
when timely (3 hours symptom onset). P275 P275
Reform of emergency services: immediate eff ects on cardiac car
unit and ICU patient intake
AO Alaspää, T Valpas, V Rautava, A Palomäki
Kanta-Häme Central Hospital, Hämeenlinna, Finland
Critical Care 2013, 17(Suppl 2):P275 (doi: 10.1186/cc12213) P277 1. Rivers E, et al.: N Engl J Med 2001, 345:1368-1377. Reform of emergency services: immediate eff ects on cardiac care
unit and ICU patient intake Introduction Following patients referred by the French emergency
dispatching centre (SAMU 15) in diff erent hospital services (emergency
or ICU) is not always easy. The purpose of this study was to analyze each
call arriving in the emergency dispatching centre, defi ne resources for
transportation, the host hospital department, and analyze the short-
term outcome of these patients. p
Methods We conducted a prospective, observational study, by
collecting all data recorded at the emergency dispatching centre
in a continuous 24-hour period. Each call creating a medical fi le
was included. We identifi ed the type of call, various administrative
information, the purpose of call, resources triggered, medical advice,
hospital department admission, and clinical evolution. p
p
Results A total of 877 patients were included; 67.80% calling from
home, 15.63% calling from a public place. Firemen ambulances were
sent in 38.89%, private ambulance in 24.65% and an emergency
medical ambulance (SMUR) in 11.23%. Simple medical advice without
transportation was given in 13.19% of cases. In total, 22.18% of patients
were entrusted to the family; 2.89% of patients refused transportation;
69.94% of triggered fi remen ambulances were done by a centre 18
call, without any medical regulation by an emergency physician. A
total of 68.59% patients were referred to the emergency department,
1.2% in ICU, and 1.8% in cardiac ICU. In the emergency room, 50.78%
of patients received a simple medical consultation with biological Figure 1 (abstract P275). Combined patient fl ow to the ICU and CCU
before and after the reformation of the ED. S104 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Results The total decrease in monthly patient infl ow from ED to the ICU
and CCU was 30.1% (P = 0.003); that is, from the mean of 47.7 ± 8.2 to
33.3 ± 8.3 patients (Figure 1). The 7.1% decrease in patients taken into
the ICU (12.9 ± 4.1 to 12.0 ± 4.6 patients) was not statistically signifi cant. However, the intake from the ED to the ICU decreased by 38.6% (from
34.8 ± 6.5 to 21.3 ± 5.1 patients) (P = 0.002). Conclusion Overall there were no statistical diff erences between the
observation years either for overtriage or undertriage. L Escobar Alvaro1, J Jimenez Gonzalez1, A Pelaez Ballesta1,
A Corbatón Anchuelo2 Overtriage and undertriage in a prehospital system over 7 years
L Carenzo, F Barra, A Messina, D Colombo, T Fontana, F Della Corte
Universitè del Piemonte Orientale A. Avogadro, Novara, Italy
Critical Care 2013, 17(Suppl 2):P276 (doi: 10.1186/cc12214) Introduction On 11 May 2011 two moderate magnitude earthquakes
hit the city of Lorca, southeast Spain. They caused 11 deaths, including
two pregnant women and their babies, more than 350 injured, and
moderate or severe damage to 80% of the building in the city, including
Lorca’s public hospital that had to be evacuated.i Introduction The Novara 118 emergency medical system (EMS) dispatch
center manages medical emergency calls coming from a region that
spreads out over 1,400 km2 and includes 88 towns and a population of
385,000 people; inhabitant density is 275 inhabitants/km2. p
p
y
Methods Data collection from 1 January 2005 to 31 December 2011
was obtained (EMS software SaveOnLine® Suite 4; REGOLA s.r.l., Turin,
Italy). Triage used was the Medical Priority Dispatch System and severity
was defi ned by color codes. Methods A descriptive study of the fi les of Lorca’s hospital and the
clinical records of patients that attended our service in the 20 hours
following the second earthquake on 11 May. i
y
Results During the study period the Novara 118 dispatch center
managed a total of 122,384 EMS interventions. Median (interquartile)
overestimation (regardless of severity) was 10.0% (2.4 to 14.1%) while
median underestimation was 1.5% (0.8 to 2.5%). See Tables 1 and 2. Results A total of 225 patients were relocated. Simultaneously 224
patients were treated by the emergency service, until the evacuation
was completed and after then, from 10:00 pm to 3:00 pm the next day,
47 more in a fi eld hospital placed just outside the hospital building. See
Figures 1 and 2. Table 1 (abstract P276). Overtriage (%)
Overtriage
2005
2006
2007
2008
2009
2010
2011
Other
3.03
2.47
5.30
4.95
4.06
4.58
4.17
Cardiac
13.23
15.30
27.06
23.72
19.28
19.17
20.14
Drunk
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Poisoning
10.19
18.92
17.91
8.47
8.85
15.20
11.81
Oncological
0.00
2.78
1.05
1.03
3.03
2.47
0.00
Neurologic
9.97
10.05
15.59
15.18
14.57
13.32
13.32
Not identifi ed
13.03
13.15
11.84
11.07
10.33
11.79
7.23
Mental
1.30
0.41
2.09
2.34
2.30
1.27
2.35
Respiratory
10.67
10.89
22.09
18.63
16.29
18.23
14.12
Trauma
12.97
12.68
10.42
8.59
6.78
7.74
7.56
Table 2 (abstract P276). Reform of emergency services: immediate eff ects on cardiac care
unit and ICU patient intake When observed
individually, trauma showed the only signifi cant overtriage reduction
over time; there were no individual modifi cations in undertriage over
time. Reference Reference Reference
1. Neely KW, Eldurkar JA, Drake ME: Acad Emerg Med 2000, 7:174-180. 1. Neely KW, Eldurkar JA, Drake ME: Acad Emerg Med 2000, 7:174-180. Conclusion According to our results the reform of the ED may have a
signifi cant role in the total infl ow of patients to the CCU and ICU. Reference P278 P278
War injuries and civilian accidents in Afghan children
R Kedzierewicz1, P Ramiara2, M Puidupin3, S Mérat4
1CMA des Alpes, Barby, France; 2HIA Sainte Anne, Toulon, France;
3HIA Desgenettes, Lyon, France; 4HIA Begin, Saint Mandé, France
Critical Care 2013, 17(Suppl 2):P278 (doi: 10.1186/cc12216) P278
War injuries and civilian accidents in Afghan children
R Kedzierewicz1, P Ramiara2, M Puidupin3, S Mérat4
1CMA des Alpes, Barby, France; 2HIA Sainte Anne, Toulon, France;
3HIA Desgenettes, Lyon, France; 4HIA Begin, Saint Mandé, France
Critical Care 2013, 17(Suppl 2):P278 (doi: 10.1186/cc12216) p
Conclusion The large number of casualties during a major disaster is a
global problem, even in the developed countries. When the role of the
intensivist is reviewed, many roles were verifi ed to be important; that
is, as a leader of a medical team or triage offi cer as well as a professional
in the fi eld of specifi c intensive care. However, there are many problems
to be solved in the fi elds of disaster medicine. In order to solve the
diversifi cation or the various medical problems, it is necessary to
compile or systematize a disaster medicine of the world version. The
concept of the compendium and our process of trial are shown in
relation to intensive care. Introduction We intended to compare paediatric traumatic injuries
due to war and others related to civilian accidents (CA). Methods We conducted an observational epidemiologic retrospective
study on paediatric cases (<18 years old) seen at the emergency room
of Kabul International Airport NATO role 3 medical treatment facility
between 2009 and 2012. Introduction We intended to compare paediatric traumatic injuries
due to war and others related to civilian accidents (CA). Methods We conducted an observational epidemiologic retrospective
study on paediatric cases (<18 years old) seen at the emergency room
of Kabul International Airport NATO role 3 medical treatment facility
between 2009 and 2012. Results During 3 years, 341 children were admitted to the emergency
room (ER) with a mean age of 10 years (9.49 to 10.51). Eighty-eight
per cent of children (301) were traumatized, 39% of them (118) due to
war injuries (WI) and 61% (183) due to CA. Forty-three per cent of WI
(gunshot wound 34%, explosion 66%) but only 19% of injuries due to
CA (falls 24%, transport accidents 56%, burns 9%, penetrating injuries
9%, unknown cause 2%) involved polytraumas defi ned as a New Injury
Severity Score (NISS) >15 (P <0.001). References Intensive care and disaster medicine: the role of a compendium
Y Haraguchi1, Y Tomoyasu1, H Nishi1, M Hoshino1, T Tsubata2, M Sakai1,
E Hoshino1
1Disaster Medicine Compendium Team, Japan, Tokyo, Japan; 2Keiyo Hospital,
Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P279 (doi: 10.1186/cc12217) 1. Hill AD, Vingilis E, Martin CM, Hartford K, Speechley KN: Interhospital transfer
of critically ill patients: demographic and outcomes comparison with
nontransferred intensive care unit patients. J Crit Care 2007, 22:290-295. 2. Flabouris A, Hart GK, George C: Outcomes of patients admitted to tertiary
intensive care units after interhospital transfer: comparison with patients
admitted from emergency departments. Crit Care Resusc 2008, 10:97-105. 3. Duke GJ, Green JV: Outcome of critically ill patients undergoing
interhospital transfer. Med J Aust 2001, 174:122-125. 1. Hill AD, Vingilis E, Martin CM, Hartford K, Speechley KN: Interhospital transfer
of critically ill patients: demographic and outcomes comparison with
nontransferred intensive care unit patients. J Crit Care 2007, 22:290-295. 1. Hill AD, Vingilis E, Martin CM, Hartford K, Speechley KN: Interhospital transfer
of critically ill patients: demographic and outcomes comparison with
nontransferred intensive care unit patients. J Crit Care 2007, 22:290-295. p
2. Flabouris A, Hart GK, George C: Outcomes of patients admitted to tertiary
intensive care units after interhospital transfer: comparison with patients
admitted from emergency departments. Crit Care Resusc 2008, 10:97-105. 3. Duke GJ, Green JV: Outcome of critically ill patients undergoing
interhospital transfer. Med J Aust 2001, 174:122-125. 3. Duke GJ, Green JV: Outcome of critically ill patients undergoing
interhospital transfer. Med J Aust 2001, 174:122-125. Introduction There were several catastrophes in the last decades. The
make-up of systematic measuring and life-saving medical systems,
including intensive care, is thought to be an urgent and essential issue. Our eff orts for establishing a disaster medicine and education system
are presented. L Escobar Alvaro1, J Jimenez Gonzalez1, A Pelaez Ballesta1,
A Corbatón Anchuelo2 Undertriage (%)
Undertriage
2005
2006
2007
2008
2009
2010
2011
Other
1.01
0.77
2.87
1.66
1.44
1.48
1.31
Cardiac
2.16
1.44
1.64
1.60
2.15
1.60
1.14
Drunk
0.00
1.89
0.00
0.00
0.00
0.00
0.00
Poisoning
8.33
0.00
0.00
1.69
0.00
1.60
0.00
Oncological
95.74
0.56
0.00
1.03
3.03
2.47
0.00
Neurologic
2.42
2.32
2.96
3.67
3.05
1.40
2.46
Not identifi ed
1.72
2.13
1.87
2.20
2.04
2.74
3.50
Mental
1.74
0.83
1.05
1.17
0.00
0.42
0.39
Respiratory
2.30
4.16
3.76
3.75
2.57
2.72
2.46
Trauma
1.07
0.92
1.50
1.41
1.36
1.31
1.46 Table 1 (abstract P276). Overtriage (%) Table 1 (abstract P276). Overtriage (%) g
Figure 1 (abstract P277). Hospital evacuation. Figure 2 (abstract P277). Emergency service taking care of patients
during evacuation. Table 1 (abstract P276). Overtriage (%)
Overtriage
2005
2006
2007
2008
2009
2010
2011
Other
3.03
2.47
5.30
4.95
4.06
4.58
4.17
Cardiac
13.23
15.30
27.06
23.72
19.28
19.17
20.14
Drunk
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Poisoning
10.19
18.92
17.91
8.47
8.85
15.20
11.81
Oncological
0.00
2.78
1.05
1.03
3.03
2.47
0.00
Neurologic
9.97
10.05
15.59
15.18
14.57
13.32
13.32
Not identifi ed
13.03
13.15
11.84
11.07
10.33
11.79
7.23
Mental
1.30
0.41
2.09
2.34
2.30
1.27
2.35
Respiratory
10.67
10.89
22.09
18.63
16.29
18.23
14.12
Trauma
12.97
12.68
10.42
8.59
6.78
7.74
7.56
Table 2 (abstract P276). Undertriage (%) Figure 1 (abstract P277). Hospital evacuation. Figure 1 (abstract P277). Hospital evacuation. Figure 1 (abstract P277). Hospital evacuation. Figure 2 (abstract P277). Emergency service taking care of patients
during evacuation. Figure 2 (abstract P277). Emergency service taking care of patients
during evacuation. Undertriage
2005
2006
2007
2008
2009
2010
2011
Other
1.01
0.77
2.87
1.66
1.44
1.48
1.31
Cardiac
2.16
1.44
1.64
1.60
2.15
1.60
1.14
Drunk
0.00
1.89
0.00
0.00
0.00
0.00
0.00
Poisoning
8.33
0.00
0.00
1.69
0.00
1.60
0.00
Oncological
95.74
0.56
0.00
1.03
3.03
2.47
0.00
Neurologic
2.42
2.32
2.96
3.67
3.05
1.40
2.46
Not identifi ed
1.72
2.13
1.87
2.20
2.04
2.74
3.50
Mental
1.74
0.83
1.05
1.17
0.00
0.42
0.39
Respiratory
2.30
4.16
3.76
3.75
2.57
2.72
2.46
Trauma
1.07
0.92
1.50
1.41
1.36
1.31
1.46 Figure 2 (abstract P277). Emergency service taking care of patients
during evacuation. L Escobar Alvaro1, J Jimenez Gonzalez1, A Pelaez Ballesta1,
A Corbatón Anchuelo2 S105 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Results Our research indicated that disaster medicine should be
established systematically or it is necessary to compile a compendium
of disaster medicine from a broad perspective or from a bird’s-eye and
long-term view. The Japanese version was tentatively completed with
22 volumes as of the fi nancial year 2005, of which nearly three-quarters
are written in Japanese. Although this worked partly during the above-
shown catastrophe in Japan 2011, several problems are left to be
solved; that is, the insuffi cient operation system of the Japan DMAT or
Disaster Medical Assistant Team that seemed to have caused a large
number of preventable deaths. Results Our research indicated that disaster medicine should be
established systematically or it is necessary to compile a compendium
of disaster medicine from a broad perspective or from a bird’s-eye and
long-term view. The Japanese version was tentatively completed with
22 volumes as of the fi nancial year 2005, of which nearly three-quarters
are written in Japanese. Although this worked partly during the above-
shown catastrophe in Japan 2011, several problems are left to be
solved; that is, the insuffi cient operation system of the Japan DMAT or
Disaster Medical Assistant Team that seemed to have caused a large
number of preventable deaths. Conclusion In less than 3 hours 225 patients were evacuated and 224
were given attention by the emergency service of Lorca’s hospital,
with the support of personnel from other services of the hospital. The
emergency service of the hospital continues to be operative in the
building until evacuation is completed and in a fi eld hospital later. P278 Patients experiencing a traumatic
injury related to war had lower Paediatric Trauma Scores (PTS) than
those injured in a CA (PTS = 6.4 (5.7 to 7.0) vs. PTS = 9.0 (8.7 to 9.4)
respectively, P <0.00001), higher ISS (ISS = 16.7 (13.8 to 19.6) vs. ISS = 8.9
(7.5 to 10.2) respectively, P <0.00001), NISS (P <0.0001), mortality rate
predicted by the Trauma Injury Severity Score (TRISS, P <0.02) or A
Severity Characterization of Traumas (ASCOT, P <0.001). The result is
longer overall hospitalization of patients having WI (P < 0.001) and a
higher number of surgeries (P <0.02). After the ER, 54% of patients with
WI were hospitalized in the ICU (86% of them after surgery) but only
26% of patients involved in a CA (71% after surgery). As many patients
with WI as involved in a CA (40%) were admitted to the ward (89% of
patients with WI after surgery but only 63% of patients with injuries
due to a CA). Thirty-three per cent of patients involved in a CA returned
home and one was transferred, whereas only three patients with WI
returned home after being in the ER, three patients were transferred
and one died in the operating room. Observed paediatric mortality in
our medical treatment facility was 2.9% (10 children out of 341): three
children died of WI, three due to a CA and one of septic shock due to a
medical cause. Factors involved in ICU mortality following medical retrieval:
identifying diff erences between ICU survivors and nonsurvivors
M Kennedy1, P Visser1, L Harriss2 y
1Ambulance Victoria, Melbourne, Australia; 2Monash University, Melbourne,
Australia y
1Ambulance Victoria, Melbourne, Australia; 2Monash University, Melbourne,
Australia
l
l
d Critical Care 2013, 17(Suppl 2):P280 (doi: 10.1186/cc12218) Introduction The study aimed to determine factors related to ICU
mortality in critically ill patients transferred by Adult Retrieval Victoria
(ARV) medical staff . Patients who died in the ICU after interhospital
transfer were compared with those who survived. Methods This is a retrospective cohort study of ARV cases between
1 January 2009 and 30 June 2010. Retrieval data were linked with data
from the ANZICS CORE APD (Australia and New Zealand Intensive Care
Society Centre for Outcome and Resource Evaluation Adult Patient
Database). Victoria Data Linkage (VDL) performed linkage of the data. Data included demographic and clinical data obtained during transfer
and data related to outcome measures in the ICU. Results Of the 601 cases transferred by ARV during the study period,
549 were eligible for linkage to 25,543 ANZICS APD case records for
the same period. VDL matched 460 of these cases (83.8%). Logistic
regression analysis revealed the variables associated with mortality
were advanced age (OR = 1.02, 95% CI = 1.00 to 1.04, P = 0.02), cardiac
principal referral problems (OR = 1.84, 95% CI = 1.02 to 3.32, P = 0.04),
lower mean arterial blood pressure (OR = 0.97, 95% CI = 0.95 to 0.99,
P = 0.005) and tachycardia (OR = 1.02, 95% CI = 1.00 to 1.03, P = 0.008)
on arrival at the destination hospital. Conclusion War injuries are more prone to cause polytrauma than CA. According to the PTS, ISS, NISS, TRISS and ASCOT, children experiencing
WI have higher severity scores and predicted mortality rate than others,
stay longer in the hospital and have more surgeries. Conclusion Advanced age, lower mean arterial blood pressure and
tachycardia towards the completion of transfer were associated with
increased ICU mortality in this population. Clinicians should be aware
of the additional risk for cardiac patients. P282 P282
Are physicians required during HEMS winch rescue missions? PB Sherren, C Hayes-Bradley, C Reid, B Burns, K Habig
Greater Sydney Area HEMS, Sydney, Australia
Critical Care 2013, 17(Suppl 2):P282 (doi: 10.1186/cc12220) Introduction A winch-capable helicopter emergency medical service
(HEMS) off ers several advantages over standard rescue operations. Little is known about the benefi t of physician winching in addition
to a highly trained paramedic. We analysed the mission profi les and
interventions performed during rescues involving the winching of a
physician in the Greater Sydney Area HEMS (GSA-HEMS). y
g y
p y
p
Methods A suggested algorithm for TCA was developed based on the
Greater Sydney Area Helicopter Emergency Medical Service’s standard
operating procedures and current available evidence. p
g p
Results An algorithm for the general management of TCA can be seen in
Figure 1. In TCA, priority should be given to catastrophic haemorrhage
control (tourniquets, direct pressure, haemostatic agents, pelvic
and long bone splintage) and volume resuscitation. Simultaneous
oxygena tion optimisation should occur with proactive exclusion of
tension pneumothoraces with bilateral open thoracostomies. Cardiac
ultrasound (US) should be used to help exclude cardiac tamponade
and assist in prognostication. The US presence of true cardiac standstill
versus low pressure state/pseudo-PEA, and an ETCO2 <1.3 kPa carries
a grave prognosis in TCA. Given the high incidence of hypovolaemia,
hypoxia and obstructive shock prior to TCA, the role of adrenaline and
chest compressions are limited. Figure 2 shows a suggested algorithm
for the management of penetrating TCA requiring prehospital
thoracotomy. y
y
y
Methods All winch missions involving a physician from August 2009
to January 2012 were identifi ed from the prospectively completed
GSA-HEMS electronic database. A structured case-sheet review
for a predetermined list of demographic data and physician-only
interventions (POI) was conducted. Results We identified 130 missions involving the winching of a
physician, of which 120 case sheets were available for analysis. The
majority of patients were traumatically injured (90%) and male (85%)
with a median age of 37 years. Seven patients were pronounced life
extinct on the scene. A total of 63 POI were performed on 48 patients. Administration of advanced analgesia was the most common POI
making up 68.3% of interventions. Patients with abnormal RTSc2
scores were more likely to receive a POI when compared with those
with normal RTSc2 (P = 0.03). The performance of POI had no effect
on median scene times (45 vs. 1.
Lockey D, et al.: Ann Emerg Med 2006, 48:240-244. Algorithm for the resuscitation of traumatic cardiac arrest patients
in a physician-staff ed helicopter emergency medical service
PB Sh
C R d
b
B B Methods Mega-disasters or catastrophes are researched basically on
actual medical experience; that is, the 9/11 attack in 2001, Hurricane
Katrina in 2005, Indian Ocean earthquake with tsunami in 2004, the
Chernobyl incident in 1986 as well as the Higashinihon earthquakes
with tsunami, which was followed by the severest degree of nuclear
disaster in 2011. f
PB Sherren, C Reid, K Habig, B Burns g
Greater Sydney Area HEMS, Sydney, Australia y
y
, y
y,
Critical Care 2013, 17(Suppl 2):P281 (doi: 10.1186/cc12219) Introduction Survival rates following traumatic cardiac arrest (TCA)
are known to be poor but resuscitation is not universally futile [1]. S106 Critical Care 2013, Volume 17 Suppl 2
htt //
f
/
l
t /17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P281). Traumatic cardiac arrest algorithm. Figure 1 (abstract P281). Traumatic cardiac arrest algorithm. There are distinct diff erences in the pathophysiology between medical
cardiac arrests and TCA. Traumatic pathologies associated with an
improved chance of successful resuscitation include hypoxia, tension
pneumothorax and cardiac tamponade [1]. The authors believe a
separate algorithm is required for the management of out-of-hospital
TCA attended to by a highly trained physician and paramedic team. P282 43 minutes; P = 0.51). See Tables 1
and 2. Conclusion The suggested algorithm is designed for a highly trained
physician-led prehospital team and aims to maximise the number of
neurologically intact survivors in out-of-hospital TCA. Reference 1. Lockey D, et al.: Ann Emerg Med 2006, 48:240-244. Conclusion Our high POI rate of 40% coupled with long rescue times
and the occasional severe injuries supports the argument for winching
doctors. Not doing so would deny a signifi cant proportion of patients
time-critical interventions, advanced analgesia and procedural
sedation. S107 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 2 (abstract P281). Thoracotomy algorithm. Figure 2 (abstract P281). Thoracotomy algorithm. Figure 2 (abstract P281). Thoracotomy algorithm. P283
Improving maintenance of critical care land and aeromedical
transfer equipment
D Ashton-Cleary, N Boyd
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P283 (doi: 10.1186/cc12221)
Introduction The aim was to assess the content and state of repair of
equipment carried for transfer of critical care patients to other hospitals. By chance, several items of date-expired stock were identifi ed in the
transfer kit whilst moving a patient to a tertiary centre. This raised the
possibility of a more extensive problem with the equipment bags. Due to the geographical location of our district general hospital we
undertake around 70 transfers of critical care patients to other hospitals
per year (16% by air) and it is clearly important that our equipment is
well maintained for these journeys. Table 1 (abstract P282). Demographic data, timings and Coded Revised
Trauma Score (RTSc2)
Total (n = 120)
Male
102 (85)
Median (IQR) age (years)
37 (26 to 53)
Received a physician-only intervention
48 (40)
Pronounced life extinct on arrival
7 (5.8)
RTSc2
7.8408
104 (86.7)
7.0001 to 7.8407
2 (1.7)
6.0001 to 7.0000
4 (3.3)
5.0001 to 6.0000
4 (3.3)
4.0001 to 5.0000
0 (0)
<4.0001
6 (5)
Table 2 (abstract P282). Interventions performed
Physician-only intervention
Number of interventions (n = 63)
Analgesia/procedural sedation
Intravenous ketamine
42 (66.7)
Intravenous fentanyl
1 (1.6)
Fascia iliaca compartment block
1 (1.6)
Airway management
Rapid sequence induction and intubation
4 (6.3)
Surgical airway
1 (1.6)
Circulatory support
Adult intraosseous access
1 (1.6)
Blood transfusion
2 (3.2)
Orthopaedic manipulation of joint/limb
6 (9.5)
Thoracostomy
1 (1.6)
Diagnostic ultrasound
1 (1.6)
Hypertonic saline administration
3 (4.8)
Data presented as n (%). Safety standards for intrahospital transfer of critical care patients
D Ashton-Cleary
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P284 (doi: 10.1186/cc12222) Introduction The aim was to assess care of patients during intrahospital
transfer. The UK Royal College of Anaesthetists has defi ned auditable
standards for the care of patients and the training of escorting medical
and nursing staff in this context [1]. p
y
y
Conclusion Prehospital hyperoxemia did not infl uence the functional
neurological outcome. One of the reasons for this fi nding could be
the short arrival time to the trauma center where repeated analyses of
arterial blood gases were performed. Therefore, correction of fraction
of inspired oxygen according to the arterial blood gas analysis shortens
the time of hyperoxemia, thus reducing neuronal brain damage. References f
Methods Patients in a 27-bed combined general and neurosurgical
critical care unit were studied in January 2011 and May 2012. Patients
undergoing radiology department imaging or intervention were
identifi ed from the electronic imaging library. Records of these transfers
were sought in the critical care electronic notes and the standards of
documentation graded on a fi ve-point scale (very good, good, average,
minimal, absent). Documentation of the grade and training of escorting
staff was also sought. Between the two study periods, a transfer safety
checklist was introduced. 1. Beynon et al.: Brain tissue oxygen monitoring and hyperoxic treatment in
patients with traumatic brain injury. J Neurotrauma 2012, 29:2109-2123. 1. Beynon et al.: Brain tissue oxygen monitoring and hyperoxic treatment in
patients with traumatic brain injury. J Neurotrauma 2012, 29:2109-2123. 1. Beynon et al.: Brain tissue oxygen monitoring and hyperoxic treatment in
patients with traumatic brain injury. J Neurotrauma 2012, 29:2109-2123. 2. Brenner et al.: Association between early hyperoxia and worse outcomes
after traumatic brain injury. Arch Surg 2012, 16:1-5. p
j
y
,
2. Brenner et al.: Association between early hyperoxia and worse outcomes
after traumatic brain injury. Arch Surg 2012, 16:1-5. Results A total of 20.9% of 143 patients underwent one or more
transfers in January 2011 (40 transfers). In May 2012, 26.4% of 151
patients underwent 57 transfers. In the fi rst period, documentation
was graded as minimal (limited to a statement that the patient had
left the critical care unit) or absent in 77.5% of transfers. In the 62.5%
of patients transferred whilst on invasive ventilation, 88.0% had no
documentation by the doctor and in 84.0% it was not known which
doctor had escorted the patient. P285 Prehospital hyperoxemia does not infl uence the functional
neurological outcome in polytraumatized patients with traumatic
head injury
V Vujanovic Popovic1, T Pelcl1, M Spindler1, Z Klemenc Ketis2, M Strnad1
1ZD dr. Adolfa Drolca Maribor, Slovenia; 2University of Maribor, Slovenia
Critical Care 2013, 17(Suppl 2):P285 (doi: 10.1186/cc12223) g
g
g
Results A total of 13.9% of drug items and 29.2% of equipment items
had expired or would do so within 30 days of the initial assessment. The
combined weight of one equipment and one drug bag was reduced
from 14 to 9 kg (36% reduction) by introducing the new inventory. At
reassessment in November 2012, only 10 items of equipment (3.2%)
were expired or near to expiry and there were no expired drug items
(4.1% near to expiry). In total, 0.3 kg (26 small items) of extraneous
equipment had been added through over-restocking and was removed. Conclusion These bags are designed for a clinician to manage a patient
when an emergency arises during transfer of a critical care patient. By
the introduction of simple measures, the risks posed by expired items
or cluttered equipment bags have almost been eradicated. Signifi cant
weight savings have been made; this off ers improved ergonomics
for staff and is also an important consideration for aeromedical
operations. Our department was surprised to discover the extent of
decline of our equipment and it may be that other departments would
fi nd themselves in a similar position. The anaesthetic registrars who
routinely escort the transfer patients have a vested interest to maintain
this equipment and this has secured their buy-in to the new checking
procedure with clear results. P284 Safety standards for intrahospital transfer of critical care patients
D Ashton-Cleary
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P284 (doi: 10.1186/cc12222) P286 Innate immune response-mediated late increase in SuPAR in
multi-trauma patients
K Timmermans, M Kox, M Vaneker, P Pickkers
Radboud University Nijmegen Medical Center, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P286 (doi: 10.1186/cc12224) P282 Table 2 (abstract P282). Interventions performed
Physician-only intervention
Number of interventions (n = 63)
Analgesia/procedural sedation
Intravenous ketamine
42 (66.7)
Intravenous fentanyl
1 (1.6)
Fascia iliaca compartment block
1 (1.6)
Airway management
Rapid sequence induction and intubation
4 (6.3)
Surgical airway
1 (1.6)
Circulatory support
Adult intraosseous access
1 (1.6)
Blood transfusion
2 (3.2)
Orthopaedic manipulation of joint/limb
6 (9.5)
Thoracostomy
1 (1.6)
Diagnostic ultrasound
1 (1.6)
Hypertonic saline administration
3 (4.8)
Data presented as n (%). Table 2 (abstract P282). Interventions performed Table 1 (abstract P282). Demographic data, timings and Coded Revised
Trauma Score (RTSc2) transfer kit whilst moving a patient to a tertiary centre. This raised the
possibility of a more extensive problem with the equipment bags. Due to the geographical location of our district general hospital we
undertake around 70 transfers of critical care patients to other hospitals
per year (16% by air) and it is clearly important that our equipment is
well maintained for these journeys. Introduction The aim was to assess the content and state of repair of
equipment carried for transfer of critical care patients to other hospitals. By chance, several items of date-expired stock were identifi ed in the S108 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Methods We maintain two identical sets of equipment (syringes,
fl uid, airway management items, and so forth) and drug bags to take
on transfers; one equipment and one drug bag taken on each trip. The contents of all four bags were checked and itemised. By careful
consideration of the aims of the bags (to provide emergency equipment
and drugs for managing one patient during an en-route emergency) a
new inventory was devised. Excess items were removed to lighten the
bags and improve accessibility to the essential items. Expired stock was
removed. A daily checking procedure and tamper-proof seals on the
bags were instigated and the bags were reassessed 12 months later. hospital protocols and specifi c training courses to improve standards
of transfer medicine locally. The study also identifi ed our portable head
CT scanner to have the potential to reduce transfers by 52% and so this
has been strongly promoted. Reference Safety standards for intrahospital transfer of critical care patients
D Ashton-Cleary
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P284 (doi: 10.1186/cc12222) There was only slight improvement
in the second period (71.9% minimal or absent documentation, 80.0%
no documentation by the doctor, 72.0% not known which doctor
escorted). In the documentation available, six severe complications
were noted during the second period (including episodes of severe
bradycardia, hypotension and pupil dilatation). 3. Davis et al.: Both hypoxemia and extreme hyperoxemia may be
detrimental in patients with severe traumatic brain injury. J Neurotrauma
2009, 26:2217-2223. Prehospital hyperoxemia does not infl uence the functional
neurological outcome in polytraumatized patients with traumatic
head injury y
V Vujanovic Popovic1, T Pelcl1, M Spindler1, Z Klemenc Ketis2, M Strnad1
1ZD dr. Adolfa Drolca Maribor, Slovenia; 2University of Maribor, Slovenia
Critical Care 2013, 17(Suppl 2):P285 (doi: 10.1186/cc12223) Introduction The association between hyperoxemia and neurological
outcome in trauma patients is not clear. We examined the association
between prehospital hyperoxemia and neurological outcome in
polytraumatized patients. Introduction The association between hyperoxemia and neurological
outcome in trauma patients is not clear. We examined the association
between prehospital hyperoxemia and neurological outcome in
polytraumatized patients. p y
p
Methods This was a retrospective study of polytraumatized patients
with traumatic head injury who were endotracheal intubated and
ventilated with supplemental oxygen (100%) in the prehospital
emergency setting. Arterial partial oxygen pressure (PaO2) was
measured after arrival at the hospital trauma center. We included
patients with initial PaO2 above 160 mmHg (hyperoxemia group). The severity of the trauma was determined upon the admission to
the hospital by the Injury Severity Scale (ISS) and the outcome was
assessed at the discharge from the hospital using the Glasgow Coma
Scale (GCS), Glasgow Outcome Scale (GOS) and Cerebral Performance
Categories scale (CPC). Mann–Whitney’s test was used for data analysis. Results Sixty patients were involved in the study. Forty-eight (80%) of
them were men and 86.7% sustained blunt trauma. Hyperoxemia was
present in 41.6% of patients. Initial average ISS was 38, in patients with
normoxemia 32.5 and in patients with hyperoxemia 35.4. Discharge
GCS, GOS and CPC in the hyperoxemia group compared with the
normoxemia group were 9.86 versus 9.33 (P = 0.503), 2.52 versus
2.24 (P = 0.613) and 3.10 versus 3.19 (P = 0.936) with the duration of
hospitalization of 26.64 days versus 27.72 days (P = 0.984).l Reference 1. Morgan G: Critical care services. In Raising the Standard: A Compendium of
Audit Recipes. 2nd edition. London: Royal College of Anaesthetists; 2006. P287 Single-dose estrogen infusion can amplify brain levels of Sonic
hedgehog, a signal protein for neuro stem cells and repair following
the indirect brain injury resulting after severe torso burns
PE Pepe1, JG Wigginton1, JW Gatson1, J Simpkins2, D Maass1,
K AbdelFattah1, AH Idris1, V Warren1, JP Minei1
1University of Texas Southwestern Medical Center, Dallas, TX, USA; 2University
of North Texas, Fort Worth, TX, USA
Critical Care 2013, 17(Suppl 2):P287 (doi: 10.1186/cc12225) Introduction Many proposed resuscitative therapies for cardiac arrest
and trauma will require the earliest possible intervention and would
occur under volatile circumstances, making true informed consent for
clinical trials unfeasible. The purpose here was to report our experience
using exception to informed consent during the inaugural pilot study
of infusing estrogen for acute injury, the so-called RESCUE Shock study. Methods Fifty patients were enrolled in RESCUE Shock in which
estrogen or placebo was infused as soon as possible in the emergency
department for trauma patients with a low systolic blood pressure
(<90 mmHg) at two level I trauma centers. They were all treated with
a single-dose estrogen or placebo infusion within 2 hours using
exception from informed consent following US federal guidelines. Introduction Severe burn patients are often noted to have subsequent
neurocognitive problems. Experimentally, we have found striking,
prolonged elevations of infl ammatory markers in the brain (for
example, IL-6) even when the injury occurs in a remote anatomic
location. This neuroinfl ammatory response can also be signifi cantly
blunted by a single post-burn dose of estrogen. Sonic hedgehog (SHH),
an important signaling protein found in the brain, controls and directs
diff erentiation of neural stem cells, infl uencing brain regeneration and
repair by generating new neurons throughout life. As estrogens not
only blunt infl ammation but also exert an infl uence on a variety of stem
cells, we hypothesized that 17β-estradiol (E2) might aff ect levels of SHH
in the post-burn rat brain. Results Investigator-initiated exception from informed consent
studies is feasible, with our FDA IND approval obtained in 31 days,
IRB 1 approval in 25 days, and IRB 2 approval in 24 days. Community
consultation/notifi cation was successfully accomplished with no one
opting out and 47/50 enrolled patients or their legal representatives
were notifi ed of participation (one died unidentifi ed, two died with no
known contact). Innate immune response-mediated late increase in SuPAR in
multi-trauma patients p
K Timmermans, M Kox, M Vaneker, P Pickkers
Radboud University Nijmegen Medical Center, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P286 (doi: 10.1186/cc12224) K Timmermans, M Kox, M Vaneker, P Pickkers
Radboud University Nijmegen Medical Center, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P286 (doi: 10.1186/cc12224) Introduction The soluble form of urokinase-type plasminogen activator
(suPAR) has been identifi ed as a marker for immune activation and is
demonstrated to accurately predict outcome in patients with sepsis or
infectious diseases. In multi-trauma patients a considerable immunological
response also occurs that is related to multiple organ failure and patient
outcome. We investigated the kinetics of suPAR, correlation with the
immune response and outcome in multi-trauma patients. Conclusion On average our unit conducts nearly two critical care
transfers each day. Severe complications seem to complicate at least
10% of these, stressing the risk, need for good care and ongoing
training. The intervention made in this audit had little impact on the
standard of documentation. However, it has raised the issue within the
consciousness of the staff . It is important to identify interventions that
have failed to reach a gold standard to provide the impetus to seek
other solutions. As a result of this study, the author has devised new Methods Blood was obtained from adult multi-trauma patients (n = 63)
on arrival at the emergency room (ER) of the Radboud University S109 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Nijmegen Medical Centre and days 1, 3, 5, 7, 10 and 14 following trauma. Plasma concentrations of TNFα, IL-6, IL-10, IFNγ, IL-8 and MCP-1 were
determined by Luminex, and SuPAR concentrations using ELISA. Clinical
data were collected from electronic patient fi les. Concentrations, areas
under the curve (AUC) and regression coeffi cients were statistically
analyzed. Spearman correlation coeffi cients were calculated and
diff erences between survival/nonsurvival groups were analyzed using
unpaired Student t tests. Methods Male rats (n = 44) were assigned randomly into three groups:
controls/no burn (n = 4); burn/placebo (n = 20); and burn/E2 (n = 20). Burned rats received a 40% 3° TBSA dorsal burn, fl uid resuscitation and
one dose of E2 or placebo (0.5 mg/kg intraperitoneally) 15 minutes post
burn. Innate immune response-mediated late increase in SuPAR in
multi-trauma patients Eight animals from each of the two burn groups (burn/placebo
and burn/E2) were sacrifi ced at 24 hours and at 7 days, respectively
(sham group at 7 days only), with four each of the two burn groups
sacrifi ced at 45 days. Brain tissue samples were analyzed by ELISA for
SHH. p
Results SuPAR values at admission to the ER were higher in nonsurvivors
compared with survivors (n = 16, mean ± SEM 4.1 ± 0.6 ng/ml vs. n = 40,
3.0 ± 0.2 ng/ml, P = 0.03). SuPAR levels increased in time. An increase of
suPAR did not predict or precede death, however. SuPAR AUC from ER
to day 5 tended to correlate with injury severity score (r = 0.5, P = 0.07). Plasma cytokines in the ER did not correlate with suPAR measured at
the same time (for example, TNFα: r = 0.2, P = 0.37, IL-10: r = –0.02,
P = 0.91), while cytokine concentrations at the ER did correlate with
suPAR levels at days 3 (TNFα: r = 0.6, P <0.01, IL-10: r = 0.5, P = 0.02) and
5 (TNFα: r = 0.7, P <0.01). Results Mean levels of SHH levels were signifi cantly elevated within
24 hours as much as 45 days post injury in burned animals receiving
the 17β-estradiol (>1,200 pcg/mg) as compared with the placebo-
treated burned animals (<700 pg/mg) and controls (<300 pcg/mg). See
Figure 1. g
Conclusion Early, single-dose estrogen administration following
severe burn injury signifi cantly elevated levels of SHH in brain tissue. This fi nding may represent an extremely novel and important pathway
for both neuroprotection and neuroregeneration in burn patients. Conclusion Plasma concentrations of SuPAR measured at admission
to the ER are associated with overall survival of multi-trauma patients. Furthermore, suPAR concentrations increased during hospital
admission, with most pronounced increases found in patients that
suff ered more serious injury and related to the innate immune response
determined in the ER. These results indicate that suPAR is an innate
immune response-induced late mediator in multi-trauma patients. Feasibility and experience of using exception from informed
consent in a pilot study of immediate estrogen infusion for
hypotensive trauma patients hypotensive trauma patients
JG Wigginton1, PE Pepe1, V Warren1, K AbdelFattah1, JW Gaston1,
J Simpkins2, JP Minei1, D Maass1, AH Idris1
1University of Texas Southwestern Medical Center, Dallas, TX, USA; 2University
of North Texas, Fort Worth, TX, USA
Critical Care 2013, 17(Suppl 2):P288 (doi: 10.1186/cc12226) P288 Feasibility and experience of using exception from informed
consent in a pilot study of immediate estrogen infusion for
hypotensive trauma patients
JG Wigginton1, PE Pepe1, V Warren1, K AbdelFattah1, JW Gaston1,
J Simpkins2, JP Minei1, D Maass1, AH Idris1
1University of Texas Southwestern Medical Center, Dallas, TX, USA; 2University
of North Texas, Fort Worth, TX, USA
Critical Care 2013, 17(Suppl 2):P288 (doi: 10.1186/cc12226) P287 The average number of days to verbal notifi cation of
patients or advocates was 6.55 days (range 0 to 51 days) as the study
team began notifi cation only after the patient or family was able to
reasonably understand information about the study. No one decided
against continued follow-up. Overall, patients and their families were
very enthusiastic about participation and the data safety monitoring
board had no safety concerns after reviewing all study data. Figure 1 (abstract P287). y
y
Conclusion Although delayed notice of participation occurs for many
justifi able reasons, the use of exception from informed consent for
novel, time-sensitive resuscitation studies is not only crucial, but
can be feasible, and well accepted by patients, their advocates and
communities at large. Level of lipopolysaccharide-binding protein and microbial
landscape with account of severity of sepsis syndromes in
polytrauma Level of lipopolysaccharide-binding protein and microbial
landscape with account of severity of sepsis syndromes in
polytrauma β
Methods A total of 168 male rats were assigned randomly to one of
three groups: (1) Sham burn (no actual injury) group (n = 8); (2) Placebo
(no treatment) burn group (n = 80); and (3) E2 (estrogen treatment)
burn group (n = 80). Groups 2 and 3 had 40% TBSA third-degree dorsal
burns, early fl uid resuscitation and, 15 minutes post burn, 0.5 mg/kg
intraperitoneal estrogen (Group 3) or placebo (Group 2). From each
group of 80, eight animals were sequentially sacrifi ced (and cardiac
tissue was sampled for IL-6, TNFα, IL-1β) at one of 10 respective time
points: 0, 0.5, 1, 2, 4, 6, 8, 18, 24 hours and day 7 post burn (at day 7 only
for the eight shams). The markers were measured by ELISA method.i Introduction The aim was estimation of the clinical and predictive
signifi cance of the level of lipopolysaccharide-binding protein (LBP) in
blood serum and examination of microbial landscape with account of
severity of sepsis syndromes in polytrauma. Methods Clinical examination was performed, which included 99
patients with polytrauma according to sepsis syndrome: SIRS (n = 18),
local infection (n = 36), sepsis (n = 27), severe sepsis (n = 12), septic shock
(n = 6) to the criteria of АССР/SССМ. The microorganism identifi cation
was performed using iEMS Reader MF (Labsystems) with the La Chema
multimicrotests. The content of LBP in blood serum was assessed
with IMMULITE ONE (USA) with the reagents DPC (USA). Statistical
analysis of the data was performed with Statistica 6.0. The numerical
characteristics of variables are presented as Me (LQ to UQ). The analysis
of diff erences was carried with the Kruskal–Wallis test for multiple
comparison of independent groups, with Friedman one-way analysis
of variance. The diff erences were statistically signifi cant with P <0.05. g
y
Results In the burned rats, 17β-estradiol signifi cantly decreased the
cardiac levels of TNFα at all time points through 45 days post burn, with
the sham animal levels (30 pg/mg) more comparable with the estradiol
group (70 pg/mg), and signifi cantly less than the placebo animals
(332 pg/mg). Similarly, IL-6 levels in the sham animals (70 pg/mg) were
comparable with the estradiol group (86.5 pg/mg), and signifi cantly
less than the placebo group (730 pg/mg), even at 45 days post burn. Risk factors for the development of complications following blunt
chest trauma: a new risk stratifi cation tool
C Battle1, H Hutchings2, S Lovett2, P Evans2
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK
Critical Care 2013 17(Suppl 2):P292 (doi: 10 1186/cc12230) C Battle , H Hutchings , S Lovett , P Evans
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK ,
Critical Care 2013, 17(Suppl 2):P292 (doi: 10.1186/cc12230) p
p
Results One hundred patients with BAT presented; 71 had complete
data. The accuracy of FAST in BAT was 59.2%; in these, 31 (43.7%) were
confi rmed by CT and 11 (15%) by laparotomy. There were 29 (40.8%)
inaccurate FAST scans, all confi rmed by CT. FAST had a specifi city of
94.7% (95% CI: 0.75 to 0.99) and sensitivity of 46.2% (95% CI: 0.33 to
0.60). A positive predictive value of 0.96 (0.81 to 0.99) and negative
predictive value of 0.39 (0.26 to 0.54). Fisher’s exact test shows positive
FAST is signifi cantly associated with intra-abdominal pathology
(P = 0.001). Cohen’s chance corrected agreement was 0.3. Twenty-
one out of 28 who underwent laparotomies had positive FAST results,
indicating accuracy of 75% (95% CI: 57 to 87%). Critical Care 2013, 17(Suppl 2):P292 (doi: 10.1186/cc12230 Introduction The aim of the study was to investigate the risk factors
for the development of complications following blunt chest trauma
and to develop a risk stratifi cation tool to assist in the management of
this patient group. The diffi culties in the management of this patient
group in the emergency department (ED) due to the development of
late complications are well recognised in the literature [1]. g
Methods Between 2009 and 2011 a total of 276 patients were admitted
to hospital from the ED of a regional trauma centre in Wales, with the
primary diagnosis of blunt chest trauma. Patients with immediate life-
threatening injuries were excluded. Data were collected retrospectively
and included risk factors (age, number of rib fractures, comorbidity,
pre-injury anticoagulant use, smoking status, oxygen saturations and
respiratory rate on initial assessment in the ED), and outcomes (mortality,
any pulmonary morbidity, length of stay of 7 days or more and need
for ICU admission). Development of complications was defi ned as the
occurrence of one or more of the outcomes investigated. Multivariable
logistic regression using fractional polynomials was used to identify Conclusion Patients with false negative scans requiring therapeutic
laparotomy is concerning. In unstable patients, FAST may help in
triaging and identifying those requiring laparotomy. Negative FAST
scans do not exclude abdominal injury. Further randomised control
trials are recommended if the role of FAST is to be better understood. Reference 1. P289 Early administration of a single dose of parenteral estrogen
decreases infl ammation in the heart for 45 days after severe burns
PE Pepe, JG Wigginton, JW Gatson, AH Idris
University of Texas Southwestern Medical Center, Dallas, TX, USA
Critical Care 2013, 17(Suppl 2):P289 (doi: 10.1186/cc12227) Introduction Patients with severe burn injury experience a rapid
elevation in multiple circulating proinfl ammatory cytokines, with the
levels correlating with both injury severity and outcome. In animal S110 Critical Care 2013, Volume 17 Suppl 2
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landscape with account of severity of sepsis syndromes in
polytrauma f
y
gi
Results In 81% of the critically ill patients with polytrauma the post-
traumatic period was accompanied with development of infectious
complications, Gram-negative (K. pneumoniae, Acinetobacter spp.,
E. coli) and Gram-positive (S. Epidermidis, S. aureus). Sepsis was diagnosed
on 8 to 10 days in 45% of the patients. The signifi cant increase of LPS-BP
was found in the fi rst 3 days of the follow-up, compared with the control
values (6.7 times higher in SIRS group (χ2(n = 18, df = 3) = 52.8666,
P <0.001); 9.9 times higher in the group with local infection (χ2(n = 36,
df = 3) = 91.6629, P <0.001); 15.2 times higher in the sepsis group; 20.5
times higher in the severe sepsis group; 47.3 times higher in the septic
shock group (χ2(n = 6, df = 3) = 11.0339, P = 0.0115)), whereas the fi rst
positive results of the microbiological examination were obtained only
on 5 to 7 days. The diagnostic sensitivity of threshold concentration of
LBP in blood serum (335 mkg/ml) was 84%, diagnostic specifi city was
88% (ROC curve: 0.88). Conclusion Following severe burn injury in an animal model, an early
single dose of estrogen can decrease the prolonged let alone the early
onset of cardiac infl ammation. Based on these data, clinical studies
of estrogen infusions should be seriously entertained as estrogen
may not only be an inexpensive, simple adjunctive therapy in burn
management, it may also obviate the need for many subsequent
interventions altogether and even diminish mortality. P290 P290
Accuracy of FAST scan in blunt abdominal trauma in a major
London trauma centre
S Fleming1, R Bird2, K Ratnasingham3, S Sarker3, M Walsh2, B Patel3
1Royal National Orthopaedic Hospital Stanmore, London, UK; 2Royal London
Hospital, London, UK; 3Barts Cancer Institute, London, UK
Critical Care 2013, 17(Suppl 2):P290 (doi: 10.1186/cc12228) Conclusion The high incidence of the diagnostic levels of LBP in blood
serum in patients with sepsis in the early term, before microbiological
confi rmation of infection, allows one to use this parameter as an early
marker of development of purulent septic complications conditioned
by Gram-negative microfl ora. Introduction Blunt abdominal trauma (BAT) is a leading cause of
morbidity and mortality. Rapid diagnosis and treatment with the
Advanced Trauma Life Support guidelines are vital, leading to the
development of focused assessment with sonography in trauma (FAST). Methods A retrospective study carried out from January 2007 to 2008
on all patients who presented with BAT and underwent FAST scan. All
patients subsequently had a CT scan within 2 hours of admission or a
laparotomy within 2 days. The presence of intraperitoneal free fl uid was
interpreted as positive. P291 models, accumulations of these cytokines have been observed in
remote organs, including the heart, brain and lungs. However, data
are lacking regarding the long-term levels of cytokines in the heart
following severe burn injury and also how infusion of parenteral
estrogen, a powerful anti-infl ammatory agent, would aff ect these
levels. Using a rat model, we studied the eff ects of a full-thickness third-
degree burn on cardiac levels of IL-6 and TNFα over 45 days with and
without 17β-estradiol infusion. Level of lipopolysaccharide-binding protein and microbial
landscape with account of severity of sepsis syndromes in
polytrauma
IM Ustyantseva, OI Khokhlova, OV Petukhova, YA Zhevlakova
Federal Scientifi c Clinical Center of Miners’ Health Protection, Leninsk-
Kuznetsky, Russia
Critical Care 2013, 17(Suppl 2):P291 (doi: 10.1186/cc12229) Prediction score of indications for whole body computed
tomography in blunt trauma patients Prediction score of indications for whole body computed
tomography in blunt trauma patients
H Oda, R Sasaki, A Hagiwara, A Kimura
National Center For Global Health and Medicine, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P294 (doi: 10.1186/cc12232) Prediction score of indications for whole body computed
tomography in blunt trauma patients g
p y
p
H Oda, R Sasaki, A Hagiwara, A Kimura
National Center For Global Health and Medicine, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P294 (doi: 10.1186/cc12232) g
p y
p
H Oda, R Sasaki, A Hagiwara, A Kimura
National Center For Global Health and Medicine, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P294 (doi: 10.1186/cc12232) Introduction Whole body computed tomography (WBCT) appears to
be useful for the early detection of clinically occult injury, although its
indications have been controversial. The purpose of this study was to
develop a clinical prediction score to clarify the indications for blunt
trauma patients with multiple injuries (MI) who require WBCT. g g
p
p
Conclusion The results of this study highlight the risk factors for the
development of complications following blunt chest trauma. A risk
stratifi cation tool has also been developed that could assist in the
prediction of poor outcomes in this patient group. The next stage is to
complete a prospective validation study. p
p
j
q
Methods We conducted a retrospective study of 173 patients with
blunt trauma who underwent WBCT at our emergency center between
June 2011 and July 2012. We chose the presence or absence of MI
(Injury Severity Score ≥15) in need of surgical intervention as the
outcome variable. We used bivariate analyses to identify variables
potentially predicting the presentation of MI. The predictor variables
were confi rmed by multivariate logistic regression analyses. We
assigned a score based on the corresponding coeffi cients. Reference g
gfi
Results Among the 173 patients enrolled, 53 were in the MI group. Four
predictors were found to be independently signifi cant by the logistic
analysis: (1) body surface wound ≥3 regions, (2) positive focused
assessment with sonography for trauma, (3) white blood cell count
≥11,000/μl, and (4) D-dimer ≥8 μg/ml. Score 1 was assigned to predictor
(1), score 2 was assigned to predictors (2), (3) and (4). A prediction score
was calculated for each patient by adding these scores. The area under
the receiver operating characteristic curve was 0.89. Prediction score of indications for whole body computed
tomography in blunt trauma patients No patients with a
score of 1 or less had MI (Figures 1 and 2). P293 P293
Guidelines for chest drain insertion do not protect relevant
anatomical structures
J Bowness1, P Kilgour1, S Whiten1, I Parkin1, J Mooney2, P Driscoll1
1St Andrews University, St Andrews, Fife, UK; 2University of Salford, UK
Critical Care 2013, 17(Suppl 2):P293 (doi: 10.1186/cc12231) Guidelines for chest drain insertion do not protect relevant
anatomical structures Conclusion In patients with a score of 1 or 0, the presence of MI is less
likely. These patients may not require WBCT, and selective CT scans of
body parts based on clinical presentation should be considered. Figure 1 (abstract P294). Prediction versus MI probability (validation
group). Figure 2 (abstract P294). Clinical scorings. Multivariate, logistic
regression analyses. Introduction We have reported the risk of chest drain insertion inferior
to the diaphragm when using current international guidelines [1]. Another complication is damage to signifi cant peripheral nerves,
such as the long thoracic nerve causing winging of the scapula [2]. We assessed these risks using: the European Trauma Course method, a
patient’s handbreadth below their axilla just anterior to the midaxillary
line; the British Thoracic Society safe triangle [3]; and the Advanced
Trauma Life Support (ATLS) course guidance [4]. Figure 1 (abstract P294). Prediction versus MI probability (validation
group). Methods We used the above guidelines to place markers (representing
chest drains) in the thoracic wall of 16 cadavers bilaterally (32 sides),
1 cm anterior to the midaxillary line. Subsequent dissection identifi ed
the course and termination of the long thoracic nerve, the site of lateral
cutaneous branches of intercostal nerves, and their relation to the
markers. Results The long thoracic nerve was found in the fi fth intercostal space
in 16 of 32 cases, always in or posterior to the midaxillary line. Contrary
to the description in Grays’ Anatomy (40th edition) it terminated before
the inferior border of serratus anterior. Most commonly it was found
to end by branching in the fourth (right) or fi fth (left) intercostal space
(range third to sixth). Lateral cutaneous branches of intercostal nerves
were found in the fi fth intercostal space in 25 of 32 cases. Contrary to
the description in Last’s Anatomy (12th edition) they always passed
anterior to the midaxillary line (and marker). Figure 1 (abstract P294). Prediction versus MI probability (validation
group). Figure 2 (abstract P294). Clinical scorings. Multivariate, logistic
regression analyses. P294 risk factors and develop a risk stratifi cation tool. The signifi cant risk
factors in the model were selected using backward elimination with
the Akaike Information Criterion (AIC) at a signifi cance level of 0.05. The
c index and the Hosmer–Lemeshow (H–L) test were calculated to assess
discrimination and calibration of the risk stratifi cation tool respectively. Results A total of 161 patients out of the 276 admitted developed
complications following blunt chest trauma. Implementation of
backward elimination using AIC values resulted in a fi nal model based
on the signifi cant risk factors; age, oxygen saturations, number of rib
fractures, presence of chronic lung disease and pre-injury anticoagulant
use (all P <0.05). The c index for the tool was 0.80 and the H–L score was
9.22 (P = 0.32), indicating good predictive capabilities of the tool. Risk factors for the development of complications following blunt
chest trauma: a new risk stratifi cation tool
C Battle1, H Hutchings2, S Lovett2, P Evans2
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK
Critical Care 2013 17(Suppl 2):P292 (doi: 10 1186/cc12230) Smith J: Focused assessment with sonography in trauma (FAST): should its
role be reconsidered? Postgrad Med J 2010, 86:285-291. S111 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 1.
Ahmad MA, et al.: Assessment of severity of chest trauma. Injury 2010,
41:981-983. P296 P296
Severe hyperkalemia is prevalent and can be detected by
quantitative electrocardiographic fi ndings and medication history
of calcium channel blocker among the patients with symptomatic
or extreme bradycardia
SB Chon
Kangwon National University Hospital, Chuncheon, South Korea
Critical Care 2013, 17(Suppl 2):P296 (doi: 10.1186/cc12234) P296
Severe hyperkalemia is prevalent and can be detected by
quantitative electrocardiographic fi ndings and medication history
of calcium channel blocker among the patients with symptomatic
or extreme bradycardia
SB Ch Severe hyperkalemia is prevalent and can be detected by
quantitative electrocardiographic fi ndings and medication history
of calcium channel blocker among the patients with symptomatic
or extreme bradycardia
h Conclusion Despite the controversies over the role of fl uids in the
prehospital environment, the carriage and use of blood is both feasible
and safe in a physician-led helicopter emergency medical service. Reference Kangwon National University Hospital, Chuncheon, South Korea
Critical Care 2013, 17(Suppl 2):P296 (doi: 10.1186/cc12234) Introduction Severe hyperkalemia (serum potassium ≥6.0 mmol/l with
electrocardiography (ECG) change) should be detected and treated
immediately [1,2]. Among symptomatic or extreme bradycardia
patients, we sought its prevalence and prediction rule by history, vital
sign, and ECG [3,4]. 1. Davenport R, et al.: J Trauma 2011, 70:90-95. Table 1 (abstract P295). Demographics of patients receiving a prehospital
blood transfusion Table 1 (abstract P295). P296 Demographics of patients receiving a prehospital
blood transfusion
Total (n = 147)
Male (%)
102 (69.3)
Age (years), median (IQR)
34.5 (22 to 52)
Mechanisms of injury (%)
Motor vehicle collision
87 (59.1)
Motor bike collision
20 (13.6)
Pedestrian versus car
9 (6.1)
Gunshot wound/stabbing
9 (6.1)
Fall from a height
5 (3.4)
Recreational
6 (4.1)
Other
11 (7.5)
Number of patients trapped on arrival (%)
45 (30.6)
Scene time (minutes), mean (SD)
49.9 (27.8)
Time from tasking to arrival at hospital (minutes), mean (SD)
126.5 (51.3)
Heart rate, median (IQR)
115 (90 to 130)
Systolic blood pressure (mmHg), median (IQR)
80 (65 to 105)
RTSo
2, median (IQR)
5.967 (4.083 to 6.904)
Total number of PRBC units transfused
382
Total number of PRBC units wasted
66
Volume of crystalloid (ml), median (IQR)
500 (0 to 1,500)
Pronounced life extinct on scene
22 (15.0) Methods A retrospective cross-sectional study was performed on
patients with symptomatic (heart rate (HR) ≤50/minute with dyspnea,
chest pain, altered mentality, dizziness/syncope/presyncope, general
weakness, oliguria, or shock) or extreme (HR ≤40/minute) bradycardia
at an ED of an academic hospital from June 2008 to March 2012. Risk factors of severe hyperkalemia were chosen by multiple logistic
regression analysis among history (gender, age, comorbidity, and
current medication), vital sign, and ECG (maximal precordial T wave
amplitude, PR and QRS intervals). Scoring index was derived by
summing up of simplifi ed regression coeffi cients of independent risk
factors. Results A total of 169 cases were enrolled. Mean age was 71.2 years (SD,
12.5 years). Females numbered 87 (51.5%). Thirty-six cases (21.3%) had
severe hyperkalemia. Four variables were independent risk factors of
severe hyperkalemia (simplifi ed scores in parentheses): medication of
calcium channel blocker (CCB: 2); maximal precordial T ≥8.5 mV (2); PR
(atrial fi brillation or junctional bradycardia: 1); and HR ≤40/minute (1). (Nagelkerke R2 = 0.503, AUC = 0.849 (95% CI 0.786 to 0.899).) Sensitivity
and specifi city reached 0.75 and 0.83 when total score was ≥3. For score
≥4, positive likelihood ratio reached 5.54 (sensitivity 0.50, specifi city
0.91). Conclusion Severe hyperkalemia is prevalent among symptomatic or
extreme bradycardia patients and could be detected immediately by
a scoring index composed of quantitative ECG parameters and history
of medication of CCB. Prediction score of indications for whole body computed
tomography in blunt trauma patients Conclusion Placement 1 cm anterior to the midaxillary line minimises
risk to the long thoracic nerve and lateral cutaneous branches of
intercostal nerves. We therefore conclude that not all areas of the British
Thoracic Society safe triangle are indeed safe, and anteroposterior
placement should follow the European Trauma Course and ATLS
guidelines: just anterior to the midaxillary line (for example, 1 cm). References 1. Bowness J, et al.: It’s higher than you think: chest drains and the 5th ICS. Clin Anatomy, in press. 2. Kesienne EB, et al.: Tube thoracostomy: complications and its management. Pulm Med 2012:256878. 3. Laws D, et al.: BTS guidelines for the insertion of a chest drain. Thorax 2003,
58:ii53-ii59. 4. American College of Surgeons Committee on Trauma: Advanced Trauma Life
Support for Doctors, Course Manual. Chicago: American College of Surgeons;
2008:108. Figure 2 (abstract P294). Clinical scorings. Multivariate, logistic
regression analyses. Figure 2 (abstract P294). Clinical scorings. Multivariate, logistic
regression analyses. S112 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Table 2 (abstract P295). Interventions performed P295 Prehospital blood transfusion: 5-year experience of an Australian
helicopter emergency medical service
PB Sherren, B Burns
Greater Sydney Area HEMS, Sydney, Australia
Critical Care 2013, 17(Suppl 2):P295 (doi: 10.1186/cc12233) Introduction There is an emerging body of evidence suggesting that
early packed red blood cell transfusion accompanied by fresh frozen
plasma, while limiting crystalloids, confers a survival benefi t in major
trauma [1]. Prehospital blood transfusion has been infrequently
described, and concerns over expense, transfusion reactions, risk of
disease transmission, short shelf half-life and diffi cult storage have
limited the interest of prehospital providers. p
p
p
Methods All Greater Sydney Area HEMS (GSA-HEMS) prehospital
missions involving a blood transfusion over a 66-month period were
identifi ed and reviewed. The prospectively completed GSA-HEMS
electronic database was utilised to identify patients and extract data.i Results We identifi ed 158 missions involving a prehospital blood
transfusion, of which 147 patient datasets were complete. The majority
of patients had a blunt mechanism of injury (93.9%) and were male
(69.3%) with a median (IQR) age of 34.5 (22 to 52) years (Table 1). The
majority of patients were haemodynamically unstable, with a median
(IQR) heart rate and systolic blood pressure of 115 (90 to 130) and 80
(65 to 105) mmHg, respectively. Twenty-two patients (15.0%) were
pronounced life extinct on the scene. A total of 382 units of packed
red blood cells were transfused, with a median of 3 units (range 1 to
6). No early transfusion reactions were noted. A variety of prehospital
interventions accompanied the transfusions, ranging from rapid
sequence intubation through to thoracotomies (Table 2).l References References
1. Huber-Wgner S, et al.: Lancet 2009, 373:1455-1461. 2. Kimura A, et al.: Acad Emerg Med 2012, 19:734-735. References
1. Huber-Wgner S, et al.: Lancet 2009, 373:1455-1461. 2. Kimura A, et al.: Acad Emerg Med 2012, 19:734-735. Table 2 (abstract P295). Interventions performed
Intervention
Total (n = 147)
Rapid sequence intubation
96 (65.3)
Cold endotracheal intubation
15 (10.2)
Surgical airway
1 (0.7)
Thoracostomy (open or tube)
59 (40.1)
Thoracotomy
3 (2.0)
Pelvic binder or fracture splintage
89 (60.5)
Intraosseous insertion sites
22 (15.0)
Humerus
19
Tibia
10
Femur
1
Tourniquet application
15 (10.2)
Hypertonic saline administration
16 (10.9)
E-FAST performed
27 (18.4)
Negative results
9
Positive results (fi ndings)
18
Abdominal free fl uid
14
Pneumothorax
2
Haemothorax
1
Thoracic ultrasound only; all negative
2 (1.3)
Cardiac ultrasound only; no cardiac motion
1 (0.7) Focused cardiac ultrasound in resuscitation E Fish, L Fuchs, G Gulati, D Talmor, A Oren-Grinberg E Fish, L Fuchs, G Gulati, D Talmor, A Oren Grinberg
Beth Israel Deaconess Medical Center, Boston, MA, USA g
Beth Israel Deaconess Medical Center, Boston, MA, USA Results Multivariate logistic regression analysis indicated the following
three prehospital variables as signifi cant high-ranking factors for
predicting favorable 1-month outcomes: shockable initial rhythm
(odds ratio (OR), 5.87; 95% CI, 5.23 to 6.60), witnessed arrest (OR, 3.05;
95% CI, 2.83 to 3.28), and age (≥18 to <71 years; OR, 3.24; 95% CI, 2.97
to 3.53). Using recursive partitioning analysis for development cohort
data (2005 to 2008, n = 307,896), we stratifi ed prehospital risk: if OHCA
with shockable initial rhythm was witnessed, the probability of CPC 1 to
2 was 20.0% (age, ≥18 to <71 years; grade 1) or 10.3% (age, ≥71 years;
grade 2); if OHCA with shockable initial rhythm was unwitnessed, the
probability was 6.8% (age, <81 years; grade 3) or 1.8% (age, ≥81 years;
grade 4a); if OHCA with unshakable initial rhythm was witnessed, the
probability was 1.4% (grade 4b); and if OHCA with unshakable initial
rhythm was unwitnessed, the probability was 0.3% (grade 5). The
c-statistics for risk stratifi cation of the development and validation
cohorts (2009 external data, n = 82,330) were 0.853 (95% CI, 0.846 to
0.859) and 0.875 (95% CI, 0.865 to 0.885), respectively. The odds ratios
for CPC 1 to 2 at 1 month between patients at very high and very low
were 79.5 (95% CI, 72.5 to 87.4) and 122.1 (95% CI, 102.0 to 147.3),
respectively, in these cohorts. ,
,
,
Critical Care 2013, 17(Suppl 2):P299 (doi: 10.1186/cc12237) Introduction Focused cardiac ultrasound is recognized as a vital tool
in critical care medicine. Few studies, however, have examined the
utility of this modality in resuscitation. While integration of ultrasound
can supplement resuscitation by potentially establishing the etiology
of cardiac arrest, it must be done safely to avoid interruption of
compressions. The aim of this study was to examine the integration of
focused cardiac ultrasound into resuscitations in our hospital. Methods We performed a retrospective observational study of patients
undergoing resuscitation at Beth Israel Deaconess Medical Center,
Boston, USA, from 2009 to 2012. Inclusion criteria were age >17 years Figure 1 (abstract P299). Ultrasound fi ndings. References . Nyirenda MJ, et al.: Hyperkalaemia. BMJ 2009, 339:b4114. . Alfonzo AV, et al.: Potassium disorders – clinical spectrum and emergency Alfonzo AV, et al.: Potassium disorders – clinical spectrum and emergency S113 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 management. Resuscitation 2006, 70:10-25. management. Resuscitation 2006, 70:10-25. Methods We performed a prospective study between January 2011
and December 2012 at a university cancer reference hospital in
Brazil. Consecutive patients undergoing cardiac arrest and receiving
CPR were identifi ed, and a database was used to register baseline
characteristics of patients, data for CPR and 30-day survival. During the
fi rst 12 months of the study (phase 1), patients received CPR according
to the institution’s protocol guided by diff erent physicians. During
the last 11 months of the study (phase 2), a multidisciplinary Medical
Emergency Team System consisting of a specialized team of ICU nurses,
physiotherapists and physicians using an electronic calling system was
responsible for the CPR. management. Resuscitation 2006, 70:10 25. 3. Neumar RW, et al.: Part 8: adult advanced cardiovascular life support: 2010
American Heart Association Guidelines for Cardiopulmonary
Resuscitation and Emergency Cardiovascular Care. Circulation 2010,
122:S729-S767. 4. Severi S, et al.: Heart rate response to hemodialysis-induced changes in
potassium and calcium levels. J Nephrol 2001, 14:488-496. Focused cardiac ultrasound in resuscitation p
y
Conclusion Prehospital risk stratifi cation (grade 1 to 5) of patients
after OHCA using three prehospital factors (shockable initial rhythm,
witnessed arrest, and age) accurately classifi es the severity of OHCA
and may help clinicians guide in-hospital strategies. Reference 1. Rerisfeld GM, et al.: Survival in cancer patients undergoing in-hospital
cardiopulmonary resuscitation: a meta-analysis. Resuscitation 2006,
71:152-160. i
p
Methods We analyzed data for 390,226 adult patients (age ≥18 years)
after nontraumatic OHCA, from a prospectively recorded nationwide
Utstein-style Japanese database for 2005 to 2009. The endpoint was
1-month survival with favorable neurological outcome (Cerebral
Performance Category (CPC) 1 to 2). Prehospital risk stratifi cation following out-of-hospital cardiac
arrest Prehospital risk stratifi cation following out-of-hospital cardiac
arrest
Y Goto1, T Maeda1, Y Goto2
1Kanazawa University Hospital, Kanazawa, Japan; 2Yawata Medical Center,
Komatsu, Japan
Critical Care 2013, 17(Suppl 2):P297 (doi: 10.1186/cc12235) p
Results A total of 182 patients experienced in-hospital cardiac arrest
and received CPR. Most patients were in hospital due to medical
admission (82%), had solid tumors (85%), and had localized disease
(60%). Pulseless electric activity was the predominant arrest rhythm
(54%) and 45% of patients were resuscitated on the ICU. Overall 30-day
survival was 9.3%. In the phase 1 of the study, 30-day survival was 3.7%,
and in phase 2, survival was 14% (P = 0.017). There were no diff erences
in the two phases of the study regarding baseline characteristics of
patients, rhythm and place of CPR. Introduction Multivariate analyses have identifi ed factors that have
enabled the development of sophisticated equations and scoring
models with the ability to predict outcomes following out-of-hospital
cardiac arrest (OHCA). However, implementation of such outcome
prediction models in research and clinical practice has been slow. The more crucial aspect of these predictions is the lack of prehospital
risk stratifi cation for OHCA patients. Prehospital risk stratifi cation for
patients after OHCA may help clinicians guide in-hospital strategies,
particularly in the emergency department. The purpose of this study
was to develop a simple and generally applicable prehospital risk-
stratifi cation scheme for patients after OHCA. Conclusion Overall survival from CPR in cancer patients compares
favorably with survival rates in noncancer patients. A multidisciplinary
Medical Emergency Team System increases survival in cancer patients
undergoing cardiopulmonary resuscitation. P298 P298
Multidisciplinary Medical Emergency Team System increases
survival in cancer patients undergoing cardiopulmonary
resuscitation
L Hajjar1, F Galas2, S Vieira2, J Almeida2, E Osawa2, C Park2, J Fukushima2,
E Angelo2, A Pinheiro2, J Auler Jr2
1Heart Institute, São Paulo, Brazil; 2Cancer Institute of University of São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P298 (doi: 10.1186/cc12236) Prehospital epinephrine administration and survival among
patients with unshockable initial rhythm after out-of-hospital
cardiac arrest g
Methods Our patient, with a medical history of a laparoscopic repair
of a symptomatic diaphragmatic hernia 6 months prior, presented
herself at the emergency department with pain in the upper abdomen
and nausea. The physical examination, laboratory tests and X-ray
of the thorax were normal and she was sent home. Twenty-four
hours later paramedics were summoned to our patient because of
increased complaints. On arrival of the paramedics she had a normal
electrocardiogram (ECG) and during the transfer from her bed to the
stretcher she collapsed due to pulseless electric activity (PEA), for which
cardiopulmonary resuscitation was started. Sinus rhythm and output
was regained after several minutes and the patient was transported to
the hospital. At arrival in the hospital, the X-ray of the thorax showed
an intrathoracic stomach and a signifi cant mediastinal shift to the right. Results After emergency laparotomy, which concerned correcting the
gastric herniation and resection of an ischemic part of stomach, the
patient remained hemodynamically stable. Cardiac ischemia was ruled
out based on ECG, laboratory fi ndings, cardiac ultrasound and cardiac
computed tomography. The ultrasound in the emergency department
did show a distended right ventricle and normal left function, which
disappeared later (after repositioning the stomach), which is evidence
for the mediastinal shift as a cause for the PEA.i Y Goto1, T Maeda1, Y Goto2 Y Goto1, T Maeda1, Y Goto2 Y Goto1, T Maeda1, Y Goto2 1Kanazawa University Hospital, Kanazawa, Japan; 2Yawata Medical Center,
Komatsu, Japan 1Kanazawa University Hospital, Kanazawa, Japan; 2Yawata Medical Center,
Komatsu, Japan Critical Care 2013, 17(Suppl 2):P302 (doi: 10.1186/cc12240) Critical Care 2013, 17(Suppl 2):P302 (doi: 10.1186/cc12240) Introduction Epinephrine has been a cornerstone of cardiac
resuscitation and advanced cardiac life support since the 1960s. However, there is little evidence from clinical trials that epinephrine
administration after out-of-hospital cardiac arrest (OHCA) improves
long-term survival. There would be subsets of patients for whom
epinephrine administration is in fact benefi cial. Our objective was to
determine whether prehospital epinephrine administration would
improve survival at 1 month in OHCA patients with unshockable initial
rhythm. y
Methods We analyzed data for 383,045 adult OHCA patients with
unshockable initial rhythm, from a prospectively recorded nationwide
Utstein-style Japanese database for 2007 to 2010. We divided these
patients into two cohorts: prehospital epinephrine administration
cohort (n = 30,237) and non-epinephrine administration cohort
(n = 352,808). Multidisciplinary Medical Emergency Team System increases
survival in cancer patients undergoing cardiopulmonary
resuscitation Methods We designed a before-and-after study based on a 3-month
retrospective assessment of the adult victims of OOH-CA in 2009, before
the implementation of the ALERT protocol in the Liege dispatching
centre, and the prospective evaluation of the same 3-month period in
2011, immediately after the implementation of this protocol. Data were
extracted from ambulance, paramedical and medical intervention
teams fi les, as well as the audio recordings of the dispatching centre. Results There were 233 OOH-CAs detected in the fi rst period and 235
in the second. Victims were predominantly male (59%, both periods),
aged 66 and 64 years, respectively. Callers were family members in 52%
in 2009 and 64% in 2011. In 2009, only 9.9% victims benefi ted from
bystander CPR, while there were 22.5% in 2011 (P < 0.0002). Reasons for
protocol underuse were: assistance not off ered (42.3%), caller remote
from the victim (20.6%) or emotionally distressed (15.5%). Mean no-
fl ow time decreased from 253 seconds in 2009 to 168 seconds in 2011
(NS). Ten victims were admitted in ROSC to hospital in 2009 and 13 in
2011 (P = 0.09). p
p
g
p
g
Methods We designed a before-and-after study based on a 3-month
retrospective assessment of the adult victims of OOH-CA in 2009, before
the implementation of the ALERT protocol in the Liege dispatching
centre, and the prospective evaluation of the same 3-month period in
2011, immediately after the implementation of this protocol. Data were
extracted from ambulance, paramedical and medical intervention
teams fi les, as well as the audio recordings of the dispatching centre.i i
g
p
g
Results There were 233 OOH-CAs detected in the fi rst period and 235
in the second. Victims were predominantly male (59%, both periods),
aged 66 and 64 years, respectively. Callers were family members in 52%
in 2009 and 64% in 2011. In 2009, only 9.9% victims benefi ted from
bystander CPR, while there were 22.5% in 2011 (P < 0.0002). Reasons for
protocol underuse were: assistance not off ered (42.3%), caller remote
from the victim (20.6%) or emotionally distressed (15.5%). Mean no-
fl ow time decreased from 253 seconds in 2009 to 168 seconds in 2011
(NS). Ten victims were admitted in ROSC to hospital in 2009 and 13 in
2011 (P = 0.09). Multidisciplinary Medical Emergency Team System increases
survival in cancer patients undergoing cardiopulmonary
resuscitation Conclusion Focused cardiac ultrasound is a feasible adjunct to ALS
resuscitation and may assist in the early identifi cation of reversible
causes of cardiac arrest. Care must be taken to ensure no interruptions
to cardiac compressions are made by performance during pulse checks. Further studies are needed to examine the outcomes associated with
its integration into resuscitations. Further studies are needed to examine the outcomes associated with
its integration into resuscitations. Conclusion Using the ALERT protocol in the Liege dispatching centre
signifi cantly improved the numbers of patients in whom bystander CPR
was attempted. Dispatchers must embrace this new opportunity to
help callers and be encouraged to accept the responsibility of initiating
such assistance. Reference Acute intrathoracic gastric herniation as a rare cause of cardiac arrest
DW Hoelen, AL Van Duijn, CL Meuwese, JP Ruurda, MA Sikma
UMC Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P300 (doi: 10.1186/cc12238) Multidisciplinary Medical Emergency Team System increases
survival in cancer patients undergoing cardiopulmonary
resuscitation L Hajjar1, F Galas2, S Vieira2, J Almeida2, E Osawa2, C Park2, J Fukushima2,
E Angelo2, A Pinheiro2, J Auler Jr2
1Heart Institute, São Paulo, Brazil; 2Cancer Institute of University of São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P298 (doi: 10.1186/cc12236) Introduction In the last years, ICU admission has been high in
cancer patients due to increased survival related to advances in
treatment. Patients with cancer reportedly have poor outcomes from
cardiopulmonary resuscitation (CPR). The goal of this study was to
evaluate the eff ectiveness of a multidisciplinary Medical Emergency
Team System in the CPR outcomes in cancer patients. Figure 1 (abstract P299). Ultrasound fi ndings. S114 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Introduction Early bystander cardiopulmonary resuscitation (CPR)
is a key factor in improving survival from out-of-hospital cardiac
arrest (OOH-CA). The ALERT algorithm, a simple and eff ective
compression-only telephone CPR protocol, has the potential to help
bystanders initiate CPR. This study evaluates the eff ectiveness of the
implementation of this protocol in the Liege dispatching centre. and performance of focused cardiac ultrasound during resuscitation. Recorded variables included admission data, code demographics and
diagnosis, therapy outside standard ALS protocol, time to return of
spontaneous circulation (ROSC), and outcomes data. p
Results Of 33 eligible patients, 12 were excluded due to incomplete cardiac
ultrasound reports; 21 patients were enrolled. Cohort demographics
included: 57.1% male, average age 64.3 years, average BMI 28.5 kg/
m2, average Charlson score 5.5. Resuscitations took place on the wards
(52.4%), ICU (42.9%), or operating room (4.8%). Most patients had an initial
unshockable rhythm (71.5%). The most common ultrasound fi nding was
cardiac standstill (47.6%) (Figure 1). The most common intervention as a
result of the ultrasound was initiation of a pressor infusion (33.3%), of which
71.4% were ionotropes. Additional therapies included blood transfusion
(4.8%), heparin (9.5%), tPA (4.8%), cardiac catheterization (4.8%), and
surgery (9.5%). ROSC was achieved in 37.5% of patients; average time to
ROSC was 13 minutes. A total 33.3% of patients who underwent ALS were
alive at hospital discharge and 28.6% at 1 year. p e
e tat o o t s p otoco
t e
ege d spatc
g ce t e. References 1. Deakin CD, et al.: Resuscitation 2010, 81:1305-1352. 1. Deakin CD, et al.: Resuscitation 2010, 81:1305-1352 2. Robson R. Resuscitation 2010, 81:1453-1454. 3. Breitkreutz R. et al.: Crit Care Med 2007, 35:S150-S161. Acute intrathoracic gastric herniation as a rare cause of cardiac arrest
DW Hoelen, AL Van Duijn, CL Meuwese, JP Ruurda, MA Sikma
UMC Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P300 (doi: 10.1186/cc12238) 1. Ghuysen A, Collas D, Stipulante S, Donneau AF, Hartstein G, Hosmans T,
Vantroyen B, D’Orio V: Dispatcher-assisted telephone cardiopulmonary
resuscitation using a French-language compression-only protocol in
volunteers with or without prior life support training: a randomized trial. Resuscitation 2011, 82:57-63. Introduction In this case report, we describe a patient who presented
with a cardiac arrest as a result of an obstructive shock, which
progressed into cardiac arrest, caused by an acute para-esophageal
gastric herniation. P302 Prehospital epinephrine administration and survival among
patients with unshockable initial rhythm after out-of-hospital
cardiac arrest The endpoints were 1-month survival after OHCA,
prehospital return of spontaneous circulations (ROSCs), and 1-month
survival with favorable neurological outcome (Cerebral Performance
Category (CPC) scale, categories 1 to 2) at 1 month. Conclusion We are the fi rst to describe a patient requiring
cardiopulmonary resuscitation for progressive obstructive shock, due
to an intrathoracic stomach. Especially after a laparoscopic repair of a
diaphragmatic hernia, this is a rare cause for shock and cardiac arrest,
which requires a diff erent medical approach. y
Results The rate of 1-month survival was 3.72% for the epinephrine
administration cohort and 2.49% for the non-epinephrine adminis-
tration cohort, 17.9% versus 3.0% for prehospital ROSC, and 0.57%
versus 0.77% for CPC 1 to 2 (all P <0.0001). Positive associations were
observed between epinephrine administration and 1-month survival
(adjusted odds ratio (aOR), 1.18; 95% CI, 1.11 to 1.27), and prehospital
ROSC (aOR, 5.50; 95% CI, 5.29 to 5.72; all P <0.0001). Negative association
was observed between epinephrine administration and CPC 1 to 2
(aOR, 0.56; 95% CI, 0.48 to 0.66; P <0.0001). Multivariate logistic analysis
revealed that age (<66 years; aOR, 4.31; 95% CI, 2.47 to 8.01), total dose Impact of termination of resuscitation for out-of-hospital
cardiopulmonary arrest in Japan Results A total of 430 cardiopulmonary arrest events occurred in-
hospital between January 2010 and December 2011. A total of 326
patients were excluded because 252 occurred in the ICU, 70 in the
coronary care unit and four in the stork unit. In total, 104 patients
were enrolled; 50.9% where female and 49.1% male. Median age was
61 years. A total 41.3% of the arrests were due to respiratory arrest
and 58.7% due to cardiac arrest. Out of 104 patients who developed
cardiopulmonary arrest, 47 (45.19%) occurred during regular working
hours, and 57 (54.81%) occurred after regular working hours (P = 0.0081). The event survival was 87.23% during regular working hours compared
with 47.37% for the patients who developed cardiopulmonary arrest
after regular working hours (P <0.0001). In total, 53.19% of those who
developed cardiopulmonary arrest during regular working hours were
discharged alive from the ICU compared with 26.32% of those who
developed cardiopulmonary arrest after regular working hours.i Introduction What can one anticipate from the introduction of
termination of resuscitation (TOR) for patients suff ering out-of-hospital
cardiopulmonary arrest (OHCA) in Japan? Irrespective of whether
patients have made a living will requesting that medics do not attempt
resuscitation, eff orts are made to resuscitate over 90% of OHCA patients
in Japan [1,2]; the number of people resuscitated exceed 120,000
every year. The 2010 American Heart Association (AHA) Guidelines for
cardiopulmonary resuscitation (CPR) and emergency cardiovascular
care (ECC) defi ned the criteria for TOR; this initiative may help reduce
the number of unnecessary hospital transports by 40 to 60% and hold
down medical costs [3]. p
p
y
g
g
Conclusion Event survival and survival to discharge were signifi cantly
higher in patients who developed cardiopulmonary arrest during
regular working hours. Methods This was a single-center retrospective cohort study of
patients who suff ered OHCA and were transported to our hospital
between April 2009 and March 2011. We investigated the patients’
characteristics, whether they met the TOR criteria, and their outcome
at the time of hospital discharge. g
Reference 1. Peberdy MA,Ornato JP, Larkin L, et al.: Survival from in-hospital cardiac arrest
during nights and weekends. JAMA 2008, 299:785-792. P305 Results A total of 195 patients (mean age, 69 years), 67% of whom
were male, were transported to our hospital after suff ering OHCA. Cardiopulmonary arrest was witnessed in 52 cases (27%). P301 Implementation of dispatcher-assisted cardiopulmonary
instructions using the ALERT protocol: preliminary results in
Belgium
A Ghuysen1, S Stipulante1, M El FAssi1, A Donneau2, V D’Orio1, R Tubes1
1CHU – Ulg Liège, Belgium; 2Liège University, Liège, Belgium
Critical Care 2013, 17(Suppl 2):P301 (doi: 10.1186/cc12239) A Ghuysen1, S Stipulante1, M El FAssi1, A Donneau2, V D’Orio1, R Tubes1
1CHU – Ulg Liège, Belgium; 2Liège University, Liège, Belgium
Critical Care 2013, 17(Suppl 2):P301 (doi: 10.1186/cc12239) S115 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 of epinephrine (1 mg; aOR, 3.65; 95% CI, 2.61 to 5.18), call-response
time (<5 minutes; aOR, 3.58; 95% CI, 1.98 to 6.69), witnessed arrest (aOR,
2.17; 95% CI, 1.51 to 3.16), and pulseless electrical activity as an initial
rhythm (aOR, 2.02; 95% CI, 1.46 to 2.80) were signifi cantly associated
with CPC 1 to 2 at 1 month in the epinephrine administration cohort. of epinephrine (1 mg; aOR, 3.65; 95% CI, 2.61 to 5.18), call-response
time (<5 minutes; aOR, 3.58; 95% CI, 1.98 to 6.69), witnessed arrest (aOR,
2.17; 95% CI, 1.51 to 3.16), and pulseless electrical activity as an initial
rhythm (aOR, 2.02; 95% CI, 1.46 to 2.80) were signifi cantly associated
with CPC 1 to 2 at 1 month in the epinephrine administration cohort. detection and resuscitation of cardiopulmonary arrest are crucial for
better clinical outcome. We conducted our study to evaluate event
survival of in-hospital cardiopulmonary arrest after regular working
hours in nonmonitored areas of a tertiary-care center. of epinephrine (1 mg; aOR, 3.65; 95% CI, 2.61 to 5.18), call-response
time (<5 minutes; aOR, 3.58; 95% CI, 1.98 to 6.69), witnessed arrest (aOR,
2.17; 95% CI, 1.51 to 3.16), and pulseless electrical activity as an initial
rhythm (aOR, 2.02; 95% CI, 1.46 to 2.80) were signifi cantly associated
with CPC 1 to 2 at 1 month in the epinephrine administration cohort. Conclusion In OHCA patients with unshockable initial rhythm,
prehospital epinephrine administration signifi cantly increased the rate
of survival at 1 month after cardiac arrest. The best single predictor for
favorable neurological outcomes at 1 month following prehospital
epinephrine administration after cardiac arrest was age (<66 years)
followed by total dose of epinephrine (1 mg) and then by call-response
time (<5 minutes). p
p
References 1. Berdowski J, et al.: Resuscitation 2010, 81:1479-1487. 2. Fukuda T: J Jpn Assoc Acute Med 2010, 23:101-108. 3. Morrison LJ, et al.: N Engl J Med 2006, 355:478-487. 4. Fukuda T: J Jpn Soc Intensive Care Med 2012, 19:275. Results TH was initiated in 93% (64/69) of OOHCA patients and 87%
(40/46) with ischaemic cardiac aetiology underwent PCI. ICU survival
was 58% (40/69); 65% (26/40) scoring CPC 1 to 2 and 35% (14/40) CPC 3
to 4 at ICU discharge. Two patients with CPC 2 improved to CPC 1 during
their hospital stay. All patients with CPC 1 to 2 survived to hospital
discharge; two required general rehabilitation before returning home. Only 43% (6/14) of CPC 3 to 4 patients survived to hospital discharge;
none returned home. Two went into hospice care, one was repatriated
to another hospital and three went to neuro-rehab. No CPC 3 to 4
patients improved CPC scores after ICU discharge. Overall hospital
survival was 46%. See Table 1. Cognitive performance and cap
out-of-hospital cardiac arrestl Introduction Before the introduction of primary percutaneous coronary
intervention (PCI) and therapeutic hypothermia (TH) to out-of-hospital
cardiac arrest (OOHCA) management, survival to hospital discharge
with intact neurological function was poor [1,2]. We aimed to quantify
the survival and degree of neurological impairment in OOHCA patients
admitted to our ICU since the adoption of post-OOHCA bundles. Conclusion In our study, 65% of the patients who were transported to
the hospital after OHCA met the criteria for TOR. Outcomes for patients
who met the TOR criteria were signifi cantly worse than those who did
not meet the criteria (2.4% vs. 14.5%, P <0.005). In Japan, eff orts are
made to resuscitate almost all individuals who suff er OHCA, but 75%
of those patients die within a day. In light of the fact that even the
medical cost for each of these patients who die within a day amounts
to US$1,500 [4], the introduction of TOR will have a particularly strong
impact in Japan. Methods Sixty-nine consecutive OOHCA patients admitted to the ICU
at Hammersmith Hospital from 1 January 2011 to 30 June 2012 were
identifi ed and reviewed from hospital databases. Cognitive status was
scored using Cerebral Performance Category (CPC); 1 to 2 normal–mild
and 3 to 4 moderate–severe neurological impairment. Scores were
determined from ICU summaries, occupational and physiotherapy
reports. Hospital discharge outcomes were determined from hospital
databases. Impact of termination of resuscitation for out-of-hospital
cardiopulmonary arrest in Japan The 2010
AHA Guidelines for CPR and ECC regarding the criteria for TOR were
applied in 126 cases (65%), of whom 113 (90%) were dead on arrival,
and 13 were successfully resuscitated and admitted. The outcomes for
these 13 patients were as follows: 10 died in the hospital, two patients
were discharged with a Glasgow Pittsburgh Cerebral Performance
Category (CPC) score of 1, and one patient was transferred to another
hospital with a CPC score of 3. P301 Methods A retrospective chart review of all adult patients who
developed in-hospital cardiopulmonary arrest between January
2010 and December 2011. Working hours are defi ned as 07:00 to
17:00 Saturday to Wednesday. Event survival is defi ned as return
of spontaneous circulation (ROSC) for more than 20 minutes. Adult patients 18 years and above who suff ered from in-hospital
cardiopulmonary arrest were included. Patients were excluded if
they had cardiopulmonary arrest in the emergency department, had
implantable cardiovascular device, arrested in monitored areas or had
pre-existing DNR orders. Data analysis was accomplished using SAS,
version 9.3 (SAS institute, Inc., Cary, NC, USA). Conclusion In OHCA patients with unshockable initial rhythm,
prehospital epinephrine administration signifi cantly increased the rate
of survival at 1 month after cardiac arrest. The best single predictor for
favorable neurological outcomes at 1 month following prehospital
epinephrine administration after cardiac arrest was age (<66 years)
followed by total dose of epinephrine (1 mg) and then by call-response
time (<5 minutes). P303 Impact of termination of resuscitation for out-of-hospital
cardiopulmonary arrest in Japan
T Fukuda, N Ohashi, M Gunshin, T Matsubara, S Nakajima, Y Kitsuta,
N Yahagi
Graduate School of Medicine, The University of Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P303 (doi: 10.1186/cc12241) References 1. Stub et al.: Am J Cardiol 2011, 107:522-527. 2. Girotra et al.: N Engl J Med 2012, 367:1912-1920. 3. Herlitz et al.: Heart 2003, 89:25-30. 4. Cronier et al.: Crit Care 2011, 15:122. 1. Stub et al.: Am J Cardiol 2011, 107:522-527. 2. Girotra et al.: N Engl J Med 2012, 367:1912-1920. 3. Herlitz et al.: Heart 2003, 89:25-30. 4. Cronier et al.: Crit Care 2011, 15:122. 2.
MacLeod et al.: J Cardiothorac Vasc Anesth 2012, 26:1007-1014. patients: does it mean something? p
g
C Genbrugge, I Meex, S Scheyltjens, J Dens, C De Deyne
ZOL, Genk, Belgium
Critical Care 2013, 17(Suppl 2):P306 (doi: 10.1186/cc12244) p
g
C Genbrugge, I Meex, S Scheyltjens, J Dens, C De Deyne
ZOL, Genk, Belgium
Critical Care 2013, 17(Suppl 2):P306 (doi: 10.1186/cc12244) p
g
C Genbrugge, I Meex, S Scheyltjens, J Dens, C De Deyne Introduction During cardiopulmonary resuscitation (CPR) monitoring
possibilities are limited. Parnia and colleagues investigated the
feasibility and role of near-infrared spectroscopy (NIRS) during CPR
in cardiac arrest patients (CA) [1]. NIRS could have a role in predicting
return of spontaneous circulation (ROSC). Recently, the Equanox® with
four wavelengths sensor was validated to provide absolute data on
regional cerebral saturation [2]. We measured cerebral oxygenation
(rSO2) during CPR with NIRS technology and analyzed the diff erences
between initial cerebral saturations in patients achieving ROSC
compared with patients without ROSC. p
g
y
Conclusion As previously reported [2], IHCA was associated with a
worse prognosis than OHCA. The OHCA survival rate was better than
reported elsewhere [3]. The percentage of IHCA attributed to MI was
low. Only one OHCA patient was referred for emergency PCI. Routine
coronary angiography with ad hoc PCI in VF OHCA has been associated
with increased survival [4]. Greater availability of PCI post OHCA could
further improve mortality in patients with a primary cardiac pathology. Further investigation should include management of noncardiogenic
cardiac arrest. p
p
Methods With IRB approval, rSO2 was measured with NIRS during
resuscitation in 18 out-of hospital CA patients. The Equanox® Advance
(NONIN), a NIRS monitoring device that measures absolute rSO2 values,
was applied on the right side of the patient’s forehead when the
medical emergency team arrived in a resuscitation setting. Placement
of the probe did not interfere with the advanced life support algorithm. The sensor remained on the patient’s forehead during resuscitation,
and if ROSC was reached the probe was removed on arrival at the
emergency department. If ROSC was not achieved, the probe was
removed prehospital. ROSC was defi ned as ROSC during more than
20 minutes. The Mann–Whitney test was utilized for comparison of
survivor and nonsurvivor data. Student’s t test was performed to
compare the initial rSO2. 1.
Parnia et al.: Resuscitation 2012, 83:982-985. P307 P307
Intensive care admission after cardiac arrest: cardiac versus
noncardiac causes and consequences for treatment
T Hargreaves1, H Kingston1, M Crews1, M Mogk2, I Welters3
1Royal Liverpool University Hospital, Liverpool, UK; 2Moredata GmbH, Giessen,
Germany; 3Liverpool University, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P307 (doi: 10.1186/cc12245) Intensive care admission after cardiac arrest: cardiac versus
noncardiac causes and consequences for treatment T Hargreaves1, H Kingston1, M Crews1, M Mogk2, I Welters3
1Royal Liverpool University Hospital, Liverpool, UK; 2Moredata GmbH, Giessen,
Germany; 3Liverpool University, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P307 (doi: 10.1186/cc12245) Introduction In-hospital (IHCA) and out-of-hospital cardiac arrest
(OHCA) are associated with high mortality [1]. Studies suggest that
up to 68% of OHCA is due to acute coronary syndrome, with 38%
requiring percutaneous coronary intervention (PCI) [1]. However,
revascularisation may not always be available or address the underlying
pathology. This study aimed to establish the prevalence of diff erent
aetiologies for IHCA and OHCA, and the use of emergency treatment
for these patients. disability (CPC 3 to 4) was seen in 19%, greater than previously reported
[1]. A higher proportion (35%) of patients discharged from the ICU had
moderate–severe neurological disability; most subsequently died
in hospital (62%). These fi gures may represent better ICU outcomes
subsequent to adoption of OOHCA bundles but suggest further work is
required in neuro-disabled survivors. Methods A retrospective case-note review of all patients admitted
between 2008 and 2011 to the ICU of an inner-city university hospital
after OHCA or IHCA. Biometric data, presenting cardiac rhythm,
presumed cause of arrest, management and outcomes were recorded. The Kruskal–Wallis test was used for numerical data analysis and chi-
square test for categorical data. Survival from cardiopulmonary arrest after regular working hours
in a tertiary-care hospital: retrospective study Introduction Detection and treatment of cardiopulmonary arrest and
their antecedents may be less eff ective at night and weekend than
weekdays because of hospital staffi ng and response factors [1]. Early Conclusion OOHCA patients admitted to our ICU had a 46% chance
of surviving to hospital discharge. Most patients left hospital with
good neurological status (CPC 1 to 2); moderate–severe neurological S116 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 disability (CPC 3 to 4) was seen in 19%, greater than previously reported
Table 1 (abstract P305). Discharge and neurological status of OOHCA patients
Discharge
Hospital discharge destination
Neuro-
Score
ICU
Hospital
Home
rehab
Repatriation Hospice
CPC 1
20
22
19
0
3
0
CPC 2
6
4
4
0
0
0
CPC 3
4
3
0
3
0
0
CPC 4
10
3
0
0
1
2
Total
40
32
23
3
4
2 disability (CPC 3 to 4) was seen in 19%, greater than previously reported
[1]. A higher proportion (35%) of patients discharged from the ICU had
moderate–severe neurological disability; most subsequently died
in hospital (62%). These fi gures may represent better ICU outcomes
subsequent to adoption of OOHCA bundles but suggest further work is
required in neuro-disabled survivors. References
1. Cobbe SM, et al.: BMJ 1999, 312:1633-1637. 2. Holzer M, et al.: N Engl J Med 2002, 346:549-556. P306
Start value of cerebral saturation in prehospital cardiac arrest
patients: does it mean something? C Genbrugge, I Meex, S Scheyltjens, J Dens, C De Deyne
ZOL, Genk, Belgium
Critical Care 2013, 17(Suppl 2):P306 (doi: 10.1186/cc12244)
Table 1 (abstract P305). Discharge and neurological status of OOHCA patients
Discharge
Hospital discharge destination
Neuro-
Score
ICU
Hospital
Home
rehab
Repatriation Hospice
CPC 1
20
22
19
0
3
0
CPC 2
6
4
4
0
0
0
CPC 3
4
3
0
3
0
0
CPC 4
10
3
0
0
1
2
Total
40
32
23
3
4
2 Table 1 (abstract P305). Discharge and neurological status of OOHCA patients
Discharge
Hospital discharge destination 1.
Parnia et al.: Resuscitation 2012, 83:982-985. References 1. Cobbe SM, et al.: BMJ 1999, 312:1633-1637. 1. Cobbe SM, et al.: BMJ 1999, 312:1633-1637. 2. Holzer M, et al.: N Engl J Med 2002, 346:549-556. q
g
Results Data were analysed for 64 patients – 44 (69%) following OHCA
and 20 (31%) after IHCA. The median APACHE score for OHCA was
17 and for IHCA was 23.5 (P = 0.001). Hospital survival rate was 10%
(n = 2) for IHCA and 38.6% (n = 17) for OHCA (P <0.02). A total of 34.1%
(n = 15) OHCA were due to myocardial infarction (MI) compared with
10% (n = 2) of IHCA (P <0.05). The most prevalent aetiologies were MI
(n = 17), hypoxia (n = 10), cardiac other (n = 5), sepsis (n = 4), arrhythmia
(n = 3) and PE (n = 3). In two IHCA patients more than one likely cause
of arrest was reported and in 19 cases no cause was identifi ed. The
presenting rhythm was ventricular fi brillation (VF) in 45.3% (n = 29),
pulseless electrical activity in 32.8% (n = 21) and asystole in 20.3%
(n = 13). A total of 9.4% (n = 6) were thrombolysed and one (1.6%)
patient was referred for emergency PCI. 2. Holzer M, et al.: N Engl J Med 2002, 346:549-556. 1.
Parnia et al.: Resuscitation 2012, 83:982-985.
2.
MacLeod et al.: J Cardiothorac Vasc Anesth 2012, 26:1007-1014. P308 P308
Comparison of 1-year neurological outcome between intra-hospital
and extra-hospital cardiac arrest survivors submitted to mild
therapeutic hypothermia in a community-based setting in Brazil
CA Abreu Filho1, A Andrade2, A Neto2, S Santos2, M Bracco2, E Silva1,
A Baruzzi1, J Teixeira2
1Hospital Israelita Albert Einstein, São Paulo, Brazil; 2Hospital Municipal Dr. Moysés Deutsch, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P308 (doi: 10.1186/cc12246) 2
Results Of the 18 patients, nine patients had ROSC (survivors). The
initial rhythm was the same in both groups, six patients in each
group had asystole as initial rhythm. In the group of survivors were
six female patients, in the nonsurvivors were two female patients. The
mean age in ROSC and no-ROSC groups is respectively 75.8 years (SD
±12.8) and 69.4 years (SD ±22.9, P = 0.48). The mean rSO2 at arrival of
the emergency medical team was 31.56% (SD ±29.4) and 12.78% (SD
±12.7) respectively in the ROSC group and no-ROSC group (P = 0.1). The mean time between collapse and start of CPR (basic life support
of bystanders) was 6.9 minutes (SD ±8.2) in the no-ROSC group and
8.2 minutes (SD ±7.08, P = 0.69) in the ROSC group. Introduction Mild therapeutic hypothermia (MTH) is the most powerful
therapy to improve survival and neurologic outcome after out-of-
hospital cardiac arrest. Such benefi t may also occur for unconscious
patients after in-hospital cardiac arrest. The aim is to compare 1-year
evolution of neurological outcomes of patients treated with MTH after
in-hospital versus out-of-hospital cardiac arrest. Conclusion Initial rSO2 values in out-of hospital CA patients with ROSC
showed a tendency towards higher values compared with nonsurvivors,
but no signifi cant diff erence could be demonstrated, probably related
to the small number of patients included in this preliminary report. References Methods A prospective study of patients treated with MTH after
cardiac arrest in a community hospital in São Paulo, Brazil. After return
of spontaneous circulation, unconscious survivors received MTH using
topical ice and cold saline infusions in order to achieve a 32 to 34°C
goal temperature, within 6 hours of cardiac arrest, and maintained P310 P310
Infrared pupillometry for outcome prediction after cardiac arrest
and therapeutic hypothermia
T Suys, N Sala, AO Rossetti, M Oddo
Lausanne University Hospital, Lausanne, Switzerland
Critical Care 2013, 17(Suppl 2):P310 (doi: 10.1186/cc12248) Conclusion Midterm neurological outcome of MTH after in hospital
cardiac arrest seems to be not as good as after out-of-hospital cardiac
arrest. Delay in hypothermia initiation, older age and associated
comorbidities could explain the worse evolution of this group of
patients. Introduction Sedation and therapeutic hypothermia (TH) modify
neurological examination and alter prognostic prediction of coma after
cardiac arrest (CA). Additional tools, such as EEG and evoked potentials,
improve prediction of outcome in this setting, but are not widely
available and require signifi cant implementation. i
Methods Using a new device for infrared pupillometry, we examined the
value of quantitative pupillary light reactivity (PLR) to predict outcome
in comatose post-CA patients treated with TH. Twenty-four comatose
CA patients treated with TH (33°C, 24 hours) were prospectively
studied. The percentage variation in PLR was measured during TH
(12 hours from CA), using the NeuroLight Algiscan® (IDMED, Marseille,
France). For each patient, three consecutive measures were performed
and the best value was retained for analysis. The relationship of PLR P309 Primary percutaneous coronary angioplasty and therapeutic
hypothermia in out-of-hospital cardiac arrest
R Hunt, M Holl, A Bailey, P Macnaughton
Derriford Hospital, Plymouth, UK
Critical Care 2013, 17(Suppl 2):P309 (doi: 10.1186/cc12247) Introduction The benefi t of primary percutaneous intervention (PCI)
in the management of out-of-hospital cardiac arrest (OHCA) is not
clear cut [1]. It has historically been used in patients with ST elevation
on post-resuscitation electrocardiogram (ECG) although this is a poor
predictor of acute coronary occlusion after cardiac arrest [2]. This
study investigates the benefi t of PCI regardless of post-resuscitation
ECG. Benefi t is widely claimed for therapeutic hypothermia, so cooling
parameters were included. Table 1 (abstract P310). False-positive and false-negative rates for outcome
(% of patients) p
Methods We analysed all 41 consecutive adults admitted post OHCA to
a university hospital ICU between January 2010 and December 2011. Patients received PCI regardless of ECG changes. A Cox proportional
hazards model was used to determine the relationship between PCI,
cooling and survival to discharge. Routinely collected data such as
demographics and details of resuscitation (OHCA Utstein data) were
also included. Results Survival to hospital discharge was 41% with 29% of survivors
discharged to a neurological rehabilitation centre. References 1. Kern KB: Optimal treatment of patients surviving out-of-hospital cardiac
arrest. JACC Cardiovasc Interv 2012, 5:597-605. 2. Spaulding CM, et al.: Immediate coronary angiography in survivors of out-
of-hospital cardiac arrest. N Engl J Med 1997, 336:1629-1633. 2. Spaulding CM, et al.: Immediate coronary angiography in survivors of out-
of-hospital cardiac arrest. N Engl J Med 1997, 336:1629-1633. 3. Dumas F, et al.: Immediate percutaneous coronary intervention is
associated with better survival after out-of-hospital cardiac arrest. Circ
Cardiovasc Interv 2010, 3:200-207. 3. Dumas F, et al.: Immediate percutaneous coronary intervention is
associated with better survival after out-of-hospital cardiac arrest. Circ
Cardiovasc Interv 2010, 3:200-207. References S117 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 with other studies suggesting benefi t for primary PCI regardless of the
post-resuscitation ECG [3]. Cooling was not found to improve survival
to discharge but further analysis is required to determine impact on
neurological function. for 24 hours. Esophageal temperature was monitored; continuous
intravenous sedation-analgesia was maintained for 48 hours after
initiation of MTH. The Glasgow Outcome Scale (GOS) was used to
analyze the neurological outcomes after hospital discharge. for 24 hours. Esophageal temperature was monitored; continuous
intravenous sedation-analgesia was maintained for 48 hours after
initiation of MTH. The Glasgow Outcome Scale (GOS) was used to
analyze the neurological outcomes after hospital discharge. Results From January 2009 to April 2012, 84 patients submitted to MTH
were divided into two groups: Group 1, 54 patients presented out-
of-hospital cardiac arrest; and Group 2, 30 patients had intra-hospital
cardiac arrest. Both groups were similar regarding to gender; Group
2 tended to be older (mean age 44.3 years vs. 33.5 years, P = 0.07),
and had more frequently asystole as the cardiac arrest rhythm (45%
vs. 15 %, P = 0.10). Group 2 had shorter duration of resuscitation
(12.3 minutes vs. 33.7 minutes, P = 0.03), longer time to hypothermia
initiation (309.3 minutes vs. 212.8 minutes, P = 0.04), longer hospital
stay after cardiac arrest (50.7 days vs. 32.4 days, P = 0.01) and worse
neurological outcome, characterized by the presence of GOS ≤3 at 30
days (68.4% vs. 35.1%, P = 0.03). Hospital mortality was 5.5% in Group 1
and 13.3% in Group 2 (P = 0.21). The 1-year survival rate was 85.1% in
Group 1 and 83.3% in Group 2 (P = 0.43); after 1 year follow-up, GOS
≤3 was present in 30.4% of Group 2 patients and in 11.1% of Group 1
patients (P = 0.04). Eff ect of moderate hyperventilation and induced hypertension on
cerebral tissue oxygen saturation in comatose post-cardiac arrest
patients treated with hypothermia
T Suys, N Sala, M Oddo
Lausanne University Hospital, Lausanne, Switzerland
Critical Care 2013, 17(Suppl 2):P311 (doi: 10.1186/cc12249) l
Methods We performed an observational study in 10 comatose
patients after cardiac arrest. Patients were treated with hypothermia for
24 hours. Blood viscosity was measured ex vivo using a Contraves LS300
viscometer. Mean fl ow velocity in the middle cerebral artery (MFVMCA)
was measured by transcranial Doppler (TCD) at the same time points. T Suys, N Sala, M Oddo T Suys, N Sala, M Oddo
Lausanne University Hospital, Lausanne, Switzerland
Critical Care 2013, 17(Suppl 2):P311 (doi: 10.1186/cc12249) Lausanne University Hospital, Lausanne, Switzerland y
p
Critical Care 2013, 17(Suppl 2):P311 (doi: 10.1186/cc12249) Introduction Maintenance of adequate brain perfusion is an essential
component of post-resuscitation care. Mean arterial pressure (MAP)
and PaCO2 are important determinants of brain perfusion; however, no
precise guidelines exist for optimal MAP and PaCO2 targets in comatose
post-cardiac arrest (CA) patients. an essential
ssure (MAP)
however, no
n comatose
ive cerebral
erventilation
ents treated
dy including
hospital CA
face cooling
NIRS system
erventilation
PaCO2 from
zation, MAP
infusion of
SctO2 were
19 minutes)
Results are
nt reduction
5), P <0.001). and IH tests (n = 10 patients). Figure 1 (abstract P312). Figure 1 (abstract P312). p
p
Methods Using NIRS, we examined changes in non-invasive cerebral
tissue oxygen saturation (SctO2) following moderate hyperventilation
(HV) and induced hypertension (IH) in comatose CA patients treated
with therapeutic hypothermia (TH). A prospective pilot study including
comatose patients successfully resuscitated from out-of-hospital CA
treated with TH (33°C for 24 hours, using cold saline and surface cooling
device), monitored for continuous SctO2 with the Foresight NIRS system
(CAS Medical Systems, Branford, CT, USA). Moderate hyperventilation
was induced for approximately 30 minutes by decreasing PaCO2 from
~40 to ~30 mmHg, at stable MAP. After PaCO2 normalization, MAP
was increased from ~70 to ~90 mmHg by intravenous infusion of
norepinephrine, at stable PaCO2. Eff ects of MV and IH on SctO2 were
analyzed with a paired t test. Figure 1 (abstract P312). Figure 1 (abstract P312). y
p
Results Ten patients (mean age, 69.5; mean time to ROSC, 19 minutes)
were studied during the stable TH maintenance phase. Results are
summarized in Figure 1. MV was associated with a signifi cant reduction
of SctO2 from baseline (75% (73 to 76) to 69% (67.5 to 71.5), P <0.001). 0 patients). Figure 1 (abstract P311). Changes in SctO2 after moderate HV and IH tests (n = 10 patients). Figure 1 (abstract P311). P312f Eff ects of viscosity on cerebral blood fl ow after cardiac arrest
L Bisschops, G Pop, S Teerenstra, J Van der Hoeven, C Hoedemaekers
Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P312 (doi: 10.1186/cc12250) p
g
Conclusion Quantitative PLR appears highly accurate and superior to
standard neurological examination to predict outcome in patients with
post-CA coma. Further study is warranted to confi rm these promising
fi ndings. Acknowledgements Supported by Grants from the Swiss National
Science Foundation (FN 320030_138191) and the European Critical
Care Research Network (ECCRN). Introduction After cardiac arrest, microcirculatory reperfusion dis orders
develop despite adequate cerebral perfusion pressure. Increased blood
viscosity strongly hampers the microcirculation, resulting in plugging
of the capillary bed, arteriovenous shunting and diminished tissue
perfusion. The aim of the present study was to assess blood viscosity in
relation to cerebral blood fl ow in patients after cardiac arrest. P310 Multivariate
analysis using a Cox proportional hazards model showed PCI to be an
independent predictive factor of survival, unrelated to ECG (hazards
ratio, 0.0583; 95% CI, 0.0076 to 0.4485). Cooling had no signifi cant
impact on patient survival. See Figure 1. p
p
g
Conclusion In this small retrospective study primary PCI appears to
be an independent predictor of survival after OHCA. This is consistent Figure 1 (abstract P309). Comparison of hazards ratios post Cox analysis. Figure 1 (abstract P309). Comparison of hazards ratios post Cox analysis. S118 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 No signifi cant changes in SctO2 were found after IH (74 (72 to 76) vs. 75
(73 to 75.5), P = 0.24). with survival and neurological outcome (CPC scores) at 3 months was
analyzed, and the predictive value of PLR was compared with that of
standard clinical examination (motor response and brainstem refl exes)
performed at 48 hours from CA. Conclusion Moderate HV was associated with signifi cant reduction
in cerebral saturation, whilst IH may be detrimental after CA and TH,
whilst increasing MAP to supranormal levels with vasopressors does
not improve cerebral oxygenation. These data stress the importance of
strict control of PaCO2 following CA and TH to avoid secondary cerebral
ischemic insults. Results Quantitative PLR was strongly associated with survival (median
left-eye PLR 14% (11 to 16%) variation in survivors vs. 5.5% (4 to 8.5%)
in nonsurvivors, P < 0.0001) and 3-month neurological outcome (14%
(11 to 21%) in patients with CPC 1 to 2 vs. 5.5% (4 to 8.5%) in those
with CPC 3 to 5, P <0.0001). Comparable fi ndings were obtained using
right-eye PLR. A PLR >10% was 100% predictive of patient prognosis,
with false-positive and false-negative rates of 0% for outcome. Clinical
examination was signifi cantly associated with outcome; however,
motor response (MR) and brainstem refl exes (BRS) yielded higher false-
positive and false-negative rates than PLR (Table 1). Acknowledgements MO is supported by Grants from the Swiss
National Science Foundation (FN 320030_138191) and The European
Critical Care Research Network (ECCRN). P312f Eff ect of moderate hyperventilation and induced hypertension on
cerebral tissue oxygen saturation in comatose post-cardiac arrest
patients treated with hypothermia Eff ect of moderate hyperventilation and induced hypertension on
cerebral tissue oxygen saturation in comatose post-cardiac arrest
patients treated with hypothermia
T Suys, N Sala, M Oddo
Lausanne University Hospital, Lausanne, Switzerland
Critical Care 2013, 17(Suppl 2):P311 (doi: 10.1186/cc12249) Association between oxygenation and 6-month mortality during
post-cardiac arrest care p
M Skrifvars1, J Vaahersalo1, M Reinikainen2, S Bendel3, J Kurola3, M Tiainen1,
R Raj1, V Pettilä1, T Varpula1, FinnResusci Study Group1
1Helsinki University Hospital, HUS, Finland; 2North Karelia Central Hosital,
Joensuu, Finland; 3Kuopio University Hospital, Kuopio, Finland
Critical Care 2013, 17(Suppl 2):P315 (doi: 10.1186/cc12253) M Skrifvars1, J Vaahersalo1, M Reinikainen2, S Bendel3, J Kurola3, M Tiainen1
R Raj1, V Pettilä1, T Varpula1, FinnResusci Study Group1
1Helsinki University Hospital, HUS, Finland; 2North Karelia Central Hosital,
Joensuu, Finland; 3Kuopio University Hospital, Kuopio, Finland
Critical Care 2013, 17(Suppl 2):P315 (doi: 10.1186/cc12253) g
g
y
g
g
y
Results We had 297 CA patients (168 SR, 129 NR) in whom CPR was
done. Return of spontaneous circulation was achieved in 90 patients. TMH was induced in 57 patients (33 SR, 24 NR). Survival to hospital
discharge was observed in 27 patients (18/33 (54.5%) SR, 9/24 (37.5%)
NR), out of which 18 patients (10/33 (30%) SR, 8/24 (33%) NR) had good
neurological outcome. On analysis, TMH was found to be associated
with increased odds of survival to hospital discharge (although
statistically not signifi cant) in SR patients compared with NR patients
(odds ratio (OR) = 2.00; 95% CI = 0.68 to 5.85; P = 0.2837), but it was not
associated with any better neurological outcome in terms of CPC score
in patients presenting with SR rather than NR (OR = 0.87; 95% CI = 0.28
to 2.68; P = 1.0000). Rather, the odds for good neurological outcome
were more in favour of NR (pulseless electrical activity/asystole). Introduction Optimal oxygenation level during post-cardiac arrest
(PCA) care is currently undefi ned, and studies have suggested
harm from hyperoxia exposure [1]. We aimed to assess the optimal
oxygenation level and possible associations of time-weighted exposure
to hyperoxia on outcome in patients during PCA care. Introduction Optimal oxygenation level during post-cardiac arrest
(PCA) care is currently undefi ned, and studies have suggested
harm from hyperoxia exposure [1]. We aimed to assess the optimal
oxygenation level and possible associations of time-weighted exposure
to hyperoxia on outcome in patients during PCA care. Methods We conducted a prospective observational cohort study in
21 ICUs in Finland between 2009 and 2010. The Utstein Guidelines
were used for collecting resuscitation and PCA care data, such as
initial rhythm and delay to return of spontaneous circulation (ROSC). P313 p
Results Agreement between the diff erent sources of temperature
measurement was generally high, with less than 10% outliers beyond
±1.96 SD. The best agreement was seen between intravascular
tempera ture measurement and bladder temperature measurement
(SD = 0.14°C), with vaginal measurements showing less agreement
(SD = 0.23°C), and rectal measurements showing the least (SD = 0.31°C)
with intravascular. Eff ect of moderate hyperventilation and induced hypertension on
cerebral tissue oxygen saturation in comatose post-cardiac arrest
patients treated with hypothermia
T Suys, N Sala, M Oddo
Lausanne University Hospital, Lausanne, Switzerland
Critical Care 2013, 17(Suppl 2):P311 (doi: 10.1186/cc12249) Changes in SctO2 after moderate HV and IH tests (n = 10 patients). S119 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Results The median viscosity on admission was 9.12 (8.19 to 11.19) mPa. second, and remained stable at 3 and 6 hours after admission. From
6 hours after admission, viscosity decreased signifi cantly to 3.66 (3.12
to 4.04) mPa.second (P <0.001). Median MFVMCA was low (27.0 (23.8
to 30.5) cm/second) on admission, and signifi cantly increased to 63.0
(51.0 to 80.0) cm/second at 72 hours (P <0.001). There was a signifi cant
association between viscosity and the MFVMCA (P = 0.0019). See
Figure 1. where temperature is measured. Measurements of temperature can
be made rectally, intravascularly, vaginally, or from the bladder, but
agreement between these sites is often uncertain. We measured
temperature at multiple sites during experimental induction and
reversal of hypothermia in swine with a novel esophageal cooling
device, hypothesizing that agreement between sites would fall within
standard acceptance criteria (average diff erence ± 1.96 standard
deviation of the diff erence) in Bland–Altman analyses. g
Conclusion Viscosity decreases in the fi rst 3 days after cardiac arrest
and is strongly associated with an increase in cerebral blood fl ow. Since viscosity is a major determinant of cerebral blood fl ow, repeated
measurements may guide therapy to help restore cerebral oxygenation
after cardiac arrest. f
y
Methods Five female Yorkshire swine (mean weight 65 kg) were
anesthetized and cooled for 24 hours, then gradually rewarmed, with
a novel esophageal heat transfer device powered by an external chiller
(Gaymar MediTherm III). Swine temperature was measured intra-
vascularly, rectally, and either vaginally or in the bladder. Comparisons
between temperature readings were then made via Bland–Altman
plots. Reference 1. Arrich J, Holzer M, Herkner H, Mullner M: Hypothermia for neuroprotection
in adults after cardiopulmonary resuscitation. Cochrane Database Syst Rev
2009, 4:CD004128. Methods We reviewed medical records of all CA patients (in-hospital
or out-of-hospital arrest) in whom cardiopulmonary resuscitation (CPR)
was performed at our hospital from 1 February 2011 to 31 January 2012
(12 months). The following information was collected: fi rst documented
rhythm, whether TMH done or not, and two outcome measures
including survival to hospital discharge and neurological outcome
at the time of hospital discharge. A measure of good neurological
outcome was Cerebral Performance Category score 1 or 2 (CPC, fi ve-
point scale; 1 = good cerebral performance to 5 = brain death). Then
we quantifi ed the association of TMH with SR as well as NR for both
the parameters of outcome – that is, survival to hospital discharge and
good neurological outcome – by logistic regression analysis. Association between oxygenation and 6-month mortality during
post-cardiac arrest care Measured arterial blood oxygen values during the fi rst 24 hours from
admission to the ICU were divided into the following predefi ned
oxygenation categories: low (<10 kPa), normal (10 to 19 kPa),
intermediate (20 to 29 kPa), and high (>30 kPa). Exposure to hyperoxia
was defi ned as paO2 levels higher than 40 kPa [1]. Time spent in
diff erent oxygenation categories and the highest, lowest and median
oxygen values during the fi rst 24 hours were calculated and included
in separate multivariate regression models along with age, delay to
ROSC, initial rhythm and the use of therapeutic hypothermia for the
prediction of 6-month mortality. Conclusion Although TMH might be associated with better survival
chances in patients presenting with SR, neurological outcome was no
better (rather worse) in this group of patients when compared with
patients with NR as the fi rst documented rhythm. Therapeutic mild hypothermia after cardiac arrest in shockable
and nonshockable rhythms: does it improve both survival and
neurological outcome? A Gupta Fortis Escorts Heart Institute, New Delhi, India
Critical Care 2013, 17(Suppl 2):P313 (doi: 10.1186/cc12251) Introduction Although therapeutic mild hypothermia (TMH) after
resuscitation from cardiac arrest (CA) has been postulated and studied
to be associated with good outcome of the patients, there is no dearth
of data that does not favour TMH. Our aim was to fi nd out whether
TMH is associated with good outcome after CA in shockable rhythm
(SR) compared with nonshockable rhythm (NR), in terms of survival as
well as neurological outcome. Conclusion Bladder temperature measurements demonstrated the
best agreement with intravascular temperature measurements during
cooling with an esophageal heat transfer device; however, reasonable
agreement was demonstrated with rectal and vaginal temperature
measurements, suggesting that these sites are also acceptable for use. Reference Diff erences of calcium binding protein immunoreactivities in the
young hippocampal CA1 region from the adult following transient
ischemic damage Results NSE >45 occurred in 24/68 patients (35.3%) and invariably
correlates with bad outcome. The positive predictive value (PPV) NSE
>45 for bad outcome is 100%. No patient in this group ever had a GCS
≥12. NSE >33 and <45 occurred in 16/68 patients (23.5%). Thirteen
out of 16 patients (81.2%) had bad outcome. However, 7/16 patients
(43.8%) woke up at some time (GCS ≥12). NSE <33 occurred in 28/68
patients (41.2%), 17/28 patients (60.7%) had good outcome and 23/28
patients (88.4%) had GCS ≥12 at some time. The PPV NSE <33 for good
outcome is 60.7%. The BIS and SR were measured in only 28 patients. Initial BIS ≤10 occurred in 13/28 patients (46.4%) and correlates with
bad outcome in 12/13 patients (92.3%). BIS >30 occurred in nine
patients, 6/9 (66.7%) had good outcome. Initial SR ≥75 occurred in
11/28 patients (39.3%) and invariably correlates with bad outcome. NSE >25 and SR >60 occurred in 15/28 patients (53.6%) and invariably
correlates with bad outcome. g
CW Park, HY Lee, JH Cho, MH Won Introduction It has been reported that the young are much more
resistant to transient cerebral ischemia than the adult. Introduction It has been reported that the young are much more
resistant to transient cerebral ischemia than the adult. Methods In the present study, we compared the chronological
changes of calcium binding proteins (CBPs) (calbindin 28k (CB-D 28k),
calretinin (CR) and parvalbumin (PV)) immunoreactivities and levels
in the hippocampal CA1 region of the young gerbil with those in the
adult following 5 minutes of transient cerebral ischemia induced by the
occlusion of both the common carotid arteries. Results In the present study, we examined that about 90% of CA1
pyramidal cells in the adult gerbil hippocampus died at 4 days post
ischemia; however, in the young hippocampus, about 56% of them
died at 7 days post ischemia. We compared immunoreactivities and
levels of CBPs, such as CB-D 28k, CR and PV. The immunoreactivities
and protein levels of all the CBPs in the young sham were higher than
those in the adult sham. In the adult, the immunoreactivities and
protein levels of all the CBPs were markedly decreased at 4 days post
ischemia; however, in the young, they were apparently maintained. At
7 days post ischemia, they were decreased in the young; however, they
were much higher than those in the adult. p
References Introduction Neuron-specifi c enolase (NSE) values >33 μg/ml [1] and
low bispectral index (BIS) [2] values correlate with bad outcome after
cardiac arrest (CA). 1. Wijdicks et al.: Neurology 2006, 67:203-210. 2. Al Thenayan et al.: Neurology 2008, 71:1535-1537. 3. Rossetti et al.: Ann Neurol 2010, 67:301-307. 4. Bisschops et al.: Resuscitation 2011, 82:696-701. 5. Cronberg et al.: Neurology 2011, 77:623-630. 6. Samaniego et al.: Neurocrit Care 2011, 15:113-119. 7. Fugate et al.: Ann Neurol 2010, 68:907-914. 1. Wijdicks et al.: Neurology 2006, 67:203-210. 2. Al Thenayan et al.: Neurology 2008, 71:1535-1537. 3. Rossetti et al.: Ann Neurol 2010, 67:301-307. 4. Bisschops et al.: Resuscitation 2011, 82:696-701. 5. Cronberg et al.: Neurology 2011, 77:623-630. 6. Samaniego et al.: Neurocrit Care 2011, 15:113-119. 7. Fugate et al.: Ann Neurol 2010, 68:907-914. Methods In this nonblinded prospective study, we observed all CA
patients from February 2011 until September 2012 surviving at least
24 hours. NSE was measured between 24 and 72 hours after CA. From
October 2011 onward, we recorded BIS and suppression ratio (SR)
values as soon as possible after arrival in the ICU. Patients treated
with therapeutic hypothermia (TH) (33°C for 24 hours) received
cisatracurium. Cerebral Performance Category (CPC) [3] 1 and 2 were
considered good outcome, CPC 3 to 5 bad outcome and were recorded
after 3 months. Statistical analysis was performed using SPSS statistics
19. Diff erences of calcium binding protein immunoreactivities in the
young hippocampal CA1 region from the adult following transient
ischemic damage Conclusion NSE >45 uniformly correlates with bad outcome after CA. However, we urge caution for the use of intermediate values (33 to 45). In preliminary data, we report that SR >75 might correlate with bad
outcome and that combining NSE and SR might improve the predictive
value. Also, low NSE and good initial BIS values correlate with preserved
cerebral potential and should encourage the clinician. 1. Zandbergen et al.: Neurology 2006, 66:62-68. 1. Zandbergen et al.: Neurology 2006, 66:62-68. 2. Leary et al.: Resuscitation 2010, 81:1133-1137. Conclusion In brief, the immunoreactivities and levels of CBPs were not
decreased in the ischemic CA1 region of the young 4 days after transient
cerebral ischemia. This fi nding indicates that the longer maintenance
of CBPs may contribute to a less and more delayed neuronal death/
damage in the young. 3. Ajam et al.: Scand J Trauma Resusc Emerg Med 2011, 19:38. Neuron-specifi c enolase and bispectral index/suppression ratio for
prognostication after cardiac arrest i
Conclusion Motor scores at day 3 post cardiac arrest of extension
or worse do not reliably predict poor neurological outcome when
therapeutic hypothermia has been used. Clinical neurological fi ndings
may not be valid predictors of poor neurological outcome after
therapeutic hypothermia. g
Van Laer1, K Deschilder2, P Lormans2, J Gillet1, W Stockman2 ,
,
,
,
1UZ Leuven, Belgium; 2HHRM, Roeselare, Belgium 1UZ Leuven, Belgium; 2HHRM, Roeselare, Belgium g
g
Critical Care 2013, 17(Suppl 2):P316 (doi: 10.1186/cc12254) g
g
Critical Care 2013, 17(Suppl 2):P316 (doi: 10.1186/cc12254) Introduction Neuron-specifi c enolase (NSE) values >33 μg/ml [1] and
low bispectral index (BIS) [2] values correlate with bad outcome after
cardiac arrest (CA). Agreement between temperature measurements during cooling
and warming through the esophagus Conclusion Motor scores at day 3 post cardiac arrest of extension
or worse do not reliably predict poor neurological outcome when
therapeutic hypothermia has been used. Clinical neurological fi ndings
may not be valid predictors of poor neurological outcome after
therapeutic hypothermia. 2
Conclusion In this multicentre observational study we were unable
to defi ne an optimal oxygenation level during PCA care, but hypoxia
seemed to be more harmful than hyperoxia. Exposure to hyperoxia was
less common than in previous trials, and we were unable to confi rm
previous fi ndings indicating an association with mortality. Reference Methods A review of existing literature was undertaken to examine
whether the utility of motor scores to predict poor neurological
outcome is infl uenced by the use of therapeutic hypothermia.i l
Results Six studies were identifi ed [2-7] that investigated the use of
motor scores on day 3 post cardiac arrest in patients who had received
therapeutic hypothermia. False positive rates (defi ned as 1 – specifi city)
for predicting poor neurological outcome were calculable in fi ve of the
six studies [2-6] and were 14%, 24%, 11%, 25% and 12% respectively. In all studies the FPR for motor scores of extension or worse were
signifi cantly higher than the 0% (0 to 3% 95% CIs) in the AAN guidelines. 1. Kilgannon JH, et al.: JAMA 2010, 303:2165-2171. Motor scores, therapeutic hypothermia and neurological outcome
after cardiac arrest M Davidson Agreement between temperature measurements during cooling
and warming through the esophagus False positive rates (defi ned as 1 – specifi city)
for predicting poor neurological outcome were calculable in fi ve of the
six studies [2-6] and were 14%, 24%, 11%, 25% and 12% respectively. In all studies the FPR for motor scores of extension or worse were
signifi cantly higher than the 0% (0 to 3% 95% CIs) in the AAN guidelines. Conclusion Motor scores at day 3 post cardiac arrest of extension
or worse do not reliably predict poor neurological outcome when
therapeutic hypothermia has been used. Clinical neurological fi ndings
may not be valid predictors of poor neurological outcome after
therapeutic hypothermia. (22 kPa vs. 20 kPa, P = 0.054). In separate multivariate models neither
time spent in the low or the intermediate categories, or the median,
highest or lowest paO2 was found to correlate with mortality. widespread use of therapeutic hypothermia. The American Association
of Neurology guidelines [1] currently recommend that absent or
extensor motor scores on day 3 post arrest are reliable indicators or
poor neurological outcome with a false positive rate of 0 to 3%. widespread use of therapeutic hypothermia. The American Association
of Neurology guidelines [1] currently recommend that absent or
extensor motor scores on day 3 post arrest are reliable indicators or
poor neurological outcome with a false positive rate of 0 to 3%. desp ead use o t e apeut c ypot e
a. e
e ca
ssoc at o
of Neurology guidelines [1] currently recommend that absent or
extensor motor scores on day 3 post arrest are reliable indicators or
poor neurological outcome with a false positive rate of 0 to 3%. Methods A review of existing literature was undertaken to examine
whether the utility of motor scores to predict poor neurological
outcome is infl uenced by the use of therapeutic hypothermia. Results Six studies were identifi ed [2-7] that investigated the use of
motor scores on day 3 post cardiac arrest in patients who had received
therapeutic hypothermia. False positive rates (defi ned as 1 – specifi city)
for predicting poor neurological outcome were calculable in fi ve of the
six studies [2-6] and were 14%, 24%, 11%, 25% and 12% respectively. In all studies the FPR for motor scores of extension or worse were
signifi cantly higher than the 0% (0 to 3% 95% CIs) in the AAN guidelines. Agreement between temperature measurements during cooling
and warming through the esophagus p
y
Results A total of 489 patients were included. The average number of
paO2 measurements during the fi rst 24 hours was eight per patient. A total of 6% of patients experienced paO2 values higher than 40 kPa
at any time during the fi rst 24 hours. Average times spent in each
time oxygenation category during the fi rst 24 hours were as follows:
low 14%, normal 69%, intermediate 14%, and high 2% of the time. Survivors spent less time in the low band (P = 0.029) and more time
in the intermediate band (P = 0.029) compared with nonsurvivors. The
median paO2 during the fi rst 24 hours was higher in survivors than in
nonsurvivors (15 kPa vs. 14 kPa, P = 0.016) but there was no diff erence
in lowest (11 kPa vs. 10 kPa, P = 0.162) or the highest paO2 values and warming through the esophagus
EB Kulstad1, DM Courtney2, P Shanley3, A Metzger3, T Matsuura3, J Rees4,
K Lurie3, S McKnite5
1Advocate Christ Medical Center, Oak Lawn, IL, USA; 2Northwestern University,
Chicago, IL, USA; 3University of Minnesota Medical Center, Minneapolis, MN,
USA; 4Advanced Circulatory Systems, Minneapolis, MN, USA; 5Minneapolis
Medical Research Foundation, Minneapolis, MN, USA
Critical Care 2013, 17(Suppl 2):P314 (doi: 10.1186/cc12252) Introduction Measurement of temperature during treatment with
therapeutic hypothermia can yield variable values depending on Introduction Measurement of temperature during treatment with
therapeutic hypothermia can yield variable values depending on S120 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (22 kPa vs. 20 kPa, P = 0.054). In separate multivariate models neither
time spent in the low or the intermediate categories, or the median,
highest or lowest paO2 was found to correlate with mortality. widespread use of therapeutic hypothermia. The American Association
of Neurology guidelines [1] currently recommend that absent or
extensor motor scores on day 3 post arrest are reliable indicators or
poor neurological outcome with a false positive rate of 0 to 3%. Methods A review of existing literature was undertaken to examine
whether the utility of motor scores to predict poor neurological
outcome is infl uenced by the use of therapeutic hypothermia. Results Six studies were identifi ed [2-7] that investigated the use of
motor scores on day 3 post cardiac arrest in patients who had received
therapeutic hypothermia. P319 Plymouth Hospitals NHS Trust, Plymouth, UK Plymouth Hospitals NHS Trust, Plymouth, UK P319
Early hypothermia improves survival and reduces the rise of serum
biomarkers after traumatic brain injury in swine
M Kumar, AD Goldberg, M Kashiouris, L Keenan, A Rabinstein
Mayo Clinic, Rochester, MN, USA
Critical Care 2013, 17(Suppl 2):P319 (doi: 10.1186/cc12257) y
p
y
Critical Care 2013, 17(Suppl 2):P317 (doi: 10.1186/cc12255) Critical Care 2013, 17(Suppl 2):P317 (doi: 10.1186/cc12255) Introduction Accurate prediction of neurological outcome after
cardiac arrest is desirable to prevent inappropriate withdrawal of life-
sustaining therapy in patients who could have a good neurological
outcome, and to limit active treatment in patients whose ultimate
neurological outcomes are poor. Established guidelines to predict
neurological outcome after cardiac arrest were developed before the y
,
,
,
Critical Care 2013, 17(Suppl 2):P319 (doi: 10.1186/cc12257) Introduction Poor outcomes in clinical trials on the use of therapeutic
hypothermia following traumatic brain injury (TBI) may be due to the S121 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 delay in reaching target temperature [1]. We hypothesize that early
and rapid induction of hypothermia will mitigate neuronal injury and
improve survival in a swine model of TBI. Results Table 1 shows the magnitude of the problem, highlighting the
leading causes of TBI in Egypt in 2010. Male sex was predominantly
aff ected, 79% of cases. Moderate and severe injuries account for 17.2%
of all TBI presented cases. delay in reaching target temperature [1]. We hypothesize that early
and rapid induction of hypothermia will mitigate neuronal injury and
improve survival in a swine model of TBI. Methods Twenty domestic cross-bred pigs (34 to 35 kg) were subjected
to a 5 ATM (100 ms) lateral fl uid percussion TBI. The brain temperature
and ICP were measured using Camino®. Serum biomarkers for neuronal
injury – S-100β, neuron-specifi c enolase, glial fi brillary acid protein
(GFAP), and neurofi laments heavy chain – were measured daily using
enzyme-linked immunosorbent assay. Twelve of the injured animals
were rapidly cooled to 32°C within 90 minutes of the injury using
a transpulmonary hypothermia technique [2]. Hypothermia was
maintained for 48 hours. Eight injured control animals were maintained
at 37°C. In both groups, anesthesia (isofl urane 1%) was discontinued
and the animals were weaned off the ventilator after 48 hours. Five days
post injury, the surviving animals were euthanized and necropsied. Alcohol intoxication impedes the recognition of traumatic brain
injury in the prehospital setting and may worsen 6-month outcome
R Raj, J Siironen, R Kivisaari, M Kuisma, M Skrifvars
Helsinki University Central Hospital, Helsinki, Finland
Critical Care 2013, 17(Suppl 2):P321 (doi: 10.1186/cc12259) Alcohol intoxication impedes the recognition of traumatic brain
injury in the prehospital setting and may worsen 6-month outcome
R Raj, J Siironen, R Kivisaari, M Kuisma, M Skrifvars
Helsinki University Central Hospital, Helsinki, Finland
Critical Care 2013, 17(Suppl 2):P321 (doi: 10.1186/cc12259) y
g
g
Results Ten of the 12 hypothermia and four of the eight normothermia
animals survived to the end of the 5-day study (χ2 = 2.597, df = 1,
P = 0.1071). Although the probability of type I error between survival
curves was 11%, the study was clinically signifi cant and showed a clear
trend toward improved survival with hypothermia. The intracranial
pressures were signifi cantly (P <0.05) lower in the hypothermia group. Both interventions – that is, general anesthesia and hypothermia –
mitigated the rise of serum biomarkers following TBI. However, the
suppression of biomarkers was sustained during the recovery period
only in the hypothermia group. With the exception of the GFAP levels,
the curves of all biomarkers were signifi cantly diff erent between the
groups.i Introduction Transport directly to a neurosurgical trauma center has
shown to reduce mortality in patients with traumatic brain injury (TBI). We hypothesized that alcohol intoxication may impede prehospital
recognition of TBI, resulting in transportation to a non-neurosurgical
hospital. Methods A retrospective analysis of TBI patients admitted to a
designated neurosurgical trauma center’s ICU in 2009/10 and primarily
treated by the Emergency Medical Service (EMS). Patients were divided
into two categories based either direct or indirect trauma transfer by
the EMS. Directly transferred patients are directly transported to the
neurosurgical trauma center from the injury scene while indirectly
transported patients are initially transported to another non-
neurosurgical hospital before re-transfer to the trauma center. Data
from patient journals and EMS forms were extracted. The blood alcohol
level (BAL) was measured by the EMS using an alcohol breath-test. Logistic regression modeling was used to identify variables present at
scene associated with transport destination. Conclusion Our preliminary fi ndings show early initiation, rapid
induction, and prolonged maintenance (48 hours) of cerebral
hypothermia to lower intracranial pressure, blunt the rise in serum
biomarkers, and improve survival following TBI. 1. Clifton GL, et al.: Lancet Neurol 2011, 10:131-139. 2. Kumar M., et al.: Anesth Analg 2012, 114:S-160. 1. Clifton GL, et al.: Lancet Neurol 2011, 10:131-139. 2. Kumar M., et al.: Anesth Analg 2012, 114:S-160. P319 The
data were analyzed using a log-rank (Mantel–Cox) test, and ANOVA. Conclusion TBI is a serious public health problem in Egypt. Further
data interpretation over wider periods of time should be conducted
for better understanding of TBI prevalence is highly recommended to
develop eff ective injury prevention programs. 1.
Clifton GL, et al.: Lancet Neurol 2011, 10:131-139.
2.
Kumar M., et al.: Anesth Analg 2012, 114:S-160. Alcohol intoxication impedes the recognition of traumatic brain
injury in the prehospital setting and may worsen 6-month outcome
R Raj, J Siironen, R Kivisaari, M Kuisma, M Skrifvars
Helsinki University Central Hospital, Helsinki, Finland
Critical Care 2013, 17(Suppl 2):P321 (doi: 10.1186/cc12259) Results Totally 470 patients met the inclusion criteria; 60% were
transported directly and 40% indirectly. In the direct group 15% of
patients had a positive BAL, compared with 26% for those indirectly
transported. In the logistic regression model, factors associated
with direct transport were: BAL ≥2.3‰ (OR: 0.06, CI: 0.01 to 0.36),
male gender (OR: 0.35; CI: 0.16 to 0.76), GCS 13 to 15 (OR: 0.28; CI
0.10 to 0.74), high-energy trauma (OR: 9.42; CI 2.15 to 41.20), major
extracranial injury (OR: 7.92; CI 2.57 to 24.41), EMS physician telephone
consultation (OR: 6.02; CI 2.51 to 14.11) or presence on scene (OR: 8.63;
CI 3.50 to 21.26) and incident at a public place outside (OR: 3.05; CI:
1.34 to 6.4) and inside (OR: 2.92; CI: 1.07 to 8.01). Median time delay
to trauma center admission was 1:07 hours (IQR: 0.52 to 1:28) for
directly transported patients and 4:06 (IQR: 2.54 to 5:43) for those
indirectly transported (P <0.001). There was a clear trend towards
poorer neurological outcome for patients with delayed trauma center
admission in univariate analysis (P = 0.001). P320 P320
Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) P320
Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) P320
Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) P322
Predictive ability of the IMPACT-TBI prognostic model is improved
by adding markers of coagulation
R Raj, J Siironen, R Kivisaari, L Handolin, J Hernesniemi, M Skrifvars
Helsinki University Central Hospital, Helsinki, Finland
Critical Care 2013, 17(Suppl 2):P322 (doi: 10.1186/cc12260) Predictive ability of the IMPACT-TBI prognostic model is improved
by adding markers of coagulation Methods A prospective cohort of STBI patients monitored with
cerebral microdialysis (CMD) and brain tissue oxygen (PbtO2) was
studied. Brain lactate metabolism was assessed by quantifi cation
of elevated CMD lactate samples (>4 mmol/l). These were matched
to pyruvate and PbtO2, and dichotomized as hyperglycolytic (CMD
pyruvate >119 μmol/l) versus nonhyperglycolytic or as hypoxic (PbtO2
<20 mmHg) versus nonhypoxic. Data were expressed as percentages
per patient. Global brain perfusion (categorized as oligemic, normal or
hyperemic) was assessed with CT perfusion (CTP). Introduction One of the most used prognostic models for traumatic
brain injury is the IMPACT-TBI model, which predicts 6-month mortality
and unfavorable outcome. Our aim was to study whether adding
markers of coagulation improves the model’s predictive power when
accounting for extracranial injury. yp
p
Results Twenty-four patients (total 1,782 CMD samples) were studied. Samples with elevated CMD lactate were frequently observed (41 ± 8%
SEM of individual samples). Brain lactate elevations were predominantly
hyperglycolytic (73 ± 8.2%), whilst only 14 ± 6.3% of them were hypoxic. Trends over time of both lactate patterns are shown in Figure 1. On CTP
(n = 17; average 48 hours from TBI) hyperglycolytic lactate was always
associated with normal or hyperemic CTP, whilst hypoxic lactate was
associated with oligemic CTP (Table 1). Methods Patients with a TBI admitted to a designated trauma center
in 2009/10 were screened retrospectively and included according
to the IMPACT study criteria. The predictive outcome was calculated
for included patients using the full IMPACT-TBI model. To assess
coagulopathy and extracranial injury we used the prothrombin time
percentage (PT), platelet count (109), and injury severity score (ISS). PT, platelets and ISS were added to the IMPACT model, creating
new models. The predictive power of the new models was tested by
comparing their area under the receiver operating characteristic curve
(AUC) with the original model. Figure 1 (abstract P323). g
Results Totally 342 were included. Six-month mortality was 32% and
unfavorable outcome 57%. The IMPACT model’s predictive power
for mortality and neurological outcome; AUC: 0.85, CI: 0.81 to 0.89;
and AUC: 0.81, CI: 0.76 to 0.86. P320
Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) Nonsurvivors and patients with an
unfavorable outcome had lower platelets (159; 165 vs. 191; 198), PT
(76; 77 vs. 86; 90) and higher ISS (50; 44% ISS >25 vs. 34; 32% ISS >25)
compared with survivors and favorable outcome (P <0.05). Addition of
PT but not ISS nor platelets resulted in a signifi cant improvement of the
IMPACT model’s predictive power for 6-month mortality; AUC: 0.87, CI:
0.81 to 0.89, P = 0.017 (Figure 1). In multivariate analysis PT remained
an independent signifi cant predictor of outcome when adjusting for
IMPACT prognosis and ISS (OR: 0.985, CI: 0.975 to 0.996). p
g
Conclusion Addition of PT to the IMPACT model improves the
predictive power for 6-month mortality but not neurological outcome
when adjusting for ISS. p
p
y
g
when adjusting for ISS. References
1. Steyerberg EW, et al.: PLoS Med 2008, 5:e165. 2. Marmarou A, et al.: J Neurotrauma 2007, 24:239-250. P323
Nonischemic endogenous lactate production in humans with
severe traumatic brain injury
N Sala1, T Suys1, JB Zerlauth1, J Bloch1, P Magistretti2, R Meuli1, M Oddo1
1Lausanne University Hospital, Lausanne, Switzerland; 2EPFL, Lausanne,
Switzerland
Critical Care 2013, 17(Suppl 2):P323 (doi: 10.1186/cc12261)
Introduction Evidence suggest that endogenous lactate, produced by
aerobic glycolysis, is an important substrate for neurons, particularly in
Figure 1 (abstract P322). P320
Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) Epidemiology of moderate and severe traumatic brain injury in
Cairo University Hospital in 2010
T Montaser1, A Hassan2
1Shobra General Hospital, Cairo, Egypt; 2King Khaled University Hospital,
Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) T Montaser1, A Hassan2 y
Critical Care 2013, 17(Suppl 2):P320 (doi: 10.1186/cc12258) Introduction Traumatic brain injury (TBI) is a contributing factor
to approximately one-third of all injury-related deaths in the USA
annually. Updated statistical records for TBI in Egypt are lacking. The
current research is aiming to estimate the prevalence of TBI in Egypt in
order to develop a comprehensive TBI prevention program. p
p
p
p
g
Methods A 1-year period (one calendar month every quarter of
2010) descriptive epidemiological study of moderate and severe TBI
cases admitted to the emergency department, Cairo main university
hospital. The data collection sheet included personal data (age, sex and
residency), incident-related data (cause, nature and time of injury) and
both clinical and radiological fi ndings. y
Conclusion Heavily alcohol intoxicated TBI patients are commonly
initially transported to a non-neurosurgical trauma center and this may
worsen 6-month neurological outcome. References 1. Hartl R, et al.: J Trauma 2006, 60:1250-1256. 1. Hartl R, et al.: J Trauma 2006, 60:1250-1256. 2. Shahin H, et al.: J Trauma 2010, 69:1176-1181. 2. Shahin H, et al.: J Trauma 2010, 69:1176-1181. Table 1 (abstract P320). Causes of TBI in diff erent age groups in Egypt
NO = 844/
Abuse/
Violence/
Struck by/
Other causes/
Age (years)
4 months
FFH
MVC
neglect
assaults
against
not known
1 to 5
54 (6%)
20 (37%)
10 (19%)
8 (15%)
–
8 (15%)
8 (14%)
6 to 18
91 (11%)
28 (31%)
20 (22%)
5 (6%)
6 (5 %)
22 (25%)
10 (11%)
19 to 29
150 (18%)
49 (33%)
45 (30%)
–
14 (9%)
18 (12%)
24 (16%)
30 to 55
381 (45%)
114 (30%)
134 (35%)
–
19 (5%)
50 (13%)
64 (17%)
Above 55
168 (20%)
47 (28%)
41 (24%)
11 (7%)
5 (3%)
25 (15%)
39 (23%) Table 1 (abstract P320). Causes of TBI in diff erent age groups in Egypt S122 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P322 conditions of increased energy demand. This study aimed to examine
brain lactate metabolism in patients with severe traumatic brain injury
(STBI). j
References 1. Steyerberg EW, et al.: PLoS Med 2008, 5:e165. 1. Steyerberg EW, et al.: PLoS Med 2008, 5:e165. 2. Marmarou A, et al.: J Neurotrauma 2007, 24:239-250. 2. Marmarou A, et al.: J Neurotrauma 2007, 24:239-250. Figure 1 (abstract P322). Table 1 (abstract P323). Lactate elevations and brain perfusion (
)
p
Oligemia (%)
Normal (%)
Hyperemia (%)
HG lactate
0
58
42
HX lactate
100
0
0
HG, hyperglycolytic; HX, hypoxic. Conclusion Our fi ndings suggest predominant nonischemic lactate
release after TBI and identify, for the fi rst time, an association between
cerebral hyperglycolytic lactate production and normal to supranormal
brain perfusion. Our data support the concept that lactate may be used
as energy substrate by the injured human brain. gy
y
j
Acknowledgement Supported by the Swiss National Science
Foundation. Figure 1 (abstract P322). P323 P323
Nonischemic endogenous lactate production in humans with
severe traumatic brain injury
N Sala1, T Suys1, JB Zerlauth1, J Bloch1, P Magistretti2, R Meuli1, M Oddo1
1Lausanne University Hospital, Lausanne, Switzerland; 2EPFL, Lausanne,
Switzerland
Critical Care 2013, 17(Suppl 2):P323 (doi: 10.1186/cc12261) Nonischemic endogenous lactate production in humans with
severe traumatic brain injury Yagi, N Saito, Y Hara, H Hisashi Matumoto, K Mashiko Introduction In the prehospital setting, it is diffi cult to use the
Glasgow Coma Scale (GCS) to evaluate the consciousness state using
in pediatric patients with severe trauma. The Japan Coma Scale (JCS) is
a consciousness scale used widely in Japan and, with its four grades, is
simpler and quicker to use than the GCS. This study examined whether Introduction In the prehospital setting, it is diffi cult to use the
Glasgow Coma Scale (GCS) to evaluate the consciousness state using
in pediatric patients with severe trauma. The Japan Coma Scale (JCS) is
a consciousness scale used widely in Japan and, with its four grades, is
simpler and quicker to use than the GCS. This study examined whether Introduction Evidence suggest that endogenous lactate, produced by
aerobic glycolysis, is an important substrate for neurons, particularly in S123 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods In this prospective observational study, measurements of
OSND were carried out in 21 patients of a polyvalent 10-bed adult
ICU before and 2 minutes after changes of PEEP from a baseline of 5
with 1 cmH2O increments, to a maximum of 12. Two measurements
were performed with a 7.5 MHz probe (MicroMaxx; Fujifi lm Sonosite
Inc., USA) in each eye before and after PEEP changes. Demographic
data (APACHE II score, age, sex, diagnosis) Ppeak and Pm were also
recorded. Exclusion criteria were intracranial pathology, ocular neuritis
and trauma. Normality with P–P plots and the Kolmogorov–Smirnov
test were calculated followed by bivariate and single linear regression
analysis (IBM SPPS v17, signifi cance P <0.05). the JCS can predict clinical traumatic brain injury (TBI) and outcome in
pediatric patients aged 3 to 15 years in the prehospital setting. Methods This retrospective study analyzed data from the Japan Trauma
Data Bank. P323 Registered pediatric patients aged 3 to 15 years with severe
trauma (maximum Abbreviated Injury Scale score ≥3 or Injury Severity
Score ≥9) were divided into four groups according to JCS score in the
prehospital setting (Grade 0: alert, Grade 1: possible eye-opening, not
lucid, Grade 2: possible eye-opening upon stimulation, Grade 3: no
eye-opening and coma). TBI was defi ned as maximum head AIS ≥3. y
gi
Results A total of 1,562 patients were included (Grade 0: 673, Grade 1:
410, Grade 2: 230, Grade 3: 249). Victims of blunt trauma accounted for
98.1%. Median age was 9 (interquartile range: 7 to 12) years, median
ISS was 16 (9 to 21). There was strong agreement between the JCS in
prehospital setting and GCS scores on arrival at hospital (r = –0.745,
P <0.001). Multivariate analysis adjusted for age and ISS revealed that
the JCS was independently associated with TBI (odds ratio (OR): 2.5;
95% CI: 2.1 to 2.8, P <0.001) and hospital mortality (OR: 3.8; 95% CI 2.4
to 6.0, P <0.001). See Table 1. y
gi
Results Seventy-two measurements were included for further analysis. Mean ± SD values were: PEEP (7.9 ± 2.3), Ppeak (24.35 ± 5), Pm
(12.03 ± 3.16), OSNDright (4.03 ± 0.79), OSNDleft (4.1 ± 0.79), ΔOSND
(0.24 ± 0.2), while values for age and APACHE II score were 62 ± 4 years
and 14.4 ± 1.4 respectively. PEEP and OSND seem to have a moderate
relation (tendency equation: OSND = 2.454 + 0.581×PEEP) compared
with the weaker eff ect of Ppeak and Pm (Table 1). Table 1 (abstract P325). Correlations (r2) between airway pressures and OSND
OSNDright
OSNDleft
PEEP
0.581*
0.571*
Ppeak
0.467*
0.448*
Pm
0.491**
0.467**
*P <0.01, **P <0.05. Table 1 (abstract P324)
TBI, n (%)
Mortality (%)
JCS grade 0
213 (31.6)
0.1
JCS grade 1
225 (62.2)
0.7
JCS grade 2
184 (80.0)
2.6
JCS grade 3
230 (92.4)
15.7 Table 1 (abstract P325). Correlations (r2) between airway pressures and OSND Conclusion Our study identifi ed a moderate relation between PEEP
and OSND and a weaker one between Ppeak, Pm and OSND. Thus,
in selected cases OSND could serve as a bedside marker of eff ect of
airway pressure to ICP. Yet, larger studies are needed to come to a safer
conclusion. Conclusion There was strong association between JCS score and
clinical outcome in pediatric patients with severe trauma. P325f 1. Geeraerts T, et al.: Int Care Med 2008, 34:2062-2067. P325
Eff ect of positive end-expiratory pressure on ultrasound
measurement of optical sheath nerve diameter: preliminary study
T Aslanidis, E Anastasiou, E Geka, E Efthimiou, A Myrou, A Kontos,
D Papadopoulos, E Boultoukas, M Giannakou-Peftoulidou
A.H.E.P.A. University Hospital, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P325 (doi: 10.1186/cc12263) P325
Eff ect of positive end-expiratory pressure on ultrasound
measurement of optical sheath nerve diameter: preliminary study
T Aslanidis, E Anastasiou, E Geka, E Efthimiou, A Myrou, A Kontos,
D Papadopoulos, E Boultoukas, M Giannakou-Peftoulidou
A.H.E.P.A. University Hospital, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P325 (doi: 10.1186/cc12263) P326 P326
National survey of current protocols and management of the
traumatic brain injury patients in UK ICUs
B Lewinsohn, S Panchatsharam, S Wijayatilake, A Billini, G Delacedra,
R Jain, J Khan, R Shetty, A Lewinsohn
Queens Hospital, Romford, UK
Critical Care 2013, 17(Suppl 2):P326 (doi: 10.1186/cc12264) P323 The results
support the use of the JCS in the prehospital transport destination
criteria for children. Reference Reference National survey of current protocols and management of the
traumatic brain injury patients in UK ICUs j
y p
B Lewinsohn, S Panchatsharam, S Wijayatilake, A Billini, G Delaced
R Jain, J Khan, R Shetty, A Lewinsohn
Queens Hospital, Romford, UK
Critical Care 2013, 17(Suppl 2):P326 (doi: 10.1186/cc12264) Introduction Non-invasive ocular ultrasonography has recently been
proposed to detect elevated intracranial pressure (ICP). On the other
hand, the eff ect of positive end-expiratory pressure (PEEP) on ICP is
well documented. The aim of the present ongoing study is to record the
eff ect of changes of airway pressures on optical sheath nerve diameter
(OSND). Introduction Following primary neurological insult, initial manage ment
of traumatic brain-injured (TBI) patients has a clearly defi ned pathway
[1]. However, after arrival at tertiary centers, further manage ment is not Figure 1 (abstract P326). Figure 1 (abstract P326). S124 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 computed for longer periods. Signifi cantly better outcome (Table 1)
was observed in patients for whom optimal CPP derived from LAx was
maintained. Table 1 (abstract P327)
Nonsurvivors
Survivors
P value
Time within CPPopt-%
18.3 (15.5; 24.4)
24.8 (19.7; 28.7)
0.0004
|CPP-CPPopt| (mmHg)
7.05 (4.98; 9.73)
5.37 (4.49; 6.82)
0.0024 Table 1 (abstract P327) standardized. Intracranial hypertension (ICH), systemic hypotension,
hypoxia, hyperpyrexia and hypocapnia have all been shown to
independently increase mortality [2]. Despite numerous studies, there
is currently no level 1 evidence to support any specifi c management
[3]. Our objective was to provide an overview of the current clinical
management protocols in the UK. g
p
Methods Thirty-one ICUs managing patients with severe TBI were
identifi ed from the RAIN (Risk Adjustment In Neurocritical care) study,
and a telephone survey was conducted. computed for longer periods. Signifi cantly better outcome (Table 1)
was observed in patients for whom optimal CPP derived from LAx was
maintained. Results A total 97% of units used a cerebral perfusion pressure
protocol for the initial management, with 83% targeting pressures of
60 to 70 mmHg and 17% aimed for >70 mmHg. Ninety-one percent of
units monitored CO2 routinely with 61% targeting CO2 of 4.5 to 5 kPa
(Figure 1). Regarding osmotherapy, mannitol was still the preferred
agent, with 48% of units using it as fi rst line; 32% used hypertonic saline,
while 20% of units used either depending on clinicians’ preference. g
References 1. NICE: Head Injury; Triage, Assessment, Investigation and Early Management of
Head Injury. London: National Collaborating Centre for Acute Care;
September 2007. g
Methods A retrospective analysis of rcSO2 utility in AMS. rcSO2 data were
collected every 30 seconds for AMS patients who had a head CT. Patients
with a negative head CT were compared with those with an abnormal
head CT. ROC analysis was performed to fi nd the AUC for each summary
statistic and performance characteristics. Subgroup analysis was done to
determine whether rcSO2 predicted injury and location. 2. Wijayatilake et al.: Updates in the management of intracranial pressure in
traumatic brain injury. Curr Opin Anaesthesiol 2012, 25:540-547. 3. Guidelines for the Management of Severe Traumatic Brain Injury. 3rd edition. New York: Brain Trauma Foundation; 2007. 3. Guidelines for the Management of Severe Traumatic Brain Injury. 3rd edition. New York: Brain Trauma Foundation; 2007. c
2 p
j
y
Results rcSO2 readings across 5, 15, 20, and 30 minutes were stable
(Figure 1). rcSO2 readings with one or both sides <50% or a wide
diff erence between L and R cerebrum was predictive of an abnormal
CT scan. A mean diff erence of 4.2 was 82% sensitive for detecting a CT
lesion with 62% specifi city, 88% PPV, and 52% NPV; a mean diff erence
of 12.2 was 100% specifi c for an abnormal head CT. Lower mean rcSO2
readings localized to the CT pathology side, and higher rcSO2 readings
trend toward the EDH group. 4. Barochia et al.: Bundled care for septic shock. Crit Care Med 2010, 38:668-678. 4. Barochia et al.: Bundled care for septic shock. Crit Care Med 2010, 38:668-678. 4. Barochia et al.: Bundled care for septic shock. Crit Care Med 2010, 38:668-678. National survey of current protocols and management of the
traumatic brain injury patients in UK ICUs Sixteen percent questioned were currently enrolled on the Eurotherm
hypothermia trial, while 16% never used hypothermia and one unit
used prophylactic hypothermia routinely. The remaining 65% of units
used hypothermia only to manage refractory ICH. P327
f Low-frequency autoregulation index for calculation of optimal
cerebral perfusion pressure in severe traumatic brain injury
F Guiza1, G Meyfroidt1, M Schuhmann2, G Van den Berghe1, I Piper3,
B Depreitere1
1University Hospitals Leuven, Belgium; 2University Hospital Tübingen,
Germany; 3Southern General Hospital, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P327 (doi: 10.1186/cc12265) Cerebral rSO2 monitoring in pediatric altered mental status patients
T Abramo, I Kane
Vanderbilt School of Medicine, Nashville, TN, USA
Critical Care 2013, 17(Suppl 2):P328 (doi: 10.1186/cc12266) Vanderbilt School of Medicine, Nashville, TN, USA Critical Care 2013, 17(Suppl 2):P328 (doi: 10.1186/cc12266) Conclusion There is no clear consensus on the initial targets used. The
surviving sepsis campaign showed that protocol-led care can reduce
mortality [4]. Perhaps it is time for a similar approach to be adopted,
with specialists coming to together to review the evidence and
formulate guidelines that can then be tested. Introduction Pediatric patients with altered mental status (AMS)
present with poor histories resulting in delayed testing and potential
poor outcomes. Non-invasive detection for altered cerebral physiology
related to TBI would improve resuscitation and outcome. Cerebral
rSO2 (rcSO2) studies demonstrate its utility in certain neurological
emergencies. References References
1. Aries M, et al.: Crit Care Med 2012, 40:2456-2463. 2. Piper I, et al.: Acta Neurochirurgica 2003, 145:615-628. 1. Aries M, et al.: Crit Care Med 2012, 40:2456-2463. 2. Piper I, et al.: Acta Neurochirurgica 2003, 145:615-628. 1. Aries M, et al.: Crit Care Med 2012, 40:2456-2463. 1. Aries M, et al.: Crit Care Med 2012, 40:2456 2463. 2. Piper I, et al.: Acta Neurochirurgica 2003, 145:615-628. Low-frequency autoregulation index for calculation of optimal
cerebral perfusion pressure in severe traumatic brain injury
F Guiza1, G Meyfroidt1, M Schuhmann2, G Van den Berghe1, I Piper3,
B Depreitere1 Conclusion Cerebral rcSO2 monitoring can non-invasively detect
altered cerebral physiology and pathology related to TBI as the cause
for pediatric altered mental status. The utility of rcSO2 monitoring has
shown its potential for localizing and characterizing intracranial lesions
among these altered children. Further studies utilizing rcSO2 monitoring
as an adjunct tool in pediatric altered mental status evaluation and
management are ongoing. Introduction Traumatic brain injury (TBI) is a major cause of permanent
disability and death in young patients. Controversy exists regarding the
optimal cerebral perfusion pressure (CPP) required in TBI management. A tool for monitoring autoregulation and determining an optimal
CPP is the pressure reactivity index (PRx), defi ned as a moving
correlation coeffi cient between the mean arterial blood pressure
(MAP) and intracranial pressure (ICP) at a frequency of at least 60 Hz. This requirement of high frequency has constrained its use to a few
academic centers. An association was shown between outcome and
continuous optimal CPP based on 4 hours of PRx [1]. We present a
novel low-frequency autoregulation index (LAx), based on correlations
between ICP and MAP at a standard minute-by-minute time resolution. Methods A total of 182 patients from the Brain-IT [2] multicentre
European database had registered outcome and ICP and MAP for
the fi rst 48 ICU hours. Twenty-one TBI patients admitted to the
university hospitals of Leuven, Belgium and Tubingen, Germany were
continuously monitored using ICM+ software (Cambridge Enterprise)
allowing for continuous PRx calculation. Autoregulation indices versus
CPP plots for PRx and LAx were computed to determine optimal CPP
every minute during the fi rst 48 ICU hours [1]. Intravenous paracetamol for fever control in acute brain-injured
patients: cerebral and hemodynamic eff ects E Picetti1, I Rossi1, P Ceccarelli1, S Risolo1, P Schiavi2, V Donelli1, A Crocamo1,
M Antonini1, M Caspani1
1I Servizio Anestesia e Rianimazione, Azienda Ospedaliero-Universitaria di
Parma, Italy; 2Neurochirurgia e Neurotraumatologia, Azienda Ospedaliero-
Universitaria di Parma, Italy
Critical Care 2013, 17(Suppl 2):P329 (doi: 10.1186/cc12267) Introduction Fever is a dangerous secondary insult for the injured
brain [1]. We investigated the cerebral and hemodynamic eff ects of
intravenous (i.v.) paracetamol administration for the control of fever in
neurointensive care unit (NICU) patients. Methods The i.v. paracetamol (1 g in 15 minutes) was administered to
NICU patients with a body temperature (Temp.) >37.5°C. Its eff ects on
mean arterial pressure (MAP), heart rate (HR), intracranial pressure (ICP),
cerebral perfusion pressure (CPP), jugular venous oxygen saturation
(SjVO2) and Temp. were recorded at the start of paracetamol infusion
(T0) and after 30 (T30), 60 (T60) and 120 (T120) minutes. Interventions
for the maintenance of CPP >60 mmHg or ICP <20 mmHg were
recorded. i
Results On the Brain-IT database, LAx resulted in an optimal CPP for
90% of the fi rst 48 hours. Table 1 shows recommendations with respect
to outcome. In the Leuven–Tübingen database, PRx and LAx resulted in
44% and 92% recommendations respectively. The average diff erence
between methods was 5.26 mmHg.f Conclusion The diff erences in optimal CPPs derived from PRx and LAx
were not clinically signifi cant. LAx allowed for recommendations to be Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S125 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Results Fifteen NICU patients (nine subarachnoid hemorrhage, fi ve
traumatic brain injury, mean age 54.9 ± 16.8, seven (50%) males, median
GCS 7) were prospectively studied. We analyzed the administration
of one dose of paracetamol for each patient (total 14 cases). After
infusion of paracetamol we found a decrease of Temp. (from 37.8 ± 0.3
to 37.4 ± 0.4°C, P <0.001), MAP (from 94.7 ± 9.9 to 86.1 ± 6.7 mmHg,
P = 0.008), CPP (from 79.6 ± 13.1 to 70.8 ± 7.6 mmHg, P = 0.011) and
HR (from 71.5 ± 14.9 to 63.8 ± 16.3 bpm, P <0.001) with respect to the
starting value (ANOVA for repeated measures), whereas ICP and SjVO2
remained unchanged (Figure 1). In fi ve cases norepinephrine infusion
was started for CPP <60 mmHg. In another two cases, for the same
reason, the norepinephrine dosage was augmented. 1.
Greer DM, Funk SE, Reaven NL, Ouzounelli M, Uman GC: Impact of fever on
outcome in patients with stroke and neurologic injury. Stroke 2008,
39:3029-3035. Intravenous paracetamol for fever control in acute brain-injured
patients: cerebral and hemodynamic eff ects We evaluated sociodemographic and clinical factors
associated with readmissions using a logistic regression model.i P330
Retrospective analysis of the hemodynamic eff ects of induction of
barbiturate coma in patients with refractory elevated intracranial
pressure
S D’Hollander1, M Vander Laenen1, JB Gillet2, C De Deyne1, R Heylen1,
W Boer1, F Jans1
1ZOL Genk, Belgium; 2UZ Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P330 (doi: 10.1186/cc12268) g
g
g
Results Among 18,342 adult patients with TBI identifi ed in the registry,
14,777 patients were included among which 2,363 had severe, 1,106
moderate and 11,308 mild traumatic brain injury. Most patients were
young (mean age: 52 ± 23 years) and had no comorbidity (73.6%). Overall, 1,032 patients (7.0%) were readmitted within 30 days, 12.7%
within 3 months and 17.6% within 6 months. At 30 days post discharge,
311 (30.1%) were readmitted for a complication. The median length of
stay was 8 days (Q1 to Q3: 3 to 20). More than 10% of patients aged
≥75 years with ≥1 comorbidity or with ≥1 admission prior to index
hospitalisation were readmitted. The severity of the TBI was not an
independent predictor of readmission. Age, highest AIS, number of
comorbidities, number of admissions prior to index hospitalization,
level of index trauma center and discharge destination were associated
with readmissions on multivariate analysis. Introduction Standard clinical practice in patients with elevated
intracranial pressure (ICP) is to keep cerebral perfusion pressure (CPP)
at (or above) 60 mmHg: we performed this retrospective study to
analyze whether induction of barbiturate coma impedes compliance
to this target CPP value. It has been repetitively shown that, although
proven as an effi cient therapy for refractory elevated ICP, barbiturate
coma may cause a decrease in mean arterial pressure (MAP) and CPP
[1]. Methods All patients that had received sodium thiopental in our ICU
during 2011 were identifi ed and their medical and nursing records
were retrospectively analyzed. The eff ect of administration of sodium
thiopental (loading dose 3 to 5 mg/kg, followed by a continuous
infusion adjusted to obtain an EEG burst suppression ratio between 40
and 80%) on MAP, ICP and CPP was evaluated. Values are reported as
mean ± SD. The changes over time in ICP, MAP and CPP are reported as
means with their 95% CI. y
Conclusion Readmissions in the months following TBI are frequent, but
were not found to be associated with the TBI severity. Intravenous paracetamol for fever control in acute brain-injured
patients: cerebral and hemodynamic eff ects The proportion of
patients who had infusion of norepinephrine increased from 42.8% at
T0 to 78.6% at T120 (P = 0.02, chi-square for trends). Conclusion Use of i.v. paracetamol is eff ective in the maintenance
of normothermia in acute brain-injured patients. However, adverse
hemodynamic eff ects, which could represent a secondary insult for the
injured brain, must be rapidly recognized and treated. Reference
1. Greer DM, Funk SE, Reaven NL, Ouzounelli M, Uman GC: Impact of fever on
outcome in patients with stroke and neurologic injury. Stroke 2008,
39:3029-3035. Figure 1 (abstract P328). Figure 1 (abstract P329). *P <0.05 versus T0. Figure 1 (abstract P328). Figure 1 (abstract P329). *P <0.05 versus T0. reason, the norepinephrine dosage was augmented. The proportion of
patients who had infusion of norepinephrine increased from 42.8% at
T0 to 78.6% at T120 (P = 0.02, chi-square for trends).f Results Fifteen NICU patients (nine subarachnoid hemorrhage, fi ve
traumatic brain injury, mean age 54.9 ± 16.8, seven (50%) males, median
GCS 7) were prospectively studied. We analyzed the administration
of one dose of paracetamol for each patient (total 14 cases). After
infusion of paracetamol we found a decrease of Temp. (from 37.8 ± 0.3
to 37.4 ± 0.4°C, P <0.001), MAP (from 94.7 ± 9.9 to 86.1 ± 6.7 mmHg,
P = 0.008), CPP (from 79.6 ± 13.1 to 70.8 ± 7.6 mmHg, P = 0.011) and
HR (from 71.5 ± 14.9 to 63.8 ± 16.3 bpm, P <0.001) with respect to the
starting value (ANOVA for repeated measures), whereas ICP and SjVO2
remained unchanged (Figure 1). In fi ve cases norepinephrine infusion
was started for CPP <60 mmHg. In another two cases, for the same q
Conclusion Use of i.v. paracetamol is eff ective in the maintenance
of normothermia in acute brain-injured patients. However, adverse
hemodynamic eff ects, which could represent a secondary insult for the
injured brain, must be rapidly recognized and treated. Reference S126 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P330 admission, and readmissions in the 12 following months. Primary
outcome was unplanned readmission 30 days, 3 months and 6 months
post discharge. Intravenous paracetamol for fever control in acute brain-injured
patients: cerebral and hemodynamic eff ects Further studies
evaluating reasons for readmission are warranted in order to develop
strategies to prevent such events. Results In 2011, a total of 20 patients were treated in our ICU with
barbiturate coma for refractory elevated ICP. In fi ve patients, systemic
hypothermia was simultaneously induced. These patients were
excluded from further analysis. Six patients died during their stay in the
ICU and nine patients could be discharged to the neurosurgical ward. The mean peak ICP value before induction of barbiturate coma was
26 ± 3 mmHg, and the ICP value 6 hours later was 20 ± 6 mmHg. The
MAP at those same time points was 91 ± 12 mmHg and 81 ± 11 mmHg,
respectively. The CPP, calculated from both previous values, was
66 ± 13 mmHg and 61 ± 11 mmHg, respectively. At both time points,
the dose of noradrenaline was comparable (10.1 ± 9.6 μg/minute and
11.5 ± 13.2 μg/minute, respectively). Analysis of the change in ICP,
MAP and CPP over the 6-hour period showed a signifi cant decrease
in ICP of –5.8 mmHg (–9.3; –2.3) (mean, 95% CI). The MAP signifi cantly
decreased by –10.4 mmHg (–19.1; –1.7). The decrease in CPP was not
signifi cant (–4.6 mmHg (–14.2; 5.0)). P332 Risk factors of pituitary disorders following traumatic brain injury
F Lauzier1, O Lachance1, I Cote1, B Senay1, P Archambault1, F Lamontagne2
A Boutin1, M Shemilt1, L Moore1, F Bernard3, C Gagnon1, D Cook4,
AF Turgeon1
1Universite Laval, Quebec, Canada; 2Universite de Sherbrooke, Canada;
3Universite de Montreal, Canada; 4McMaster University, Hamilton, Canada
Critical Care 2013, 17(Suppl 2):P332 (doi: 10.1186/cc12270) Risk factors of pituitary disorders following traumatic brain injury
F Lauzier1, O Lachance1, I Cote1, B Senay1, P Archambault1, F Lamontagne2
A Boutin1, M Shemilt1, L Moore1, F Bernard3, C Gagnon1, D Cook4,
AF Turgeon1 g
1Universite Laval, Quebec, Canada; 2Universite de Sherbrooke, Canada;
3Universite de Montreal, Canada; 4McMaster University, Hamilton, Canada
Critical Care 2013, 17(Suppl 2):P332 (doi: 10.1186/cc12270) g
1Universite Laval, Quebec, Canada; 2Universite de Sherbrooke, Canada;
3Universite de Montreal, Canada; 4McMaster University, Hamilton, Canada
Critical Care 2013, 17(Suppl 2):P332 (doi: 10.1186/cc12270) Introduction Pituitary disorders following traumatic brain injury
(TBI) are frequent, but their determinants are poorly understood. We performed a systematic review to assess the risk factors of TBI-
associated pituitary disorders. p
y
Methods We searched MEDLINE, Embase, Scopus, The Cochrane
Library, BIOSIS, and Trip Database, and references of narrative reviews
for cohort, cross-sectional and case–control studies enrolling at least
fi ve adults with TBI in whom ≥1 pituitary axis was tested and one
potential predictor reported. Two independent investigators selected
citations, extracted data and assessed the risk of bias. We pooled the
data from all studies assessing a specifi c predictor, regardless of the
pituitary axis being evaluated. When more than one pituitary axis was
assessed, we used the data related to hypopituitarism or the data from
the most defective axis. When a pituitary axis was evaluated several
times, we used assessment farthest from the injury. A meta-analysis
was performed using random eff ect models and I2 was used to evaluate
heterogeneity. Studies were considered at low risk of bias if the authors
defi ned inclusion/exclusion criteria, did not use voluntary sampling,
and tested > 90% of patients with proper detailed diagnostic criteria. i
Conclusion This retrospective study indicates that inducing a thiopental
coma in patients with refractory elevated ICP eff ectively reduces ICP. Although the concomitant reduction in MAP was signifi cant, the
resulting decrease in CPP was small and nonsignifi cant. Moreover, the
target CPP of 60 mmHg was maintained after induction of barbiturate
coma. Reference 1. 1.
Haddad et al.: Scand J Trauma Resusc Emerg Med 2012, 20:12-27. P332 Haddad et al.: Scand J Trauma Resusc Emerg Med 2012, 20:12-27. P331
H
i S127 Critical Care 2013, Volume 17 Suppl 2
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Vasopressin reduces the cumulative epinephrine dose in
hypothermic swine with traumatic brain injury
AD Goldberg, MG Kashiouris, L Keenan, A Rabinstein, M Kumar
Mayo Clinic, Rochester, MN, USA
Critical Care 2013, 17(Suppl 2):P333 (doi: 10.1186/cc12271) Introduction Severe traumatic brain injury (TBI) is a major cause of death
and of severe neurologic sequelae. Long-term functional outcome
of TBI and its best timing of assessment are not well understood,
and may be evaluated too prematurely in clinical studies because of
resources required to do so without too much missing data. Hence,
we conducted a systematic review of studies in severe TBI patients
to evaluate the long-term functional outcome. We hypothesized that
functional impact measured by the Glasgow Outcome Scale (GOS),
or the extended version (GOSe), may plateau after several months in
patients with severe TBI. Introduction Severe traumatic brain injury (TBI) is a major cause of death
and of severe neurologic sequelae. Long-term functional outcome
of TBI and its best timing of assessment are not well understood,
and may be evaluated too prematurely in clinical studies because of
resources required to do so without too much missing data. Hence,
we conducted a systematic review of studies in severe TBI patients
to evaluate the long-term functional outcome. We hypothesized that
functional impact measured by the Glasgow Outcome Scale (GOS),
or the extended version (GOSe), may plateau after several months in
patients with severe TBI. Introduction Prevention of secondary neurologic injuries is paramount
for improved neurologic outcomes after traumatic brain injury (TBI). Evidence suggests that although therapeutic hypothermia (TH) lowers
intracranial pressure and attenuates secondary cerebral insults after TBI
[1], it also induces hypotension. Brief episodes of mild hypotension in
brain-injured patients can trigger secondary injuries, which have been
associated with increased mortality in patients with TBI [2]. Vasopressin
mitigates hypotension in septic shock and improves coronary perfusion
in hypothermic cardiac arrest models [3]. We hypothesized that a low-
dose vasopressin infusion may reduce the cumulative epinephrine
dose in hypothermic, brain-injured swine. Methods We performed a systematic review of randomized controlled
trials and cohort studies (prospective and retrospective) in patients
with severe TBI. We searched MEDLINE, Embase, Cochrane Central,
BIOSIS, CINAHL and Trip Database from their inception to December
2011. References of included studies were searched for additional
studies. P331
H
i P331
Hospital readmissions following traumatic brain injury
A Boutin, K Francisque, L Moore, F Lauzier, X Neveu, A Turgeon
Université Laval, Québec, Canada
Critical Care 2013, 17(Suppl 2):P331 (doi: 10.1186/cc12269) p
p
p
g
Results Among 13,559 citations, we included 26 studies (1,708 patients). Increased age was associated with pituitary disorders (weighted mean
diff erence = 3.2, 95% CI = 0.3 to 6.1, 19 studies, 1,057 patients, I2 = 59%). This fi nding was no longer signifi cant when only considering studies
with low risk of bias. TBI severity was associated with an increased risk
of developing pituitary disorders (RR = 1.49, 1.24 to 1.77, I2 = 17% for
moderate/severe vs. mild TBI; RR = 1.78, 1.09 to 2.91, I2 = 80% for severe
vs. mild/moderate TBI), while sex was not (RR for male = 1.05, 0.98 to
1.13, 15 studies, 870 patients, I2 = 0%). Among CT scan fi ndings, only
skull fractures were associated with pituitary disorders (RR = 1.75, 1.13
to 2.70, six studies, 345 patients, I2 = 47%). An insuffi cient number of
studies with low risk of bias assessing the association between GCS,
CT scan fi ndings and pituitary disorders was retrieved to perform
meta-analysis. Introduction Evaluating resource utilization is paramount in critically
ill patients with traumatic brain injury (TBI), but little is known
on readmissions after hospital discharge. We evaluated rates and
determinants of unplanned readmission following TBI. Methods We conducted a multicenter retrospective cohort study from
April 1998 to March 2009. Data were obtained from a Canadian provincial
trauma system, based on mandatory contribution from 59 trauma
centres, and a hospital discharge database. Patients aged ≥16 years
with TBI (ICD-9 or ICD-10 codes of 850-854 and S06, respectively) were
included. Patients who died during the index hospitalization, who
lived outside the province, who could not be linked with the hospital
discharge database were excluded. We collected baseline and trauma
characteristics, hospital admissions in the 12 months preceding index Conclusion Age, TBI severity and skull fractures are associated with an
increased risk of pituitary disorders. Further studies are necessary to
identify additional factors that will help developing targeted screening
strategies. References 1. Polderman KH, et al.: Intensive Care Med 2002, 28:1563-1573. 2. Fearnside MR, et al.: Br J Neurosurg 1993, 7:267-279. 2. Fearnside MR, et al.: Br J Neurosurg 199 Results All 31 (100%) units used ICP monitoring and parenchymal ICP
bolt was the most widely used method (77%). Thirty (96.7%) units used
a cerebral perfusion pressure protocol to guide therapy. Twenty-eight
(90.3%) units used continuous capnography. Twelve (38.7%) units
used transcranial Doppler. Eight (25.8%) units used partial pressure of
oxygen in brain tissue. Four (12.9%) units used microdialysis. Only one
unit used jugular bulb oximetry. None of the units used near-infrared
spectrometry or optic nerve sheath diameter. Eight units (25.8%) used
a cerebral function monitor and seven (22.5%) units used the bispectral
index for guiding depth of sedation. 3. Voelckel WG, et al.: Anesth Analg 2000, 91:627-634. Monitoring of severe traumatic brain injury patients in UK ICUs:
a national survey Monitoring of severe traumatic brain injury patients in UK ICUs:
a national survey
S Panchatsharam, B Lewinsohn, G De La Cerda, D Wijayatilake
Queen’s Hospital, Romford, UK
Critical Care 2013, 17(Suppl 2):P335 (doi: 10.1186/cc12273) S Panchatsharam, B Lewinsohn, G De La Cerda, D Wijayatilake
Queen’s Hospital, Romford, UK Introduction Prevention of secondary brain injury is the cornerstone
in the management of patients with severe traumatic brain injury (TBI)
and raised intracranial pressure (ICP). Although a variety of monitoring
methods are available, due to lack of strong evidence their use varies
considerably [1]. The objective of this survey was to provide an
overview of the current practice in monitoring of patients with severe
TBI in all neuro-ICUs across the UK. Results The median cumulative epinephrine dose in the animals that
received the vasopressin infusion was 715 mg with a 25th to 75th
interquartile range (IQR) of 320 to 930 mg. The median cumulative
epinephrine dose in the control group was 2,044 mg (IQR 1,640 to
2,344 mg). This was statistically signifi cant (P = 0.003), based on the
Wilcoxon rank-sum test. Conclusion A low-dose infusion of vasopressin can signifi cantly
reduce vasopressor requirements and improves hemodynamics in
hypothermic, brain-injured swine. This hemodynamic stability may
improve neurological outcomes. Methods The ICUs managing adult patients with severe TBI were
identifi ed from the RAIN (Risk Adjustment In Neurocritical care) study
sites, and the Society of British Neurosurgeons. Thirty-one centers were
identifi ed, and a telephonic survey was conducted by the investigators. Data were collected using a single-page questionnaire containing 18
questions. P331
H
i Two reviewers independently determined study eligibility and
collected data. The primary outcome measure was the proportion of
unfavourable functional outcome (GOS 1 to 3 or GOSe 1 to 4) at 6 to
12 months, 12 to 18 months, 18 to 24 months and more than 24 months
after severe TBI. We calculated Freeman Tukey-type arcsine square-
root transformations and pooled data using random-eff ect models. Heterogeneity was assessed with the I2 test and sensitivity analyses
were based on a priori hypotheses. Methods Six domestic cross-bred pigs were subjected to epinephrine
infusion after general anesthesia, standardized TBI and transpulmonary
hypothermia (32°C for 48 hours). All animals received the same care,
aiming for a mean arterial pressure >60 mmHg. At hour 24, animals
received additional vasopressin infusion at 0.04 units/minute. We
measured the cumulative epinephrine dose for each animal pre
and post vasopressin infusion (Figure 1) and performed a two-
sample Wilcoxon rank-sum test, comparing the median cumulative
epinephrine doses in the two groups. Results In total, 4,432 studies were assessed for eligibility; 209 studies
(n = 31,540) were included. In the 188 studies using the GOS, a poor
functional outcome was observed in 56.6% (95% CI = 54.0 to 59.1%,
I2 = 91%), 51.9% (95% CI = 38.0 to 59.0%, I2 = 84%), 57.0% (95% CI = 48.2
to 55.5%, I2 = 93%) and 56.9% (95% CI = 48.2 to 65.1%, I2 = 93%) of
patients at 6 to 12 months, 12 to 18 months, 18 to 24 months and
beyond 24 months, respectively. In the 18 studies using GOSe, a
poor functional outcome was observed in 62.9% of patients at 6
to 12 months (95% CI = 55.9 to 69.2%, I2 = 90%) and 54.6% at 12 to
18 months (95% CI = 43.2 to 65.8%, I2 = 90%). Heterogeneity was
present in most analyses and was not entirely explained by the planned
sensitivity analyses. Figure 1 (abstract P333). Eff ect of vasopressin on cumulative amount of
epinephrine administered. Conclusion Considering that the incidence of patients with an
unfavourable outcome remained constant at diff erent assessments,
a follow-up of severe TBI patients longer than 12 months does not
provide incremental information. Functional outcomes measured
longer than 12 months after the injury may not be warranted in clinical
studies. Survival analysis after head injury: is a normal INR reassuring?
K Leclerc-Gagne
Hôpital Sacré-Coeur de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P336 (doi: 10.1186/cc12274) Critical Care 2013, 17(Suppl 2):P337 (doi: 10.1186/cc12275) Introduction Intracranial bleeding after head injury is an important
issue in the emergency department (ED), and elevated INR increases
this risk [1]. A retrospective study showed an association between
diff erent levels of normal INRs (<1.6) and elevated risk of intracranial
bleeding, with a signifi cant elevation of this risk over an INR threshold
of 1.3 [2]. The objective of the study was to evaluate clinical impact
of an INR within the normal range in patients with head trauma. We
compared mortality between patients with INR <1.3 and those with INR
≥1.3 to <1.6. Introduction Traumatic brain injury (TBI) is a leading cause of death
and disability worldwide. Because mortality seriously underestimates
the impact of TBI, an outcome tool including vegetative state and
severe disability should be used. Our purpose was to study the long-
term outcome of a small cohort of severe TBI patients in Brazil. Methods This was a retrospective analysis of a prospectively collected
database. We included 34 consecutive adult patients admitted with
severe TBI to a tertiary private hospital in Rio de Janeiro, Brazil between
2009 and 2011. We analyzed data on demographics, admission, clinical
scores, imaging and complications, as well as hospital mortality and
1-year extended Glasgow Outcome Scale (eGOS) of all patients. Methods A post hoc analysis of prospective data collected from 3,356
patients seen in a tertiary-care ED from March 2008 to February 2011. We included patients aged 16 years old and over with an INR <1.6 and
a head CT performed within 24 hours of the trauma. We followed these
patients until December 2012, performed a chi-square test between
mortality of the two groups and calculated the hazard ratio (HR) from
survival analysis using Cox regression. y
Results We analyzed 33 patients with severe TBI. Mean age was 36 (± 20)
years and 79% were male. All patients were mechanically ventilated
and 70% underwent ICP monitoring. Overall hospital mortality was
36% and 60% had unfavorable outcome (eGOS <4) after 1 year. In
univariate analysis, higher APACHE II scores, the presence of midriasis
and intracranial hypertension were more frequent in nonsurvivors
compared with survivors. g
References 1. Lingsma HF, et al.: Early prognosis in traumatic brain injury: from
prophecies to predictions. Lancet Neurol 2010, 9:543-554. 1. Lingsma HF, et al.: Early prognosis in traumatic brain injury: from
h
i
di i
L
N
l 2010 9 543 554 1. Lingsma HF, et al.: Early prognosis in traumatic brain injury: from
prophecies to predictions Lancet Neurol 2010 9:543-554 1. Wijayatilake et al.: Updates in the management of intracranial pressure in
traumatic brain injury. Curr Opin Anaesthesiol 2012, 25:540-547. tilake et al.: Updates in the management of intracranial pressure in Wijayatilake et al.: Updates in the management of intracranial pres 1. Wijayatilake et al.: Updates in the management of intracranial pressure in
traumatic brain injury. Curr Opin Anaesthesiol 2012, 25:540-547. 2. Chauny JM, et al.: Non Elevated INR and Post Traumatic Intracranial Bleeding. Day of Research in Neurotraumatology. Montreal: Hôpital du Sacré-Cœur; 22
June 2012. 2. Chesnut et al.: A trial of intracranial-pressure monitoring in traumatic brain
injury. N Engl J Med 2012, 367:2471-2481. 2. Chesnut et al.: A trial of intracranial-pressure monitoring in traumatic brain
injury. N Engl J Med 2012, 367:2471-2481. P337 One-year extended Glasgow Outcome Scale and hospital mortality
predictors in patients with severe traumatic brain injury in Brazil
R Turon1, FR Ferreira1, D Prado1, P Kurtz2, M Damasceno1
1Hospital de Clínicas de Niteroi, Brazil; 2Clinica São Vicente, Rio de Janeiro,
Brazil
Critical Care 2013, 17(Suppl 2):P337 (doi: 10.1186/cc12275) P338 P338
Early respiratory deterioration predicts poor outcome after
aneurysmal subarachnoid hemorrhage
E Van Lummel, L Vergouw, M Van der Jagt
Erasmus MC, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P338 (doi: 10.1186/cc12276) One-year extended Glasgow Outcome Scale and hospital mortality
predictors in patients with severe traumatic brain injury in Brazil
R Turon1, FR Ferreira1, D Prado1, P Kurtz2, M Damasceno1 One-year extended Glasgow Outcome Scale and hospital mortality
predictors in patients with severe traumatic brain injury in Brazil
R Turon1, FR Ferreira1, D Prado1, P Kurtz2, M Damasceno1 Survival analysis after head injury: is a normal INR reassuring?
K Leclerc-Gagne
Hôpital Sacré-Coeur de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P336 (doi: 10.1186/cc12274) Furthermore, older age, higher APACHE II
score and worse GCS on admission were associated with unfavorable
outcome after 1 year, as measured by eGOS <4.i y
g
g
Results Patient mean age was 55.1 years (SD ±23), 65% were men and
mean follow-up duration was 3.1 years (SD ±0.8). A total of 115 patients
(15.9%) died during follow-up: 16 (36.4%) in the group with INR ≥1.3
and 99 (14.6%) in the group with INR <1.3 (P <0.001). Results showed a Figure 1 (abstract P336). Survival analysis of patients with INR <1.3 and
with INR ≥1.3. y
y
Conclusion Our descriptive results suggest a signifi cant burden of
severe TBI in a small cohort of young patients. Moreover, severity
of the primary disease, age and pupil refl exes, as well as intracranial
hypertension seemed to be associated with worse neurological
outcome. Long-term functional outcome in adults with severe TBI:
a meta-analysis y
M Asselin1, Y Lachance1, G Lalonde1, A Boutin1, M Shemilt1, L Moore1, F Lauzier1,
R Zarychanski2, P Archambault1, F Lamontagne1, D Fergusson3, A Turgeon1
1Université Laval, Québec, Canada; 2University of Manitoba, Winnipeg,
Canada; 3University of Ottawa, Canada
Critical Care 2013, 17(Suppl 2):P334 (doi: 10.1186/cc12272) M Asselin1, Y Lachance1, G Lalonde1, A Boutin1, M Shemilt1, L Moore1, F Lauzier1,
R Zarychanski2, P Archambault1, F Lamontagne1, D Fergusson3, A Turgeon1
1Université Laval, Québec, Canada; 2University of Manitoba, Winnipeg,
Canada; 3University of Ottawa, Canada
Critical Care 2013, 17(Suppl 2):P334 (doi: 10.1186/cc12272) S128 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Conclusion This survey shows that there is no clear consensus on
what types of monitoring should be used to guide management of
patients with TBI. A CPP protocol, based on measurement of ICP, is the
most widely used method, although a recent randomized control trial
did not show any benefi t in outcome [2]. Other invasive monitoring
methods, although they may help in individualized care, are still not
yet popular to due lack of strong evidence. Larger multicentre portfolio
studies are needed to establish their benefi ts. References signifi cantly higher risk of death in the group with INR ≥1.3: HR = 2.99
(95% CI = 1.8 to 5.1); P <0.001 (Figure 1). g
Conclusion In patients with head injury and normal INR (<1.6), there
is an association between an INR ≥1.3 and higher risk of death. Therefore, it would be useful to request an INR in patients presenting
with a head injury when bleeding is suspected, even in the absence of
anticoagulant. f P339 Evaluation of proximal–distal velocity gradient in spastic middle
cerebral artery after aneurismal subarachnoid hemorrhage
M Gotti1, F Stretti1, S Piff eri1, V Conte1, M Zara1, N Stocchetti2
1Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan,
Italy; 2Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan
University, Milan, Italy
Critical Care 2013, 17(Suppl 2):P339 (doi: 10.1186/cc12277) Results Twenty-fi ve patients (age 57 ± 12 years, WFNS 1 to 5) were
included in the analysis. Overall increased levels of MPs were observed
at early phase after aSAH when compared with controls. Contrarily
to our hypothesis, in the early phase no signifi cant diff erences of
MP level were observed in WFNS 4 to 5 (n = 9) when compared with
WFNS 1 to 2 (n = 16) patients. Interestingly, patients with most severe
ECI and DCI showed the highest values of MPs, but this trend did not
reach statistical signifi cance, probably because of the small number of
patients included in the study. Introduction Delayed cerebral ischemia (DCI) worsens neurological
outcome in patients with aneurysmal subarachnoid hemorrhage
(aSAH). DCI pathogenesis is multifactorial and not completely
understood, but vasospasm plays a central role. Transcranial color-
coded duplex sonography (TCCDS) is a non-invasive bedside tool to
explore cerebral vessels, but specifi city is still suboptimal. The aim
of this study was to evaluate the proximal–distal gradient of mean
cerebral blood fl ow velocity (CBF-V) in middle cerebral artery (MCA) as
a possible indicator of critical vasospasm. Conclusion Microparticles and microthrombosis may increase the
severity of early and delayed ischemic damage after aSAH. p
p
Methods Consecutive aSAH patients (WFNS 1 to 5, age 18 to 80 years)
admitted to NeuroICU between November 2011 and September 2012
were included in this study. TCCDS was used to assess CBF-V in MCA of
both sides: we defi ned TCCDS vasospasm as CBF-V >120 cm/second. Magnetic resonance angiography (MRA, 3D TOF HR) was performed to
evaluate vasospasm at early (<3 days) and delayed (7 to 10 days) time
points. When patients underwent MRA, in the same clinical conditions,
we recorded CBF-V with TCCDS at two levels: at the origin (proximal
CBF-V) and at the end (distal CBF-V) of the MCA. Then we calculated the
absolute value of proximal–distal CBF-V gradient for MCA of both sides. We defi ned clinical vasospasm as the appearance of a new neurological
defi cit confi rmed by imaging. P338
Early respiratory deterioration predicts poor outcome after
aneurysmal subarachnoid hemorrhage
E Van Lummel, L Vergouw, M Van der Jagt
Erasmus MC, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P338 (doi: 10.1186/cc12276) Introduction Pulmonary complications are frequently occurring
medical complications after aneurysmal subarachnoid hemorrhage
(aSAH) [1]. Early respiratory deterioration (ERD) may be associated
with delayed cerebral ischemia (DCI) or outcome and would then be a
potential target for therapeutic interventions. We investigated whether
respiratory deterioration within the fi rst 72 hours after admission
predicted DCI or poor outcome. Methods We conducted a retrospective study in 137 consecutively
admitted patients with aSAH, admitted between October 2007 and
October 2011 to the ICU of a university hospital. ERD was defi ned as
increased need for ventilatory support the second or third day after
admission (Table 1). Elective intubation for a surgical procedure was
not included as ERD. Inclusion criteria were availability of detailed
information on respiratory status and level of support, admission
within 48 hours after hemorrhage and age ≥18 years. Multivariable Figure 1 (abstract P336). Survival analysis of patients with INR <1.3 and
with INR ≥1.3. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S129 Table 1 (abstract P338). Categories of respiratory support
Level of respiratory support
Category
(compared with fi rst admission day)
1
Up to 5 l O2
2
>5 l O2 by nasal canulla or Venturi mask
3
Nonrebreathing mask or non-invasive positive-pressure
ventilation
4
Invasive mechanical ventilation
When mechanically
Decreased PaO2/FiO2 ratio
terminated Results We included 26 consecutive aSAH patients (WFNS 1 to 5, age
57 ± 12 years). The absolute value of MCA CBF-V gradient was higher in:
(1) vessels aff ected by TCCDS vasospasm when compared with vessels
not aff ected (63 cm/second, IR 23 to 85 and 15 cm/second IR 7 to 23
respectively, P <0.001); (2) vessels aff ected by MRA vasospasm when
compared with vessels not aff ected (43 cm/second, IR 18 to 78 and
13 cm/second IR 6 to 22, P <0.001); and (3) in patients who developed
clinical vasospasm than in patients who did not (56 cm/second IR 11
to 124 and 16 cm/second IR 8 to 30 respectively, P <0.01). Considering
only the subset of MCAs aff ected by MRA vasospasm, the CBF-V
gradient was higher when clinical vasospasm was also present (75 cm/
second IR 56 to 124 and 23 cm/second IR 17 to 30 respectively, P <0.05). Conclusion Proximal–distal CBF-V gradient of MCA might be a reliable
indicator of critical vasospasm but further studies are warranted to
defi ne threshold values and specifi city in aSAH patients. P338
Early respiratory deterioration predicts poor outcome after
aneurysmal subarachnoid hemorrhage
E Van Lummel, L Vergouw, M Van der Jagt
Erasmus MC, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P338 (doi: 10.1186/cc12276) survival analysis was used to investigate associations of DCI, death
and Glasgow Outcome Scale (GOS) with ERD adjusted for condition
on admission, Hijdra score, treatment of ruptured aneurysm and
pulmonary comorbidity. GOS was assessed at 3 to 6 months after the
bleed. DCI was defi ned as described recently [2]. P340 P340
Microparticles in aneurismal subarachnoid hemorrhage: role in
acute and delayed cerebral ischemia
V Conte1, A Artoni1, S Magnoni1, M Leoni1, L Capuano1, V Civelli1,
N Stocchetti2
1Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan,
Italy; 2Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan
University, Milan, Italy
Critical Care 2013, 17(Suppl 2):P340 (doi: 10.1186/cc12278) i
y
Results Mean age of the patients was 55.9 (± 12.7) and 63.5% was
female. A total 46.7% of the patients developed DCI. Mortality was
25.6%. Forty percent of the patients were classifi ed as having ERD. ERD
was not associated with DCI (adjusted HR = 1.48; 95% CI = 0.88 to 2.49;
P = 0.14). ERD showed a trend towards an association with mortality
(adjusted HR = 2.22; 95% CI = 0.96 to 5.14; P = 0.06; additionally adjusted
for age, and rebleed). A clear association was found between absence
of ERD and functional outcome with ordinal logistic regression analysis
(0.98 point increase in GOS score at 3 to 6 months; 95% CI = 0.14 to 1.82;
P = 0.02; additionally adjusted for age and rebleed). Introduction Microthrombosis has been demonstrated in early and
delayed cerebral ischemia after aneurysmal subarachnoid hemorrhage
(aSAH). Markers of coagulation activation as microparticles (MPs) are an
established risk factor for thrombosis. Our hypothesis was that levels
of microparticles might correlate with aSAH severity, early cerebral
ischemia (ECI) and delayed cerebral ischemia (DCI). y
j
g
Conclusion ERD within 72 hours after admission is associated with
increased risk of poor functional outcome after aSAH, but not DCI. Further investigations are required to assess whether prevention of
ERD may improve outcome. Methods Consecutive aSAH patients (age 18 to 80) admitted to our
department between November 2011 and September 2012 were
enrolled in the study. Total MPs (anxV+), platelet MPs (PLT-MPs, anx+/
CD41+), tissue factor MPs (TF-MPs, anx+/CD142+), and endothelial
MPs (E-MPs, anx+/CD144+) were assessed in venous blood samples. Measurement of MPs levels were obtained at early (<3 days) and delayed
phase (7 to 10 days). Multiparameter MRI (T2; FLAIR; T1; AngioRM 3D TOF
HR; DWI) was performed to evaluate ischemic damage and vasospasm
at the same time points. Qualitative evaluation of severity of ischemic
damage was performed on DWI sequences. Levels of MPs were evaluated
comparing diff erent groups in terms of SAH severity (World Federation
of Neurological Surgeons score or WFNS), ECI severity and occurrence of
delayed complications (vasospasm and DCI).i References References
1. Friedman JA, et al.: Neurosurgery 2003, 52:1025-1031. 2. Vergouwen MD, et al.: Stroke 2010, 41:2391-2395. P339 The relationships between the absolute
value of TCCDS CBF-V gradient and (1) TCCDS vasospasm, (2) MRA
vasospasm and (3) clinical vasospasm were explored. P341 P341
Intracranial pressure after subarachnoid hemorrhage
b d
1
l
1
2
h 1
S
h
1 Intracranial pressure after subarachnoid hemorrhage
A Lombardo1, T Zoerle1, ER Zanier2, L Longhi1, N Stocchetti1
1Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, University of
Milan, Italy; 2Mario Negri Institute, Milan, Italy
Critical Care 2013, 17(Suppl 2):P341 (doi: 10.1186/cc12279) Introduction Elevated intracranial pressure (ICP) may have deleterious
eff ects on cerebral metabolism and mortality after aneurysmal
subarachnoid hemorrhage (SAH) [1,2], but its relevance has not yet
been well explored. Aims of this study are to track ICP changes after
SAH, to identify clinical factors associated with it and to explore the
relationship between ICP and outcome. Introduction Elevated intracranial pressure (ICP) may have deleterious
eff ects on cerebral metabolism and mortality after aneurysmal
subarachnoid hemorrhage (SAH) [1,2], but its relevance has not yet
been well explored. Aims of this study are to track ICP changes after
SAH, to identify clinical factors associated with it and to explore the
relationship between ICP and outcome. S130 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods A total of 116 consecutive SAH patients with ICP monitoring
were enrolled. Episodes of ICP >20 mmHg for at least 5 minutes and the
mean ICP value for every 12-hour interval were analyzed. The highest
mean ICP collected in every patient was identifi ed. ICP values were
analyzed in relation to clinical and CT fi ndings; 6-month outcome and
ICU mortality were also introduced in multivariable logistic models. with aSAH. Studies including more than 50% of non-aneurismal SAH
were excluded. Studies were considered at low risk of bias if the authors
defi ned inclusion/exclusion criteria, avoided voluntary sampling, and
tested >90% of included patients with proper detailed diagnostic
criteria. Studies testing all pituitary axes were considered as evaluating
hypopituitarism, which was defi ned as the dysfunction of ≥1 axis. We used a Freeman Tukey-type arcsine square-root transformation
and pooled prevalences using the DerSimonian–Laird random-eff ect
method. We determined the degree of heterogeneity with I2 values. Results Among 12,363 citations, we included 28 studies (1,628
patients). Patients were mostly female (64%) aged 50.5 ± 5.4. Sixteen
studies reported the severity of aSAH, 14 reported the procedure for
securing the aneurysm and 13 reported the location of aneurysm. Overall, hypopituitarism was observed in 53.4% of patients at short-
term (<3 months), 36.5% at mid-term (3 to 12 months) and 34.2% at
long-term (>12 months) (Table 1). Correlation between the timing of endovascular coiling or
microsurgical clipping and long-term outcomes of patients after
aneurysmal subarachnoid haemorrhage
N Ojukwu, R Jain, S Wijayatilake, A Bellini, R Shetty, J Khan, G De la Cerda
Queen’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P343 (doi: 10.1186/cc12281) Correlation between the timing of endovascular coiling or
microsurgical clipping and long-term outcomes of patients after
aneurysmal subarachnoid haemorrhage
N Ojukwu, R Jain, S Wijayatilake, A Bellini, R Shetty, J Khan, G De la Cerda
Queen’s Hospital, London, UK
Critical Care 2013, 17(Suppl 2):P343 (doi: 10.1186/cc12281) References 1. Heuer GG, et al.: Relationship between intracranial pressure and other
clinical variables in patients with aneurismal subarachnoid hemorrhage. J
Neurosurg 2004, 101:408-416. 1. Heuer GG, et al.: Relationship between intracranial pressure and other
clinical variables in patients with aneurismal subarachnoid hemorrhage. J
Neurosurg 2004, 101:408-416. 2. Nagel A, et al.: Relevance of intracranial hypertension for cerebral
metabolism in aneurismal subarachnoid hemorrhage. Clinical article. J
Neurosurg 2009, 111:94-101. 2. Nagel A, et al.: Relevance of intracranial hypertension for cerebral
metabolism in aneurismal subarachnoid hemorrhage. Clinical article. J
Neurosurg 2009, 111:94-101. P341
Intracranial pressure after subarachnoid hemorrhage
b d
1
l
1
2
h 1
S
h
1 There was an insuffi cient number of
studies with low risk of bias to perform sensitivity analyses according
to study quality. y
Results Eighty-one percent of patients had at least one episode of
elevated ICP and 36% had a highest mean ICP >20 mmHg. The number
of patients with highest mean ICP >20 mmHg or with episodes of
HICP was maximum at day 3 after SAH and decreased only after day 7. Neurological status, aneurysmal rebleeding, amount of blood on
CT and CT ischemic lesion occurred within 72 hours from SAH were
signifi cantly related to highest mean ICP >20 mmHg in a multivariable
model. Patients with highest mean ICP >20 mmHg showed signifi cantly
higher mortality in ICU. However, ICP is not an independent predictor
of 6 months unfavorable outcome. Conclusion Elevated intracranial pressure is a common complication in
the fi rst week after SAH. It is associated with early brain injury severity
and ICU mortality. Conclusion The exact prevalence of pituitary disorders following aSAH
remains uncertain, mainly due to high heterogeneity and the small
number of studies with low risk of bias. However, the prevalence seems
to decrease during the recovery phase. The prevalence, risk factors and
clinical signifi cance of pituitary disorders in aSAH will require further
rigorous evaluation. y
References Prevalence of pituitary disorders associated with aneurysmal
subarachnoid hemorrhage g
F Lauzier, I Cote, A Bureau, M Shemilt, A Boutin, L Moore, C Gagnon,
AF Turgeon
Universite Laval, Quebec, Canada
Critical Care 2013, 17(Suppl 2):P342 (doi: 10.1186/cc12280) Introduction
Aneurysmal
subarachnoid
haemorrhage
(aSAH)
is associated with a high morbidity and mortality. Although UK
Anaesthesia Guidelines advocate early coiling or clipping of the
aneurysm within the fi rst 72 hours of admission for all grades of aSAH,
the optimal timing of treatment and whether this is linked with better
neurological long-term outcome are a subject of debate [1]. We aimed
to investigate whether the timing of the occlusion of the aneurysm
translates into better outcome. Introduction Pituitary disorders are an often-neglected consequence
of aneurysmal subarachnoid hemorrhage (aSAH). We systematically
reviewed their prevalence, aiming particularly at studies with low risk
of bias. Methods We searched Embase, MEDLINE, The Cochrane Library, Trip
Database, references of included studies and narrative reviews. We
included cohort studies, cross-sectional studies and RCTs published
in any language that tested the integrity of ≥1 pituitary axis in adults Methods A retrospective analysis of prospective collected data in
a tertiary neuroscience centre from January to September 2012. Neuroprotective eff ect of a new synthetic aspirin–decursinol
adduct in a rat model of ischemic stroke Neuroprotective eff ect of a new synthetic aspirin–decursino
adduct in a rat model of ischemic stroke
YH Lee, CW Park, JH Cho, MH Won
Kangwon National University, Chuncheonsi, South Korea
Critical Care 2013, 17(Suppl 2):P345 (doi: 10.1186/cc12283) patients were managed according to the local Guidelines for the
management of aSAH. Outcome was assessed at 3 months using the
extended Glasgow Outcome Scale (GOSE) defi ning good recovery as a
GOSE ≥7 and poor outcome as GOSE ≤6. Introduction Stroke is a major cause of death. This study investigated
the preventative eff ect of a new synthetic drug on brain function in
experimentally induced ischemic stroke. Introduction Stroke is a major cause of death. This study investigated
the preventative eff ect of a new synthetic drug on brain function in
experimentally induced ischemic stroke. Results A total of 28 patients were included within the study period. Three patients were not expected to survive the fi rst 24 hours and were
not included in the study. Seventeen patients were classifi ed as good
grade aSAH (WFNS I to III) and eight as poor grade (IV to V). Twenty-two
patients underwent successful coiling while the other three required
clipping due to unsuccessful coiling. We did not fi nd any correlation
between the timing of coiling/clipping and the 3-month GOSE
(Figure 1). A total 44% of the patients had a poor 3-month GOSE while
56% had a good long-term functional outcome. The overall mortality
rate was 21%. Methods Male Sprague–Dawley rats were administered aspirin (ASA),
decursinol (DA) or ASA-DA before and after ischemic insults. Brain
and neuronal damage were examined by TTC staining, PEP-CT, NeuN
immunohistochemistry and F-J B histofl uorescence. Gliosis was also
observed by GFAP and iba-1 immunohistochemistry. y
y
Results Pretreatment with 20 mg/kg, but not 10 mg/kg, of ASA-DA
protected against ischemic neuronal death and damage, and its
neuroprotective eff ect was much more pronounced than that of ASA
or DA alone. In addition, treatment with 20 mg/kg ASA-DA reduced the
ischemia-induced activation of astrocytes and microglia.i Conclusion Overall mortality in patients with aSAH is low when
aneurysm is treated early post rupture of aneurysm. We did not fi nd
any correlation between the timing of occlusion of aneurysm and the
3-month functional outcome. Prevalence of pituitary disorders associated with aneurysmal
subarachnoid hemorrhage All Table 1 (abstract P342)
Hypopituitarism
GH
ACTH
TSH
Gonadal
ADH
Short term
All studies
53.4% (20.5 to 84.7) 30.4% (11.5 to 53.2)
13.2% (4.4 to 25.1)
49.2% (0.1 to 17.7)
26.1% (8.8 to 48.0)
6.3% (2.7 to 11.0)
n (patients)
5 (235)
8 (346)
13 (537)
7 (327)
8 (351)
3 (481)
I2
96.0%
93.5%
90.2%
89.1%
93.0%
61.1%
Studies of low risk of bias (patients)
2(56)
–
–
2(56)
1(26)
–
Mid-term
All studies
36.5% (13.6 to 62.8)
7.3% (0.6 to 18.3)
8.9% (1.3 to 21.0)
8.8% (2.4 to 18.0)
3.9% (0.0 to 11.9)
3.7% (0.6 to 13.5)
n (patients)
4 (109)
6 (158)
7 (208)
5 (176)
6 (178)
2(56)
I2
85.7%
70.9%
79.4%
59.4%
66.8%
29.0%
Studies with low risk of bias (patients)
1(40)
–
–
1(40)
–
–
Long-term
All studies
34.2% (18.7 to 51.4)
17.1% (9.4 to 26.4)
11.2% (4.8 to 19.4)
2.3% (0.4 to 5.3)
3.2% (0.7 to 6.8)
0% (0 to 1.5)
n (patients)
12 (486)
13 (531)
13 (531)
12 (486)
13(351)
3 (142)
I2
92.5%
82.9%
82.9%
49.0%
62.9%
0%
Studies with low risk of bias (patients)
2(40)
2(40)
2(40)
2(40)
1(10)
– Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S131 Results All WFNSpt 1 to 2 patients completed neuropsychological tests
at each time point. WFNSpt 3 and WFNS 4 patients were testable in 80%
and 50% of the cases respectively at early time points. WFNS 5 patients
were not testable at any time point. In all testable patients, cognitive
functions were severely impaired at early time points. At 3 months in
WFNS 1 to 3 a good recovery of language defi cits while only a partial
recovery of attention, memory and executive functions were observed;
at the same time point 70% of WFNS 4 patients became testable, but
they had a worse recovery of all cognitive functions. At 1 month after
SAH less than 30% of patients return to work, at 3 months approximately
50%. Despite a good recovery of everyday life activities at 3 months, for
all patients quality of life was lower than a normal population.f Figure 1 (abstract P343). Time of coiling/clipping versus GOSE boxplot. Prevalence of acute critical neurological disease in children: a global
epidemiological assessment (PANGEA) Prevalence of acute critical neurological disease in children: a global
epidemiological assessment (PANGEA)
E Fink1, R Tasker2, J Beca3, MJ Bell1, RS Clark1, J Hutchison4,
M Vavilala5, RS Watson1, L Weissfeld6, PM Kochanek1, DC Angus7,
PANGEA Investigators1
1Children’s Hospital of Pittsburgh, PA, USA; 2Children’s Hospital Boston, MA,
USA; 3Starship Children’s Hospital, Auckland, New Zealand; 4The Hospital for
Sick Children, Toronto, Canada; 5Seattle Children’s Hospital, Seattle, WA, USA;
6University of Pittsburgh, PA, USA; 7University of Pittsburgh Medical Center,
Pittsburgh, PA, USA
Critical Care 2013, 17(Suppl 2):P346 (doi: 10.1186/cc12284) Prevalence of pituitary disorders associated with aneurysmal
subarachnoid hemorrhage p
q
y
p p
Conclusion Cognitive dysfunction has diff erent time courses after
aSAH: signifi cant defi cits in diff erent cognitive domains, worse quality
of life and diffi culties in return to work persist in more than 50% of
patients at 3 months following SAH. P344 Time course of neuropsychological functioning after aneurysmal
subarachnoid hemorrhage
B Zarino1, G Bertani1, V Conte1, S Magnoni1, A Di Cristofori1, N Stocchetti2
1Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan,
Italy; 2Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan
University, Milan, Italy
Critical Care 2013, 17(Suppl 2):P344 (doi: 10.1186/cc12282) Reference 1. van der Schaaf I, et al.: Stroke 2006, 37:572-573. 1. van der Schaaf I, et al.: Stroke 2006, 37:572-573. Neuroprotective eff ect of a new synthetic aspirin–decursinol
adduct in a rat model of ischemic stroke Conclusion Our fi ndings indicate that ASA-DA, a new synthetic drug,
prevents against transient focal cerebral ischemia, which provides a
resource for the development of its clinical application for stroke. Time course of neuropsychological functioning after aneurysmal
subarachnoid hemorrhage An aff ordable and reliable bedside
test could change practice to an individual tailored dosing of LMWH. The aim of our study was to evaluate whether a prophylactic dose
regimen of 40 mg enoxaparine in cardiac surgical patients increases
the anti-Xa activity to the level necessary for effi cient prevention of a
thromboembolic event [1]. Secondarily we tested whether there was a
reliable correlation between a bedside anti-Xa measurement compared
with a two-stage chromogenic assay at the laboratory [2]. assay remain controversial in daily clinical practice. One of the major
obstacles is the cost of such a test. An aff ordable and reliable bedside
test could change practice to an individual tailored dosing of LMWH. The aim of our study was to evaluate whether a prophylactic dose
regimen of 40 mg enoxaparine in cardiac surgical patients increases
the anti-Xa activity to the level necessary for effi cient prevention of a
thromboembolic event [1]. Secondarily we tested whether there was a
reliable correlation between a bedside anti-Xa measurement compared
with a two-stage chromogenic assay at the laboratory [2]. Results Of 3,113 subjects screened, 1,009 (32%) met enrollment criteria. The mean number of subjects enrolled per site for each study day was
2.4. Most sites were dedicated pediatric ICUs with a mean number of
22 ICU beds (range 3 to 72). ICUs had resources to invasively monitor
intracranial pressure (93%), continuous electroencephalography (14%),
invasive and non-invasive brain tissue oxygenation (14% and 57%),
and somatosensory evoked potentials (39%). There were on average
11 ICU faculty and six fellows per site, and nearly one-half reported
a neurocritical care ICU team. Subjects were 58% male and 52%
white, and 60% had normal pre-admission PCPC scores (85%). Status
epilepticus and cardiac arrest (both 21%) had the highest prevalence. Sixty-one per cent of subjects were mechanically ventilated during ICU
admission. ICU length of stay was a mean 29 days (median 43 days) and
hospital LOS was a mean 43 days (median 61 days). Survival at hospital
discharge was 87% with 58% of subjects discharged home and 17% to
inpatient rehabilitation.i y
y
Methods This was an open, single-centre, prospective, nonrandomized
clinical trial at a university hospital. All patients that needed
prophylactic dosing of enoxaparine after cardiac surgery were duly
informed and after giving written consent we included 44 patients with
a mean Euroscore of 1.66. Eff ective dosage of enoxaparin for intensive care patients
S Robinson, A Zincuk, U Larsen, P Toft
Odense University Hospital, Odense, Denmark
Critical Care 2013, 17(Suppl 2):P349 (doi: 10.1186/cc12287) Eff ective dosage of enoxaparin for intensive care patients
S Robinson, A Zincuk, U Larsen, P Toft
Odense University Hospital, Odense, Denmark
Critical Care 2013, 17(Suppl 2):P349 (doi: 10.1186/cc12287) gi
Results In total, 943 patients met the inclusion criteria with a mean
follow-up period of 1 year. The mean age was 74.3 years and 50.4%
were men. A total of 108 deaths (11.5%) were recorded within the
follow-up period; eight (25%) for dabigatran compared with 44 (12.6%)
for warfarin and 56 (9.9%) for aspirin. The mortality risk for patients
on dabigatran was signifi cantly higher than for patients on warfarin:
HR = 2.1 (95% CI: 1.0 to 4.5), P = 0.05 after controlling for age. Aspirin
had a lower (but not statistically signifi cant) mortality risk compared
with warfarin; HR = 0.75 (95% CI: 0.50 to 1.14), P = 0.18 after controlling
for age. y
p
Critical Care 2013, 17(Suppl 2):P349 (doi: 10.1186/cc12287) Introduction We hypothesized that higher doses of enoxaparin would
improve thromboprophylaxis without increasing the risk of bleeding. Critically ill patients are predisposed to venous thromboembolism,
leading to increased risk of adverse outcome [1]. Peak anti-factor
Xa (anti-Xa) levels of 0.1 to 0.4 IU/ml, 4 hours post administration of
enoxaparin, refl ect adequate thromboprophylaxis for medico-surgical
patients. Conclusion The results showed higher overall mortality in patients
who presented to the ED with a bleeding complication and were
taking dabigatran compared with warfarin or aspirin. Physicians
should be aware of the potential higher mortality with dabigatran over
warfarin when treating a bleeding patient. However, this was a single-
centre retrospective analysis with a small number of patients taking
dabigatran (n = 32), and further studies are needed to corroborate the
results. p
Methods The sample population consisted of 78 patients, randomized
to receive subcutaneous (s.c.) enoxaparin: 40 mg ×1 (control group),
versus 30 mg ×2, 40 mg ×2 or 1 mg/kg ×1 (test groups) for a period
of 3 days. Anti-Xa activity was measured at baseline, and at 4, 12, 16
and 24 hours post administration on each day. Patients did not diff er
signifi cantly between groups. i
Results On day 1 of administration, doses of 40 mg ×1 and 40 mg
×2 yielded similar mean peak anti-Xa of 0.20 IU/ml and 0.17 IU/ml
respectively, while a dose of 30 mg ×2 resulted in subtherapeutic
levels of anti-Xa (0.08 IU/ml). Time course of neuropsychological functioning after aneurysmal
subarachnoid hemorrhage B Zarino1, G Bertani1, V Conte1, S Magnoni1, A Di Cristofori1, N Stocchetti2
1Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan,
Italy; 2Fondazione IRCCS Ca’ Granda-Ospedale Maggiore Policlinico, Milan
University, Milan, Italy
Critical Care 2013, 17(Suppl 2):P344 (doi: 10.1186/cc12282) Introduction Early and delayed cognitive dysfunctions are an under-
studied issue after aneurysmal subarachnoid hemorrhage (aSAH),
irrespective of neurological outcome. The aim of this study was to
describe early and delayed changes of cognitive functions, activities of
everyday life, and quality of life in aSAH patients. Introduction Acute neurological injury is a leading cause of morbidity
and mortality in children. Global prevalence and regional disparities
of etiology, interventions, and outcomes are unknown. The aim of
this point-prevalence study was to measure the burden of pediatric
neurological injury and to describe variations in interventions and
outcomes in ICUs. Methods Consecutive aSAH patients admitted to our ICU between
November 2011 and September 2012 were prospectively studied. Patients underwent neuropsychological evaluation at early (<3 days,
10 days) and delayed time points (1 month, 3 months). Patients were
tested for language, verbal fl uency, short-term and long-term memory,
attention, executive functions, praxis, and neglect. Impairments in
activities of everyday life were assessed using the Activities of Daily
Living scale and the Instrumental Activities of Daily Living scale. The SF-36 was used to assess the quality of life at 3 months. Since
complications of aneurysm treatment in addition to aSAH severity may
signifi cantly aff ect cognitive status, patients were evaluated according
to the World Federation of Neurological Surgeons score after treatment
(WFNSpt). Methods One hundred and three ICUs on six continents enrolled
subjects on 4 specifi c days in a 1-year period. Included subjects were
between ages 7 days and 17 years who were diagnosed with acute
traumatic brain injury, stroke, cardiac arrest, central nervous system
infection or infl ammation, status epilepticus, spinal cord lesion,
hydrocephalus, or brain mass. Sites completed a secure web-based
case report form that included subject and hospital demographics,
details about the neurological disease, interventions, length of stay,
and Pediatric Cerebral Performance Category (PCPC) score (good
outcome = PCPC 1 to 3) and mortality at hospital discharge. S132 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 assay remain controversial in daily clinical practice. One of the major
obstacles is the cost of such a test. P347
Survival analysis of patients taking dabigatran who consulted an
emergency department for acute bleeding: a fi rst alarm
OA Arlet, EN Notebaert, JP Paquet, RD Daoust, MV Vincent, JC Chauny
Hôpital du Sacré-Coeur de Montréal, Université de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P347 (doi: 10.1186/cc12285) P347
Survival analysis of patients taking dabigatran who consulted an
emergency department for acute bleeding: a fi rst alarm
OA Arlet, EN Notebaert, JP Paquet, RD Daoust, MV Vincent, JC Chauny
Hôpital du Sacré-Coeur de Montréal, Université de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P347 (doi: 10.1186/cc12285) Survival analysis of patients taking dabigatran who consulted an
emergency department for acute bleeding: a fi rst alarm OA Arlet, EN Notebaert, JP Paquet, RD Daoust, MV Vincent, JC Chauny
Hôpital du Sacré-Coeur de Montréal, Université de Montréal, Canada
Critical Care 2013, 17(Suppl 2):P347 (doi: 10.1186/cc12285) Introduction The newly approved oral anticoagulant dabigatran has
no eff ective antidote. We therefore suspected an overall increase in
mortality in patients presenting to the emergency department (ED)
with a bleeding complication on dabigatran compared with warfarin
or aspirin. Conclusion The bedside anti-Xa activity assay with a Hemochron device
tends to show some correlation with the two-stage chromogenic assay,
but insuffi cient to be used as an alternative, in this small but uniform
patient population. Use of a standard dosing protocol for enoxaparine
administration is prone for underdosage in post-cardiac surgery
patients and may increase postoperative morbidity. R f p
Methods We conducted a post hoc analysis on a database of all patients
admitted to a tertiary-care ED with any kind of bleeding or suspicion
of one from March 2011 to August 2012 who were taking dabigatran,
warfarin, or aspirin. The primary endpoint was long-term survival. Patients were censored at death or at the end of the study period
(7 December 2012). We performed a Cox proportional hazard model,
controlled for age, to calculate the hazard ratio (HR) for dabigatran
versus warfarin and one for warfarin versus aspirin. Statistical
signifi cance was set at α = 0.05 and results are presented with 95% CI. Results In total, 943 patients met the inclusion criteria with a mean
follow-up period of 1 year. The mean age was 74.3 years and 50.4%
were men. A total of 108 deaths (11.5%) were recorded within the
follow-up period; eight (25%) for dabigatran compared with 44 (12.6%)
for warfarin and 56 (9.9%) for aspirin. The mortality risk for patients
on dabigatran was signifi cantly higher than for patients on warfarin:
HR = 2.1 (95% CI: 1.0 to 4.5), P = 0.05 after controlling for age. Aspirin
had a lower (but not statistically signifi cant) mortality risk compared
with warfarin; HR = 0.75 (95% CI: 0.50 to 1.14), P = 0.18 after controlling
for age. Methods We conducted a post hoc analysis on a database of all patients
admitted to a tertiary-care ED with any kind of bleeding or suspicion
of one from March 2011 to August 2012 who were taking dabigatran,
warfarin, or aspirin. The primary endpoint was long-term survival. Eff ective dosage of enoxaparin for intensive care patients
S Robinson, A Zincuk, U Larsen, P Toft
Odense University Hospital, Odense, Denmark
Critical Care 2013, 17(Suppl 2):P349 (doi: 10.1186/cc12287) Patients receiving 1 mg/kg enoxaparin
achieved near-steady-state levels from day 1 with mean peak anti-Xa
levels of 0.34 IU/ml. Steady-state anti-Xa was achieved for all doses of
enoxaparin at day 3. At steady state, mean peak anti-Xa levels of 0.13 IU/
ml and 0.15 IU/ml were achieved with doses of 40 mg ×1 and 30 mg ×2
respectively. This increased signifi cantly to 0.33 IU/ml and 0.40 IU/ml for
doses of 40 mg ×2 and 1 mg/kg enoxaparin respectively (P = 0.0000)
(Figure 1). A dose of 1 mg/kg enoxaparin yielded therapeutic anti-Xa
levels for over 80% of the study period. There were no adverse eff ects. Survival analysis of patients taking dabigatran who consulted an
emergency department for acute bleeding: a fi rst alarm Patients were censored at death or at the end of the study period
(7 December 2012). We performed a Cox proportional hazard model,
controlled for age, to calculate the hazard ratio (HR) for dabigatran
versus warfarin and one for warfarin versus aspirin. Statistical
signifi cance was set at α = 0.05 and results are presented with 95% CI. 1. Ribic C, et al.: J Crit Care 2009, 24:197-205. 1. Ribic C, et al.: J Crit Care 2009, 24:197-205. 1. Ribic C, et al.: J Crit Care 2009, 24:197-205. 2. Gehrie E, et al.: Am J Hematol 2012, 87:194-196. Time course of neuropsychological functioning after aneurysmal
subarachnoid hemorrhage The demographic specifi cations, medical
and surgical history of all patients were collected. Anti-Xa activity was
measured at three diff erent points in time. We determined baseline,
peak and trough anti-Xa activity: preoperatively, and respectively
4 hours after the third dose of enoxaparine and 30 minutes before the
fourth dose. Each measurement was done with both techniques, the
two-stage chromogenic assay at the laboratory (Biophen®) and the
bedside assay (Hemochron® Jr). Conclusion Acute neurological disease is a signifi cant pediatric health
issue. These data suggest a vital need for increased research and
healthcare resources to assist in the challenge of improving outcomes
for these children. Results Our dose regimen of enoxaparine achieved in one-half of
the included patients a suffi cient anti-Xa activity for prevention of
thromboembolic events. One-half of the patients with insuffi cient anti-
Xa activity had a body mass index over 30 kg/m2. Comparison of the
bedside assay with the two-stage chromogenic assay by means of the
Pearson’s correlation coeffi cient showed correlation of the two tests if
no variables were taken into account. In the Bland–Altman analysis we
could not confi rm this correlation. P348 P348
Bedside anti-Xa measurement for therapeutic assessment of a
prophylactic anticoagulation regimen
B Dewulf1, I Herck2, F De Somer2, K Francois2, J Decruyenaere2
1AZ Sint-Jan, Brugge, Belgium; 2University Hospital, Gent, Belgium
Critical Care 2013, 17(Suppl 2):P348 (doi: 10.1186/cc12286) P351 The fi rst-in-class hepcidin
antagonist NOX-H94, a PEGylated anti-hepcidin L-RNA oligonucleotide,
is in development for targeted treatment of anemia of infl ammation. We investigated whether NOX-H94 prevents the infl ammation-induced
serum iron decrease during experimental human endotoxemia. Methods A randomized, double-blind, placebo-controlled trial in
24 healthy young men. At T = 0 hours, 2 ng/kg E. coli endotoxin was
administered intravenously (i.v.), followed by 1.2 mg/kg NOX-H94 or
placebo i.v. at T = 0.5 hours. Blood was drawn serially after endotoxin
administration for measurements of infl ammatory parameters, Introduction Anemia is very frequently encountered on the ICU. Increased hepcidin production is one of the cornerstones of the
pathophysiology of anemia of infl ammation. The fi rst-in-class hepcidin
antagonist NOX-H94, a PEGylated anti-hepcidin L-RNA oligonucleotide,
is in development for targeted treatment of anemia of infl ammation. We investigated whether NOX-H94 prevents the infl ammation-induced
serum iron decrease during experimental human endotoxemia. Introduction Unfractionated heparin is preferred over LMWH in
ICU patients but LMWH is used more frequently in many European
ICUs. Thromboprophylaxis with standard doses of nadroparin and
enoxaparin has been shown to result in signifi cantly lower anti-Xa in
ICU patients when compared with medical patients [1,2]. p
p
p
Methods ICU patients (SAPS 44 ± 16, MV, n = 44; pressors n = 32)
received 7,500 IU (Group 1, n = 25) or 5,000 IU dalteparin s.c. (Group 2,
n = 29). Twenty-nine medical patients receiving 5,000 IU dalteparin
served as controls (Group 3). g
p
Methods A randomized, double-blind, placebo-controlled trial in
24 healthy young men. At T = 0 hours, 2 ng/kg E. coli endotoxin was
administered intravenously (i.v.), followed by 1.2 mg/kg NOX-H94 or
placebo i.v. at T = 0.5 hours. Blood was drawn serially after endotoxin
administration for measurements of infl ammatory parameters, p
Results Group 2 had signifi cantly lower areas under the Xa curve
(AUC) compared with Groups 1 and 3 (Table 1). Diff erences were not
signifi cant between Groups 1 and 3. Peak anti-Xa activities (Cmax-anti-Xa) Table 1 (abstract P351). g
Referencesfl 1. Dörffl er-Melly J, deJonge E, dePont AC, et al.: Bioavailability of subcutaneous
low-molecular-weight heparin to patients on vasopressors. Lancet 2002,
359:849-850. 1. Cook D, et al.: N Engl J Med 2011, 364:1305-1314. 2. Priglinger U, Delle Karth G, Geppert A, et al.: Prophylactic anticoagulation
with enoxaparin: is the subcoutaneous route appropriate in the critically
ill? Crit Care Med 2003, 31:1405-1409. 2. Priglinger U, Delle Karth G, Geppert A, et al.: Prophylactic anticoagulation
with enoxaparin: is the subcoutaneous route appropriate in the critically
ill? Crit Care Med 2003, 31:1405-1409. P350
Abstract withdrawn P350
Abstract withdrawn P351 Anti-Xa activities following 5,000 IU and 7,500 IU of s.c. dalteparin in
critically ill patients and 5,000 IU in medical patients: a prospective
randomized study
GH Heinz1, U Priglinger2, I Pabinger3
1University Clinic of Internal Medicine II, Vienna, Austria; 2University Clinic of
Emergency Medicine, Vienna, Austria; 3University Clinic of Internal Medicine I,
Vienna, Austria
Critical Care 2013, 17(Suppl 2):P351 (doi: 10.1186/cc12289) Randomized double-blind placebo-controlled PK/PD study on the
eff ects of a single intravenous dose of the anti-hepcidin Spiegelmer
NOX-H94 on serum iron during experimental human endotoxemia
L Van Eijk1, DW Swinkels1, A John1, F Schwoebel2, F Fliegert2, L Summo2,
S Vauleon2, JM Laarakkers1, K Riecke2, P Pickkers1
1Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands;
2NOXXON Pharma AG, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P352 (doi: 10.1186/cc12290) Anti-Xa activities following 5,000 IU and 7,500 IU of s.c. dalteparin in
critically ill patients and 5,000 IU in medical patients: a prospective
randomized study Randomized double-blind placebo-controlled PK/PD study on the
eff ects of a single intravenous dose of the anti-hepcidin Spiegelmer
NOX-H94 on serum iron during experimental human endotoxemia
L Van Eijk1, DW Swinkels1, A John1, F Schwoebel2, F Fliegert2, L Summo2,
S Vauleon2, JM Laarakkers1, K Riecke2, P Pickkers1
1Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands;
2NOXXON Pharma AG, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P352 (doi: 10.1186/cc12290) Randomized double-blind placebo-controlled PK/PD study on the
eff ects of a single intravenous dose of the anti-hepcidin Spiegelmer
NOX-H94 on serum iron during experimental human endotoxemia
L Van Eijk1, DW Swinkels1, A John1, F Schwoebel2, F Fliegert2, L Summo2,
S Vauleon2, JM Laarakkers1, K Riecke2, P Pickkers1
1Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands;
2NOXXON Pharma AG, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P352 (doi: 10.1186/cc12290) GH Heinz1, U Priglinger2, I Pabinger3
1University Clinic of Internal Medicine II, Vienna, Austria; 2University Clinic of
Emergency Medicine, Vienna, Austria; 3University Clinic of Internal Medicine I,
Vienna, Austria
Critical Care 2013, 17(Suppl 2):P351 (doi: 10.1186/cc12289) GH Heinz1, U Priglinger2, I Pabinger3
1University Clinic of Internal Medicine II, Vienna, Austria; 2University Clinic of
Emergency Medicine, Vienna, Austria; 3University Clinic of Internal Medicine I,
Vienna, Austria
Critical Care 2013, 17(Suppl 2):P351 (doi: 10.1186/cc12289) Introduction Anemia is very frequently encountered on the ICU. Increased hepcidin production is one of the cornerstones of the
pathophysiology of anemia of infl ammation. Bedside anti-Xa measurement for therapeutic assessment of a
prophylactic anticoagulation regimen Introduction Dose adjustments of low molecular weight heparin
(LMWH) based on daily anti-Xa measurement by chromogenic S133 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P349). Peak anti-Xa over 3 days with varying doses of enoxaparin. Figure 1 (abstract P349). Peak anti-Xa over 3 days with varying doses of enoxaparin. Conclusion Current standard s.c. doses of 40 mg ×1 enoxaparin
(Europe) or 30 mg ×2 (North America) yield levels of anti-Xa thought to
be subtherapeutic for critically ill patients. A weight-based dose yielded
the best anti-Xa levels, allowed the establishment of near-steady-state
levels from the fi rst day of enoxaparin administration, and may thus be
more appropriate, convenient, and eff ective. A new study using 1 mg/
kg enoxaparin s.c. with clinical endpoints has recently been approved
by the Danish Ministry of Health. R f were delayed (tmax-anti-Xa) in Group 2 compared with Groups 1 and 3
(Table 1). Conclusion In ICU patients a s.c. dose of 5,000 IU dalteparin results
in signifi cantly lower Xa activities when compared with normal ward
patients. A s.c. dose of 7,500 IU dalteparin in ICU patients resulted in
kinetics and peak anti-Xa activities comparable with medical patients
receiving 5,000 IU dalteparin. P351 Pharmacokinetics
Group 1 (n = 25)
Group 2 (n = 29)
Group 3 (n = 29)
ICU, 7,500 IU
ICU, 5,000 IU
Normal ward, 5,000 IU
P overall
AUC-anti-Xa0–12 hours (U/l*hour)
2.51 (1.15 to 4.61)
1.27 (1.15 to 4.4)
2.58 (1.45 to 4.87)
<0.001*
AUC-anti-Xa12_00 (U/l*hour)
1.37 (0.58 to 13)
1.47 (0.65 to 6.3)
0.89 (0.35 to 3.88)
0.003**
Cmax-anti-Xa (U/l)
0.29 (0.10 to 0.52)
0.14 (0.1 to 0.43)
0.33 (0.14 to 0.65)
<0.001***
tmax-anti-Xa (hours)
3 (3 to 12)
4.5 (1 to 12)
3 (3 to 6)
0.017§
Cmax, peak anti-Xa level; tmax, time of anti-Xa peak. Medians and range. *Group 1 versus 2, P = 0.001; Group 1 versus 3, P = 0.51. **Group 1 versus 2, P = 0.96; Group 1
versus 3, P = 0.045; Group 2 versus 3, P = 0.13. ***Group 1 versus 2, P = 0.001; Group 1 versus 3, P = 0.29. §Group 1 versus 3, P = 0.09; Group 1 versus 2, P = 0.8; Group 2
versus 3, P = 0.018. Table 1 (abstract P351). Pharmacokinetics Cmax, peak anti-Xa level; tmax, time of anti-Xa peak. Medians and range. *Group 1 versus 2, P = 0.001; Group 1 versus 3, P = 0.51. **Group 1 versus 2, P = 0.96; Group 1
versus 3, P = 0.045; Group 2 versus 3, P = 0.13. ***Group 1 versus 2, P = 0.001; Group 1 versus 3, P = 0.29. §Group 1 versus 3, P = 0.09; Group 1 versus 2, P = 0.8; Group 2
versus 3, P = 0.018. S134 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 cytokines, NOX-H94 pharmacokinetics, total hepcidin-25, and iron
parameters. The diff erence of serum iron change from baseline at
T = 9 hours was defi ned as the primary endpoint.ll Introduction According to many authors, acute necrotizing pancreatitis
(ANP) still remains one of the diffi cult problems of abdominal surgery. The
complexity of the pathogenesis of the disease, features of the pancreas
pathomorphology, abdominal hypertension, and high mortality (30 to
70%) necessitate a search for new ways to treat this disease. i
y
Results Endotoxin administration led to fl u-like symptoms. Infl am-
matory parameters (CRP, body temperature, leucocytes, and plasma
levels of TNFα, IL-6, IL-10, and IL-1RA) peaked markedly and similarly in
both treatment groups. P351 NOX-H94 was well tolerated. Plasma concen-
trations peaked at 0.7 ± 0.4 hours after the start of administration, after
which they declined according to a two-compartment model, with a
T1/2 of 22.5 ± 4.28 hours. In the placebo group, serum iron increased
from 19.0 ± 7.6 μg/l at baseline to a peak at T = 3 hours, returned close to
baseline at T = 6 hours and decreased under the baseline concentration
at T = 9 hours, reaching its lowest point at T = 12 hours. In the NOX-H94
group, serum iron concentrations rose until T = 6 hours and then slowly
declined until T = 24 hours. From 6 to 12 hours post LPS, the serum iron
concentrations in NOX-H94-treated subjects were signifi cantly higher
than in placebo-treated subjects (P <0.0001, ANCOVA). Methods The study was conducted in 44 patients with ANP, who were
divided into two groups according to type of analgesia: epidural or opioids. Patients from the fi rst group (n = 23) had epidural analgesia by ropivacaine
6 to 14 mg/hour during 7 to 10 days, and from the second group (n = 21)
opioid analgesia by trimeperidine 20 mg three times a day during the
same period. We monitored the level of septic and thrombohemorrhagic
complications by clinical and instrumental data, during the month after
treatment starting. The hemostatic system was evaluated using indicators
of hemoviscoelastography (Mednord-01M analyzer). g p y
y
Results It was found that all patients with ANP initially have
hypercoagulation and fi brinolysis inhibition. Levels of hemostatic
disorders correlate with the level of septic complications, treatment
in the ICU, and mortality. In the fi rst group we noted a deep vein
thrombosis, two pneumonia, seven pseudopancreatic cysts and
abscesses, two deaths and time of stay in the ICU as 15.4 days. In the
second group: three cases of deep vein thrombosis, four pneumonia, 10
pseudopancreatic cysts and abscesses, two episodes of gastroduodenal
bleeding, fi ve deaths and time of stay in the ICU as 27.8 days. Conclusion Experimental human endotoxemia induces a robust
infl am matory response and a subsequent decrease in serum iron. Treatment with NOX-H94 had no eff ect on innate immunity, but
eff ectively prevented the infl ammation-induced drop in serum iron
concentrations. These fi ndings demonstrate the clinical potential of the
anti-hepcidin drug NOX-H94 for further development to treat patients
with anemia of infl ammation. Hemoviscoelastography: a new method of hemostasis monitoring
O Tarabrin, S Galich, D Gavrychenko, G Mazurenko, A Suhanov
Odessa National Medical University, Odessa, Ukraine
Critical Care 2013, 17(Suppl 2):P355 (doi: 10.1186/cc12293) Introduction The association between haemoglobin concentrations
and mortality has been studied in patients with various comorbidities
[1,2]. This study aims to determine the association between haemo-
globin levels on admission to intensive care and patient length of stay
and mortality. Introduction Deep vein thrombosis of lower extremities and
pulmonary embolism occupy an important place in the structure of
postoperative morbidity and mortality. Introduction Deep vein thrombosis of lower extremities and
pulmonary embolism occupy an important place in the structure of
postoperative morbidity and mortality. y
Methods A retrospective collection of data from patient admissions
to a single fi ve-bed ICU over a 15-year period identifi ed 5,826 patients
between April 1994 and November 2012. Patients were split into groups
according to haemoglobin concentration on admission. The data were
analysed to determine whether there was any relationship between
haemoglobin concentration at ICU admission and any of our outcome
measures (unit and hospital mortality, unit and hospital length of stay). Results Patients with haemoglobin concentrations ≤10 g/dl and
>10.1 g/dl were used in mortality comparisons. Patients with a
haemo globin concentration ≤10 g/dl had an increase in ICU mortality
compared with those with haemoglobin levels >10 g/dl (OR = 1.36, 95%
CI = 1.30 to 1.71, P <0.0001). A similar diff erence was seen with hospital
mortality (≤10 g/dl 37.7% vs. >10 g/dl 27.3%, P <0.0001). Unit length
of stay was signifi cantly longer in patients with admission Hb ≤10 g/
dl (5.34 days) compared with an admission Hb >10 g/dl (4.08 days),
P <0.0001. Hospital length of stay was also signifi cantly longer in
patients with Hb ≤10 g/dl versus Hb >10 g/dl (21.6 days vs. 15.5 days,
P <0.0001). There was seen to be an inverse correlation between
haemoglobin concentration and patient age (r = –0.174; P <0.0001). Methods After ethics approval and informed consent, we studied the
functional state of hemostasis in a group of 57 healthy volunteers,
who were not receiving drugs aff ecting coagulation, and 43 patients
with postphlebothrombotic syndrome (PPTS). In the PPTS patients
we conducted baseline studies of coagulation state and daily
monitoring of dynamic changes in the functional state of hemostasis,
a comparative evaluation of performance low-frequency piezoelectric
vibration hemoviscoelastography (LPVH) and platelet aggregation test
(PAT), standard coagulation tests (SCT), and thromboelastogram (TEG). Results We found that LPVH correlated with SCT, PAT and TEG. P355 Hemoviscoelastography: a new method of hemostasis monitoring
O Tarabrin, S Galich, D Gavrychenko, G Mazurenko, A Suhanov
Odessa National Medical University, Odessa, Ukraine
Critical Care 2013, 17(Suppl 2):P355 (doi: 10.1186/cc12293) P353 Haemoglobin concentration on admission to intensive
care infl uences hospital mortality rates and length of stay:
a retrospective study
M Adams, P Dean, P Doherty, S Noble, A Mackay
Victoria Infi rmary, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P353 (doi: 10.1186/cc12291) P356 P356
Thromboelastographic monitoring of fondaparinux in surgical patients
B Martinez1, R Giacomello1, R Paniccia2
1University Hospital of Udine, Italy; 2University of Florence, Italy
Critical Care 2013, 17(Suppl 2):P356 (doi: 10.1186/cc12294) P356
Thromboelastographic monitoring of fondaparinux in surgical patients
B Martinez1, R Giacomello1, R Paniccia2
1University Hospital of Udine, Italy; 2University of Florence, Italy
Critical Care 2013, 17(Suppl 2):P356 (doi: 10.1186/cc12294) Hemoviscoelastography: a new method of hemostasis monitoring
O Tarabrin, S Galich, D Gavrychenko, G Mazurenko, A Suhanov
Odessa National Medical University, Odessa, Ukraine
Critical Care 2013, 17(Suppl 2):P355 (doi: 10.1186/cc12293) However,
our proposed method is more voluminous: indexes ICC (the intensity
of the contact phase of coagulation), t1 (the time for the contact
phase of coagulation), and AO (initial rate of aggregation of blood)
are consistent with PAT; indexes ICD (the intensity of coagulation
drive), CTA (a constant thrombin activity) and CP (the clot intensity of
the polymerization) are consistent with SCT and TEG. In addition, the
advantage of this method is to determine the intensity of fi brinolysis –
with the indicator IRIS (the intensity of the retraction and clot lysis). Conclusion Haemoglobin concentrations ≤10 g/dl on admission to the
ICU are associated with an increase in ICU mortality, hospital mortality,
unit length of stay and hospital length of stay when compared with
patients admitted with haemoglobin concentrations >10 g/dl. References y
y
Conclusion LPVH allows one to make a total assessment of all parts
of hemostasis: from initial viscosity and platelet aggregation to
coagulation and lysis of clots, as well as their interaction. These fi gures
are objective and informative, as evidenced by close correlation with
the performance of traditional coagulation methods. 1. Zhao Y, et al.: Crit Care 2007, 11:P234. 2. Sekhon MS, et al.: Crit Care 2012, 16:R128. P351 Conclusion Using epidural anesthesia in patients with ANP reduced
the number of septic complications on 36.6%, and reduced the
mortality rate from 23.8% (second group) to 8.7% (fi rst group). We
think that violations of blood coagulation and microcirculation are the
basis for ischemia, necrosis in tissues and septic complications. Epidural
analgesia is an eff ective method to decrease the level of septic and
thrombohemorrhagic complications and mortality in ANP patients. Infl uence of epidural anesthesia on the hemocoagulation disorders
and quantity of septic complications in patients with acute
necrotizing pancreatitis O Tarabrin, S Shcherbakov, A Tkachenko, O Kushnir, I Grychushenko
Odessa National Medical University, Odessa, Ukraine
Critical Care 2013, 17(Suppl 2):P354 (doi: 10.1186/cc12292) O Tarabrin, S Shcherbakov, A Tkachenko, O Kushnir, I Grychushenko
Odessa National Medical University, Odessa, Ukraine
Critical Care 2013, 17(Suppl 2):P354 (doi: 10.1186/cc12292) Introduction Fondaparinux is currently the fi rst therapeutic choice
in several clinical settings such as deep venous thrombosis (DVT) S135 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 8.7 ± 3.4 minutes vs. 12.2 ± 1.9 minutes, P <0.001; α-angle 59.7 ± 9.4 vs. 47.2 ± 11.8, P <0.01). This reduction of fi brinolysis was more evident at
a urokinase concentration of 160 UI/ml (Figure 1). Conclusion UKIF-TEG could be a feasible point-of-care method to
evaluate fi brinolysis in critically ill patients. Reference
1
Gallimore MJ et al : J Thromb Haemost 2005 3:2506-2513
Figure 1 (abstract P357). prophylaxis in orthopedics or in cases of acute coronary syndromes. The main drawback of FOND is that routine monitoring is not currently
available. This could be a problem during the management of critical
and surgical patients, especially in cases of old patients and renal failure. The aim of this study is to evaluate the ability of thromboelastography
(TEG) to determine the level of anticoagulation due to FOND in a
surgical population. prophylaxis in orthopedics or in cases of acute coronary syndromes. The main drawback of FOND is that routine monitoring is not currently
available. This could be a problem during the management of critical
and surgical patients, especially in cases of old patients and renal failure. The aim of this study is to evaluate the ability of thromboelastography
(TEG) to determine the level of anticoagulation due to FOND in a
surgical population. Figure 1 (abstract P357). Methods We prospectively analyzed all patients to whom elective
major orthopedic surgery was consecutively performed in a 2-month
period. All the patients received FOND 2.5 mg in the postoperative
period according to ACCP 2012 Guidelines. Native and heparinase
(hep) TEG (Haemoscope Corporation, Niles, IL, USA) tests activated
with kaolin were performed using whole blood citrated samples
at four times: T0, before FOND administration; T1, 2 hours after
administration; T2, 17 hours after administration (half-life); T3, 24 hours
after administration. The following native and hep TEG parameters
were analyzed: reaction time (R), α angle, maximum amplitude (MA)
and coagulation index (CI). Infl uence of epidural anesthesia on the hemocoagulation disorders
and quantity of septic complications in patients with acute
necrotizing pancreatitis These parameters were compared with
levels of anti-Xa. Unvariate analysis and Spearman’s test were applied
to our data. 8.7 ± 3.4 minutes vs. 12.2 ± 1.9 minutes, P <0.001; α-angle 59.7 ± 9.4 vs. 47.2 ± 11.8, P <0.01). This reduction of fi brinolysis was more evident at
a urokinase concentration of 160 UI/ml (Figure 1). Conclusion UKIF-TEG could be a feasible point-of-care method to
evaluate fi brinolysis in critically ill patients. Reference
1. Gallimore MJ, et al.: J Thromb Haemost 2005, 3:2506-2513. Results Eighteen patients were analyzed. Ten patients met the
inclusion criteria. The mean R value increased from T1 to T3. The mean
R parameter was in the normal range at any phase of the study and
there was no signifi cant diff erences between the R mean value at the
diff erent phases. The lowest value of R was at T1, which coincides with
plasmatic peak concentration of FOND. This value did not correlate with
anti-Xa mean value at T1, which showed the highest value at that time. There was signifi cant diff erence between the mean native and hep R
value only at T2 (P <0.05), native and hep α angle at T3, MA and MA hep
at T2 (P <0.01) and CI and CI hep at T3 (P <0.02). Only the parameter MA
had signifi cant variation over time (P <0.02). 1. Gallimore MJ, et al.: J Thromb Haemost 2005, 3:2506-2513. P358 p
References References 1. Falck-Ytter Y, et al.: Chest 2012, 141:2. 1. Falck-Ytter Y, et al.: Chest 2012, 141:2. 2. FUTURA/OASIS Trial Group, Steg PG, Jolly SS, Mehta SR, et al.: Low Methods In this retrospective study, we investigated 1,737 consecutive
insertions of central venous catheters on 1,444 patients in a large
university hospital during 2009 to 2010. Pre-procedural coagulation
status, blood component use, type of catheter, insertion site, and
complications during insertion were recorded and compared with
bleeding complications documented in electronic charts. 2. FUTURA/OASIS Trial Group, Steg PG, Jolly SS, Mehta SR, et al.: Low-dose vs
standard dose unfractioned heparin for percutaneous coronary
intervention in acute coronary syndromes treated with fondaparinux: the
FUTURA/OASIS randomized trial. JAMA 2010, 304:1339-1349. py
TK Kander, US Schött y
TK Kander, US Schött TK Kander, US Schött Lund University, Lund, Sweden y
Critical Care 2013, 17(Suppl 2):P358 (doi: 10.1186/cc122 Conclusion R represents the time necessary to thrombin formation
and in the presence of FOND we hypothesized a prolonged R time. In
our population, TEG performed with citrated kaolin-activated whole
blood was not able to detect prophylatic doses of FOND in every
phase. On the contrary, levels of anti-Xa were able to reveal the exact
pharmacokinetics of the drug. Further studies including a large number
of patients are necessary. Introduction Bleeding complications in association with insertion of
central venous catheters are reported in 0.5 to 1.6% of cases, are rarely
fatal, but sometimes require intervention. The aim of this study was
to map pre-procedural variables at the insertion of a central venous
catheter and to investigate whether any independent variable could
be identifi ed as an independent risk factor for bleeding complications
associated with insertion of central venous catheters. Urokinase-induced fi brinolysis in thromboelastography (UKIF-TEG)
to assess fi brinolysis in critically ill patients Urokinase-induced fi brinolysis in thromboelastography (UKIF-TEG)
to assess fi brinolysis in critically ill patients
L Zacchetti, M Panigada, M Cressoni, T Marchesi, L Gattinoni
Fondazione IRCCS Ca’ Granda Ospedale Policlinico Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P357 (doi: 10.1186/cc12295) Results No serious bleeding complications were recorded in connection
with the insertions of central venous catheters. Sixteen out of 1,769
(0.9%) insertions caused grade 2 bleeding, being defi ned as a bleeding
requiring prolonged compression at the insertion site. Insertion of a
large-bore central dialysis catheter was found to be an independent risk
factor for bleeding complications. Neither conventional coagulation
tests nor accidental arterial puncture or the number of needle passes
could predict bleeding complications in this study. i
y
y
L Zacchetti, M Panigada, M Cressoni, T Marchesi, L Gattinoni L Zacchetti, M Panigada, M Cressoni, T Marchesi, L Gattinoni Fondazione IRCCS Ca’ Granda Ospedale Policlinico Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P357 (doi: 10.1186/cc12295) Fondazione IRCCS Ca’ Granda Ospedale Policlinico Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P357 (doi: 10.1186/cc12295) p
itical Care 2013, 17(Suppl 2):P357 (doi: 10.1186/cc12295) Introduction Coagulopathy, particularly a trend toward hyper-
coagula bility and hypofi brinolysis, is common in critically ill patients
and correlates with worse outcome. Available laboratory coagulation
tests to assess fi brinolysis are expensive and time demanding. We
investigated whether a modifi ed thromboelastography with the
plasminogen activator urokinase (UKIF-TEG) [1] may be able to evaluate
fi brinolysis in a population of critically ill patients.i Conclusion This retrospective study shows that serious bleeding
complications in association with central line insertion are uncommon
and that insertion of dialysis catheter is an independent risk factor
for mild bleeding complications. Coagulopathy, measured with
conventional coagulation tests, is not an independent risk factor for
bleeding complication at insertion of central venous catheters. i
y
y
Methods UKIF-TEG was performed as follows: fi rst urokinase was
added to citrate blood to give fi nal concentrations of 160 UI/ml, then
thromboelastography (TEG) analysis was started after kaolin activation
and recalcifi cation with calcium chloride. Basal TEG (no addition of
urokinase) was also performed. Fibrinolysis was determined by the
loss of clot strength after the maximal amplitude (MA), and recorded as
Ly30 (percentage lysis at 30 minutes after MA) and as Ly60 (percentage
lysis at 60 minutes after MA). P359
Effi cacy of tranexamic acid in decreasing blood loss during cesarean
section V Zaporozhan, O Tarabrin, D Gavrychenko, G Mazurenko, O Saleh,
I Lyoshenko Results UKIF-TEG was performed on 17 healthy volunteers and 18
critically ill patients. Ly60 was predicted by Ly30 according to an
exponential function, so we used Ly30 as an indicator of clot lysis. Basal
TEG showed increased coagulability and a trend toward less fi brinolysis
in critically ill patients compared with healthy volunteers (reaction time Introduction Despite signifi cant progress in obstetric care, the
problem of bleeding during labor remains unsolved. Annually in the
world, 125,000 women die from obstetric hemorrhage. Introduction Despite signifi cant progress in obstetric care, the
problem of bleeding during labor remains unsolved. Annually in the
world, 125,000 women die from obstetric hemorrhage. S136 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods We performed a randomized, double-blind study in 37
patients who underwent cesarean section. Patients were divided into
two groups: the fi rst group (n = 19) received preoperative (30 minutes
before operation) tranexamic acid 10 mg/kg; the second group (n = 18)
received preoperative placebo. The condition of hemostasis was
monitored by haemoviscoelastography. Methods High and low concentrations of both fi brinogen and FXIII
were used to reverse coagulopathy induced by 1:1 dilution in vitro with
5% albumin of blood samples from healthy volunteers, monitored by
rotational thromboelastometry (ROTEM).i y (
)
Results Haemodilution with albumin signifi cantly attenuated EXTEM
maximum clot fi rmness (MCF), α angle (AA), clotting time (CT) and
clot formation time (CFT), and FIBTEM MCF (P <0.001). Following
haemodilution, both doses of fi brinogen signifi cantly corrected all
ROTEM parameters (P ≤0.02), except the lower dose did not correct
AA. Compared with the lower dose, the higher dose of fi brinogen
signifi cantly improved FIBTEM MCF and EXTEM MCF, AA and CFT
(P <0.001). The lower dose of FXIII did not signifi cantly correct any of
the ROTEM parameters, and the high dose only improved EXTEM CT
(P = 0.004). All combinations of high/low concentrations of fi brinogen/
FXIII signifi cantly improved all ROTEM parameters examined (P ≤0.001). Fibrinogen concentration generally had a greater eff ect on each
parameter than did FXIII concentration; the best correction of ROTEM
parameters was achieved with high-dose fi brinogen concentrate and
either low-dose or high-dose FXIII. 1Hôpital Jeanne de Flandre, Lille, France; 2Centre hospitalier regional
universitaire Lille, France Critical Care 2013, 17(Suppl 2):P362 (doi: 10.1186/cc12300) Critical Care 2013, 17(Suppl 2):P362 (doi: 10.1186/cc12300) Introduction Postpartum hemorrhage (PPH) remains the leading
cause of maternal mortality. After IOCS introduction in our north of
France tertiary-care obstetric unit, the consecutive cohort of severe
PPH was collected prospectively and compared regarding the use of
IOCS or not. y
Methods Citrated venous blood from 10 healthy volunteers was
diluted by 33% using 130/0.42 hydroxyethyl starch or Ringer’s acetate. ROTEM was used to evaluate the eff ect of addition of either fi brinogen
concentrate corresponding to 4 g/70 kg, or this fi brinogen dose
combined with factor XIII equivalent to 20 IU/kg. Blood was analyzed at
33 or 37°C with ROTEM ExTEM and FibTEM reagents.i Methods A case–control prospective open-label study comparing the
management and outcome of a consecutive cohort of 324 severe PPHs
in elective and emergency hemorrhagic caesarean sections (CS) over
3 years. Results A signifi cant dilutive response was shown in both groups:
hypocoagulation was greater in the starch group. Hypothermia
lengthened the following: ExTEM clotting time (CT), clot formation
time and α angle; FibTEM maximal clot formation (MCF). Irrespective
of temperature, fi brinogen overcorrected Ringer’s acetate’s eff ects on
all ROTEM parameters and partially reversed starch’s eff ects on ExTEM
CT and FibTEM MCF. FibTEM demonstrated that factor XIII provided an
additional procoagulative eff ect in the Ringer’s acetate group at both
temperatures but not the starch group. The only ExTEM parameter to
be improved by addition of factor XIII was MCF at 33°C.i Results IOCS was used in 70 severe PPHs and 254 severe PPH controls
were managed without IOCS. Placenta accreta can be selected as
the best indication for RBC restitution. In the 1,500 to 3,000 ml PPH,
allogeneic transfusion was decreased in the IOCS group: 17.6 versus
56.3% (P = 0.006); PRBC: 0 (0 to 3) versus 3 (0 to 6) (P = 0.045). IOCS
spared 87 blood bank PRBC (17,374 ml); that is, 24.2% of the total
transfusion need. No amniotic fl uid embolism has been observed in
the group with IOCS whereas one case appeared in the control group
without IOCS. Conclusion Regarding the literature [1-4] and our study, IOCS could be
used safely in PPH during CS. A leukocyte fi lter for retransfusion has
been recommended and Rhesus isoimmunization must be precluded
and monitored by repeated fetal RBC testing. In vitro correction of hypothermic and dilutive crystalloid and
colloid rotational thromboelastography-monitored coagulopathy
with fi brinogen and factor XIII i
g
D Winstedt, O Thomas, US Schött Intraoperative cell savage for life-threatening haemorrhagic
caesarean section Lund University, Lund, Sweden AS Ducloy1, C Barre-Drouard2, E Jaillette2, S Depret-Mosser2, B Wibaut2,
B Vallet2, S Susen2 1Hôpital Jeanne de Flandre, Lille, France; 2Centre hospitalier regional
universitaire Lille, France References y
Conclusion ROTEM shows that fi brinogen concentrate can reverse
dilutive coagulation defects induced by colloid and crystalloid at both
33 and 37°C. Some additional reversal was provided by factor XIII:
higher doses of both fi brinogen and factor XIII may counteract starch’s
eff ects on clot structure. 1. Rainaldi et al.: Br J Anaesth 1998, 80:195-198. P361 2. Bernstein et al.: Anesth Analg 1997, 85:831-833. Dose–response of fi brinogen and factor XIII concentrate for
correcting albumin-induced coagulopathy
US Schött, DW Winstedt, JH Hanna
Lund University, Lund, Sweden
Critical Care 2013, 17(Suppl 2):P361 (doi: 10.1186/cc12299) Dose–response of fi brinogen and factor XIII concentrate for
correcting albumin-induced coagulopathy
US Schött, DW Winstedt, JH Hanna
Lund University, Lund, Sweden
Critical Care 2013, 17(Suppl 2):P361 (doi: 10.1186/cc12299) 3. Catling et al.: Int J Obstet Anesth 1999, 8:79-84. g
4. Waters et al.: Anesthesiology 2000, 92:1531-1536. P359
Effi cacy of tranexamic acid in decreasing blood loss during cesarean
section y
g
p y
Results All patients included in the study before surgery had moderate
hypercoagulation and normal fi brinolysis: increasing the intensity of
clot formation (ICF) to 11.4% compared with normal rates; the intensity
of the retraction and clot lysis (IRCL) was 16.45 ± 1.40 in both groups. At the start of the operation in patients (Group 1), ICF decreased by
9.7% (P <0.05), and IRCL decreased by 27.6% (P <0.05) compared with
preoperatively. In Group 2, there was ICF decrease by 8.8% (P <0.05), and
IRCL increase by 11.4% (P <0.05) compared with preoperatively. At the
end of the operation, the condition of hemostasis in both groups came
almost to the same value – moderate hypocoagulation, depressed
fi brinolysis. In both groups there were no thrombotic complications. Intraoperative blood loss in the fi rst group was 300 ± 40.5 and in the
second was 500 ± 60.6.i Conclusion Using of tranexamic acid before surgery signifi cantly reduces
intraoperative blood loss by 60%, without thrombotic complications. Conclusion Fibrinogen concentrate successfully corrected initiation,
propagation and clot fi rmness defi cits induced by haemodilution with
albumin, and FXIII synergistically improved fi brin-based clot strength. Reference AS Ducloy1, C Barre-Drouard2, E Jaillette2, S Depret-Mosser2, B Wibaut2,
B Vallet2, S Susen2 Introduction Rotational thromboelastography (ROTEM) can detect
dilutive and hypothermic eff ects on coagulation and evaluate
corrective treatments. The aim of this in vitro study was to study
whether fi brinogen concentrate alone or combined with factor XIII
could reverse colloid-induced and crystalloid-induced coagulopathies
in the presence and absence of hypothermia. 1Hôpital Jeanne de Flandre, Lille, France; 2Centre hospitalier regional
universitaire Lille, France In vitro correction of hypothermic and dilutive crystalloid and
colloid rotational thromboelastography-monitored coagulopathy
with fi brinogen and factor XIII
D Winstedt, O Thomas, US Schött
Lund University, Lund, Sweden
Critical Care 2013, 17(Suppl 2):P360 (doi: 10.1186/cc12298) 1. Anesth Analg 2008, 106:1360-1365. P363 primary composite endpoint – all-cause mortality, cardiovascular
complications, acute respiratory distress syndrome, acute kidney injury
requiring renal replacement therapy, septic shock or reoperation at
30 days – occurred in 19.6% of the patients in the liberal strategy group
and in 35.6% in the restrictive group (P = 0.012). The liberal strategy
group had a signifi cantly lower 30-day mortality rate as compared with
the restrictive group (8.2% (95% CI, 4.2 to 15.4%) vs. 22.8% (95% CI, 15.7
to 31.9%), respectively, P = 0.005). The occurrence of cardiovascular
complications was lower in the liberal group than in the restrictive
group (5.2% (95% CI, 2.2 to 11.5%) vs. 13.9% (95% CI, 8.4 to 21.9%),
respectively, P = 0.038). The restrictive strategy group had a higher 60-
day mortality rate as compared with the liberal group (23.8% (95% CI,
16.5 to 32.9%) vs. 11.3% (95% CI, 6.5 to 9.2%), respectively, P = 0.022). Conclusion The liberal RBC transfusion strategy with a hemoglobin
trigger of 9 g/dl was associated with fewer major postoperative
complications in patients undergoing major cancer surgery compared
with the restrictive strategy. Methods Data from 240 cardiac surgery patients were retrospectively
analysed. Of these, 157 had routine management with a bolus of
protamine to correct the activated clotting time and then expectant
management of subsequent bleeding, and 47 had the same but also
a protamine infusion of 10 to 80 mg/hour for between 3 and 8 hours
postoperatively. Blood loss was measured at 1, 6, 12 and 24 hours. In
all, excessive bleeding was investigated using thromboelastography
(TEG). Rebound heparinisation was determined by a ratio of R-times
(heparinase/plain) <0.8. The Mann–Whitney U test and the chi-squared
test were used to assess statistical signifi cance.if p
y
Conclusion The liberal RBC transfusion strategy with a hemoglobin
trigger of 9 g/dl was associated with fewer major postoperative
complications in patients undergoing major cancer surgery compared
with the restrictive strategy. gi
Results There was no signifi cant diff erence in blood loss between the
two groups. Blood loss at 1 hour in the infusion and non-infusion group
was 145 and 88 ml, respectively (P = 0.06); at 6 hours: 450 and 392 ml
(P = 0.5); at 12 hours: 620 and 595 ml (P = 0.62); and at 24 hours: 971
and 872 ml (P = 0.12). P364 P364
Transfusion Requirements in Surgical Oncology Patients (TRISOP):
a randomized, controlled trial
J Almeida1, F Galas1, E Osawa1, J Fukushima1, S Moulin1, C Park1,
E Almeida1, S Vieira1, J Vincent2, A Rhodes3, M Balzan1, J Inacio1,
H Palomba1, R Nakamura1, F Bergamin1, A Sandrini1, U Ribeiro Jr1,
J Auler Jr1, L Hajjar1
1Instituto do Cancer do Estado de São Paulo, Brazil; 2Erasme Hospital,
Université libre de Bruxelles, Brussels, Belgium; 3St George’s Healthcare NHS
Trust and St George’s University of London, UK
Critical Care 2013, 17(Suppl 2):P364 (doi: 10.1186/cc12302) p
Results One hundred and thirty-six patients (66.3%) were exposed
to RBC transfusion. In the intraoperative room, 63.4% of patients
received at least one RBC unit, and in the ICU, 11.2% of children were
transfused. From all patients, 66 (32.1%) presented the combined
endpoint. Patients with complications had higher RACHS-1 score, were
younger (69 months (0 to 137) vs. 73 months (37 to 138), P <0.001),
had a lower weight (13 kg (3 to 23) vs. 20 kg (12 to 36), P <0.001), a
longer time of surgery (475 minutes (410 to 540) vs. 353 (275 to 433),
P <0.001), a longer duration of cardiopulmonary bypass (205 minutes
(175 to 235) vs. 106 minutes (73 to 123), P = 0.003), a lower SVO2 at
the end of surgery (59% (IQR 39 to 80) vs. 78% (71 to 83), P <0.001), a
higher arterial lactate at the end of surgery (6.9 mmol/l (4.3 to 9.2) vs. 2.7 mmol/l (73 to 123), P = 0.003), a lower intraoperative hematocrit
(26.2 ± 5.6% vs. 29.5 ± 6% (P <0.001)) and a lower hematocrit at the end
of surgery (33.4 ± 6.7% vs. 36.9 ± 6.9% (P <0.001)) as compared with
patients without complications. Patients with complications were more
exposed to RBC transfusion in the intraoperative room (75% vs. 57%,
P = 0.011) and in the ICU (21% vs. 6.4%, P = 0.002). In an adjusted model
of logistic regression, RBC transfusion is an independent risk factor of
combined endpoint (OR 4.25 (95% CI, 1.359 to 13.328), P = 0.013). Conclusion Blood transfusion after pediatric cardiac surgery is a risk
factor for worse outcome including 30-day mortality. P363 Does protamine infusion reduce postoperative blood loss and
heparin rebound in cardiac surgery patients? A retrospective
analysis of 240 patients on the cardiac ICU p
p
p
heparin rebound in cardiac surgery patients? A retrospectiv Introduction Natural colloid albumin induces a lesser degree of
dilutional coagulopathy than synthetic colloids. Fibrinogen concentrate
has emerged as a promising strategy to treat coagulopathy, and factor
XIII (FXIII) works synergistically with fi brinogen to correct coagulopathy
following haemodilution with crystalloids. Objectives were to examine
the ability of fi brinogen and FXIII concentrates to reverse albumin-
induced dilutional coagulopathy. analysis of 240 patients on the cardiac ICU E Helme, H Meeran, N Al-Subaie St George’s Hospital, London, UK g
p
Critical Care 2013, 17(Suppl 2):P363 (doi: 10.1186/cc12301) Introduction Cardiac surgery is associated with signifi cant blood
loss. Teoh and colleagues demonstrated a reduction in postoperative S137 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 primary composite endpoint – all-cause mortality, cardiovascular
complications, acute respiratory distress syndrome, acute kidney injury
requiring renal replacement therapy, septic shock or reoperation at
30 days – occurred in 19.6% of the patients in the liberal strategy group
and in 35.6% in the restrictive group (P = 0.012). The liberal strategy
group had a signifi cantly lower 30-day mortality rate as compared with
the restrictive group (8.2% (95% CI, 4.2 to 15.4%) vs. 22.8% (95% CI, 15.7
to 31.9%), respectively, P = 0.005). The occurrence of cardiovascular
complications was lower in the liberal group than in the restrictive
group (5.2% (95% CI, 2.2 to 11.5%) vs. 13.9% (95% CI, 8.4 to 21.9%),
respectively, P = 0.038). The restrictive strategy group had a higher 60-
day mortality rate as compared with the liberal group (23.8% (95% CI,
16.5 to 32.9%) vs. 11.3% (95% CI, 6.5 to 9.2%), respectively, P = 0.022). Conclusion The liberal RBC transfusion strategy with a hemoglobin
trigger of 9 g/dl was associated with fewer major postoperative
complications in patients undergoing major cancer surgery compared
with the restrictive strategy. bleeding with the use of a protamine infusion and an abolishment of
heparin rebound [1]. The aim of this study was to see whether the use
of postoperative protamine infusions in our cardiac ITU was associated
with a reduction in heparin rebound and blood loss. 1.
Teoh K, et al.: J Thorac Cardiovasc Surg 2004, 128:211-219.
2.
Teoh K, et al.: Circulation 1993, 88:420-425. References 1. Teoh K, et al.: J Thorac Cardiovasc Surg 2004, 128:211-219. 2. Teoh K, et al.: Circulation 1993, 88:420-425. y
Methods We performed a prospective cohort study of 205 patients
undergoing cardiac surgery for congenital heart disease. We
recorded baseline characteristics, RACHS-1 score, intraoperative data,
transfusion requirement and severe postoperative complications as
need for reoperation, acute kidney injury, arrhythmia, severe sepsis,
septic shock, bleeding, stroke, and death during 30 days. We performed
univariate analysis using baseline, intraoperative and postoperative
variables. Selected variables (P <0.10) were included in a forward
stepwise multiple logistic regression model to identify predictive
factors of a combined endpoint including 30-day mortality and severe
complications. Conclusion Blood transfusion after pediatric cardiac surgery is a risk
factor for worse outcome including 30-day mortality. Avoiding blood
transfusion must be a goal of best postoperative care. P365 Red blood cell transfusion is a predictor of mortality and morbidity
in children undergoing cardiac surgery
C Colognesi, R Maia, L Hajjar, F Bergamin, J Fukushima, E Osawa,
J Almeida, L Camara, S Zeferino, V Guimaraes, F Galas
Heart Institute, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P365 (doi: 10.1186/cc12303) Introduction Red blood cell (RBC) transfusion is associated with
morbidity and mortality in critically ill patients. Congenital cardiac
surgeries are associated with high rates of bleeding and consequently
with high rates of allogeneic transfusion. We aimed to evaluate the
association of transfusion with worse outcomes in children undergoing
cardiac surgery. P363 There was also no signifi cant diff erence in those
getting heparin rebound with 40% in the infusion group and 47% in
the non-infusion group (P = 0.54). gy
Acknowledgement ClinicalTrials.gov NCT 01560494. Acknowledgement ClinicalTrials.gov NCT 01560494. g
Conclusion Unlike Teoh and colleagues [1], we did not fi nd an
advantage in using protamine infusions. That there were still cases
of heparin rebound in the infusion group suggests that the infusion
was not as eff ective as expected and/or the dose was inadequate. However, previous studies assessed heparin rebound using isolated
clotting parameters [1,2]. Here, we used TEG. As TEG measures the
thrombodynamic properties of whole blood coagulation, perhaps it is
a more reliable indicator of heparin activity? As a retrospective study,
there are limitations; namely, the nonstandardised management of the
patients and the potential bias in the anaesthetists’ selection of patients
for an infusion. This group may be inherently higher risk for bleeding. However, heparin rebound is common and protamine is a simple,
relatively safe and low-cost intervention compared with transfusion
and so further study is needed. 1.
Teoh K, et al.: J Thorac Cardiovasc Surg 2004, 128:211-219.
2
Teoh K et al : Circulation 1993 88:420-425 P364 Avoiding blood
transfusion must be a goal of best postoperative care Introduction The purpose of this study was to evaluate whether a
restrictive strategy of red blood cell (RBC) transfusion was superior to
a liberal one for reducing mortality and severe clinical complications
among patients undergoing major cancer surgery. Introduction The purpose of this study was to evaluate whether a
restrictive strategy of red blood cell (RBC) transfusion was superior to
a liberal one for reducing mortality and severe clinical complications
among patients undergoing major cancer surgery. Methods The trial was designed as a phase III, randomized, controlled,
parallel-group, superiority trial. The inclusion criteria were adult patients
with cancer who were undergoing major abdominal surgery requiring
postoperative care in an ICU. The patients were randomly allocated to
treatment with either a liberal RBC transfusion strategy (transfusion
when hemoglobin levels decreased below 9 g/dl) or a restrictive RBC
transfusion strategy (transfusion when hemoglobin levels decreased
below 7 g/dl). The primary outcome was a composite endpoint of death
or severe complications. The patients were monitored for 30 days. Results A total of 1,521 patients were screened for eligibility and 248
met the inclusion criteria. After exclusions for medical reasons or a
lack of consent, 198 patients were included in fi nal analysis, with 101
allocated to the restrictive group and 97 to the liberal group. The Conclusion Blood transfusion after pediatric cardiac surgery is a risk
factor for worse outcome including 30-day mortality. Avoiding blood
transfusion must be a goal of best postoperative care. S138 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 patients in the MT group (23.7%; 27/114) and 87 in the non-MT group. The MT group was signifi cantly higher in the ratio of females (P <0.001),
ISS (P <0.01), PT-INR (P <0.001), APTT (P <0.05), ABCs (P <0.001) and
TASHs (P <0.001) than in the on-MT group. On the other hand, the
MT group was signifi cantly lower in Ps (P <0.05) and fi brinogen
level (P <0.001) than the non-MT group. In the receiver operating
characteristics (ROC) analysis, the area under the curve (AUC) to
distinguish a MT was the highest for TASHs (0.831, P <0.001), followed
by fi brinogen (0.758, P <0.001), and ABCs (0.732, P <0.001). P367i i
Results During the study period, 913 admissions were documented,
843 of which were included in the study. Among them, 144 (17%) were
transfused. When comparing TCs with nontransfused cases (NTCs),
the odds ratio (OR) of new or progressive multiple organ dysfunction
syndrome (NPMODS) was 2.39 (95% CI = 1.58 to 3.62, P <0.001). This
association remained statistically signifi cant in the multivariate analysis
for children with admission PRISM score ≤5 (OR = 2.41, 95% CI = 1.08 to
5.37, P = 0.032). TCs were ventilated longer than NTCs (14.1 ± 32.6 days
vs. 4.3 ± 9.6 days, P <0.001). This diff erence was still signifi cant after
adjustment using a Cox model. Moreover, we observed an adjusted
dose–eff ect relationship between RBC transfusions and length of
mechanical ventilation. The PICU length of stay was signifi cantly
increased for TCs (12.4 ± 26.2 days vs. 4.9 ± 10.2 days, P <0.001), even
after multivariate adjustment (hazard ratio of PICU discharge for TCs:
0.61, 95% CI = 0.5 to 0.74, P <0.001). We also observed an adjusted
dose–eff ect relationship between RBC transfusions and PICU length of
stay. The paired analysis for comparison of pre-transfusion and post-
transfusion values showed that the arterial partial pressure in oxygen
was signifi cantly reduced after the fi rst transfusion (mean diff erence
42.8 mmHg, 95% CI = 27.2 to 58.3, P <0.001). The paired analysis also
showed an increased proportion of renal replacement therapy, while
the proportions of sepsis, severe sepsis and septic shock did not diff er. Conclusion RBC transfusions were associated with prolonged
mechanical ventilation and prolonged PICU stay. The risk of NPMODS
was increased in some transfused children. Moreover, our study
questions the ability of stored RBCs to improve oxygenation in critically
ill children. These results should help to improve transfusion practice
in the PICU. P364 Fibrinogen
was only a predictor of a MT without a scoring system such as ABCs and
TASHs, and the optimal cutoff value was 205 mg/dl. P366 P366
Red blood cell transfusion results in worse outcomes in patients
with left ventricular dysfunction undergoing on-pump cardiac
surgery
R Miadda, C Colognesi, L Hajjar, M Sundin, L Camara, S Zeferino,
F Bergamin, A Leme, V Guimaraes, F Galas
Heart Institute, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P366 (doi: 10.1186/cc12304) Introduction Cardiac surgeries are associated with high rates of
bleeding and consequently with high rates of allogeneic transfusion. Patients with left ventricular dysfunction usually present a higher
incidence of complications after cardiac surgery, including low output
syndrome and organ dysfunction. It is a matter of controversy whether
this subgroup of patients need higher hemoglobin levels to optimize
oxygen delivery. We aimed to evaluate the association of transfusion
with worse outcomes in patients with left ventricular dysfunction. pf
g
Conclusion We found that the level of fi brinogen was the most
valuable predictor of a MT in the coagulation or fi brinolysis markers. It is certain that the level of fi brinogen at admission was not as useful
as the TASHs about predicting a MT in this study. Whereas the scoring
systems require the assessment of several factors, the measurement
of fi brinogen is simple, easy and quick. We strongly suggest that the
level of fi brinogen will be a useful predictor of a MT at in severe blunt
traumatic patients. p
y
Methods A prospective cohort study was conducted in 215 patients
undergoing cardiac surgery, presenting ejection fraction <35%. We
recorded baseline characteristics, intraoperative data, transfusion
requirement and severe postoperative complications as need for
reoperation, acute kidney injury, arrhythmia, severe sepsis, septic
shock, bleeding, stroke, and death during 30 days. We performed
univariate analysis using baseline, intraoperative and postoperative
variables associated with severe postoperative complications. Selected
variables (P <0.10) were included in a forward stepwise multiple logistic
regression model to identify predictive factors of a combined endpoint
including 30-day mortality and severe complications.i p
References 1. Timothy C, et al.: J Trauma 2009, 66:346-352. 2. Yücel N, et al.: J Trauma 2006, 60:1228-1236. 3. Sørensen B, et al.: Semin Thromb Hemost 2012, 38:268-273. 1. Timothy C, et al.: J Trauma 2009, 66:346-352. 1. Timothy C, et al.: J Trauma 2009, 66:346-352. 2. Yücel N, et al.: J Trauma 2006, 60:1228-1236. 3. Sørensen B, et al.: Semin Thromb Hemost 2012, 38:268-273. 1. Timothy C, et al.: J Trauma 2009, 66:346 352. 2. Yücel N, et al.: J Trauma 2006, 60:1228-1236. 3. Sørensen B, et al.: Semin Thromb Hemost 2012, 38:268-273. 2. Yücel N, et al.: J Trauma 2006, 60:1228-1236. 3. Sørensen B, et al.: Semin Thromb Hemost 2012, 38:268-273. Association between red blood cell transfusions and clinical
outcome in critically ill children y
P Demaret1, M Tucci2, T Ducruet2, H Trottier2, J Lacroix2 Results Two hundred and fi fteen patients were included in the study. The mean ejection fraction was 31%. From all patients, 129 (54.4%)
presented the combined endpoint. The incidence of RBC transfusion
was higher in the group with complications compared with the
group without complications both intraoperative and postoperative,
P <0.001. Patients with complications were older, had lower initial
hemoglobin levels and had higher EuroSCORE. At multivariate analysis,
age (OR = 1.07, 95% CI = 1.013 to 1.137, P = 0.016) and perioperative
RBC transfusion (OR = 7.78, 95% CI = 2.13 to 28.41, P = 0.002) were
independent predictors of severe postoperative complications after
cardiac surgery. Introduction Red blood cell (RBC) transfusions are frequent in critically
ill children. Their benefi ts are clear in several situations. However,
issues surrounding their safety have emerged in the past decades. It is
important to identify the potential complications associated with RBC
transfusions, in order to evaluate their risk–benefi t ratio better. i
Methods A single-center prospective observational study of all
children admitted to the pediatric intensive care unit (PICU) over a
1-year period. The variables possibly related to RBC transfusions were
identifi ed before the study was initiated, and their presence was
assessed daily for each child. In transfused cases (TCs), a variable was
considered as a possible outcome related to the transfusion only if it
was observed after the fi rst transfusion. g
y
Conclusion In patients with left ventricular dysfunction, red blood
transfusion after cardiac surgery is a risk factor for worse outcome
including 30-day mortality. Defi ning the need for a massive transfusion in severe blunt
traumatic patients T Umemura, H Ishikura, Y Nakamura, K Hoshino, T Nishida, T Kamitani
Fukuoka University, Fukuoka, Japan
Critical Care 2013, 17(Suppl 2):P367 (doi: 10.1186/cc12305) Introduction We do not have enough criteria to make a judgment
of the need for a massive transfusion (MT) in severe blunt traumatic
patients. As a scoring system to predict the need for a MT, we usually
use the Assessment Blood Consumption score (ABCs) and/or the
Trauma-Associated Severe Hemorrhage score (TASHs). However, for
these scoring systems, the procedure is slightly complicated. The aim
of this study was to establish a predictor of a MT using coagulation or
fi brinolysis markers. i
Methods A retrospective analysis of MT was conducted in patients
with severe blunt traumatic injury, which was defi ned as Injury Severity
Score (ISS) of 16 or more admitted to the ICU between 1 June 2009
and 31 December 2010. Blood samples were collected from patients
immediately after arriving at our level I trauma center. We defi ned
the patients who received more than 10 unit packed red blood cells
(PRBCs) within the fi rst 24 hours as a MT group and who received less
than 9 units PRBCs as a non-MT group. After the demographic data,
number of units of PRBCs and the need for massive transfusions were
recorded and analyzed in each groups, we compared data between
two groups. Results There were 114 patients who met the inclusion criteria. Fifty
patients received blood transfusions (43.9%; 50/114). There were 27 Critical Care 2013, Volume 17 Suppl 2
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Microcirculation and blood transfusions during sepsis:
leukoreduced (LR) versus non-LR red blood cells
A Donati1, E Damiani1, R Domizi1, C Scorcella1, A Carsetti1, S Tondi1,
R Castagnani1, N Mininno1, P Pelaia1, C Ince2
1Università Politecnica delle Marche, Ancona, Italy; 2Academic Medical Center,
Amsterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P369 (doi: 10.1186/cc12307) P369 density and fl ow were assessed with sidestream dark-fi eld imaging
sublingually. Thenar tissue O2 saturation (StO2) was measured
using near-infrared spectroscopy and a vascular occlusion test was
performed. Results The De Backer score (P = 0.02), total vessel density (P = 0.08),
perfused vessel density (P = 0.04), proportion of perfused vessels
(P = 0.01), and microvascular fl ow index (P = 0.04, Figure 1) increased
only in Group 2. The StO2 upslope (Figure 2) during reperfusion
increased in both groups (P <0.05). Massive blood transfusion for obstetric haemorrhage S Simmons1, WE Pollock1, L Phillips2, S McDonald3 p
1Mercy Hospital for Women, Heidelburg, Australia; 2Monash University,
Prahran, Australia; 3La Trobe University, Bundoora, Australia
Critical Care 2013, 17(Suppl 2):P370 (doi: 10.1186/cc12308) Figure 1 (abstract P369). Blood transfusion and microvascular fl ow. Prahran, Australia; 3La Trobe University, Bundoora, Australia
Critical Care 2013, 17(Suppl 2):P370 (doi: 10.1186/cc12308) Introduction Obstetric haemorrhage remains a leading cause
of maternal mortality and severe morbidity. Cardiovascular and
haemostatic physiology alters in pregnancy and massive transfusion
protocols have been implemented for obstetric haemorrhage based
on limited evidence. The objective of this study was to examine the
pattern and rate of blood products used in massive transfusion for
obstetric haemorrhage in a tertiary obstetric hospital. Methods Massive transfusion was defi ned as 5 or more units of red
blood cells within 4 hours in accordance with the Australian Massive
Transfusion Registry defi nition. Following ethics approval, all cases
fi lling this criterion were identifi ed in the hospital’s birthing and blood
bank systems. Data were extracted from the medical histories and
analysed using SPSS. P <0.05 was considered statistically signifi cant. i
Results Twenty-eight women in three years (2009 to 2011) underwent
a massive transfusion for obstetric haemorrhage, with nine receiving
more than 10 units of RBCs in 24 hours. Eleven (39%) were admitted to
the ICU and 11 underwent a hysterectomy, of which six were admitted
to the ICU. The median estimated blood loss was 4,335 ml (IQR 3,000
to 5,000). Median blood product delivery was RBC 8 units (IQR 6 to 13);
FFP 4 units (IQR 4 to 8); platelets 4 units (IQR 0 to 8) and cryoprecipitate
3 units (IQR 0 to 10). One-half of the women received the fi rst four units
of RBCs in less than 34 minutes. Other blood products were started
a median of 49 minutes, 44 minutes and 75 minutes after the RBC
transfusion commenced, respectively. Eight women had a fi brinogen
level <0.8 g/l on the initial coagulation test during the haemorrhage. The remaining 20 women had a median fi brinogen level of 3.7 g/l (IQR
3.15 to 4.9). There was no diff erence in the transfusion of RBCs (P = 0.20),
FFPs (P = 0.96) and platelets (P = 0.48) in women who showed an initial
low fi brinogen and those who did not, although there was a diff erence
in the number of units of cryoprecipitate (P <0.05). Massive blood transfusion for obstetric haemorrhage The median lowest
Hb during the haemorrhage was 66 g/l (IQR 51 to 80) and median
discharge Hb was 103 g/l (IQR 96 to 113). No blood product reaction
was noted and there was one death. Figure 1 (abstract P369). Blood transfusion and microvascular fl ow. Figure 2 (abstract P369). Blood transfusion and microvascular reactivity. Conclusion Massive transfusion for obstetric haemorrhage involved
rapid blood product administration with no consistent pattern in the
ratio of products administered. Defi ning the need for a massive transfusion in severe blunt
traumatic patients In Group 1 the baseline StO2 and
StO2 downslope during ischemia increased, probably refl ecting a lower
O2 consumption. Introduction Microcirculatory alterations during sepsis impair tissue
oxygenation, which may be further worsened by anemia. Blood
transfusions proved not to restore O2 delivery during sepsis [1]. The
impact of storage lesions and/or leukocyte-derived mediators in red
blood cell (RBC) units has not yet been clarifi ed [2]. We compared
the eff ects of leukoreduced (LR) versus nonLR packed RBCs on
microcirculation and tissue oxygenation during sepsis. 2
p
Conclusion Unlike nonLR RBCs, the transfusion of fresh LR RBCs seems
to improve microvascular perfusion and might help to restore tissue
oxygenation during sepsis. yg
References 1. Marik PE, et al.: JAMA 1993, 269:3024-3029. Methods A prospective randomized study. Twenty patients with
either sepsis, severe sepsis or septic shock requiring RBC transfusion
randomly received nonLR (Group 1, n = 10) or LR (Group 2, n = 10) fresh
RBCs (<10 days old). Before and 1 hour after transfusion, microvascular 2. Sparrow RL: Blood Transfus 2010, 8(Suppl 3):s26-s30. 3
Blood transfusion practices in the ICU of a level 1 trauma centre and
tertiary cardiac unit P373 P373
Incidence, risk factors and outcome of TRALI after cardiac surgery
A Dijkhuizen, R De Bruin, S Arbous
Leiden University Medical Center, Leiden, the Netherlands
Critical Care 2013, 17(Suppl 2):P373 (doi: 10.1186/cc12311) Introduction Transfusion-related acute lung injury (TRALI) is a
syndrome that presents as a sudden onset of respiratory distress 6
hours after transfusion of blood products. The diagnosis is based on
clinical and radiographic fi ndings. Particularly at risk for TRALI are
cardiac surgery patients. However, specifi c patient risk factors and
data on outcome are largely unknown. The aim of this study was to
investigate incidence, risk factors and outcome of TRALI in cardiac
surgical patients on cardiopulmonary bypass. g
p
p
Results A total 175 units of PRC were transfused in the ICU, over
105 episodes during the 2-month period (excluding immediately
postoperative transfusions). Ninety-four units (53.7%) administered in
64 transfusion episodes (61.0%) occurred contrary to the guidelines. In 89.3% of these cases the recorded reason for transfusion was an
apparently low Hb level. The median (IQR (range)) Hb level at which
patients were transfused: within guidelines was 6.9 g/dl (6.6 to 7.7 (4.8
to 13.9)); within guidelines, excluding cases of acute blood loss, was
6.6 g/dl (6.5 to 6.8 (5.5 to 6.9)); and outside the guidelines was 7.7 g/dl
(7.4 to 8.2 (7.0 to 9.7)). One unit of PRC was transfused in 57 episodes
(54.3%), 2 units of PRC were transfused in 36 episodes (34.3%), and 3 to
6 units were transfused in 12 episodes (11.4%), with two-thirds of the
latter due to acute haemorrhage. g
y y
Methods All thoracic surgery patients from a university hospital in the
Netherlands of 18 years and older admitted to the ICU from January
2009 until December 2011 were screened. Included patients were
observed during surgery and the fi rst 24 hours on the ICU for the onset
of possible TRALI. The Canadian Consensus Conference TRALI defi nition
was used. Two independent physicians blinded to the predictor
variables scored the chest radiographs for the onset of bilateral
interstitial abnormalities on 2K monitors. When interpretation diff ered,
chest radiographs were reviewed by a third physician to achieve
consensus. The European System for Cardiac Operative Risk Evaluation
(EURO score) and the American Association of Anesthesiology (ASA)
were scored before surgery. 3
Blood transfusion practices in the ICU of a level 1 trauma centre and
tertiary cardiac unit Y Mustafa, B Pouchet
University Hospitals Birmingham NHS Foundation Trust, Birmingham, UK
Critical Care 2013, 17(Suppl 2):P371 (doi: 10.1186/cc12309) Introduction Blood transfusions are associated with longer ICU and
hospital inpatient durations, and an increase in mortality [1]. This study
was undertaken to investigate whether the practice of packed red cell Figure 2 (abstract P369). Blood transfusion and microvascular reactivity. S140 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 4.9 ± 0.5 (P = 0.09). Cetor® also reduced BALF levels of MIP-2 from 214
(54) to 127 (22) pg/ml (P <0.01). KC and IL-6 levels were not aff ected. Conclusion In a model of antibody-mediated TRALI, C1 inhibitor
attenuated pulmonary infl ammation. C1 inhibition may be a potential
benefi cial intervention in TRALI. 4.9 ± 0.5 (P = 0.09). Cetor® also reduced BALF levels of MIP-2 from 214
(54) to 127 (22) pg/ml (P <0.01). KC and IL-6 levels were not aff ected. Conclusion In a model of antibody-mediated TRALI, C1 inhibitor
attenuated pulmonary infl ammation. C1 inhibition may be a potential
benefi cial intervention in TRALI. (PRC) transfusions in the ICU was in accordance with the best clinical
evidence. A number of studies, most notably the TRICC study [2], have
shown that indications for ICU blood transfusions are a haemoglobin
(Hb) level of <7 g/dl or evidence of acute haemorrhage [3]. These
criteria were therefore employed. (PRC) transfusions in the ICU was in accordance with the best clinical
evidence. A number of studies, most notably the TRICC study [2], have
shown that indications for ICU blood transfusions are a haemoglobin
(Hb) level of <7 g/dl or evidence of acute haemorrhage [3]. These
criteria were therefore employed. y
Methods This study prospectively examined episodes of PRC unit
transfusions over a 2-month period in the ICU of a large level 1 trauma
centre and a tertiary cardiac unit. The number of PRC units transfused
in each episode was recorded by nurses, along with the proposed
indication and concurrent Hb level. The data were analysed to assess
the number of transfusions administered contrary to the guidelines,
along with the average Hb level at which a PRC unit was transfused and
the average number of units administered per episode. P372 P372
C1 inhibitor attenuates pulmonary infl ammation in an in vivo model
of transfusion-related acute lung injury
M Müller1, PR Tuinman1, G Jongsma1, AM Tuip-de Boer1, L Boon2,
SS Zeerleder1, NP Juff ermans1
1Academic Medical Center, Amsterdam, the Netherlands; 2Bioceros, Utrecht,
the Netherlands
Critical Care 2013, 17(Suppl 2):P372 (doi: 10.1186/cc12310) 3
Blood transfusion practices in the ICU of a level 1 trauma centre and
tertiary cardiac unit By calculating the Acute Physiology and
Chronic Health Evaluation (APACHE) II and IV scores the severity of
illness was determined on arrival in the ICU. Conclusion Our results indicate a liberal transfusion threshold
currently exists in the ICU. Patients are frequently receiving excessive
PRC transfusions for Hb levels above the recommended concentration. In the 2-month study period, these were associated with a cost of
approximately £12,220. We recommend increased staff awareness of
the guidelines to reduce the number of unnecessary transfusions. This
would decrease exposure of ICU patients to unnecessary risks of blood
transfusion, reduce cost of treatment and help to preserve a valuable
resource. f
1Academic Medical Center, Amsterdam, the Netherlands; 2Bioceros, Utrecht,
the Netherlands 1Academic Medical Center, Amsterdam, the Netherlands; 2Bioceros, Utrecht,
the Netherlands 1Academic Medical Center, Amsterdam, the Netherlands; 2Bioceros, Utrecht,
the Netherlands Critical Care 2013, 17(Suppl 2):P372 (doi: 10.1186/cc12310) Introduction Transfusion-related acute lung injury (TRALI) has a
high incidence in critically ill and surgical patients and contributes
to adverse outcome, while specifi c therapy is absent. Recently it was
demonstrated that complement activation plays a pivotal role in TRALI. We aimed to determine whether a C1 inhibitor is benefi cial in a two-hit
mouse model of antibody-mediated TRALI. 1. Vlaar APJ, et al.: Crit Care Med 2010, 38:771-778. References Results In total, 1,787 cardiac surgical patients were included. A total of
19 (1.1%) patients developed TRALI within 24 hours following surgery. Patients developing TRALI were older compared with patients not
developing TRALI, mean age respectively 71 and 65 years (P = 0.035). Furthermore, patients developing TRALI had higher APACHE II, APACHE
IV, EURO and ASA score (P = 0.000, P = 0.000, P = 0.000 and P = 0.37). Compared with patients not developing TRALI, patients developing
TRALI had a longer cardiopulmonary bypass time. Of the 19 patients
developing TRALI, four patients (21.1%) died during ICU stay, in total six
patients (31.6%) died during hospital stay. Patients developing TRALI
were ventilated longer and had a longer ICU and hospital stay. 1. Corwin HL, et al.: Crit Care Med 2004, 32:39-52. 1. Corwin HL, et al.: Crit Care Med 2004, 32:39-52. 1. Corwin HL, et al.: Crit Care Med 2004, 32:39-52. 2. Hebert PC: Br J Anaesth 1998, 81(Suppl 1):25-33. 3. Napolitano et al.: Crit Care Med 2009, 37:3124-3157. 2. Hebert PC: Br J Anaesth 1998, 81(Suppl 1):25-33. 3. Napolitano et al.: Crit Care Med 2009, 37:3124-3157. g
j
y
M Müller1, PR Tuinman1, G Jongsma1, AM Tuip-de Boer1, L Boon2,
SS Zeerleder1, NP Juff ermans1 g
g
p
y
Conclusion Older age, higher APACHE II, IV, EURO and ASA scores and
a longer time on cardiopulmonary bypass and ventilation time are risk
factors for the development of TRALI. The ICU and hospital stay are
longer, and the prognosis is bad for patients developing TRALI. Reference Albumin-induced coagulopathy is less severe and more
eff ectively reversed with fi brinogen concentrate than is synthetic
colloid-induced coagulopathy y
Methods BALB/c mice were primed with lipopolysaccharide (LPS, from
E. coli 0111:B4) that was administered intraperitoneally in a dose of
0.1 mg/kg, after which TRALI was induced by injecting MHC-I antibody
against H2Kd (IgG2a,k) at a dose of 2 mg/kg. Mice infused with PBS or
LPS served as controls. Concomitantly, mice infused with the MHC-I
antibody were treated with C1 inhibitor (Cetor®; Sanquin, Amsterdam,
the Netherlands) in a dose of 400 IU/kg intravenously. After infusion,
mice were mechanically ventilated with a lung-protective pressure-
controlled mode for 2 hours and then sacrifi ced, after which a
bronchoalveolar lavage (BAL) was done. Statistics were analyzed by
one-way ANOVA, values expressed as mean and standard deviation. Results Injection of LPS and MHC-I antibodies resulted in TRALI,
indicated by increased levels of protein in the BAL fl uid, wet/dry
ratios and levels of KC, MIP-2 and IL-6. C1 inhibitor Cetor® signifi cantly
reduced total protein in BAL fl uid from 792 (169) to 421 (230) μg/ml
(P <0.001) and tended to reduce the wet/dry ratio from 5.3 ± 0.3 to DW Winstedt, JH Hanna, US Schött
Lund University, Lund, Sweden
Critical Care 2013, 17(Suppl 2):P374 (doi: 10.1186/cc12312) Introduction Volume resuscitation is essential to restore normovolemia
during hemorrhagic shock, burns and sepsis. However, synthetic
colloids cause dilutional coagulopathy. The aims were to determine
whether the natural colloid albumin induces a lesser degree of
coagulopathy compared with synthetic colloids, and the comparative
eff ectiveness of fi brinogen concentrate to reverse coagulopathy
following dilution with these solutions. Methods Rotational thromboelastometry-based tests were used
to examine coagulation parameters in samples from 10 healthy
volunteers, in undiluted blood and samples diluted 1:1 with saline, S141 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Ringer’s acetate, hydroxyethyl starch (HES) 130/0.4, buff ered HES
130/0.4, 3% dextran 60, 6% dextran 70 or 5% albumin. Samples were
analyzed before and after addition of 2 mg/ml fi brinogen concentrate. Results EXTEM maximum clot fi rmness (MCF), clot formation time
(CFT) and α angle (AA) decreased signifi cantly upon dilution with
all colloid solutions (P <0.001), although a signifi cantly greater
coagulopathic eff ect was seen for samples diluted with synthetic
colloid solutions versus albumin (P ≤0.001). Albumin-induced coagulopathy is less severe and more
eff ectively reversed with fi brinogen concentrate than is synthetic
colloid-induced coagulopathy A signifi cant reduction in
the platelet component of clot strength (EXTEM MCF – FIBTEM MCF)
was seen for samples diluted with synthetic colloids (P <0.001) but
not albumin (P = 0.10). Following addition of fi brinogen, FIBTEM MCF,
EXTEM MCF and EXTEM AA were signifi cantly higher, and EXTEM CFT
was signifi cantly shorter in samples diluted with albumin versus those
treated with HES or dextran (P ≤0.001). P375 P375
Network meta-analysis of clinical trials of fl uid treatments in sepsis
demonstrates improved survival with albumin compared with
crystalloid and hydroxyethyl starch
M Bansal1, A Farrugia2, G Martin3
1Plasma Protein Therapeutics Association, Annapolis, MD, USA; 2University of
Western Australia, Crawley, Australia; 3Emory University School of Medicine,
Atlanta, GA, USA
Critical Care 2013, 17(Suppl 2):P375 (doi: 10.1186/cc12313) Critical Care 2013, 17(Suppl 2):P375 (doi: 10.1186/cc12313) Critical Care 2013, 17(Suppl 2):P375 (doi: 10.1186/cc12313) Introduction Fluid resuscitation is widely practiced in treating sepsis. Comparative assessment of the diff erent fl uid modalities is hampered
by a paucity of direct trials. We present a network meta-analysis for
assessing the relative eff ectiveness of two fl uid treatments in sepsis
when they have not been compared directly in a randomized trial but
have each been compared with a common treatment. p
g
Results The economic model shows the potential cost saving by
switching from open to closed albumin administration systems. It is
measured in occurrence of BSI in the ICU, estimated occurrence of BSI
in the albumin population and incidence of overall mortality in the ICU
and cost of BSI per albumin patient. Finally, the model describes the
potential cost savings by switching to a closed system. Using closed
albumin delivery systems may overall provide a cost saving for the
hospital and healthcare sector. Assuming 800 patients treated in an ICU
per year with all in all 4,800 ml albumin, the total annual cost saving
would be £24,000 switching from open to closed albumin infusions. Methods A systematic review of trials sepsis yielded 13 trials for
assessment in network meta-analysis. The indirect comparison between
albumin, HES and crystalloid was conducted using Bayesian methods
for binomial likelihood, fi xed-eff ects network meta-analysis with a
Monte Carlo Gibbs sampling method. Studies in septic patients with
crystalloid as a reference treatment compared with any formulation of
the colloid treatments albumin or HES were included, as were direct
head-to-head trials between the two colloids.f g
Conclusion Switching from an open to a closed albumin infusion
system may result in a reduction in the risk of external contamination,
resulting in less infection and may provide a cost saving for the
healthcare system. Conclusion Switching from an open to a closed albumin infusion
system may result in a reduction in the risk of external contamination, resulting in less infection and may provide a cost saving for the
healthcare system. P376 Assessing the economic impact of catheter-related bloodstream
infections when switching from open to closed system delivery of
human albumin solutions in ICUs
F Axelsen1, P Riss2
1Baxter Healthcare, Zurich, Switzerland; 2Medical University of Vienna, Austria
Critical Care 2013, 17(Suppl 2):P376 (doi: 10.1186/cc12314) y
References Results Odds ratios between the diff erent treatments were obtained
(Figure 1). Ranking the interventions [1] demonstrated that albumin
ranked highest in lowering mortality at a 96.4% probability compared
with 3.6% and 0.01% for crystalloid and HES, respectively.l 1. Maki et al.: Infect Control Hosp Epidemiol 2011, 32:50-58. 2. Tacconelli et al.: J Hosp Infection 2009, 72:97-103. 3. Franzetti et al.: Epidemiol Infect 2009, 137:1041-1048. 1. Maki et al.: Infect Control Hosp Epidemiol 2011, 32:50-58. 1. Maki et al.: Infect Control Hosp Epidemiol 2011, 32:50-58. 2
Tacconelli et al : J Hosp Infection 2009 72:97-103 1. Maki et al.: Infect Control Hosp Epidemiol 2011, 32:50-58. 2. Tacconelli et al.: J Hosp Infection 2009, 72:97-103. 3. Franzetti et al.: Epidemiol Infect 2009, 137:1041-1048. 2. Tacconelli et al.: J Hosp Infection 2009, 72:97-103. Conclusion Albumin as a fl uid therapy in sepsis is associated with the
lowest mortality of the three modalities studied. Reference . Salanti G, Ades AE, Ioannidis JP: Graphical methods and numerical
summaries for presenting results from multiple-treatment meta-analysis:
an overview and tutorial. J Clin Epidemiol 2011, 64:163-171. Assessing the economic impact of catheter-related bloodstream
infections when switching from open to closed system delivery of
human albumin solutions in ICUs ,
1Baxter Healthcare, Zurich, Switzerland; 2Medical University of Vienna, Austria
Critical Care 2013, 17(Suppl 2):P376 (doi: 10.1186/cc12314) Introduction Hospital-acquired infections create a signifi cant burden
to the healthcare system. To understand the economic impact, a model
was developed to assess the cost of diff erences in catheter-related
bloodstream infections (BSI) in ICUs based on either closed or open
albumin systems. Conclusion Hemodilution using albumin induced a lesser degree of
coagulopathy compared with the synthetic colloids HES and dextran. In addition, albumin-induced coagulopathy was more eff ectively
reversed following addition of fi brinogen concentrate compared with
coagulopathy induced by synthetic colloids. y
Methods A model was developed with the aim of showing the
economic consequences of diff erences between intravenous (i.v.)
infusions of albumin via open or closed infusion containers. The model
took a healthcare perspective in the UK. The impact of central venous
catheter-related BSI was evaluated. The model took diff erences in
BSI rates and the associated cost consequences for the hospital into
account. The model bases on the evidence that closed system delivery
helps to reduce the risk of external contamination, which decreases the
BSI rate and reduces overall mortality [1-3]. We assessed the total costs
using public list prices and literature. The model assumes that infection
rates for using open and closed albumin delivery systems are similar to
any other closed or open nonalbumin i.v. fl uid delivery system used in
the ICU. The model looks at reduction in central-line-associated BSI and
the potential cost saving of this reduction. P375 Procedural sedation in the emergency department Methods An electronic-dedicated database was created to retro-
spectively collect volume, type of fl uids infused and plasmatic acid–
base balance variations in postoperative ICU patients from admission
to 9:00 am of the day after. SIDin was calculated as the average SID of all
fl uids infused during the whole study period (crystalloids, colloids and
blood products). Arterial base excess variation (ΔBEa) was computed
as the diff erence between values at 9:00 am on the day after and those
at entry. We report data from all patients admitted in 2006 and 2007
(650 patients). MS Shah1, FS Shah2, KP Pope3, AS Abbas4
1Luton and Dunstable Hospital, London, UK; 2Birmingham Hospitals NHS
Trust, Birmingham, UK; 3Chesterfi eld NHS Trust, Chesterfi eld, UK; 4Colchester
Hospital, Colchester, UK
Critical Care 2013, 17(Suppl 2):P380 (doi: 10.1186/cc12318) MS Shah , FS Shah , KP Pope , AS Abbas
1Luton and Dunstable Hospital, London, UK; 2Birmingham Hospitals NHS
Trust, Birmingham, UK; 3Chesterfi eld NHS Trust, Chesterfi eld, UK; 4Colchester
Hospital, Colchester, UK
Critical Care 2013 17(Suppl 2):P380 (doi: 10 1186/cc12318) p
,
,
Critical Care 2013, 17(Suppl 2):P380 (doi: 10.1186/cc12318) Introduction Are safety guidelines being followed when administering
procedural sedation in the emergency department? Between
November 2004 and November 2008, the NPSA received 498 alerts
of patients being given the wrong dose of midazolam for procedural
sedation [1]. In the fi rst 5 years of midazolam use there were 86 deaths,
most related to procedural sedation [2]. Introduction Are safety guidelines being followed when administering
procedural sedation in the emergency department? Between
November 2004 and November 2008, the NPSA received 498 alerts
of patients being given the wrong dose of midazolam for procedural
sedation [1]. In the fi rst 5 years of midazolam use there were 86 deaths,
most related to procedural sedation [2]. p
Results Nine patients not receiving intravenous infusions were
excluded. The remaining population was divided into three groups
according to SIDin distribution (Group 1, 18 ± 12; Group 2, 47 ± 6;
Group 3, 55 ± 0 mEq/l). We observed a progressive increment in
ΔBEa between the groups (1.1 ± 2.0 vs. 2.8 ± 2.9 vs. 3.0 ± 2.8 mmol/l,
P <0.001). We further subdivided each group by the median value of
baseline HCO3-a (24.3 (22.3 to 26.1) mmol/l) and we analyzed the ΔBEa:
we observed a greater increase in patients with lower baseline HCO3-a
(Group 1, 1.8 ± 2.9 vs. P379 (SAP), SV and CO were recorded directly before the administration of
any colloid (T0) and every 5 minutes for the next 1 hour (T1 to T12). Kolmogorov–Smirnov was used to test normal distribution of data and
ANOVA was used for the statistical analysis. P <0.05 was considered
statistically signifi cant. P379
Microcirculatory response to fl uid challenge: should we prefer
balanced colloids to rebalance tissue perfusion? A Donati, E Damiani, R Domizi, C Scorcella, A Carsetti, S Tondi,
R Castagnani, N Mininno, V Monaldi, P Pelaia
Università Politecnica delle Marche, Ancona, Italy
Critical Care 2013, 17(Suppl 2):P379 (doi: 10.1186/cc12317) y
gi
Results Demographic data and ASA classifi cation did not diff er
statistically signifi cant among the six groups of the study. CO, SV, HR
and SAP did not show any statistically signifi cant evolution compared
with their baseline value during the study period. Moreover, there were
no statistically signifi cant diff erences among the six study groups with
regard to any of the recorded parameters. Introduction Fluid resuscitation should improve tissue oxygenation in
hypovolemia, besides restoring macrohemodynamic stability [1]. We
evaluated the microvascular response to fl uid challenge with diff erent
colloid solutions and its relation to macrohemodynamics.l y
Conclusion According to our results, volume replacement with the
six colloids tested in our study did not result in any hemodynamic
response. Within comparison of these six colloids did not reveal any
statistically signifi cant diff erence in any of the parameters recorded
according to our protocol. Methods An observational study of patients receiving a fl uid challenge
(500 ml colloids in 30 minutes) according to the attending physician’s
decision. Before and after the infusion, sublingual microcirculation was
evaluated with sidestream dark-fi eld imaging (Microscan; Microvision
Medical, Amsterdam, the Netherlands). Microvascular fl ow and density
were assessed for small vessels [2]. The cardiac index (CI), intrathoracic
blood volume index (ITBVI) and extravascular lung water index (ELWI)
were measured in seven patients with PiCCO2 (Pulsion Medical System,
Munich, Germany). Results Ten patients (two sepsis, four trauma, three intracranial
bleeding, one post surgery) received either saline-based hydroxyethyl
starch (HES) 130/0.4 (Amidolite®; B.BraunSpA; n = 5) or balanced HES
130/0.42 (Tetraspan®; B.BraunSpA; n = 5). The CI (P = 0.02) and ITBVI
(P = 0.07) tended to increase, the EVLWI did not change. Microvascular
fl ow and density improved in the whole sample. No correlation was
found between macro-circulatory and micro-circulatory parameters. P378 Strong ion diff erence and arterial bicarbonate concentration as
cornerstones of the impact of fl uid therapy on acid–base balance
D Ottolina, M Ferrari, L Zazzeron, E Scotti, M Stanziano, C Rovati,
C Marenghi, L Gattinoni, P Caironi
Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico Università
degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P378 (doi: 10.1186/cc12316) g
Conclusion Balanced HES may be more effi cacious than saline-based
HES in recruiting the microcirculation, thereby improving tissue O2
delivery. Introduction The biochemical characteristics of infused fl uids may be
important in regulating acid–base balance, by modifying plasmatic
volume and strong ion diff erence. In vitro and animal studies [1,2] have
shown that volume and strong ion diff erence of infused fl uids (SIDin)
as well as the arterial baseline bicarbonate concentration (HCO3-a)
infl uence acid–base variations. Our aim was to verify these changes in
critically ill patients after surgery. Procedural sedation in the emergency department 0.2 ± 2.6, mmol/l, P <0.001; Group 2, 4.0 ± 2.7
vs. 1.5 ± 2.6, mmol/l, P <0.001; Group 3, 4.4 ± 2.8 vs. 1.7 ± 2.0 mmol/l,
P <0.001), as compared with those with higher baseline levels. When
the study population was divided into quartiles of the diff erence
between SIDin and HCO3-a, ΔBEa appeared to increase with the rise of
such diff erence (P <0.001).f Methods We searched through the controlled drugs book in
resuscitation over a 2-month period and found a list of patients who
had received midazolam or fentanyl. From this, we could make a search
for the relevant A and E notes for these patients. From these notes, we
looked for (see shorthand in Table 1): verbal consent documentation
(consent), past medical history recorded (pmhx), safe initial dose of
midazolam (midaz), pre-procedure monitoring (pre), post-procedure
monitoring (post), and monitoring for 1 hour before discharge (1hr). Following introduction of a reminder in the controlled drugs book/
sedation room and staff education, the case notes were analysed over
another 2-month period (24 sets of notes) to assess practise against
safety guidelines. f
Conclusion SIDin aff ects the acid–base status per se and in relation-
ship with HCO3-a. We verifi ed this hypothesis in critically ill patients,
highlighting the importance of the diff erence between SIDin
and HCO3-a, which better describes and predicts the acid–base
modifi cations to fl uid therapy. y g
Results See Table 1 (key for shorthand in Methods). Conclusion The re-audit notices within the procedural sedation room
and controlled drug book front cover served as a reminder of good
practise. The visibility of this reminder (within the CD book) helped
ensure better adherence to the audit standard. This reminder will now
be kept within the CD book. i
References il
References 1. Carlesso E, et al.: The rule regulating pH changes during crystalloid
infusion. Intensive Care Med 2011, 37:461-468. Carlesso E, et al.: The rule regulating pH changes during crystalloid infusion. Intensive Care Med 2011, 37:461-468. P377 Figure 1 (abstract P375). Forest plot of results of Bayesian fi xed-eff ect
network meta-analysis of mortality. Comparing the hemodynamic eff ects of six diff erent colloids
N Kteniadakis, V Grosomanidis, E Oloktsidou, B Fyntanidou, C Nouris,
K Kotzampasi, D Vasilakos
Aristotle Medical School, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P377 (doi: 10.1186/cc12315) Comparing the hemodynamic eff ects of six diff erent colloids
N Kteniadakis, V Grosomanidis, E Oloktsidou, B Fyntanidou, C Nouris,
K Kotzampasi, D Vasilakos
Aristotle Medical School, Thessaloniki, Greece
Critical Care 2013, 17(Suppl 2):P377 (doi: 10.1186/cc12315) Introduction Despite the fact that colloids are widely used, doubt still
remains as to which colloid is the best and what is the ideal volume of
administration. The aim of our study was to compare the hemodynamic
eff ects of six diff erent colloids. Introduction Despite the fact that colloids are widely used, doubt still
remains as to which colloid is the best and what is the ideal volume of
administration. The aim of our study was to compare the hemodynamic
eff ects of six diff erent colloids. ff
Methods A total of 180 patients were enrolled in our study undergoing
general surgery procedures, who were assigned to six groups (A, B, C, D,
E, F). After fasting fl uid replacement and induction of anesthesia, 500 ml
Gelofusine, Haemaccel, Voluven, HES 6%, HES 10% and Hemohes were
administered to each group respectively. An oesophageal Doppler
monitor probe was inserted into the patients for measuring stroke
volume (SV) and cardiac output (CO). Standard monitoring included
ECG, IBP, ETCO2 and SpO2, and heart rate (HR). Systemic arterial pressure Figure 1 (abstract P375). Forest plot of results of Bayesian fi xed-eff ect
network meta-analysis of mortality. S142 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 References 1. Futier et al.: Crit Care 2011, 15:R214. 2. De Backer et al.: Crit Care 2007, 11:R101. P379 Balanced HES led to a greater increase in capillary density than NaCl
HES (Figure 1).fi p
References 2. Langer T, et al.: In vivo conditioning of acid–base equilibrium by crystalloid
solutions: an experimental study on pigs. Intensive Care Med 2012,
38:686-693. [http://www.npsa.nhs.uk/corporate/news/
reducing-risk-of-midazolam-overdose-in-adults] [http://www.npsa.nhs.uk/corporate/news/
reducing-risk-of-midazolam-overdose-in-adults] S143 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http //ccfor m com/s pplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P379). Microvascular response to fl uid challenge: eff ects of diff erent colloids. Figure 1 (abstract P379). Microvascular response to fl uid challenge: eff ects of diff erent colloids. Figure 1 (abstract P379). Microvascular response to fl uid challenge: eff ects of diff erent colloids. 2. Epstein B: Data Retrieval Unit HFD-737. Department of Health and Human
Services, Offi ce of Epidemiology and Biostatistics, Center for Drug Evaluation
and Research; 27 June 1989. P381
Impact of an age, kidney and liver function adjusted sedation
protocol in critically ill patients
S Haddad, H Tamim, C Gonzales, A Rishu, A Deeb, Y Arabi
King Abdulaziz Medical City, Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P381 (doi: 10.1186/cc12319)
Introduction Daily sedation interruption and protocol implementation
have been recommended to reduce excessive sedation; however,
their use has been inconsistent. We hypothesized that the use of an
age, kidney and liver function adjusted sedation protocol would be
Table 1 (abstract P380). Before and after education programme and
reminder in the controlled drugs book
Before (%)
After (%)
Yes
No
Yes
No
Consent
35
65
50
50
Pmhx
55
45
67
33
Midaz
58
42
88
12
Pre
25
75
54
46
Post
10
90
71
29
1hr
55
45
54
46 Table 1 (abstract P380). Before and after education programme and
reminder in the controlled drugs book associated with reduced doses and improved outcomes compared
with a standard protocol. Table 1 (abstract P380). Before and after education programme and
reminder in the controlled drugs book
Before (%)
After (%)
Yes
No
Yes
No
Consent
35
65
50
50
Pmhx
55
45
67
33
Midaz
58
42
88
12
Pre
25
75
54
46
Post
10
90
71
29
1hr
55
45
54
46 Methods This was a prospective cohort study comparing 3 months
of a standard protocol (control group) with 3 months of an adjusted
protocol (intervention group). Benzodiazepines induce hyperglycemia in rats by aff ecting
peripheral disposal of glucose VS Salice, FV Valenza, MP Pizzocri, LV Valenti, GC Chevallard, MU Umbrello,
SG Gatti, SF Fargion, GI Iapichno, LG Gattinoni
Università degli Studi Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P385 (doi: 10.1186/cc12323) Results We retrieved 902 citations. Among them, nine RCTs fulfi lled
inclusion criteria. PS was compared with the standard practice in one
RCT, DIS with the standard practice in four, PS with DIS in two, and PS
with PS plus DIS (PS/DIS) in two. PS or DIS reduced the duration of
mechanical ventilation by 28.9% and ICU stay by 27.8% compared with
the standard practice. Neither the duration of mechanical ventilation
nor ICU stay diff ered among DIS, PS and PS/DIS. Mortality did not diff er
among the three strategies. DIS increased the daily dose of sedatives
and the workload of nurses in one recent RCT. Introduction In light of the interest in the relationship between
glycemia control in critically ill subjects and outcome, we set up a study
to investigate whether benzodiazepine, commonly used in anesthesia
and ICUs, interferes with glucose metabolism and to explore the
mechanism. Introduction In light of the interest in the relationship between
glycemia control in critically ill subjects and outcome, we set up a study
to investigate whether benzodiazepine, commonly used in anesthesia
and ICUs, interferes with glucose metabolism and to explore the
mechanism. Methods A total of 40 sedated and paralyzed Sprague–Dawley rats
(301 ± 55 g) were investigated in four consecutive studies. (1) To
investigate the eff ects of diazepam on blood glucose, 15 rats were
randomly assigned to intraperitoneal anesthesia with tiopenthal
80 mg/kg (DZP0), tiopenthal 40 mg/kg + diazepam 5 mg/kg (DPZ5)
or tiopenthal 40 mg/kg + diazepam 15 mg/kg (DZP15). Blood levels of
glucose (GEM premier 3000; IL) were measured at time intervals over
2 hours. (2) Ten animals randomized to DZP0 or DZP5 underwent an
intravenous glucose tolerance test with glucose bolus (0.5 g/kg). Acute
insulin response, the mean value of blood insulin (Insulin ELISA kit;
Millipore) from 2 to 10 minutes after glucose bolus, was measured as
index of insulin secretion. (3) A hyperinsulinemic euglycemic clamp
obtained by a continuous intravenous infusion of insulin (130 mUI/
kg/minute) was run in 10 animals randomized to DZP0 or DZP5 and
the glucose infusion rate (GIR, mg/kg/minute) was assessed [1]. Benzodiazepines induce hyperglycemia in rats by aff ecting
peripheral disposal of glucose (4) The
eff ect of midazolam on blood glucose was tested in fi ve additional
animals (M5: tiophental 40 mg/kg + midazolam 5 mg/kg). Data are
presented as mean ± SEM. Statistical analysis (ANOVA, t test) was
conducted with Sigma Stat 3.1 (Systat Software). Conclusion PS or DIS decreased the duration of mechanical ventilation
and ICU stay. PS seems to be superior to DIS based on the doses of
sedatives and the workload of nurses. P383 Chemical and physical compatibility of continuous intravenous
drug infusion combinations used in paediatric intensive care
C Cole1, A Fox1, M Van Der Merwe2, L Dickson2, K Ball2, A Bevan1,
A Hocking1, J Pappachan1
1University Hospitals Southampton, UK; 2University of Portsmouth, UK
Critical Care 2013, 17(Suppl 2):P383 (doi: 10.1186/cc12321) Introduction The aim of this research was to provide clinically relevant
evidence for Y-site compatibility of drug infusion combinations used in
the PICU. Pharmacists and clinicians regularly have to interpret limited
published data, particularly when more than two drugs are Y-sited. The risk of potential incompatibility must be balanced against that of
additional line insertion. Results (1) Diazepam was associated with higher levels of blood
glucose in a dose–response fashion: DZP0 128 ± 7 mg/dl, DZP5
166 ± 7.3, DZP15 197 ± 7 (P <0.05). (2) The acute insulin response
to intravenous glucose tolerance test was similar in DZP0 and DZP5
(DZP0 3.97 ± 0.42 ng/ml, DZP5 3.68 ± 0.44, P = 0.68), while blood
glucose levels were diff erent (DZP0 192 ± 5 mg/dl, DZP5 217 ± 5,
P <0.05). (3) During hyperinsulinemic euglycemic clamp, blood
glucose levels were similar (109 ± 3 mg/dl, P = 0.2), but the DZP5
group showed a trend through lower values of GIR (DZP0 30.8 ± 2 mg/
kg/minute, DZP5 24.7 ± 2, P = 0.08). (4) Infusion of midazolam was
associated with higher blood glucose levels (DZP0 128 ± 5 mg/dl, M5
151 ± 6, P <0.05).i Methods A full 28-factorial design (total 256 combinations) was
used to investigate chemical and physical compatibility of fi ve drugs
(clonidine, morphine, ketamine, midazolam and furosemide). The
drugs were studied at their highest commonly infused concentrations
and exposed to three variations in environmental conditions (diluent:
sodium chloride 0.9% or glucose 10%; temperature 25 or 37°C;
and normal room lighting or blue light phototherapy). Chemical
stability was assessed using HPLC; >10% reduction in concentration
indicated incompatibility. Benzodiazepines induce hyperglycemia in rats by aff ecting
peripheral disposal of glucose Physical incompatibility was confi rmed by
precipitation, pH or colour change.f Conclusion Both diazepam and midazolam signifi cantly alter plasma
glucose levels in rats. Peripheral disposal of glucose rather than altered
pancreas secretion of insulin explains the benzodiazepine-associated
hyperglycemia. Results Environmental conditions had no eff ect on the drug mixtures. The precipitation observed in incompatible combinations was due to
either a change in pH, or with ketamine the presence of benzethonium
chloride. Of 31 possible drug combinations, 12 were incompatible. A
further three combinations were incompatible at extreme pH, or were
of concern and so should be avoided. The incompatible formulations p
References We hypothesized that the use of an
age, kidney and liver function adjusted sedation protocol would be S144 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 adjusted protocol group (18% vs. 36%, P = 0.004) with no signifi cant
diff erences in MVD, ICU and hospital LOS, and hospital mortality. Conclusion The use of an age, kidney and liver function adjusted
sedation protocol is associated with lower doses of analgesics and
sedatives, lower risk of agitation and lower ICU mortality. all contained furosemide. All combinations of the sedative agents
studied were chemically and physically compatible. adjusted protocol group (18% vs. 36%, P = 0.004) with no signifi cant
diff erences in MVD, ICU and hospital LOS, and hospital mortality. y
y
y
Conclusion This work provides evidence for Y-site compatibility of
morphine, midazolam, clonidine and ketamine in any combination,
which will potentially reduce the need for extra intravenous lines. Furosemide is incompatible with any of these sedative drugs and must
be infused via a separate line. These results will aid clinical decision-
making and help satisfy the requirements of recent UK Department of
Health legislation relating to the mixing of medicines. Reference Conclusion The use of an age, kidney and liver function adjusted
sedation protocol is associated with lower doses of analgesics and
sedatives, lower risk of agitation and lower ICU mortality. P382 Risk and benefi t of protocol sedation and daily interruption of
sedation for mechanically ventilated patients: a systematic review
K Hosokawa1, M Egi2, M Nishimura3
1Kyoto Prefectural Yosanoumi Hospital, Kyoto, Japan; 2Okayama University
Hospital, Okayama, Japan; 3Tokushima University Hospital, Tokushima, Japan
Critical Care 2013, 17(Suppl 2):P382 (doi: 10.1186/cc12320) DeFronzo: Am J Physiol 1979, 273:E214-E223. p
References In the adjusted protocol, patients
were divided into three categories: category 1 (age <60 years, and
normal kidney and liver function), category 2 (age = 60 to 70 years,
or moderate kidney or liver function impairment), and category 3
(age >70 years, or severe kidney or liver function impairment). The
upper limits of analgesics and sedatives doses were determined
by age, and kidney and liver function, being lowest in category
3, and lower in category 2 than category 1. All consecutive adults
mechanically ventilated patients who required infusion of analgesics
and/or sedatives for >24 hours were included in the study. We
compared the main outcomes of both groups including average daily
doses of analgesics and sedatives; average Sedation–Agitation Scale
(SAS), pain and GCS scores; mechanical ventilation duration (MVD);
sedation-related complications during ICU stay; ICU and hospital
length of stay (LOS), and ICU and hospital mortality. y
y
Results Two hundred and four patients were included in the study
(control group = 105; adjusted protocol group = 99). There was no
diff erence in baseline characteristics between the two groups. The
adjusted protocol group, compared with the control group, received
signifi cantly lower average daily doses of fentanyl (2,162 ± 2,110 μg
vs. 3,650 ± 3,253 μg, P = 0.0001), nonsignifi cant lower average daily
doses of midazolam and dexmedetomidine, and a trend toward
higher average daily doses of propofol. Pain score was higher in the
adjusted protocol group (0.98 ± 0.72 vs. 0.16 ± 0.35, P <0.0001) with
no diff erence in SAS or GCS scores. Sedation-related complications
during ICU stay were not diff erent between the two groups; however,
agitation (SAS = 5) was less frequent in the adjusted protocol group
(3% vs. 30%, P <0.0001). ICU mortality was signifi cantly lower in the P381
Impact of an age, kidney and liver function adjusted sedation
protocol in critically ill patients
S Haddad, H Tamim, C Gonzales, A Rishu, A Deeb, Y Arabi
King Abdulaziz Medical City, Riyadh, Saudi Arabia
Critical Care 2013, 17(Suppl 2):P381 (doi: 10.1186/cc12319) Introduction Daily sedation interruption and protocol implementation
have been recommended to reduce excessive sedation; however,
their use has been inconsistent. Reference 1. Mixing of Medicines Prior to Administration in Clinical Practice: Medical
and Non-medical Prescribing [http://www.dh.gov.uk/en/
Publicationsandstatistics/Publications/PublicationsPolicyAndGuidance/
DH_116361] 1. Mixing of Medicines Prior to Administration in Clinical Practice: Medical
and Non-medical Prescribing [http://www.dh.gov.uk/en/
Publicationsandstatistics/Publications/PublicationsPolicyAndGuidance/
DH_116361] 1. Mixing of Medicines Prior to Administration in Clinical Practice: Medical
and Non-medical Prescribing [http://www.dh.gov.uk/en/
Publicationsandstatistics/Publications/PublicationsPolicyAndGuidance/
DH_116361] g
1Kyoto Prefectural Yosanoumi Hospital, Kyoto, Japan; 2Okayama University
Hospital, Okayama, Japan; 3Tokushima University Hospital, Tokushima, Japan
Critical Care 2013, 17(Suppl 2):P382 (doi: 10.1186/cc12320) Introduction Protocol-directed sedation (PS) and daily interruption of
sedation (DIS) are two major strategies to optimize sedation depth for
mechanically ventilated patients. To clarify the clinical risk and benefi ts
of PS and DIS, we performed a systematic review of randomized
controlled trials (RCTs) in previous literature. Introduction Protocol-directed sedation (PS) and daily interruption of
sedation (DIS) are two major strategies to optimize sedation depth for
mechanically ventilated patients. To clarify the clinical risk and benefi ts
of PS and DIS, we performed a systematic review of randomized
controlled trials (RCTs) in previous literature. P384
Abstract withdrawn P384
Abstract withdrawn P384 P384
Abstract withdrawn P384
Abstract withdrawn Methods We searched the MEDLINE database from January 1990
to October 2012 for studies on PS and DIS in critically ill patients. English-language manuscripts were included when they assessed
the impact of PS or DIS on outcomes in critically ill patients requiring
mechanical ventilation for >48 hours. PS is defi ned as sedation
practices implemented by ICU staff using a sedation scale and a
written protocol. DIS is defi ned as strategies with cessation of the
continuous sedatives. P387 Involvement of mitochondrial ATP-sensitive K+ channels in fentanyl-
induced mitochondrial dysfunction of cultured human hepatocytes
S Djafarzadeh, M Vuda, J Takala, SM Jakob
Bern University Hospital (Inselspital) and University of Bern, Switzerland
Critical Care 2013, 17(Suppl 2):P387 (doi: 10.1186/cc12325) Introduction Pharmacological agents used to treat critically ill patients
may alter mitochondrial function. The aim of the present study was
to investigate whether fentanyl, a commonly used analgesic drug,
interacts with hepatic mitochondrial function. Introduction We report our experience in the use of isofl urane for
prolonged sedation in severe ARDS patients. Prolonged sedation in
the ICU may be diffi cult because of tolerance, drug dependence and
withdrawal, drug interactions and side eff ects. Inhaled anesthetics
have been proposed for sedation in ventilator-dependent ICU patients. AnaConDa is a device that allows a safety and easy administration of
inhaled anesthetics in the ICU. Methods The human hepatoma cell line HepG2 was exposed to
fentanyl at 0.5, 2 or 10 ng/ml for 1 hour, or pretreated with naloxone
(an opioid receptor antagonist) at 200 ng/ml or 5-hydroxydecanoate
(5-HD; a specifi c inhibitor of mitochondrial ATP-sensitive K+ (KATP)
channels) at 50 μM for 30 minutes, followed by incubation with
fentanyl at 2 ng/ml for an additional hour. The mitochondrial complex
I-dependent, II-dependent and IV-dependent oxygen consumption
rates of the permeabilized cells were measured using a high-resolution
oxygraph (Oxygraph-2k; Oroboros Instruments, Innsbruck, Austria). The respiratory electron transfer capacity of intact cells was evaluated
using FCCP (carbonyl cyanide p-trifl uoromethoxyphenylhydrazone) to
obtain the maximum fl ux. Methods From January 2009 to June 2012, 15 patients were sedated
with isofl urane by means of the AnaConDa device. We consider
administration of isofl urane as a washout period from common
sedative drugs in patients with (at least one of): high sedative drug
dosage (propofol ≥300 mg/hour or midazolam ≥8 mg/hour) to reach
the target Richmond Agitation Sedation Score (RASS) or inadequate
paralysis; two or more hypnotic drugs to reach the target RASS
(propofol, midazolam, hydroxyzine, haloperidol, diazepam, quetiapine);
and hypertriglyceridemia. During isofl urane administration previous
hypnotic drugs were interrupted. We retrospectively collected data
before, during and after administration of isofl urane: hemodynamic
parameters, renal and hepatic function, level of sedation (RASS)
and sedative drug dosage. All data are reported as mean ± standard
deviation, otherwise as median (minimum to maximum). P387 l
Results Incubation of HepG2 cells with fentanyl (1 hour, 2 ng/ml)
induced a reduction in complex II-dependent and IV-dependent
respiration (Figure 1). Cells pretreated with 5-HD before the addition
of fentanyl exhibited no signifi cant changes in complex activities in
comparison with controls. Pretreatment with naloxone tended to
abolish the fentanyl-induced mitochondrial dysfunction. Treatment
with fentanyl led to a reduction in cellular ATP content (0.24 ± 0.06 in
controls vs. 0.17 ± 0.14 μmol/mg cellular protein in stimulated cells;
P = 0.02). We did not observe any diff erence in basal or FCCP-uncoupled
respiration rates of cells treated with fentanyl at 2 ng/ml compared
with controls (data not shown). Results Mean age was 43 ± 12 years and SAPS II was 35.7 ± 11; 13
patients were treated with ECMO for severe ARDS and four had a
history of drug abuse; median ICU length of stay was 41 (15 to 127)
days and they were ventilated for 41 (12 to 114) days. Due to severe
critical illness, target RASS was –4 for all patients, most of which were
also paralysed. Isofl urane was administered in nine patients because
of a high level of common sedative drugs, in fi ve patients due to the
use of two or more hypnotic drugs and in one patient because of
hypertrigliceridemia. Isofl urane administration lasted 5.6 ± 1.8 days. During isofl urane administration no alteration in renal function or
hemodynamic instability was recorded. After the isofl urane washout
period we observed a reduction in sedative drug dosage in 10 patients
while two patients were quickly weaned from mechanical ventilation
and the target RASS raised to 0. In two patients isofl urane was
precautionarily interrupted because of concomitant alteration of liver
function and suspected seizures respectively. Conclusion Fentanyl reduces cultured human hepatocyte mito-
chondrial respiration by a mechanism that is blocked by a KATP channel
antagonist. In contrast, antagonism with naloxone does not seem to
completely abolish the eff ect of fentanyl. Acknowledgement This study was supported by the Swiss National
Science Foundation (grant number 32003B_127619). P388f Reference S145 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P386 P386
Prolonged sedation in ARDS patients with inhaled anesthetics:
our experience
S Redaelli, P Mangili, V Ormas, S Sosio, L Peluso, F Ponzoni, N Patroniti,
A Pesenti
University of Milan-Bicocca, Monza, Italy
Critical Care 2013, 17(Suppl 2):P386 (doi: 10.1186/cc12324) P389 Conjunctival administration of S+ketamine in rabbits
L Hess1, J Malek2, A Kurzova2, Z Simunkova1, M Votava2
1Institute for Clinical and Experimental Medicine, Prague, Czech Republic;
23rd Medical Faculty of Charles University, Prague, Czech Republic
Critical Care 2013, 17(Suppl 2):P389 (doi: 10.1186/cc12327) Results During the study period 75 patients underwent procedural
sedation and were discharged from the ED. The median age was 40
years and 57% were male. The commonest procedure performed was
shoulder reduction (52%). In the propofol group (n = 20) the mean
LOS was 100 minutes compared with 165 minutes in those receiving
midazolam (n = 40) and 141 minutes in those receiving a combination
(n = 15), P = 0.004. There was no diff erence in adverse events between
groups. See Figure 1. Introduction Delivering analgesics via conjunctival application could
provide rapid and convenient pain relief in disaster medicine. There are
sporadic reports from the USA concerning inhalation administration
of aerosol with various drugs producing a wide variety of eff ects from
anxiolysis, sedation, and loss of aggressiveness to immobilisation. We
attempted to determine in an animal experiment whether conjunctival
administration of S+ketamine could produce signifi cant eff ect without
side eff ects. g
p
g
Conclusion Propofol is increasingly used in EDs for procedural sedation
due to its short duration of action. This study suggests that a shorter
duration of action and faster recovery may result in a reduced LOS in
the ED. f
Methods After ethic committee approval, 10 rabbits were administered
conjunctival S+ketamine 2.5 mg/kg. Measured parameters were SpO2,
blood pressure (BP) and heart rate (HR) before administration and in
1-minute intervals and immobilisation time (loss of righting refl ex). The
measurements were performed for 20 minutes. Conjunctival irritation
was measured 1, 10 and 20 minutes after administration according to
modifi ed technical standard EN IS O 10993-10. ANOVA test was used for
statistical analysis of hemodynamic parameters. Figure 1 (abstract P390). Length of stay (minutes) in the ED. Figure 1 (abstract P390). Length of stay (minutes) in the ED. Results The immobilisation time was 207 ± 60 seconds. P388
Eff ects of clonidine on microcirculatory alterations during
endotoxemia
K S h
idt C Phili
b
A Zi k
i
S H f P388
Eff ects of clonidine on microcirculatory alterations during
endotoxemia
K Schmidt, C Philipsenburg, A Zivkovic, S Hofer
Universitätsklinikum Heidelberg, Germany
Critical Care 2013, 17(Suppl 2):P388 (doi: 10.1186/cc12326) Conclusion Inhaled anesthetics could be successfully used in the ICU
especially in case of an inadequate sedation plan; for example, in
patients with a history of drug abuse or young severe ARDS patients
that required deep sedation and paralysis for a long period. Introduction
Endothelial
dysfunction
during
endotoxemia
is
responsible for the functional breakdown of microvascular perfusion
and microvessel permeability. The cholinergic anti-infl ammatory
pathway (CAP) is a neurophysiological mechanism that regulates
the infl ammatory response by inhibiting proinfl ammatory cytokine
synthesis, thereby preventing tissue damage. Endotoxemia-induced Figure 1 (abstract P387). Cellular oxygen consumption after incubation with fentanyl, naloxone or 5-HD. S146 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 eff ect of ketamine or due to the method of administration. There
were no signs of conjunctival irritation in any animal (S+ketamine is a
preservative-free solution). microcirculatory dysfunction can be reduced by cholinergic CAP
activation. Clonidine improves survival in experimental sepsis [1] by
reducing the sympathetic tone, resulting in the parasympathetic-
mediated CAP activation. The aim of this study was to determine the
eff ects of clonidine on microcirculatory alterations during endotoxemia. Methods Using fl uorescent intravital microscopy, we determined
the venular wall shear rate, macromolecular effl ux and leukocyte
adhesion in mesenteric postcapillary venules of male Wistar rats. Endotoxemia was induced over 120 minutes by intravenous infusion
of lipopolysaccharide (LPS). Control groups received an equivalent
volume of saline. Clonidine 10 μg/kg was applied as i.v. bolus in
treatment groups. Animals received either (i) saline alone, (ii) clonidine
10 minutes prior to saline administration, (iii) clonidine 45 minutes
prior to LPS administration, (iv) clonidine 10 minutes prior to LPS
administration, (v) clonidine 30 minutes after LPS administration or
(vi) LPS alone. p
Conclusion Conjunctival S+ketamine 2.5 mg/kg in rabbits produced
rapid onset without changes in cardiorespiratory parameters and
without signs of irritation of the eye. The results of our project warrant
further research to increase the variety of drugs and methods of
their administration for anxiolysis, sedation and analgesia in disaster
medicine. Acknowledgement The study was supported by scientifi c grant IGA NT
11284. Reference 1. Malek J, et al.: Conjunctival administration of sufentanil in rabbits. P388
Eff ects of clonidine on microcirculatory alterations during
endotoxemia
K S h
idt C Phili
b
A Zi k
i
S H f Eur J Pain
2011, Suppl 5:212. 1. Malek J, et al.: Conjunctival administration of sufentanil in rabbits. Eur J Pain
2011, Suppl 5:212. Results All LPS groups (iii to vi) showed a signifi cantly reduced venular
wall shear rate compared with the saline group after 120 minutes. There
were no signifi cant diff erences between the numbers of adhering
leukocytes in the clonidine/LPS groups (iii, iv, v) and the LPS group
after 120 minutes. Macromolecular effl ux signifi cantly increased in all
groups over the time period of 120 minutes. After 120 minutes there
was no diff erence between the LPS group and the clonidine 10 minutes
prior to LPS administration group (iv) whereas all other groups (i, ii, iii, v)
showed a signifi cantly reduced macromolecular effl ux compared with
the LPS group.f Use of propofol for procedural sedation reduces length of stay in
the emergency department
J Millar, F Adamson, P O’Connor, R Wilson, E Ferrie, R McLaughlin
Royal Victoria Hospital, Belfast, UK
Critical Care 2013, 17(Suppl 2):P390 (doi: 10.1186/cc12328) Use of propofol for procedural sedation reduces length of stay in
the emergency department
J Millar, F Adamson, P O’Connor, R Wilson, E Ferrie, R McLaughlin
Royal Victoria Hospital, Belfast, UK
Critical Care 2013, 17(Suppl 2):P390 (doi: 10.1186/cc12328) Introduction Procedural sedation is used in the emergency department
(ED) to facilitate short but painful interventions. Many patients are
suitable for discharge after completion. Ideally, the agent used to
achieve sedation should not have a prolonged eff ect, allowing safe
discharge in the shortest time frame. We hypothesised that propofol,
with its short onset and off set, may reduce length of stay (LOS) in
comparison with traditional benzodiazepines. Conclusion Clonidine has no positive eff ect on microhemodynamic
alterations and leukocyte–endothelial interaction during endotoxemia. The reduction of capillary leakage in clonidine-treated groups
depends on the time interval relative to the initiation of endotoxemia. Endothelial permeability and leukocyte activation are regulated by
diff erent pathways when stimulated by clonidine during endotoxemia. We conclude that clonidine might have an important time-dependent
anti-infl ammatory and protective eff ect on endothelial activation
during infl ammation. Methods Data from a prospective registry were analysed for the
period 1 August 2011 to 31 January 2012. Patients who underwent
procedural sedation and who were discharged from the ED were
identifi ed. Individuals were grouped as having received propofol,
midazolam or a combination of the two. All were discharged when
fully alert and able to eat and drink. Demographic details and the type
of procedure undertaken were extracted. ANOVA was performed to
identify diff erences in the length of stay between groups, in addition
to descriptive analysis. Reference Reference 1. Hofer S, et al.: Crit Care 2009, 13:R11. 1. Hofer S, et al.: Crit Care 2009, 13:R11. 1. Hofer S, et al.: Crit Care 2009, 13:R11. P393 Use of physical restraint in Dutch ICUs: prevalence and motives
RJ Raijmakers1, RL Vroegop1, H Tekatli1, M Van den Boogaard2,
AW Van der Kooi1, AJ Slooter1
1University Medical Center Utrecht, the Netherlands; 2Radboud University
Nijmegen Medical Centre, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P393 (doi: 10.1186/cc12331) Use of physical restraint in Dutch ICUs: prevalence and motives
RJ Raijmakers1, RL Vroegop1, H Tekatli1, M Van den Boogaard2,
AW Van der Kooi1, AJ Slooter1
1University Medical Center Utrecht, the Netherlands; 2Radboud University
Nijmegen Medical Centre, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P393 (doi: 10.1186/cc12331) Results Data were collected from 54 adults; 50 received an infusion for
≥24 hours. We noted a signifi cant CK rise in 11 (22%) of our patients
that could be attributed to propofol alone. They all had raised TG
(≥1.7 IU/l); six (12%) were markedly raised (≥3.4 IU/l). No patients
became signifi cantly acidotic, developed cardiac arrhythmias or renal
failure as a result of propofol. In the 11 patients who met our criteria,
highest daily propofol dose varied from 1.5 to 4 mg/kg/hour. Pre-PRIS
developed from between 2 and 6 days of infusion. The mean propofol
dose in patients displaying Pre-PRIS was 2.2 mg/kg/hour whilst in those
not meeting our criteria it was 2.0 mg/kg/hour. No patients developed
sequelae of full PRIS. See Table 1. Introduction Physical restraints are used to facilitate essential care
and prevent secondary injuries. However, physical restraint may be
regarded as humiliating. It may lead to local injury and increase the risk
of delirium and post-traumatic stress syndrome. Research on physical
restraint is scarce. The aim of this study is to investigate the scope of
physical restraint use. Methods Twenty-one ICUs ranging from local hospitals to academic
centres were each visited twice and 327 patients were included. We
recorded characteristics of restrained patients, motives and awareness
of nurses and physicians. Table 1 (abstract P391)
Total
Trauma
Nontrauma
≥24 hours infusion
50
34
16
CK + TG ≥3.4
6
6
0
CK + TG ≥1.7
5
3
2 Results Physical restraint was applied in 74 (23%) patients, ranging
from 0 to 54% in diff erent hospitals. Frequent motives for restraint
use were ‘possible threat to airway’ (36%) and ‘pulling lines/probes’
(31%). Restrained subjects more often had a positive CAM-ICU (34%
vs. 16%, P <0.001), could less frequently verbally communicate (14%
vs. 49%, P <0.001), and received more often antipsychotics (49% vs. P394 Factors infl uencing the use of physical restraints in Canadian ICUs
B Sneyers1, E Luk2, M Perreault3, D Williamson3, L Rose2, S Mehta4, L Burry4
1Université Catholique de Louvain, Brussels, Belgium; 2Lawrence S. Bloomberg
Faculty of Nursing, University of Toronto, Ontario, Canada; 3Université de
Montréal, Canada; 4Mount Sinai Hospital, University of Toronto, Ontario,
Canada
Critical Care 2013, 17(Suppl 2):P394 (doi: 10.1186/cc12332) Factors infl uencing the use of physical restraints in Canadian ICUs
B Sneyers1, E Luk2, M Perreault3, D Williamson3, L Rose2, S Mehta4, L Burry4
1Université Catholique de Louvain, Brussels, Belgium; 2Lawrence S. Bloomberg
Faculty of Nursing, University of Toronto, Ontario, Canada; 3Université de
Montréal, Canada; 4Mount Sinai Hospital, University of Toronto, Ontario,
Canada
Critical Care 2013, 17(Suppl 2):P394 (doi: 10.1186/cc12332) P389 There were
no changes in cardiorespiratory parameters (initial, 10 and 20 minutes
after administration: HR 255.5 ± 24.7, 265.8 ± 26.0 and 267.8 ± 20.4,
systolic BP 114.3 ± 8.6, 108.2 ± 10.7 and 113.9 ± 11.9 mmHg, diastolic
BP 66.8 ± 11.6, 68.4 ± 10.6 and 69.7 ± 9.3 mmHg and SpO2 99.2 ± 1.0,
98.2 ± 0.6 and 98.7 ± 0.5). These results are in contrast to conjunctival
administration of sufentanil 5 μg/kg that caused a signifi cant decrease
of SpO2 and HR [1]. We can speculate that the reason for stability of
cardiorespiratory parameters was due to the sympathoadrenergic S147 Critical Care 2013, Volume 17 Suppl 2
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Pre-PRIS? Prospective monitoring for early markers of propofol
infusion syndrome
M Stovell, S Smith, M Udberg, W Loh, P Nair
Walton Centre Neurology & Neurosurgery, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P391 (doi: 10.1186/cc12329) P391 Conclusion The study shows a 1% adverse event rate. This supports use
of propofol sedation by emergency physicians but within the limits of a
strict governance framework. Our safety analysis using the World SIVA
adverse events tool provides a reference point for further studies. R f P391
Pre-PRIS? Prospective monitoring for early markers of propofol
infusion syndrome 1. Green SM, Yealy DM: Procedural sedation goes Utstein: the Quebec
guidelines. Ann Emerg Med 2009, 53:436-438. 1. Green SM, Yealy DM: Procedural sedation goes Utstein: the Quebec
guidelines. Ann Emerg Med 2009, 53:436-438. 1. Green SM, Yealy DM: Procedural sedation goes Utstein: the Quebec
guidelines. Ann Emerg Med 2009, 53:436-438. Introduction The use of propofol for sedation in intensive care has been
associated with the propofol infusion syndrome (PRIS) characterised by
cardiac dysfunction, metabolic acidosis, renal failure, rhabdomyolysis
and hyperlipidaemia. We prospectively monitor biochemical markers
that we believe demonstrate early signs of this dangerous, often fatal
syndrome. When this Pre-PRIS state is identifi ed, propofol is withdrawn
whilst the syndrome is still reversible. 2. Green SM, Krauss B: Barriers to propofol use in emergency medicine. Ann
Emerg Med 2008, 52:392-398. 3. Mason KP, Green SM, Piacevoli Q; International Sedation Task Force: Adverse
event reporting tool to standardize the reporting and tracking of adverse
events during procedural sedation: a consensus document from the
World SIVA International Sedation Task Force. Br J Anaesth 2012, 108:13-20. 3. Mason KP, Green SM, Piacevoli Q; International Sedation Task Force: Adverse
event reporting tool to standardize the reporting and tracking of adverse
events during procedural sedation: a consensus document from the
World SIVA International Sedation Task Force. Br J Anaesth 2012, 108:13-20. y
Methods We prospectively audited our monitoring of these markers
over a 4-month period in propofol-sedated patients: propofol infusion
rate, creatine kinase (CK), triglycerides (TG), creatinine, lactate, pH
and base defi cit. We defi ned the criteria for Pre-PRIS as requiring a
CK ≥320 mmol/l that had doubled from its base level and a rise in TG
≥1.7 IU/l; both that followed a trend with propofol dose. P393 28%, P <0.001), or benzodiazepines (55% vs. 36%, P = 0.003). The use
of physical restraint was registered in the patient’s fi les in 48% of cases. Of the 310 interviewed nurses, 258 (83%) were familiar with a physical
restraint protocol and 89 (29%) used it in any situation. Thirty percent
of the 60 interviewed physicians were aware of the physical restraint
status of their patients. Conclusion We propose that a paired rise in CK and TG that can be
attributed to propofol alone represents a Pre-PRIS state that is at risk
of developing into full PRIS. We noted this in 22% of our patients, all
on modest doses of propofol. It is unclear what proportion of patients
will develop the full syndrome as it is not ethically possible to continue
propofol in this situation. We advocate daily monitoring of CK and TG
to identify Pre-PRIS so that propofol can be reduced or substituted to
avoid the morbidity and mortality of the full syndrome. Conclusion Physical restraint is frequently used in Dutch ICUs, but the
frequency diff ers strongly between diff erent ICUs. Attending physicians
are often not aware of physical restraint use. P392
Safety profi le of 1,008 cases of propofol sedation in the emergency
department Safety profi le of 1,008 cases of propofol sedation in the emergency
department S Bradburn, G Lloyd, B Newstead
Royal Devon and Exeter Hospital, Exeter, UK
Critical Care 2013, 17(Suppl 2):P392 (doi: 10.1186/cc12330) Introduction Until recently there were no guidelines for the reporting
of adverse events (AEs) during procedural sedation [1,2]. A consensus
document released in 2012 by the world SIVA International Sedation
Task Force proposed a benchmark for defi ning AEs [3]. We analysed
1,008 cases of procedural sedation in the emergency department. Introduction Physical restraint (PR) use in critically ill patients has
been associated with delirium, unplanned extubation, prolonged ICU
length of stay, and post-traumatic stress disorder. Our objectives were
to defi ne prevalence of PR use, and to examine patient, treatment, or
institutional factors associated with their use in Canadian ICUs. Introduction Physical restraint (PR) use in critically ill patients has
been associated with delirium, unplanned extubation, prolonged ICU
length of stay, and post-traumatic stress disorder. Our objectives were
to defi ne prevalence of PR use, and to examine patient, treatment, or
institutional factors associated with their use in Canadian ICUs. Methods The study is based on 1,008 patients who received procedural
sedation with propofol in the emergency department between
December 2006 and March 2012. Patients were selected and sedated
to a strict protocol by ED consultant staff . We applied the AE tool
by performing a search through patient records, discussion with
consultants performing the sedation and consensus opinion.i Methods We conducted an analysis of a database of sedative, analgesic,
antipsychotic and neuromuscular blocker prescribing practices and PR
use in 711 mechanically ventilated (MV) adults in 51 Canadian ICUs
(3,619 patient-days). Data were collected during a 2-week period (2008
to 2009). Logistic and Poisson regression analysis were used to identify
factors associated with PR use and number of days of use respectively. Results PR was used on 1 or more days in 52.6% (374/711) of patients. For patients restrained, the mean number of days restrained was Results From 1,008 cases we identifi ed 11 sentinel (six of hypotension,
fi ve cases of hypoxia), 34 moderate, 25 minor and three minimal risk
adverse events. S148 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 4.1 ± 4.0 and the mean number of restraint applications per patient was
1.2 ± 0.5. Current perspectives, beliefs and practices concerning delirium in
critically ill patients: a multicenter survey among Dutch healthcare
professionals Z Trogrlic, M Van der Jagt, P Van der Voort, J Bakker, E Ista, on behalf of
iDECePTIvE Study Group Results We assessed 1,576 patients, corresponding with 8,150 ICU
treatment days with a median length of stay of 3 days (IQR 2 to 5). The mean age of the patients was 62 years (SD = 16) and 58% were
male. Delirium occurred in 23% (356/1,576) of patients with a median
duration of 3 days (IQR 2 to 7) and ranged from 11 to 40% for each ICU. Delirium assessment with the CAM-ICU at any point during ICU stay
was performed in 38% of all patients. Screening with CAM-ICU was
applied in three ICUs, in 29 to 96% of the patients in these centers. Of
3,564 documented screening days with the CAM-ICU, it was positive
in 1,459 (41%); in only 120 (8%) of these CAM-ICU-positive days there
was a documented action or treatment started for delirium. However,
patients still received haloperidol on 52% (n = 766) of all CAM-ICU-
positive days. Patients received benzodiazepines in 49% (n = 1,141)
of patient sedation days. Delirium preventive interventions were
physiotherapy (19% of 8,150 ICU days), mobilization (10%), glasses
use (2.6%) and hearing aid use (0.3%). Presence of hearing or visual
impairment at admission was not documented in 65% of patients. Erasmus MC University Medical Center, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P397 (doi: 10.1186/cc12335) Introduction The beliefs, knowledge and practices regarding ICU
delirium among ICU professionals may vary. This may interfere with
the implementation of the Dutch ICU Delirium guideline. We aimed to
get insight into potential barriers and facilitators for delirium guideline
implementation that may help to fi nd an eff ective implementation
strategy. Methods An online survey was sent to healthcare professionals from
the six participating ICUs. Respondents included ICU physicians, nurses
and delirium experts (psychiatrists, neurologists, geriatricians, nurse
experts). The survey consisted of statements on beliefs, knowledge
and practices towards ICU delirium. Agreement with statements by
more than 75% of respondents were regarded as facilitating items and
agreement lower than 50% as barriers for implementing protocolled
care. Conclusion Daily screening for ICU delirium with a validated screening
instrument is applied in less than one-half of the time in critically ill
patients and management of delirium is often not guided by this
screening. Haloperidol was used as the fi rst-choice medication. P395 Current practices in ICU delirium management: a prospective
multicenter study in the Netherlands
Z Trogrlic, E Ista, A Slooter, J Bakker, M Van der Jagt, on behalf of
iDECePTIvE Study Group
Erasmus MC University Medical Center, Rotterdam, the Netherlands
Critical Care 2013, 17(Suppl 2):P395 (doi: 10.1186/cc12333) Results Of 768 eligible patients, complete data on delirium could
be collected in 738 patients. The incidence of delirium was 14.2% in
the dexamethasone group and 14.9% in the placebo group (odds
ratio = 0.95, 95% CI = 0.63 to 1.43). No signifi cant diff erence was found
on the duration of delirium between the intervention (median = 2 days,
interquartile range 1 to 3 days) and placebo (median = 2 days,
interquartile range 1 to 2 days) group (P = 0.45). The use of restrictive
measures and administration of sedatives, haloperidol, benzodiazepine
and opiates were comparable between both groups. Introduction As part of a multicenter prospective study on barriers
and facilitators for implementation of protocolled care for delirious
critically ill patients, we aimed to describe current practices in delirium
management in a representative cohort of ICU patients. p
p
g
p
Conclusion Intraoperative injection of dexamethasone seems not
to aff ect the incidence or duration of delirium in the fi rst 4 days after
cardiac surgery, suggesting this regimen is safe to use in the operative
setting with respect to psychiatric adverse events. Reference Methods We included consecutively admitted patients during a
4-month period in a prospective multicenter study in six ICUs (one
academic, teaching and nonteaching hospitals) in the southwest of
the Netherlands. We assessed: percentage of patients screened for
delirium with validated screening tools; pharmacological treatment
with haloperidol or other antipsychotic drugs; psychohygiene (use of
hearing aids and or glasses, preventing sleep disturbances); and access
to early mobilization and physiotherapy. Delirium was pragmatically
defi ned as administration of haloperidol and/or delirium reported by
physicians or ICU nurses in patient records, as assessed by a designated
research nurse. Diff erences between centers were tested with non-
parametric tests. 1. Dieleman JM, et al.: JAMA 2012, 308:1761-1767. 1. Dieleman JM, et al.: JAMA 2012, 308:1761-1767. Intraoperative dexamethasone and delirium after cardiac surgery:
a randomized clinical trial
AC Sauër, AJ Slooter, DS Veldhuijzen, MM Van Eijk, D Van Dijk
University Medical Center Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P396 (doi: 10.1186/cc12334) Introduction The objective of this study is to investigate the eff ect of
intraoperative administration of dexamethasone versus placebo on the
incidence of delirium in the fi rst four postoperative days after cardiac
surgery. Introduction The objective of this study is to investigate the eff ect of
intraoperative administration of dexamethasone versus placebo on the
incidence of delirium in the fi rst four postoperative days after cardiac
surgery. Methods Within the context of the large multicenter Dexamethasone
for Cardiac Surgery (DECS) trial [1] for which patients were randomized
to 1 mg/kg dexamethasone or placebo at induction of anesthesia, a
monocenter substudy was conducted. The primary outcome of this
study was the incidence of delirium in the fi rst four postoperative
days. Secondary outcomes were duration of delirium, use of restrictive
measures and sedative, antipsychotic and analgesic requirements. Delirium was assessed daily by trained research personnel, using the
Richmond Agitation Sedation Scale and the Confusion Assessment
Method. Medical, nursing and medication charts were evaluated for
signs of delirium and use of prespecifi ed medication. Analysis was by
intention to treat. Conclusion We identifi ed multiple factors associated with PR use
and number of days of PR use. Particularly among those related to
treatment, strategies such as daily sedation interruption or use of
antipsychotics are associated with PR use and increasing the number
of days of use. P392
Safety profi le of 1,008 cases of propofol sedation in the emergency
department Treatment characteristics associated with PR use were: daily
benzodiazepine dose (OR = 1.01, 95% CI: 1.00 to 1.01), daily opiate dose
(OR = 1.00, 95% CI: 1.00 to 1.01), daily sedation interruption (OR = 1.84,
95% CI: 1.24 to 2.72), use of antipsychotics (OR = 3.42, 95% CI: 1.98 to
5.91) and agitation (Sedation–Agitation Scale (SAS) >4, OR = 2.55, 95%
CI: 1.20 to 5.44). Additionally, number of days of PR use was increased
according to the following: daily sedation interruption (IRR = 4.52,
95% CI: 2.09 to 9.75), use of antipsychotics (IRR = 18.65, 95% CI: 7.95
to 43.77), the presence of a heavily sedated score (SAS <3, IRR = 2.57,
95% CI: 1.06 to 6.24) or the presence of an adverse event, such as self-
extubation (IRR = 10.46, 95% CI: 2.55 to 42.92). Hospital characteristics
associated with PR use included the proportion of MV patients in
the ICU. University hospitals were associated with reduced PR use,
compared with community hospitals (OR = 0.272, 95% CI: 0.15 to 0.50). Nonmodifi able patient characteristics (age, sex, APACHE score, patient
category, prior substance abuse, prior psychotropic medication, pre-
existing psychiatric condition or dementia) were not associated with
PR use. mobilization) were carried only in a small minority or were not
documented. To implement protocolled delirium care in the region at
study, a multifaceted tailored implementation program is needed. P396 Intraoperative dexamethasone and delirium after cardiac surgery:
a randomized clinical trial
AC Sauër, AJ Slooter, DS Veldhuijzen, MM Van Eijk, D Van Dijk
University Medical Center Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P396 (doi: 10.1186/cc12334) International validation of the delirium prediction model for ICU
patients (PREDELIRIC): a multicenter observati onal study Methods We included patients in whom days with delirium could
be compared with days without delirium, based on the Confusion
Assessment Method for the ICU and inspection of medical records. Patients with conditions aff ecting thermal regulation, including
infectious diseases, and those receiving therapies aff ecting body
temperature were excluded. Twenty-four ICU patients were included
after screening 334 delirious ICU patients. Daily temperature variability
was determined by computing the mean absolute second derivative
of the temperature signal. Per patient, temperature variability
during delirious days was compared with nondelirium days using a
Wilcoxon signed-rank test. With a linear mixed model, diff erences
between delirium and nondelirium days with regard to temperature
variability were analysed adjusted for daily mean Richmond Agitation
and Sedation Scale scores, daily maximum Sequential Organ Failure
Assessment score, and within-patient correlation. M van den Boogaard1, L Schoonhoven2, E Maseda3, C Plowright4,
C Jones5, A Luetz6, PV Sackey7, P Jorens8, LM Aitken9, F van Haren10,
JG van der Hoeven1, P Pickkers1 M van den Boogaard1, L Schoonhoven2, E Maseda3, C Plowright4,
C Jones5, A Luetz6, PV Sackey7, P Jorens8, LM Aitken9, F van Haren10,
JG van der Hoeven1, P Pickkers1 1Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands;
2Radboud University Nijmegen Medical Centre, Scientifi c Institute for Quality
of Healthcare, Nijmegen, the Netherlands; 3Hospital Universitario La Paz,
Madrid, Spain; 4Medway Maritime Hospital, Kent, UK; 5Whiston Hospital,
Prescot, UK; 6Charité – Universitaetsmedizin Berlin, Germany; 7Karolinska
University Hospital, Stockholm, Sweden; 8University of Antwerpen, Belgium;
9Princess Alexandra Hospital, Woolloongabba, Australia; 10Canberra Hospital,
Canberra, Australia Critical Care 2013, 17(Suppl 2):P398 (doi: 10.1186/cc12336) Critical Care 2013, 17(Suppl 2):P398 (doi: 10.1186/cc12336) Critical Care 2013, 17(Suppl 2):P398 (doi: 10.1186/cc12336) Introduction Delirium is a serious and frequent disorder in ICU patients. The aim of this study was to internationally validate the existing
Prediction of Delirium ICU (PREDELIRIC) model to predict delirium in
ICU patients. Results Temperature variability was increased during delirium days
compared with days without delirium (mean diff erence = –0.007, 95%
CI = –0.004; –0.011, P <0.001). Adjusting for confounders did not alter
our fi ndings (adjusted mean diff erence = –0.005, 95% CI = –0.008;
–0.002, P <0.001). Methods A prospective multicenter cohort study was performed in
eight ICUs in six countries (Table 1). The 10 predictors (age, APACHE
II score, urgent and admission category, infection, coma, sedation,
morphine use, urea level, metabolic acidosis) were collected within
24 hours after ICU admission. P398 International validation of the delirium prediction model for ICU
patients (PREDELIRIC): a multicenter observati onal study
M van den Boogaard1, L Schoonhoven2, E Maseda3, C Plowright4,
C Jones5, A Luetz6, PV Sackey7, P Jorens8, LM Aitken9, F van Haren10,
JG van der Hoeven1, P Pickkers1 Current perspectives, beliefs and practices concerning delirium in
critically ill patients: a multicenter survey among Dutch healthcare
professionals Measures aimed at delirium prevention (psychohygiene and early Results Of the 565 surveys distributed, 360 were completed (63.7%). The majority of respondents were ICU nurses (79%). Delirium was
considered a major problem (83%) that requires adequate treatment
(99%) and is underdiagnosed (81%). Respondents considered that S149 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 routine screening of delirium can improve prognosis (95%). However,
only a minority (20%) answered that delirium is preventable. Only
39% of the respondents had received any training about delirium
in the previous 3 years and 77% of them found training useful. The
mean delirium knowledge score was 6.6 out of 10 (SD = 1.54). When
all groups were mutually compared, nurses scored lower than delirium
experts (ANOVA, P = 0.013). The respondents (58%; n = 210) from three
ICUs indicated that CAM-ICU assessment was department policy. However, 50% (n = 106) of these respondents felt unfamiliar with CAM-
ICU and only 47% (n = 99) of them indicated that a positive CAM-ICU
was used for treatment decisions. Haloperidol was the fi rst-choice
pharmacological treatment. Only 21% of all respondents knew that a
national ICU delirium guideline existed, but in-depth knowledge was
generally low. was 82 ± 16% and inter-rater reliability 0.87 ± 0.17. Overall delirium
incidence was 19.9% (range 27.2%). Despite signifi cant diff erences
between centres on all 10 predictors, the overall area under the receiver
operating characteristic curve (AUROC) of the eight centers was 0.77
(95% CI: 0.74 to 0.79). was 82 ± 16% and inter-rater reliability 0.87 ± 0.17. Overall delirium
incidence was 19.9% (range 27.2%). Despite signifi cant diff erences
between centres on all 10 predictors, the overall area under the receiver
operating characteristic curve (AUROC) of the eight centers was 0.77
(95% CI: 0.74 to 0.79). Conclusion The overall predictive value of the PREDELIRIC model
was good, indicating that the prediction model can now be used
internationally. P399 Delirium in ICU patients is associated with increased temperature
variability
AW Van der Kooi, TH Kappen, RJ Raijmakers, IJ Zaal, AJ Slooter
University Medical Center Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P399 (doi: 10.1186/cc12337) Delirium in ICU patients is associated with increased temperature
variability Delirium in ICU patients is associated with increased temperature
variability y
AW Van der Kooi, TH Kappen, RJ Raijmakers, IJ Zaal, AJ Slooter
University Medical Center Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P399 (doi: 10.1186/cc12337) y
AW Van der Kooi, TH Kappen, RJ Raijmakers, IJ Zaal, AJ Slooter
University Medical Center Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P399 (doi: 10.1186/cc12337) y
Conclusion Our survey showed that healthcare professionals
considered delirium an important but underdiagnosed form of
organ failure. In contrast, screening tools for delirium are scarcely
used, knowledge can be improved and protocolled treatment based
on positive screening is often lacking. These results suggest that the
focus of implementation of ICU delirium management should not be
on motivational aspects, but on knowledge improvements, training
in screening tools and implementation of treatment and prevention
protocols. Introduction Delirium is an acute disturbance of consciousness
and cognition. It is a common disorder in the ICU and associated
with impaired long-term outcome [1,2]. Despite its frequency and
impact, delirium is poorly recognized by ICU physicians and nurses
using delirium screening tools [3]. A completely new approach to
detect delirium is to use monitoring of physiological alterations. Temperature variability, a measure for temperature regulation, could
be an interesting parameter for monitoring of ICU delirium, but this has
never been investigated before. The aim of this study was to investigate
whether temperature variability is aff ected during ICU delirium. International validation of the delirium prediction model for ICU
patients (PREDELIRIC): a multicenter observati onal study CAM-ICU was used to identify ICU
delirium. Conclusion Temperature variability is increased during delirium
in ICU patients, which refl ects the encephalopathy that underlies
delirium. Opportunities for delirium monitoring using temperature
variability should be further explored. Particularly, in combination with
electroencephalography it could provide the input for an objective tool
to monitor delirium. Results A total of 2,852 adult ICU patients were screened and 1,824
(64%) were included. Main exclusion reasons were length of stay <1 day
(19.1%) and sustained coma (4.1%). Mean ± CAM-ICU compliance Gastrointestinal symptoms in the ICU: a comparison of primary and
secondary acute gastrointestinal injury A Reintam Blaser, J Starkopf ,
p
University of Tartu, Tartu University Hospital, Tartu, Estonia y
y
Critical Care 2013, 17(Suppl 2):P401 (doi: 10.1186/cc12339) Introduction We aimed to clarify the diff erences between primary and
secondary acute GI injury. Methods A total of 2,690 consecutive adult patients were retro-
spectively studied during their fi rst week in the ICU. Pathology in the
GI system or laparotomy defi ned the primary GI insult. If GI symptoms
developed without primary GI insult it was considered secondary GI
injury. Absent bowel sounds (BS), vomiting/regurgitation, diarrhoea,
bowel distension, GI bleeding, and high gastric residuals (GRV
>1,000 ml/24 hours) were recorded daily. ventilation with relative risk of 9 to 27% and with mortality of 25 to
50% [1,2]. One of the promoting factors of VAP is the increased pH of
the gastric acid, which occurs when H2-receptor antagonists (H2RA)
or proton pump inhibitors (PPI) are used for stress ulcer prophylaxis. The aim of this study was to investigate eff ects of PPI versus H2RA
on the occurrence of VAP and, as second endpoint, the occurrence of
gastrointestinal bleeding (GIB). y
Results In total, 2,690 patients (60.4% male), median age 59 years
(range 16 to 92), were studied. Eighty-four per cent of them were
ventilated, 72% received vasopressor/inotrope. Median (IQR) APACHE
II score was 14 (9 to 21) and SOFA on the fi rst day was 6 (3 to 10). A
total 35.5% had primary GI pathology. During the fi rst week 34% of
patients had absent BS, 38% vomiting/regurgitation, 9% diarrhoea,
7% bowel distension, 6% high GRV and 5% GI bleeding. All symptoms
except diarrhoea occurred more often (<0.001) in patients with primary
GI insult. Eighty-fi ve per cent of patients with primary GI insult versus
46% without developed at least one GI symptom. The incidence of GI
symptoms was signifi cantly higher in nonsurvivors. ICU mortality was
lower in patients with primary than secondary GI injury (43.6% vs. 61.2%, P = 0.046). Nonsurvivors without primary GI insult developed GI
symptoms later (Figure 1).f Methods After ethics committee approval mechanically ventilated
(>48 hours) adults were recruited in a prospective randomized
controlled, double-blind clinical trial and randomised into PPI or H2RA
groups. Patients with admission diagnosis of pneumonia or other
acute infl ammatory pulmonary disease, severe sepsis/septic shock,
and/or regular PPI/H2RA users were excluded. References 1. Pisani MA, et al.: Am J Respir Crit Care Med 2009, 180:1092-1097. 2. Ely EW, et al.: JAMA 2004, 291:1753-1762. 1. Pisani MA, et al.: Am J Respir Crit Care Med 2009, 180:1092-1097. 2
El EW t l JAMA 2004 291 1753 1762 Table 1 (abstract P398). PREDELIRIC AUROC of diff erent centers Table 1 (abstract P398). PREDELIRIC AUROC of diff erent centers Table 1 (abstract P398). PREDELIRIC AUROC of diff erent centers
Delirium (%)
AUROC
95% CI
Belgium (%)
86 (15.2)
0.79
0.74 to 0.84
Germany (%)
60 (26.9)
0.85
0.80 to 0.91
Spain (%)
23 (18.0)
0.88
0.81 to 0.94
Sweden (%)
30 (39.0)
0.71
0.60 to 0.83
Australia Brisbane (%)
42 (12.8)
0.81
0.75 to 0.88
Australia Canberra (%)
23 (11.8)
0.80
0.72 to 0.89
UK_Prescot (%)
73 (30.8)
0.65
0.57 to 0.73
UK_Kent (%)
26 (36.6)
0.76
0.65 to 0.88
PREDELIRIC overall
363 (19.9)
0.77
0.74 to 0.79 y
3. van Eijk MM, et al.: Am J Respir Crit Care Med 2011, 184:340-344. P400
Long-term outcome of delirium in critically ill patients
A Wolters, D Van Dijk, O Cremer, D De Lange, A Slooter
UMC Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P400 (doi: 10.1186/cc12338) Long-term outcome of delirium in critically ill patients
A Wolters, D Van Dijk, O Cremer, D De Lange, A Slooter
UMC Utrecht, the Netherlands
Critical Care 2013, 17(Suppl 2):P400 (doi: 10.1186/cc12338) Introduction In ICU patients, little research has been performed on
the relationship between delirium and long-term outcome, including
health-related quality of life (HRQoL), cognitive functioning and
mortality. In addition, results seem to be inconsistent. Furthermore,
in studies that reported increased mortality in delirious patients,
no proper adjustments were made for severity of illness during ICU
admission. This study was conducted to investigate the association S150 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Figure 1 (abstract P401). Daily dynamics of the number of GI symptoms. P <0.001 between survivors and nonsurvivors in all time points. between ICU delirium and long-term HRQoL, cognitive functioning
and mortality. The hypothesis was that delirious patients have worse
long-term outcome in comparison with nondelirious patients. between ICU delirium and long-term HRQoL, cognitive functioning
and mortality. The hypothesis was that delirious patients have worse
long-term outcome in comparison with nondelirious patients. Methods A prospective observational cohort study was conducted. Gastrointestinal symptoms in the ICU: a comparison of primary and
secondary acute gastrointestinal injury The diagnostic criteria
for VAP were: leukocytosis, increased PCT level, fever, purulent tracheal
secretion, positive microbiological result of tracheal sample, and new/
increased infi ltrate on chest X-ray. Data are presented as median
(interquartile range, IQR). For statistical analysis Mann–Whitney U test
and chi-square tests were used. q
Results Out of 94 patients recruited 14 were excluded due to early
weaning/death (n = 9) or protocol violation (n = 6). The remaining
79 patients were analysed (PPI, n = 38; H2RA, n = 41). There was
no signifi cant diff erence (P >0.05) between the groups regarding
demographics: age 67 (56 to 77) versus 72 (58 to 79) years; male/
female: 23/15 versus 25/16; length of mechanical ventilation: 5 (3 to
9) versus 5 (2 to 8) days; APACHE II score: 28 (22 to 32) versus 26 (21 to
36) (PPI vs. H2RA, respectively). There was no signifi cant diff erence in
the number of cases with VAP in the PPI versus H2RA groups: 9 (24%)
versus 10 (24%). None of the patients developed GI bleeding during
their stay on the ICU. Conclusion Primary and secondary acute GI injury have diff erent
incidence, dynamics and outcome. References A median of 12 months after ICU discharge, questionnaires were sent
to all survivors. HRQoL and cognitive functioning were measured with
the EuroQol-6D. Age, gender and severity of illness were considered
relevant covariates. Severity of illness was estimated using the APACHE
IV score and the maximal SOFA score during admission. HRQoL was
investigated with linear regression analysis, cognitive functioning
using logistic regression and mortality with Cox regression analysis. y
y
Results The patient population consisted of 690 patients admitted to
the ICU, subdivided into delirious (n = 257) and nondelirious patients
(n = 433). During follow-up, 181 (26%) patients died. The response rate
of the questionnaire was 70.6%. After adjusting for the predefi ned
covariates, delirium was signifi cantly associated with a lower HRQoL
(adjusted β: –0.137; 95% CI = –0.140 to –0.005) and more mild and
severe cognitive impairment (adjusted odds ratio: respectively: 2.3; 95%
CI = 1.3 to 4.2 and 5.8; 95% CI = 1.3 to 15.2). No signifi cant association
between delirium and long-term mortality was found (adjusted hazard
ratio: 1.0; 95% CI = 0.7 to1.4). Conclusion Delirium during ICU admission was associated with
lower HRQoL and worse cognitive functioning, 1 year after discharge. Furthermore, delirium on the ICU was not associated with long-term
mortality after adjusting for relevant covariates, including severity of
illness during ICU admission. Early diagnosis of liver failure in septic patients using the
maximal liver function capacity test (LiMAx test): comparison with
conventional methods M Kaff arnik, M Stockmann, J Lock
Charite Berlin Campus Virchow, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P403 (doi: 10.1186/cc12341) Introduction Patients with bacterial sepsis often suff er from multiorgan
failure. No reliable parameter exists for the exact estimation of the liver
function. The indocyaningreen test (ICG test) showed a correlation
between liver function and mortality rate. The new maximal liver
function capacity test (LiMAx test) is a real-time method to investigate
the liver function. g
p
pp
Results HVPG increased in Group 1 from 3.7 to 5.4 mmHg (P <0.05) and
decreased in Group 2 (4.8 to 4.3 mmHg, P = NS) (Table 1). Liver function
as assessed by ICG-PDR decreased in Group 1 by day 1 (18.5 to 15.3,
P <0.05) and remained stable in Group 2 (25.5 to 26.8, P = NS). Renin,
aldosterone, ADH, adrenaline, noradrenaline and dopamine increased
signifi cantly (P <0.05) in Group 1 during operation. Group 2 showed
a signifi cant rise only in ADH and dopamine. Acute kidney injury
occurred in fi ve of 13 patients in Group 1 (P <0.05). Methods Thirty septic patients were prospectively included in the
study. The LiMAx test was performed on days 0, 2, 5 and 10 after sepsis
onset and was compared with ICG test and liver-specifi c laboratory
parameters. Primary endpoint was the mortality rate after 90 days. Secondary endpoint was the comparison with the ICG test. Results The LiMAx test showed low results initially with increasing
values on days 5 and 10. The 90-day mortality rate in patients with a
low LiMAx test on day 2 (<100 μg/kg/hour) was signifi cantly higher
than in patients with a LiMAx test >100 μg/kg/hour. The LiMAx test
was comparable with the ICG test. Patients with indication for renal
replacement therapy during hospital treatment showed signifi cantly
lower LiMAx values than patients without dialysis. See Figure 1. Table 1 (abstract P404)
Group 1
Group 2
Renin
36.4
7.9
PostOP
189.7*
30.5
Day 1
258
31.3
ADH 3.49
2.59
PostOP
16.11*
10.8*
Day 1
10.8
2.39
Creatine
0.8
0.83
Day 1
1.25*
0.91
*P <0.05 compared with preOP value. p
y
g
Conclusion With the LiMAx test we can detect a liver dysfunction in
septic patients early on days 0 to 2. P403 P403
Early diagnosis of liver failure in septic patients using the
maximal liver function capacity test (LiMAx test): comparison with
conventional methods
M Kaff arnik, M Stockmann, J Lock
Charite Berlin Campus Virchow, Berlin, Germany
Critical Care 2013, 17(Suppl 2):P403 (doi: 10.1186/cc12341) Early diagnosis of liver failure in septic patients using the
maximal liver function capacity test (LiMAx test): comparison with
conventional methods The LiMAx test is equal with the
ICG test and a result of <100 μg/kg/hour on day 2 suggests a low
probability of survival in septic patients. 1. Stockmann et al.: New liver function test for prediction of postoperative
outcome in liver surgery. HPB 2010, 12:139. 2. Lock et al.: Early diagnosis of primary nonfunction and indication for
reoperation after liver transplantation. Liver Transpl 2010, 16:172. 2. Lock et al.: Early diagnosis of primary nonfunction and indication for
reoperation after liver transplantation. Liver Transpl 2010, 16:172. Figure 1 (abstract P403). Kaplan–Meier survival curve of patients with
high and low LiMAx results. Conclusion Depending on resection size liver resection acutely
increases portal venous pressure and induces neurohumoral activation
resulting in compromised renal function and increased risk of
developing AKI. P402f Eff ects of proton pump inhibitor versus H2-receptor antagonist
stress ulcer prophylaxis on ventilator-associated pneumonia:
a pilot study
JF Fogas, KK Kiss, FG Gyura, ZT Tóbiás, ZM Molnár
University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P402 (doi: 10.1186/cc12340) Eff ects of proton pump inhibitor versus H2-receptor antagonist
stress ulcer prophylaxis on ventilator-associated pneumonia:
a pilot study
JF Fogas, KK Kiss, FG Gyura, ZT Tóbiás, ZM Molnár
University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P402 (doi: 10.1186/cc12340) University of Szeged, Hungary
Critical Care 2013, 17(Suppl 2):P402 (doi: 10.1186/cc12340) y
Conclusion The results of this pilot study suggest that there may
be no diff erence in the incidence of VAP and GI bleeding if stress
ulcer prophylaxis is performed by H2RA or PPI. As the latter is more
expensive, its use as fi rst choice in critical care should be questioned. Introduction Ventilator-associated pneumonia (VAP) is the most
frequent ICU-acquired infection among patients receiving mechanical S151 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 However, the completion of the study on the planned 198 patients is
required to come to the fi nal conclusions. References
1. Efrati S, et al.: J Clin Monit Comput 2010, 24:161-168. 2. Torres A, et al.: Intensive Care Med 2009, 35:9-29. However, the completion of the study on the planned 198 patients is
required to come to the fi nal conclusions. References P Biesenbach, T Grünberger, B Payer, A Ferlitsch, W Druml, E Fleischmann,
F Luf, J Mühlbacher
Medical University of Vienna, Austria
Critical Care 2013, 17(Suppl 2):P404 (doi: 10.1186/cc12342) Introduction Liver dysfunction – in correlation to the severity of
functional impairment – but also any increase in portal pressure per
se (hepatorenal refl ex) can induce alterations in renal function and
ultimately result in hepatorenal syndrome (HRS). In this prospective
investigation we determined the impact of liver resection on portal
venous pressure by measuring the hepatic venous pressure gradient
(HVPG), on concentrations of vasoactive peptides and on renal function. Methods Twenty patients (mean age 66.3 years) undergoing elective
liver resection surgery because of malignant tumor were assessed
and grouped according to resection size: (Group 1) hemihepatectomy,
n = 13 versus (Group 2) segmentectomy, n = 7. HPVG was measured
before and after resection by canulation of a hepatic vein under
fl uoroscopic guidance, liver function was assessed by the indocyanine
green plasma disappearance rate (ICG-PDR). However, the completion of the study on the planned 198 patients is
required to come to the fi nal conclusions.
References
1.
Efrati S, et al.: J Clin Monit Comput 2010, 24:161-168.
2.
Torres A, et al.: Intensive Care Med 2009, 35:9-29. Impact of liver resection on portal venous pressure and renal
function p
p
p
function
P Biesenbach, T Grünberger, B Payer, A Ferlitsch, W Druml, E Fleischmann,
F Luf, J Mühlbacher
Medical University of Vienna, Austria
Critical Care 2013, 17(Suppl 2):P404 (doi: 10.1186/cc12342) 1. Efrati S, et al.: J Clin Monit Comput 2010, 24:161-168. 2. Torres A, et al.: Intensive Care Med 2009, 35:9-29. Does just fungal evidence increase the mortality of crit
patients with liver cirrhosis? A retrospective analysis patients with liver cirrhosis? A retrospective analysis T Lahmer, U Mayr, M Messer, B Saugel, C Schultheiss, R Schmid, W Huber
University Hospital Munich, Germany Introduction Patients with liver cirrhosis (LC) are susceptible to a
variety of complications. Especially, infection disorders can reduce their
life expectancy markedly. The aim of this retrospective analysis was
to detect the incidence and prognosis in patients with LC and fungal
evidence (FE). Table 1 (abstract P407)
Variable
AUC
Standard error
95% CI
APACHE II score
0.934
0.023
0.889 to 0.980
SOFA score
0.908
0.026
0.857 to 0.960
Number OF
0.901
0.026
0.849 to 0.952
AUC, areas under the curves. Methods A 3-year retrospective analysis of ICU patients with LC. Patients with LC with/without FE were compared with a matched
control group based on comparable age and APACHE II score. Specimens were collected based on clinical and/or laboratory
(infection signs) reasons and were analyzed by microbiological
standard methods. Results A total of 101 patients with LC were enrolled in this analysis. Basic data for both groups: Child–Pugh Score: 4*A, 18*B, 79*C; reason
for ICU admission: 28*hepatorenal syndrome, 18*SBP, 20*GI bleeding,
19*sepsis, 16*pneumonia; etiology of LC: 83*alcoholic, 8*NASH, 8*viral
(hepatitis B/C), 2*PSC. Fifty-fi ve (54%) patients with LC had FE (mean age:
57.6 ± 14 years, sex M/F: 40/15, APACHE II score 27 ± 2). Compared with
the 46 patients with LC but without FE (mean age: 59.2 ± 12 years, sex
M/F: 35/11, APACHE II score 23 ± 4), these patients have a signifi cantly
higher APACHE II score (27 ± 2 vs. 23 ± 4; P <0.001) and signifi cantly
higher mortality (43/55 (78%) vs. 15/46 (33%); P <0.001). Multivariate
analysis showed that FE is associated with a higher mortality (P <0.001)
but is not associated with age, sex or CHILD score. Seventeen (31%) of
the 55 patients in the control group had FE (screening patients n = 147,
mean age 65.8 ± 12 years, sex M/F: 35/20, APACHE II score 26.5 ± 3,
reason for ICU admission: 19*pneumonia, 16*sepsis, 9*intracerebral
haemorrhage, 7*acute abdomen, 4*lymphatic disorder). The mortality
rate was signifi cantly lower (27/55 (49%) vs. 43/55 (78%); P = 0.003) as
compared with patients with LC and FE. The main localisation of fungal
was in both groups the lung (46 vs. 14), followed by urine (18 vs. 2),
blood culture (6 vs. 1) and ascites (4 vs. 0). P405 P405
Value of the preoperative Model for End-stage Liver Disease
score in predicting postoperative outcome in living donor liver
transplantation recipients
Y Nassar, K Omar, A Battah, A Mwafy
Cairo University, Giza, Egypt
Critical Care 2013, 17(Suppl 2):P405 (doi: 10.1186/cc12343) P405
Value of the preoperative Model for End-stage Liver Disease
score in predicting postoperative outcome in living donor liver
transplantation recipients
Y Nassar, K Omar, A Battah, A Mwafy
Cairo University, Giza, Egypt
Critical Care 2013, 17(Suppl 2):P405 (doi: 10.1186/cc12343) P405
Value of the preoperative Model for End-stage Liver Disease
score in predicting postoperative outcome in living donor liver
transplantation recipients Cairo University, Giza, Egypt Cairo University, Giza, Egypt y
gyp
Critical Care 2013, 17(Suppl 2):P405 (doi: 10.1186/cc12343) Introduction Partial liver grafting predisposes recipients to a unique
set of potential technical and anatomic complications that are
not prevalent in whole deceased donor grafts. We aimed to assess
the preoperative Model for End-stage Liver Disease (MELD) score,
postoperative 30-day clinical and laboratory follow-up, and detection
of the value of MELD score in predicting 30-day postoperative outcome
of LDLT recipients. Figure 1 (abstract P403). Kaplan–Meier survival curve of patients with
high and low LiMAx results. Methods A prospective multicenter registry involving 142 patients who
underwent LDLT from October 2004 to December 2010. Preoperative S152 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 and long lengths of stay [1]. Survival among patients with predicted
SAP at admission has been shown to correlate with the duration of
organ failure (OF) [2]. The systemic determinant of severity in a new
classifi cation of acute pancreatitis (AP) is also based on identifi cation of
patients with transient or persistent OF [3]. MELD score was calculated: (0.378×log (bilirubin) (mg/dl) + 1.120×log
(INR) + 0.957×log (creatinine) (mg/dl) + 0.643)×10 then rounded to
the nearest integer. Postoperative daily recording of all the clinical and
laboratory data was done for a period of 30 days. Results The survival rate recorded in our study was 86.62%. The
most frequent complications were renal complications (86.6%),
pulmonary complications (73.9%), neurological complications (14%),
cardiovascular complications (12.6%), infectious complications (13.3%),
intra-abdominal infections (10.5%) and immunosuppressant toxicity
(7.7%). MELD score proved to be a good predictor of perioperative
outcome, patients with MELD score (>12) was associated with less
favorable outcome, accuracy was (80.3%). P407 0
Predictors of persistent organ failure in ICU patients with severe
acute pancreatitis P408
Increased plasma levels of thrombopoietin in patients with severe
acute pancreatitis 1Città della Salute e della Scienza Hospital, Torino, Italy; 2University of Torino,
Italy y
Critical Care 2013, 17(Suppl 2):P408 (doi: 10.1186/cc12346) Introduction The aim of this study was to evaluate the accuracy of
thrombopoietin (TPO) plasma levels as a biomarker of clinical severity
in patients with acute pancreatitis (AP). TPO is a humoral growth factor
that stimulates megakaryocyte proliferation and diff erentiation [1]. Furthermore, it favors platelet aggregation and polymorphonuclear
leukocyte activation [2]. Elevated plasmatic concentrations of TPO have
been shown in patients with critical diseases, including ACS, burn injury
and sepsis [2]. In particular, clinical severity is the major determinant of
elevated TPO levels in patients with sepsis [3]. AP is a relatively common
disease whose diagnosis and treatment are often diffi cult, especially in
the clinical setting of the emergency department (ED). About 20% of
patients with AP develop a severe form of the disease. In order to early
identify those patients aff ected by severe AP, several biomarkers have
been studied. No data regarding TPO plasma levels in patients with AP
are currently available. y
p
Conclusion ICU patients with LC have a high incidence of FE. The
mortality rate is therefore signifi cantly higher as compared with
patients with LC without FE and also as compared with a control
group without LC. Therefore early antimycotic treatment should be
considered. P405 Methods The aim of the study was to retrospectively determine the
predictors of early persistent OF in ICU patients with SAP. The analysis
involved 152 patients. The median time interval between the onset of
AP and admission was 24 (9; 48) hours. The patients were divided into
two groups: the fi rst group (n = 46) had transient OF and the second
group (n = 106) had persistent OF. The ability of the APACHE II score,
total SOFA score and number of organ/system failure to discriminate
transient from persistent OF was explored with receiver operating
characteristic (ROC) curves.i Conclusion MELD score >12 was associated with less favorable
outcome with an accuracy of 80.3%. Results Hospital mortality was signifi cantly higher in the second
group as compared with the fi rst group (45% vs. 7%, P = 0.000); while
infectious complications were 39% versus 11% (P = 0.001) and median
lengths of ICU stay were 8 (5; 18) days for the second group and 6 (4; 7)
days for the fi rst group (P = 0.001). Optimum cutoff levels (by ROC curve
analysis) were APACHE II score ≥12 (sensitivity 0.790; 1 – specifi city
0.119), total SOFA score ≥4 (sensitivity 0.829; 1 – specifi city 0.190), and
failure ≥2 organs/systems (sensitivity 0.819; 1 – specifi city 0.214). See
Table 1. Does just fungal evidence increase the mortality of crit
patients with liver cirrhosis? A retrospective analysis Predominantly candida
species were detected (especially C. albicans/glabrata), fi ve patients
in the LC group had aspergillus fumigatus. In LC patients, FE could be
detected in many cases in several compartments. Conclusion Persistent OF in ICU patients with SAP is associated with the
highest risk of hospital mortality. Early identifi cation of patients with a
high likelihood of persistent OF (APACHE II score ≥12, total SOFA score
≥4, failure ≥2 organs/systems) is an important goal in determining
optimal management of ICU patients with SAP. p
References References
1. Harrison DA, et al.: Crit Care 2007, 11(Suppl 1):S1. 2. Johnson CD, et al.: Gut 2004, 53:1340-1344. 3
D lli
EP t l A
S
2012 [E
b h
d f
i t] 3. Dellinger EP, et al.: Ann Surg 2012. [Epub ahead of print] P411
Urinary neutrophil gelatinase-associated lipocalin as an early
predictor of acute kidney injury in cardiac surgery patients
T García Rodríguez San Miguel
Hospital Santa Creu i Sant Pau, Barcelona, Spain
Critical Care 2013, 17(Suppl 2):P411 (doi: 10.1186/cc12349) Urinary neutrophil gelatinase-associated lipocalin as an early
predictor of acute kidney injury in cardiac surgery patients
T García Rodríguez San Miguel
Hospital Santa Creu i Sant Pau, Barcelona, Spain
Critical Care 2013, 17(Suppl 2):P411 (doi: 10.1186/cc12349) Introduction To evaluate whether urinary neutrophil gelatinase-
associated lipocalin (uNGAL) detects acute kidney injury (AKI) earlier
than the estimated glomerular fi ltration rate (eGFR) in cardiac surgery
patients. Introduction The objective was to evaluate the eff ect of the renal
dysfunction measured by pRIFLE on clinical outcome in children
admitted to the pediatric intensive care unit (PICU). Introduction The objective was to evaluate the eff ect of the renal
dysfunction measured by pRIFLE on clinical outcome in children
admitted to the pediatric intensive care unit (PICU). p
Methods Two-hundred and seventy-four adult patients undergoing
cardiac surgery were consecutively included from February to
December 2011. Exclusion criteria were absence of diuresis due to
end-stage renal disease or chronic renal failure and a previous cardiac
catheterism with i.v. contrast use the week before surgery. Four serial
blood and urine samples immediately before (PRE) and after (POST)
surgery, and 1 day (1d) and 2 days (2d) after surgery were obtained. uNGAL was measured in an Architect 6200 (Abbott Diagnostics). AKIN
criteria were used to diagnose AKI. The study was approved by the
local ethics committee and all patients gave informed consent. Delta
uNGAL was defi ned as the diff erence between the PRE and the POSTs
concentrations. Methods A retrospective cohort involving all children admitted
between August 2009 and July 2010 to a Brazilian PICU located in a
university-affi liated hospital. Patients were classifi ed according to the
pRIFLE at admission as well the maximum pRIFLE during the PICU
stay. The endpoints were: length of stay (LOS) in the PICU, LOS on
mechanical ventilation, mortality rate quoted by predicted mortality of
the Pediatric Index of Mortality (PIM2). y
Results In total, 375 children were included in the study. A total 169
(45%) of them presented renal dysfunction according to the pRIFLE. Renal injury and renal failure were associated with higher LOS in the
PICU and length of mechanical ventilation. The observed and predicted
mortality by PIM2 did not show diff erences. However, the more severe
renal presentation (Injury and Failure) was associated with higher
mortality rate and was not predicted by PIM2 (8% vs. 11%, P <0.01). Thorough evaluation for the new acute kidney injury criteria by
Kidney Disease Improving Global Outcomes Thorough evaluation for the new acute kidney injury criteria by
Kidney Disease Improving Global Outcomes Thorough evaluation for the new acute kidney injury criteria by
Kidney Disease Improving Global Outcomes y
J Izawa, S Uchino, T Fujii, T Arii, T Fukushima, S Kawano
Jikei University School of Medicine, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P410 (doi: 10.1186/cc12348) J Izawa, S Uchino, T Fujii, T Arii, T Fukushima, S Kawano
Jikei University School of Medicine, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P410 (doi: 10.1186/cc12348) Introduction Two previous classifi cations of acute kidney injury (AKI)
that are known as RIFLE criteria and AKIN criteria have shown that
AKI is associated with increased morbidity and mortality. Diff erences
in predicting ability for prognosis, however, have been reported. In
2012, Kidney Disease Improving Global Outcomes (KDIGO) created the
new AKI criteria, combining RIFLE and AKIN criteria. However, such a
combination might cause inconsistency among each defi nition in the
criteria. We have investigated all of the defi nitions in the new KDIGO
criteria in detail. Methods This is a retrospective historical cohort study including adult
patients admitted to the ICU (Jikei University, Tokyo, Japan) between
January 2010 and October 2011. Patients undergoing chronic dialysis
were excluded. KDIGO criteria were applied to all patients to diagnose
AKI. Hospital mortality of patients with AKI diagnosed by the 10
defi nitions in the criteria was compared. Results We studied 41 patients with AP (17 severe and 24 mild
pancreatitis). No diff erences for gender and age were detected
between patients with mild and severe disease. TPO plasma levels
were signifi cantly higher in patients with severe AP (99.33 ± 23.68 pg/
ml) than in those with mild AP (50.81 ± 6.73 pg/ml) (Figure 1). The
ROC curve led us to calculate a cutoff value of 51.55 pg/ml. This value
identifi ed correctly 12 out of the 17 patients aff ected by severe AP. i
Results A total of 2,399 patients were evaluated. AKI occurred in 26.6%
with standard defi nition of KDIGO; 18.0% with creatinine criteria alone;
15.5% with urine output alone. By multivariable analysis, each AKI
stage was associated with hospital mortality: 5.6%, odds ratio 2.95, for
Stage 1; 10.1%, odds ratio 5.52, for Stage 2; 30.2%, odds ratio 21.30,
for Stage 3. Crude hospital mortality stratifi ed by the 10 defi nitions
showed increasing trends with stage progression. P411
Urinary neutrophil gelatinase-associated lipocalin as an early
predictor of acute kidney injury in cardiac surgery patients
T García Rodríguez San Miguel
Hospital Santa Creu i Sant Pau, Barcelona, Spain
Critical Care 2013, 17(Suppl 2):P411 (doi: 10.1186/cc12349) Results One-hundred and eighty-one patients (66.1%) were men;
mean age was 68.2 ± 12.2 years. Valve replacement was performed
in 123, coronary artery bypass graft (CABG) in 81, valve surgery +
CABG in 48, cardiac transplant in fi ve, aorta aneurism surgery in nine,
and other procedures in eight patients. ICU and hospital stays were
6.7 ± 8.1 and 15.7 ± 13.9 days, respectively. Renal replacement therapy
(RRT) was required in 16 patients (5.8%) within 48 hours of ICU stay and
in 28 patients (10.2%) within 4 weeks. Mortality at 28 days was 2.9%. Eighty-six patients (31.4%) were diagnosed with AKI within 48 hours of
surgery. Area under the ROC curve of POST uNGAL for AKI diagnosis
was 0.72 (0.66 to 0.79) (P <0.0001) at an optimal cutoff value of 180 μg/l, Conclusion Renal dysfunction measured by pRIFLE is a very common
fi nding (45%) among pediatric patients admitted to the PICU. The more
severe renal aff ection is associated with higher mortality, length of MV
and length of PICU stay. Thorough evaluation for the new acute kidney injury criteria by
Kidney Disease Improving Global Outcomes Mortality of the
three defi nitions in Stage 1 was from 4.2% to 8.8%. Stage 2 had two
defi nitions and their mortality was 12.1% and 12.9%. Stage 3 had fi ve
defi nitions and their mortality was from 20.5% to 55.6%.i i
yf
y
Conclusion TPO may be proposed as biomarker of clinical severity
in patients with AP at the time of fi rst evaluation in the ED. Further
studies, involving larger numbers of patients, are needed in order to
validate these preliminary data. References 1. Kaushansky K: J Thromb Haemost 2003, 1:1587-1592. 2. Lupia E, et al.: Mediators Infl amm 2012, in press. 3. Zakynthinos SG, et al.: Crit Care Med 2004, 32:1004-1010. i
Conclusion AKI defi ned by the new KDIGO criteria was associated
with increased hospital mortality. In addition, defi nitions in the KDIGO
criteria seem to be appropriate because of clear relations between
mortality and stage progression. P409 P409
Severe renal dysfunction measured by pRIFLE in children is an
independent factor associated with death and not predicted by
PIM2
J Piva1, P Lago2, G Pedro3, F Cabral3
1UFRGS, Porto Alegre, Brazil; 2Hospital de Clinicas de Porto Alegre, Brazil;
3PUCRS, Porto Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P409 (doi: 10.1186/cc12347) Predictors of persistent organ failure in ICU patients with severe
acute pancreatitis Predictors of persistent organ failure in ICU patients with severe
acute pancreatitis
I Aleksandrova, M Ilynskiy, S Rei, V Kiselev, G Berdnikov, L Marchenkova
Hospital Research Institute for Emergency Medicine N.V. Sklifosovsky, Moscow,
Russia
Critical Care 2013, 17(Suppl 2):P407 (doi: 10.1186/cc12345) I Aleksandrova, M Ilynskiy, S Rei, V Kiselev, G Berdnikov, L Marchenkova
Hospital Research Institute for Emergency Medicine N.V. Sklifosovsky, Moscow,
Russia Critical Care 2013, 17(Suppl 2):P407 (doi: 10.1186/cc12345) Methods We enrolled patients with AP at the moment of the fi rst
clinical evaluation in the ED. AP patients were classifi ed as having mild
or severe forms of AP on the basis of the APACHE II score (≥8). TPO
concentrations were determined by ELISA. Introduction Severe acute pancreatitis (SAP) requiring admission to an
ICU is associated with high mortality (hospital mortality reached 42%) S153 Critical Care 2013, Volume 17 Suppl 2 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Figure 1 (abstract P408). Plasma TPO levels in patients with AP. 2.
Hassinger AB, Becker CL: Predictive power of serum cistatin C. Pediatr Crit
Care Med 2012, 13:435-440. 1.
Picca S, Picardo S: Acute kidney injury in an infant after cardiopulmonary
bypass. Semin Nephrol 2008, 28:470-476. P413 Perioperative sodium bicarbonate to prevent acute kidney injury
after cardiac surgery: a multicenter double-blind randomized
controlled trial
A Haase-Fielitz1, M Haase1, M Plass2, P Murray3, M Bailey4, R Bellomo5,
S Bagshaw6 Perioperative sodium bicarbonate to prevent acute kidney injury
after cardiac surgery: a multicenter double-blind randomized
controlled trial Atrial natriuretic peptide attenuates metabolic acidosis and
infl ammation of the kidney, lung and heart in a rat model of renal
ischemia–reperfusion injury Reference 1. Vollmar AM: The role of atrial natriuretic peptide in the immune system. Peptide 2005, 26:1086-1094. Introduction Renal ischemia–reperfusion injury (IRI) is a common
cause of acute kidney injury and occurs in various clinical conditions
including shock and cardiovascular surgery. Renal IRI releases
proinfl ammatory cytokines within the kidney. Atrial natriuretic peptide
(ANP) has natriuretic, diuretic and anti-infl ammatory eff ects [1] and
plays an important role of regulating blood pressure and volume
homeostasis. The hypothesis was that renal IRI induces infl ammation
not only in the kidney but also in remote organs such as the lung and
heart and ANP attenuates renal injury and infl ammation in the kidney,
lung and heart. References 1. Picca S, Picardo S: Acute kidney injury in an infant after cardiopulmonary
bypass. Semin Nephrol 2008, 28:470-476. 1. Picca S, Picardo S: Acute kidney injury in an infant after cardiopulmonary
bypass. Semin Nephrol 2008, 28:470-476. 2. Hassinger AB, Becker CL: Predictive power of serum cistatin C. Pediatr Crit
Care Med 2012, 13:435-440. S154 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 with 78.7% specifi city, 64% sensitivity and 74.1% accuracy. uNGAL
advanced diagnosis of AKI in 44 patients (51.2%), whereas diagnosis
was achieved at the same time as AKI criteria in 11 patients; AKI criteria
outperformed uNGAL in only 36% of cases. Accordingly, uNGAL was
useful to diagnose postoperative AKI in 63.9% of cases. Median delta
uNGAL was 12.5 (from –1.9 to 71.1) and 154.5 (from 16.6 to 484.5) μg/l
in non-AKI and AKI patients, respectively (P <0.0001) and its area under
the ROC curve for AKI prediction was 0.70 (0.63 to 0.77) (P <0.0001). measured. The kidney, lung and heart were immunostained to examine
the localization of IL-6 and NF-κB and assigned an expression score. The
wet/dry ratio of the lung was also measured. Results Renal IRI induced metabolic acidosis, pulmonary edema, mRNA
expression of IL-6 in the kidney, lung and heart. Renal IRI increased
immunohistochemical localization of IL-6 in the proximal convoluted
tubule of the left kidney and NF-κB in the bronchial epithelial cells of
the lung. ANP attenuated metabolic acidosis, pulmonary edema and
expression of IL-6 mRNA in the kidney, heart, and lung. ANP decreased
immunohistochemical localization of IL-6 in the left kidney and NF-κB
in the lung. p
Conclusion Compared with AKIN criteria, a urinary NGAL concentration
>180 μg/l anticipates AKI diagnosis in more than 50% of cardiac surgery
patients in the fi rst 24 to 48 hours after intervention. Conclusion These fi ndings suggested that infl ammation within the
kidney after renal IRI was extended into the lung and heart. ANP
attenuated metabolic acidosis and infl ammation in the kidney, lung
and heart in a rat model of renal IRI. ANP may attenuate organ cross-
talk between the kidney, lung and heart. P412 P412
Atrial natriuretic peptide attenuates metabolic acidosis and
infl ammation of the kidney, lung and heart in a rat model of renal
ischemia–reperfusion injury
M Khin Hnin Si, C Mitaka, M Tulafu, S Abe, S Ikeda
Tokyo Medical and Dental University Graduate School, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P412 (doi: 10.1186/cc12350) P414 p
Methods We connected the urinary catheter of 39 ICU patients to
a quasi-continuous urine analyzer (Kidney INstant monitorinG®),
allowing measurement of pH (pHU), sodium, chloride, potassium
and ammonium concentrations (Na+U, Cl–U, K+U, NH4+U) every
10 minutes. The study period lasted 3 hours after a single intravenous
bolus of furosemide (time 0). In 13 patients receiving two or more
administrations over a longer period (46 (26 to 49) hours), according to
clinical needs, we reviewed data on fl uid therapy, hemodynamics and
acid–base balance from the beginning to the end of the observation. Introduction Here, we compare the value of acute tubular damage
markers measured early in the course after cardiac surgery for
prediction of postoperative AKI compared with conventional markers
used in clinical routine for risk assessment of acute renal function loss. Methods One hundred adult patients undergoing cardiac surgery in
the control arm of a RCT (NCT00672334) were analyzed. We quantifi ed
the following biomarkers in urine: NGAL, hepcidin, midkine, IL-6, α1-
microglobulin; in plasma: NGAL, hepcidin, haptoglobin, CK, CKMB, CRP,
leukocytes, lactate, urea and creatinine. Introduction Here, we compare the value of acute tubular damage
markers measured early in the course after cardiac surgery for
prediction of postoperative AKI compared with conventional markers
used in clinical routine for risk assessment of acute renal function loss. Results Ten minutes after furosemide administration, Na+U and
Cl–U rose from 65 ± 6 to 140 ± 5 and from 109 ± 6 to 150 ± 5 mEq/l
respectively, while K+U fell from 60 ± 5 to 39 ± 4 mEq/l (P <0.001 for
all electrolytes vs. time 0) with a consequent increase in urinary anion
gap (AGU = Na+U + Cl–U – K+U). Urinary output increased from 10 (5
to 19) to 53 (29 to 71) ml/10 minutes (P <0.05). After the fi rst hour Cl–U
remained higher than Na+U, which progressively decreased, leading
to a reduction in AGU and pHU over time. In parallel, a progressive
increment in NH4+U was observed. In patients receiving more than
one administration we observed an increase in arterial base excess
(1.8 ± 0.8 vs. 5.0 ± 0.6 mmol/l, P <0.001) and plasmatic strong ion
diff erence (SIDpl) (31 (30 to 33) vs. 35 (34 to 36) mEq/l, P = 0.01) during
the study period. These changes were due to a decrease in plasmatic
Cl– concentration (109.0 ± 1.1 vs. P414 P414
Tubular damage biomarkers linked to infl ammation or iron
metabolism predict acute kidney injury
A Haase-Fielitz1, S Westphal1, R Bellomo2, P Devarajan3, M Westerman4,
P Mertens1, M Haase1
1Otto von-Guericke University, Magdeburg, Germany; 2Austin Hospital,
Melbourne, Australia; 3Cincinnati Children’s Hospital, Cincinnati, OH, USA;
4Intrinsic LifeSciences, La Jolla, CA, USA
Critical Care 2013, 17(Suppl 2):P414 (doi: 10.1186/cc12352) balance in response to the administration of furosemide. Methods We connected the urinary catheter of 39 ICU patients to
a quasi-continuous urine analyzer (Kidney INstant monitorinG®),
allowing measurement of pH (pHU), sodium, chloride, potassium
and ammonium concentrations (Na+U, Cl–U, K+U, NH4+U) every
10 minutes. The study period lasted 3 hours after a single intravenous
bolus of furosemide (time 0). In 13 patients receiving two or more
administrations over a longer period (46 (26 to 49) hours), according to
clinical needs, we reviewed data on fl uid therapy, hemodynamics and
acid–base balance from the beginning to the end of the observation. Results Ten minutes after furosemide administration, Na+U and
Cl–U rose from 65 ± 6 to 140 ± 5 and from 109 ± 6 to 150 ± 5 mEq/l
respectively, while K+U fell from 60 ± 5 to 39 ± 4 mEq/l (P <0.001 for
all electrolytes vs. time 0) with a consequent increase in urinary anion
gap (AGU = Na+U + Cl–U – K+U). Urinary output increased from 10 (5
to 19) to 53 (29 to 71) ml/10 minutes (P <0.05). After the fi rst hour Cl–U
remained higher than Na+U, which progressively decreased, leading
to a reduction in AGU and pHU over time. In parallel, a progressive
increment in NH4+U was observed. In patients receiving more than
one administration we observed an increase in arterial base excess
(1.8 ± 0.8 vs. 5.0 ± 0.6 mmol/l, P <0.001) and plasmatic strong ion
diff erence (SIDpl) (31 (30 to 33) vs. 35 (34 to 36) mEq/l, P = 0.01) during
the study period. These changes were due to a decrease in plasmatic
Cl– concentration (109.0 ± 1.1 vs. 106.6 ± 0.9 mEq/l, P = 0.009). Plasmatic
sodium and potassium concentrations did not change. In these
patients, considering the total amount of administered fl uids and urine,
a negative water and chloride balance was observed (–460 ± 403 ml
and –48 ± 48 mEq, respectively). P415 increase in urinary NGAL in patients receiving bicarbonate infusion
was observed compared with control (P = 0.011). The incidence of
postoperative RRT was similar but hospital mortality was increased
in patients treated with bicarbonate compared with chloride (11/174
(6.3%) vs. 3/176 (1.7%), OR 3.89 (1.07 to 14.2), P = 0.031). See Figure 1. Conclusion On this basis of our fi ndings we do not recommend the
use of perioperative infusions of sodium bicarbonate to reduce the
incidence or severity of AKI in this patient group. Renal response and acid–base balance alterations during
furosemide administration Renal response and acid–base balance alterations during
furosemide administration
L Zazzeron, D Ottolina, E Scotti, M Ferrari, M Stanziano, C Rovati, F Zadek,
C Marenghi, L Gattinoni, P Caironi
Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P415 (doi: 10.1186/cc12353) Renal response and acid–base balance alterations during
furosemide administration L Zazzeron, D Ottolina, E Scotti, M Ferrari, M Stanziano, C Rovati, F Zadek,
C Marenghi, L Gattinoni, P Caironi
Fondazione IRCCS Ca’ Granda – Ospedale Maggiore Policlinico, Milan, Italy
Critical Care 2013, 17(Suppl 2):P415 (doi: 10.1186/cc12353) Introduction Furosemide is one of the most employed diuretics in
the ICU for its ability to induce negative water balance. However, one
common side eff ect is metabolic alkalosis [1]. We aimed to describe the
time course of urinary excretion and changes in plasmatic acid–base
balance in response to the administration of furosemide. P414 106.6 ± 0.9 mEq/l, P = 0.009). Plasmatic
sodium and potassium concentrations did not change. In these
patients, considering the total amount of administered fl uids and urine,
a negative water and chloride balance was observed (–460 ± 403 ml
and –48 ± 48 mEq, respectively). Methods One hundred adult patients undergoing cardiac surgery in
the control arm of a RCT (NCT00672334) were analyzed. We quantifi ed
the following biomarkers in urine: NGAL, hepcidin, midkine, IL-6, α1-
microglobulin; in plasma: NGAL, hepcidin, haptoglobin, CK, CKMB, CRP,
leukocytes, lactate, urea and creatinine. y
Results Preoperatively, no biomarker predicted AKI. At ICU arrival, four
urinary tubular damage markers showed good discriminatory ability
(AUC ≥0.80) for subsequent development or absence of AKI (Figure 1). An excellent predictive value was found for uNGAL/uhepcidin ratio
(AUC 0.90, Figure 1). This ratio combines an AKI prediction marker
(NGAL) and a marker of protection from AKI (hepcidin), potentiating
their individual discriminatory values. Contrarily, at ICU admission,
none of the plasma biomarkers was a good early AKI predictor with
AUC-ROC ≥0.80. Conclusion Several urinary markers of acute tubular damage predict
AKI after cardiac surgery and the biologically plausible combination of
NGAL and hepcidin provides excellent AKI prediction. Conclusion Furosemide acts immediately after administration, causing
a rise in urinary output, Na+U and Cl–U concentrations. Loop-diuretic-
induced metabolic alkalosis may be due to an increased urinary
chloride loss and the associated increase in SIDpl. Figure 1 (abstract P414). 1. Wilcox CS: Metabolic and adverse eff ects of diuretics. Semin Nephrol 1999,
19:557-568. Perioperative sodium bicarbonate to prevent acute kidney injury
after cardiac surgery: a multicenter double-blind randomized
controlled trial A Haase-Fielitz1, M Haase1, M Plass2, P Murray3, M Bailey4, R Bellomo5,
S Bagshaw6 g
1Otto von-Guericke University, Magdeburg, Germany; 2German Heart Center,
Berlin, Germany; 3University College, Dublin, Ireland; 4ANZIC, Melbourne,
Australia; 5Austin Hospital, Melbourne, Australia; 6University of Alberta, Canada
Critical Care 2013, 17(Suppl 2):P413 (doi: 10.1186/cc12351) g
Methods Male Sprague–Dawley rats were anesthetized with pento-
barbital. Tracheostomy was performed and rats were ventilated at VT
10 ml/kg with 5 cmH2O PEEP. The right carotid artery was catheterized
for blood sampling and continuous blood pressure measurements. The
right femoral vein was catheterized for infusion of saline or ANP. Rats
were divided into three groups; IRI group (n = 10), left renal pedicle was
clamped for 30 minutes; IRI+ANP group (n = 10), left renal pedicle was
clamped for 30 minutes, ANP (0.2 μg/kg/minute, for 3 hours 25 minutes)
was started 5 minutes after clamp; and Sham group (n = 6), the sham-
operated rats. Hemodynamics, arterial blood gas, and plasma lactate
levels were measured at baseline and at 1 hour, 2 hours and 3 hours after
declamp. The mRNA expression of IL-6 in the kidney, lung, and heart were Introduction Evidence suggests a nephroprotective eff ect of urinary
alkalinization in patients at risk of acute kidney injury (AKI). y
y
Methods In a multicenter, double-blind, RCT we enrolled 350 adult
cardiac surgery patients. At induction of anesthesia, patients received
either 24 hours of intravenous infusion of sodium bicarbonate
(5.1 mmol/kg) or sodium chloride (5.1 mmol/kg). The primary endpoint
was the proportion of patients developing AKI. Results Sodium bicarbonate increased urinary pH (from 6.0 to 7.5,
P <0.001). More patients in the bicarbonate group (83/174 (47.7%))
developed AKI compared with control (64/176 (36.4%), OR = 1.60
(95% CI, 1.04 to 2.45); unadjusted P = 0.032). A greater postoperative Figure 1 (abstract P413). Figure 1 (abstract P413). Figure 1 (abstract P413). S155 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P416 Red cell distribution width is not a predictor of mortality in acute
kidney injury
P Purvis1, J Kinsella1, R Docking2
1University of Glasgow, UK; 2Glasgow Royal Infi rmary, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P416 (doi: 10.1186/cc12354) Value of Tc-99m DMSA renal scan in assessing the prognosis and
outcome of acute renal failure due to sepsis Methods A retrospective observational study of renal function in all
patients admitted to a London teaching hospital ICU for ≥5 days and
surviving to hospital discharge in 2011. We excluded cases with current
or new diagnosis of end-stage renal disease or renal transplant. We
assessed AKI in ICU by KDIGO 1 criteria and hospital discharge eGFR by
the CKD-EPI equation. For comparison we assumed a normal GFR in a
healthy individual as 120 ml/minute/1.73 m2 at age 20 decreasing by
0.8 per year over age 20. Value of Tc-99m DMSA renal scan in assessing the prognosis and
outcome of acute renal failure due to sepsis S Fawzi, K Mashour, A Amin, H Gamal Cairo University, Cairo – Manial, Egypt y
gyp
Critical Care 2013, 17(Suppl 2):P417 (doi: 10.1186/cc12355) Introduction Acute renal failure (ARF) is a common complication in
patients admitted to the ICU. Sepsis is also a well-known risk factor for
the development of ARF. The combination of ARF and severe sepsis was
reported to carry a mortality up to 70% whereas the mortality of ARF
alone is 40 to 45%. The aim of the study is to evaluate the role of renal
perfusion scanning in detecting the prognosis and outcome of patients
with acute renal failure due to sepsis. Results We identifi ed 282 patients, 180 of whom had AKI. Median age
was 50 and 68% were male. Median hospital discharge serum creatinine
was 57 μmol/l (range 16 to 654), median eGFR was signifi cantly higher
than predicted normal GFR for age at 115 versus predicted 95 (P <0.001,
median diff erence 16). In patients who had not had AKI discharge the
eGFR was 119 versus normal predicted 98 (P <0.001, median diff erence
19), suggesting that eGFR could be overestimating true GFR in our
population by at least a factor of 1.23 (Figure 1). Applying this correction
factor to eGFRs of patients who had recovered from AKI resulted in 44%
more diagnoses of CKD (eGFR <60) at hospital discharge (36 vs. 25). p
Methods Forty patients with acute renal failure due to sepsis, aged
between 15 and 74 years, were enrolled in the study. They were
monitored for their ICU prognosis and outcome after doing renal
perfusion scanning. All patients were subjected to routine ICU and
laboratory investigations including APACHE II and SOFA score. Results Thirty patients had normal renal scan and 10 patients had
abnormal renal scan. Red cell distribution width is not a predictor of mortality in acute
kidney injury Red cell distribution width is not a predictor of mortality in acute
kidney injury P Purvis1, J Kinsella1, R Docking2
1University of Glasgow, UK; 2Glasgow Royal Infi rmary, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P416 (doi: 10.1186/cc12354) Introduction Given the signifi cant morbidity and mortality associated
with acute kidney injury (AKI), there is a need to fi nd factors to help
aid decision-making regarding levels of therapeutic support. As
a prognostic biomarker, the red cell distribution width (RDW) has
attracted interest in the setting of critical care when added to existing
scoring systems [1]. By examining RDW in a previously studied AKI
cohort, we aimed to evaluate the utility of this routine blood test. Methods A cohort of 209 mixed critical care patients who received renal
replacement therapy for AKI had their demographic and biochemical
data retrieved from electronic databases. Outcomes were gathered for
ICU and hospital mortality. Incomplete datasets were discarded, leading
to 153 complete sets. RDW data were taken from the fi rst sample after
admission to the ICU, as were all other biochemical values apart from
pre-RRT creatinine and potassium. Overall cohort characteristics were
gathered, and two groups were created: those with a RDW value within
normal range (≤14.5%) and those with a greater than normal value Figure 1 (abstract P414). S156 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 (>14.5%). We then further subgrouped RDW to assess the correlation
between rising levels and ICU mortality. Figure 1 (abstract P418). eGFR at hospital discharge versus expected
normal GFR for age in 280 ICU survivors. Results A total 77.1% of our cohort had a RDW greater than the normal
laboratory range at time of ICU admission. Key baseline characteristics
(age, APACHE II score, length of stay, ICU mortality) did not diff er
signifi cantly between patients with normal and abnormal RDW. When
subgroup analysis was performed, no statistically signifi cant correlation
between rising RDW and ICU mortality was found (Spearman
correlation = 0.426, P = 0.233). Conclusion In this cohort of critically ill patients with AKI, RDW was
not found to be a predictor of mortality. Our results contradict those of
recent studies [1,2]. However, both groups of RDW patients in our study
suff ered a higher ICU mortality than in other studies. Red cell distribution width is not a predictor of mortality in acute
kidney injury To further explain
these fi ndings, we intend to perform multivariate logistic regression
analysis and assess the eff ect of social deprivation on RDW. R f 1. Hunziker et al.: Red cell distribution width improves the simplifi ed acute
physiology score for risk prediction in unselected critically ill patients. Crit
Care 2012, 16:R89. 1. Hunziker et al.: Red cell distribution width improves the simplifi ed acute
physiology score for risk prediction in unselected critically ill patients. Crit
Care 2012, 16:R89. Figure 1 (abstract P418). eGFR at hospital discharge versus expected
normal GFR for age in 280 ICU survivors. 2. Oh et al.: Red blood cell distribution width is an independent predictor of
mortality in acute kidney injury patients treated with continuous renal
replacement therapy. Nephrol Dial Transplant 2012, 27:589-594. 2. Oh et al.: Red blood cell distribution width is an independent predictor of
mortality in acute kidney injury patients treated with continuous renal
replacement therapy. Nephrol Dial Transplant 2012, 27:589-594. to survivors of critical illness who may have reduced muscle mass. We
hypothesized that eGFR may underestimate rates and severity of CKD
in ICU survivors. Varying models of intra-abdominal hypertension and their eff ect on
renal function in a porcine model Varying models of intra-abdominal hypertension and their eff ect on
renal function in a porcine model
R Wise1, R Rodseth1, I De Laet2, L Luis3, L Correa-Martin4, M Garcia4,
G Castellanos5, M Malbrain2
1University of Kwazulu Natal, Durban, South Africa; 2ZNA Stuivenberg,
Antwerp, Belgium; 3Neuron NPh, S.A., Granada, Spain; 4Jesús Usón Minimally
Invasive Surgery Centre, Cacères, Spain; 5‘Virgen de la Arrixaca’ University
Hospital, Murcia, Spain
Critical Care 2013, 17(Suppl 2):P419 (doi: 10.1186/cc12357) ,
1University of Kwazulu Natal, Durban, South Africa; 2ZNA Stuivenberg,
Antwerp, Belgium; 3Neuron NPh, S.A., Granada, Spain; 4Jesús Usón Minimally
Invasive Surgery Centre, Cacères, Spain; 5‘Virgen de la Arrixaca’ University
Hospital, Murcia, Spain
Critical Care 2013, 17(Suppl 2):P419 (doi: 10.1186/cc12357) 1. Ympa YP, et al.: Am J Med 2005, 118:827-832. Introduction Intra-abdominal hypertension (IAH) is an independent
predictor of renal impairment and mortality [1]. Organ dysfunction
caused by the pressure eff ect of IAH is well understood, but how this
is modifi ed in the presence of bowel obstruction is unclear. The aim
of this study was to determine how diff erent IAH models cause renal
dysfunction in a pig model. 1. Ympa YP, et al.: Am J Med 2005, 118:827-832. Value of Tc-99m DMSA renal scan in assessing the prognosis and
outcome of acute renal failure due to sepsis The mortality percentage was higher among
abnormal renal scan cases (three out of 10, 30%) compared with cases
with normal renal scan (seven out of 30, 23.3%) with nonsignifi cant
P value: 0.6. The median length of stay/day in ICU was longer among
nonsurvivors than survivors 15.5 ± 10, 11.5 ± 8, P value: 0.058
(approaching signifi cance). APACHE II score was higher in nonsurvivors
than survivors 23.9± 3.2, 19.6± 4.2, P value: 0.0001. The percentage of
mortality among cases that needed mechanical ventilation was higher
(nine out of 16, 56.3%) compared with mortality cases that did not
need mechanical ventilation (one out of 24, 4.2% with P value: 0.0001). Conclusion ARF may exert an independent adverse eff ect on outcome
in septic and septic shock patients. It is also a risk factor for mortality. Tc-99m DMSA scanning is useful for detecting renal dysfunction and
help to predict the outcome and prognosis. R f Conclusion eGFR may overestimate renal function in survivors of critical
illness confounding identifi cation of CKD in this at-risk population. Prospective studies with measurement of actual GFR are required to
assess the burden of CKD in survivors of critical illness. Use, type and duration of cardiopulmonary bypass are related to
acute kidney injury occurrence and urinary NGAL concentrations
T García Rodríguez San Miguel Use, type and duration of cardiopulmonary bypass are related to
acute kidney injury occurrence and urinary NGAL concentrations
T García Rodríguez San Miguel Use, type and duration of cardiopulmonary bypass are related to
acute kidney injury occurrence and urinary NGAL concentrations
T García Rodríguez San Miguel
Hospital Santa Creu i Sant Pau, Barcelona, Spain
Critical Care 2013, 17(Suppl 2):P422 (doi: 10.1186/cc12360) Methods We included medical ICU patients with sepsis and oliguria (urine
output <1 ml/kg/hour) who were prescribed i.v. furose mide. Patients with
known chronic renal failure, hypovolemia (CVP <10 mmHg) or severe
hypotension (MAP <80 mmHg) were excluded. Resistive index (1 − (end
diastolic velocity / maximum systolic velocity)×100) was measured
in at least three segmental arteries of both kidneys, the average of all
measurements was reported as the result. Repeated assessments were
viewed as independent if separated by more than 24 hours. Furosemide
was given intravenously in the dose of 40 mg after RI measurement. Positive response to furosemide was defi ned as doubling of hourly dieresis
or achieving urine output >1.5 ml/kg/hour after drug administration. Introduction To analyze whether variables related to cardiopulmonary
bypass (CPB) infl uence acute kidney injury (AKI) occurrence and urinary
neutrophil gelatinase-associated lipocalin (uNGAL) in cardiac surgery
patients. Introduction To analyze whether variables related to cardiopulmonary
bypass (CPB) infl uence acute kidney injury (AKI) occurrence and urinary
neutrophil gelatinase-associated lipocalin (uNGAL) in cardiac surgery
patients. Methods A total of 274 adult cardiac surgery patients were consecu-
tively included from February to December 2011. Exclusion criteria
were absence of diuresis due to end-stage renal disease or chronic
renal failure and cardiac catheterism with i.v. contrast in the week
before surgery. CPB, when performed, was used as standard CPB
(SCPB) or MiniCPB. We obtained four serial blood and urine samples,
immediately before (PRE) and after (POST) surgery, and 1 day (1d)
and 2 days (2d) after surgery. uNGAL was measured by Architect 6200
(Abbott Diagnostics). AKIN criteria were used to diagnose AKI. The
study was approved by the local ethics committee and all patients gave
informed consent. g
g
g
Results We included 47 patients with a total of 59 measurements. In
28 cases patients had positive response to furosemide. Median RI in
responders was 0.67 (range 0.55 to 0.78) and in nonresponders 0.79
(range 0.58 to 0.81); P = 0.027. Estimated glomerular fi ltration rate based on hospital discharge
creatinine may signifi cantly overestimate renal function and
underestimate chronic kidney disease in survivors of critical illness
l
d ll
l Methods Twenty-four pigs were divided into three groups; a control
group (n = 5), a pneumoperitoneum (Pn) (n = 10), and an intestinal
occlusion (Oc) model (n = 10). IAP was maintained for 3 hours at
20 mmHg during which time creatinine, urea, urine output, potassium,
and glomerular fi ltration pressure (GFP) were measured. Statistical
analysis was performed using repeated-measures ANOVA.ii Introduction Acute kidney injury (AKI) complicates over 50% of ICU
admissions. Episodes of AKI are a major risk factor for development
or progression of chronic kidney disease (CKD); however, methods of
estimated glomerular fi ltration rate (eGFR) may be poorly calibrated Results Over the fi rst 3 hours there was a statistically signifi cant
diff erence between the control group and both IAH models for S157 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 potassium (P = 0.002), urea (P = 0.045), creatinine (P = 0.012), GFP
(P <0.001), and urine output (P = 0.003). Over the full 5-hour period there
was a statistically signifi cant diff erence in the potassium measurement
between the Pn and the Oc model (P = 0.01). The other parameters did
not show a signifi cant diff erence: urea (P = 0.171), creatinine (P = 0.074),
GFP (P = 0.141), and urine output (P = 0.242).i potassium (P = 0.002), urea (P = 0.045), creatinine (P = 0.012), GFP
(P <0.001), and urine output (P = 0.003). Over the full 5-hour period there
was a statistically signifi cant diff erence in the potassium measurement
between the Pn and the Oc model (P = 0.01). The other parameters did
not show a signifi cant diff erence: urea (P = 0.171), creatinine (P = 0.074),
GFP (P = 0.141), and urine output (P = 0.242).i Methods A prospective single-center cohort study started in May 2012
in an academic cardiac surgery ICU. Blood and urine were collected
preoperatively (baseline), at ICU admission and at several time points
until 48 hours after ICU admission. Patients of age <18 years were
excluded, as well as patients with AKI based on the KDIGO sCr and urine
output criteria, patients with chronic kidney disease stage 5 and/or
with recent renal transplantation (≤3 months before cardiac surgery). AKI was defi ned on KDIGO sCr criteria only. Estimated glomerular fi ltration rate based on hospital discharge
creatinine may signifi cantly overestimate renal function and
underestimate chronic kidney disease in survivors of critical illness
l
d ll
l Baseline sCr was assessed as
the sCr concentration measured before start of cardiac surgery. Urinary
CHI3L1 was normalized to urinary creatinine. For both serum and urine
CHI3L1, we measured changes from baseline. Data are expressed as
median (interquartile range), and number (%). Conclusion As expected the IAH models resulted in signifi cantly worse
renal function after 3 hours. This early renal dysfunction may be as a
result of an early infl ammatory process that has been associated with
the pathophysiology of acute kidney injury. Potassium was signifi cantly
elevated in the Pn group as compared with the Oc group. Early changes
in potassium levels with IAH may be a marker of early renal dysfunction
and the usefulness of other renal biomarkers, such as NGAL, prompts
further investigation. Results Data for 17 patients (nine males, eight females) were generated
for interim analysis. Five patients (29.4%), of which four were females,
developed AKI KDIGO class 1. AKI patients were older compared
with non-AKI patients (82.0 years (77.5 to 83.5) vs. 66.5 (60.3 to 75.8);
P = 0.002). The baseline sCr did not diff er signifi cantly between the
AKI (1.12 mg/dl (0.80 to 1.15)) and non-AKI (0.90 mg/dl (0.86 to 1.01);
P = 0.383) group. Baseline serum CHI3L1 was highly variable in both
AKI and non-AKI patients. Moreover, no trend towards a higher serum
CHI3L1 level in AKI versus non-AKI patients was observed either at
baseline or during the study follow-up. In contrast, the increase in the
urinary CHI3L1 to urinary creatinine ratio from baseline was higher in
the AKI versus non-AKI group at time points 4 hours and 6 hours after
admission to the ICU (2.00 ng/mg Cr (1.38 to 3.03) vs. 0.26 (0.05 to 0.88);
P = 0.002, resp. 2.47 (1.09 to 7.26) vs. 0.23 (–0.18 to 0.74); P = 0.006)
and, more importantly, occurred before the diagnosis of AKI by sCr
(>16 hours after ICU admission). Use, type and duration of cardiopulmonary bypass are related to
acute kidney injury occurrence and urinary NGAL concentrations
T García Rodríguez San Miguel Construction of receiver operating
characteristic curve showed 83% sensitivity and 81% specifi city for the
cutoff RI 0.73. No other measured patient characteristic was found to
be predictive of response to diuretic treatment. Conclusion Our results show that the RI could be used to guide diuretic
treatment in nonhypovolemic, nonhypotensive septic patients. Further
studies are needed to confi rm those preliminary results. Results One hundred and eighty-one patients (66.1%) were men; mean
age was 68.2 ± 12.2 years. ICU and hospital stays were 6.7 ± 8.1 and
15.7 ± 13.9 days, respectively. Twenty-eight-day mortality was 2.9%. Eighty-six patients (31.4%) were diagnosed with AKI within 48 hours
after surgery. In total, 219 patients required CPB (195 SCPB, 24 MiniCPB)
and 55 did not (no-CPB). Seven no-CPB patients (12.7%) developed AKI
and their median uNGAL POST was 330 (42.6 to 489.9) μg/l compared
with 13.6 (6.9 to 38.3) μg/l in the 48 patients without AKI (P <0.0001). Of the 195 patients undergoing SCPB, 76 (38.9%) developed AKI and
119 did not; POST uNGAL was 204 (34.8 to 575.7) μg/l and 44.5 (13.2 to
175.8) μg/l (P <0.0001), respectively. In the 24 patients under MiniCPB,
POST uNGAL was 113 (58.8 to 211.8) μg/l in those (three patients, 12.5%)
developing AKI and 19.1 (9.2 to 41.8) μg/l in those without AKI (P = 0.01). Aortic clamp time (r = 0.31, P <0.0001) and cardiopulmonary bypass
time (r = 0.30, P <0.0001) correlated with POST uNGAL concentrations. Reference 1. Malbrain ML, et al.: Incidence and prognosis of intraabdominal
hypertension in a mixed population of critically ill patients: a multiple-
center epidemiological study. Crit Care Med 2005, 33:315-322. 1. Malbrain ML, et al.: Incidence and prognosis of intraabdominal
hypertension in a mixed population of critically ill patients: a multiple-
center epidemiological study. Crit Care Med 2005, 33:315-322. Renal resistive index can predict response to furosemide in septic
patients with oliguria I Gornik, A Godan, V Gasparovic y
p
g
,
Critical Care 2013, 17(Suppl 2):P420 (doi: 10.1186/cc12358) Introduction Oliguria is common in septic patients and is frequently
therapeutically addressed with loop diuretics; that is, furosemide. Diuretic treatment in shock and hypovolemia is not rational, but can
be tried in oliguric patients with normovolemia or hypervolemia and
without hypotension. In such patients it still does not always increase
dieresis and can also be harmful. The resistive index is a measure of
pulsatile blood fl ow that refl ects the resistance to blood fl ow caused by
the microvascular bed distal to the site of measurement. It can refl ect
functional status of the tissue distal to the point of measurement. We
investigated whether measuring the renal resistive index (RI) could
be helpful in determining which patients will respond to furosemide
treatment. Conclusion This interim analysis of an ongoing pilot study, with 300
ICU patients in three cohorts (general ICU, pediatric and adult cardiac
surgery ICU), shows promising results for urinary CHI3L1 (ng/mg Cr) as
an early diagnostic biomarker for AKI in adult ICU patients that undergo
elective cardiac surgery. P422 P421 P421
Urinary chitinase 3-like 1, a novel biomarker for acute kidney injury
in adult cardiac intensive care patients: a pilot study
J De Loor1, E Hoste2, I Herck2, K Francois2, L De Crop2, C Clauwaert2,
K Demeyere1, E Meyer1
1Faculteit diergeneeskunde, Merelbeke, Belgium; 2UZ Gent, Belgium
Critical Care 2013, 17(Suppl 2):P421 (doi: 10.1186/cc12359) P423
Symmetrical dimethylarginine is a more sensitive biomarker of
renal dysfunction than creatinine t e apy (
). Stat st cs: OC
UC. S SS 0. Results There were 91 patients (34 female, 57 male), 65 ± 14 years,
APACHE II score 20.6 ± 8.3, SOFA score 9.3 ± 4.2. Etiology: 10 ARDS,
36 sepsis, 22 cirrhosis, six pancreatitis, eight cardiogenic shock, two
CNS-aff ections, seven various. Mortality was best predicted by SOFA
(ROC-AUC: 0.747; P <0.001) and APACHE II (AUC: 0.705; P <0.001). Among admission parameters of renal function only NGAL signifi cantly
predicted mortality (AUC: 0.647; P = 0.023), whereas creatinine
(P = 0.290) and BUN (P = 0.067) were not predictive. ROC-AUCs for
NGAL further increased after 12 hours (0.659), 24 hours (0.691; P = 0.01)
and 48 hours (0.728; P = 0.004). NGAL on admission also predicted
requirement of RRT during the ICU stay (AUC: 0.678; P = 0.015), which
was also predicted by creatinine (AUC 0.688; P = 0.010) and BUN
(AUC: 0.649; P = 0.043). Among baseline TPTD parameters, only the
pulmonary vascular permeability index (PVPI) signifi cantly predicted
mortality (AUC: 0.700; P = 0.007). The EVLWI (AUC 0.628; P = 0.083)
slightly failed signifi cance. Cardiac index, global end-diastolic volume
index and heart rate were not predictive. Furthermore, PVPI was the
only TPTD-derived parameter to predict requirement of RRT (AUC:
0.693; P = 0.009).i Symmetrical dimethylarginine is a more sensitive biomarker of
renal dysfunction than creatinine
JJ Dixon1, K Lane1, RN Dalton2, IA MacPhee1, BJ Philips1
1Acute Kidney Injury Research Group, St George’s Hospital and University of
London, UK; 2King’s College, University of London, UK
Critical Care 2013, 17(Suppl 2):P423 (doi: 10.1186/cc12361) Symmetrical dimethylarginine is a more sensitive biomarker of
renal dysfunction than creatinine y
JJ Dixon1, K Lane1, RN Dalton2, IA MacPhee1, BJ Philips1
1Acute Kidney Injury Research Group, St George’s Hospital and University of
London, UK; 2King’s College, University of London, UK
Critical Care 2013, 17(Suppl 2):P423 (doi: 10.1186/cc12361) y
JJ Dixon1, K Lane1, RN Dalton2, IA MacPhee1, BJ Philips1
1Acute Kidney Injury Research Group, St George’s Hospital and University of
London, UK; 2King’s College, University of London, UK
Critical Care 2013, 17(Suppl 2):P423 (doi: 10.1186/cc12361) Introduction Current defi nitions of acute kidney injury (AKI) are based
upon changes in serum creatinine (SCr) concentration and urine
output: both have limitations in patients with AKI. Diagnosis may be
delayed if using these criteria alone. Assessment of neutrophil gelatinase-associated lipocalin as a
biomarker of acute kidney injury in acute heart failure patients Results Mean GFR measured by single injection was 78.7 ± 28.5 ml/
minute/1.73 m2, and 78.9 ± 28.6 ml/minute/1.73 m2 when measured
by CIVLDI (P = 0.82). Mean SDMA concentration was similar on both
occasions (641 ± 38 vs. 623 ± 22 nmol/l; P = 0.68). Tubular reabsorption
of SDMA was lower in subjects with GFR <60 ml/minute/1.73 m2
(12 ± 8% vs. 18 ± 8% in subjects with GFR >60 ml/minute/1.73 m2;
P = 0.0002), and tubular secretion of SCR was higher in subjects with
GFR <60 ml/minute/1.73 m2 (29% vs. 23%; P <0.0001). The third-order
polynomial equation (r = 0.93) estimated GFR better than quadratic
(r = 0.92) and linear (r = 0.87) equations. Bland–Altman comparison
revealed no bias when the third-order equation was used (precision:
±20 ml/minute/1.73 m2). p
Critical Care 2013, 17(Suppl 2):P425 (doi: 10.1186/cc12363) Introduction Growing evidence hints that bidirectional interaction
between heart failure and kidney disease and renal insuffi ciency
is a strong predictor of mortality as well as causally linked to the
progression of heart failure. Neutrophil gelatinase-associated lipocalin
(NGAL) is an early predictor of acute kidney injury (AKI). We evaluated
the impact of NGAL on morbidity and mortality in patients with acute
heart failure. Methods Seventy-six patients presenting with symptoms consistent
with acute heart failure (median age 72 years, 56% male) were enrolled. Plasma NGAL levels were measured by an ELISA at admission and
compared with the glomerular fi ltration rate (eGFR) and B-natriuretic
peptide (BNP) levels. The primary outcome was AKI development
defi ned by RIFLE criteria (fall in GFR >25% or creatinine rise ≥50%
from baseline, or a fall in urine output <0.5 ml/kg/hour) and secondary
outcomes were duration of hospital stay and in-hospital mortality. Conclusion SDMA appears to be an accurate and precise estimate of
GFR and a more sensitive biomarker of renal dysfunction than SCr. We
predict SDMA will perform better than SCr as a biomarker of AKI. This
forms the basis of a future study. P425 Assessment of neutrophil gelatinase-associated lipocalin as a
biomarker of acute kidney injury in acute heart failure patients
H Michalopoulou, H Michalopoulou, P Stamatis, J Xenogiannis, D Stamatis
Metaxa Hospital, Athens, Greece
Critical Care 2013, 17(Suppl 2):P425 (doi: 10.1186/cc12363) Urinary chitinase 3-like 1, a novel biomarker for acute kidney injury
in adult cardiac intensive care patients: a pilot study
J D L
1 E H
2 I H
k2 K F
2 L D C
2 C Cl
2 Introduction As a proof of concept, the potential added value of chitinase
3-like 1 (CHI3L1) as a more early and specifi c diagnostic parameter for
acute kidney injury (AKI) was investigated in adult ICU patients that
underwent elective cardiac surgery. Introduction As a proof of concept, the potential added value of chitinase
3-like 1 (CHI3L1) as a more early and specifi c diagnostic parameter for
acute kidney injury (AKI) was investigated in adult ICU patients that
underwent elective cardiac surgery. S158 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Incidence of AKI in patients without CPB or with MiniCPB was identical
and signifi cantly lower (P <0.0001) than in SCPB patients. Conclusion The incidence of AKI was higher in patients undergoing
SCPB. Aortic clamp and cardiopulmonary bypass time correlated with
the POST uNGAL concentration. The higher values found in patients
without AKI undergoing SCPB suggest that a subclinical AKI, only
detectable by uNGAL, could exist in this group. with mortality. Regarding a delayed increase in creatinine, other
biomarkers of AKI including neutrophil gelatinase-associated lipocalin
(NGAL) have been suggested. It was the aim of our study to compare
the predictive capabilities of NGAL and transpulmonary thermodilution
(TPTD)-derived parameters within 48 hours after ICU admission. Incidence of AKI in patients without CPB or with MiniCPB was identical
and signifi cantly lower (P <0.0001) than in SCPB patients. i
Conclusion The incidence of AKI was higher in patients undergoing
SCPB. Aortic clamp and cardiopulmonary bypass time correlated with
the POST uNGAL concentration. The higher values found in patients
without AKI undergoing SCPB suggest that a subclinical AKI, only
detectable by uNGAL, could exist in this group. p
Methods Urinary NGAL, serum creatinine and BUN as well as TPTD-
derived parameters were measured 0 hours, 12 hours, 24 hours and
48 hours after ICU admission. Primary endpoint: prediction of ICU
mortality. Secondary endpoint: requirement of renal replacement
therapy (RRT). Statistics: ROC-AUC. IBM SPSS 20. P423
Symmetrical dimethylarginine is a more sensitive biomarker of
renal dysfunction than creatinine We have previously validated a
method of measuring the glomerular fi ltration rate (GFR) intended for
use in patients with AKI (a continuous infusion of very low dose iohexol;
CIVLDI). In this study we compare the performance of symmetrical
dimethylarginine (SDMA) against SCr and CIVILDI. SDMA is the
structural isomer of the endogenous nitric oxide inhibitor asymmetrical
dimethylarginine. SDMA increases in parallel with SCr. Despite this,
no formula exists to estimate GFR from SDMA concentration. Tubular
secretion of SCr may be as high as 40%. SDMA may be a more sensitive
biomarker of renal dysfunction than SCr; however, the tubular
reabsorption of SDMA is unknown. The aims were to compare the
performance of SDMA against SCr and accurately measured GFR, and
to derive a formula to estimate GFR from SDMA concentration. Conclusion Urinary NGAL on admission signifi cantly predicts mortality,
whereas creatinine and BUN were not predictive. The predictive
capabilities of NGAL further increased after 12 hours, 24 hours and
48 hours, with the ROC-AUC of NGAL 48 hours exceeding the AUC of the
APACHE II score. NGAL on admission also predicts requirement of RRT. Among TPTD-derived parameters, PVPI on admission is signifi cantly
associated with mortality and requirement of RRT. Methods Seventeen volunteers had GFR measured twice via measuring
the clearance of a 5 ml i.v. bolus of iohexol, or CIVLDI. Serum and urine
iohexol, Cr and SDMA were measured by high-performance liquid
chromatography/tandem mass spectrometry (HPLC-ms/ms) at 10 time
points. Fractional excretion of SDMA and SCr were calculated using
iohexol as the denominator. SDMA was plotted against measured GFR,
and estimated GFR equations were derived from linear, quadratic and
third-order polynomial plots. P424 Prognostic value of neutrophil gelatinase-associated lipocalin
and transpulmonary thermodilution-derived parameters within
48 hours after admission
W Huber, J Rauch, B Saugel, S Mair, M Messer, T Lahmer, C Schultheiss,
P Luppa, RM Schmid
Klinikum Rechts der Isar, Technical University of Munich, II. Medizinische Klinik,
Munich, Germany
Critical Care 2013, 17(Suppl 2):P424 (doi: 10.1186/cc12362) Prognostic value of neutrophil gelatinase-associated lipocalin
and transpulmonary thermodilution-derived parameters within
48 hours after admission
W Huber, J Rauch, B Saugel, S Mair, M Messer, T Lahmer, C Schultheiss,
P Luppa, RM Schmid
Klinikum Rechts der Isar, Technical University of Munich, II. Medizinische Klinik,
Munich, Germany
Critical Care 2013, 17(Suppl 2):P424 (doi: 10.1186/cc12362) Results AKI developed in 11 patients (14.4%). The subgroup that during
hospitalization developed renal dysfunction had increased NGAL
values with respect to patients with preserved renal function (146 ± 42
vs. 84 ± 24 ng/ml, P <0.05). NGAL signifi cantly associated with admission
eGFR (r = 0.72, P <0.01) and BNP levels (r = 0.80, P <0.01). For prediction
of AKI, NGAL >140 ng/ml has been related to renal insuffi ciency with
sensitivity of 83% and specifi city of 74% with area under the receiver
operator curve of 0.70 (0.58 to 0.82). In the follow-up setting, NGAL was
associated with recurrent hospital admission for heart failure (HR: 3.1,
CI: 1.5 to 6.2) and cardiac death (HR: 2.2, CI: 1.8 to 4.5). Neutrophil gelatinase-associated lipocalin predicts postoperative
fl uid overload after cardiac surgery Survivors of acute kidney injury requiring renal replacement
therapy rarely receive follow-up: identifi cation of an unmet need
CJ Kirwan, R Taylor, JR Prowle Survivors of acute kidney injury requiring renal replacement
therapy rarely receive follow-up: identifi cation of an unmet need
CJ Kirwan, R Taylor, JR Prowle
The Royal London Hospital, Barts Health NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P427 (doi: 10.1186/cc12365) l
g
y
M Haase1, P Devarajan2, P Michael3, R Bellomo4, A Haase-Fielitz1
1Otto von-Guericke University, Magdeburg, Germany; 2Cincinnati Children’s
Hospital Medical Center, Cincinnati, OH, USA; 3German Heart Center, Berlin,
Germany; 4Austin Hospital, Melbourne, Australia
Critical Care 2013, 17(Suppl 2):P426 (doi: 10.1186/cc12364) Introduction Acute kidney injury (AKI) occurs in more than 50% of
ICU admissions, requiring renal replacement therapy (RRT) in around
10% of cases. There is now increasing evidence that AKI is a risk factor
for the development and progression of chronic kidney disease
(CKD); however, when AKI occurs as a complication of critical illness
appropriate follow-up may be neglected. Accordingly, we reviewed the
follow-up of renal function in all patients who received RRT on our ICU
and survived to hospital discharge. Introduction Neutrophil gelatinase-associated lipocalin (NGAL),
measured early after cardiac surgery, has been demonstrated to predict
postoperative acute kidney injury (AKI). Fluid overload potentially
masks a subsequent acute renal function loss through dilution of
serum creatinine and maintenance of urine output just above AKI-
defi ning criteria. g
Methods A retrospective audit of patients who received RRT in a central
London adult critical care unit during 2011. i
Methods We investigated the early postoperative value of NGAL
versus that of simultaneously measured serum creatinine to
predict subsequent fl uid overload. We studied 100 adult cardiac
surgery patients in the control arm of a RCT (NCT00672334). Severe
postoperative fl uid overload was defi ned as positive fl uid balance
>10% of preoperative body weight within 48 hours after surgery.l Results Of 921 patients admitted, 203 received RRT with 109 surviving
to hospital discharge. We excluded 52 patients who had end-stage
renal disease, renal transplant or known glomerular disease. Of the
remaining 57 AKI patients, median age was 60 (range: 18 to 77) and
37 (65%) were male. Median discharge creatinine was 74.5 μmol/l
(27 to 662). Forty-two (74%) were off ered follow-up, but in only six
cases (11%) was this to nephrology services. Neutrophil gelatinase-associated lipocalin predicts postoperative
fl uid overload after cardiac surgery Twenty-eight attended
follow-up (fi ve to nephrology) at a median time of 6 weeks; however,
creatinine was measured at in only 14 and in six of these it had risen (by
median 16.5 μmol/l). In addition, 14 patients had creatinine measured
3 to 6 months post discharge and in eight it had risen (by median
31.5 μmol/l). y
y
Results Severe postoperative fl uid overload was present in 5% of
patients with a mean positive fl uid balance of 15.8 ± 9.5 l. At ICU
admission, urine NGAL predicted severe fl uid overload (AUC-ROC 0.82
(95% CI = 0.70 to 0.94)) (Figure 1) and mortality (AUC 0.88 (0.78 to 0.97)). Serum creatinine measured at the same time did not predict severe
fl uid overload (AUC 0.52 (0.26 to 0.79)) or mortality (AUC 0.61 (0.16 to
0.99)).l Conclusion Early NGAL-guided adjustments to fl uid management
may reduce organ edema after cardiac surgery. Findings should be
validated in larger cohorts. Conclusion Follow-up of patients who received RRT for AKI in the
ICU was poor and they were rarely referred to nephrologists. Where
renal function was measured after discharge, there was evidence of
progressive renal dysfunction; however, renal function was often not
assessed. We propose an algorithm for clinicians to guide follow-up. See Figure 1. Figure 1 (abstract P426). NGAL at ICU admission predicts fl uid overload
of >10% of body weight. Figure 1 (abstract P426). NGAL at ICU admission predicts fl uid overload
of >10% of body weight. P427 P426
Neutrophil gelatinase-associated lipocalin predicts postoperative
fl uid overload after cardiac surgery
M Haase1, P Devarajan2, P Michael3, R Bellomo4, A Haase-Fielitz1
1Otto von-Guericke University, Magdeburg, Germany; 2Cincinnati Children’s
Hospital Medical Center, Cincinnati, OH, USA; 3German Heart Center, Berlin,
Germany; 4Austin Hospital, Melbourne, Australia
Critical Care 2013, 17(Suppl 2):P426 (doi: 10.1186/cc12364) Prognostic value of neutrophil gelatinase-associated lipocalin
and transpulmonary thermodilution-derived parameters within
48 hours after admission W Huber, J Rauch, B Saugel, S Mair, M Messer, T Lahmer, C Schultheiss,
P Luppa, RM Schmid
Klinikum Rechts der Isar, Technical University of Munich, II. Medizinische Klinik,
Munich, Germany
Critical Care 2013, 17(Suppl 2):P424 (doi: 10.1186/cc12362) Introduction Outcome of ICU patients is predicted by multifactorial
scores such as APACHE II and SOFA. Furthermore, markers of single
organ failure such as the extravascular lung water index (EVLWI) and
several biomarkers of acute kidney injury (AKI) have been associated Conclusion NGAL is emerging as a promising biomarker of AKI in the
setting of acute heart failure and elevated NGAL levels indicate a poor
prognosis in this population regarding morbidity and mortality. S159 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P426 Eff ect of continuous high-volume hemofi ltration on patients with
acute respiratory distress syndrome Eff ect of continuous high-volume hemofi ltration on patients with
acute respiratory distress syndrome
A Kuzovlev1, E Tishkov2, O Bukaev2
1V.A. Negovsky Scientifi c Research Insitute of General Reanimatology RAMS,
Moscow, Russia; 2A.I. Evdokimov Moscow State University of Medicine and
Dentistry, Moscow, Russia
Critical Care 2013, 17(Suppl 2):P431 (doi: 10.1186/cc12369) Eff ect of continuous high-volume hemofi ltration on patients with
acute respiratory distress syndrome
A Kuzovlev1, E Tishkov2, O Bukaev2
1V.A. Negovsky Scientifi c Research Insitute of General Reanimatology RAMS,
Moscow, Russia; 2A.I. Evdokimov Moscow State University of Medicine and
Dentistry, Moscow, Russia
Critical Care 2013, 17(Suppl 2):P431 (doi: 10.1186/cc12369) A Kuzovlev1, E Tishkov2, O Bukaev2i ,
,
1V.A. Negovsky Scientifi c Research Insitute of General Reanimatology RAMS,
Moscow, Russia; 2A.I. Evdokimov Moscow State University of Medicine and
Dentistry, Moscow, Russia ,
,
1V.A. Negovsky Scientifi c Research Insitute of General Reanimatology RAMS,
Moscow, Russia; 2A.I. Evdokimov Moscow State University of Medicine and
Dentistry, Moscow, Russia Critical Care 2013, 17(Suppl 2):P431 (doi: 10.1186/cc12369) Introduction The aim of this study was to investigate the eff ects of
intermittent venovenous high-volume hemofi ltration (IHVHF) on
extravascular lung water, respiratory function and cytokine removal
in patients with acute respiratory distress syndrome (ARDS) associated
with postoperative multiple organ dysfunction syndrome in severe
abdominal infection patients. Results The average quantity of the substitutive renal therapy
procedures in the CG was 13.4 ± 0.7, in the IG it was 8 ± 0.6 (P <0.05). The recuperation of the renal excretory functions was on 19 ± 1 day in
12 patients of the CG, and on 11 ± 1.3 day in the IG, from the moment
of substitutive renal therapy start (P <0.05). Lethality in the CG was
43% (nine patients), and in the IG it was 29% (six patients, P <0.05). The
duration of the MLV in the CG and IG was 21 ± 1.2 days and 16 ± 1.2,
respectively (P <0.05). In the IG the duration of the ICU was lower by
23%, hospitality duration was lower by 17% (P <0.05).fi Methods Thirty patients with severe abdominal infection and
ARDS were randomized into the IHVHF group (n = 15) and control
group (n = 15). All patients were mechanically ventilated. The IHVHF
technique with a rate of 100 to 120 ml/kg/hour during 6 to 8 hours was
used. Can treatment with the molecular adsorbent recirculation system
be the solution for type-1 hepatorenal syndrome?
L Lavayssiere
University Hospital, Toulouse, France
Critical Care 2013, 17(Suppl 2):P430 (doi: 10.1186/cc12368) Can treatment with the molecular adsorbent recirculation system
be the solution for type-1 hepatorenal syndrome? L Lavayssiere
University Hospital, Toulouse, France
Critical Care 2013, 17(Suppl 2):P430 (doi: 10.1186/cc12368) University Hospital, Toulouse, France Introduction The aim of this retrospective study was to determine
whether the molecular adsorbent recirculation system (MARS) can
improve renal function in HSR1 patients. Methods Thirty-two patients with chronic liver disease and HRS1 were
treated by MARS sessions that were performed every other day. The
endpoint was renal function improvement by 28 days after diagnosis
of HRS1 that was defi ned as a serum creatinine level <133 μmol/l. Partial renal recovery was defi ned as a 10% decrease in baseline serum
creatinine level. y
Conclusion Individuals who develop dialysis-dependent AKI in the
ICU setting in general terms either die or recover. Sepsis is the most
common association with death. The need for mechanical ventilation
and inotropic therapy are both associated with increased incidence of
death. Results The mean number of MARS sessions required by each patient
was 3.5 ± 1.5. The median time between admission and the start
of MARS therapy was 3 (0 to 15) days. Of the total patients, 13 (40%)
had improved renal function. Among these, nine (28%) had complete
renal recovery. Among the patients that survived, only 40% (6/15) had
improved renal function. When MARS was started, 53% of patients had
failed renal failure according to RIFLE criteria. Seven patients received
a liver transplant after diagnosis of HRS. Of these, four had complete
or partial recovery after transplantation (57%) versus nine of the 25
patients who did not undergo liver transplantation (36%), P = NS. The
28-day survival rate was 47%. P429fi P429
Effi ciency of substitutive renal therapy in the complex intensive
care of multiple organ failure in patients with polytrauma
S Kravtsov, A Shatalin, V Agadzhanyan, D Skopintsev
Federal State Budgetary Medical Prophylactic Institution ‘Scientifi c Clinical
Center of the Miners’ Health Protection’, Leninsk-Kuznetsky, Russia
Critical Care 2013, 17(Suppl 2):P429 (doi: 10.1186/cc12367) Introduction This study was to evaluate the effi ciency of the early start
of intermittent substitutive renal therapy in patients with polytrauma
complicated by multiple organ failure syndrome. 1. Nadim MK, Genyk YS, Tokin C, et al.: Impact of etiology of acute kidney
injury on outcomes following liver transplantation: acute tubular necrosis
versus hepatorenal syndrome. Liver Transpl 2012, 18:539-548. 2. Cholongitas E, Senzolo M, Patch D, Shaw S, O’Beirne J, Burroughs AK:
Cirrhotics admitted to intensive care unit: the impact of acute renal failure
on mortality. Eur J Gastroenterol Hepatol 2009, 21:744-750. 1. Nadim MK, Genyk YS, Tokin C, et al.: Impact of etiology of acute kidney
injury on outcomes following liver transplantation: acute tubular necrosis
versus hepatorenal syndrome. Liver Transpl 2012, 18:539-548. p
y
p
g
y
Methods Forty-two patients with polytrauma complicated by
multiple organ failure syndrome were included in the study. The age
of the patients was from 20 to 60 years (38.3 ± 1.6 years average). All
patients were divided into two equal groups. In the control group
(CG) the criteria for the start of the substitutive renal therapy were:
hyperkalemia ≥6 mmol/l, plasma creatinine ≥280 μmol/l, diuresis
≤20 ml/hour. In the investigation group (IG) there were subtests to
carry out the substitutive renal therapy, allowing one to start it in
the earlier period of the multiple organ failure progression. These are
increase of Na+ >150 mmol/l, osmolarity >300 mOsm/l, elevation of
the plasma toxicity according to the average molecule concentration
≥1.0, diuresis decrease ≤40 ml/hour. These were examined: lethality,
quantity of the substitutive renal therapy procedures, mechanical lung
ventilation duration (MLV), intensive care and hospital duration. The
substitutive renal therapy was carried out by AK-200-Ultra apparatus
(Gambro, Sweden). The statistical analysis was realized using Statistica
6.1 and the Mann–Whitney U test. 2. Cholongitas E, Senzolo M, Patch D, Shaw S, O’Beirne J, Burroughs AK:
Cirrhotics admitted to intensive care unit: the impact of acute renal failure
on mortality. Eur J Gastroenterol Hepatol 2009, 21:744-750. 2. References 1. Metnitz PG, et al.: Eff ect of acute renal failure requiring renal replacement
therapy on outcome in critically ill patients. Crit Care Med 2002,
30:2051-2058. 2. Uchino S, et al.: Acute renal failure in critically ill patients: a multinational,
multicenter study. JAMA 2005, 294:813-818. 2. Uchino S, et al.: Acute renal failure in critically ill patients: a multinational,
multicenter study. JAMA 2005, 294:813-818. y
Conclusion MARS therapy improved renal function in only very few
patients with HRS1. Even if liver transplantation is the best option, MARS
could be used as a bridge until transplantation. Some authors argue
that prolonged duration of dialysis (>1 month) before transplantation
may result in chronic renal failure probably because there are two
etiologies of AKI in end-stage liver disease: acute tubular necrosis (ATN)
and real functional renal failure. ATN before liver transplantation has
been identifi ed as a risk factor for patients’ mortality and chronic renal
failure at 1 and 5 years post liver transplantation. Further prospective
controlled studies including large number of patients are required. References P429fi Cholongitas E, Senzolo M, Patch D, Shaw S, O’Beirne J, Burroughs AK:
Cirrhotics admitted to intensive care unit: the impact of acute renal failure
on mortality. Eur J Gastroenterol Hepatol 2009, 21:744-750. Aetiology and outcomes for dialysis-dependent acute kidney injury
patients on the ICU M Hameed1, P Carmichael2
1Salford Royal NHS Foundation Trust, Salford, UK; 2The Royal Wolverhampton
Hopsitals NHS Trust, Wolverhampton, UK
Critical Care 2013, 17(Suppl 2):P428 (doi: 10.1186/cc12366) Introduction AKI is a common occurrence in sick hospitalized patients,
in particular those admitted to intensive care. Published data suggest
that 4 to 5% of all critically ill patients develop severe AKI and require
initiation of renal replacement therapy (RRT) [1,2]. Such patients have
high mortality rates often exceeding 60% [2]. We aimed to review
the outcomes of patients admitted to the ICU and required renal
replacement therapy for AKI. We examined whether aetiology of AKI,
comorbidity burden, hospital length of stay and treatment in ICU had
any signifi cant association with survival in the study cohort.i i
Methods During 2009, 56 patients were identifi ed to have received RRT
with AKI who were admitted to the ICU at the Royal Wolverhampton
Hospitals NHS Trust. Computerised and paper-based case records were
examined for these patients to collect the data. AKIN classifi cation was
used to classify the severity of AKI. Figure 1 (abstract P426). NGAL at ICU admission predicts fl uid overload
of >10% of body weight. Figure 1 (abstract P426). NGAL at ICU admission predicts fl uid overload
of >10% of body weight. Figure 1 (abstract P427). A suggested follow-up pathway for AKI survivors. S160 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Results Median age at admission was 66 years (27 to 85) with 29 males
and 27 females. Thirty-one (55.4%) patients had sepsis and 20 (35.7%)
patients had ATN as the main cause of AKI. Thirty-two patients (57%)
had three organ failures at the time of commencement of RRT. Forty-
six patients (82.1%) received haemofi ltration only. Thirty-two (57%)
patients died, with more than 80% of these occurring in the ITU. There
was no signifi cant diff erence in survival when compared with duration
of haemofi ltration, length of stay, number of organs failed and number
of comorbidities. However, signifi cantly more patients that died had
AKI due to sepsis (P = 0.003) or if they received mechanical ventilation
(P = 0.48) or inotropes (0.04). Of the 27 patients who survived until
discharge from hospital, 18 (66.7%) had normal renal function, eight
(29.6%) had AKIN stage I and only one patient required maintenance
haemodialysis. Dialyzer clearance of myoglobin with middle molecule fi lter and
low blood fl ow CVVHD in patients with rhabdomyolysis-associated
acute kidney injury y
j
y
I Leppanen1, T Ahonen1, J Tenhunen2
1Tampere University Hospital, Tampere, Finland; 2Uppsala University Hospital,
Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P432 (doi: 10.1186/cc12370) y
j
y
I Leppanen1, T Ahonen1, J Tenhunen2
1Tampere University Hospital, Tampere, Finland; 2Uppsala University Hospital,
Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P432 (doi: 10.1186/cc12370) Figure 1 (abstract P433). Daily fl uid balance and 95% CIs among
survivors and nonsurvivors. Introduction Rhabdomyolysis-associated acute kidney injury can be
treated with various modes of renal replacement therapy (RRT) [1,2]. We have used continuous venoveno hemodialysis (CVVHD) with
middle molecule fi lter and low extracorporeal blood fl ow (100 to
180 ml/minute) to avoid albumin loss and hemodynamic instability.f y
y
Methods We treated nine patients suff ering from rhabdomyolysis and
acute kidney injury with various causes. We used a CVVHD mode with
low (100 to 180 ml/minute) blood fl ow and a middle molecule fi lter
(Ultrafl ux EMiC2; 1.8 m2 surface area, polysulfone membrane, cutoff at
40 kDa, blood fi lling volume 130 ml, blood fl ow range 100 to 350 ml/
minute, maximum dialysate fl ow 1,000 ml/minute). Myoglobin and
albumin concentrations were measured from prefi lter and postfi lter
samples and dialyzer clearances were calculated (CLdial = blood
fl ow×Cpre – Cpost/Cpre + UF×Cpre/Cpost). Measurements were taken
at 0, 15 minutes, 30 minutes, 4 hours, 12 hours, 24 hours, 36 hours and
48 hours from the start of the CRRT or until CRRT was no longer needed. Results The baseline and prefi lter plasma albumin concentrations varied
from 8 to 29 g/l. Postfi lter and post-treatment albumin concentrations
remained comparable. Prefi lter concentrations of myoglobin (17.8 kDa)
varied from 96,109 to 747 μg/l and the dialyzer clearance of myoglobin
from 50.6 to 0 ml/minute. The mean dialyzer clearance was 23.1 ml/
minute and the median clearance 22.2 ml/minute. Maximal changes
between corresponding prefi lter and postfi lter samples were: absolute
concentration change 27,091 μg/l, percentage 34%, dialyzer clearance
50.6 ml/minute. Clearances were achieved with low extracorporeal
blood fl ow between 100 and 180 ml/minute, most commonly 120 ml/
minute. Ultrafi ltration (UF) was used only at fi ve of the 56 time points
calculated, because of hemodynamic instability. All patients required
either vasopressor or vasopressor and inotrope support. P432 P432
Dialyzer clearance of myoglobin with middle molecule fi lter and
low blood fl ow CVVHD in patients with rhabdomyolysis-associated
acute kidney injury
I Leppanen1, T Ahonen1, J Tenhunen2
1Tampere University Hospital, Tampere, Finland; 2Uppsala University Hospital,
Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P432 (doi: 10.1186/cc12370) Dialyzer clearance of myoglobin with middle molecule fi lter and
low blood fl ow CVVHD in patients with rhabdomyolysis-associated
acute kidney injury The highest
dialyzer clearance of myoglobin (50.6 ml/minute) was measured at the
prefi lter myoglobin of 29,266 μg/l, blood fl ow 180 ml/ml and no UF. Figure 1 (abstract P433). Daily fl uid balance and 95% CIs among
survivors and nonsurvivors. Conclusion Among critically ill patients who are receiving CRRT, daily
fl uid balance is an independent predictor of mortality, even after
adjusting for the daily severity of illness and comorbidities. References Conclusion Among critically ill patients who are receiving CRRT, daily
fl uid balance is an independent predictor of mortality, even after
adjusting for the daily severity of illness and comorbidities. References 1. Bagshaw SM, et al.: Crit Care 2008, 12:169. 2. Bellomo R, et al.: N Engl J Med 2009, 361:1627. 3. Bellomo R, et al.: Crit Care Med 2012, 40:1753-1760. Eff ect of continuous high-volume hemofi ltration on patients with
acute respiratory distress syndrome The PiCCOplus system was used to monitor cardiac output (CO),
extravascular lung water index (EVLWI) and intrathoracic blood volume
index (ITBVI). Arterial partial pressure of oxygen (PaO2), arterial partial
pressure of carbon dioxide (PaCO2), oxygenation index (PaO2/FiO2)
and dynamic lung compliance (Cdyn), oxygen delivery and oxygen
consumption were measured. Serum concentrations of TNFα, IL-6, IL-8 Conclusion The effi ciency of the substitutive renal therapy depends
directly on the hydroelectrolytic and metabolic changes and toxicosis
degrees in the polytrauma complicated by multiple organ failure
syndrome. The early start of the dialysis methods treatment allows
one to achieve the earlier recuperation of the renal functions and to
decrease the lethality level by 14%. S161 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P433l and lactate were measured by ELISA method. The APACHE II scores
before and after IHVHF therapy were detected.i Daily fl uid balance predicts hospital mortality in critically ill patients
receiving continuous renal replacement therapy
M Kashiouris, A Akhoundi, S Chaudhary, V Velagapudi, A Goldberg,
K Kashani
Mayo Clinic, Rochester, MN, USA
Critical Care 2013, 17(Suppl 2):P433 (doi: 10.1186/cc12371) Results All indexes in the control group did not show any signifi cant
improvement before and after treatment (P <0.05). There was a
signifi cant decrease of serum concentrations of TNFα, IL-6, IL-8 and
lactate, increase of CO and decrease of EVLWI 24 hours after IHVHF
(P <0.05). The indexes of oxygen delivery, consumption, PaO2, and Cdyn
improved signifi cantly, PaO2/FiO2 increased markedly compared with
those before IHVHF and the control group. Signifi cant diff erences were
detected in all of the hemodynamics and oxygen indexes (P <0.05). Introduction It has been suggested that fl uid balance is a biomarker
in critically ill patients [1]. There is a paucity of randomized trials
examining the eff ect of daily fl uid balance on outcomes in patients
on continuous renal replacement therapy (CRRT). The RENAL trial did
not fi nd mortality diff erence with higher CRRT dose [2], but did not
investigate the eff ect of daily fl uid balance on patient outcomes. A post
hoc analysis suggested survival benefi t in patients with negative fl uid
balance [3]. In this study, we hypothesize that daily fl uid balance is an
independent predictor of mortality in critically ill patients. y
yg
Conclusion IHVHF presents with a signifi cant benefi cial eff ect on
respiratory function in patients with ARDS as a result of removal
of cytokines, decrease in EVLWI, improvement of hemodynamic
parameters, and correction of disturbances in oxygen balance. IHVHF
adjuvant treatment for ARDS reduces pulmonary edema, improves
PaO2/FiO2 and Cdyn and mechanical ventilation parameters. IHVHF
may be a promising treatment for ARDS in postoperative multiple
organ dysfunction syndrome in severe abdominal infection patients. Reference Methods We conducted a retrospective cohort study in eight ICUs of
a tertiary academic center. We constructed a robust clustered linear
regression model of daily fl uid balance and all-cause hospital mortality
among 595 critically ill patients receiving CRRT. We adjusted the model
for the Charlson comorbidity score, the daily SOFA scores in the fi rst
week after initiation of CRRT as well the type of ICU. 1. 1.
Honoré P, et al.: Anestesiol Reanimatol 2008, 6:4-11. 2.
Sorrentino SA, et al.: Crit Care Med 2011, 39:184-186. 1.
Albert C, et al.: Crit Care Resusc 2012, 14:159-162. P433l Honoré P, et al.: Anestesiol Reanimatol 2008, 6:4-11. Results After adjusting for the type of ICU and the daily severity
of illness, patients who died had on average 779 ml higher daily
fl uid balance compared with patients who survived (P <0.001, 95%
CI = 385 to 1,173 ml, Figure 1). Severity of illness predicted daily fl uid
accumulation; each additional point of the SOFA score predicted an
additional 57 ml of extra daily fl uid (P = 0.002). P432 Fluid balance and renal outcomes in patients requiring renal
replacement therapy in the ICU prefi lter myoglobin of 29,266 μg/l, blood fl ow 180 ml/ml and no UF. Conclusion In a rhabdomyolysis-associated kidney injury a middle
molecule fi lter and a low blood fl ow CVVHD off er a safe and eff ective
treatment choice for patients requiring vasopressor or vasopressor and
inotrope for hemodynamic support. Introduction Fluid overload is associated with mortality in critically
ill patients with acute kidney injury (AKI) [1-3]. We explored this
relationship in patients with AKI who received renal replacement
therapy (RRT) in the ICU to investigate the relationship between fl uid 1.
Albert C, et al.: Crit Care Resusc 2012, 14:159-162.
2
Sorrentino SA et al : Crit Care Med 2011 39:184-186 Pharmacokinetics of gentamicin and vancomycin during
continuous venovenous hemofi ltration in critically ill septic patients
with acute kidney injury p
Results A total of 522 patients were included (319 male, mean age
64 years); 264 (50.6%) died in hospital. Survivors and those who died
were similar with respect to age, weight and incidence of heart failure,
liver cirrhosis and primary renal diagnoses. Independent risk factors for
hospital mortality were higher Sequential Organ Failure Assessment
score at RRT initiation (odds ratio (OR) = 1.18, 95% confi dence
interval = 1.11 to 1.25), higher Charlson comorbidity index (OR = 1.22,
1.09 to 1.37), lower baseline creatinine (OR = 0.984 per 10 μmol/l,
0.970 to 0.998), lower minimum mean arterial pressure (MAP) on day 1
(OR = 0.81 per 10 mmHg, 0.66 to 0.95) and more positive fl uid balance
for the fi rst 7 days (OR 1.03 per 100 ml, 1.02 to 1.05). Of 172 hospital
survivors, 57 (33.1%) were RRT dependent at discharge. Although
univariable analysis suggested lower SOFA scores and higher baseline
serum creatinine levels in those who remained RRT dependent,
no factor was independently associated with RRT dependence at
discharge in a multivariate model. y
j
y
N Petejova1, J Duricova1, A Martinek1, J Zahalkova2
1University Hospital, Ostrava, Czech Republic; 2Hospital, Sternberk, Czech
Republic Introduction Acute kidney injury (AKI) is a common complication of
critical illness and sepsis [1]. Dosing of antibacterial agents in septic
patients is complicated by altered pharmacokinetics due to both
acute renal failure and critical illness [2]. Current dosing regimens for
administration of gentamicin and vancomycin to septic patients with
AKI on continuous venovenous hemofi ltration (CVVH) at a fi ltration rate
of 45 ml/kg/hour are missing. Methods Seventeen septic patients with AKI treated with vancomycin
and seven patients with gentamicin on CVVH were included. In the
vancomycin group, patients received the fi rst dose of 1.0 g intravenously
followed by 1.0 g/12 hours if not adjusted. In the gentamicin group,
patients received a loading dose of 240 mg followed by a maintenance
dose every 24 hours. The vancomycin maintenance dose was optimized
to achieve AUC0–24/MIC ≥400 (Cmin >10 mg/l), gentamicin target was
Cmax/MIC of 8 to 10. Maintenance doses were adjusted according to
drug level simulation using a pharmacokinetic programme. g
Conclusion In this cohort of patients with AKI requiring RRT, a more
positive fl uid balance over 1 week and lower initial minimum MAP
were associated with mortality. References 1. Uchino S, et al.: JAMA 2005, 294:813-818. 2. Roberts JA, et al.: Crit Care Med 2009, 37:840-851. Methods Esomeprazole 40 mg was administered intravenously
once daily in nine intensive care patients and ranitidine 50 mg was
administered intravenously three times daily in nine intensive care
patients with acute renal failure undergoing CVVHDF who required
stress ulcer prophylaxis. The concentration of esomeprazole and
ranitidine in serum and ultradiafi ltrate was determined by high-
performance liquid chromatography. The intragastric pH was obtained
via intragastric pH-metry.i Pharmacokinetics and pharmacodynamics of esomeprazole and
ranitidine during continuous venovenous hemodiafi ltration pi
Conclusion CVVH at a fi ltration rate of 45 ml/kg/hour leads to high
removal of both antibiotics. Due to rapid change in patient’s clinical
status it was impossible to predict a fi xed dosage regimen. We
recommend administration of unreduced loading dose and: blood
sampling as early as 6 hours after fi rst vancomycin dose; blood
sampling 30 to 60 minutes after gentamicin administration and before
the next dose; and the maintenance dose should be based on drug-
level monitoring. Introduction Esomeprazole, a proton pump inhibitor, and ranitidine, a
histamine-2 receptor antagonist are frequently used in intensive care
patients requiring stress ulcer prophylaxis. Continuous venovenous
hemodiafi ltration (CVVHDF) is an important extracorporeal renal
replacement therapy in critically ill patients suff ering from multiple
organ failure. This study investigates the pharmacokinetics and
pharmacodynamics of esomeprazole and ranitidine in anuric critically
ill patients undergoing CVVHDF. p
References S162 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 balance and intradialytic hypotension with mortality and recovery of
renal function. balance and intradialytic hypotension with mortality and recovery of
renal function. intragastric pH >5 and median intragastric pH decreased signifi cantly
in the ranitidine group over time (P <0.05) but not in the esomeprazole
group. Methods We conducted a retrospective cohort study among patients
aged ≥16 years who had RRT initiated and continued for ≥2 days in a
level 2 or 3 ICU at two academic centres, and had fl uid balance data
available. Patients with end-stage kidney disease, within 1 year of
a renal transplant or who had RRT initiated to treat a toxic ingestion
were excluded. We used multivariable logistic regression to determine
the relationship between mean daily fl uid balance over the fi rst
7 days following RRT initiation and the outcomes of mortality and RRT
dependence in survivors. g
Conclusion Although pharmacokinetics of esomeprazole and raniti-
dine are only little infl uenced by CVVHDF, we found a signifi cant
decrease of intragastric pH in patients treated with ranitidine. Esome-
prazole may be superior for long-term stress ulcer prophylaxis in
critically ill patients undergoing CVVHDF. P436 P436 P436
Pharmacokinetics of gentamicin and vancomycin during
continuous venovenous hemofi ltration in critically ill septic patients
with acute kidney injury
N Petejova1, J Duricova1, A Martinek1, J Zahalkova2
1University Hospital, Ostrava, Czech Republic; 2Hospital, Sternberk, Czech
Republic
Critical Care 2013, 17(Suppl 2):P436 (doi: 10.1186/cc12374) P435 P435
Pharmacokinetics and pharmacodynamics of esomeprazole and
ranitidine during continuous venovenous hemodiafi ltration
V Fuhrmann1, P Schenk2, W Jaeger3, M Miksits3, R Kitzberger4, U Holzinger1,
C Madl5
1Medical University Vienna, Austria; 2Landesklinikum Thermenregion
Hochegg, Grimmenstein, Austria; 3University Vienna, Austria; 4Kaiser Franz
Josef Krankenhaus, Vienna, Austria; 5Krankenanstalt Rudolfstiftung Wien,
Vienna, Austria
Critical Care 2013, 17(Suppl 2):P435 (doi: 10.1186/cc12373) Pharmacokinetics and pharmacodynamics of esomeprazole and
ranitidine during continuous venovenous hemodiafi ltration
V Fuhrmann1, P Schenk2, W Jaeger3, M Miksits3, R Kitzberger4, U Holzinger1,
C Madl5 Pharmacokinetics of gentamicin and vancomycin during
continuous venovenous hemofi ltration in critically ill septic patients
with acute kidney injury Among survivors, a less positive fl uid
balance was not associated with increased risk of RRT dependence at
discharge, suggesting that conservative fl uid management does not
signifi cantly attenuate renal recovery. gi
y
References References
1. Bouchard J, et al.: Kidney Int 2009, 76:422-427. 2. Grams ME, et al.: Clin J Am Soc Nephrol 2011, 6:966-973. 3. Payen D, et al.: Crit Care 2008, 12:R74. Results The median vancomycin total clearance (Cltot) was 0.89 and
0.55 ml/minute/kg on the fi rst and second day of the study. CRRT
clearance accounted for about 50 to 60% of vancomycin Cltot found
in a population with normal renal function (0.97 ml/minute/kg). Vancomycin serum concentrations after the fi rst dose were below the
required target of 10 mg/l as early as 6 hours in 10 patients. AUC0–24/
MIC ≥400 ratio was achieved in 67% of patients on the fi rst day. The
median gentamicin Cltot was 0.68 and 0.79 ml/minute/kg on the fi rst
and second day of the study. CRRT clearance accounted for about 50%
of gentamicin Cltot found in a population without renal impairment
(0.73 ml/minute/kg). The target Cmax/MIC ratio was achieved in 78% of
patients after the fi rst dose. Muscle wasting in the ICU can be reliably monitored using
ultrasound
H Jørgensen1, B Nedergaard2, T Gilsaa1
1Lillebaelt Hospital, Kolding, Denmark; 2Odense University Hospital,
Svendborg, Denmark
Critical Care 2013, 17(Suppl 2):P439 (doi: 10.1186/cc12377) g
Critical Care 2013, 17(Suppl 2):P439 (doi: 10.1186/cc12377) Introduction The aim of this study was to establish the intraobserver
and interobserver variation of ultrasonographic measurements of the
rectus femoris muscle cross-section area (RF-CSA). Muscle wasting is
frequent in the ICU, aff ecting more than one-half of the patients with
severe sepsis [1]. Muscle mass reduces rapidly, and 15 to 20% is lost
within the fi rst week [1]. To monitor muscle mass, ultrasound has the
benefi ts of being both readily available in the ICU and non-invasive. Ultrasonographic measurement of RF-CSA has an almost perfect
correlation with MRI (mean interclass correlation (ICC) = 0.999) [2] and
RF-CSA is linearly related to maximum voluntary contraction strength
in both healthy subjects and COPD patients (r = 0.78) [3]. Conclusion Adipogenesis and accumulation of M2-macrophages are
hallmarks of critical illness, irrespective of nutritional management
in humans and mice. Critical illness evokes macrophage polarization
to the M2-state not only in adipose tissue but also in liver and lungs,
which is further accentuated by fasting. P440 Critical illness induces nutrient-independent adipogenesis and
accumulation of alternatively activated tissue macrophages
M Marques, S Vander Perre, A Aertgeerts, S Derde, F Guiza, M Casaer,
G Hermans, G Van den Berghe, L Langouche
KU Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P440 (doi: 10.1186/cc12378) P437
ff P437
Eff ects of two diff erent regional citrate anticoagulation CVVH
protocols on acid–base status and phosphate supplementation
S Morabito1, V Pistolesi1, V Pistolesi1, L Tritapepe2, L Tritapepe2, E Vitaliano3,
E Vitaliano3, L Zeppilli1, L Zeppilli1, F Polistena1, F Polistena1, E Fiaccadori4,
E Fiaccadori4, A Pierucci1, A Pierucci1
1Umberto I, Policlinico di Roma, ‘Sapienza’ University, Rome, Italy; 2Umberto
I, Policlinico di Roma, Rome, Italy; 3Pertini Hospital, Rome, Italy; 4University of
Parma Medical School, Parma, Italy
Critical Care 2013, 17(Suppl 2):P437 (doi: 10.1186/cc12375) Results The mean peak prefi lter concentration, AUC0–24, volume of
distribution, half-life, total clearance and hemodiafi ltration clearance of
esomeprazole were 6.9 μmol/l, 30.6 μmol.hour/l, 53.3 l, 2.0 hours, 1.4 l/
hour and 0.2 l/hour, respectively. In the ranitidine group the mean peak
prefi lter concentration, AUC0–8, volume of distribution, half-life, total
clearance and hemodiafi ltration clearance were 5.5 μmol/l, 12.0 μmol. hour/l, 91.4 l, 14.4 hours, 5.8 l and 1.3 l, respectively. Median time of Introduction Regional citrate anticoagulation (RCA) is increasingly
used in high bleeding risk patients undergoing CRRT. Regardless of
anticoagulation protocol, hypophosphatemia occurs frequently in Introduction Regional citrate anticoagulation (RCA) is increasingly
used in high bleeding risk patients undergoing CRRT. Regardless of
anticoagulation protocol, hypophosphatemia occurs frequently in S163 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 distance from the anterior superior iliac spine to the superior patellar
border [1]. RF-CSA was calculated by planimetry. At each scan, three
measurements were made. For intraobserver variation, the 3×3
scans were analyzed using the interclass correlation coeffi cient. For
interobserver variation, the three measurements from each observer
were averaged and compared using Bland–Altman statistics. Results Intraobserver variation: 15 healthy adults, age 39.6 ± 2.4 years,
weight 66.8 ± 2.3 kg, sex three male/12 female. ICC: 0.996 (95% CI: 0.990
to 0.998). Interobserver variation: 15 ICU patients, age: 77 ± 8.3 years,
weight: 71.3 ± 9.1 kg, sex nine male/six female. Bland–Altman: bias:
–0.07 cm2, 95% limits of agreement –0.188 to 0.048 cm2. Conclusion Ultrasonographic measurement of RF-CSA is easily distance from the anterior superior iliac spine to the superior patellar
border [1]. RF-CSA was calculated by planimetry. At each scan, three
measurements were made. For intraobserver variation, the 3×3
scans were analyzed using the interclass correlation coeffi cient. References
1.
Langouche L, et al.: Am J Respir Crit Care Med 2010, 182:507-516.
2.
Casaer MP, et al.: N Engl J Med 2011, 365:506-517. References 1. Parry SM, et al.: BMJ Open 2012, e001891. doi:10.1136/bmjopen
2. Reeves ND, et al.: Eur J Appl Physiol 2004, 91:116-118. 3. Seymour JM, et al.: Thorax 2009, 64:418-423. g
p
Results In 30 cardiac surgery patients with AKI, acid–base status
and electrolytes have been evaluated comparing protocol A (20
patients, running time 5,283 hours) versus protocol B (10 patients,
1,170 hours) (median (IQR)): pH 7.40 (7.36 to 7.44) versus 7.43 (7.41
to 7.47) (P <0.0001), bicarbonate 22.1 (20.9 to 23.5) versus 24.4 (23.2
to 25.6) mmol/l (P <0.0001), base excess –3.1 (–4.6 to –1.15) versus 0
(–1.5 to 1.1) (P <0.0001), systemic Ca2+ 1.16 (1.1 to 1.23) versus 1.14
(1.08 to 1.19) mmol/l (P <0.0001), phosphate 0.7 (0.5 to 1) versus
1.1 (0.9 to 1.4) mmol/l (P <0.0001). Protocol A required bicarbonate
infusion in 90% of patients (6 ± 6.4 mmol/hour) and sodium phosphate
supplementation in all cases (4.1 ± 2.4 g/day). A lower amount of
phosphate supplementation (0.9 ± 2 g/day) was needed in 30% of
patients undergoing protocol B while bicarbonate infusion was never
required. Filter life was comparable (51.8 ± 36.5 vs. 46.8 ± 30.3 hours,
P = NS).f P437
ff For
interobserver variation, the three measurements from each observer
were averaged and compared using Bland–Altman statistics. CRRT. The aim was to evaluate the eff ects on electrolyte and acid–base
status of a new RCA-CVVH protocol using an 18 mmol/l citrate solution
combined with a phosphate-containing replacement fl uid, compared
with a previously adopted RCA-CVVH protocol combining a 12 mmol/l
citrate solution with a conventional replacement fl uid. CRRT. The aim was to evaluate the eff ects on electrolyte and acid–base
status of a new RCA-CVVH protocol using an 18 mmol/l citrate solution
combined with a phosphate-containing replacement fl uid, compared
with a previously adopted RCA-CVVH protocol combining a 12 mmol/l
citrate solution with a conventional replacement fl uid. distance from the anterior superior iliac spine to the superior patellar
border [1]. RF-CSA was calculated by planimetry. At each scan, three
measurements were made. For intraobserver variation, the 3×3
scans were analyzed using the interclass correlation coeffi cient. For
interobserver variation, the three measurements from each observer
were averaged and compared using Bland–Altman statistics. pl
Methods Until September 2011, RCA-CVVH was routinely performed
in our centre with a 12 mmol/l citrate solution and a postdilution
replacement fl uid with bicarbonate (HCO3
– 32, Ca2+ 1.75, Mg2+ 0.5, K+
2 mmol/l) (protocol A). In cases of metabolic acidosis, not related to
inappropriate citrate metabolism and persisting after optimization of
RCA-CVVH parameter setting, bicarbonate infusion was scheduled. Starting from September 2011, in order to optimize buff er balance
and to reduce the need for phosphate supplementation, a new
RCA-CVVH protocol has been designed using an 18 mmol/l citrate
solution combined with a recently introduced phosphate-containing
replacement fl uid with bicarbonate (HCO3
– 30, phosphate 1.2, Ca2+ 1.25,
Mg2+ 0.6, K+ 4 mmol/l) (protocol B). g
g
Results Intraobserver variation: 15 healthy adults, age 39.6 ± 2.4 years,
weight 66.8 ± 2.3 kg, sex three male/12 female. ICC: 0.996 (95% CI: 0.990
to 0.998). Interobserver variation: 15 ICU patients, age: 77 ± 8.3 years,
weight: 71.3 ± 9.1 kg, sex nine male/six female. Bland–Altman: bias:
–0.07 cm2, 95% limits of agreement –0.188 to 0.048 cm2. g
Conclusion Ultrasonographic measurement of RF-CSA is easily
learned and quickly performed. It has a very low intraobserver and
interobserver variation and can be recommended as a reliable method
for monitoring muscle wasting in the ICU. R f Critical illness induces nutrient-independent adipogenesis and
accumulation of alternatively activated tissue macrophages
M Marques, S Vander Perre, A Aertgeerts, S Derde, F Guiza, M Casaer,
G Hermans, G Van den Berghe, L Langouche
KU Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P440 (doi: 10.1186/cc12378) Critical Care 2013, 17(Suppl 2):P440 (doi: 10.1186/cc12378) Introduction In artifi cially fed critically ill patients, adipose tissue
reveals an increased number of small adipocytes and accumulation of
M2-type macrophages [1]. We hypothesized that nutrient-independent
factors of critical illness explain these fi ndings, and also that M2-
macrophage accumulation during critical illness may not be limited to
adipose tissue. Conclusion Protocol B provided a buff er balance more positive than
protocol A and allowed one to adequately control acid–base status
without additional bicarbonate infusion and in the absence of alkalosis,
despite the use of a standard bicarbonate concentration replacement
solution. Furthermore, the combination of a phosphate-containing
replacement fl uid appeared eff ective to prevent hypophosphatemia. p
Methods We performed a randomized investigation in a septic mouse
model of critical illness and a study of ICU patient biopsies. In the
critically ill mouse, we compared the eff ect of parenteral nutrition
(n = 13) with fasting (n = 11) on body composition, adipocyte cell
size, and macrophage accumulation in adipose tissue, liver and lung. Fed healthy control mice (n = 11) were studied for comparison. In
vivo adipose tissue was harvested after 1 week of illness from human
patients (n = 40) who participated in a RCT on early parenteral nutrition
versus tolerating nutrient restriction [2], adipose tissue morphology
was characterized and compared with healthy controls (n = 13). P438
Abstract withdrawn P438
Abstract withdrawn P438 Results Irrespective of nutritional intake, critically ill mice lost body
weight, total fat and fat-free mass. Part of the fat loss was explained
by reduced ectopic fat accumulation. Adipocyte cell number and
the adipogenic markers peroxisome proliferator-activated receptor
γ and CCAT/enhancer binding-protein β increased with illness, again
irrespective of nutritional intake. Macrophage accumulation with
predominant M2-phenotype was observed in adipose tissue, liver and
lungs of critically ill mice, further accentuated by fasting in visceral
tissues. Macrophage M2-markers correlated with chemoattractant
factor expression in all studied tissues. In human subcutaneous
adipose tissue biopsies of critically ill patients, increased adipogenic
markers and M2 macrophage accumulation were present irrespective
of nutritional intake. P439
Muscle wasting in the ICU can be reliably monitored using
ultrasound
H Jørgensen1, B Nedergaard2, T Gilsaa1
1Lillebaelt Hospital, Kolding, Denmark; 2Odense University Hospital,
Svendborg, Denmark
Critical Care 2013, 17(Suppl 2):P439 (doi: 10.1186/cc12377) P444
Strong ion gap can be accurately estimated with a simple bedside
equation g
y
y
Methods Obstetric patients were identifi ed from the ICU admissions
database and divided into two groups – those treated with magnesium
(for suspected pre-eclampsia) and those admitted for other obstetric
indications (postpartum hemorrhage, infection, etc.). The baseline
calcium values were compared, as well as the lowest and discharge
values. Albumin and magnesium values were also compared. All
comparisons used Student’s t test. Introduction The anion gap (AG) is used routinely in the assessment
of metabolic acidosis, but can be misleading in patients with
hypoalbuminemia and other disorders commonly encountered in
intensive care. This approach to acid–base analysis relies on assessment
of pH, pCO2, sodium, bicarbonate and chloride, and can lead to
underestimation or overestimation of the true electrochemical status
of a patient, as it does not include important ions such as lactate,
calcium, magnesium, and albumin. The strong ion gap (SIG) is an
alternative to the AG and is based upon Stewart’s physical chemistry
approach. However, the SIG is cumbersome to calculate. As such, a
number of shortcut equations have been developed in an eff ort to
approximate the SIG. We sought to compare three such equations, the
Kellum corrected anion gap (KellAGc), the Moviat equation, and EZSIG,
in an eff ort to evaluate precision and accuracy [1-3]. Results Data were collected on 88 parturients admitted over 2 years
including 40 (45%) who received magnesium and 48 (55%) who did not. Magnesium-treated women were younger (age: 31 ± 7 vs. 36 ± 5 years,
P = 0.02). The baseline calcium concentrations were similar for the
two groups (2.2 ± 0.2 vs. 2.2 ± 0.1 mmol/l, P = 0.85). Patients receiving
magnesium had signifi cantly higher magnesium concentrations
(2.1 ± 0.4 vs. 0.7 ± 0.2 mmol/l, P <0.001), and signifi cantly lower
calcium concentrations during therapy (1.6 ± 0.3 vs. 1.9 ± 0.3 mmol/l,
P <0.001). At discharge, the calcium levels were closer (magnesium
treated 1.9 ± 0.2 vs. untreated 2.1 ± 0.1 mmol/l, P = 0.02). The albumin
concentrations did not diff er between the two groups (magnesium
treated 27 ± 13 vs. nontreated 33 ± 23 g/l, P = 0.134). Normal values:
calcium 2.15 to 2.55 mmol/l, magnesium 0.7 to 0.95 mmol/l, albumin
35 to 50 g/l. f
y
Methods We conducted a retrospective chart review of consecutive
patients admitted to the ICU of George Washington University Medical
Center from September 2010 to March 2011. References 1. Kellum JA: Crit Care 2005, 9:500-507. 2. Moviat M, et al: Crit Care 2003, 7:R41-R45. 3. Busse L, et al.: Chest J 2011, 140(4_MeetingAbstracts):1012A-1012A. 3. Busse L, et al.: Chest J 2011, 140(4_MeetingAbstracts):1012A-1012A. P444
Strong ion gap can be accurately estimated with a simple bedside
equation Of the 1,516 patients
screened, 200 met inclusion criteria, which included availability of all
laboratory components to calculate the SIG, obtained within 1 hour of
each other. Demographic data and serum values for pH, pCO2, albumin,
lactate, sodium, potassium, chloride, bicarbonate, magnesium,
phosphate, and calcium were collected. The AG, SIG, KellAGc, EZSIG,
and Moviat equations were subsequently calculated and compared
using Pearson correlation and Bland–Altman analysis. Conclusion Magnesium therapy was associated with hypocalcemia. Potential causative mechanisms include a renal excretion interaction
and magnesium-induced suppression of parathyroid hormone
secretion. Physicians should be aware of the potential for symptomatic
hypocalcemia during magnesium therapy. g
y
Results The mean SIG was 3.25 ± 3.5. Mean values for KellAGc, Moviat,
and EZSIG were 4.5 ± 5.0, 1.77 ± 2.2, and 3.6 ± 3.7, respectively. Pearson
correlation coeffi cients for KellAGc, Moviat, and EZSIG when compared
with the SIG were r = 0.77, P = 0.0001; r = 0.88, P = 0.001; and r = 0.89,
P = 0.001, respectively. In Bland–Altman analysis, the mean bias for the
test equations versus the SIG were: KellAGc (1.25), Moviat (–1.48), and
EZSIG (0.40). References Methods The study had two purposes: to determine the intraobserver
variation for RF-CSA by one observer scanning 15 healthy adult
volunteers three times each at 2-day intervals; and to determine the
interobserver variation for RF-CSA by two observers each scanning 15
adult ICU patients on the same day. Patients were in a supine position,
legs in passive extension. The transducer was placed perpendicular
to the long axis of the right thigh over the RF, two-thirds of the 1. Langouche L, et al.: Am J Respir Crit Care Med 2010, 182:507-516. 2. Casaer MP, et al.: N Engl J Med 2011, 365:506-517. P441
Abstract withdrawn P441
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http://ccforum.com/supplements/17/S2 P442 of low serum sodium levels, or if hyponatremia is simply a marker for
‘sicker’ patients, is not known. R f of low serum sodium levels, or if hyponatremia is simply a marker for
‘sicker’ patients, is not known. Reference P442
Hypocalcemia during magnesium therapy for obstetric ICU
admissions
PD Levin, A Szalat, M Vizana, CL Sprung, R Zaguri, Y Haviv
Hadassah Hebrew University Medical Center, Jerusalem, Israel
Critical Care 2013, 17(Suppl 2):P442 (doi: 10.1186/cc12380) 1. Asadollahi et al.: Hyponatraemia as a risk factor for hospital mortality. QJM
2006, 99:877-880. Introduction Intravenous magnesium sulfate is commonly used
in obstetric patients with pre-eclampsia. Following a case of acute
symptomatic hypocalcemia we retrospectively examined a cohort of
patients to investigate the frequency of hypocalcemia. P445 Relative adrenal insuffi ciency in burns
A Mokline, L Gharsallah, I Rahmani, H Oueslati, B Gasri, S Tlaili,
R Hammouda, A Ksontini, A Ghanem, AA Messadi
Burn and Trauma Center, Tunis, Tunisia
Critical Care 2013, 17(Suppl 2):P445 (doi: 10.1186/cc12383) Relative adrenal insuffi ciency in burns Hyponatremia and intensive care outcome Hyponatremia and intensive care outcome
P Dean, R Docking, D Govenden, A Davidson, A Mackay
Victoria Infi rmary, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P443 (doi: 10.1186/cc12381) i
y
g
Critical Care 2013, 17(Suppl 2):P443 (doi: 10.1186/cc12381) Introduction Disorders of sodium (Na+) and water homeostasis are
common in hospitalised patients. Hyponatremia in particular has
been associated with worse hospital outcome and length of stay
[1]. We aimed to defi ne the incidence of hyponatremia (serum Na+
≤134 mmol/l) in our intensive care population and to determine
whether it was associated with ICU outcome or length of stay. Conclusion While all three equations correlated highly with the SIG,
the EZSIG and Moviat outperformed the KellAGc in Pearson and Bland–
Altman analysis. The EZSIG had a smaller bias than the Moviat equation
and a slightly better correlation (0.89 vs. 0.88). In the assessment of
critically ill patients, EZSIG is a candidate scanning equation for the
measurement of the SIG when all SIG components are not available. f Methods Demographics, APACHE II score, outcome data and admission
sodium were retrieved from the Ward Watcher system in the Victoria
Infi rmary ICU for 2,440 consecutive admissions from January 2005 to
present. We divided patients into three groups depending on serum
Na+ (≤134 mmol/l, 135 to 144 mmol/l, ≥145 mmol/l) and compared
APACHE II score, length of stay and ICU outcome between patients
with a low versus a normal serum Na+. Data were analysed using the
chi-squared test, Student’s t test and the Mann–Whitney test where
appropriate. P444 P444
Strong ion gap can be accurately estimated with a simple bedside
equation
L Busse, L Chawla, R Panchamia, D Choi, E Nobakht, E Brasha-Mitchell,
M Seneff
George Washington University Medical Center, Washington, DC, USA
Critical Care 2013, 17(Suppl 2):P444 (doi: 10.1186/cc12382) Etiology of low testosterone levels in mal
sepsis requiring mechanical ventilation
h A Bech, H Van Leeuwen, H De Boer A Bech, H Van Leeuwen, H De Boer Rijnstate Hospital, Arnhem, the Netherlands yp
Results Data from 1,038 patients were included. Admission iCa
was available for 976 (94%) patients. A total of 539 (55%) were
hypocalcaemic (iCa <1.1 mmol/l). adjCa was available for 1,031 (99%),
with 602 (58%) classifi ed as hypocalcaemic (adjCa <2.2 mmol/l). adjCa
<2.2 mmol/l had sensitivity of 78% and specifi city of 63% for detecting
iCa <1.1 mmol/l. The best previously published formula had an AUC of
0.81 (95% CI = 0.78 to 0.83), compared with an AUC of 0.78 (0.75 to 0.81)
for the locally validated formula [1]. It was hypothesized that albumin,
pH or phosphate derangements could aff ect the proportion of albumin-
bound calcium and the performance of adjustment formulae. Albumin
concentration was not correlated with the diff erence between adjCa
and iCa. Weak but statistically signifi cant correlations were found for pH
(r = 0.164; P <0.001) and phosphate (r = 0.135; P <0.001). The sensitivity
and specifi city of adjCa <2.2 mmol/l for detecting hypocalcaemia did
not improve when looking at patients with normal pH (sensitivity
60.4%; specifi city 69%) or phosphate (32.2%; 78.9%). j
p
Critical Care 2013, 17(Suppl 2):P448 (doi: 10.1186/cc12386) Introduction Low testosterone levels are frequently found in critically
ill male patients. The etiology and clinical signifi cance is still poorly
understood. In the present study we have investigated the kinetics
and pathophysiology of altered gonadal hormone synthesis in male
patients with severe sepsis and respiratory failure. Methods All male patients with severe sepsis and respiratory failure
who were admitted to the ICU of a large teaching hospital in the
Netherlands between September 2011 and June 2012 were included. Steroid hormone levels were measured on days 1, 3 and 7. y
Results In total, 18 patients were included. The mean age was
69 ± 2 years, mean weight 76 ± 2 kg, APACHE II score 23 ± 2 and most
patients suff ered from pneumosepsis. On the fi rst day of intubation,
total and free testosterone levels were extremely low in most patients
and remained low during the fi rst week (Figure 1). 17β-Estradiol levels
were elevated on day 1 and decreased during the fi rst week. LH and i
Conclusion adjCa is a relatively poor predictor of iCa in critically
ill patients. Table 1 (abstract P447) Table 1 (abstract P447)
Low
Intermediate
High
Tertile
(n = 216)
(n = 219)
(n = 220)
P value
25(OH)D (ng/ml)
10.4 (± 3.6)
17.6 (± 5.1)
30.7 (± 11.4)
<0.001
SAPS 2
32 (± 15)
28 (± 16)
28 (± 16)
0.007
Hospital mortality
58 (26.9%)
48 (21.2%)
29 (13.2%)
0.002
Lethal sepsis
11 (5.1%)
9 (4.1%)
0 (0.0%)
0.005
Conclusion Low 25(OH)D status is predictive of all-cause and sepsis
mortality in the critically ill. Interventional studies are needed to
investigate the eff ect of vitamin D on mortality and sepsis incidence
and outcomes. Low
Intermediate
High
Tertile
(n = 216)
(n = 219)
(n = 220)
P value
25(OH)D (ng/ml)
10.4 (± 3.6)
17.6 (± 5.1)
30.7 (± 11.4)
<0.001
SAPS 2
32 (± 15)
28 (± 16)
28 (± 16)
0.007
Hospital mortality
58 (26.9%)
48 (21.2%)
29 (13.2%)
0.002
Lethal sepsis
11 (5.1%)
9 (4.1%)
0 (0.0%)
0.005 T Steele , R Kolamunnage-Dona , C Downey , C Toh , I Welters
1University of Liverpool, UK; 2Royal Liverpool University Hospital, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P446 (doi: 10.1186/cc12384) Introduction Hypocalcaemia is common in critical illness and accurate
assessment is crucial. Small studies have shown that albumin-adjusted
calcium (adjCa) does not accurately predict the ionised calcium (iCa)
concentration in critically ill patients, yet adjCa continues to be widely
used [1]. We investigated the reliability of using adjCa to identify
hypocalcaemia in a large, diverse population requiring intensive care. Introduction Hypocalcaemia is common in critical illness and accurate
assessment is crucial. Small studies have shown that albumin-adjusted
calcium (adjCa) does not accurately predict the ionised calcium (iCa)
concentration in critically ill patients, yet adjCa continues to be widely
used [1]. We investigated the reliability of using adjCa to identify
hypocalcaemia in a large, diverse population requiring intensive care. Methods In a retrospective study of patients admitted to the ICUs of a
tertiary care hospital between January 2008 and 2012, iCa and pH were
extracted from routine blood gas results and total calcium, albumin
and phosphate from routine biochemistry results. adjCa was calculated
using a formula derived from and validated on the local population
[2]. Sensitivity, specifi city, positive and negative predictive values
(PPV and NPV) and area under the curve (AUC) of adjCa for predicting
hypocalcaemia (iCa <1.1 mmol/l) were calculated. Vitamin D status and overall and sepsis mortality in critically ill
patients Table 1 (abstract P445). Characteristics of the two groups
G1
G2
P value
Age
38 ± 13
34 ± 12
NS
TBSA
57.8 ± 30
26 ± 22
0.019
UBS
158 ± 100
70 ± 99
NS
ABSI
8 ± 3
6 ± 3
NS
CS T0
37 ± 11
19 ± 14
0.01
D CS
2.2 ± 1.8
19 ± 6
<0.001
Shock
6 (86%)
4 (36%)
0.04 Table 1 (abstract P445). Characteristics of the two groups y
,
Critical Care 2013, 17(Suppl 2):P447 (doi: 10.1186/cc123 Introduction Vitamin D status may aff ect hospital mortality and
infectious complications in critical illness. Methods A total of 655 mixed critically ill patients with available
25(OH)D levels hospitalized between 2008 and 2010 were included. Cox regression analysis adjusted for SAPS 2, age and gender was
performed. 25(OH)D levels were categorized by month-specifi c tertiles
(high, intermediate, low) to refl ect seasonal variation. Primary endpoint
was the correlation between 25(OH)D and hospital mortality. Secondary
endpoints were sepsis mortality and blood culture positivity.i y
y
Results Hospital mortality was signifi cantly higher in patients in the
low (HR = 1.98) and the mid-range tertile (HR = 1.88) compared with
the highest tertile. Vitamin D levels were signifi cantly lower in patients
with lethal sepsis compared with other causes of death (12.3 ± 5.0
vs. 18.2 ± 11.2, P = 0.02). Blood culture positivity rates did not diff er
between the groups (23.0% vs. 26.8% vs. 17.3%, P = 0.361). See Table 1. Conclusion RAI is common in severely burned patients during the
acute phase, and is associated with shock. Further prospective
controlled studies will be necessary to establish risk factors of RAI in
severely burned patients and its impact on their prognosis. Relative adrenal insuffi ciency in burns With direct iCa measurements now readily available
on ward-based analysers, adjCa should not be used to determine
calcium status in critical illness. phosphate or pH. With direct iCa measurements now readily available
on ward-based analysers, adjCa should not be used to determine
calcium status in critical illness. university-affi liated teaching hospital in Tunis. Patients admitted within
the fi rst 24 hours post burn with greater than 10% total body surface
area (TBSA) burned were enrolled in this study from 1 January 2009
to 30 June 2010. Exclusion criteria were pregnancy, history of adrenal
insuffi ciency, or steroid therapy within 6 months prior to burns. A
short corticotrophin test (250 μg) was performed, and cortisol levels
were measured at baseline (CS T0) and 60 minutes post test. Adrenal
insuffi ciency was defi ned by a response ≤9 μg/dl. Relative adrenal
insuffi ciency was further defi ned by a baseline cortisol >20 μg/dl. Results Patients were assigned into two groups: G1 (RAI, n = 7) and G2
(absence AI, n = 11). Comparative study of the two groups shows the
results presented in Table 1. 1. Slomp J, et al.: Albumin-adjusted calcium is not suitable for diagnosis of
hyper- and hypocalcemia in the critically ill. Crit Care Med 2003,
31:1389-1393. 1. Slomp J, et al.: Albumin-adjusted calcium is not suitable for diagnosis of
hyper- and hypocalcemia in the critically ill. Crit Care Med 2003,
31:1389-1393. 2. James M, et al.: Derivation and internal validation of an equation for
albumin-adjusted calcium. BMC Clin Pathol 2008, 8:12. Relative adrenal insuffi ciency in burns Relative adrenal insuffi ciency in burns pp
p
Results Of the 2,440 patients studied, 1,993 had APACHE II data and
serum Na+ recorded and so were included for analysis. In total, 453
patients (22.7%) had a serum Na+ ≤134 mmol/l and 1,388 patients
(67.1%) had a serum Na+ of 135 to 144 mmol/l. Patients with a low Na+
had a higher mortality (OR = 1.48, 95% CI = 1.16 to 1.90, P <0.001), a
higher APACHE II score (22 vs. 19, P <0.001) and higher mean age
(60 years vs. 58 years, P <0.001) than patients with a normal serum Na+. Mean length of stay of patients with low serum Na+ was also longer
(5.1 days vs. 4.6 days) although this was not statistically signifi cant
(P = 0.09). Introduction Relative adrenal insuffi ciency (RAI) is an uncommon
disorder among burn patients, which can often go unrecognized. However, there are no or almost no data about the incidence of this
disease in burns. The goal of the current study is to evaluate the
incidence of RAI in burn patients during the acute phase. Methods A prospective study, approved by our Institutional
Ethics Committee, was conducted in a 20-bed adult burn ICU at a Conclusion In summary, hyponatremia is a useful index of severity of
illness in our ICU population. Whether this is a direct adverse eff ect S165 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 university-affi liated teaching hospital in Tunis. Patients admitted within
the fi rst 24 hours post burn with greater than 10% total body surface
area (TBSA) burned were enrolled in this study from 1 January 2009
to 30 June 2010. Exclusion criteria were pregnancy, history of adrenal
insuffi ciency, or steroid therapy within 6 months prior to burns. A
short corticotrophin test (250 μg) was performed, and cortisol levels
were measured at baseline (CS T0) and 60 minutes post test. Adrenal
insuffi ciency was defi ned by a response ≤9 μg/dl. Relative adrenal
insuffi ciency was further defi ned by a baseline cortisol >20 μg/dl. Results Patients were assigned into two groups: G1 (RAI, n = 7) and G2
(absence AI, n = 11). Comparative study of the two groups shows the
results presented in Table 1. phosphate or pH. Table 1 (abstract P447) Methods In a retrospective study of patients admitted to the ICUs of a
tertiary care hospital between January 2008 and 2012, iCa and pH were
extracted from routine blood gas results and total calcium, albumin
and phosphate from routine biochemistry results. adjCa was calculated
using a formula derived from and validated on the local population
[2]. Sensitivity, specifi city, positive and negative predictive values
(PPV and NPV) and area under the curve (AUC) of adjCa for predicting
hypocalcaemia (iCa <1.1 mmol/l) were calculated. Etiology of low testosterone levels in mal
sepsis requiring mechanical ventilation
h Unpaired t test showed a P value of 0.07. Conclusion Calcium replacement with 40 ml of 10% calcium gluconate
is eff ective when the adjusted calcium is <2.0 mmol/l. Replacement
with 20 ml of 10% calcium gluconate when the adjusted calcium
is between 2.0 and 2.15 mmol/l is not much more eff ective than no
replacement. This suggests that either replacement is not needed or
there is under-replacement in mild hypocalcaemia. Adjusted calcium
may be giving falsely low calcium results in mild hypocalcaemia. The
measurement of ionised calcium is a possible solution [2,3]. References
1. Zaloga GP: Crit Care Med 1992, 20:251-262. 2. Slomp J, et al.: Crit Care Med 2003, 31:1389-1393. 3. Byrnes MC, et al.: Am J Surg 2005, 189:310-314. P450
Endogenous melatonin in critically ill patients
VS Salice, IG Galluccio, BS Salihovic, IP Piva, FM Marazzo, CV Villa,
MT Taverna, MU Umbrello, GM Mistraletti, GI Iapichino
Figure 1 (abstract P448). Figure 2 (abstract P448). Figure 1 (abstract P448). the adjusted calcium result and the calcium replacement were
documented. Adequate replacement is defi ned as 40 ml of 10%
calcium gluconate if the adjusted calcium is <2.0 mmol/l and 20 ml of
10% calcium gluconate if the adjusted calcium result is between 2.0
and 2.15 mmol/l. Figure 2 (abstract P448). Figure 2 (abstract P448). Results Eighty-eight adjusted calcium results were <2.0 mmol/l. In 74
results adequate replacement was given and the average increase in
calcium was 0.2 mmol/l. This led to an increase in the adjusted calcium
to >2.15 mmol/l in 36% of results and improvement to >2.0 mmol/l
in 89% of results. In 14 results no replacement was given and the
average increase in calcium was 0.07 mmol/l, which equated to 57%
of calcium results remaining <2.0 mmol/l. Unpaired t test showed a P
value of 0.0006. A total of 150 adjusted calcium results were between
2.0 and 2.15 mmol/l. In 111 results adequate replacement was given
and the average increase was 0.006 mmol/l. The adjusted calcium
fell to <2.0 mmol/l in 26% of results and normalised to >2.15 mmol/l
in 23%. Where the adjusted calcium fell despite replacement, 51%
of results were below 2.05 mmol/l. In 39 results no replacement was
given, leading to an average fall in calcium of 0.03 mmol/l. Thirty-
three percent of calcium results fell to <2.0 mmol/l and 20% of results
normalised. Unpaired t test showed a P value of 0.07. References 1. Zaloga GP: Crit Care Med 1992, 20:251-262. 2. Slomp J, et al.: Crit Care Med 2003, 31:1389-1393. 3. Byrnes MC, et al.: Am J Surg 2005, 189:310-314. Etiology of low testosterone levels in mal
sepsis requiring mechanical ventilation
h FSH levels were inappropriately low. All lipoprotein fractions and their
apo-proteins were reduced as well as 17-OH-progesterone, DHEA and
DHEAS. In contrast, androstenedione (adione) levels were elevated. This suggests preferential and stimulated synthesis of androstenedione
(Figure 2). The high 17β-estradiol levels indicate that androstenedione
is shunted into the estrogen pathway, a process that requires high
aromatase activity. The high estradiol/total testosterone ratio supports
this conclusion. p
Conclusion Calcium replacement with 40 ml of 10% calcium gluconate
is eff ective when the adjusted calcium is <2.0 mmol/l. Replacement
with 20 ml of 10% calcium gluconate when the adjusted calcium
is between 2.0 and 2.15 mmol/l is not much more eff ective than no
replacement. This suggests that either replacement is not needed or
there is under-replacement in mild hypocalcaemia. Adjusted calcium
may be giving falsely low calcium results in mild hypocalcaemia. The
measurement of ionised calcium is a possible solution [2,3]. Conclusion Hyperestrogenic hypotestosteronemia is a frequent
fi nding in the acute phase of severe sepsis in male patients with
respiratory failure. It is suggested to be caused by decreased androgen
production and shunting of androgen to estrogen synthesis as a result
of increased aromatase activity. The clinical relevance of gonadal
hormone substitution needs further study. Effi cacy of calcium replacement in hypocalcaemia
H Chatha, K Sim
Whiston Hospital, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P449 (doi: 10.1186/cc12387 Effi cacy of calcium replacement in hypocalcaemia
H Chatha, K Sim
Whiston Hospital, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P449 (doi: 10.1186/cc12387) Etiology of low testosterone levels in mal
sepsis requiring mechanical ventilation
h This cannot be explained by abnormalities of albumin, S166 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 http://ccforum.com/supplements/17/S2 FSH levels were inappropriately low. All lipoprotein fractions and their
apo-proteins were reduced as well as 17-OH-progesterone, DHEA and
DHEAS. In contrast, androstenedione (adione) levels were elevated. This suggests preferential and stimulated synthesis of androstenedione
(Figure 2). The high 17β-estradiol levels indicate that androstenedione
is shunted into the estrogen pathway, a process that requires high
aromatase activity. The high estradiol/total testosterone ratio supports
this conclusion. Conclusion Hyperestrogenic hypotestosteronemia is a frequent
fi nding in the acute phase of severe sepsis in male patients with
respiratory failure. It is suggested to be caused by decreased androgen
production and shunting of androgen to estrogen synthesis as a result
of increased aromatase activity. The clinical relevance of gonadal
hormone substitution needs further study. P449
Effi cacy of calcium replacement in hypocalcaemia
H Chatha, K Sim
Whiston Hospital, Liverpool, UK
Critical Care 2013, 17(Suppl 2):P449 (doi: 10.1186/cc12387)
the adjusted calcium result and the calcium replacement were
documented. Adequate replacement is defi ned as 40 ml of 10%
calcium gluconate if the adjusted calcium is <2.0 mmol/l and 20 ml of
10% calcium gluconate if the adjusted calcium result is between 2.0
and 2.15 mmol/l. Results Eighty-eight adjusted calcium results were <2.0 mmol/l. In 74
results adequate replacement was given and the average increase in
calcium was 0.2 mmol/l. This led to an increase in the adjusted calcium
to >2.15 mmol/l in 36% of results and improvement to >2.0 mmol/l
in 89% of results. In 14 results no replacement was given and the
average increase in calcium was 0.07 mmol/l, which equated to 57%
of calcium results remaining <2.0 mmol/l. Unpaired t test showed a P
value of 0.0006. A total of 150 adjusted calcium results were between
2.0 and 2.15 mmol/l. In 111 results adequate replacement was given
and the average increase was 0.006 mmol/l. The adjusted calcium
fell to <2.0 mmol/l in 26% of results and normalised to >2.15 mmol/l
in 23%. Where the adjusted calcium fell despite replacement, 51%
of results were below 2.05 mmol/l. In 39 results no replacement was
given, leading to an average fall in calcium of 0.03 mmol/l. Thirty-
three percent of calcium results fell to <2.0 mmol/l and 20% of results
normalised. 50
Endogenous melatonin in critically ill patients The higher melatonin
peak in renal failure may be due to an increased distribution volume;
greater AUC in patients with liver failure could be due to a less effi cient
removal of the hormone from the systemic circulation. The fi nding of
increased peak and AUC in nonsurvivor patients could be due to a
hormonal response increased by the body stress reaction, potentially
similar to cortisol [1], or to a higher production of a physiological
antioxidant [2] with a decreased ability to use it. References
1. Venkatesh B, et al.: Best Practice Res Clin End Met 2011, 25:719-733. 2. Flaring UB, et al.: Intensive Care Med 2005, 31:1072-1078. P451
Succinate ameliorates mitochondrial oxygen consumption of
metformin-intoxicated human platelets
A Protti1, M Monti2, A Lecchi1, A Artoni1, N Greppi1, L Gattinoni1
1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Methods Platelet-rich-plasma was incubated for 72 hours with
metformin at a fi nal concentration of 0 mg/l (control), 1.66 mg/l
(therapeutic dose) or 166 mg/l (toxic dose). Platelet VO2 was then
measured with a Clark-type electrode, in the presence of glutamate plus
malate (complex I electron donors) (fi nal concentration: 20 mmol/l for
both) or succinate (complex II electron donor) (30 mmol/l), before and
after adding cyanide (40 mmol/l). Mitochondrial (cyanide-sensitive)
and extra-mitochondrial (cyanide-insensitive) VO2 were corrected for
platelet count. Results The main results, from four preliminary experiments, are
shown in Figure 1. In the presence of glutamate plus malate, only
platelets incubated with a high dose of metformin had a mitochondrial
VO significantly lower than controls. In the presence of succinate,
Figure 1 (abstract P450). Figure 1 (abstract P451). Mitochondrial oxygen use of human platelets
incubated with metformin. Figure 1 (abstract P450). this study is to describe the endogenous blood melatonin values in ICU
patients and their correlation with clinical parameters. Figure 1 (abstract P451). Mitochondrial oxygen use of human platelets
incubated with metformin. Methods Seventy-three high-risk critically ill patients mechanically
ventilated for >48 hours were enrolled. Blood samples for melatonin
assay were collected between the 3rd and the 8th day of the ICU stay. Melatonin was determined by radioimmunoassay and ELISA. The
peak and the area under the curve (AUC) calculated for each patient
were correlated with the clinical parameters using the regression for
quantiles test. 50
Endogenous melatonin in critically ill patients Endogenous melatonin in critically ill patients
VS Salice, IG Galluccio, BS Salihovic, IP Piva, FM Marazzo, CV Villa,
MT Taverna, MU Umbrello, GM Mistraletti, GI Iapichino
Università degli Studi Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P450 (doi: 10.1186/cc12388) Introduction The aim of the study is to examine the effi cacy of calcium
replacement in patients with hypocalcaemia in the ICU. Hypocalcaemia
aff ects between 15 and 50% of admissions to the ICU [1]. Introduction The aim of the study is to examine the effi cacy of calcium
replacement in patients with hypocalcaemia in the ICU. Hypocalcaemia
aff ects between 15 and 50% of admissions to the ICU [1]. Introduction Melatonin could have a meaningful role in critically ill
patients, because of its immunomodulatory, antioxidant and sleep
regulation properties; it is reduced in critical illness. The purpose of Introduction Melatonin could have a meaningful role in critically ill
patients, because of its immunomodulatory, antioxidant and sleep
regulation properties; it is reduced in critical illness. The purpose of f
Methods Adjusted calcium results <2.0 mmol/l were followed up
from admission for 5 days or until discharge or death. On each day S167 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 this study is to describe the endogenous blood melatonin values in ICU
patients and their correlation with clinical parameters. Methods Seventy-three high-risk critically ill patients mechanically
ventilated for >48 hours were enrolled. Blood samples for melatonin
assay were collected between the 3rd and the 8th day of the ICU stay. Melatonin was determined by radioimmunoassay and ELISA. The
peak and the area under the curve (AUC) calculated for each patient
were correlated with the clinical parameters using the regression for
quantiles test. Results Endogenous melatonin was found lower in critically ill patients
compared with healthy subjects (Figure 1), although it showed a
great individual variability and it generally maintained a night-time
increase. In the univariate analysis the peak was found related to: blood
creatinine (P = 0.034); patients in coma (P = 0.024); hospital mortality
(P = 0.016). The AUC was found related to: SAPS II (P = 0.047); creatinine
(P <0.001); AST (P <0.001); ALT (P <0.001); hospital mortality (P <0.022). Peak and AUC were found higher in nonsurvivor patients. Conclusion In accord with previous studies, the endogenous blood
melatonin was found reduced in ICU patients. References 1. Venkatesh B, et al.: Best Practice Res Clin End Met 2011, 25:719-733. 2. Flaring UB, et al.: Intensive Care Med 2005, 31:1072-1078. 50
Endogenous melatonin in critically ill patients Results Endogenous melatonin was found lower in critically ill patients
compared with healthy subjects (Figure 1), although it showed a
great individual variability and it generally maintained a night-time
increase. In the univariate analysis the peak was found related to: blood
creatinine (P = 0.034); patients in coma (P = 0.024); hospital mortality
(P = 0.016). The AUC was found related to: SAPS II (P = 0.047); creatinine
(P <0.001); AST (P <0.001); ALT (P <0.001); hospital mortality (P <0.022). Peak and AUC were found higher in nonsurvivor patients. g
p
Conclusion In accord with previous studies, the endogenous blood
melatonin was found reduced in ICU patients. The higher melatonin
peak in renal failure may be due to an increased distribution volume;
greater AUC in patients with liver failure could be due to a less effi cient
removal of the hormone from the systemic circulation. The fi nding of
increased peak and AUC in nonsurvivor patients could be due to a
hormonal response increased by the body stress reaction, potentially
similar to cortisol [1], or to a higher production of a physiological
antioxidant [2] with a decreased ability to use it. Figure 1 (abstract P451). Mitochondrial oxygen use of human platelets
incubated with metformin. Methods Platelet-rich-plasma was incubated for 72 hours with
metformin at a fi nal concentration of 0 mg/l (control), 1.66 mg/l
(therapeutic dose) or 166 mg/l (toxic dose). Platelet VO2 was then
measured with a Clark-type electrode, in the presence of glutamate plus
malate (complex I electron donors) (fi nal concentration: 20 mmol/l for
both) or succinate (complex II electron donor) (30 mmol/l), before and
after adding cyanide (40 mmol/l). Mitochondrial (cyanide-sensitive)
and extra-mitochondrial (cyanide-insensitive) VO2 were corrected for
platelet count. 1.
Venkatesh B, et al.: Best Practice Res Clin End Met 2011, 25:719-733.
2.
Flaring UB, et al.: Intensive Care Med 2005, 31:1072-1078. Does chronic metformin therapy off er cardiomyoprotection in
coronary artery bypass grafting surgery patients? A case–control
study A Delaporte, JP Hacquebard, P Origer, I Pastijn, M Maatouk, O Germay,
E Stevens, P Kapessidou, A Roman
CHU Saint-Pierre, Brussels, Belgium
Critical Care 2013, 17(Suppl 2):P452 (doi: 10.1186/cc12390) g
Results Ten patients were selected, fi ve males and fi ve females, with a
mean age of 72.2. On admission, nine of 10 patients had a framework
of general illness and acute renal failure; one patient appeared with
a probable acute abdomen not subsequently confi rmed on CT. Eight
patients had a state of dehydration resulting from gastroenteritis,
diarrhea, fever; two patients had taken NSAIDs in the days prior to
hospitalization. No patient, in these conditions, has discontinued
treatment with metformin. All patients showed a critical clinical
framework with a SAPS II mean score of 77.5. Cardiovascular and
respiratory support was required in all cases. The hemogas analysis
showed that patients had a severe metabolic acidosis (mean pH = 6.96)
with increased plasma lactate (mean lactate = 15.93 mmol/l). Prothrombin activity was normal in eight of 10 patients. The overall
mortality was 70%. Introduction Metformin, widely used as an antidiabetic drug, activates
the AMP activated protein kinase, a key regulator of the metabolism
providing protection under fuel defi ciency. Chronic metformin therapy
has been shown in long-term follow-up clinical studies to reduce
cardiovascular mortality [1]. In animal experiments, acute metformin
pretreatment has been shown to reduce ischemia–reperfusion injury
on cardiomyocytes [2]. We want to evaluate whether outcomes are
aff ected in coronary artery bypass grafting (CABG) surgery. f
Methods We performed a retrospective case–control study on 341
patients admitted exclusively for CABG surgery with cardiopulmonary
bypass between January 2006 and April 2012. Eighty-one patients
chronically on metformin were matched 1:3 with nonmetformin-
treated ones using gender, age, body mass index and number of
arterial bypass grafts. Global mortality, ICU-free 28 postoperative days
and creatinine kinase MB (CKMB) at postoperative baseline, 8 and
18 hours post baseline were compared.i Conclusion Patient education on correct use of metformin is essential
to prevent MALA, specially in those clinical conditions of increased risk
(for example, acute renal failure), as recommended by the AIFA 2011
guidelines [4]. In our study, a higher plasma concentration of lactate
represents the main negative prognostic factor, as pointed out by
other studies [5]. P452 Does chronic metformin therapy off er cardiomyoprotection in
coronary artery bypass grafting surgery patients? A case–control
study
A Delaporte, JP Hacquebard, P Origer, I Pastijn, M Maatouk, O Germay,
E Stevens, P Kapessidou, A Roman
CHU Saint-Pierre, Brussels, Belgium
Critical Care 2013, 17(Suppl 2):P452 (doi: 10.1186/cc12390) Multiscale modeling of acute insulin resistance in critical care Critical Care 2013, 17(Suppl 2):P454 (doi: 10.1186/cc12392) Critical Care 2013, 17(Suppl 2):P454 (doi: 10.1186/cc12392) Introduction Stress hyperglycemia in the critically ill is a complex
process in which insulin signaling is systematically hijacked to
provide energy substrate for metabolic priorities such as cell healing
or infection containment. Fluctuating levels of plasma glucose are
associated with increased mortality in the ICU [1]. We develop a
multiscale mathematical model that can characterize the severity of
stress hyperglycemia based on a fundamental understanding of the
signaling molecules involved. Conclusion These data suggest that chronic preoperative treatment
with metformin is associated with lower CKMB levels at postsurgery
baseline, 8 and 18 hours post baseline with a shorter ICU stay in patients
receiving CABG surgery under extracorporeal circulation. References g
g
Methods Insulin resistance following insult has been shown to be
driven primarily by the immune response via the cytokine IL-6 [2]. We
created a multiscale mathematical model that links circulating glucose
and insulin concentration dynamics from the extended minimal
model [3] to a cellular insulin response model [4] that captures insulin-
mediated glucose uptake in an insulin-responsive cell. p
References References . UK Prospective Diabetes Study Group: Lancet 1998, 352:854-865. 2. Lalau JD: Drug Saf 2010, 33:727-740. 3. Le Gall JR, et al.: JAMA 1993, 270:2957-2963. 4. Italian Medicines Agency 2011 [http://www.agenziafarmaco.gov.it/it/content/
raccomandazioni-sull%E2%80%99utilizzo-dei-medicinali-base-di-metformina-
nella-gestione-del-diabete-m] Table 1 (abstract P452). Case–control analysis Table 1 (abstract P452). Case–control analysis Table 1 (abstract P452). Case–control analysis Table 1 (abstract P452). Case–control analysis
Statistics,
Metformin
Nonmetformin
P value
Mortality
1/81
8/260
0.24
28 days free ICU (days)
23.4
22.9
<0.04
CKMB T 0 hours (ng/ml)
21
26
<0.003
CKMB T+8 hours (ng/ml)
22
53
<0.004
CKMB T+18 hours (ng/ml)
21.5
27.5
<0.012 5. Dell’Aglio DM, et al.: Ann Emerg Med 2009, 54:818-823. 6. Peters N, et al.: Crit Care 2008, 12:R149. Succinate ameliorates mitochondrial oxygen consumption of
metformin-intoxicated human platelets Succinate ameliorates mitochondrial oxygen consumption of
metformin-intoxicated human platelets
A Protti1, M Monti2, A Lecchi1, A Artoni1, N Greppi1, L Gattinoni1
1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P451 (doi: 10.1186/cc12389) p
Results The main results, from four preliminary experiments, are
shown in Figure 1. In the presence of glutamate plus malate, only
platelets incubated with a high dose of metformin had a mitochondrial
VO2 signifi cantly lower than controls. In the presence of succinate,
mitochondrial VO2 of controls did not change signifi cantly whereas
that of platelets incubated with metformin did. The eff ect of succinate
tended to become larger as the dose of metformin was increased from
0 up to 166 mg/l (0.3 ± 0.2 vs. 0.6 ± 0.3 vs. 1.0 ± 0.3 nmol/minute*106
cells) (P = 0.068). Even so, mitochondrial VO2 of platelets incubated with
the highest dose of metformin did not return to the levels of controls. Extra-mitochondrial VO2 was always the same. A Protti1, M Monti2, A Lecchi1, A Artoni1, N Greppi1, L Gattinoni1
1Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy;
2Università degli Studi di Milano, Milan, Italy
Critical Care 2013, 17(Suppl 2):P451 (doi: 10.1186/cc12389) Introduction Metformin intoxication inhibits mitochondrial complex
I and oxygen consumption (VO2). Succinate bypasses complex I by
donating electrons to complex II. The aim of this study was to clarify
whether succinate ameliorates mitochondrial VO2 of metformin-
intoxicated human platelets. S168 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Conclusion Succinate ameliorates (but does not return to normal)
mitochondrial VO2 of human platelets incubated with a toxic dose of
metformin. predisposing conditions, the use of metformin may result in metformin-
associated lactic acidosis (MALA), a rare adverse event associated with a
high mortality rate [2]. The aim of this study is to assess risk factors and
prognostic factors in patients with MALA. Methods We conducted a retrospective study of patients with MALA
admitted to the ICU of ASL 2 Chieti between 1 January 2008 and
30 September 2012. The eligibility criteria were: diagnosis of diabetes
mellitus type 2, treatment with oral hypoglycemic drugs containing
metformin, increased anion gap metabolic acidosis (pH <7.35, HCO3
–
<22 mmol/l, lactate >5 mmol/l). For each patient we evaluated: sex,
age, home care, SAPS II score [3], blood tests. Does chronic metformin therapy off er cardiomyoprotection in
coronary artery bypass grafting surgery patients? A case–control
study The prothrombin activity, which is considered to be a
decisive prognostic factor in the study of Peters and colleagues [6], was
not impaired in patients with poor outcome. Results Metformin-treated patient mortality was not signifi cantly
lower than in nonmetformin (1.2% vs. 3.1%, P = 0.24, Fischer exact test). ICU-free 28 postoperative days were higher in the metformin group
(median 23.4 days; 95% CI = 22.5 to 24.3) than in the nonmetformin
group (median 22.9 days; 95% CI 22.3 to 23.5, P <0.04, Wilcoxon test). Median CKMB levels at postoperative baseline were 21 versus 26 ng/ml
(P <0.003), at 8 hours were 22 versus 53 ng/ml (P <0.004) and 18 hours
post baseline were 21.5 versus 27.5 ng/ml (P <0.012). See Table 1. Impact of a real-time electronic persistent hyperglycemia alert on
glucose control in the ICU g
K Colpaert, S Oeyen, T van den Driessche, C Danneels, J Decruyenaere
Ghent University Hospital, Ghent, Belgium
Critical Care 2013, 17(Suppl 2):P455 (doi: 10.1186/cc12393) Introduction Hyperglycemia is frequently encountered in critically
ill patients, and associated with adverse outcome. Improvement of
glucose protocol adherence may be accomplished using electronic
alerts. We confi gured a non-intrusive real-time electronic alert, called a
GLYC sniff er, as part of our Intensive Care Information System (ICIS) that
continuously evaluates the occurrence of persistent hyperglycemia
and hypoglycemia. The GLYC sniff er is confi gured in such a way to
give an alert: if two consecutive glucose values are >150 mg/dl, with
a minimum interval of 60 minutes; or if a glucose value is <80 mg/dl. We wanted to evaluate whether the GLYC sniff er would improve the
glucose control. p p
Conclusion The MPC–MHE algorithm achieves targeted glucose
control in response to changing patient dynamics and multiple
measured disturbances for a pilot population of 10 VPs. Furthermore,
the MHE scheme updates patient parameters in real time in response
to changing patient dynamics. P457 Methods A single-center, prospective intervention study during a
6-month period in our 22-bed surgical ICU. Two study phases were
compared: a 3-month pre-alert phase with no alerting to the nurses,
and a 3-month intervention phase where the GLYC sniff er was alerting
through the Clinical Notifi cation System of our ICIS.f Hyperglycemia in intensive care identifi es patients at high risk of
developing diabetes: preliminary data of a multicentre study
E Lukic, I Gornik, M Grgic-Medic, D Milicic, V Vegar, V Ivancan, M Peric,
V Gasparovic, I Pavlic Renar
University Hospital Centre Zagreb, Croatia
Critical Care 2013, 17(Suppl 2):P457 (doi: 10.1186/cc12395) i
y
Results A total of 652 diff erent patients having a total of 699 admission
episodes was recorded during the study period. There were no
signifi cant diff erences between the two study groups regarding
baseline demographic data, fi rst glucose value upon admission. A
total of 2,335 GLYC sniff er alerts were recorded during the whole study
period: 84.3% persistent hyperglycemia alerts (1,021 in the pre-alert
group, 948 in the alert group), and 15.7% hypoglycemia alerts (139
vs. 227). A signifi cantly lower percentage of glucose values in the alert
group were hyperglycemic (19.5% vs. 26.5%, P <0.001). The proportion
of persistent hyperglycemic values (i.e. P455 Results MPC controller performance on one VP is shown in Figure 1. Across a population of 10 VPs, the average [Gbl] under MPC is
6.31 mmol/l, the average minimum is 4.62 mmol/l, the population
individual minimum is 3.49 mmol/l and the average absolute average
residual error is 0.83 mmol/l from a 5.6 mmol/l target. With standard
intervention, the 10 VPs have an average [Gbl] of 9.32 mmol/l, an
average minimum [Gbl] of 3.77 mmol/l, and a population minimum
[Gbl] of 2.78 mmol/l. Algorithm performance deteriorates signifi cantly
if the imputed sampling time exceeds 30 minutes, underlining
the importance of dynamic variations in insulin sensitivity in this
population. y
References References
1. Van den Berghe G: N Engl J Med 2001, 345:1359-1367. 2. Thorell A: Clin Nutr 1996, 15:75-79. 3. Roy A: Diabetes Technol Ther 2006, 8:617-626. 4. Sedaghat A: Am J Physiol Endocrinol Metab 2002, 283:E1084-E1101. References
1. Van den Berghe G: N Engl J Med 2001, 345:1359-1367. 2. Thorell A: Clin Nutr 1996, 15:75-79. 3. Roy A: Diabetes Technol Ther 2006, 8:617-626. 4. Sedaghat A: Am J Physiol Endocrinol Metab 2002, 283:E1084-E1101. References
1. Van den Berghe G: N Engl J Med 2001, 345:1359-1367. 2. Thorell A: Clin Nutr 1996, 15:75-79. 3. Roy A: Diabetes Technol Ther 2006, 8:617-626. 4. Sedaghat A: Am J Physiol Endocrinol Metab 2002, 283:E1084-E1101. 3. Roy A: Diabetes Technol Ther 2006, 8:617-626. 4. Sedaghat A: Am J Physiol Endocrinol Metab 2002, 283:E1084-E1101. P453
Metformin-associated lactic acidosis: risk factors and prognostic
factors Metformin-associated lactic acidosis: risk factors and prognostic
factors
V Di Falco, A Milano, M Battilana, F Araosta, A Grosso, D Albanese,
F Petrini, L Di Liberato
ASL Chieti SS. Annunziata, Chieti, Italy
Critical Care 2013, 17(Suppl 2):P453 (doi: 10.1186/cc12391) V Di Falco, A Milano, M Battilana, F Araosta, A Grosso, D Albanese,
F Petrini, L Di Liberato
ASL Chieti SS. Annunziata, Chieti, Italy
Critical Care 2013, 17(Suppl 2):P453 (doi: 10.1186/cc12391) Results Inhibitory dynamics driven by IL-6 were incorporated into the
cellular model to attenuate an insulin signaling intermediate (insulin
receptor substrate 1) according to the proposed biological mechanisms. The percentage reduction in glucose uptake as a function of IL-6
concentration was fi t to data from patients who underwent elective
abdominal surgery [2], shown in Figure 1. The overall multiscale model
captures decreased insulin signaling as a result of increased IL-6 levels
and the subsequent hyperglycemia that may ensue. y
Critical Care 2013, 17(Suppl 2):P453 (doi: 10.1186/cc12391) Introduction Metformin, an oral hypoglycemic drug, belongs to the
biguanide class and is now generally accepted as fi rst-line treatment in
type 2 diabetes mellitus, especially in overweight patients [1]. In some Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S169 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P454). Reduced glucose uptake driven by increased
plasma IL-6 levels. group (9.9% vs. 15.4%, P <0.001). The patients in the alert group spend
signifi cantly more time within the set target glucose interval of 80 to
150 mg/dl (82.3% vs. 75.0%, P = 0.009). A signifi cantly lower proportion
of patients experienced a new-onset hypoglycemic event (<80 mg/dl)
in the alert group (19.3% vs. 26.2%, P = 0.04). g
p
Conclusion A real-time electronic persistent glycemia sniff er resulted
in a signifi cantly higher proportion of normoglycemia, without
increasing the variability. Furthermore, hypoglycemic events occurred
less frequently, and were resolved more timely. Smart alerting is able to
improve quality of care, while diminishing the problem of alert fatigue. P456 P456
Individualized targeted glucose control to avoid hypoglycemia
SP Gawel1, G Clermont2, T Ho1, BM Newman3, B Yegneswaran2, RS Parker1
1University of Pittsburgh, PA, USA; 2University of Pittsburgh Medical Center,
Pittsburgh, PA, USA; 3Iowa State University, Ames, IA, USA
Critical Care 2013, 17(Suppl 2):P456 (doi: 10.1186/cc12394) Introduction Hyperglycemia and hypoglycemia have been linked to
worse outcomes in critically ill patients. While there is controversy as
to the optimal tightness of glucose control in critically ill patients, there
is agreement that an upper limit to safe glucose levels exists and that
avoiding hypoglycemic episodes should be prioritized. Our algorithm
can assist clinicians in maintaining blood glucose ([Gbl]) within a
desired target range while avoiding hypoglycemia. Figure 1 (abstract P454). Reduced glucose uptake driven by increased
plasma IL-6 levels. Conclusion A multiscale model has been developed to describe the
inhibitory eff ects of IL-6 on insulin-mediated glucose uptake. Cellular
inhibitory dynamics were shown to capture reduced insulin sensitivity
on the macroscale, which could then be used to characterize insulin
sensitivity and to provide insulin treatment advice to reduce glucose
variability. Methods Our model predictive control (MPC) algorithm uses insulin
and glucose as control inputs and a linearized model of glucose–
insulin–fatty acid interactions. To allow the controller model to learn
from data, a moving horizon estimation (MHE) technique tailored
the tissue sensitivity to insulin to individual responses. Patient data
([Gbl] measurements, insulin and nutritional infusion rates) were from
the HIDENIC database at the University of Pittsburgh Medical Center. [Gbl] measurements, typically hourly, were interpolated to impute
a measurement every 5 minutes. The model captured patient [Gbl]
via nonlinear least squares by adjusting insulin sensitivity (SI) and
endogenous glucose production (EGP0). The resulting virtual patient
(VP) is used to evaluate the performance of the MPC–MHE algorithm. Impact of a real-time electronic persistent hyperglycemia alert on
glucose control in the ICU consecutive glucose values
exceeding the limit of 150 mg/dl) was signifi cantly lower in the alert Introduction A recent study showed that hyperglycaemia (blood
glucose ≥7.8 mmol/l) in nondiabetic patients hospitalised in a medical
ICU is associated with increased risk of diabetes [1]. We investigated a
large mixed ICU population to confi rm these results. Methods This study retrospectively included patients with negative
history of diabetes admitted to ICUs during the year 2007. We excluded
patients receiving steroids, with newly diagnosed diabetes and those
with end-stage disease. Patients were followed-up 5 years after index S170 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P456). Figure 1 (abstract P456). Figure 1 (abstract P456). Figure 1 (abstract P456). admission. Diagnosis of diabetes within 6 months from the index
admission was presumed as revealing DM at inclusion, which excluded
the patient. Patients who were taking glucocorticoids during the follow-
up period were excluded. The rest of the patients, if consented, were
assessed for their diabetic status with a single HbA1c measurement. Introduction The objective of the study was to determine the
association between average glucose level (AVG) and hospital
outcomes in critically ill patients. Methods We performed a chart review of the MIMICII and HIDENIC
databases, prospective cohorts of over 32,000 and 46,000 patients
admitted to the ICUs at Beth Israel Deaconess Medical Center between
2001 and 2007 and the ICUs at University of Pittsburgh Medical Center
between 1991 and 2008, respectively. All admissions of patients
≥18 years without the diagnosis of DKA and NKHS were included. Diabetics were identifi ed from ICD-9 documentation. Propensity score
for death (pDead) was computed from either SAP1 (MIMICII) or APACHE
III (HIDENIC) to assess the risk of death. Hypoglycemia was defi ned as
AVG ≤60 mg/dl. AVG was computed as the area under the glucose
curve throughout ICU admission. Mortality was examined within
bins (each bin is categorized by a 10 mg/dl increase in AVG) and was
compared between adjacent categories using a chi-square test. The
same method was repeated among diabetics, nondiabetics, patients
with lower (pDead greater than median) and higher (pDead lower than Results A total of 2,696 patients were included in the study: 1,441
medical and 1,255 surgical, hyperglycaemia (≥7.8 mmol/l) was
registered in 1,419 (52%). Impact of a real-time electronic persistent hyperglycemia alert on
glucose control in the ICU During the 5 years from index admission 409
died and 38 were taking glucocorticoids, and 10 refused participation. Diabetes was diagnosed in 189 patients during the 6 months after index
admission. The remaining cohort of 2,320 patients had 1,169 patients
who were hyperglycaemic during hospitalisation. Diabetes was already
diagnosed in 132 patients, of whom 112 were hyperglycaemic in the
ICU; prediabetes was already diagnosed in 121 patients, of whom
83 had ICU hyperglycaemia. Since 125 patients (55 hyperglycaemic)
refused HbA1c testing, it was performed on 1,852 patients and revealed
diabetes in 136 patients (107 had hyperglycaemia) and prediabetes
in 103 patients (75 were hyperglycaemic in ICU). Overall, patients
who had hyperglycaemia had 19.8% cumulative 5-year incidence of
diabetes and 14.3% of prediabetes. Nonhyperglycaemic ICU patients
had cumulative incidences of 4.5% and 6.6%, respectively. Relative risk
for developing diabetes during 5 years after ICU admission is 4.2 (95%
CI = 3.2 to 5.9) for patients who did have hyperglycaemia.i Figure 1 (abstract P458). Figure 1 (abstract P458). p
yp
g y
Conclusion The results of this study confi rm in a large mixed medical/
surgical ICU population that hyperglycaemia occurring in severe
illness or the perioperative period is associated with increased risk
of developing diabetes. Suggesting lifestyle changes to reduce the
risk and regular follow-up should be implemented at least for these
patients. p
Reference p
Reference 1. Gornik I, et al.: A prospective observational study of the relationship of
critical illness associated hyperglycaemia in medical ICU patients and
subsequent development of type 2 diabetes. Crit Care 14:R130. Association between average glucose levels and hospital mortality
among critically ill patients B Yegneswaran1, R Parket2, S Gawel2, A Pritchard-Bell2, T Ho2, G Clermont2
1UPMC Hamot, Erie, PA, USA; 2University of Pittsburgh, PA, USA
Critical Care 2013, 17(Suppl 2):P458 (doi: 10.1186/cc12396) Figure 1 (abstract P458). Figure 1 (abstract P458). S171 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 with increased risk of mortality (M). Heightened attention to identifi ed
risk factors (RF) and causes of hypoglycemic events (HE) may lead to
improvement in glycemic control. A limited literature describes RF for
H but a detailed root cause analysis (RCA) of individual HE has not been
published. median) risk of death. Statistical signifi cance was noted following a
Bonferroni correction. Results MIMICII database: 21,209 admissions met inclusion criteria. Mean age: 62.5 ± 16.4 years; female: 9,070 (42.8%); diabetes: 5,496
(25.9%); mean admission SAP1 score: 13.8 ± 5.2; mean LOS: 9 days (IQR
6.15); hospital mortality: 10.3%. HIDENIC database: 38,872 admissions
met inclusion criteria. Mean age: 59.4 ± 17.3 years; female: 16,675
(42.9%); diabetes: 11,326 (29.1%); mean admission APACHE III score:
80.2 ± 14; mean LOS: 11 days (IQR 7.22); hospital mortality: 14.3%. In
both databases, association between average glucose and hospital
mortality followed a bathtub shape with mortality nadir between
80 and 130 mg/dl. No mortality diff erence was observed between
adjacent AVG glucose bins in the bottom of the bathtub (Figure 1). This relationship was preserved in nondiabetics, and for patients with
higher risk of death. p
Methods This is a retrospective analysis of prospectively collected data
including 835 patients admitted consecutively between 1 February
2012 and 31 October 2012 to the 16-bed medical–surgical ICU of a
university-affi liated teaching hospital. The blood glucose (BG, mg/dl)
target was 90 to 120 and the monitoring frequency was every 3 hours,
or every 1 hour when the patient received i.v. IN infusion. HE data were
collected by the bedside nurse, using a cutoff of BG <60, and information
was abstracted from the unit’s database. RF included: shock (SH), renal
insuffi ciency (RI), hepatic failure (HF), mechanical ventilation (MV), and
diabetes mellitus (DM). Attributable causes included: IN treatment,
improper monitoring frequency (MF), interruption of feeding (FE),
and spontaneous (SP). P461 Continuous arterial and venous glucose monitoring by quenched
chemical fl uorescence in ICU patients after cardiac surgery
S Bird1, L Macken1, O Flower1, F Bass1, N Hammond1, S Webb1, N Kennedy1,
A Baker1, E Yarad1, C Chau2, M Librande2, P Strasma2, S Finfer1
1Royal North Shore Hospital, Sydney, Australia; 2Glumetrics, Irvine, CA, USA
Critical Care 2013, 17(Suppl 2):P461 (doi: 10.1186/cc12399) Results Two groups had similar characteristics at baseline. A total of
1,060 CGM/ABG pairs were analyzed and reference glucose ranged
from 4.3 to 18.8 mmol/l. Median (IQR) absolute relative deviation was
lower in the FC arm (7.0% (3.5, 13.0) vs. 12.8% (6.3, 21.8), FC vs. MC,
P <0.001). Similarly, the percentage of points in the Clarke error grid
zone A points meeting ISO criteria were higher with FC (87.8% vs. 70.2%). Sensor bias (median (IQR)) was signifi cantly lower in the FC
arm (–0.1 (–0.7, 0.4) mmol/l vs. –1.1 (–2.3, –0.1) mmol/l, P <0.001). The
median (IQR) interval between calibrations in FC arm was 169 (122,
213) minutes. Introduction Continuous glucose monitoring (CGM) in ICUs has the
potential to improve patient safety outcomes. The GluCath Intravascular
CGM System uses a novel quenched chemical fl uorescence sensing
mechanism to measure glucose concentration in venous or arterial
blood (BG). This is the fi rst report of this system deployed for 48 hours
in both arteries and veins of ICU patients. Methods This ongoing clinical study evaluates up to two sensors per
subject in 20 patients undergoing cardiac surgery. An arterial sensor is
deployed via a standard 20 G radial artery catheter inserted for routine
care and an optional venous sensor is deployed percutaneously in an
upper arm vein. Data are presented from the fi rst fi ve patients. Outcome
measures are qualitative (ease of use, workfl ow fi t) and quantitative
(accuracy vs. reference analyzer). Sensors were inserted shortly after
ICU admission, with ultrasound guidance for venous sensor insertion. In vivo calibration was performed at 1 and 2 hours, then each morning. Glucose values were recorded every 10 seconds by the system. Hourly
arterial reference samples were analyzed via Radiometer ABL 800 Flex
Blood Gas Analyzer (BGA).i Conclusion CGM accuracy in the MC arm was comparable with accuracy
in subjects with type 1 diabetes. Further signifi cant improvements to
CGM accuracy in critical care are possible by increasing the frequency
of calibrations. References References
1. Weinstein RL, et al.: Diabetes Care 2007, 30:1125-1130. 2. Kovatchev B, et al.: Diabetes Care 2008, 31:1160-1164. Results Arterial sensors were successfully deployed in all fi ve patients
and did not interfere with clinical care, blood pressure monitoring
or sampling. One arterial catheter failed resulting in sensor removal
at 36 hours. The venous sensor was deployed in three patients, but
removed from two patients due to thrombosis identifi ed during
surveillance ultrasound examinations. A total of 202 reference BG
samples ranging from 5.3 to 11.3 mmol/l were collected. Precision
between arterial and venous sensors (n = 2) was 9.3% CV. Arterial
sensor accuracy compared with BGA was 5.5% MARD. One hundred Association between average glucose levels and hospital mortality
among critically ill patients We measured rates of severe (<40) and mild (40
to 69) H, and compared them as well as percentage of values in the
optimum range (70 to 139) and hyperglycemic range (>139) with data
from the preceding 3 months (PRE). Conclusion Low and high average glucose are associated with
mortality. P461 Such accurate CGM may provide valuable information
to guide insulin therapy in critically ill subjects. f Continuous glucose monitoring in critically ill adults: comparison of
two diff erent calibration protocols p
g
Results Sixty-three (7.5%) patients sustained a total of 79 HE. They were
older, more likely to be diabetic, and had higher APACHE IV predicted
(37.3% vs. 15.4%, P <0.0001) and observed (25.4% vs. 8.6%, P <0.0001)
M than those without H. RF: SH 24.1%; RI 24.1%; HF 8.9%; MV 46.8%; DM
36.5%. Attributable causes: IN 58.2%; MF 8.9%; FE 6.3%; SP 36.7%. BG
<40, 40 to 69 and >139 decreased 78.6%, 39.0% and 9.3% (P = 0.0006,
P <0.0001, P <0.0001). BG 70 to 139 increased 5.5% (P <0.0001). In
total, 4.2% and 22.4% of PRE patients sustained severe and mild H. In total, 0.7% and 14.1% of RCA patients sustained severe and mild H
(P = 0.0029, P = 0.0003). two diff erent calibration protocols
L Leelarathna1, S English2, H Thabit1, K Caldwell1, J Allen1,
K Kumareswaran1, M Wilinska1, M Nodale1, J Mangat1, M Evans1,
R Burnstein2, R Hovorka1
1University of Cambridge, UK; 2Addenbrookes Hospital, Cambridge, UK
Critical Care 2013, 17(Suppl 2):P459 (doi: 10.1186/cc12397) f
p
L Leelarathna1, S English2, H Thabit1, K Caldwell1, J Allen1, Introduction We evaluated the clinical and numerical accuracy of the
Freestyle Navigator continuous glucose monitoring (CGM) system, in
critically ill adults using two diff erent methods of calibration. Studies
looking at intensive glucose control have yielded confl icting results
with increased rates of iatrogenic hypoglycemia. Availability of accurate
real-time glucose information may improve safety and effi cacy of
glucose control in the critical care unit. Conclusion IN treatment was associated with barely more than
one-half of HE in this population of intensively monitored patients. Implementation of an initiative mandating RCA of every HE, including
identifi cation of RF and attributable causes, was associated with a
signifi cant improvement in glycemic control. g
Methods In a randomized prospective trial, paired CGM and reference
glucose (hourly arterial blood gas (ABG)) were collected from 24 adults
with critical illness (age 60 ± 14 years, BMI 29.6 ± 9.3 kg/m2, APACHE
score range 6 to 19) and hyperglycemia (glucose ≥10 mmol/l or treated
with intravenous insulin), over 48 hours. In 12 subjects, CGM was force-
calibrated at variable 1 to 6 hourly intervals using ABG glucose (FC
arm). In the other 12 subjects, the sensor was calibrated according to
the manufacturer’s instructions (1, 2, 10, 24 hours after insertion) using
arterial blood and built-in glucometer (MC arm). P461 Performance of the Medtronic Sentrino® continuous glucose
management system in the cardiac ICU Performance of the Medtronic Sentrino® continuous glucose
management system in the cardiac ICU
M Kosiborod1, R Gottlieb2, J Sekella2, D Peterman1, A Grodzinsky1,
P Kennedy1, M Borkon1
1Saint Luke’s Mid America Heart Institute, Kansas City, MO, USA; 2Medtronic
Minimed, Northridge, CA, USA
Critical Care 2013, 17(Suppl 2):P462 (doi: 10.1186/cc12400) 1. Lazar HL, et al.: Ann Thorac Surg 2009, 87:663-669. P460 P460
Root cause analysis of hypoglycemic events in critically ill patients
A McDonald, J Krinsley
Stamford Hospital, Stamford, CT, USA
Critical Care 2013, 17(Suppl 2):P460 (doi: 10.1186/cc12398) Introduction Hypoglycemia (H) is a complication of intensive insulin
(IN) therapy of critically ill patients and is independently associated S172 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
htt //
f
/
l
t /17/S2 http://ccforum.com/supplements/17/S2 Figure 1 (abstract P461). Arterial sensor ISO-modifi ed Bland–Altman plot. Figure 1 (abstract P461). Arterial sensor ISO-modifi ed Bland–Altman plot. Figure 1 (abstract P461). Arterial sensor ISO-modifi ed Bland–Altman plot. percent (202/202) of arterial sensor measurements met ISO 15197
criteria (within =/–20% of reference measurements if BG ≥4.2 mmol/l;
Figure 1). was 12.2%. No diff erences in CGM system accuracy were seen within
subgroups of low versus high Society of Thoracic Surgeons (STS)
score (MARD 12.1% and 10.6% for STS >8% vs. ≤8%, respectively)
or hemodynamic status (MARD 12.0% and 12.4% for compromised
vs. stable hemodynamics). Consensus grid analysis showed >99%
of SG values within A/B zones, and 0% in D/E zones. No device or
study-related adverse events were reported. In total, 80% and 100%
of clinicians found Sentrino® easy to use after one and two patients,
respectively. Conclusion The GluCath System measured glucose concentration
continuously in cardiac surgery ICU patients. Arterial catheter
deployment did not appear to compromise line function or patient
care. Percutaneous venous deployment was feasible, but may be
associated with risk of local venous thrombosis. p
y
Conclusion The Sentrino® CGM system demonstrated good analytic
and clinically relevant accuracy, excellent reliability and safety in
critically ill cardiac patients; and was easy to use and integrate in the
cardiac ICU. Future studies are needed to determine whether CGM can
improve BG control and reduce hypoglycemia in this patient group. Reference P463
Intensive insulin therapy in critically ill children: impact on blood
glucose dynamics and its relation with mortality
M Van Tornout, M Gielen, T Van Herpe, D Vlasselaers, L Desmet,
I Vanhorebeek, G Van den Berghe, D Mesotten
University Hospitals Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P463 (doi: 10.1186/cc12401) Introduction Blood glucose (BG) control reduces morbidity and length
of stay, and is standard practice in patients undergoing cardiac surgery
[1]. However, maintaining BG in the target range, while avoiding
hypoglycemia, is challenging. Continuous glucose monitoring (CGM)
is a promising technology that may help address these challenges. We investigated the performance and safety of Medtronic Sentrino®, a
newly developed CGM for critically ill adults, in the cardiac ICU. P464 P464 Implementation of evidence-based protocols improves survival:
a 15-year surgical ICU experience in 10,172 patients
SA Nasraway
Tufts Medical Center, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P464 (doi: 10.1186/cc12402) Tufts Medical Center, Boston, MA, USA Table 1 (abstract P465)
Admitted
Admitted
with
without
decubitus
decubitus
P
Variable
(n = 180)
(n = 2,543)
value
Age 65 or older
106 (58.9%)
1,062 (41.8%)
<0.001
APACHE IV score on admission
80.2 (28.2%)
58.7 (30.7)
<0.001
Diabetes
53 (29.4%)
420 (16.5%)
<0.001
Dialysis
16 (8.9%)
58 (2.3%)
<0.001
Hepatic failure
3 (1.7%)
56 (2.2%)
1.000
Mechanically ventilated during
123 (68.3%)
1,457 (57.3%)
hospitalization
Admission source to the ICU
Direct admit
10 (5.6%)
154 (6.1%)
ER
84 (46.7%)
1,206 (47.4%)
Floor
25 (13.9%)
214 (8.4%)
0.017
OR
19 (10.6%)
459 (18.1%)
Other location
42 (23.3%)
510 (20.1%)
Outcomes
ICU LOS (days)
4.9 (4.9)
3.3 (4.7)
<0.001
Hospital LOS (days)
15.6 (20.7)
10.5 (11.1)
<0.001
ICU mortality
30 (16.7%)
308 (12.1%)
0.073
Hospital mortality
58 (32.2%)
464 (18.3%)
<0.001
Readmission rate for
20 out of 112
173 out of 2,079
0.002
survivors
(16.4%)
(8.3%)
Conclusion The presence of decubiti on admission to the ICU is
associated with longer hospitalizations even after adjusting for age,
acuity, and organ supportive therapies. DU on admission to ICU provide
a unique, unambiguous marker of increased resource utilization. Introduction There has been enormous interest in measuring
ICU performance in terms of mortality and resource use, owing to
increased public and health insurer scrutiny. We elected to describe the
performance of our surgical ICU, using a standardized mortality ratio
(SMR = observed / predicted mortality). y
Methods The primary cohort was all patients admitted to the surgical
ICU from March 2010 through February 2012 and related outcomes. The change in SMR was longitudinally determined from the latest 15-
year study period (1997 to 2011) comprised of 10,172 patients. y
y
Results There were 1,799 ICU admissions in the primary cohort. Hospital
mortality, observed and predicted by APACHE IV, was measured. Crude
hospital mortality was 8.4%. The hospital SMR (observed / predicted
mortality) was 0.58 (95% CI: 0.49 to 0.65). The SMR decreased by 20%
from 0.73 to 0.58 over the 15-year study period, an absolute 1% per year
decrease (P = 0.039; 95% CI: –0.02 to –0.002), as shown in Figure 1. (
;
),
g
Conclusion Mortality was less than predicted and steadily declined
during the previous 15 years. Intensive insulin therapy in critically ill children: impact on blood
glucose dynamics and its relation with mortality Introduction A large RCT showed that tight glucose control (TGC),
targeting age-adjusted normal fasting blood glucose levels with insulin
infusion, decreased morbidity and mortality in critically children [1]. However, the incidence of hypoglycemia increased substantially in the
TGC group. We aimed to assess the eff ect of TGC on the three domains
of blood glucose dynamics (hyperglycemia, hypoglycemia and blood
glucose variability) and their independent association with mortality
in the pediatric ICU. Methods Adult patients with actual or planned cardiac ICU admission
at a single tertiary center were approached for participation and
signed consent. Other inclusion criteria were treatment with i.v. insulin
(target BG <140 mg/dl) and life expectancy >96 hours. After initiation
of i.v. insulin, Sentrino® subcutaneous glucose sensors were inserted
into patients’ anterior thighs with planned study participation of 72
to 96 hours. Reference BG was collected according to ICU protocol,
obtained from central venous catheter and analyzed with bedside
blood gas analyzer (BGA; i-STAT®, Abbott, USA). Sensor glucose
(SG) results were displayed, and its predictive alerts and alarms fully
enabled. Additional reference BGs were obtained during alarms and
calibration. All treatment decisions were based on BGA data, not on SG
values. p
Methods This is a preplanned substudy of a published RCT in one
10-bed pediatric ICU. Seven hundred patients (age 1 to 16 years),
admitted to the PICU between October 2004 and December 2007, were
randomized to either TGC (50 to 80 mg/dl in infants, 70 to 100 mg/dl in
children) or to the usual care tolerating hyperglycemia up to 215 mg/
dl (UC). Patients with at least two arterial blood glucose measurements
were included (UC n = 349; TGC n = 348). Results A total of 21 patients were enrolled; all successfully completed
the study. Mean age was 65 years, 38% were women, 67% had diabetes. Types of surgery were CABG (38%), valve replacement (29%), combined
CABG and valve (19%) and cardiac transplant (14%). SG was displayed
95% of the time during the study, and 864 paired BG–SG points were
used for analysis. Overall mean absolute relative diff erence (MARD) Results Mean blood glucose levels were lowered from 143 ± 34 mg/dl
in the UC group to 104 ± 24 mg/dl (P <0.0001). The median number
of samples per patient did not diff er between UC (21 (13 to 48)) and
TGC (22 (15 to 49)). Decubitus ulcers are associated with prolonged length of stay in
critically ill patients Decubitus ulcers are associated with prolonged length of stay in
critically ill patients Decubitus ulcers are associated with prolonged length of sta
critically ill patients
WT McGee1, BH Nathanson2, E Lederman1, TL Higgins1
1Baystate Medical Center, Springfi eld, MA, USA; 2OptiStatim LLC,
Longmeadow, MA, USA
Critical Care 2013, 17(Suppl 2):P465 (doi: 10.1186/cc12403) y
WT McGee1, BH Nathanson2, E Lederman1, TL Higgins1
1Baystate Medical Center, Springfi eld, MA, USA; 2OptiStatim LLC,
Longmeadow, MA, USA
Critical Care 2013, 17(Suppl 2):P465 (doi: 10.1186/cc12403) WT McGee1, BH Nathanson2, E Lederman1, TL Higgins1
1Baystate Medical Center, Springfi eld, MA, USA; 2OptiStatim LLC,
Longmeadow, MA, USA Introduction How decubitus ulcers (DU) diagnosed on admission
to the ICU aff ect resource use is unknown. We hypothesize that this
easily identifi able clinical fi nding would be associated with longer
hospitalizations. Methods Clinical assessment was used to identify patients with DU on
admission to the ICU. The association of DU on hospital and ICU length
of stay (LOS) was assessed with generalized linear models that adjusted
for ICU admission source, age, APACHE IV score, diabetes, sepsis,
hepatic failure, dialysis and mechanical ventilation. i
Conclusion TGC substantially reduced the hyperglycemic index, at the
expense of increased hypoglycemia incidence, but without aff ecting
blood glucose variability. Statistical analysis suggested strictly avoiding
hyperglycemia in order to bring about outcome benefi t with insulin in
critically ill children. p
y
Results DU was present on admission in 180 of 2,723 (6.6%) during a
19-month period in 2011 to 2012 at a single teaching hospital in New
England. Patients with decubiti were sicker on admission (mean (SD)
APACHE IV 80.2 (28.2) vs. 58.7 (30.7), P <0.001), more likely to be age 65
or older (58.9% vs. 41.8%, P <0.001) and require more organ supportive
therapies (Table 1). DU patients had a higher in-hospital mortality rate
(32.2% vs. 18.3%, P <0.001) and longer mean ICU and hospital LOS
(4.9 vs. 3.3 days and 15.6 vs. 10.5 days, respectively (P <0.001)). After
multivariate adjustment, DU patients stayed 3.2 days, 95% CI (1.6,
4.8) longer in the hospital than non-DU patients and stayed 2.5 days
longer, 95% CI (0.7, 0.43) among survivors. DU patients had longer ICU
stays (1.3 days, 95% CI (0.3, 1.3)) in general but were statistically similar
among survivors. Reference 1. Vlasselaers D, et al.: Lancet 2009, 373:547-556. 1. Vlasselaers D, et al.: Lancet 2009, 373:547-556. Intensive insulin therapy in critically ill children: impact on blood
glucose dynamics and its relation with mortality TGC lowered the hyperglycemic index, a marker of S173 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 hyperglycemia over time, from median 44 (IQR 29 to 66) mg/dl in UC to
12 (7 to 23) mg/dl, as well as the glycemic penalty index, an aggregate
measure of dysglycemia, from median 44 (32 to 58) in UC to 22 (16
to 30) (both P <0.0001). Despite frequent hypoglycemia <40 mg/dl
(1.4% in UC, 25.0% in TGC, P <0.0001), TGC did not result in increased
blood glucose variability, as refl ected by the standard deviation of
the blood glucose time series (TGC: 36 (24 to 50) mg/dl, UC 34 (26
to 48) mg/dl, P = 0.82). In multivariable logistic regression analysis,
adjusting for baseline risk factors (age, severity of illness, malignancy,
type of admission), hypoglycemia (OR = 1.6, 95% CI = 0.19 to 2.22) and
blood glucose variability (OR = 1.0, 95% CI = 0.97 to 1.02, per mg/dl)
were not associated with PICU mortality. In contrast, hyperglycemic
index (OR = 1.02, 95% CI = 1.01 to 1.04, per mg/dl) was independently
associated with mortality and its lowering statistically explained the
outcome benefi t of the intervention. P464 Mortality predictors in septic HIV patients in the ICU
P Vidal-Cortés1, P Lameiro-Flores1, A Aller-Fernández2, M Mourelo-Fariña2,
P Fernández-Ugidos1, R Gómez-López1, M Alves-Pérez1, E Rodríguez-García1
1CHU Ourense, Spain; 2CHU A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P468 (doi: 10.1186/cc12406) Introduction Sepsis in HIV patients carries a high mortality. Our
purpose is to identify mortality predictors in this population. Introduction Sepsis in HIV patients carries a high mortality. Our
purpose is to identify mortality predictors in this population. Methods A retrospective study of HIV patients admitted to our
ICU between January 2005 and December 2009. We identify septic
patients and analyzed demographic factors, etiology, organ failure
and outcome. We used logistic regression to calculate the relative
mortality risk for each qualitative variable and the beta (β) coeffi cient
for quantitative variables. Conclusion Depending on the patient and the cause of admission,
ICU admission may represent an excellent opportunity as a screening
method to determine HIV status. Given the greater effi cacy of HAART
at present, most patients with medical or surgical conditions unrelated
to HIV infection will be eligible to join the ICU. People with HIV can and
should benefi t from using reasonable and individualized care in an ICU. References q
Results Sixty-two septic patients were admitted to our ICU during
the study period (71% men, mean age: 40.59 ± 8.12). A total of
56.5% came from the ED, mean hospital stay before ICU admission:
6.14 ± 10.96 days. A total of 66.1% had a history of intravenous drug
use, other comorbidities: COPD (9.7%), cirrhosis (8.1%), solid or
hematologic malignancy (12.9%). A total of 40.3% were under HAART. Mean CD4 count at admission: 219.62 ± 353.93 cells/mm3. Mean viral
load: 4.57 ± 3.25 log. Mean albumin levels: 2.3 ± 0.53 g/dl. APACHE
II: 21.98 ± 7.97. The lung was the most frequent focus of infection
(65.6%) followed by the CNS. The most common pathogen was
S. pneumoniae (28.8%), followed by P. jirovecii (13.6%). In total, 62.9%
needed vasopressors, 79% mechanical ventilation and 19.4% renal
replacement. Mean ICU and hospital length of stay: 10.43 ± 10.52 and
34.76 ± 29.64 days. ICU mortality: 33.9%; hospital mortality: 41.9%. P464 The SMR can be used to track success
when new quality measures are introduced or changes in the delivery
of care are made. ICUs should report their standardized mortality ratios
to evaluate performance. Figure 1 (abstract P464). Tufts surgical ICU, 15-year outcome
standardized hospital mortality ratio. Conclusion The presence of decubiti on admission to the ICU is
associated with longer hospitalizations even after adjusting for age,
acuity, and organ supportive therapies. DU on admission to ICU provide
a unique, unambiguous marker of increased resource utilization. Figure 1 (abstract P464). Tufts surgical ICU, 15-year outcome
standardized hospital mortality ratio. Figure 1 (abstract P464). Tufts surgical ICU, 15-year outcome
standardized hospital mortality ratio. S174 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P466
HIV patients admitted to an ICU of a university hospital –
experience of 15 years: 1995 to 2009
A Agrifoglio, M Arce, M Hernández, P Millan, C Guallar, I Pozuelo,
M Jiménez
Hospital Universitario La Paz, Madrid, Spain
Critical Care 2013, 17(Suppl 2):P466 (doi: 10.1186/cc12404) Results We included 139 patients with HIV/AIDS admitted to the
ICU. Median age was 40 (31 to 48) years and 71% were male. Severe
malnutrition was common (34%). The CD4 cell count was 84 (25 to 274)
cells/mm3 and viral load was 17,733 (67 to 174,214) copies/ml; 57% had
at least one opportunistic infection; 55% had used antiretroviral therapy
previous to ICU admission. Mechanical ventilation was used by 46% of
patients and hospital mortality was 42%. Total SOFA score was 5 (2 to
9) points. Cardiovascular dysfunction was the most common on the
fi rst day of stay (51%), followed by respiratory (42%), neurological (40%),
renal (35%), hematological (27%) and hepatic (17%). Cardiovascular and
renal dysfunctions presented with higher rate of severe dysfunction
(30% and 15%, respectively). Rates of neurological (P = 0.002), renal
(P = 0.009) and hematological (P = 0.003) dysfunctions were higher
in nonsurvivors. Age, CD4 cell count, malnutrition, and opportunistic
infections were included in the multivariate analysis. Neurological
dysfunction was the independent risk factor for hospital mortality
(odds 3.2 (1.4 to 7.2)). The presence of neurological dysfunction was
dichotomized: associated or not with primary neurological diagnosis;
survival was lower in the patients with neurological dysfunction and
without primary neurological diagnosis (log-rank test 0.001 in the
7-day and 0.02 in the 28-day analysis). P464 Sixty-day survival was similar
in primary and secondary neurological dysfunction, but it remained
lower than in patients without neurological impairment. P466 Introduction HIV infection is a major public health problem in the
world. The use of prophylaxis against opportunist infection and
the introduction of HAART in 1996 increased life expectancies. The
therapeutic use of ICU resources for HIV patients has been controversial,
questioning the admission of these patients especially in advanced
stages of the disease, given the poor prognosis. The aim of this study
was to determine the experience of the past 15 years in relation to the
income of these patients in an ICU. Methods A retrospective case series consisting of patients with
diagnosis of HIV infection (known or unknown) admitted between
January 1995 and December 2009. We collected demographic and
epidemiological data, process of acquisition of the disease, infection
status: known or unknown patient infected, whether or not receiving
antiretroviral therapy and whether it was eff ective (undetectable viral
load at the time of admission), APACHE II, cause of admission, need
for mechanical ventilation (MV), pathology related or unrelated to HIV
infection and ICU mortality. Conclusion Neurological dysfunction was independently associated
with hospital survival, mainly in those AIDS critically patients without
primary neuropathy. y
Results During this period 12,607 patients were admitted to the
ICU, 188 (1.5%) HIV-positive. Mean APACHE II score 17.6, median age
39 years, 73% men and 90% Spanish nationality. Principal risk behavior:
addiction drugs injection (67%). Seventeen percent of patients did
not know who was infected with HIV at the time of admission to the
ICU. Fifty-three percent were not receiving HAART. Of the 88 patients
treated, 93% were receiving HAART (eff ective in 88% of cases). Sixty
percent of the patients came from the emergency department of the
hospital. Main admission diagnoses: acute respiratory failure caused
by infection (Streptococcus pneumoniae and Pneumocystis jirovecii),
neurological disorders (coma for illicit drugs and psychotropic) and
septic shock. Seventy percent required MV. Of patients whose HIV
infection was not known, 93.5% were admitted for related pathology. In patients of known infection, the pathology associated with HIV was
30%. Average length of stay 9 was days. ICU mortality was 35%. Most
frequent causes of death: septic shock and multiple organ failure. P464 ICU
mortality increases for each day of hospital stay before ICU admission
(β = 1.062, 95% CI = 1.009 to 1.118, P = 0.022), each point of: cardiac
frequency at admission (β = 1.059, 95% CI = 1.024 to 1.096, P = 0.001),
FiO2 (β = 1.034, 95% CI = 1.011 to 1.057, P = 0.003), pCO2 (per mmHg)
(β = 1.052, 95% CI = 1.007 to 1.098, P = 0.023); and decreases for each
point of: albumin (g/dl) (β = –0.307, 95% CI = 0.095 to 0.986, P = 0.047),
mean arterial pressure (mmHg) (β = 0.951, 95% CI = 0.904 to 0.999,
P = 0.048), pH (β = 0.002, 95% CI = 0.0 to 0.172, P = 0.006), fi rst ICU
24-hour diuresis (ml) (β = 0.999, 95% CI = 0.998 to 1.000, P = 0.004) and
base excess (β = 0.912, 95% CI = 0.836 to 0.995, P = 0.039). Mortality
increases threefold (β = 3.134, 95% CI = 1.051 to 9.345, P = 0.04) if
patients do not come from the ED, almost 10-fold if the patients
need vasopressors for more than 24 hours (β = 9.975, 95% CI = 2.054
to 48.451, P = 0.004) and fi vefold if patients need renal replacement
(β = 5.692, 95% CI = 1.466 to 22.099, P = 0.012). 1. Huang L, et al.: N Engl J Med 2006, 355:173-181. 1. Huang L, et al.: N Engl J Med 2006, 355:173-181. g
g
2. Akgun K, et al.: J Intensive Care Med 2011, 26:151-164. 2. Akgun K, et al.: J Intensive Care Med 2011, 26:151-164. 2. Akgun K, et al.: J Intensive Care Med 2011, 26:151-164. 2.
Akgun K, et al.: J Intensive Care Med 2011, 26:151-164. Prevalence and prognosis of organ dysfunctions of AIDS critically ill
patients AJ Japiassu, RA Amâncio, D Medeiros, EM Mesquita, EM Motta, FA Bozza
Oswaldo Cruz Foundation, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P467 (doi: 10.1186/cc12405) AJ Japiassu, RA Amâncio, D Medeiros, EM Mesquita, EM Motta, FA Bozza
Oswaldo Cruz Foundation, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P467 (doi: 10.1186/cc12405) Introduction The aim was to analyze the prognosis of AIDS patients
with organ dysfunctions at ICU admission. Introduction The aim was to analyze the prognosis of AIDS patients
with organ dysfunctions at ICU admission. y
Methods A prospective cohort study, including all patients with HIV/
AIDS diagnosis, who were admitted to a specialized ICU from November
2009 until May 2012. Patients with less than 24 hours of ICU stay were
excluded. Demographics and nutritional status were collected. The
organ dysfunctions were classifi ed according to the SOFA score, and
categorized as absent (0 SOFA point), mild (1 to 2 points) and severe (3 to
4 points). We expressed numeric variables as median and interquartile
interval (25% to 75%). We performed a multivariate analysis of possible
variables associated with hospital mortality (P <0.2), and we explored
the 7-day, 28-day and 60-day survival of patients with and without
independent risk factors. S175 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Conclusion Neither immune status-related variables nor comorbidity
or infection focus are mortality predictors. Poor nutritional status,
delayed ICU admission, shock or renal failure increase the ICU relative
mortality risk. Tachycardia, hypotension, hypercapnia, acidosis, and
oliguria in the fi rst ICU 24 hours increase signifi cantly ICU mortality. Mechanical ventilation is not a mortality predictor. Methods A retrospective study was conducted in consecutive medical
and surgical older patients with cancer admitted to a university hospital
ICU in São Paulo, between 2007 and 2012. Univariate and multivariate
analysis were performed to identify associated and independent
factors related to hospital mortality. p
y
Results From 8,976 patients with cancer requiring ICU at the period,
600 patients were 80 years old or higher. Most patients were male
(53%), had solid neoplasm (86%), were from medical admission (54%)
and 37% had metastatic disease. The mean age was 84 years (± 3). The ICU and hospital mortality rates were 30% and 44%, respectively. Identifying risk factors for readmission to the ICU: a qualitative
approach Results From 597 patients included in the study, 477 were medical
admissions (79.8%) and 120 were surgical admissions (20.2%). Four
hundred and twenty (70%) patients had metastasis, 222 patients (37%)
required the ICU because of respiratory failure and 121 (20%) because
of septic shock. The ICU and hospital mortality rates were 53.9% and
64%, respectively. In the univariate analysis, variables associated
with hospital mortality were diagnosis of nonsmall-cell lung cancer,
higher Charlson morbidity index, medical admission, active neoplasm,
vasopressor need at admission to and at 24 hours of ICU, acute renal
failure at admission, non-invasive ventilation or mechanical ventilation
need at admission to and at 24 hours of ICU and a higher admission
arterial lactate. By multivariate analysis, risk factors of hospital mortality
were diagnosis of nonsmall-cell lung cancer (OR = 3.32; 95% CI, 2.08
to 5.29, P <0.001), medical admission (OR = 2.89; 95% CI, 1.75 to 4.77,
P <0.001), acute renal failure at admission (OR = 4.09; 95% CI, 1.49 to
11.21, P <0.001), non-invasive ventilation at 24 hours of ICU (OR = 2.50;
95% CI, 1.48 to 4.23, P = 0.001) and mechanical ventilation at 24 hours
of ICU (OR = 3.68; 95% CI, 1.87 to 7.26, P <0.001). DG Strange, K Tjelle, A Lindhardt, K Jensen Introduction Readmission to the ICU within 48 hours is an indicator of
quality of intensive care and is associated with an increase in mortality. During the last years several groups have published data based on
multivariate logistic regression analysis to describe characteristics of
patients who needed readmission to the ICU. Older age, comorbid
conditions and severity of illness (APACHE score) have been among
the strongest predictors for readmission. In our ICU most patients are
in the groups formerly identifi ed as risk groups, which means that
stratifi cation and prediction of readmission is diffi cult. Because of the
unusual high severity of acute and pre-existing illnesses we could
not fi nd a data match on comparable patient groups. To investigate
whether we could reduce our rate of readmission we therefore decided
to perform a qualitative investigation to identify risk factors related to
readmission. After identifi cation of the risk factors we will take actions
to optimize care and perform ongoing control of the implemented
actions to secure that they decreases the rate of readmission. Conclusion Hospital survival in patients with lung cancer requiring
ICU admission was 36%. P469 Outcome of intensive care treatment for lung cancer patients
L Hajjar1, F Galas2, D Goto2, A Tavares2, M Santos2, A Freitas2, T Oliveira2,
P Tonini2, L Cavichio2, M Bazan2, J Almeida2, M Cavalcante2, J Fukushima2,
S Vieira2, E Osawa2
1Heart Institute, São Paulo, Brazil; 2Cancer Institute of University of São Paulo,
Brazil
Critical Care 2013, 17(Suppl 2):P469 (doi: 10.1186/cc12407) Introduction Patients with lung cancer commonly require the ICU
for a variety of acute illnesses related to the underlying malignancy,
treatment, or comorbid conditions. ICU admission of patients with
nonresectable lung cancer has been criticized based on the high
mortality rate in this population. However, recent advances in critical
care may have changed this scenario. The aim of this study was to
identify factors associated with hospital mortality in this group of
patients. Conclusion Hospital survival in older patients with cancer requiring
ICU admission is acceptable. Our results provide evidence that ICU
management may be appropriate in older patients with cancer and
appoint risk factors for mortality, helping to better triage cancer
patients who will benefi t from ICU care. Methods A retrospective study was conducted in consecutive medical
and surgical patients with lung cancer admitted to a university hospital
ICU in São Paulo, between 2007 and 2012. A univariate analysis was
performed to identify associated variables with hospital mortality. Selected variables were included in the multivariate model. Prevalence and prognosis of organ dysfunctions of AIDS critically ill
patients In the univariate analysis, variables associated with hospital mortality
were diagnosis of lung cancer, medical admission, active neoplasm,
vasopressor need at 24 hours of ICU, acute renal failure at admission,
mechanical ventilation need at admission to and at 24 hours of ICU and
a higher admission arterial lactate. By multivariate analysis, risk factors
of hospital mortality were diagnosis of lung cancer (OR = 2.32; 95% CI,
1.08 to 3.29, P <0.001), medical admission (OR = 6.473; 95% CI, 2.76 to
15.15, P <0.001), acute renal failure at admission (OR = 3.09; 95% CI, 1.49
to 9.21, P <0.001), mechanical ventilation at 24 hours of ICU (OR = 5.68;
95% CI, 2.87 to 7.26, P <0.001) and lactate levels at admission (OR = 1.02;
95% CI, 1.006 to 1.051, P <0.001). Identifying risk factors for readmission to the ICU: a qualitative
approach Our results provide evidence that ICU
management may be appropriate in patients with nonresectable lung
cancer and appoint risk factors for mortality, helping to better triage
cancer patients who will benefi t from ICU care. Methods Retrospective data on patients readmitted to the ICU within
48 hours during an 8-month period (November 2011 to June 2012) were
drawn from the Critical Information System (CIS) at ICU ZIT, Bispebjerg
Hospital, Denmark. ZIT is a multidisciplinary unit with 10 beds and 500
to 600 admissions/year and a median SAPS II score of 47. A group of
consultants, junior doctors and nurses from the ICU and the ward each
read the patient fi les with focus on pattern recognition and suggested
trigger points to focus on. Data on trigger points were then drawn from
the CIS system and re-evaluated. Finally, fi ve key points were identifi ed
and serves as basis for future actions. Complementarity of prognosis models SAPS 3 and EuroSCORE in
cardiac surgery Discrimination
was assessed using the area under the ROC curve. With the standardized
mortality ratio (SMR) we evaluated the agreement between predicted
and observed mortality. We used multiple logistic regression for
multivariate analysis Introduction The aim of this study is to evaluate the performance of
SAPS 3 and EuroSCORE and whether there is complementarity between
them in ICU patients admitted after cardiac surgery. Methods An observational, prospective and multicenter study of
patients included in the ARIAM registry of adult cardiac surgery. We
analyzed clinical variables, surgery data and postoperative compli-
cations, outcomes and scores (SAPS 3 and EuroSCORE). Discrimination
was assessed using the area under the ROC curve. With the standardized
mortality ratio (SMR) we evaluated the agreement between predicted
and observed mortality. We used multiple logistic regression for
multivariate analysis. y
Results A total of 5,642 patients were included, admitted to any fi ve
hospitals, consecutively between 2008 and 2012. Mean age was
63.62 ± 12.80 years. Surgery room mortality was 1.4%, ICU mortality
was 7.9% and in-hospital mortality was 11.2%. SAPS 3 was calculated
in 5,383 patients. SAPS 3 was 40.91 ± 10.46, and the expected rate of
mortality was 11.09% (with the southwestern Europe equation), with
an observed hospital mortality of 9.6%, SMR = 0.87 (0.79 to 0.94). With
the general equation, the expected mortality was 10.47%, SMR = 0.917
(0.843 to 0.996). Discrimination of SAPS 3 was evaluated using the area
under the ROC curve, it was 0.77 (0.75 to 0.79). The 30-day mortality was
9.6%, with an expected 30-day mortality predicted by EuroSCORE of
7.86% (SMR = 1.22; CI = 1.12 to 1.32). The area under the ROC curve was
0.734 for EuroSCORE (0.712 to 0.755). Multivariate analysis with logistic
regression showed complementarity between SAPS 3 and logistic
EuroSCORE (vers. I). There is a relationship between hospital mortality
and the probability of dying predicted by SAPS 3 (general equation),
OR 1.05 (1.04 to 1.06), and logistic EuroSCORE (vers. I), OR 1.024 (1.015
to 1.032). The model with both variables has a similar discrimination to
SAPS 3 alone. The area under the ROC curve of the combined model
was 0.778 (0.758 to 0.799) and 0.77 (0.75 to 0.79) using SAPS 3 only. Reference 1. Breslow MJ, Badawi O: Severity scoring in the critically ill: part 1 –
interpretation and accuracy of outcome prediction scoring systems. Chest
2012, 141:245-252. P474 g
Methods A prospective registry involving patients who underwent LDLT
in the period from October 2004 to December 2010. Postoperative 30-
day clinical and laboratory data included: clinical BP, HR, temperature, ICU
stay, liver functions AST, ALT, GGT, ALP, TotPtn, Alb, TBil, DBil, INR, PTT, CRP,
kidney functions Crea, Na, K, blood picture Hb, Hct, TLC and Plt count. Complementarity of prognosis models SAPS 3 and EuroSCORE in
cardiac surgery
MJ Delgado-Amaya1, E Curiel-Balsera1, MD Arias-Verdu1,
E Castillo-Lorente2, FJ Carrero-Gómez3, E Aguayo-DeHoyos4,
A Herruzo-Avilés5
1Hospital Regional Carlos Haya, Málaga, Spain; 2Complejo Hospitalario
de Jaén, Spain 3Puerta del Mar Hospital, Cádiz, Spain; 4Virgen de las Nieves
Hospital, Granada, Spain; 5Virgen del Rocio Hospital, Seville, Spain
Critical Care 2013, 17(Suppl 2):P474 (doi: 10.1186/cc12412) Are APACHE II scores better predictors of mortality than routine
laboratory values? y
Z Baykara, H Özocak, A Kuş, Z Arslan, B Yüksel, C Aksu, M Ertagın, M Solak,
K Toker
University of Kocaeli, Turkey
Critical Care 2013, 17(Suppl 2):P473 (doi: 10.1186/cc12411) Introduction Even though more accurate and updated versions such
as APACHE III and IV have been developed, APACHE II has remained the
most widely used severity scoring system in the ICU [1]. In the present
study, we aimed to develop a model mainly based on the laboratory
data available upon admission to predict mortality and to compare its
performance with that of APACHE II in a mixed ICU in Turkey. g
y
Conclusion There is complementarity between the SAPS 3 model and
EuroSCORE. However, a joint model with both prognosis scores has no
appreciable improvement in discrimination, compared with the SAPS
3 model. In our study, SAPS 3 shows very useful to rule out that the
superior observed mortality (compared with the mortality predicted
by EuroSCORE) is not attributable to inadequate care of patients in the
ICU, the observed mortality being even slightly below the predicted
mortality for patients undergoing cardiac surgery at the reference
hospitals in which the SAPS 3 system was developed. Methods A total of 645 adult patients who were over 18 years old
and who stayed in the ICU for more than 24 hours were included in
this study. Using stepwise multivariate logistic regression analysis, a
mortality prediction model was developed based on the laboratory
data, diagnostic category and age. The adjusted probability of death,
according to the diagnostic category of the APACHE II score (adj-APACHE
II), was calculated. The ability of the laboratory model and the adj-
APACHE II model to discriminate between survivors and nonsurvivors
was assessed using receiver operating characteristic (ROC) curves. The
calibration of the laboratory model and the adj-APACHE II model was
assessed by the Hosmer and Lemeshow goodness-of-fi t test. Complementarity of prognosis models SAPS 3 and EuroSCORE in
cardiac surgery p
Conclusion Predictors of mortality were INR >3.06 (accuracy 87.5%)
followed by PTT >53.3 (79.2%), HR >94 (76.8%), ICU stay >8 days
(76.2%), Na >142.9 (72.9%), TBil >15 (71.3%), ALT >379 (67.5%), TProt
<4.6 (63.5%), AST >145 (57.4%), Hct <24.4 (54.2%), CRP >12 (Acc 51.6%)
and GGT >22 (51.4%). Complementarity of prognosis models SAPS 3 and EuroSCORE in
cardiac surgery g
y
MJ Delgado-Amaya1, E Curiel-Balsera1, MD Arias-Verdu1, MJ Delgado-Amaya1, E Curiel-Balsera1, MD Arias-Verdu1,
E Castillo-Lorente2, FJ Carrero-Gómez3, E Aguayo-DeHoyos4,
A Herruzo-Avilés5 y
Results Our study involved 142 recipients, 124 (87%) males and 18
(13%) females, who underwent living donor liver transplantation;
mean age was 53 ± 14.8 years, ranging from 24 to 66 years; 29 cases
(20.43%) at Kasr Al Aini University Hospital and 113 cases (79.57%) at
the International Medical Center. The 30-day mortality rate was 13.38%. Clinical and laboratory parameters showed gradual resolution towards
normal values in survivors or deterioration in nonsurvivors, allowing
one to predict mortality at various cutoff points. Clinical parameters: HR
>94 cutoff (Acc 76.8%, Sens 89.5%, Spec 62.6%, PPV 27, NPV 97) and ICU
stay >8 days (Acc 76.2%, Sens 75.3%, Spec 95.7%, PPV 95.7, NPV 35.3). Laboratory parameters: INR >3.06 cutoff (Acc 87.5%, Sens 92.9%, Spec
78.6%, PPV 96.3, NPV 64.7), followed by PTT >53.3 (Acc 79.2%, Sens
78.6%, Spec 75%, PPV 93.6, NPV 42.9), Na >142.9 (Acc 72.9%, Sens 80%,
Spec 72.3%, PPV 26.7, NPV 96.6), TBil >15 (Acc 71.3%, Sens 95.5%, Spec
50%, PPV 92.4, NPV 63.6), ALT >379 (Acc 67.5%, Sens 72.2%, Spec 73.8%,
PPV 28.9, NPV 94.7), TProt <4.6 (Acc 63.5%, Sens 55.6%, Spec 73.3%, PPV
24.4, NPV 91.4), AST >145 (Acc 57.4%, Sens 50%, Spec 84.4%, PPV 32.1,
NPV 92), Hct <24.4 (Acc 54.2%, Sens 47.4%, Spec 81.5%, PPV 34.6, NPV
88.2), CRP >12 (Acc 51.6%, Sens 43.7%, Spec 73.3%, PPV 90.5, NPV 17.7),
GGT >22 (Acc 51.4%, Sens 83.3%, Spec 36.9%, PPV 16.3, NPV 93.7). 1Hospital Regional Carlos Haya, Málaga, Spain; 2Complejo Hospitalario
de Jaén, Spain 3Puerta del Mar Hospital, Cádiz, Spain; 4Virgen de las Nieves
Hospital, Granada, Spain; 5Virgen del Rocio Hospital, Seville, Spain
Critical Care 2013, 17(Suppl 2):P474 (doi: 10.1186/cc12412) 1Hospital Regional Carlos Haya, Málaga, Spain; 2Complejo Hospitalario
de Jaén, Spain 3Puerta del Mar Hospital, Cádiz, Spain; 4Virgen de las Nieves
Hospital, Granada, Spain; 5Virgen del Rocio Hospital, Seville, Spain
Critical Care 2013, 17(Suppl 2):P474 (doi: 10.1186/cc12412) Introduction The aim of this study is to evaluate the performance of
SAPS 3 and EuroSCORE and whether there is complementarity between
them in ICU patients admitted after cardiac surgery. Methods An observational, prospective and multicenter study of
patients included in the ARIAM registry of adult cardiac surgery. We
analyzed clinical variables, surgery data and postoperative compli-
cations, outcomes and scores (SAPS 3 and EuroSCORE). P470 P470
Outcome of older patients with cancer requiring ICU admission
L Hajjar1, M Franca Cavalcante2, F Galas2, J T Fukushima2, M R Sundin2,
F Bergamin2, C Park2, H Palomba2, D M Goto2, A Tavares2, M Santos2,
A Freitas2, P Tonini2, L Cavichio2, J P Almeida2, M Bazan2
1Heart Institute, São Paulo, Brazil; 2Cancer Institute of University of São Paulo,
Brazil
Critical Care 2013, 17(Suppl 2):P470 (doi: 10.1186/cc12408) Results In a qualitative analysis, readmissions to the ICU are related
to the following fi ve key points – discharge outside day hours, lack of
infection control, stay in ICU <2 days, lung physiotherapy ordinated but
not eff ectuated, and several minor organ dysfunctions (atrial fi brillation
and acute kidney injury). Age, diagnosis, SAPS II score or ventilator
treatment during intensive care was not diff erent in patients with
successful discharge and patients readmitted in this group of patients. Conclusion It is possible and suitable to identify key points for future
eff orts in a given subgroup of patients using a systematic qualitative
approach. Critical Care 2013, 17(Suppl 2):P470 (doi: 10.1186/cc12408) Introduction Because the prognosis of older patients with cancer may
be poor compared with younger patients, it remains controversial
whether they benefi t from ICU treatment. The objective of this study
was to identify factors associated with hospital mortality in older
patients with cancer requiring the ICU. S176 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 P472 P472
Clinical and laboratory postoperative impact on outcome of living
donor liver transplantation recipients
Y Nassar, K Omar, A Battah, A Mwafy
Cairo University, Giza, Egypt
Critical Care 2013, 17(Suppl 2):P472 (doi: 10.1186/cc12410) 2.01 (P = 0.98) for the laboratory model and 13.2 (P = 0.10) for the adj-
APACHE II model. Conclusion If the aim is to predict mortality as accurately as APACHE
II, the mortality prediction model based mainly on routine admission
laboratory tests can achieve this using computer technology, without
labor-related costs, as soon as the patient is admitted to the ICU. However, it is necessary to perform multicenter validation studies. Reference Introduction Adult recipients of living donor liver transplantation
have a wide range of postoperative medical comorbidities altering
various clinical and laboratory parameters. We aimed to assess 30-
day postoperative clinical and laboratory changes and their value in
predicting outcome of LDLT recipients. P477 Eldicus scores for admission to our ICU: a retrospective study
G Zani, B Gemelli, S Vitali, C Villani, R Ragazzi, S Spadaro, CA Volta, R Alvisi
University of Ferrara, Italy
Critical Care 2013, 17(Suppl 2):P477 (doi: 10.1186/cc12415) Results A total of 273 charts were reviewed. In total, 242 were
categorized into A (n = 126), B (n = 79) or C (n = 37). D (n = 31) consisted
mainly of patients with hematological malignancies (n = 12) and
patients with chemotherapy or immunosuppressive treatment
(n = 14). The groups diff ered in length of stay with A< B< C. During the
fi rst 3 days the SOFA score was higher in A compared with C and in
B compared with C. The duration of antibiotic therapy was longer in
both B and C compared with A. There were no diff erences in 28-day
mortality (A: 34/126 = 27%, B: 30/79 = 38%, C: 15/37 = 40%); however,
the proportions of patients dying between days 8 and 28 were higher
in B (14/63 = 22%) and C (7/29 = 24%) compared with A (5/95 = 5%). Introduction The indication for ICU admission is poorly defi ned and
identifying patients who may benefi t from it is extremely diffi cult. As
reported in literature, the Eldicus scores require further validation in
centers in diff erent countries and with diff erent patient populations [1]. Previous studies have demonstrated that patients with intermediate
severities of illness are those that benefi t from ICU treatment; instead,
patients who are too well or too sick should not be admitted. There are
many unmeasurable factors that infl uence the triage decision and can
infl uence the subsequent outcome. The Eldicus scores could be the fi rst
triage decision rule. The aim of this study was to confi rm the ability of
the Eldicus scores to discriminate the survival status in our general ICU. Methods We retrospectively enrolled all patients >18 years admitted to
the Ferrara ICU from January to June 2012. Patients’ data were recorded
using Prosafe international data collection software and consulting
the individual medical records. Data were chosen based on variables
used in the Eldicus scores. As indicated in the Eldicus algorithm, for
each patient the initial refusal score was calculated, to evaluate the
too severe patients. Reference Results A total of 481 patients aged 80 or over were admitted between
1 January 2001 and 31 December 2010, representing 9.6% of the
admissions during this time with minimal annual change. Admission
age ranged from 80 to 96 years (median 83) with a median APACHE II
score of 20 (6 to 48, IQR 16 to 25). In total, 318 (66.1%) survived to critical
care discharge and 256 (53.2%) survived to hospital discharge; 228
(89.0%) went home rather than to a care facility. Those under 80 years
demonstrated 80.6% critical care survival, 65.5% hospital survival and a
median APACHE II score of 17 (0 to 51, IQR 13 to 23). Eighty patients died
within the 22-month follow-up period with an average life expectancy
of 7.4 months. None reached life expectancy (average curtailment of
5.6 years compared with age-matched and sex-matched population
statistics). Neither of these fi gures changed signifi cantly through the 1. Sprung CL, et al.: Crit Care Med 2012, 40:125-131. 1. Sprung CL, et al.: Crit Care Med 2012, 40:125-131. Comparing intensive care treated primary, secondary and tertiary
sepsis with respect to length of stay, organ failure and mortality:
a pilot study Conclusion Older people represent a growing proportion of the
population although their representation in the critical care population
remained constant in this 10-year study. These patients had a slightly
higher median APACHE II score and 14.5% greater critical care mortality
than the younger patients. The majority of survivors were able to
go home; however, 31% died within 22 months with signifi cant life
expectancy curtailment, surviving on average only 7.4 months after
discharge; this has not changed in the last 10 years. Those who survived
this initial period (69%) had a much better outlook. This information
may be vital to patients and physicians when discussing admission to
critical care. p
p
y
Methods The Swedish ICU registry was used to identify all adult
patients diagnosed with severe sepsis or septic shock in a single
Swedish ICU during 2006 to 2011. The charts were reviewed and
patients categorized into four groups based on whether the patient
had suff ered systemic infl ammatory-inducing insults previous to the
septic episode. The patients were categorized as: (A) primary sepsis,
no recent (<30 days) trauma/infection, ongoing systemic infl ammatory
or immunosuppressive disease or malnutrition; (B) secondary sepsis,
recent (<7 days) trauma/infection or systemic infl ammatory disease,
but no immunosuppressive disease or malnutrition; (C) tertiary
sepsis, >1 fulfi lled criteria of secondary sepsis during the last 10 days
or immunosuppressive disease or malnutrition; and (D) patients not
attributable to A to C. After the categorization, physiological and
laboratory parameters during the ICU stay as well as therapy and
outcome were extracted from the charts. Only signifi cant diff erences
(P <0.05) are presented; Mann–Whitney and chi-squared tests were
used. P477 Then we counted the fi nal triage score, to discover
whether some patients were too healthy to be admitted to the ICU. Correlation between the severity of the scores and patients’ outcome
was estimated using the Wilcoxon’s rank-sum test. Conclusion In this retrospective material it was possible to categorize
88.6% of all patients as having primary, secondary or tertiary sepsis. The categories diff ered in clinical picture at presentation as well as in
outcome. A prospective study is warranted to validate the results of this
study. P476 Comparing intensive care treated primary, secondary and tertiary
sepsis with respect to length of stay, organ failure and mortality:
a pilot study yi
Results The areas under the ROC curves of the laboratory models and
the adj-APACHE II scores for the prediction of mortality were 0.80 (95%
CI: 0.76 to 0.85) and 0.78 (95% CI: 0.74 to 0.83), respectively (P >0.05, z
statistic). The Hosmer–Lemeshow statistic had a chi-squared value of y
S Castegren1, M Jonasson1, J Sjölin2, M Castegren1
1Centre for Clinical Research Sörmland, Uppsala University, Uppsala, Sweden;
2Medical Sciences, Uppsala University, Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P475 (doi: 10.1186/cc12413) S Castegren1, M Jonasson1, J Sjölin2, M Castegren1
1Centre for Clinical Research Sörmland, Uppsala University, Uppsala, Sweden;
2Medical Sciences, Uppsala University, Uppsala, Sweden
Critical Care 2013, 17(Suppl 2):P475 (doi: 10.1186/cc12413) S177 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 decade. However, 10-year follow-up on 2001/2002 admissions showed
that, despite this early mortality eff ect, the average post-discharge
lifespan was 5.1 years with 2.2 years shortfall in life expectancy. Conclusion Older people represent a growing proportion of the
population although their representation in the critical care population
remained constant in this 10-year study. These patients had a slightly
higher median APACHE II score and 14.5% greater critical care mortality
than the younger patients. The majority of survivors were able to
go home; however, 31% died within 22 months with signifi cant life
expectancy curtailment, surviving on average only 7.4 months after
discharge; this has not changed in the last 10 years. Those who survived
this initial period (69%) had a much better outlook. This information
may be vital to patients and physicians when discussing admission to
critical care. decade. However, 10-year follow-up on 2001/2002 admissions showed
that, despite this early mortality eff ect, the average post-discharge
lifespan was 5.1 years with 2.2 years shortfall in life expectancy. Introduction It may be hypothesized that a patient’s infl ammatory
status might aff ect the clinical picture and outcome of a subsequent
septic insult. To test this hypothesis, patients were categorized
according to previous infl ammatory activity and analyzed with respect
to clinical picture and outcome in a retrospective study. Introduction It may be hypothesized that a patient’s infl ammatory
status might aff ect the clinical picture and outcome of a subsequent
septic insult. To test this hypothesis, patients were categorized
according to previous infl ammatory activity and analyzed with respect
to clinical picture and outcome in a retrospective study. Reference 1. Offi ce of National Statistics Population Data [http://www.ons.gov.uk/ons/
taxonomy/index.html?Nscl=Population] 1. Offi ce of National Statistics Population Data [http://www.ons.gov.uk/ons/
taxonomy/index.html?Nscl=Population] P476
Survival of older patients after critical care
D Ashton-Cleary, R Sinclair
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P476 (doi: 10.1186/cc12414) Survival of older patients after critical care
D Ashton-Cleary, R Sinclair
Royal Cornwall Hospital, Truro, UK
Critical Care 2013, 17(Suppl 2):P476 (doi: 10.1186/cc12414) g
Results Ninety-eight patients were fi nally enrolled. Data are presented
as median ± SD. The SAPS II score was 33 ± 13 and was similar to those
reported in the literature [1]. The Karnofsky performance status scale
was 69 ± 20. The Eldicus initial score was 56 ± 30. The fi nal score was
70 ± 48. Seventy-eight percent of the analyzed patients were alive at
hospital discharge. A signifi cant correlation between the severity scores
and in-hospital outcome was found (P <0.001). Limitations of our study
were the low sample size and the fact that patients not admitted to the
ICU were not considered. Introduction We aimed to assess the survival of older patients after
admission to critical care compared with the population in general. Introduction We aimed to assess the survival of older patients after
admission to critical care compared with the population in general. p
admission to critical care compared with the population in general. Methods We reviewed admission data for a 12-bed critical care
unit in a district general hospital in the UK between 2001 and 2010. We calculated critical care and hospital discharge survival rates and
studied the trend in these through the decade. We searched hospital
records to establish whether these patients were still alive at November
2012 to determine life-expectancy (minimum 22-month follow-up on
all patients). This was compared with UK Offi ce of National Statistics
data for the local population [1]. Discharge destination (care facility or
home address) was recorded as a surrogate marker of change in quality
of life post discharge. Conclusion The Eldicus scores appear to be useful in identifying
patients that may benefi t more from ICU admission. Further studies
should confi rm the ability of these scores in helping identify those
patients who really benefi t from ICU admission. Reference Prediction of sequences of organ dysfunction in critical patients
studied with Bayesian analysis M Sandri1, P Berchialla2, D Gregori3, L Gottin1, V Sweiger1, E Polati1,
RA De Blasi4 1University of Verona, Italy; 2University of Torino, Italy; 3University of Padova,
Italy; 4University of Rome ‘La Sapienza’, Rome, Italy
Critical Care 2013, 17(Suppl 2):P479 (doi: 10.1186/cc12417) p
p
g
p
p
Results Lower age, NT-Pro-BNP and TnI were signifi cantly associated
with 1-year survival. We built a neural network model using the training
database. We used the test database to compare our model with the
IGS 2 score. The diff erence between the AUC of the biomarker score
and the IGS 2 score was statistically signifi cant (Figure 1). Introduction The multiorgan dysfunction syndrome (MODS) is a
dynamic process involving simultaneously or consecutively two
or more organ systems [1]. The organ dysfunction’s degree can be
assessed by three severity scores (SOFA [2], MODS [3], LODS [4]), but
they have some limitations: they do not allow the evaluation of the
clinical course of a patient, they are not reliable in populations diff erent
from the reference one, and they do not support clinicians’ decisions. Because MODS implies a systemic infl ammatory reaction leading to
microcirculatory dysfunction, our hypothesis was that organ failures
follow a predictable sequence of appearance. Our aims were to verify
the presence of more likely organ failure sequences and to assess an
online method to predict the evolution of MODS in a patient. The high
mortality and morbidity rate of MODS in ICUs can in fact be reduced
only by a prompt and well-timed treatment [5]. y
gi
g
Conclusion Our new neural network model built using age, NT-PRO-
BNP and TnI succeeded the internal-validation processes and predicts
1-year mortality better than IGS 2. P481
Signifi cance of preadmission quality of life for mortality in the ICU:
a prospective cohort study
RB Bukan1, A Moeller1, M Henning1, K Mortensen2, T Klausen1, T Waldau1
1Herlev University Hospital, Herlev, Denmark; 2University of Copenhagen,
Denmark
Critical Care 2013, 17(Suppl 2):P481 (doi: 10.1186/cc12419)
Figure 1 (abstract P480). Comparison of area under ROC curves (AUC) of
the biomarker score and IGS 2. Figure 1 (abstract P480). Comparison of area under ROC curves (AUC) of
the biomarker score and IGS 2. Methods We selected 73 patients consecutively admitted to the ICU of
Sant’Andrea Hospital from January to June 2012. The inclusion criteria
were at least two organ systems with SOFA ≥2, ICU length of stay
>48 hours. Developing a screening instrument for predicting psychological
morbidity after critical illness
Ö A Schandl1, M Bottai2, E Hellgren1, Ö Sundin3, P Sackey1
1Karolinska University Hospital Solna, Stockholm, Sweden; 2Karolinska
Institutet, Stockholm, Sweden; 3Mid Sweden University, Östersund, Sweden
Critical Care 2013, 17(Suppl 2):P478 (doi: 10.1186/cc12416) Introduction The purpose was to develop a predictive screening
instrument for use at ICU discharge to identify patients at risk for post-
traumatic stress, anxiety or depression. Introduction The purpose was to develop a predictive screening
instrument for use at ICU discharge to identify patients at risk for post-
traumatic stress, anxiety or depression. S178 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods Potential risk factors for psychological problems were
prospectively collected at ICU discharge. Two months after ICU
discharge 252 ICU survivors received the questionnaires Post-
Traumatic Stress Symptom scale-10 (PTSS-10) and Hospital Anxiety
and Depression Scale (HADS) to estimate the degree of post-traumatic
stress, anxiety and depression. Sepsis-Related Problems of the European Society of Intensive Care
Medicine. Intensive Care Med 1996, 22:707-710. Marshall JC, et al.: Multiple organ dysfunction score: a reliable descriptor of
a complex clinical outcome. Crit Care Med 1995, 23:1638-1652. Le Gall JR, et al.: The logistic organ dysfunction system: a new way to assess
organ dysfunction in the intensive care unit. JAMA 1996, 276:802-810. Kilica YA, et al.: Visualizing multiple organ failure: a method for analyzing
temporal and dynamic relations between failing systems and
interventions. Crit Care 2007, 11:417. y
Results Of the 150 responders, 31% had adverse psychological
outcome, defi ned as PTSS-10 >35 and/or HADS subscales ≥8. After
analysis, six predictors with weighted risk scores were included in the
screening instrument: major pre-existing disease, being a parent to
children younger than 18 years of age, previous psychological problems,
in-ICU agitation, being unemployed or sick-listed at ICU admission and
appearing depressed in the ICU. Each predictor corresponded to a
given risk score. The total risk score, the sum of individual risk scores,
was related to the probability for adverse psychological outcome in the
individual patient. The predictive accuracy of the screening instrument,
as assessed with area under the receiver operating curve, was 0.77. When categorizing patients in three risk probability groups – low (0 to
29%), moderate (30 to 59%) and high (60 to 100%) risk – the actual
prevalence of adverse psychological outcome in respective groups was
12%, 50% and 63%. 6. Prediction of sequences of organ dysfunction in critical patients
studied with Bayesian analysis For each patient we calculated the SOFA since the beginning
of the inclusion criteria and daily for 8 days. For the statistical analysis
we used Dynamic Bayesian Networks (DBNs) [6]. DBNs were applied to
model SOFA changes in order to identify the most probable sequences
of organs failures in a patient who experienced a fi rst known failure. gi
Results We created a DBN for the analysis of MODS studying the
relations between organ failures at diff erent times. The DBN was made
so that each organ failure is dependent on the previous one. We also
considered a corrective factor to take account that not all patients
completed the observation. Using software (GeNie) we obtained the
probabilities of the organ failure sequences. Figure 1 (abstract P480). Comparison of area under ROC curves (AUC) of
the biomarker score and IGS 2. Figure 1 (abstract P480). Comparison of area under ROC curves (AUC) of
the biomarker score and IGS 2. g
Conclusion The use of DBNs, although with our limited set of data,
allowed us to identify the most likely organ dysfunction sequences
associated with a fi rst known one. Capability to predict these sequences
in a patient makes DBNs a promising prognostic tool for physicians in
order to treat patients in a timely manner, or to test a treatment effi cacy. References Developing a screening instrument for predicting psychological
morbidity after critical illness
Ö Murphy K, et al.: Modelling Gene Expression Data using Dynamic Bayesian
Networks. Technical report. University of California; 1999. Comparison of IGS 2 score and a new neural network biomarker
score performance in predicting 1-year mortality Comparison of IGS 2 score and a new neural network biomarker
score performance in predicting 1-year mortality
C Romdhani, I Labbene, M Belhadj Amor, Z Hajjej, A Massoudi, M Ferjani
Military Hospital of Tunis, Tunisia
Critical Care 2013, 17(Suppl 2):P480 (doi: 10.1186/cc12418) C Romdhani, I Labbene, M Belhadj Amor, Z Hajjej, A Massoudi, M Ferjani
Military Hospital of Tunis, Tunisia
Critical Care 2013, 17(Suppl 2):P480 (doi: 10.1186/cc12418) Introduction The aim of this study was to compare the IGS 2 score with
a predictive model built using clinical characteristics and biomarkers
associated with 1-year mortality from procalcitonin, C-reactive protein,
troponin I (TnI) and NT-Pro-BNP. Conclusion The preliminary screening instrument may aid ICU clinicians
in identifying patients at risk for adverse psychological outcome after
critical illness. Prior to wider clinical use, external validation is needed. Methods We conducted a prospective observational cohort study
in a polyvalent ICU. We calculated the IGS 2 severity score in the fi rst
24 hours. We dosed NT-Pro-BNP, TnI, procalcitonin and C-reactive
protein at admission to the ICU. To test the biomarker model, the patient
database was divided into training (70%) and test (30%) databases. A
neural network model was trained from variables associated with
1-year mortality. All statistics were done using R v.2.15.1. We used the
pROC package to plot and compare ROC curves.i P481
Signifi cance of preadmission quality of life for mortality in the ICU:
a prospective cohort study
RB Bukan1, A Moeller1, M Henning1, K Mortensen2, T Klausen1, T Waldau1
1Herlev University Hospital, Herlev, Denmark; 2University of Copenhagen,
Denmark
Critical Care 2013, 17(Suppl 2):P481 (doi: 10.1186/cc12419) Predicting physical function and mental health in trauma intensive
care patients 2 years after hospitalisation Predicting physical function and mental health in trauma intensive
care patients 2 years after hospitalisation
L Aitken1, B Macfarlane1, W Chaboyer2, M Schuetz1, C Joyce1, AG Barnett3
1Princess Alexandra Hospital, Brisbane, Australia; 2Griffi th University,
GoldCoast, Australia; 3Queensland University of Technology, Brisbane,
Australia
Critical Care 2013, 17(Suppl 2):P482 (doi: 10.1186/cc12420) gi
Results We present the results of an 18-month period, 9 months prior
to and 9 months after the introduction of the SwissDRG. Data of 1,491
and 1,492 patients were analyzed, respectively. When all patients were
included, we found prior to and after the introduction of the DRGs a
comparable length of stay on the ICU (mean ± SD of 52.1 ± 2.2 hours vs. 50.82 ± 2.2 hours), no diff erence in the severity of illness at discharge
detected by the SAPS II (mean ± SD of 27.9 ± 0.3 vs. 28.4 ± 0.3) and
the number of readmissions (91 vs. 92). There was also no signifi cant
diff erence when only percentiles 6 to 94 were included or when the
three subgroups were analyzed. p
y
p
L Aitken1, B Macfarlane1, W Chaboyer2, M Schuetz1, C Joyce1, AG Barnett3
1Princess Alexandra Hospital, Brisbane, Australia; 2Griffi th University,
GoldCoast, Australia; 3Queensland University of Technology, Brisbane,
Australia Critical Care 2013, 17(Suppl 2):P482 (doi: 10.1186/cc12420) Introduction Long-term compromise after traumatic injury is
signifi cant; however, few modifi able factors that infl uence outcome
have been identifi ed. The aim of this study was to identify acute and
early post-acute predictors of long-term recovery amenable to change
through intervention. g
p
y
Conclusion Up to now, the introduction of the SwissDRG has no
infl uence on patients’ discharge characteristics from a large ICU. Data
assessment will continue and further data analysis has to be performed. Methods Adults (>17 years) admitted to the ICU, Princess Alexandra
Hospital, Australia following injury were prospectively followed. Data were collected on demographics, pre-injury health, injury
characteristics and acute care factors. Psychosocial measures (self-
effi cacy (SE), illness perception (IP), post-traumatic stress disorder
(PTSD) symptoms and psychological distress) and health status (SF-36)
were collected via questionnaire 1, 6, 12, and 24 months post injury. Outcomes of interest were the Physical Function (PF) and Mental Health
(MH) subscales of the SF-36. Regression models were used to estimate
predictors of physical function and mental health over a 2-year period. Do ICU patients’ discharge characteristics change with Swiss
Diagnosis Related Groups? Do ICU patients’ discharge characteristics change with Swiss
Diagnosis Related Groups? Do ICU patients’ discharge characteristics change with Swiss
Diagnosis Related Groups? G Rauchenstein-Ziltener, CA Rüst, P Fodor, S Blumenthal
Tiemli City Hospital, Zurich, Switzerland
Critical Care 2013, 17(Suppl 2):P483 (doi: 10.1186/cc12421) g
G Rauchenstein-Ziltener, CA Rüst, P Fodor, S Blumenthal
Tiemli City Hospital, Zurich, Switzerland
Critical Care 2013, 17(Suppl 2):P483 (doi: 10.1186/cc12421) Results Preliminary results, based upon data from 175 included patients,
reveal an ICU mortality of 12.6% and 30-day mortality of 22.9%. No
patients were lost to follow-up. When the Physical Component of SF-36
(PCS) was used as an estimate of preadmission quality of life, the area
under the curve for model B (AUC = 0.80) was comparable with model E
(AUC = 0.81), and better than model A (AUC = 0.85). The General Health
item of SF-36 (GH), used as an estimate of preadmission quality of life,
gave an AUC = 0.76 (model D). All models were controlled for sex and
age. Introduction Swiss Diagnosis Related Groups (SwissDRG) have been
eff ective since 1 January 2012. The infl uence of this new system on
patients’ discharge characteristics from a large ICU is not known. With
the introduction of the DRG we expect patients to be discharged after
a shorter length of stay on the ICU and with higher severity of illness. Methods The ICU of the City Hospital Triemli in Zurich has an inter-
disciplinary organization with surgical and internal medical patients,
with a maximum occupancy of 18 beds and a center function for the
surrounding hospitals. In this ongoing prospective observational
study, we collect and analyze the anonymized data of all patients
discharged from our ICU prior to and after the introduction of the
SwissDRG. The primary endpoint was the length of stay on the ICU
in hours. The secondary endpoints were the severity of illness of the
patients at the time of discharge, detected by the scoring system SAPS
II as well as measured by the number of readmissions to the ICU. Initially
all patients were analyzed and in a second step only patients within
percentiles 6 to 94 were considered. We also analyzed the subgroups of
patients referred internally, patients sent back to referring hospital and
patients regionalized to a homebase hospital. The statistics have been
done with SPSS and P <0.05 was considered signifi cant. P481i P481
Signifi cance of preadmission quality of life for mortality in the ICU:
a prospective cohort study
RB Bukan1, A Moeller1, M Henning1, K Mortensen2, T Klausen1, T Waldau1
1Herlev University Hospital, Herlev, Denmark; 2University of Copenhagen,
Denmark
Critical Care 2013, 17(Suppl 2):P481 (doi: 10.1186/cc12419) Signifi cance of preadmission quality of life for mortality in the ICU:
a prospective cohort study p
p
y
RB Bukan1, A Moeller1, M Henning1, K Mortensen2, T Klausen1, T Waldau1
1Herlev University Hospital, Herlev, Denmark; 2University of Copenhagen,
Denmark 1. Levy MM, et al.: 2001 SCCM/ESICM/ACCP/ATS/SIS International Sepsis
Defi nitions Conference. Intensive Care Med 2003, 29:530-538. Levy MM, et al.: 2001 SCCM/ESICM/ACCP/ATS/SIS International Sei 2. Vincent JL, et al.: The SOFA (Sepsis-Related Organ Failure Assessment) score
to describe organ dysfunction/failure. On behalf of the Working Group on Critical Care 2013, 17(Suppl 2):P481 (doi: 10.1186/cc12419) S179 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Introduction Assessing whether a critically ill patient should be
admitted to an ICU remains diffi cult and mortality amongst ICU
patients is high. To render intensive care with no prospect of success
is an immense emotional burden for both patient and relatives, and
a great socioeconomic burden for society as well. Therefore, validated
strategies that can help identify patients who will benefi t from
intensive care are in demand. This study seeks to investigate whether
preadmission quality of life can act as a predictor of mortality amongst
patients admitted to the ICU. Predictors of PF included IP (β = –1.5, 95% CI = –3.1 to –1.1, P <0.01), SE
(β = 1.8, 95% CI = 1.3 to 2.6, P <0.01), hospital length of stay (β = –1.7,
95% CI = –2.0 to –0.8, P <0.01), never having been married (β = 1.8, 95%
CI = 0.3 to 5.5, P = 0.03), and having injury insurance (β = –2.7, 95%
CI = –6.9 to –1.9, P <0.01). Predictors of MH included PTSD symptoms
(β = –2.4, 95% CI –3.4 to –1.4, P <0.01), psychological distress (β = –6.9,
95% CI = –8.9 to –5.2, P <0.01), SE (β = 0.6, 95% CI = 0.2 to 1.1, P <0.01),
and unemployment (β = –2.3, 95% CI = –5.0 to –0.2, P = 0.04). P481i Predictors of PF included IP (β = –1.5, 95% CI = –3.1 to –1.1, P <0.01), SE
(β = 1.8, 95% CI = 1.3 to 2.6, P <0.01), hospital length of stay (β = –1.7,
95% CI = –2.0 to –0.8, P <0.01), never having been married (β = 1.8, 95%
CI = 0.3 to 5.5, P = 0.03), and having injury insurance (β = –2.7, 95%
CI = –6.9 to –1.9, P <0.01). Predictors of MH included PTSD symptoms
(β = –2.4, 95% CI –3.4 to –1.4, P <0.01), psychological distress (β = –6.9,
95% CI = –8.9 to –5.2, P <0.01), SE (β = 0.6, 95% CI = 0.2 to 1.1, P <0.01),
and unemployment (β = –2.3, 95% CI = –5.0 to –0.2, P = 0.04). p y
β
Conclusion Trauma ICU patients experience compromised physical
function and mental health 24 months after injury. Psychological
distress, self-effi cacy and illness perception infl uence outcomes and are
potentially amenable to change in response to interventions initiated
during hospital stay. p
Methods All patients (>18 years) admitted to the ICU for more than
24 hours are included. In order to assess preadmission quality of life,
the patient or close relatives complete the Short-Form 36 (SF-36)
within 72 hours after ICU admission. Mortality is evaluated from ICU
admission until 30 days hereafter. Logistic regression and receiver
operating characteristic analyses are employed to assess predictive
value for mortality using fi ve models: SF-36 Physical Component
Summary (PCS) and APACHE II (model A), SF-36 PCS (model B), SF-36
General Health (GH) and APACHE II (model C), SF-36 GH (model D),
and APACHE II (model E). Classifi cation tables are composed in order
to assess sensitivity, specifi city, positive and negative predictive values
and likelihood ratios. Do ICU patients’ discharge characteristics change with Swiss
Diagnosis Related Groups? Conclusion Preliminary results indicate that the SF-36 GH and the
SF-36 PCS are as good as APACHE II to predict mortality 30 days after
ICU admission. However, in order to conclude whether preadmission
quality of life can contribute to triage, by successfully identifying
patients suitable for intensive care, fi nal analyses, due in 2013, are
awaited. These results will clarify whether future randomized controlled
trials, in which preadmission quality of life acts as a supplement to
triage, are justifi able. P484
Do Diagnosis Related Groups change admission practice to a large
Swiss ICU?
K Homburg, CA Rüst, P Fodor, S Blumenthal
Triemli City Hospital, Zurich, Switzerland
Critical Care 2013, 17(Suppl 2):P484 (doi: 10.1186/cc12422) P484
Do Diagnosis Related Groups change admission practice to a large
Swiss ICU? K Homburg, CA Rüst, P Fodor, S Blumenthal
Triemli City Hospital, Zurich, Switzerland
Critical Care 2013, 17(Suppl 2):P484 (doi: 10.1186/cc12422) Predicting physical function and mental health in trauma intensive
care patients 2 years after hospitalisation A subject-specifi c intercept in a mixed model was used to account for
repeated data from participants over time. P484 Do Diagnosis Related Groups change admission practice to a large
Swiss ICU? Introduction The new Swiss fi nancing system by means of Diagnosis
Related Groups (DRG), eff ective 1 January 2012, forms the basis of a
performance-based system to reimburse general hospital services. There are only few data on the infl uence of DRG on ICU patients [1,2]. We expect that the introduction of the DRG in Switzerland will change
the number of admissions from external hospitals to a large ICU with
a centre function and will infl uence the severity of disease of the
admitted patients. Introduction The new Swiss fi nancing system by means of Diagnosis
Related Groups (DRG), eff ective 1 January 2012, forms the basis of a
performance-based system to reimburse general hospital services. There are only few data on the infl uence of DRG on ICU patients [1,2]. We expect that the introduction of the DRG in Switzerland will change
the number of admissions from external hospitals to a large ICU with
a centre function and will infl uence the severity of disease of the
admitted patients. p
p
p
Results Participants (n = 123) were young (median 37, IQR 22 to
55 years), predominantly male (83%) and spent on average 3 days
in the ICU and 3 weeks in hospital. Response rates were over 55% at
each follow-up, with responders similar to nonresponders except for
being generally older. PF and MH scores improved over time, although
the averages remained below the Australian norms at 24 months. S180 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Methods The ICU of the Triemli City Hospital in Zurich has an inter-
disciplinary organisation with surgical and internal medical patients,
with a maximum occupancy of 18 beds and a centre function for
the surrounding hospitals of the region. In this prospective ongoing
observational study, we collect and analyse the anonymised data of all
patients admitted to our ICU from an external hospital during 12 months
prior to (1 January to 31 December 2011) and after (1 January to 31
December 2012) the introduction of the DRG in Switzerland. Exclusion
criteria are admissions by the emergency department, self-assignments
into the hospital and internal relocations. The primary endpoint is the
number of admissions from an external hospital to our ICU. References References
1. McLaughlin AM, et al.: Intensive Care Med 2009, 35:2135-2140. 2. Rossi C, et al.: Intensive Care Med 2006, 32:545-552. References
1. McLaughlin AM, et al.: Intensive Care Med 2009, 35:2135-2140. 2. Rossi C, et al.: Intensive Care Med 2006, 32:545-552. P486 Variable cost of a Greek ICU E Tsigou1, D Karabatsou1, E Boutzouka1, V Psallida1, G Baltopoulos1,
M Tsironi2
1A. Anargiroi Hospital, Athens, Greece; 2University of Peloponnisos, Sparti,
Greece
Critical Care 2013, 17(Suppl 2):P485 (doi: 10.1186/cc12423) Critical Care 2013, 17(Suppl 2):P485 (doi: 10.1186/cc12423) Introduction The ICU consumes almost 20% of total hospital resources,
despite accounting for less than 10% of hospital beds. Medical
cost can be divided into a fi xed part, consisting of personnel and
accommodation expenses, and a variable part, determined by the
needs of each patient (medications, consumables, examinations). In
order to rationalize medical cost, estimation of expenses must be real
and individualized. The aim of this study was to assess the variable cost
of critically ill patients in a new, seven-bed, adult, general ICU, using
bottom-up costing methodology. Conclusion The latest audit follows introduction of a referral system
directly to the ICU consultant and may account for the reduction in
numbers of referrals attended by junior doctors. ED/medicine persist
as the main source of referral to the ICU. Discussion with the referring
team consultant may reduce inappropriate referrals. ICU staffi ng
should not be reduced. Methods All 138 patients who were treated during 2011, and stayed for
at least 24 hours, were included in the study. Data were retrospectively
collected from patient records and included demographics, cause
of admission, APACHE II score at admission, length of stay (LOS) and
outcome. Cost was recorded for every day and for every patient, based
on a hospital-specifi c cost catalogue and on national agreements. Data
are presented as mean ± SEM. Analysis of data was carried out using
Graph Pad Prism 5.0., applying Student’s t test. References 1. Ahmad M, et al.: Chest 1988, 93:176-179. 2. Thomas F, et al.: Chest 1987, 91:418-423. 1. Ahmad M, et al.: Chest 1988, 93:176-179. Results There were 105 occasions when a junior doctor left the unit
in 2011, totalling 118 hours and 38 minutes (5 days), a continuation of
the upward trend from the previous 2 years. In 2012 this reduced to 47
occasions totalling 40 hours and 55 minutes. Most attended referrals
were during the daytime, 54% in 2011 and 68% in 2012. The majority
of referrals originated from the ED/medicine, combined numbers
58% in 2011 and 38% in 2012. In 2011 only 30% of ED referrals had
been discussed with a consultant of the referring team and only 32%
of medical referrals. Just two of the 14 inappropriate ED referrals were
discussed with the referring consultant, and none of the six inappropriate
medical referrals were discussed with a medical consultant. In 2012 just
33% of ED/medical referrals had prior discussion with a consultant from
the referring team. At the point of leaving, the ICU was left without a
doctor on 22 occasions in 2011 and six occasions in the 2012 audit. Most
occasions occurred at either night shift/weekend (86% in 2011/100%
in 2012). Referral resulted in a patient being admitted to the ICU on 20
occasions in 2011 and 12 in 2012. 2. Thomas F, et al.: Chest 1987, 91:418-423. Audit of the medical staffi ng of and referrals to a mixed adult ICU in
a teaching hospital g
p
A Needham, T Lyndham, D O’Neil, J Greiff
University Hospitals of Leicester, UK
Critical Care 2013, 17(Suppl 2):P486 (doi: 10.1186/ Introduction Leicester Royal Infi rmary is a (funded) 15-bed ICU, annual
admission 1,102 patients per year (ICNARC data 2011 to 2012). It houses
a broad range of specialty, has over 1,000 beds and an emergency
department (ED) responsible for 297 admissions over the 2011 to 2012
period. Unit guidelines state that a doctor should be immediately
available to the unit at all times, ideally physically present on the unit. This audit looked in detail at each occasion a junior doctor left the unit,
the timings, reason, and outcome of each episode. Results We present the preliminary data for 10 months (in each case
January to October) before and after the introduction of the DRG. We observed an increase of 9.2% (391 vs. 427 patients) of admissions
to our ICU after the introduction of the DRG. The severity of disease
determined by the SAPS II score is unchanged (mean 26.7 vs. 26.0
points, P = 0.466). The severity of disease determined by the APACHE
II score is signifi cantly lower (15.4 vs. 14 points, P = 0.017). We also
noted that after the introduction of the DRG the patients were earlier
transferred from an external hospital to our ICU (mean time until
transfer 29.9 vs. 18.7 hours), but this value was not signifi cant (P = 0.55). Conclusion Up to now the introduction of the DRG in Switzerland
has had a complex infl uence on the number and the kind of patients
admitted from an external hospital to an ICU with centre function. Data
assessment and analysis will continue. g
p
Methods Between 17 October 2011 and 14 November 2011, and
between 2 July 2012 and 30 July 2012, junior doctors on the ICU
completed a data-capture form after each occasion that they left the
unit for a clinical need. Data collected were matched to the objectives. The 2011 audit results were communicated at the University Hospitals
of Leicester Trust ICU Audit Meeting. A system was introduced so
that daytime referrals were directed to the ICU consultant. The impact
of this intervention was assessed by the 2012 re-audit. Do Diagnosis Related Groups change admission practice to a large
Swiss ICU? Secondary
endpoints are the severity of the disease of the admitted patients,
detected by the scoring systems SAPS II and APACHE II as well as the
length of stay in external hospitals before admission. The statistical
analysis is descriptive. variable cost of intensive care treatment. Among drugs, antibiotics
accounted for the largest part. A medical cause of admission was
correlated with higher costs. variable cost of intensive care treatment. Among drugs, antibiotics
accounted for the largest part. A medical cause of admission was
correlated with higher costs. Impact of the time elapsed between ICU request and actual
admission on mortality and length of stay Introduction The World Bank has warned that the rapid growth of the
world’s urban population can only be accommodated safely if cities
adequately develop key infrastructure, such as the provision of acute
care resources. Yet, even basic descriptive information on urban acute
care supply and demand is extremely limited. We therefore conducted
a pilot assessment across seven diverse urban settings across the world. Methods We selected a convenience sample of seven large cities with
varying geographical and socioeconomic characteristics: Boston, Paris,
Bogota, Recife, Liaocheng, Chennai, and Kumasi. To estimate acute care
supply, we developed an instrument to collect data on acute and critical
care infrastructure. We collected data from municipal authorities and
local research collaborators. We expressed the burden of acute disease
as the number of deaths due to acute illnesses, estimated from the
2008 Global Burden of Disease Study. Results were expressed as acute
care supply and acute deaths per 100,000 population and acute care
supply per 100 acute deaths. Introduction The World Bank has warned that the rapid growth of the
world’s urban population can only be accommodated safely if cities
adequately develop key infrastructure, such as the provision of acute
care resources. Yet, even basic descriptive information on urban acute
care supply and demand is extremely limited. We therefore conducted
a pilot assessment across seven diverse urban settings across the world. Methods We selected a convenience sample of seven large cities with
varying geographical and socioeconomic characteristics: Boston, Paris,
Bogota, Recife, Liaocheng, Chennai, and Kumasi. To estimate acute care
supply, we developed an instrument to collect data on acute and critical
care infrastructure. We collected data from municipal authorities and
local research collaborators. We expressed the burden of acute disease
as the number of deaths due to acute illnesses, estimated from the
2008 Global Burden of Disease Study. Results were expressed as acute
care supply and acute deaths per 100,000 population and acute care
supply per 100 acute deaths. Results In total, 413 patients were included, 300 of which belonged to
the LWP group (65.4%). For the entire cohort, the mean APACHE II score
was 19 ± 7, the mean age was 52 ± 22 years, and 211 patients were male
(51.1%). The LWP group did not show diff erence in the APACHE II score
(19 ± 7 vs. 18 ± 8, P = 0.13), but was older (55 ± 20 vs. Impact of the time elapsed between ICU request and actual
admission on mortality and length of stay 49 ± 23, P = 0.01). LWP also had a higher incidence of primary bloodstream infection (23.8%
vs. 10.4%, P = 0.01) and catheter-associated urinary tract infection (10.2%
vs. 1.9%, P = 0.01). LWP patients had higher mortality (37.8% vs. 25.9%,
P = 0.02) and longer ICU LOS (21 ± 47 vs. 14 ± 18 days, P = 0.01). Relative
risk for death in the LWP was 1.74 (95% CI: 1.11 to 2.72). Conclusion Despite showing no signifi cant diff erences on APACHE II
scores from the SWP group, patients from the LWP group presented
greater incidence of primary bloodstream infection, catheter-
associated urinary tract infection, higher mortality outcomes and
longer ICU LOS. pp y p
Results The supply of hospital beds varied from 72.4/100,000
population in Kumasi to 245.8/100,000 in Boston. ICU beds with
capacity for invasive mechanical ventilation and intensive nursing
services ranged from 0.4/100,000 in Kumasi to 19/100,000 population
in Boston. The number of ambulances varied 70-fold between cities. The gap between cities widened when demand was estimated based
on disease burden, with a 70-fold diff erence between cities in ICU
beds/acute deaths. In general, most of the data were unavailable from
municipal authorities. P490f P490
Diff erences in severity and resource utilization for medical and
surgical ICU patients
BH Nathanson1, WT McGee2, E Lederman2, TL Higgins2
1OptiStatim LLC, Longmeadow, MA, USA; 2Baystate Medical Center,
Springfi eld, MA, USA
Critical Care 2013, 17(Suppl 2):P490 (doi: 10.1186/cc12428) Impact of the time elapsed between ICU request and actual
admission on mortality and length of stay GM Filho1, TA Silva1, AR Santana1, FB Soares1, LJ Almeida1, LG Godoy1,
TA Rodrigues1, MO Maia2, JA Neto2, AP Amorim3, EB Moura2, FF Amorim1
1Escola Superior de Ciências da Saúde, Brasília, Brazil; 2Unidade de Terapia
Intensiva Adulto do Hospital Santa Luzia, Brasilia, Brazil; 3Liga Academica de
Medicina Intensiva do Distrito Federal (LIGAMI), Brasilia, Brazil
Critical Care 2013, 17(Suppl 2):P487 (doi: 10.1186/cc12425) Results The age of participants (84 men and 54 women, 71 medical and
67 surgical) was 68.75 ± 1.18, APACHE II score was 18.64 ± 0.61, LOS was
18.46 ± 2.54, and mortality was 19%. The majority of the patients (61%)
were mechanically ventilated. The total days of ICU stay were 2,548 and
the total variable cost was €1,460,465 (€573.18 per patient and per day). Cost per patient was subdivided as follows: medication (including drugs,
fl uids, blood products, nutrition): 56.49%, examinations (including
laboratory, microbiological assays and diagnostic procedures):
22.23%, consumables: 21.26%. As for medication cost, the largest part
was comprised of antibiotics (45.69%), followed by blood products
(17.61%) and cardiovascular drugs (12.39%). Costs for medical patients
were signifi cantly higher than those for surgical patients (P <0.0001). Conclusion The total average cost per patient and per day was found
to be €573.18. Medication expenditure was responsible for the highest Introduction Measures to ensure an appropriate early treatment for
critically ill patients result in signifi cant decreases in mortality [1,2]. This
study aims to evaluate the impact of the time elapsed from request
until admission to the ICU on mortality and ICU length of stay (LOS). Methods A retrospective cohort study performed on patients in the
ICU of Hospital Regional de Samambaia over a period of 4 years, from
January 2008 to December 2011. The patients were allocated into two
groups: patients who waited longer than 6 hours, long waiting period S181 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (LWP, n = 300); and patients whose waiting time was equal to or less
than that period, short waiting period (SWP, n = 113). (LWP, n = 300); and patients whose waiting time was equal to or less
than that period, short waiting period (SWP, n = 113). Diff erences in severity and resource utilization for medical and
surgical ICU patients BH Nathanson1, WT McGee2, E Lederman2, TL Higgins2
1OptiStatim LLC, Longmeadow, MA, USA; 2Baystate Medical Center,
Springfi eld, MA, USA Methods Several disasters are studied. The Fukushima Daiichi Nuclear
plant explosion after the Higashinihon earthquake 2011 was medically
reviewed based on the total 30-day stay on-site in addition to several days
around the site. The Chernobyl incident 1986 was inspected 15 years
after the incident. Other nuclear disasters are included. p
gi
Critical Care 2013, 17(Suppl 2):P490 (doi: 10.1186/cc12428) Introduction Medical and surgical patients use the ICU diff erently. Resources may not always be allocated by severity of illness, but by
custom or habit, particularly if diff erent groups administer bed control
and triage. Specialty-specifi c diff erences may exist even when a single
team controls triage. Variability in resource utilization has important
implications for cost-containment and triage. Results Many serious problems were revealed in the medical teams,
which are as follows: inappropriate basic preparedness against large
special disasters, including nuclear disaster; lack of appropriate
education and training for medical teams against nuclear disaster –
that is, most members of Japan DMAT or the disaster medical assistance
team are still laypersons; incorrect standard/rules of Japan DMAT, which
were excessively focused upon cure of the usual type of injury and
planned short period or nearly 48 hours, which should be abandoned;
and insuffi cient consideration to the weak/vulnerable people or CWAP,
children, (pregnant) women, aged people, and the poor people/sicker
patients. Many of them died because of an insuffi cient emergency
transportation system from their contaminated houses or hospital. Methods Patients admitted to a single, closed medical/surgical
ICU with full-time intensivists and unifi ed triage control in a large,
university-affi liated hospital were evaluated during 2011 to 2012. Patients who died in the ICU were excluded. The day of discharge (D/C)
and severity using APACHE IV and its related Acute Physiology Score Figure 1 (abstract P490). y
Conclusion In order to cope with the special disasters, such as NBC
or nuclear, biological and chemical disaster, it is insuffi cient to take
makeshift measures or use cheap tricks. Working out the systematization
of disaster medicine, based upon the academic viewpoints and
philosophy/reliability, is essential to protect the people and the nation. g
References 1. Amorim FF, et al.: J Clin Med Res 2012, 4:410-414. 1. Amorim FF, et al.: J Clin Med Res 2012, 4:410-414. 2. Rivers E, et al.: N Engl J Med 2001, 345:1368-1377. 1. Amorim FF, et al.: J Clin Med Res 2012, 4:410-414. 2. Rivers E, et al.: N Engl J Med 2001, 345:1368-1377. 2. Rivers E, et al.: N Engl J Med 2001, 345:1368-1377. Medical problems: review from the major nuclear incidents Medical problems: review from the major nuclear incidents
Y Haraguchi1, Y Tomoyasu2, H Nishi3, M Hoshino4, M Sakai5, E Hoshino6
1Disaster Medicine Compendium Team, Japan, Tokyo, Japan; 2Shirahigebashi
Hospital, Tokyo, Japan; 3Tokyo Disaster Medical Center, Tokyo, Japan; 4Shisei
Hospital, Saitama, Japan; 5Tokyo Women’s Medical College, Tokyo, Japan;
6Chiba Nursing Association, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P488 (doi: 10.1186/cc12426) Medical problems: review from the major nuclear incidents
Y Haraguchi1, Y Tomoyasu2, H Nishi3, M Hoshino4, M Sakai5, E Hoshino6
1Disaster Medicine Compendium Team, Japan, Tokyo, Japan; 2Shirahigebashi
Hospital, Tokyo, Japan; 3Tokyo Disaster Medical Center, Tokyo, Japan; 4Shisei
Hospital, Saitama, Japan; 5Tokyo Women’s Medical College, Tokyo, Japan;
6Chiba Nursing Association, Chiba, Japan
Critical Care 2013, 17(Suppl 2):P488 (doi: 10.1186/cc12426) Conclusion The provision of acute care services, a key aspect of urban
infrastructure, varied substantially across the seven diverse urban
settings we studied. Furthermore, the local municipal authorities
generally appeared to have little knowledge of their acute care
infrastructure, with implications for future planning and development. Introduction Intensivists are expected to have many roles during
and after a major disaster/catastrophe; that is, triage, intensive care,
education for people, and so forth. The roles of intensivists against
special disaster or nuclear disaster are studied based on actual
experiences. P489 Variation in acute care burden and supply across diverse urban settings
S Murthy1, S Austin2, H Wunsch3, NK Adhikari4, V Karir2, K Rowan5,
ST Jacob6, J Salluh7, F Bozza8, B Du9, Y An10, B Lee2, F Wu2, C Oppong11,
R Venkataraman12, V Velayutham13, D Angus2
1Hospital for Sick Children, Toronto, Canada; 2University of Pittsburgh, PA, USA;
3Columbia University, New York, NY, USA; 4University of Toronto, Canada; 5Intensive
Care National Audit & Research Centre, London, UK; 6University of Washington,
Seattle, WA, USA; 7D’OR Institute for Research and Education, Rio de Janeiro, Brazil;
8Oswaldo Cruz Foundation, Rio de Janeiro, Brazil; 9Peking Union Medical College
Hospital, Beijing, China; 10Peking University People’s Hospital, Beijing, China;
11Komfo Anokye Teaching Hospital, Kumasi, Ghana; 12Apollo Hospital, Chennai,
India; 13Stanley Medical College Hospital, Chennai, India
Critical Care 2013, 17(Suppl 2):P489 (doi: 10.1186/cc12427) Figure 1 (abstract P490). S182 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Introduction The management of emergency medical admissions has
been a subject of recent clinical incidents. There was a high percentage
of patients that were referred to the ICU by staff in training, and 21%
of referrals were made by junior doctors. Consultant physicians had no
knowledge of the case in 57% of referrals. Introduction The management of emergency medical admissions has
been a subject of recent clinical incidents. There was a high percentage
of patients that were referred to the ICU by staff in training, and 21%
of referrals were made by junior doctors. Consultant physicians had no
knowledge of the case in 57% of referrals. (APS) component were calculated daily for the fi rst 7 days. Trend was
assessed across days by Cuzick’s test. y
y
Results A total of 719 surgical and 925 medical patients met inclusion
criteria. In total, 20.2% of surgical and 21.3% of medical patients had an
ICU LOS <1; P = 0.58. Admission severity was correlated with length of
stay, P = 0.014 for both medical and surgical patients. Medical patients
are sicker on admission and D/C from the ICU than surgical patients
(P <0.05) (Figure 1). g
Methods A prospective study of 21 cases of referrals and admissions to
the ICU was conducted at the Glasgow Victoria Infi rmary Hospital from
8 to 21 September 2012. P489 A questionnaire was produced relating to the
referrals, admissions, seniority involvement, cause of referral, and time
of patient review by the ICU consultant after ICU admission. They were
distributed to specialist registrars and the ICU consultants. All data
were electronically recorded into an Excel database. Questionnaires
that were not completely fi lled were further investigated using patient
clinical notes and contact with medical staff . Information that may
identify a patient or clinician was removed from the questionnaire for
confi dentiality purposes. g
Conclusion ICU utilization diff ered by patient type even with unifi ed
triage control within a single unit. Surgical patients were less severely
ill on admission to and D/C from the ICU. A signifi cant percentage of
medical and surgical patients are D/C within 1 day and may be more
effi ciently served in a less resource-intensive environment. The reasons
for the diff erences in ICU utilization for surgical versus medical patients
require clarifi cation and may have implications for both resource
utilization and cost. Results Twenty-one complete questionnaires were collected. Fifty-
seven percent (12/21) of cases involved admission to the ICU. Nine
percent of the cases involved contacting either a specialist registrar
or ICU consultant intensivist for assistance in practical procedures. Of
the patients admitted to the ICU, 33% (4/12) were from medical wards,
33% were admitted from A&E. Consultants were the most common
professionals who referred patients to critical care (48%; 10/21). Fourteen percent of cases (3/21) involved the referral of patients into
ICU by a junior doctor, but only one of the referrals was accepted by
the ICU intensivist. Consultants referred or were aware of the referral
in 71% (15/21) of cases. Of admissions, 58% (7/12) were accepted by
the ICU consultant and the remaining by the specialist registrars. All
accepted were acknowledged by the ICU consultant. After admission
all of the patients were reviewed by the ICU consultant and the time of
review after admission was on average 1 hour 23 minutes (25 minutes
to 3 hours 45 minutes). Educational eff ectiveness of introductory training for the Rapid
Response System in Japan p
y
p
T Kodama1, M Nakagawa2, E Kawamoto3, S Fujiwara4, H Imai3, S Fujitani5,
K Atagi6 y
T Kodama1, M Nakagawa2, E Kawamoto3, S Fujiwara4, H Imai3, S Fujitani5,
K Atagi6 g
1The University of Texas Southwestern Medical Center at Dallas, TX, USA;
2Social Insurance Kinan Hospital, Tanabe-City, Japan; 3Mie University Hospital,
Tsu-City, Japan; 4NHO Ureshino Medical Center, Ureshino-City, Japan;
5Tokyobay UrayasuIchikawa Medical Center, Urayasu-City, Japan; 6Osaka City
General Hospital, Osaka-City, Japan
Critical Care 2013, 17(Suppl 2):P491 (doi: 10.1186/cc12429) 1The University of Texas Southwestern Medical Center at Dallas, TX, USA;
2Social Insurance Kinan Hospital, Tanabe-City, Japan; 3Mie University Hospital,
Tsu-City, Japan; 4NHO Ureshino Medical Center, Ureshino-City, Japan;
5Tokyobay UrayasuIchikawa Medical Center, Urayasu-City, Japan; 6Osaka City
General Hospital, Osaka-City, Japan
Critical Care 2013, 17(Suppl 2):P491 (doi: 10.1186/cc12429) Introduction Interest in safety and clinical outcomes of inpatients
has been growing in Japan, because the 100,000 Lives Campaign was
introduced under the Japanese Patient Safety Act in 2008. In this act,
an introduction of the Rapid Response System (RRS) was one of the
mainstreams to inpatients’ care. However, many Japanese healthcare
providers cannot understand how to achieve the introduction of
the RRS, because there are few who have knowledge of the system. Therefore, we developed a new introductory training course for
the RRS. The educational eff ectiveness was analyzed through the
surveillance questionnaires after the course. Conclusion There is still an issue with junior doctors referring patients
to the ICU without the acknowledgement of consultant physicians,
resulting in unnecessary admissions and decreased time that ICU
trainees spend in the ICU. There are more appropriate ICU admissions
when there is involvement with seniority. Contact with ICU staff to
perform practical procedures outside the ICU and not about admissions
should be explored further. Methods The educational program includes a lecture series con cern-
ing the outline and management methods, introduction of facilities
that have already deployed, small group discussions, and teaching
methods-of-training for the medical emergency team using a simulator. Evaluation was made in the fi ve-point scale by 82 participants (58
physicians, 16 nurses and eight other professions) throughout seven
courses. The questionnaires are: A. understanding of RRS, B. knowledge
acquisition about patient safety, C. expectation for decreasing the
cardiopulmonary arrest by RRS, and D. expectation for decreasing the
psychological burden by RRS. P491 P491
Educational eff ectiveness of introductory training for the Rapid
Response System in Japan
T Kodama1, M Nakagawa2, E Kawamoto3, S Fujiwara4, H Imai3, S Fujitani5,
K Atagi6
1The University of Texas Southwestern Medical Center at Dallas, TX, USA;
2Social Insurance Kinan Hospital, Tanabe-City, Japan; 3Mie University Hospital,
Tsu-City, Japan; 4NHO Ureshino Medical Center, Ureshino-City, Japan;
5Tokyobay UrayasuIchikawa Medical Center, Urayasu-City, Japan; 6Osaka City
General Hospital, Osaka-City, Japan
Critical Care 2013, 17(Suppl 2):P491 (doi: 10.1186/cc12429) Educational eff ectiveness of introductory training for the Rapid
Response System in Japan Reference 1. Gillbe C, Gunning K: Standards for Consultant Staffi ng of Intensive Care Units. ICS & IBTICM Standards. Intensive Care Society; 2006. P493 Knowledge of confi dentiality, consent and information disclosure is
not aff ected by position or experience in adult critical care
M Lowings1, A Molokhia2, C Anderson3, S Cowman4, T Dixson5,
K El-Boghdadly6, L Williams2
1St Thomas’ Hospital, London, UK; 2Lewisham University NHS Trust, London,
UK; 3King’s College Hospital, London, UK; 4Great Ormond Street Hospital,
London, UK; 5Queen Elizabeth Hospital, Woolwich, UK; 6Princess Royal
University Hospital, Bromley, UK
Critical Care 2013, 17(Suppl 2):P493 (doi: 10.1186/cc12431) y
g
y
Results Seventy-three participants (89.0%) answered the question-
naires. The numbers of participants who scored more than four points
were as follows: A. was 71 (97.2%), B. was 70 (95.9%), C. was 64 (87.7%),
and D. was 68 (93.2%), respectively. The majority of participants
obtained the correct knowledge, and had a solid understanding for
the RRS. It was evident that providing abundant material and didactic
lectures traced from the introduction to management, and collecting
and resolving the questions, promoted comprehension. However,
there is a limitation of whether or not the participants introduce the
RRS into their own institutions. It is essential to improve the course and
continue to support the activities of the participants. Introduction Teaching of medical ethical issues including confi den-
tia lity and consent have long been a small part of the medical
curriculum. These issues are more complex in an ICU where patients
may lack capacity. Documents such as Good Medical Practice 1995,
Confi dentiality 2009 and the Mental Capacity Act 2005 give guidance
to medical professionals in these matters in the UK. Conclusion Our training course may promote the introduction and
dissemination of the RRS in Japan. Methods A questionnaire was distributed amongst staff in four ICUs in
South London. Results were analysed according to level of experience
and background (medical/nursing or allied health professional (AHP)). Results Of 225 questionnaires distributed, the response rate was
66% (31% doctors, 56% nurses and 13% AHP). Staff with either less
than 1 year experience or greater than 10 years experience had the
greatest exposure to the Mental Capacity Act and Data Protection
Act, suggesting a gap in knowledge in staff with intermediate
experience. Knowledge of the Caldicott principles were unaff ected
by experience, with many experienced respondents having ‘No Idea’. The majority of respondents (unaff ected by experience) felt that References 1. Leape LL, et al.: JAMA 2005, 293:2384-2390. 2. Jones D, et al.: N Engl J Med 2011, 365:139-146. References 1. Schiza et al.: J Clin Sleep Med 2012, 8:21-26. 2. Kaw et al.: Br J Anaesth 2012, 109:897-906. 1. Schiza et al.: J Clin Sleep Med 2012, 8:21-26. Survey of alcohol-related admissions to critical care and the
resource and fi nancial implications Survey of alcohol-related admissions to critical care and the
resource and fi nancial implications
M Slattery, P Temblett, M Heatley, N Singatullina, B Jones
ABMU University Trust, Swansea, UK
Critical Care 2013, 17(Suppl 2):P494 (doi: 10.1186/cc12432) Introduction Alcohol-related hospital and ICU admissions are known
to have a huge impact on healthcare resources in the UK. Excessive
use of alcohol is independently associated with sepsis, septic shock
and hospital mortality among ICU patients. This study assesses the
relationship between alcohol abuse and intensive care resource
utilisation in a mixed medical, surgical and neurosurgical ICU. gi
p
p
y
Conclusion Undiagnosed SDB is highly prevalent among cardiac
surgical patients. Clinical trajectories of individuals with severe SDB
are described by a prolonged recovery of pulmonary function, delayed
weaning and a pronounced infl ammatory response after surgery. Screening for SDB might identify patients that are susceptible for a
complicated postoperative course. Methods A prospective survey of emergency alcohol-related
admissions over a 1-year period was undertaken at a tertiary university
adult general and neurosurgical ICU. All patients were screened for
acute and chronic alcohol abuse on admission. Acute alcohol abuse was
defi ned as being intoxicated with alcohol at the time of admission and
chronic alcohol abuse was defi ned as chronic alcohol use exceeding
recommended UK national guidelines on consumption. The amount
of alcohol consumption was obtained, diagnosis on admission, ICU
and hospital mortality, length of stay, and total cost were recorded. All
patients were screened for alcohol-related comorbidities. Comparative
retrospective data were obtained for the same time period for
nonalcohol-related emergency ICU admissions. Data were analyzed
using SPSS. P492
Standards of referrals and admissions of critically ill patients to the
ICU A Ng
Glasgow Victoria Infi rmary Hospital, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P492 (doi: 10.1186/cc12430) A Ng
Glasgow Victoria Infi rmary Hospital, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P492 (doi: 10.1186/cc12430) S183 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 is signifi cantly less than previously reported. Our study reveals
interspecialty variations in demographic data, APACHE II scores,
mortality and cost of admission. Neurosurgical alcohol-related
admissions bear higher mortality and result in greater resource
utilisation relative to nonalcohol-related neurosurgical admissions. Alcohol continues to burden both our patients and critical care. when giving information to relatives face to face, relatives should be
kept fully informed. When giving information over the telephone,
most doctors felt the response should be tailored to the knowledge
of the person being spoken to whilst nurses were split between
tailoring the response, giving full information, setting up a password
system and not giving any information at all. Most respondents felt
date of birth and hospital number constituted ‘Patient Identifi able
Information’. However, experienced staff did not appreciate the
importance of unusual diagnosis and clinical photographs as also
being able to identify patients. Similarly, the majority knew that the
patient themselves identifi ed the ‘Next of Kin’ but 7% (unaff ected
by experience) felt this was decided by the family and felt the family
could decide on resuscitation status. When consent is required for an
elective procedure in a patient who lacks capacity, doctors tended
to have a better understanding of the need to delay the procedure
where possible than nurses, the majority of which felt this could be
decided by the next of kin or two consultant doctors. Most doctors
felt that ‘Acting in the Patient’s Best Interests’ would mean doing what
would give the patient the best outcome rather than doing what the
patient would have wanted (unaff ected by experience). The majority
of staff , on answering this questionnaire, felt that they lacked suffi cient
knowledge on the subject and most felt annual reminders would be
useful. P495 P495 95
Association of sleep disordered breathing with clinical trajectories
in patients undergoing cardiac surgery
J Roggenbach, B Tan, E Von der Leyen, A Weymann, M Karck, E Martin,
S Hofer
University of Heidelberg, Germany
Critical Care 2013, 17(Suppl 2):P495 (doi: 10.1186/cc12433) Introduction The prevalence of sleep disordered breathing (SDB)
is presumably high among individuals with cardiac diseases [1],
nonetheless SDB remains predominantly undiagnosed. However,
unrecognized SDB might have relevant impact on the postoperative
course of patients undergoing cardiac surgery [2]. g
g
g
y
Methods Polygraphic recordings of 181 patients, without previous
diagnosis of SDB, undergoing standard cardiac surgical procedures
with extracorporeal circulation were obtained during a preoperative
night. The apnea–hypopnea index (AHI – the number of apneas,
hypopneas per hour recorded) was determined and compared with
clinical characteristics and postoperative course. Conclusion The ICU is an environment where issues of consent,
confi dentiality and disclosure of information occur daily. Staff feel they
lack knowledge in these areas that is unaff ected by their experience. We need to ensure that all staff have the necessary knowledge to deal
with these situations. p
p
Results The prevalence of SDB was considerably high among all
examined patients. Median AHI was 20.8 (interquartile range, 10.6 to
36.4). Preoperative AHI was >30 in 32% of all examined individuals. During the fi rst three postoperative days, preoperative AHI >30 was
associated with a prolonged weaning time, a reduced oxygenation
index (arterial pO2/FiO2), an impaired kidney function, an augmented
infl ammatory response and an overall increased length of stay in the
ICU. The observed association of high preoperative AHI values with
postoperative clinical characteristics remained statistically signifi cant
throughout the fi rst three postoperative days. Can massage therapy help cardiac patients? A literature review plus
a case study
A Fenlon St Georges NHS Trust, London, UK
Critical Care 2013, 17(Suppl 2):P496 (doi: 10.1186/cc12434) Introduction A literature review was performed to assess whether
massage benefi ts patients postoperatively following coronary bypass
grafts (CABG) and or valve replacement/repair. A case study on a patient
who had suff ered a hypoxic brain post cardiac arrest was conducted. Methods A review on MEDLINE and Cochrane using search terms
massage, cardiac and ICU identifi ed nine research papers on the
benefi ts of massage postoperatively for the aforementioned patient
group. Other papers were listed but unrelated to cardiac surgery. None
of the nine papers identifi ed for this review were ICU specifi c in the title
but the ICU was mentioned in the main text body. For the purpose of
this review the selected papers are researching the eff ects of massage
on physiological parameters, anxiety, pain, calm and perceived stress
indicators in the CABG and/or valve repair/replacement. Out of these
nine papers, one is British (2002). Five are American (2006 to 2012),
two are Brazilian (2010) and one is an Indian paper (2010). All papers
are randomised control trials (RCTs). Papers written prior to 1999 were
excluded from this literature review. Results In total, 7.7% of patients were admitted with a history
of acute/chronic alcohol excess. Sixty-seven per cent of alcohol-
related admissions were due to acute alcohol excess. Neurosurgical
patients admitted due to alcohol excess had higher ITU mortality
than nonalcohol-related neurosurgical patients: 32.1% versus 14.39%
(P = 0.02), respectively. Ninety-three per cent of alcohol-related
neurosurgical admissions were caused by acute alcohol intoxication. The intensive care cost was signifi cantly higher for alcohol-related
(£12,396 per patient) compared with nonalcohol-related neurosurgical
admissions (£7,284 per patient). Of the medical patients admitted,
60% of these admissions were due to acute alcohol excess. The cost
of intensive care treatment was lower for alcohol-related medical
admissions. Conclusion This is one of the largest studies of alcohol-related
admissions to critical care. Our survey confi rms that alcohol-related
admissions to the ICU are commonplace; however, our frequency S184 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 educational program can reduce VAP incidence in the ICU [1]. In this
research we aimed to observe the reduction of VAP incidence after the
implementation of VAP care bundles to ICU medical personnel. Can massage therapy help cardiac patients? A literature review plus
a case study
A Fenlon Results Research from 1999 states there are methodological errors
in prior papers and very few large-scale studies prior to this date,
destabilising the validity and reliability of research from the papers
written before 1999. The later research suggests that any change in
the measured physiological parameters of blood pressure, heart rate
and respiratory rate are insignifi cant (P >0.05). Pain, anxiety, rest and
a calm score perform better with P >4.71 overall. Another American
study showed that the length of stay is reduced if a patient receives
healing touch postoperatively. However, in one RCT pain and anxiety
increases and in some case SpO2 decreases. A case study was chosen
by the author and the results support the benefi ts of massage with the
cardiac patient group.i Methods Inclusion criteria: all adult surgical patients (>18 years old)
who are on ventilatory support in the surgical ICU at Siriraj Hospital. There are two groups, divided into pre-educational group (group I) and
post-educational group (group II) (n = 220/group). We also observed
the adherence rate to VAP care bundles according to the educational
program. The pretest and post-test to determine the effi cacy of the
educational program were done. The VAP care bundles consisted of
weaning according to weaning protocol, sedation vacation, head-
of-bed elevation, measurement of cuff pressures four times/day, 2%
chlorhexidine use for mouth care and emptying of ventilator circuit
condensate. Conclusion There appears to be some benefi ts but larger-scale studies
are required within this and other ICUs in Britain. The author will be
inquiring at her workplace whether further studies can be performed
and whether she can initiate the research. She will identify further case
studies and trial massage on select patients. The author recommends
training for nurses interested in massage therapy so a bigger caseload
can be identifi ed. Results There were 45.38 and 25.25 episodes of VAP per 1,000 ventilator-
days in group I and group II, respectively (P = 0.020). The incidence of
VAP was 21.82% in group I and 9.09% in group II (P = 0.000). There was
signifi cant reduction in the length of ventilatory support per person
(group I = 2, group II = 1 (median), P = 0.013, 95% CI = 0.319 to 0.936)
and mortality rate (group I = 15.5%, group II = 8.2%, P = 0.017). There
was no signifi cant diff erence in LOI, LOH and ATB cost. References 1. Apisarnthanarak A, Pinitchai U, Thongphubeth K, Yeukyen C, Warren DK, Zack
JE, et al.: Eff ectiveness of an educational program to reduce ventilator-
associated pneumonia in a tertiary care center in Thailand: a 4-year study. Clin Infect Dis 2007, 45:704-711. Ischemic stroke in diff erent ICU type
M Damasceno, F Rodriguez, R Turon Adherence rate to VAP care bundles
Bundle
Pre-education
Post-education
P value
Wean
95.04
96.08
0.723
Sedate
92.26
95.28
0.467
HOB
50.10
70.36
0.017
Cuff pressure
87.29
80.78
0.278
M care
89.60
80.86
0.204
Drain
95.42
99.42
0.130 Results A total of 3,854 admissions were registered in the period in
both ICUs, with 257 (6.7%) being by ischemic stroke – 49 in GICU and
208 in NICU. Mean age of patients: 73 years in GICU and 70.1 years in
NICU. Admissions from emergency unit: 44 (89.8%) GICU, 181 (87%)
NICU. Mean SAPS 3 score: 50.7 (29 to 71) GICU, 50.6 (29 to 98) NICU. Patients admitted with infection: 6 (12.2%) GICU, 10 (4.81%) NICU. Mean
Charlson comorbidity index points: 1.61 (median 1.0) GICU, 1.1 (median
1.0) NICU. Mean length of ICU stay: 4.8 days (median 3) GICU, 4.3 days
(median 3) NICU. Mean length of hospital stay: 12.2 days (median
8) GICU, 14.5 days (median 8) NICU. Predicted hospital mortality
(mean ± SD): 20.07% ± 13.48 GICU, 20.4% ± 13.83 NICU. Hospital
mortality rate: 12.5% GICU, 5.39% NICU. Observed-to-expected (O/E)
mortality ratios: 0.62 GICU, 0.26 NICU. Conclusion The educational program and the implementation of VAP
care bundles can reduce the incidence of VAP, length of ventilatory
support and mortality rate in the ICU. Reference y
Conclusion Despite the similar proportions numbers for patients in
both ICUs, the mortality rate and the O/E mortality ratio for ischemic
stroke were higher in patients of the GICU when compared with the
NICU. Can massage therapy help cardiac patients? A literature review plus
a case study
A Fenlon The pretest
scores were 15.53 and 17.53 on average from 40 medical personnel in
group I and group II, respectively (P = 0.000). The head-of-bed elevation
adherence rate was improved after the educational program (group
I = 50.1%, group II = 70.36%, P = 0.017). But the adherence to other
bundles was not improved. See Tables 1 and 2. Ischemic stroke in diff erent ICU type
M Damasceno, F Rodriguez, R Turon Hospital de Clinicas de Niterói, Rio de Janeiro, Brazil
Critical Care 2013, 17(Suppl 2):P497 (doi: 10.1186/cc12435) Table 1 (abstract P498). Impact of the educational program on outcomes
Outcome
Pre-education
Post-education
P value
VAP
45.38
25.25
0.020
Ventday
2
1
0.013
LOI
3
3
0.054
LOH
17
17
0.960
ATB cost
2,842.50
2,098.00
0.994
% death
15.5
8.2
0.017
Table 2 (abstract P498). Adherence rate to VAP care bundles
Bundle
Pre-education
Post-education
P value
Wean
95.04
96.08
0.723
Sedate
92.26
95.28
0.467
HOB
50.10
70.36
0.017
Cuff pressure
87.29
80.78
0.278
M care
89.60
80.86
0.204
Drain
95.42
99.42
0.130
Conclusion The educational program and the implementation of VAP
care bundles can reduce the incidence of VAP, length of ventilatory
support and mortality rate in the ICU. Reference
1. Apisarnthanarak A, Pinitchai U, Thongphubeth K, Yeukyen C, Warren DK, Zack
JE, et al.: Eff ectiveness of an educational program to reduce ventilator-
associated pneumonia in a tertiary care center in Thailand: a 4-year study. Clin Infect Dis 2007, 45:704-711. P499
Development of a severe infl uenza critical care curriculum and
training materials for resource-limited settings
JV Diaz1, P Lister2, JR Ortiz3, NK Adhikari4, N Adhikari5
1World Health Organization, San Francisco, CA, USA; 2Great Ormond
Street Hospital, London, UK; 3University of Washington, Seattle, WA, USA;
4Sunnybrook Health Sciences Centre and University of Toronto, Canada;
5Health Sciences Centre and University of Toronto, Canada
Critical Care 2013, 17(Suppl 2):P499 (doi: 10.1186/cc12437) Table 1 (abstract P498). Impact of the educational program on outcomes Introduction In our hospital there are a general intensive care unit
(GICU) and a neurointensive care unit (NICU). Despite the preference
for the NICU, in both there are admissions for ischemic stroke. There
are diff erent staff for each ICU, with the same physician’s leadership. We have decided to evaluate the performance of both ICUs, analysing
whether there are diff erences in results as some authors publish best
results in specialized ICUs. Methods Using prospectively collected data, we undertook a
retrospective evaluation of all patients admitted to the GICU and NICU
of our hospital with the diagnosis of ischemic stroke, from December
2010 to November 2012. In both ICUs there are intensivists, but in the
NICU the intensivists have special expertise in neuroscience. Data were
collected from Epimedmonitor. Table 2 (abstract P498). P497 Ischemic stroke in diff erent ICU type
M Damasceno, F Rodriguez, R Turon 1.
Varelas PN, et al.: Neurocrit Care 2008, 9:293-299.
2.
Berchad EM, et al.: Neurocrit Care 2008, 9:287-292. P499
Development of a severe infl uenza critical care curriculum and
training materials for resource-limited settings j
p
CH Pisitsak, O Chaiwat Siriraj Hospital, Bangkok, Thailand JV Diaz1, P Lister2, JR Ortiz3, NK Adhikari4, N Adhikari5
1World Health Organization, San Francisco, CA, USA; 2Great Ormond
Street Hospital, London, UK; 3University of Washington, Seattle, WA, USA;
4Sunnybrook Health Sciences Centre and University of Toronto, Canada;
5Health Sciences Centre and University of Toronto, Canada
Critical Care 2013, 17(Suppl 2):P499 (doi: 10.1186/cc12437) Introduction VAP has continued to be a major cause of morbidity
and mortality in critically ill patients in Thailand for decades. Previous
research found that the implementation of VAP care bundles and the S185 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Introduction The 2009 H1N1 pandemic caused surges of severely ill
patients with viral pneumonia requiring ventilation and particularly
aff ected high-dependency medical services such as ICUs in settings
that lack suffi cient personnel and resources to provide optimal care. Ministries of Health of several countries asked the World Health
Organization (WHO) for clinical management guidance. Introduction The 2009 H1N1 pandemic caused surges of severely ill
patients with viral pneumonia requiring ventilation and particularly
aff ected high-dependency medical services such as ICUs in settings
that lack suffi cient personnel and resources to provide optimal care. Ministries of Health of several countries asked the World Health
Organization (WHO) for clinical management guidance. Figure 2 (abstract P500). Compliance with pre-ICU resuscitation bundle. g
(
)
g
g
Methods We developed (2009 to 2011) and piloted (2011 to 2012)
curriculum and training materials targeted at clinicians without formal
critical care training who care for adult and paediatric patients with
severe acute respiratory infections in ICUs in resource-limited settings. Results With contributions from 37 global experts, we developed a
3-day course including 14 learning units in early recognition, patho-
physiology, oxygen therapy, infl uenza diagnostics, infection control,
resuscitation of septic shock, antimicrobial therapy, monitoring, lung-
protective ventilation for acute respiratory distress syndrome, sedation,
weaning, preventive care, quality improvement, and ethics. Teaching
techniques are appropriate for adult learners and include short
slideshow-based lectures, interactive small-group role-play sessions
and a toolkit with practical resources such as checklists. With funding
from the WHO and local Ministries of Health, the training course was
piloted among 67 clinician participants in the Caribbean and Indonesia
(29 and 38, respectively) and taught by external critical care specialists. Participants’ daily evaluations rated all units as at least very good. P501 Safety climate perception for staff nurses in an ICU
RL Rivera-Romero, M Torres-Campos, MJ Delgado-Amaya, E Curiel-Balsera,
JG Quesada-García
Hospital Regional Carlos Haya, Málaga, Spain
Critical Care 2013, 17(Suppl 2):P501 (doi: 10.1186/cc12439) Introduction Following our study of severe sepsis care across three
centres [1], we aimed to introduce a rapid feedback mechanism into
our rolling audit programme. Whilst previous audits raised awareness
of severe sepsis, only whole organisation performance was reported
and no feedback was given to individual clinicians. It is recognised that
such feedback loops can improve clinical practice [2]. Introduction When we talk about safety culture, we speak of being
aware that things can go wrong. We must be able to recognize mistakes
and learn from them, sharing that information fairly and impartially
to try to prevent its recurrence. Organizations such as the Agency for
Healthcare Research and Quality (AHRQ) have developed tools to help
organizations measure their safety culture and there is little information
about our country. Methods Patients admitted to critical care (58 beds, four units) with
a primary admission diagnosis of infection were screened for severe
sepsis. Pre-ICU care was then audited against the Surviving Sepsis
Guidelines [3]. Time zero is defi ned as when criteria for severe sepsis
were fi rst met. An individualised traffi c-light report was then generated
and emailed to the patient’s consultant and other stakeholders
involved in care (Figure 1). We aimed to report cases within 7 days of
critical care admission. A cumulative report is generated monthly to
track organisation-wide performance. Methods A descriptive survey study. We sent the Spanish version of the
questionnaire on patient safety culture (AHRQ) to the nursing staff of a
polyvalent ICU of 42 beds in a tertiary hospital. Results The questionnaire was sent to 179 nurses, receiving correctly
answered 88 surveys (response rate of 49.16%). On a scale of 0 to
10, 6.97 points was obtained to estimate the safety climate for staff
respondents. The item best scored was teamwork in the unit (65.9%). Detected as a fortress, ‘communication between nurses at shift
changes’ (76.1% positive responses). The worst rating was obtained in
the section on human resources, followed by management support in
the fi eld of patient safety.f Figure 1 (abstract P500). Example report. Figure 1 (abstract P500). Example report. Conclusion The perception of safety culture in an ICU by nursing staff is
far from optimal levels. Quality assurance in severe sepsis: an individualised audit/feedback
system results in substantial improvements at a UK teaching
hospital p
M Simmonds, E Blyth, W Robson, M Chikhani
Nottingham University Hospitals NHS Trust, Nottingham, UK
Critical Care 2013, 17(Suppl 2):P500 (doi: 10.1186/cc12438) P499
Development of a severe infl uenza critical care curriculum and
training materials for resource-limited settings Test scores to assess participant critical care knowledge improved
signifi cantly from before the training to immediately after (Caribbean,
58 to 80%; Indonesia, 56 to 75%; P <0.001 for both). Results Since November 2011, 153 cases of severe sepsis have been
audited and reported back to clinicians. Compliance with antibiotics
in <1 hour has risen from 35 to 75% and compliance with the pre-
ICU elements of the resuscitation bundle has risen from 20 to 70%
(Figure 2). Feedback from clinicians has been encouraging as our
reports highlight both positive and negative examples of practice. Conclusion Individualised feedback on sepsis care has led to
substantial improvements in guideline compliance. This concept could
be translated to other time-dependent patient pathways. References 1. Simmonds MJR, Chikhani M, Smith P, et al.: Multi-departmental system
analysis is needed for evaluation of severe sepsis care: a multi-centre
study [abstract]. Presented at the BMJ International Forum on Quality and
Safety in Healthcare; 2011; Amsterdam. Conclusion It was feasible to develop and deliver an advanced critical
care curriculum and related training materials in a short, interactive
workshop for noncritical care trained clinicians in resource-limited
settings. However, the small-scale programme was labour and time
intensive. Widespread dissemination of these materials requires
identifi cation of target countries, engagement of Ministries of Health,
and secure funding. Longer-term evaluation is necessary to determine
whether such programmes improve processes of care and clinical
outcomes for critically ill patients. 2. Jamtvedt G, Young JM, Kristoff ersen DT, et al.: Audit and feedback: eff ects on
professional practice and health care outcomes. Cochrane Database Syst
Rev 2006, 2:CD000259. 3. Dellinger RP, Levy MM, Carlet JM, et al.: Surviving Sepsis Campaign:
international guidelines for management of severe sepsis and septic
shock: 2008. Crit Care Med 2008, 36:296-327. First-hour protocol clarifi es resource management and nursing staff
education in the ICU First-hour protocol clarifi es resource management and nursing staff
education in the ICU
M Arbelius-Iltanen, R Siren, J Heinilä, P Korhonen, T Sutinen, V Jalkanen,
T Ahonen, S Karlsson
Tampere University Hospital, Tampere, Finland
Critical Care 2013, 17(Suppl 2):P503 (doi: 10.1186/cc12441) Results A 10% increase in the proportion of ICU nurses with a bachelor’s
degree or higher was associated with 2% lower odds of death while
controlling for patient and hospital characteristics. Patients cared for
in better work environments experienced 11% lower odds of risk-
adjusted death than those cared for in poorer ICU work environments. Conclusion Patients cared for in hospitals with a greater proportion of
bachelor’s prepared ICU nurses and in better ICU work environments
experienced signifi cantly lower odds of death. As the demand for
critical care services increases, attention to the education level of ICU
nurses and ICU work environment may be warranted to optimize
currently available resources and potentially yield better outcomes. M Arbelius-Iltanen, R Siren, J Heinilä, P Korhonen, T Sutinen, V Jalkanen,
T Ahonen, S Karlsson
Tampere University Hospital, Tampere, Finland
Critical Care 2013, 17(Suppl 2):P503 (doi: 10.1186/cc12441) Introduction The fi rst-hour protocol determines the patient-specifi c
resources for the start of an ICU stay [1]. Staff resources are decided
through triage. Task charts direct the start of intensive care. Our primary
goal is to improve patient care. g
p
p
Methods A triage method (red, yellow, green) is used to manage ICU
resources according to the severity of illness. For example, one doctor
and one nurse would admit a stable (green) patient coming from the
operating room for postoperative ICU care. A patient in septic shock
with multiple organ disorder (red), on the other hand, would be
admitted by a team of two doctors and three nurses. Each staff member
has a task chart in a checking-list format. Also, an admission chart is used
to improve data collection. The use of the protocol started as a pilot
study in early 2012. Simulation education for staff members started in
August 2012 and has included video recordings and debriefi ng of each
simulated ICU admission. Primary goal-directed therapy goals have
been mean arterial pressure (MAP >65 mmHg), SpO2 >94%, timing
of the laboratory tests, start of antibiotics, and blood glucose level
6 to 8 mmol/l. Quality indicators have been followed from the data
provided by The Finnish Intensive Care Consortium. P504l Methods In a survey, we evaluated the study course. Therefore, a
questionnaire was distributed among all students including the topics
course contents, learning materials, time management, supervision,
and overall impression. The students were asked to score their
agreement to the statements ‘content is well structured’, ‘content
extent is appropriate’ and ‘content is relevant for medical purposes’ on
a scale ranging from 1 (fully disagree) to 5 (fully agree). P502
Physico-technical medicine: teaching technical skills to the
intensivist Introduction The requirements for the intensivist in handling medical
technology are constantly growing. It appears necessary to acquire
technological competences particularly within the fi elds of medical
technology and physics. In the master’s degree program ‘MasterOnline
Physico-Technical Medicine’, such technical authority is conveyed. To cope with the intensive vocational situation of the physician,
this study course follows the blended-learning concept; that is, it is
conceived as an online study course with small portions of intermittent
presence phases. Within the fi rst year, technical basic skills such as
‘measurement technique’, ‘informatics’, and ‘advanced physics’ are
covered. Subsequently, two of various advanced courses in diff erent
fi elds of medical technology (‘technology in intensive care medicine’,
‘technology in surgery’, ‘technical cardiology’, ‘radiology’, and other) are
selected. Conclusion The fi rst-hour protocol has helped us in resource
management, start of the patients’ intensive care and education of
nursing staff . Reference 1. Brown R, et al.: Aust Crit Care 2012, 25:178-187. 1. Brown R, et al.: Aust Crit Care 2012, 25:178-187. 1University of Pittsburgh, PA, USA; 2University of Pennsylvania, Philadelphia,
PA, USA Critical Care 2013, 17(Suppl 2):P504 (doi: 10.1186/cc12442) Introduction Demand for critical care services is increasing yet a
comprehensive understanding of how critical care nurses – the largest
group of ICU direct care providers – impact outcomes remains unclear. The purpose of this study was to determine how critical care nurse
education (hospital proportion of bachelor’s prepared ICU nurses) and
ICU work environment infl uenced 30-day mortality of mechanically
ventilated older adults. Results The students participated actively in this study course with
highest motivation and large commitment. The students’ workload
was in the targeted range of about 10 hours/week. Content structure
was scored with 4.3 ± 0.1, content extent with 4.1 ± 0.2 and medical
relevance with 4.3 ± 0.2. Conclusion The blended-learning concept fulfi lls the requirements
for occupation-accompanying continued medical education, since it
off ers the possibility to study self-employed accessing text documents,
lecture recordings, and electronic lectures and to convert in
concentrated presence phases this knowledge into practical exercises. Methods A multi-state cross-sectional nurse survey was linked to
hospital administrative data and Medicare claims (2006 to 2008). The
fi nal sample included 55,159 mechanically ventilated older adults in 303
hospitals. Logistic regression modeling was employed to jointly assess
the relationship of critical care nurse education, work environment and
staffi ng on 30-day mortality while adjusting for hospital and patient
characteristics and accounting for clustering. First-hour protocol clarifi es resource management and nursing staff
education in the ICU Questionnaires for
the staff members have been used to evaluate opinions about the fi rst-
hour protocol. P505 Infl uence of critical care nurse education and work environment on
outcomes of mechanically ventilated older adults
DM Kelly1, LH Aiken2 1University of Pittsburgh, PA, USA; 2University of Pennsylvania, Philadelphia,
PA, USA P501 The team work dimension was identifi ed as the
most valued by workers, with the transmission of information on shift
changes the most valued item. Figure 1 (abstract P500). Example report. S186 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P502
Physico-technical medicine: teaching technical skills to the
intensivist
J Seifried, S Richter, J Guttmann, S Schumann
University Medical Center Freiburg, Germany
Critical Care 2013, 17(Suppl 2):P502 (doi: 10.1186/cc12440) nurses considered that the protocol has no eff ect, and none thought
it to be adverse for patient care. Corresponding numbers for our ICU
doctors were 87% (benefi cial n = 13/15), 13% (no eff ect n = 2/15) and
0% (adverse). Furthermore, 82.5% (n = 66/80) of the nurses replied that
education of new nurse staff members has improved because of the
fi rst-hour protocol. A total of 17.5% (n = 14/80) thought there has been
no eff ect, and none considered the protocol harmful for education. For ICU doctors the protocol did not bring either clear educational
advantages or disadvantages. The variable life-adjusted display curves
(The Finnish Intensive Care Consortium) have shown improvement in
our patient care after the implementation of the fi rst-hour protocol. However, we cannot determine whether it is a signifi cant factor in our
intensive care results.i Are general wards suffi ciently staff ed to care for level 1 patients?
G Rajendran, C Tjen, S Hutchinson, S Fletcher
Norfolk & Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P506 (doi: 10.1186/cc12444) y
gi
Results The survey was completed by 129 ICU workers. The global
prevalence of ISQ in ICU was 67.4%. ISQ was observed in 45% of
physicians and 82% of nurses and nurses assistant (P <0.001). Sleep
medication was used by 13.3% of the ICU team. Univariate analysis
showed that ISQ was signifi cantly associated with sex (73% vs. 43%, P = 0.03 in women and men, respectively), marital status (84%
vs. 61%, P = 0.01 in single and couple workers, respectively), more
than 60 hours working in the last week (76% vs. 61%, P = 0.07) and
less than 6 sleeping hours (95% vs. 54%, P <0.0001). Multivariable
analysis demonstrated that a sleep duration less than 6 hours was
independently associated with ISQ (OR = 24.5; 95% CI = 5.2 to 115.8;
P <0.0001). Furthermore, pathologic sleepiness was present in 59.3%
of ICU workers. Sleepiness was independently associated with use of
sleep medication (OR = 5.9; 95% CI = 1.2 to 28.5; P = 0.025). Introduction There are several defi nitions of level 1 (L1) care, all refer
to a group at risk of clinical deterioration on the ward [1-3]. There is
evidence that ward patients who become acutely unwell often receive
suboptimal care [4]. A regional study commissioned by Norfolk, Suff olk
& Cambridgeshire Critical Care Network (NSCCCN) found that a majority
of ward patients may be of L1 dependency and death rates appear to be
correlated with L1 status. We aim to examine the relationship between
the ward distribution of illness acuity, staffi ng and patient outcome. fi
Methods Data were collected as part of NSCCCN’s observational
prevalence study in 2010. Ward surveys included acuity of illness,
staffi ng levels and skill mix. Secondary data were obtained from the
Patient Administration System. Emergency, oncology, paediatric and
maternity units were excluded. Conclusion The prevalence of ISQ and sleepiness is very high among
ICU workers. Those disturbances are independently associated with a
sleep duration less than 6 hours, and sleep medication use, respectively. These results highlights that strategies to decrease ISQ and sleepiness
in ICU clinicians are urgently needed to improve work performance,
improve quality of care provided and prevent adverse events. Results Complete datasets were obtained from 1,402 patients in 22
wards in our university hospital over two seasons. This constitutes
98.3% of inpatients from those wards. P505 Activity and case-mix changes in a medical ICU after the
geographical transfer of a third-level university hospital
JC Cebrián, FM Monsalve, JB Bonastre
Hospital Universitario y Politecnico La Fe, Valencia, Spain
Critical Care 2013, 17(Suppl 2):P505 (doi: 10.1186/cc12443) Introduction Information about big hospital geographical transfer is
scarce in the medical literature. On 20 February 2011 our hospital (in fact,
a big university complex) was transferred from their previous location
in the north-center of our city towards a new southern peripheral,
geographical location. This transfer has been done without any changes
in assisted population or nursing/medical staff . The only change was a
slight increase in bed number (21 to 24). Our aim is to analyze changes
in activity indexes (length of stay, occupancy rate, and so forth) and case
mix (origin, previous quality of life and NYHA score, main diagnostic
groups, severity scores, in-ICU and in-hospital mortality). Results According to the questionnaire replies, 80% (n = 64/80) of our
nurses estimate that the fi rst-hour protocol has improved the starting
process of our patients’ intensive care. Twenty percent (n = 16/80) of the S187 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Methods To compare our number of admissions, related activity and
case-mix indicators 1 year before and after the geographical change
was done. We analyzed our whole number of patients admitted to the
ICU. We used the chi-square test for categorical variables and one-way
analysis of variance for quantitative data. Minitab and Statbas statistical
programs were used. We plotted activity data using the Barber–
Johnson 1 diagram. [http://www.nice.org.uk/cg50]
3. AUKUH Acuity/Dependency Tool. Association of UK University Hospitals; 2007. [http://www.aukuh.org.uk/index.php/affi liate-groups/directors-of-nursing/
patient-care-portfolio] [http://www.nice.org.uk/cg50]
3. AUKUH Acuity/Dependency Tool. Association of UK University Hospitals; 2007. [http://www.aukuh.org.uk/index.php/affi liate-groups/directors-of-nursing/
patient-care-portfolio] 4. 4. An Acute Problem? NCEPOD; 2005. [http://www.ncepod.org.uk/2005report/index.html] Results A total of 2,774 cases (63% males; mean age 61 years) were
admitted to our ICU during the period (1 year before and after the
transfer). No diff erences between both groups were founded in
demographic data, Knaus score and NYHA status. Regarding their
origin, we found more patients admitted from other hospital centers
(20 vs. 29%; P <0.001). APACHE II score increased from 17.24 to 19.08%
(P <0.001) and a slight increase change in SAPS 3 score was also found
(52.29 to 53.75; P <0.01). Work-related stress amongst doctors in intensive care, anaesthetics,
accident and emergency and general medicine g
y
g
Tuthill, MS Ahmed, G Mathew, AC Bolton, AA Molokhia JI Tuthill, MS Ahmed, G Mathew, AC Bolton, AA Molokhia
University Hospital Lewisham, London, UK Introduction Work-related stress is a potential problem among
doctors and is associated with anxiety, depression, reduced job
satisfaction, days off work, errors and near misses [1]. To compare stress
levels between diff erent groups of doctors and identify causes of stress,
we conducted a survey at University Hospital Lewisham using the UK
Health and Safety Executive’s Management Standards (HSEMS). HSEMS
is a validated tool developed to identify work conditions that warrant
interventions to reduce stress levels across organisations [2]. y
y
Conclusion Better outcome with improved SPr may be unsurprising,
although if proven conclusively would signifi cantly inform workforce
planning. Lack of correlation between staffi ng levels and occupancy or
acuity is also interesting given that we know L1 criteria are associated
with worse outcome. P505 Our in-ICU mortality remains lower (15.5
to 15.6%) whereas observed mortality decreased (22.37 to 19.88%;
P <0.001). An increase in our neurologic patients has been the most
consistent change regarding diagnostic groups. The activity indexes
show a slightly decrease in occupancy rate (79.2 vs. 76.8). 1.
Levels of Critical Care for Adult Patients. Intensive Care Society; 2002.
[http://www.ics.ac.uk/professional/standards_safety_quality/standards_and_
guidelines/levels_of_critical_care_for_adult_patients]
2.
Acutely Ill Patients in Hospital. NICE Guideline 50. NICE; 2007. P508 Work-related stress amongst doctors in intensive care, anaesthetics,
accident and emergency and general medicine
JI Tuthill, MS Ahmed, G Mathew, AC Bolton, AA Molokhia
University Hospital Lewisham, London, UK
Critical Care 2013, 17(Suppl 2):P508 (doi: 10.1186/cc12446) Work-related stress amongst doctors in intensive care, anaesthetics,
accident and emergency and general medicine 1.
Yates J: Hospital Beds: A Problem for Diagnostic and Management. Heinemann
Medical Books; 1982. Are general wards suffi ciently staff ed to care for level 1 patients?
G Rajendran, C Tjen, S Hutchinson, S Fletcher
Norfolk & Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P506 (doi: 10.1186/cc12444) The mean ward occupancy rate
was 94% (10th to 90th percentile: 85% to 100%). At least one L1 acuity
criterion was scored by 898 (64%) patients, with 25% from geriatrics
followed by orthopaedics (17%) and general surgery (10%). Each ward
had an average of eight qualifi ed nursing staff (range: 4 to 12) equating
to an average staff :patient ratio (SPr) of 0.253. There was no correlation
between ward occupancy and nursing staff (Pearson correlation, corr:
0.55), nor between prevalence of L1 criteria and staffi ng (corr: 0.34). The
admission rate to intensive care was noted to be higher if the patients
were nursed in a ward with lower than average SPr compared with
higher SPr (2.7% vs. 1.2%, P = 0.058 Fisher’s exact), but this was not
statistically signifi cant. Senior nursing (Band 6) staff were part of the
skill mix on only nine of 44 ward surveys. P508 Reference 1. Yates J: Hospital Beds: A Problem for Diagnostic and Management. Heinemann
Medical Books; 1982. Methods A survey was conducted in six Uruguayan ICUs. The sleep
quality was evaluated on the basis of the Pittsburgh score (PS), and the
sleepiness was identifi ed by the Epworth scale. ISQ was defi ned as PS
greater than 5 points and sleepiness by an Epworth scale higher than
6 points. ICU’s, patient’s, and clinician’s characteristics were assessed for
their association with the prevalence of ISQ. All variables with P <0.2
in univariate analysis were included in a model of ordinal regression. P <0.05 was considered statistically signifi cant. P507
Sleep quality among ICU workers
h 1
k
h
1
h
2 Sleep quality among ICU workers
G Burghi1, I Serkochian1, B Frache2, P Alzugaray2, K Goinheix1,
M Rodriguez Verde3, H Bagnulo1, E Azoulay4
1Hospital Maciel, Montevideo, Uruguay; 2Sanatorio Americano, Montevideo,
Uruguay; 3Hospital de Paysandú, Uruguay; 4Hôpital Saint Louis, Paris, France
Critical Care 2013, 17(Suppl 2):P507 (doi: 10.1186/cc12445) Introduction Prolonged shifts, workload, stress, and diff erent con-
fl icts are associated with burnout, loss of psychological wellbeing,
and probably with an inadequate sleep quality (ISQ). This relevant
disturbance leads to deterioration of the work performance, may impair
quality of care provided to patients and increases the incidence of
serious adverse events. The objective was to determine the prevalence
of ISQ and sleepiness among Uruguayan ICU workers, and to evaluate
risk factors associated with ISQ. Conclusion According to the previous data our ICU seems to perform
better in the new location with a decrease in the standardized mortality
rate. On the other hand, we are admitting more patients transferred
from other hospitals. A better occupancy rate was found. f P506
A P506
Are general wards suffi ciently staff ed to care for level 1 patients? G Rajendran, C Tjen, S Hutchinson, S Fletcher
Norfolk & Norwich University Hospital, Norwich, UK
Critical Care 2013, 17(Suppl 2):P506 (doi: 10.1186/cc12444) Infl uence of adverse drug events on the mortality and the length of
hospital stay in ICUs in Japan: the JADE Study
Y Ohta1, M Sakuma1, D Bates2, T Morimoto1
1Kinki University, Osaka, Japan; 2Harvard University, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P510 (doi: 10.1186/cc12448) og dad y , J
o
,
O
oc e
1Princess Royal University Hospital, London, UK; 2University Hospital
Lewisham, London, UK
Critical Care 2013, 17(Suppl 2):P509 (doi: 10.1186/cc12447) Critical Care 2013, 17(Suppl 2):P509 (doi: 10.1186/cc12447) Introduction South-east London (SEL) presents unique challenges to
healthcare providers due to its diverse demographic. The high levels of
poverty, immigration and psychiatric illness impact delivery of obstetric
care. These were identifi ed as risk factors for poor outcome in the latest
CMACE report [1]. The Intensive Care National Audit and Research
Centre (ICNARC) produced data on obstetric critical care admissions in
2007 [2]. We reviewed the obstetric critical care admissions in three SEL
hospitals and compared this with the national average determined in
the ICNARC and CMACE data. Introduction Adverse drug events (ADEs) are associated with a
substantial increase in morbidity and mortality in any setting. Because
patients in ICUs were critically ill with complex diseases and varied
organ dysfunction, the incidence of ADEs on such patients is much
more crucial than the counterparts. We thus assessed the nature of
ADEs and their infl uence in ICUs. l
Methods We conducted a prospective cohort study at ICUs at three
large tertiary-care hospitals in Japan. Trained research nurses reviewed
all medical charts, incident reports and reconciliations from the
pharmacy to identify suspected ADEs as well as the background of
patients. ADEs are any injuries that result from the use of a drug. After
suspected ADEs are collected by research nurses, physician reviewers
independently evaluated them and classifi ed them as ADEs or rule
violations. We used the validated methodology [1]. Methods All critical care admissions in three high-risk obstetric units
in SEL (1 August 2009 to 31 July 2011) were screened for patients who
were currently or recently pregnant. We compared local results with
national data by ICNARC and CMACE. Figure 1 (abstract P509). Causes for critical care admission in SEL,
CMACE and ICNARC. Results We included 459 patients with 3,231 patient-days. The median
age was 70 years and the median length of stay was 3 days. In total, 70
patients (15%) had at least one ADE during their stay in the ICU. The
median ICU stay in patients who had ADEs was 14 days while 2 days
in patients who had no ADEs (P <0.0001). The median length of the
ADE onset days since admission was 3 days. References Methods We conducted an anonymous survey of doctors working
in anaesthetics, intensive care, general medicine and accident and
emergency (A&E) departments over 6 weeks using the HSEMS S188 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 question naire. We also surveyed awareness of the Trust’s stress
management services and whether staff had a designated supervisor
or mentor. Results were analysed using the HSEMS Analysis Tool, which
rates stressors with a score from 1 to 5 (5 represents the lowest amount
of stress). We compared the Trust’s results against HSEMS national
standards. Results There were 68 obstetric critical care admissions in the SEL
hospitals within the audited time frame. The mean age was 30.05 in
ICNARC data compared with 33.93 in SEL. Average APACHE II scores
were lower in SEL compared with the ICNARC data, but length of
stay was greater in SEL (2.72 days) compared with ICNARC (1.5 days). Haemorrhage was the most common reason for admission in SEL,
whilst sepsis was the leading cause of death according to the latest
CMACE report (Figure 1). Results Seventy-two doctors completed the survey. Lowest stress
levels were found in doctors working in intensive care (n = 12, mean
3.63, SD 0.39). This was followed by medicine (n = 26, mean 3.55,
SD 0.47), anaesthetics (n = 27, mean 3.40, SD 0.44), and A&E (n = 7,
mean 3.11, SD 0.65), which had the highest stress levels. There was
no signifi cant diff erence in stress levels between diff erent grades of
doctors. When compared with HSEMS targets, staff relationships and
peer support exceeded national standards. However, management
of organisational change and demands at work need improvement. The majority of doctors (82%) had no idea what stress management
services were provided by the Trust. Seventy-nine per cent of doctors
had an allocated supervisor or mentor, 91% of those felt able to
approach their supervisor. g
Conclusion Data from national audits may guide protocol, but services
must be tailored to local circumstances. SEL has unique population
characteristics and obstetric critical care admissions diff er signifi cantly
from national statistics; in particular, haemorrhage is over-represented
in our region. Critical care services were generally required for a short
period of time; during this period, routine postpartum care may be
omitted as treatment priorities diff er. Morimoto T, et al.: Qual Saf Health Care 2004, 13:306-314. References 1. Kerr et al.: HSE management standards and stress-related work outcomes. Occup Med (Lond) 2009, 59:574-579. 1. Kerr et al.: HSE management standards and stress-related work outcomes. Occup Med (Lond) 2009, 59:574-579. 2. Health and Safety Executive Management Standards [http://www.hse.gov. uk/stress/standards/index.htm] 2. Health and Safety Executive Management Standards [http://www.hse.gov. uk/stress/standards/index.htm] References Dedicated critical care services on
the labour ward may be a way to combine postnatal care with transient
high-dependency requirements. This may enhance patient experience
and prove cost-eff ective. References Conclusion These survey results provide reassurance that stress levels
in intensive care compare well, despite critically unwell patients and
higher mortality rates. We identifi ed areas that need improvement
within the Trust and will present these results to all relevant
departments. With the support of hospital management we will initiate
HSEMS-validated measures to reduce stress. 1. Centre for Maternal and Child Enquiries: Saving mothers’ lives: reviewing
maternal deaths to make motherhood safer: 2006–08. The Eighth Report
on Confi dential Enquiries into Maternal Deaths in the United Kingdom. BJOG 2011, 118(Suppl 1):1-203. 2. Female Admissions (Aged 16–50 Years) to Adult, General Critical Care
Units in England, Wales and Northern Ireland, Reported as ‘Currently
Pregnant’ or ‘Recently Pregnant’ [http://www.oaa-anaes.ac.uk/assets/_
managed/editor/File/Reports/ICNARC_obs_report_Oct2009.pdf] References P510l Infl uence of adverse drug events on the mortality and the length of
hospital stay in ICUs in Japan: the JADE Study
Y Ohta1, M Sakuma1, D Bates2, T Morimoto1
1Kinki University, Osaka, Japan; 2Harvard University, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P510 (doi: 10.1186/cc12448) P509 Audit of obstetric critical care admissions in a high-risk population
K El-Boghdadly1, J Aron2, DN Onwochei1
1Princess Royal University Hospital, London, UK; 2University Hospital
Lewisham, London, UK
Critical Care 2013, 17(Suppl 2):P509 (doi: 10.1186/cc12447) P511
Attitude and knowledge of intensive care staff concerning donation
in Hungary: it is the fi rst step to change g
yi
p
A Smudla1, S Mihály2, J Fazakas1 A Smudla , S Mihály , J Fazakas
1Semmelweis University, Budapest, Hungary; 2Hungarian National Blood
Transfusion Service, Budapest, Hungary
Critical Care 2013, 17(Suppl 2):P511 (doi: 10.1186/cc12449) Conclusion Introduction of a SN:OD and a clinical pathway has led to
the identifi cation of previously missed potential organ donors in the
ED. Several patients have subsequently been admitted to critical care
solely to facilitate organ donation. Introduction In Hungary, despite the high level of social support, the
number of organ recoveries from deceased donors has not changed
signifi cantly. The donation activity shows a positive relationship with
the level of education of staff in ICUs as well as with their attitude
towards transplantation. The aim of this cross-sectional study is to
estimate the attitude and knowledge of intensive care specialists and
nurses as regard donation and transplantation. Infl uence of adverse drug events on the mortality and the length of
hospital stay in ICUs in Japan: the JADE Study
Y Ohta1, M Sakuma1, D Bates2, T Morimoto1
1Kinki University, Osaka, Japan; 2Harvard University, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P510 (doi: 10.1186/cc12448) Regarding the mortality,
73 patients (16%) were dead during their ICU stay: 12 deaths (17%) in
patients who had ADEs and three of 12 deaths were caused by an ADE,
and 61 deaths (16%) in counterparts (P = 0.8). There were no signifi cant
diff erences of patients’ characteristics between patients with ADEs and
without ADEs (Table 1). Conclusion ADEs were associated with longer stay and caused a part of
death in ICU (4%) although they did not increase the mortality. Because
the characteristics of patients were not associated with ADEs, early
detection and intervention for ADEs could be important to improve the
morbidity and reduce the death caused by ADEs in ICUs. Reference Figure 1 (abstract P509). Causes for critical care admission in SEL,
CMACE and ICNARC. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S189 Table 1 (abstract P510). Patient characteristics
Patients
Patients
with
without
ADEs
ADEs
Characteristic
(n = 70)
(n = 389)
P value
Age ≥65 years, n (%)
49 (70)
227 (58)
0.08
Male, n (%)
40 (57)
250 (64)
0.2
Department
Medicine
35 (50)
228 (59)
Surgery
35 (50)
161 (41)
0.2
Unconsciousness, GCS ≤8
13 (19)
91 (23)
0.4
Heart failure, NYHA = 4
37 (53)
216 (56)
0.7
Respiratory failure
44 (62)
262 (67)
0.5
Kidney failure
6 (9)
58 (15)
0.2
Clinical laboratory measurements, median (25%, 75%)
Blood urea nitrogen (mg/dl)
21 (15, 39)
19 (14, 31)
0.2
Creatinine (mg/dl)
0.8 (0.5, 1.5) 0.9 (0.7, 1.7)
0.8
Aspartate aminotransferase (IU/l)
27 (20, 47)
30 (22, 48)
0.5
Alanine transaminase (IU/l)
20 (13, 42)
20 (12, 33)
1.0
Total protein (g/dl)
6.7 (5.9, 7.4) 6.6 (5.9, 7.3)
0.8
Number of medications on admission ≥4,
38 (54)
223 (57)
0.6
n (%)
Number of medications on admission,
5 (3, 7)
5 (4, 8)
0.7
median (25%, 75%)
GCS, Glasgow Common Scale; NYHA, New York Heart Association. Table 1 (abstract P510). Patient characteristics knowledge regarding the law and ethics of donation (P <0.01), donor
management (P <0.01), living and deceased donor transplantation
(P <0.01) and joining Eurotransplant (P <0.01). Older professionals had
more information about all fi elds (P <0.01). Nurses had less knowledge
concerning donor management (P <0.01), law and ethics (P <0.01) and
deceased donor transplantation (P <0.01) than doctors. y
Reference 1. UK Organ Donation Taskforce: Organs for Transplants: A Report from the Organ
Donation Taskforce. Department of Health; 2008. Specialist nurse for organ donation in an emergency department
will increase organ donation Introduction In the UK, three people die each day awaiting trans plan-
tation, due to the unavailability of donor organs. Traditionally, donor
identifi cation has been restricted to the ICU. However, following the UK
Organ Donation Taskforce report in 2008 [1], a number of emergency
departments (EDs) have been working with specialist nurses for organ
donation (SN:OD) to identify potential donors and approach their
families for consent in the ED. We present our initial experience after
the introduction of a SN:OD to an Irish teaching hospital’s ED. Methods We conducted a retrospective review of deaths in our
ED during a 28-month period. For those who died in the ED, case
notes were reviewed to identify those suitable for organ donation. Referral and donation rates were compared in two cohorts, pre and
post introduction of a SN:OD. Fisher’s exact test was used to assess
diff erences between groups. f
Results Ninety-one deaths occurred in the study period. Following
introduction of the SN:OD, referrals increased from zero to eight. Of
the eight referred, three received consent and were transferred to the
ICU, two of whom became successful donors. The number of missed
potential donors fell from six to one (P = 0.009). Infl uence of adverse drug events on the mortality and the length of
hospital stay in ICUs in Japan: the JADE Study
Y Ohta1, M Sakuma1, D Bates2, T Morimoto1
1Kinki University, Osaka, Japan; 2Harvard University, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P510 (doi: 10.1186/cc12448) knowledge regarding the law and ethics of donation (P <0.01), donor
management (P <0.01), living and deceased donor transplantation
(P <0.01) and joining Eurotransplant (P <0.01). Older professionals had
more information about all fi elds (P <0.01). Nurses had less knowledge
concerning donor management (P <0.01), law and ethics (P <0.01) and
deceased donor transplantation (P <0.01) than doctors. Conclusion Education about organ donation needs to be part of
specialist training of intensive care staff , and refresher courses every
fi fth year as well. The course should include knowledge regarding brain
death, donor management and communication with family. This is the
fi rst step to improve the number of transplantations. Reference Sepsis in HIV patients admitted to the ICU
d l
1
l
1
l Sepsis in HIV patients admitted to the ICU
P Vidal-Cortés1, P Lameiro-Flores1, M Mourelo-Fariña2, A Aller-Fernández2,
R Gómez-López1, P Fernández-Ugidos1, M Alves-Pérez1, E Rodríguez-García1
1CHU Ourense, Spain; 2CHU A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P513 (doi: 10.1186/cc12451) Sepsis in HIV patients admitted to the ICU
P Vidal-Cortés1, P Lameiro-Flores1, M Mourelo-Fariña2, A Aller-Fernández2,
R Gómez-López1, P Fernández-Ugidos1, M Alves-Pérez1, E Rodríguez-García1
1CHU Ourense, Spain; 2CHU A Coruña, Spain
Critical Care 2013, 17(Suppl 2):P513 (doi: 10.1186/cc12451) nurses as regard donation and transplantation. Methods The self-completed questionnaire that consisted of 20 items
was completed at the Congress of Hungarian Society of Anesthesiology
and Intensive Therapy in 2011. Besides the epidemiological data, the
intensive care specialists (n = 179) and nurses (n = 103) were asked
about donation activity, participation in an organ donation course, self-
reported knowledge of joining Eurotransplant, donor management,
legislation, and transplantation. The data were analyzed by SPSS 17.0. Results A total of 53.6% of physicians and 16.7% of nurses attended
an earlier organ donation course (P <0.01). The average age of those
who participated in training was signifi cantly higher among doctors
(P <0.01). Fifty-nine percent of doctors and 65.1% of nurses did not
even want to participate in such training. Donation activity was
higher among staff who joined training (P <0.01). Independently
from accepting the presumed consent legislation (91.1%), 66% of
physicians agreed with the hospital practice that requests the adult
donor’s relatives to consent to organ recovery. This standpoint did
not depend on donation activity, participation in an organ donation
course, opinion about legislation and the nature of staff . A total
95.4% of participants consented to their organ retrieval after death. The staff who participated in an organ donation course had more g
Methods The self-completed questionnaire that consisted of 20 items
was completed at the Congress of Hungarian Society of Anesthesiology
and Intensive Therapy in 2011. Besides the epidemiological data, the
intensive care specialists (n = 179) and nurses (n = 103) were asked
about donation activity, participation in an organ donation course, self-
reported knowledge of joining Eurotransplant, donor management,
legislation, and transplantation. The data were analyzed by SPSS 17.0. Introduction Our objective was to analyze septic HIV patients admitted
to intensive care. Introduction Our objective was to analyze septic HIV patients admitted
to intensive care. Table 2 (abstract P515). Unit and hospital mortality Methods A retrospective study analyzed children with hematological
disorders admitted to our ICU between April 2005 and June 2012. All
of the included children required emergency admission and invasive
mechanical ventilation. Those who did not need intubation, or required
intubation only for therapeutic intervention and died within 24 hours
of ICU admission were excluded. The survival group was defi ned as
patients who were discharged from the ICU, and the nonsurvival
group was defi ned as those who died in the ICU or within 7 days after
discharge from the ICU. The PELOD score and PIM-II were applied as
morbidity scoring systems Patients
ICU mortality, Hospital mortality,
(n)
n (%)
n (%)
Haematology
163
52 (31.9%)
81 (49.7%)
Controls
237
73 (30.8%)
99 (41.8%)
P value
0.827
0.126 Conclusion Unit mortality of critically ill patients with HM was similar
to those without HM. Hospital mortality in patients with HM was higher
than those without HM, although not statistically signifi cant. Severity
of illness at presentation to critical care is the main determinant of
outcome in patients with HM. Results Twenty-seven patients, including 18 males and nine females,
with a median age of 6.1 years (range, 0.2 to 16.6 years) were analyzed. Sixteen patients had leukemia, fi ve had hemophagocytic syndrome,
six had solid tumors. The average predicted mortality rate was 31.3%
in PIM-II. The survival group included 15 patients (56%) and the
nonsurvival group included 12 patients (44%). When the survival group
was compared with the nonsurvival group, there were no signifi cant
diff erences in the systolic blood pressure (101.3 ± 13.9 mmHg vs. 92.8 ± 25.4 mmHg; P = 0.15), the proportion of patients requiring
continuous renal replacement therapy (33.3% vs. 50.0%; P = 0.30), and
PELOD score (15.5 ± 10.4 vs. 21.8 ± 15.4; P = 0.22). In the nonsurvival
group, the PIM-II was higher than that in the survival group (27.9 ± 10.4
vs. 35.7 ± 9.0; P = 0.06); the PaO2/FiO2 (272.5 ± 136.7 vs. 153.3 ± 123.3;
P = 0.03) and oxygenation index (6.7 ± 8.1 vs. 14.1 ± 9.5; P = 0.04) were
signifi cantly worse in the nonsurvival group than in the survival group. Conclusion The data show that respiratory failure is more strongly
associated with mortality than other organ failures in pediatric
hematology patients requiring intensive care. Sepsis in HIV patients admitted to the ICU
d l
1
l
1
l Mean viral load: 4.57 ± 3.25 log (vs. 2.257 ± 1.96, P = 0.001). Of sepsis patients, 62.1% were on their CD4 nadir (vs. 34.5% in
nonseptic patients, P = 0.009). Mean albumin levels were 2.3 ± 0.53 g/
dl (vs. 2.92 ± 0.94, P <0.001). APACHE II at admission was 21.98 ± 7.97
(vs. 18.15 ± 8.47, P = 0.046). At admission, 52.8% were on severe sepsis
and 44.3% on septic shock. The lung was the most frequent source of
infection (65.6%) followed by CNS (16.4%), UTI (4.9%) and IE (4.9%). The
most common pathogen isolated on these patients was S. pneumoniae
(28.8%), followed by P. jirovecii (13.6%), toxoplasma (8.5%), E. coli (5.1%)
and H. infl uenzae (5.1%). In total, 62.9% needed vasopressors (vs. 28.6%
in non-infected patients, P = 0.001), 79% mechanical ventilation (vs. 42.85%, P <0.001) and 19.4% renal replacement (9.5% in no septic
patients, P = 0.173). Mean ICU and hospital LOS was 10.43 ± 10.52 and
34.76 ± 29.64 days in septic patients versus 6.04 ± 8.45 (P = 0.026) and
20.54 ± 27.93 days (P = 0.016). ICU mortality: 33.9% (19% in nonseptic
patients, P = 0.098), hospital mortality: 41.9% (vs. 23.8%, P = 0.057). Conclusion Sepsis is a common reason for admission to the ICU in
HIV patients and is accompanied by high mortality. Pneumonia is the
most frequent source of infection. Septic patients are less frequently
under HAART and have a worse inmune status (lower CD4 count
and higher viral load). Despite a higher APACHE II, and a higher need
for hemodynamic and respiratory support, there is no statistically
signifi cant diff erence in ICU and hospital mortality between septic and
nonseptic patients. (2.4% in non-infected patients, P = 0.223) and 40.3% were under HAART
(64.3% in patients admitted without infection, P = 0.016). Mean CD4
count at admission: 219.62 ± 353.93 cells/mm3 (vs. 370.22 ± 362.56,
P = 0.048). Mean viral load: 4.57 ± 3.25 log (vs. 2.257 ± 1.96, P = 0.001). Of sepsis patients, 62.1% were on their CD4 nadir (vs. 34.5% in
nonseptic patients, P = 0.009). Mean albumin levels were 2.3 ± 0.53 g/
dl (vs. 2.92 ± 0.94, P <0.001). APACHE II at admission was 21.98 ± 7.97
(vs. 18.15 ± 8.47, P = 0.046). At admission, 52.8% were on severe sepsis
and 44.3% on septic shock. Table 2 (abstract P515). Unit and hospital mortality These results also suggest
that mechanical ventilation intervention in patients with respiratory
failure must occur earlier to improve the outcomes for these patients. Sepsis in HIV patients admitted to the ICU
d l
1
l
1
l The lung was the most frequent source of
infection (65.6%) followed by CNS (16.4%), UTI (4.9%) and IE (4.9%). The
most common pathogen isolated on these patients was S. pneumoniae
(28.8%), followed by P. jirovecii (13.6%), toxoplasma (8.5%), E. coli (5.1%)
and H. infl uenzae (5.1%). In total, 62.9% needed vasopressors (vs. 28.6%
in non-infected patients, P = 0.001), 79% mechanical ventilation (vs. 42.85%, P <0.001) and 19.4% renal replacement (9.5% in no septic
patients, P = 0.173). Mean ICU and hospital LOS was 10.43 ± 10.52 and
34.76 ± 29.64 days in septic patients versus 6.04 ± 8.45 (P = 0.026) and
20.54 ± 27.93 days (P = 0.016). ICU mortality: 33.9% (19% in nonseptic
patients, P = 0.098), hospital mortality: 41.9% (vs. 23.8%, P = 0.057). References 1. Hampshire PA, et al.: Crit Care 2009, 13:R137. 2
M
i
PB t l C it C
M d 2002 30 2260 2 References References References
1. Tamburro RF, et al.: Pediatr Crit Care Med 2008, 9:270-277. 2. Chima RS, et al.: Pediatr Crit Care Med 2012, 13:e336-e342. P515
Survival of critically ill patients with haematological malignancies
compared with patients without haematological malignancy
R Pugh1, P Hampshire2, P Hajimichael3
1Glan Clwyd Hospital, Rhyl, UK; 2Royal Liverpool University Hospital, Liverpool,
UK; 3Christie Hospital, Manchester, UK
Critical Care 2013, 17(Suppl 2):P515 (doi: 10.1186/cc12453) P515
Survival of critically ill patients with haematological malignancies
compared with patients without haematological malignancy
R Pugh1, P Hampshire2, P Hajimichael3
1Glan Clwyd Hospital, Rhyl, UK; 2Royal Liverpool University Hospital, Liverpool,
UK; 3Christie Hospital, Manchester, UK
Critical Care 2013, 17(Suppl 2):P515 (doi: 10.1186/cc12453) Introduction Critically ill patients with haematological malignancies
(HM) have high hospital mortality [1]. Severity of illness scores may
underestimate mortality in such patients [2]. Introduction Critically ill patients with haematological malignancies
(HM) have high hospital mortality [1]. Severity of illness scores may
underestimate mortality in such patients [2]. Methods Data collection was conducted at three hospitals from 2008
to 2011. Patients with any active HM condition were matched with
two control patients at two hospitals and with one control at Christie
Hospital. Control patients had the same APACHE II (within 2 points) and
admission diagnosis, but no HM. Readmissions and planned surgical
cases were excluded. p
p
y
Conclusion Sepsis is a common reason for admission to the ICU in
HIV patients and is accompanied by high mortality. Pneumonia is the
most frequent source of infection. Septic patients are less frequently
under HAART and have a worse inmune status (lower CD4 count
and higher viral load). Despite a higher APACHE II, and a higher need
for hemodynamic and respiratory support, there is no statistically
signifi cant diff erence in ICU and hospital mortality between septic and
nonseptic patients. Results A total of 163 patients with HM were compared with 237
control patients. Seventy-four admissions with HM were identifi ed at
two hospitals, and each was matched with two control patients. Eighty-
nine admissions with HM from Christie Hospital were identifi ed. These
were matched with 89 controls. Patients with HM spent signifi cantly
longer in hospital before ICU admission (Table 1). Unit and hospital
mortality rates were not statistically diff erent between patients with
HM and without HM (Table 2). Mortality related to respiratory failure among pediatric hematology
patients requiring intensive care
H Okuno, K Atagi, H Shimaoka
Osaka City General Hospital, Osaka, Japan
Critical Care 2013, 17(Suppl 2):P514 (doi: 10.1186/cc12452) Table 1 (abstract P515). Patient characteristics
Haematology
Controls
P value
Age, mean (SD)
56.5 (15.1)
57.1 (15.7)
0.717
APACHE II, mean (SD)
22.7 (6.1)
23.1 (6.0)
0.484
ICU LOS (median)
3.2 (1.8 to 6.2)
3.1 (1.3 to 8.4)
0.948
LOS prior (median)
6.5 (2 to 17)
2.0 (0 to 7)
<0.0001
bl
( b
P
)
i
d h
i
l
li Table 1 (abstract P515). Patient characteristics Table 1 (abstract P515). Patient characteristics H Okuno, K Atagi, H Shimaoka Introduction Although recent reports show an improvement in out-
comes for pediatric hematology patients requiring intensive care [1,2],
respiratory failure remains one of the major risks of pediatric mortality. This study was conducted to assess our hypothesis that mortality
associated with respiratory failure is higher than that for other organ
failures in pediatric hematology patients admitted to our ICU. Table 2 (abstract P515). Unit and hospital mortality Sepsis in HIV patients admitted to the ICU
d l
1
l
1
l Methods A retrospective study of HIV patients admitted to our ICU
between January 2005 and December 2009. We identify patients
admitted to the ICU with sepsis and analyze demographic factors,
etiology, organ failure and outcome, and we compare immune status,
frequency of organ failure and outcome between these patients and
those admitted for other reasons. We use Student’s t test to compare
quantitative variables, and the chi-square test for qualitative data. Methods A retrospective study of HIV patients admitted to our ICU
between January 2005 and December 2009. We identify patients
admitted to the ICU with sepsis and analyze demographic factors,
etiology, organ failure and outcome, and we compare immune status,
frequency of organ failure and outcome between these patients and
those admitted for other reasons. We use Student’s t test to compare
quantitative variables, and the chi-square test for qualitative data. Results A total of 104 HIV patients were admitted to our ICU during
the study period, 62 with sepsis (71% men, mean age: 40.59 ± 8.12). Of sepsis patients, 56.5% were admitted from the ER and 38.7% from
a medical ward; 66.1% had history of intravenous drugs use, other
comorbidities: COPD (9.7%), cirrhosis (8.1%), solid or hematologic
malignancy (12.9%); 8.1% patients were unaware of their condition Results A total of 53.6% of physicians and 16.7% of nurses attended
an earlier organ donation course (P <0.01). The average age of those
who participated in training was signifi cantly higher among doctors
(P <0.01). Fifty-nine percent of doctors and 65.1% of nurses did not
even want to participate in such training. Donation activity was
higher among staff who joined training (P <0.01). Independently
from accepting the presumed consent legislation (91.1%), 66% of
physicians agreed with the hospital practice that requests the adult
donor’s relatives to consent to organ recovery. This standpoint did
not depend on donation activity, participation in an organ donation
course, opinion about legislation and the nature of staff . A total
95.4% of participants consented to their organ retrieval after death. The staff who participated in an organ donation course had more S190 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (2.4% in non-infected patients, P = 0.223) and 40.3% were under HAART
(64.3% in patients admitted without infection, P = 0.016). Mean CD4
count at admission: 219.62 ± 353.93 cells/mm3 (vs. 370.22 ± 362.56,
P = 0.048). Automated, electronic monitoring for early detection of sepsis
S Rauch1, M Fischer1, C Martin2, A Sablotzki3 Critical Care 2013, 17(Suppl 2):P517 (doi: 10.1186/cc12455) Introduction In the year 2009, the organizational structure of the ICU
in the Kanazawa University Hospital changed from an open to a semi-
closed format ICU. The objective of this study was to evaluate the eff ect
of this organizational change on outcome in high-risk surgical patients. Methods The medical records of all consecutive high-risk surgical
patients admitted to the ICU from 2006 to 2009 (open format, n = 1,598)
and from 2009 to 2012 (semi-closed format, n = 1,521) were reviewed. Parameters studied were mortality and ICU length of stay. Introduction Early detection of sepsis is important for a suffi cient
treatment to reduce mortality. We hypothesized that using modifi ed
systemic infl ammatory response syndrome criteria over 1 hour using an
electronic software program facilitates the clinical diagnosis of sepsis. Methods After IRB approval and informed consent we enrolled in this
prospective, observational, single-center study 1,119 consecutive
patients (age 68.6 ± 16.4, female/male 476/649) admitted over a
6-month period to a surgical ICU. A total 149 of them met modifi ed
systemic infl ammatory response criteria. Patients were monitored by
an electronic software program using live data from the laboratory and
bedside monitors to detect modifi ed systemic infl ammatory response
syndrome criteria persisting over 1 hour. The physicians were blinded
to the software program alerts that notifi ed in real time when modifi ed
systemic infl ammatory response syndrome criteria were detected and
persisted over 1 hour, but did not provide treatment recommendations. Results There was a total of 149 modifi ed systemic infl ammatory
response syndrome criteria alerts. Seventy-four were confi rmed as
true sepsis cases by physicians. The overall incidence of sepsis was 7%. Patients were categorized into length of stay <24 hours, 24 to 96 hours
and >96 hours. The overall sensitivity of our system for detecting sepsis
was 68% and the specifi city was 91%. The positive predictive value is
34% and the negative predictive value is 98%. y
g
y
Results Mortality of ICU patients was 9.9% in the open format group
and 6.6% in the closed format group (P <0.05). The average length of
hospital stay was 4.9 days in the open format group and 4.8 days in the
closed format group. Outcome of critically ill patients with haematological malignancy
admitted to the ICU as an emergency T Lofaro1, S Easdale2, S Rowe2, M Ostermann2, R Carr2 1Queen Elisabeth Hospital, Woolwich, UK; 2Guys’ and St Thomas’ Hospital,
London, UK Introduction ICU admission policies regarding patients with haemato-
logical malignancy still vary despite data showing an acceptable
prognosis. Our aim was to review ICU and 6-month outcomes in this S191 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 group when requiring emergency admission to the ICU in a tertiary
cancer centre. Figure 1 (abstract P518). Starch administration between November
2008 and November 2012. Figure 1 (abstract P518). Starch administration between November
2008 and November 2012. Methods A retrospective review of medical notes between 2004 and
2012. Results A total of 249 patients were admitted, of whom 54 had more
than one admission. There were 310 episodes in total. Leukaemia n = 85;
lymphoma n = 90; myeloma n = 36. We compared the characteristics of
those who survived ICU admission with those who failed to survive to
discharge from ICU. The two populations were similar (age 51 vs. 57;
males 59% vs. 57%). Those who survived had a lower APACHE II score
on admission (19 vs. 23; P <0.001), lower mean organ failure scores (1
vs. 2; P <0.05), lower requirements of inotropes (26% vs. 50%; P = 0.001),
ventilation (31% vs. 64%; P = 0.001) and fi ltration (11% vs. 26%;
P = 0.004). There was no diff erence in the prevalence of sepsis at the
time of admission (64% vs. 70%). Both groups included patients with
prior bone marrow transplant (38% vs. 40%). Of note, ICU and 6-month
survival were 27% and 50%, respectively. These values are lower than
those reported in the literature to date. Figure 1 (abstract P518). Starch administration between November
2008 and November 2012. Methods Our e-prescribing software enables users to prescribe
intravenous fl uids from a series of menus. One of these is a template
that has several fl uids available to use as a bolus when instructed by
a clinician. We removed starch as an option from the template in April
2009. Starch could still be prescribed elsewhere on the prescribing
system. Data on the use of starch from November 2008 to November
2012 were analysed as the mean volume of starch infused per patient
per month. The mean of each set of parameters was then compared
using a Student’s t test. Outcome of critically ill patients with haematological malignancy
admitted to the ICU as an emergency Conclusion ICU and 6-month mortalities were 27% and 50%, respectively. Patients with haematological malignancy stand to benefi t from intensive
care, and should be off ered admission based on clinical need. References 1. Cuthberson et al.: The outcome of haematological malignancy in Scottish
intensive care units. J Intensive Care Soc 2008, 9:135-140. 2. Evison JM, et al.: Intensive care unit admission in patients with
haematological disease: incidence, outcome and prognostic factors. Swiss
Med Wkly 2001, 131:681-688. g
Results The mean volume of starch per patient administered before
and after electronic prescription options were altered was 480 ml and
21 ml, respectively (P = 0.004). See Figure 1. y
3. Beed et al.: Intensive care management of patients with haematological
malignancy. Conti Edu An Crit Care Pain 2010, 10:167-171. 4. Kleber et al.: Comorbidity as a prognostic variable in multiple myeloma:
comparative evaluation of common comorbidity scores and use of a novel
MM–comorbidity score. Blood Cancer J 2011, 1:e35. Conclusion Despite clinicians intending to reduce the use of starch it
was still regularly administered on our ICU. The removal of a default
prescribing option dramatically reduced the volume of starch used
whilst not restricting the ability to make a conscious choice to prescribe
it. Adjusting default options has potential to infl uence clinical decisions
and ensure more reliable, evidence-based care. Reference 5. McGrath S, et al.: ICU and 6-month outcome of oncology patients in the
intensive care unit. QJM 2010, 103:397-403. P517f 1. Schultz MJ, et al.: J Crit Care 2005, 20:199-204. Eff ect of organizational structure of the ICU on the prognosis:
open format versus semi-closed format
T Taniguchi, M Okajima
Kanazawa University Hospital, Kanazawa, Japan
Critical Care 2013, 17(Suppl 2):P517 (doi: 10.1186/cc12455) T Taniguchi, M Okajima P518 Use of defaults on an electronic prescribing tool infl uences the type
of fl uid received by patients
L Herbert, C Bordeaux, M Thomas, L Burrows, J Bewley, T Gould
Bristol Royal Infi rmary, Bristol, UK
Critical Care 2013, 17(Suppl 2):P518 (doi: 10.1186/cc12456) Use of defaults on an electronic prescribing tool infl uences the type
of fl uid received by patients Automated, electronic monitoring for early detection of sepsis
S Rauch1, M Fischer1, C Martin2, A Sablotzki3 Conclusion Our results suggest that a semi-closed format is a more
favorable setting than an open format to improve mortality in the ICU
and to warrant safe outcome in this patient group. P519 Kanazawa University Hospital, Kanazawa, Japan Automated, electronic monitoring for early detection of sepsis
S Rauch1, M Fischer1, C Martin2, A Sablotzki3
1Alb Fils Kliniken, Göppingen, Germany; 2Fa. Löser, Leipzig, Germany; 3Klinikum
Sankt Georg, Leipzig, Germany
Critical Care 2013, 17(Suppl 2):P519 (doi: 10.1186/cc12457) Use of defaults on an electronic prescribing tool infl uences the type
of fl uid received by patients l
y p
L Herbert, C Bordeaux, M Thomas, L Burrows, J Bewley, T Gould
Bristol Royal Infi rmary, Bristol, UK
Critical Care 2013, 17(Suppl 2):P518 (doi: 10.1186/cc12456) l
y p
L Herbert, C Bordeaux, M Thomas, L Burrows, J Bewley, T Gould
Bristol Royal Infi rmary, Bristol, UK yi
y
Critical Care 2013, 17(Suppl 2):P518 (doi: 10.1186/cc12456) Introduction Many evidence-based interventions are not delivered to
patients [1]. This may not be due to a clinician’s intentional decisions. The aim of this project was to compare the use of starch before and
after removing it as an option from an e-prescribing template. Conclusion Real-time alerts using an automated, electronic monitoring
of modifi ed systemic infl ammatory response syndrome criteria facili-
tate the clinical diagnosis of sepsis. S192 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 P520 score correlations between three raters were excellent (r = 0.80 to
0.94). Internal consistency in 98 videotaped patient presentations
was acceptable (α = 0.78). Application to 22 IDRs led by 14 diff erent
intensivists in three ICUs in two hospitals demonstrated that indicators
could be unambiguously rated. The staff and management of all three
ICUs that were rated had considered their IDRs to be adequately
performed, and they were surprised by these study results. score correlations between three raters were excellent (r = 0.80 to
0.94). Internal consistency in 98 videotaped patient presentations
was acceptable (α = 0.78). Application to 22 IDRs led by 14 diff erent
intensivists in three ICUs in two hospitals demonstrated that indicators
could be unambiguously rated. The staff and management of all three
ICUs that were rated had considered their IDRs to be adequately
performed, and they were surprised by these study results. Conclusion This study showed that the quality of IDRs can be reliably
assessed for patient plan of care and process. The IDR-Assessment
Scale had satisfactory inter-rater reliability, excellent overall item score
correlations, and acceptable internal consistency. Our instrument
may provide feedback for ICU professionals and managers to develop
adjustments in quality of care. Testing the IDR-Assessment Scale in
other ICUs may be required to establish general applicability. Reference P520
Admission to the ICU overnight: is it really a bad thing? Leadership training and quality improvement of interdisciplinary
rounds in the ICU EC Ten Have, JE Tulleken
Universitair Medical Center Groningen, the Netherlands
Critical Care 2013, 17(Suppl 2):P522 (doi: 10.1186/cc12460) Introduction The development of patient-centered care by inter-
disciplinary teams in the ICU has focused attention on leadership
behavior. The purpose of this intervention study was to measure
the eff ect of leadership training on the quality of performed inter-
disciplinary rounds (IDRs) in the ICU. Methods In this nonrandomized intervention study, participants
included nine intensive care medicine fellow trainees (intervention
group) and 10 experienced intensivists (control group). Participants
in the intervention and control groups previously were untrained in
leading IDRs in the ICUs. After each participant led an IDR that was
videotaped, the fellow trainees participated in a 1-day leadership
training, which was consistent with principles of adult learning and
behavioral modeling. After training, each fellow trainee led another
IDR that was videotaped. Quality of the performed IDRs was measured
by review of videotapes of the 19 IDRs lead by 19 intensivists, including
198 patient discussions subdivided into four ICUs, and assessment with
the IDR-Assessment Scale. Conclusion There is no signifi cant diff erence between the mortality of
patients admitted overnight and patients admitted during the day to
our unit. The hospital length of stay is increased in patients who are
admitted overnight to intensive care; however, ICU length of stay is
not aff ected. Adjustment for other confounders such as current bed
occupancy and staffi ng ratios during the entire patient stay may help
to understand the diff erences seen in the hospital length of stay. Reference 1. Kuijsten HA, et al.: Intensive Care Med 2010, 36:1765-1771. f
References References y
Methods A literature search was performed to identify criteria for
instruments about assessing team processes in the ICU. Then, 10
videotaped patient presentations led by diff erent intensivists were
analyzed by Delphi rounds. Appropriate and inappropriate behaviors
were highlighted. The IDR-Assessment Scale was developed and
statistically tested. The inter-rater reliability was evaluated by
rating nine randomly selected videotaped patient presentations by
three raters. Finally, the scale was applied to 98 videotaped patient
presentations during 22 IDRs in three ICUs for adults in two hospitals
in Groningen. 1. Ten Have ECM, et al.: Assessing the quality of interdisciplinary rounds in the
intensive care unit. J Crit Care 2013, in press. 1. Ten Have ECM, et al.: Assessing the quality of interdisciplinary rounds in the
intensive care unit. J Crit Care 2013, in press. 2. Miller A, et al.: Uni- and interdisciplinary eff ects on round and handover
content in intensive care units. Hum Factors 2009, 51:339-353. 3. Pronovost PJ, et al.: Perspective: physician leadership in quality. Acad Med
2009, 84:1651-1656. Reference 1. Reader TW, et al.: Developing a team performance framework for the
intensive care unit. Crit Care Med 2009, 37:1787-1793. y
g
y
Results Crude ICU and hospital mortality rates in patients admitted
during the day and overnight were examined. There was no signifi cant
diff erence in unit mortality (day 22.3% vs. night 22.7%, OR = 1.02, 95%
CI = 0.91 to 1.16, P = 0.718) or hospital mortality (day 30.7% vs. night
29.1%, OR = 0.93, 95% CI = 0.83 to 1.04, P = 0.203). The mean unit length
of stay showed no diff erence in patients admitted during daytime
compared with those admitted overnight (4.27 days vs. 4.09 days,
P = 0.162). The mean hospital length of stay was decreased in patients
admitted during daytime compared with patients admitted overnight
(19.3 days vs. 21.7 days, P = 0.004). The average age of patients was
less in those admitted out of hours (night 56.5 years vs. day 59.2 years,
P = <.0001). There was no signifi cant diff erence in APACHE II scores of
patients between the groups (day 19 vs. night 19, P = 0.580).if Assessing the quality of interdisciplinary rounds
EC Ten Have, JE Tulleken
Universitair Medical Center Groningen, the Netherlands
Critical Care 2013, 17(Suppl 2):P521 (doi: 10.1186/cc12459) Results Comparison of the intervention versus control groups shows
that the intervention group has more yes scores on the IDR-Assessment
Scale than the control group. This diff erence was signifi cant in 12 of the,
in total, 19 quality indicators. g
,
Critical Care 2013, 17(Suppl 2):P521 (doi: 10.1186/cc12459) Conclusion Quality of leadership will be reliably trained and measured
in the context of IDRs in ICUs. Training in a simulation environment, with
real-life IDR scenarios including confl icting situations, and workplace-
based feedback in the preparation and feedback phases, appears to be
eff ective to train leadership behaviour. Introduction Interdisciplinary rounds (IDRs) in the ICU are increasingly
recommended to support quality improvement and to reduce
confl icts, but uncertainty exists about assessing the quality of IDRs. We developed, tested, and applied a scoring instrument to assess the
quality of IDRs in ICUs. Use of defaults on an electronic prescribing tool infl uences the type
of fl uid received by patients M Adams, P Dean, K MacDowall, P Stenhouse, A Mackay
Victoria Infi rmary, Glasgow, UK
Critical Care 2013, 17(Suppl 2):P520 (doi: 10.1186/cc12458) Introduction Admission to hospital overnight has been shown
to increase mortality and decrease hospital length of stay [1]. The
objective of this study was to determine whether this relationship is
valid in patients admitted to our ICU, and whether length of stay was
aff ected. y
y
y
Conclusion This study showed that the quality of IDRs can be reliably
assessed for patient plan of care and process. The IDR-Assessment
Scale had satisfactory inter-rater reliability, excellent overall item score
correlations, and acceptable internal consistency. Our instrument
may provide feedback for ICU professionals and managers to develop
adjustments in quality of care. Testing the IDR-Assessment Scale in
other ICUs may be required to establish general applicability. Reference f
Methods A retrospective data collection identifi ed 5,827 patients
admitted to a fi ve-bed ICU from April 1994 to November 2012. Data
regarding patient age, sex, APACHE II score and ICU admission date
and time were collected along with the length of stay in the unit and
hospital. Defi nitions of day and night were set to local ICU standards of
9:00 am to 8:00 pm. Patients were then separated into two groups and
analysed using Analyse-It software for Excel. Let’s go round again! Quality improvement through intentional
roundingi g
Results The IDR-Assessment Scale had 19 quality indicators,
subdivided into two domains: Patient Plan of Care, and Process. The
domain Patient Plan of Care refl ects the technical performance from
the initial identifi cation of a goal to the evaluative phase. The domain
Process refl ects the team processes that are important to ensure that
the appropriate plan of care is agreed, understood, and executed as
planned by all care providers. Indicators were essential or supportive. The inter-rater reliability of nine videotaped patient presentations
among three raters was satisfactory (κ = 0.85). The overall item P Doyle, F Cox, R Tollyfi eld, A Serajii P Doyle, F Cox, R Tollyfi eld, A Serajii Harefi eld Hospital, Harefi eld, UK ii
Critical Care 2013, 17(Suppl 2):P523 (doi: 10.1186/cc12461) Introduction Harefi eld Hospital is a 150-bed cardiothoracic tertiary
referral centre with transplantation, artifi cial heart, ECMO and primary
angioplasty services. Our 35-bed critical care department consists of
18 intensive therapy unit, seven recovery and 10 high-dependency S193 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (572.08 ± 214.68 vs. 168.6 ± 97.27 seconds, P <0.001) but are also
associated with higher distractions particularly during evening shifts. Conclusion A higher percentage of nurses received handover training;
nursing handovers are longer and more inclusive of other components
of patient management; perceived importance of components of
handover varies among healthcare professionals; distractions are
common during handovers and associated with longer duration,
by nurses and in the evening shifts; and higher confi dence level in
patient’s management following the handover is associated with
better satisfaction. (572.08 ± 214.68 vs. 168.6 ± 97.27 seconds, P <0.001) but are also
associated with higher distractions particularly during evening shifts. beds. Intentional rounds or proactive patient rounds were recognised
by the Royal College of Physicians and the Royal College of Nursing [1]
as structured, evidence-based processes for nurses to carry out regular
checks with individual patients at set intervals. The senior nursing team
decided to adapt this initiative to the intensive care setting in order
to address clinical challenges and provide guidance for shift leaders to
focus on key elements of care. Methods Our intentional rounds, performed once per shift (twice daily),
include two components. First, pressure area care – this component
involves the shift leader checking whether key elements of pressure
sore prevention have been performed. P524 Prospective observational study of handover in a medical ICU
A Mukhopadhyay1, B Leong1, A Lua2, R Aroos2, J Wong2, N Koh2, N Goh2,
K See1, J Phua1, Y Kowitlawakul3
1National University Health System, Singapore; 2National University of
Singapore, Singapore; 3Alice Lee School of Nursing, National University of
Singapore, Singapore
Critical Care 2013, 17(Suppl 2):P524 (doi: 10.1186/cc12462) Results Over a 6-month period, 131 teleconsultations (114 patients)
were done. Mean age was 50.1 years, 57.1% was male and mean
APACHE II score was 24.3. A total of 64.8% originated from the ICU and
35.2% from the ED. Main consultation diagnoses were sepsis (31.3%);
stroke (29.8%); survival from cardiac arrest (6.1%); trauma (6.1%); and
acute myocardial infarction (5.3%). TM improved diagnosis in 14.5%
and infl uenced the clinical management in 85.5% of the consultations. Invasive procedures were indicated in 61.1%. Life-saving procedures
were TM related in seven patients (6.1%): stroke thrombolysis (n = 6)
and limb amputation (n = 1). Seven patients (6.1%) were transferred and
submitted to surgical procedures (heart surgery (n = 2), neurosurgery
(n = 4) and liver transplantation (n = 1)). The majority of the patients
remained at HMMD and were discharged. g p
g p
Critical Care 2013, 17(Suppl 2):P524 (doi: 10.1186/cc12462) Introduction Handovers are often associated with poor communi ca-
tion. ICU patients with multiple complex problems are ideal to study
naturally occurring handovers. However, few studies have been
conducted in the ICU. Methods We conducted questionnaires of physicians and nurses
involved and observed handovers in real time of medical ICU patients
over 1 month. Results We interviewed 580 of 672 physicians and nurses involved
(86.3%) and observed 90 real-time handovers (45 patients, 26.8%) of
168 patients. Mean duration of handover was 391.3 (± 263.6) seconds,
78.5% were face to face and 1.26 (± 1.75) distractions per handover
were noted, person-to-person calling being the commonest mode
of distraction (46.7%). Nurses received training during induction
in signifi cantly higher numbers, covered allied specialties more
and reviewed the patients early (all P <0.05). Perception of the
relative importance of diff erent components of the handover varied
signifi cantly between donors, recipients, physicians and nurses. P525 Impact of knowledge transfer through the implementation of a
telemedicine program in a community hospital in Brazil
CA Abreu Filho1, M Steinman1, A Andrade2, R Cal1, N Akamine1, J Teixeira2,
E Silva1, A Kanamura1, M Cenderoglo1, C Lottenberg1
1Hospital Israelita Albert Einstein, São Paulo, Brazil; 2Hospital Municipal Dr. Moysés Deutsch, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P525 (doi: 10.1186/cc12463) Results The incidence of pressure ulcers in the 4 months since the
initiative began has averaged 2.25 per month compared with 7.8 per
month prior to commencement of intentional rounding. Added to the
rounding tool at the end of September 2012, RRT rates in the preceding
4 months averaged 31.5 ml/kg/hour over 24 hours, an 11.9% reduction
from the previous average of 35.75 ml/kg/hour. If the pattern of RRT
was to continue, this could equate to a cost saving of UK£40,000 per
annum. 1Hospital Israelita Albert Einstein, São Paulo, Brazil; 2Hospital Municipal Dr. Moysés Deutsch, São Paulo, Brazil
C
l C
(S
l ) P
(d
/
) Introduction Emergency survival rates vary signifi cantly according
to the quality of care, which depends on human and technological
resources. Emergency and critical care medicine physicians must make
fast decisions; the presence of experienced consultants improves
survival. In developing countries, there is a shortage of skilled
doctors. The aim is to describe the fi rst Brazilian initiative of real-
time teleconferencing telemedicine (TM) providing 24/7 emergency
department (ED) and ICU coverage. Introduction Emergency survival rates vary signifi cantly according
to the quality of care, which depends on human and technological
resources. Emergency and critical care medicine physicians must make
fast decisions; the presence of experienced consultants improves
survival. In developing countries, there is a shortage of skilled
doctors. The aim is to describe the fi rst Brazilian initiative of real-
time teleconferencing telemedicine (TM) providing 24/7 emergency
department (ED) and ICU coverage. Conclusion The use of a modifi ed targeted intentional rounding tool
by the nursing shift leader can help ensure that best practice guidelines
are adhered to. This strategy can improve patient outcomes and
provide potentially signifi cant fi scal benefi ts. f 1. RCP, RCN: Ward Rounds in Medicine. Principles for Best Practice. London: Royal
College of Physicians, Royal College of Nursing; 2012. 1. RCP, RCN: Ward Rounds in Medicine. Principles for Best Practice. London: Royal
College of Physicians, Royal College of Nursing; 2012. References 1. Ye K, et al.: Handover in the emergency department: defi ciencies and
adverse eff ects. Emerg Med Australas 2007, 19:433-441 1. Ye K, et al.: Handover in the emergency department: defi ciencies and
adverse eff ects. Emerg Med Australas 2007, 19:433-441 2. Apker J, et al.: Communicating in the ‘gray zone’: perceptions about
emergency physician hospitalist handoff s and patient safety. Acad Emerg
Med 2007, 14:884-894. Let’s go round again! Quality improvement through intentional
roundingi These include completion
of the Waterlow risk assessment tool [2], noting the frequency of
repositioning, use of lateral positioning and pressure-relieving pads. Second, renal replacement therapy rates – this element was identifi ed
as an area for focus after we established that our haemofi ltration fl uid
use per hour of therapy was twice that of a near identical clinical setting. This pattern continued even after adopting similar therapy guidelines. The shift leader was guided to check whether therapy rates had been
adjusted in line with latest biochemical results. P525 g
Methods Since May 2012 a TM program has been implemented at
two hospitals in São Paulo, Brazil – Hospital Municipal Dr. Moysés
Deutsch (HMMD), a public, secondary hospital, and Hospital Israelita
Albert Einstein (HIAE), a tertiary private philanthropic entity – due to
a partnership with the Brazilian Health Ministry. TM Central Command
was located at HIAE with Endpoint 97 MXP Cisco® Solution and a
mobile Intern MXP ISDN/IP Cisco® for the remote hospital (HMMD) via
dedicated GB/sec connection. Imaging examinations were evaluated
using PACS technology. Every recruited patient was assessed by the
Central Command through TM with an experienced consultant. 2. Waterlow J: The importance of accurate risk assessment and appropriate
intervention in tissue viability. Br J Nurs 2009, 18:1090. 2. Waterlow J: The importance of accurate risk assessment and appropriate
intervention in tissue viability. Br J Nurs 2009, 18:1090. P524 Both
physicians and nurses seldom (39.7%) reviewed the available electronic
past medical records of the patient before handover, which in addition
to training in handover and overall confi dence level in the management
following handover are signifi cantly associated with better satisfaction
in univariate analysis; only the confi dence level in patient management
remained signifi cant after multivariate analysis. However, agreement
between donor and recipient on overall satisfaction was poor
(P >0.05). Nursing handovers were signifi cantly longer than physicians’ Conclusion A TM program is feasible to be implemented in a community
hospital. The major benefi t is expertise medical transfer from the
tertiary hospital to the community setting, improving diagnosis and
management of critical care patients, and avoiding routine transfer to
a major urban center. P526 P526
Utilization of telemedicine to improve burn care in a developing
country
J Knittel, G Fuzaylov
Massachusetts General Hospital, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P526 (doi: 10.1186/cc12464) Alternative to improve palliative care for all patients and families
in critical care units: development and preliminary evaluation
following MRC guidance of the King’s Psychosocial, Assessment and
Care toolf Results Since 2011 we have provided consultation on 14 patients in
Lviv, Ukraine, ranging in age from 15 months to 63 years. Each patient
had an average of six consultations. We present two of these cases as
examples of the capabilities of our telemedicine program. The fi rst
case involved a 15-month-old female with 40% TBSA from scald injury,
where telemedicine was instrumental in the primary assessment as
well as to arrange a direct assessment from a nearby burn surgeon. The
second case resulted from a house fi re with multiple casualties, where
physicians in Boston were able to utilize telemedicine to guide the
initial resuscitation and airway management of three critically burned
children, as well as to arrange for transport of one of the victims, an
11-year-old male with 87% TBSA, from Ukraine to the USA for acute
management. Multiple diffi culties were overcome in implementing the
system between the two countries including: time zone diff erences,
language barrier, and diff erent approaches to patient care. I Higginson, C Rumble, J Koff man, P Hopkins, S Heenen, W Prentice,
R Burman, S Leonard, O Dampier, J Noble, M Morgan, C Shipman
King’s Health Partners AHSC, London, UK Critical Care 2013, 17(Suppl 2):P528 (doi: 10.1186/cc12466) Introduction More than one in fi ve people admitted to an ICU will die
there. Research has highlighted concerns about support for patients
and families and decision-making in this context [1,2]. Here, we describe
the development and evaluation of a tool to improve palliative care in a
32-bed general ICU in a central London teaching hospital. Methods Medical Research Council guidance for complex interventions
Phase 0 to I comprised literature review, theoretical modelling, obser-
vation and qualitative interviews and focus groups with staff and
families exploring concerns and views of interventions identifi ed in
the literature review. Phase II comprised intervention development,
implementation and evaluation of tool feasibility and eff ects using staff
survey, observation, audit of records and relative survey.f f
Conclusion We have established a telemedicine program linking
physicians in Boston, MA, USA with City Hospital #8 in Lviv, Ukraine
to improve care in pediatric and adult burn patients. Our program has
provided consultation on 14 patients since 2011, and it highlights the
capabilities of telemedicine for acute consultation as well as triage and
transport of critically ill patients to tertiary-care facilities. Diff erent aspects of therapy limitation: a comparative study of the
nurse’s view
É O Szűcs1, G Élő1, L Szabó1, C Varga2, J Gál1, L Zubek1
1Semmelweis University, Budapest, Hungary; 2University of Kaposvár, Hungary
Critical Care 2013, 17(Suppl 2):P527 (doi: 10.1186/cc12465) O Szűcs1, G Élő1, L Szabó1, C Varga2, J Gál1, L Zubek1
1Semmelweis University, Budapest, Hungary; 2University of Kaposvár, Hungary
Critical Care 2013, 17(Suppl 2):P527 (doi: 10.1186/cc12465) Introduction During the last few years the frequency of end-of-
life decisions (EOLD) signifi cantly increased in ICUs. The method of
nurse involvement in making EOLD is diff erent worldwide [1,2]. The
purpose of this study was to analyze opinions of nurses about therapy
restriction. We have examined with a multicenter study the opinions of
the medical stuff about end-of-life care in Hungarian ICUs. Methods We performed a questionnaire evaluation among physicians
and nurses of ICUs about infl uencing factors of therapy restriction, the
method of the decision-making process, and the frequency of diff erent
EOLD. The questionnaire, containing 21 questions, was delivered
electronically to Hungarian ICUs, and then we analyzed the responses
anonymously. The retrieved 302 answers (191 physicians, 102 nurses)
were analysed using a nonparametric Student’s test. Conclusion K-PACE is a feasible tool to improve the palliative care of
patients and their families in the ICU. Further refi nement is needed and
planned, with consideration of roll-out into the wider medical centre. References 1. Azoulay E, et al.: Half the families of ICU patients experience inadequate
communication with physicians. Crit Care Med 2000, 28:3044-3049 1. Azoulay E, et al.: Half the families of ICU patients experience inadequate 1. Azoulay E, et al.: Half the families of ICU patients e communication with physicians. Crit Care Med 2000, 28:3044-3049.l Results A total 71% of the nurse responders work in university clinics,
2% in regional centrum, 24% in municipal hospital, 3% in other ICUs. The nurses found both human (2.72/5 vs. 1.98/5) and material (2.81/5
vs. 2.12/5) resources more restrictive factors during patient admission
than physicians (P = 0.025, P = 0.0024). Nurses working in municipal
hospital were more strongly infl uenced by lack of material and human
resources (3.34/5, 3.3/5) than nurses working in university clinics (2.2/5,
2.43/5), P = 0.01, P = 0.025. Younger nurses (working between 6 and
10 years) were more interested in the patient’s or surrogate’s wishes
than older nurses (working more than 10 years). P528 P528
Alternative to improve palliative care for all patients and families
in critical care units: development and preliminary evaluation
following MRC guidance of the King’s Psychosocial, Assessment and
Care tool
I Higginson, C Rumble, J Koff man, P Hopkins, S Heenen, W Prentice,
R Burman, S Leonard, O Dampier, J Noble, M Morgan, C Shipman
King’s Health Partners AHSC, London, UK
Critical Care 2013, 17(Suppl 2):P528 (doi: 10.1186/cc12466) country y
J Knittel, G Fuzaylov
Massachusetts General Hospital, Boston, MA, USA
Critical Care 2013, 17(Suppl 2):P526 (doi: 10.1186/cc12464) Introduction Our objective is to present our experience from Shriner’s
Hospital and Massachusetts General Hospital in Boston, MA, USA in S194 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 physicians slightly involved them in the end-of-life decision-making
process (2.1/5 vs. 2.4/5 P = 0.0001). using telemedicine to provide acute burn and critical care consultation
on pediatric and adult burn patients in Lviv, Ukraine, as well as in triage
and transport of critically ill patients from Lviv to a tertiary-care facility
in the USA for further management. using telemedicine to provide acute burn and critical care consultation
on pediatric and adult burn patients in Lviv, Ukraine, as well as in triage
and transport of critically ill patients from Lviv to a tertiary-care facility
in the USA for further management. Conclusion We found that the workplace, level of medical attendance,
godliness, work experience, and position in medical staff strongly
infl uenced making EOLD. While limitation of the therapy should
be team work, nurses felt their opinions were hardly taken into
consideration, although nurses seemed to be more realistic in the
decision-making process. Methods Using a new telemedicine learning center established at City
Hospital #8 in Lviv, Ukraine, consultations regarding acutely injured
burn victims occurred between physicians in Ukraine and physicians at
Shriners Hospital and Massachusetts General Hospital in Boston. After
the initial presentation, each patient was reviewed on a daily basis by
physicians in Boston. Skype, an Internet-based communication tool,
was used in communication with the Burn Center in Lviv. Radiographic
images were scanned and digitalized using an electronic scanner,
and JPEG image compression was used to facilitate the transmission
of radiographic images and patient charts. Informed consent and
HIPPA guidelines were followed in transmitting any patient-related
information. Diff erent aspects of therapy limitation: a comparative study of the
nurse’s view
É Religion did not
infl uence patient admission and forego therapy; however, religious
nurses compared with atheists and nonpracticing believers preferred
to prolong therapy against the patient’s will (P = 0.04). Nurses felt that communication with physicians. Crit Care Med l 2. Asch DA, et al.: Confl icts between physicians’ practices and patients’ wishes. Am J Respir Crit Care Med 1995, 151:288-292. P527f P527
Diff erent aspects of therapy limitation: a comparative study of the
nurse’s view
O Szűcs1, G Élő1, L Szabó1, C Varga2, J Gál1, L Zubek1
1Semmelweis University, Budapest, Hungary; 2University of Kaposvár, Hungary
Critical Care 2013, 17(Suppl 2):P527 (doi: 10.1186/cc12465) Diff erent aspects of therapy limitation: a comparative study of the
nurse’s view
É References References . Prendergast et al.: A national survey of end-of-life care for critical 1. Prendergast et al.: A national survey of end-of-life care for critic 1. Prendergast et al.: A national survey of end-of-life care for critically ill
patients. Am J Respir Crit Care Med 1998, 158:1165-1167. 2. Banbenishty et al.: Nurse involvement in end-of-life decision making: the
ETHICUS Study. Intensive Care Med 2006, 32:129-132. 2. Banbenishty et al.: Nurse involvement in end-of-life decision making: the
ETHICUS Study. Intensive Care Med 2006, 32:129-132. Alternative to improve palliative care for all patients and families
in critical care units: development and preliminary evaluation
following MRC guidance of the King’s Psychosocial, Assessment and
Care toolf y
y
Results Phase I: 47 staff and 24 family members were interviewed. The short time between decisions for treatment withdrawal and
death, plus concerns for support management, communication and
decision-making, highlighted a need to ensure excellent psychosocial
assessment for all. Phase II: as part of integrated care guidelines, we
developed the King’s Psychosocial Assessment and Care tool (K-PACE). K-PACE is used for all patients entering the ICU, completed within
24 hours of admission. It contains psychosocial assessment of the
family and patient needs, and identifi es key individuals for contact. Educational training was supported by K-PACE and was implemented
in two waves. Post-implementation survey of 95 ICU staff found that
most (80%) were aware of K-PACE. Eighty-two per cent of nurses but
only 17% of doctors had completed the tool. In total, 158/213 (74%)
family members responded to the survey (additionally three patients
responded). There were high levels of satisfaction for symptom control
and psychosocial care but concerns continued regarding explanation
of treatment and care. P529
d End-of life thoughts in the ICU: results of a survey
A Yaguchi, M Namiki, N Saito, R Nagai, M Takeda, T Harada, R Moroi
Tokyo Women’s Medical University, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P529 (doi: 10.1186/cc12467) End-of life thoughts in the ICU: results of a survey
A Yaguchi, M Namiki, N Saito, R Nagai, M Takeda, T Harada, R Moroi
Tokyo Women’s Medical University, Tokyo, Japan
Critical Care 2013, 17(Suppl 2):P529 (doi: 10.1186/cc12467) y
y
y
p
Critical Care 2013, 17(Suppl 2):P529 (doi: 10.1186/cc12467) Introduction The decision of terminal care in the ICU is a very
tough issue because the law, ethics, traditions and futility should S195 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 that withholding and withdrawing of treatment were ethically equal. A
DNR would be used most often. Withdrawal of inotropes or antibiotics
would be used more often than withdrawal of mechanical ventilation
or extubation. Termination of artifi cial hydration would be rarely used
in practice. be concerned involving the family’s will. Especially, stopping or
withdrawing therapy is a quite diffi cult operation in Japan because of
legal issues. Our hypothesis is that some diff erence exists in thoughts
between physicians and nurses for terminal patients in the ICU. The aim
of this study is to know their real thoughts. y
g
Methods A questionnaire survey was performed on physicians and
nurses in our medico-surgical ICU. The questionnaire consists of
11 questions with fi ve optional answers related to the thoughts of
participants about treatment of hopeless or brain death patients. Concretely, the questions were; whether to withhold therapy or not,
whether to accept to withdraw therapy or not and with family’s will,
whether to accept to immediately stop therapy and with family’s will,
whether to positively or not donate organs from a brain death patient,
necessity of ICU care for brain death patients, and feeling guilty and
stress for stopping or withdrawing therapy. The optional answer has
fi ve gradations from ‘Yes’ to ‘No’ for all questions. The participants were
asked to answer the questionnaire by expressing themselves without
regarding legal issues or the consensus. It was guaranteed to be
anonymous for them in the data analysis. The answers were compared
between physicians and nurses. The Mann–Whitney U test was used
for statistical analysis. P <0.05 was considered statistically signifi cant. P529
d P531 Attitudes of intensivists in the UK to withdrawal of futile therapy
M Mariyaselvam, M Irvine, J Carter, M Blunt, P Young
Queen Elizabeth Hospital, Kings Lynn, UK
Critical Care 2013, 17(Suppl 2):P531 (doi: 10.1186/cc12469) Introduction We aimed to determine the current practice and
attitudes of consultants in intensive care medicine when withdrawing
futile life-sustaining therapy. Published guidelines suggest variation
in withdrawal of futile life-sustaining therapy and are therefore not
prescriptive [1]. Although there is an awareness of diff ering practices,
the extent of these variations is not established. Methods We surveyed a convenience sample of delegates at the
Intensive Care Society (UK) State of the Art Meeting (2012) on attitudes
and practice regarding withdrawal practice. Anonymised data were
collected using surveymonkey.com. i
Results There were in total 52 participants (response rate 98.1%) with
20 physicians and 32 nurses. Withdrawing therapy was signifi cantly
accepted in nurses than in physicians (83% vs. 55%, P = 0.039), when the
family well understood. Withholding therapy should not be operated
for brain death patients for physicians (65%), while it seemed a diffi cult
judgement for nurses (23%, P = 0.021). ICU care for brain death patients
is less necessary for physicians than nurses (80% vs. 53%, P = 0.016). There were no signifi cant diff erences in other questions between
physician and nurses such as feeling guilty or stress for stopping or
withdrawing therapy.f Results Of 457 consultant attendees from the UK, 149 completed the
survey (33%). For 58% of consultants there was no formal institutional
protocol for withdrawal of futile therapy. When deciding to withdraw
therapy, 57% of consultants routinely seek and document a second
opinion. Regarding donation after cardiac death (DCD), 93% of
consultants were happy to delay withdrawal to facilitate successful
donation, 85% have already done so in their practice and 14%
routinely withdraw therapy in theatres rather than on the ICU. Even
if it would impact on the care of other patients, 48% would delay
withdrawal of therapy to facilitate DCD. For patients accepted for
DCD, 36% think that some intensivists withdraw more aggressively
(in essence, hasten death) in the hope of improving the likelihood of a
successful organ donation and 29% have felt pressurised to withdraw
therapy more quickly than their usual practice. Furthermore, 45%
experienced pressure to refer a patient for DCD when it they felt it
was not appropriate. End-of-life decisions in Slovenian ICUs: a cross-sectional survey S Grosek1, M Orazem2, M Kanic2, G Vidmar3, U Groselj1 Conclusion This survey confi rms variation in the practice and attitudes
to withdrawal of futile therapy amongst UK consultant intensivists. Formal protocols were frequently unavailable to guide withdrawal
and second opinions were often not sought. Nearly one-half of the
intensivists delay withdrawal to facilitate donation, even if this may
impact on the care of other patients. Many intensivists have felt
pressure to refer for donation when they feel this is inappropriate
and there is a perception that some intensivists may withdraw care
more aggressively in those who are accepted for DCD to improve the
likelihood of a successful donation. This survey may help inform debate
in this ethically challenging area. Critical Care 2013, 17(Suppl 2):P530 (doi: 10.1186/cc12468) Introduction The purpose of our study was to assess the attitudes of
Slovenian intensivists towards end-of-life (EOL) decision-making and to
analyze the decision-making process in their clinical practice. Methods A cross-sectional survey among Slovenian intensivists and
intensive care medicine residents from 35 diff erent ICUs was performed
using a questionnaire containing 43 questions about views on EOL
decision-making. Fisher’s exact test and the Fisher–Freeman–Halton
test were applied to cross-tabulated data; signifi cance level was set at P
≤0.001 due to the large number of tested hypotheses. Reference Reference g
yp
Results The response rate was 72.1% (267 questionnaires were
returned out of 370 distributed), which represented roughly the same
percentage of all Slovenian intensivists. Termination of futile treatment
was assessed as ethically acceptable (P <0.001). The statement that
there is no ethical distinction between withholding and withdrawing
of treatment could not be confi rmed (the answers ‘there is a diff erence’
and ‘undecided’ were less frequent, but not statistically signifi cant;
P = 0.216). A do-not-resuscitate order (DNR) was used more often than
other withholding treatment limitations (P <0.001). A DNR was used
most frequently in internal medicine ICUs (P <0.001; compared with
paediatric and surgical ICUs). Withdrawal of inotropes or antibiotics
was used more often than withdrawal of mechanical ventilation or
extubation (66.7% vs. 12.0%; P <0.001). Withdrawal of mechanical
ventilation or extubation was more often used in the paediatric ICUs
(21.7%) as compared with the internal medicine ICUs (19.6%) and the
surgical ICUs (3%) (P <0.001). Over two-thirds (70.6%) of intensivists
were against termination of hydration, which would be more often
used in the internal medicine ICUs (P <0.001). Thirty-one percent of
intensivists used written DNR orders. 1. Organ Donation after Circulatory Death 1. Organ Donation after Circulatory Death
[http://www.ics.ac.uk/professional/standards_and_guidelines/dcd] 1. Organ Donation after Circulatory Death
[http://www.ics.ac.uk/professional/standards_and_guidelines/dcd] P529
d Conclusion Some of end-of-life thoughts in the ICU showed diff erences
between physicians and nurses. P530 P530
End-of-life decisions in Slovenian ICUs: a cross-sectional survey
S Grosek1, M Orazem2, M Kanic2, G Vidmar3, U Groselj1
1University Medical Centre Ljubljana, Slovenia; 2 University of Ljubljana,
Slovenia; 3University Rehabilitation Institute, Republic of Slovenia, Ljubljana,
Slovenia
Critical Care 2013, 17(Suppl 2):P530 (doi: 10.1186/cc12468) Feasibility of neuromuscular electrical stimulation in critically ill
patients p
J Segers, G Hermans, F Bruyninckx, G Meyfroidt, D Langer, R Gosselink
UZ/KU Leuven, Belgium
Critical Care 2013, 17(Suppl 2):P535 (doi: 10.1186/cc12473) J Segers, G Hermans, F Bruyninckx, G Meyfroidt, D Langer, R Gosselink
UZ/KU Leuven, Belgium
Critical Care 2013 17(Suppl 2):P535 (doi: 10 1186/cc12473) Results Seven terminally ill patients wishing to go home for dying were
transferred home. Diagnoses varied: end-stage lung disease, cancer,
surgical complications. Ages ranged from 68 to 84 years. All patients
survived transport home, and time at home varied from a few hours to
4 days. Later contacts with patients’ families indicated that both patient
and family were grateful, and that they did not experience the patient
having pain or dyspnea at home. Introduction Survivors of critical illness often have a prolonged stay
on the ICU. These patients may suff er from ICU-acquired weakness. It
has been shown that reduction in muscle mass and muscle strength
occurs early after admission to the ICU. However, in the very early stage
on the ICU, patients are often sedated and not able to participate in
any active mobilizations. Therefore the use of neuromuscular electrical
stimulation (NMES) is becoming a treatment of interest in the ICU. The
aim was to study the feasibility and safety of NMES in a surgical and
medical ICU of a large, tertiary referral university hospital. Conclusion Sending critically ill patients home to die is not common. Anyhow, our experiences doing that are only positive. Terminally ill
patients, awake to make a decision of their own, and in a condition
making it possible, should have the choice to go home to die, with our
help in logistics, planning and transportation. Methods Fifty patients with an expected prolonged stay on the ICU
of 5 more days (judged on day 3) with no trauma or neurological
disease were included. They then received daily a NMES session (DUO
500; Gymna, Belgium) for 25 minutes on the quadriceps bilaterally
during their entire stay on the ICU. The main outcome was the ability
to produce a contraction of the quadriceps through NMES. The muscle
contraction was quantifi ed on a 5-point scale: 1 (no contraction
palpable and visible) up to 5 (contraction very well palpable and visible). Patients were classifi ed as responders when an adequate muscle bulk
was obtained in ≥75% of the sessions. Getting critical care patients home for end-of-life care
LS Nielsen
Kolding Sygehus, Kolding, Denmark
Critical Care 2013, 17(Suppl 2):P533 (doi: 10.1186/cc12471) Introduction Despite our eff orts in making patients healthy and going
home, critical illness has a mortality, in Danish ICUs, of 10 to 12%. Approximately 90% of deaths in ICUs happen after life-sustaining
therapy has been withheld or withdrawn. Although trying to provide
patients and family with what we suppose is a good death, most
patients would prefer dying at home, and sometimes patients and
family ask for the possibility of doing that. The last 2 years we transferred
seven patients from our unit to end-of-life care in their own home. Conclusion The 8-week exercise intervention resulted in statistically
signifi cant improvements in fi tness at 9 weeks while focus group
participants highlighted the positive eff ects of the exercise intervention
leading to enhanced energy levels, motivation and achievement. Psychological benefi ts of the exercise programme are apparent from
the focus group, emphasising the important link between physical and
mental health. Methods After making the decision of withholding or withdrawing
intensive care therapy, the care of the patient changes from an active,
medical, technological treatment to relief and care. In that period we
determine whether the patient’s condition is stable enough to go
home. We try to fi nd out if it is a wish for the patient to go home, and
if the family has resources to take care of the patient at home. If that is
the case, we start planning care at home, arrange for transportation,
and contact the primary care physician and nurse. Due to the patient’s
condition on the day of transferring, we plan following the patient by
either a nurse or a doctor. P535 Parent’s perception on end-of-life care in Brazil
P Lago, J Piva, G Halal, M Halal
Hospital de Clinicas de Porto Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P532 (doi: 10.1186/cc12470) Parent’s perception on end-of-life care in Brazil
P Lago, J Piva, G Halal, M Halal
Hospital de Clinicas de Porto Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P532 (doi: 10.1186/cc12470) Introduction The aim of the study is evaluate the perceptions of
parents of children who died in two Brazilian pediatric ICUs. Introduction The aim of the study is evaluate the perceptions of
parents of children who died in two Brazilian pediatric ICUs. Methods An exploratory-descriptive study with a qualitative approach
in the PICU of Hospital São Lucas and Hospital de Clinicas de Porto
Alegre involving 15 parents of children who died. Data collection was
performed through three steps: (a) the researchers contacted the
parents through a telephone call to invite them to attend the hospitals;
(b) at the hospital, the doctors who assisted the children clarifi ed doubts
about the therapy off ered; and (c) an interview was carried out by two
researchers not involved in the care. Data analysis was performed using
the technique of thematic content analysis. Conclusion Termination of futile treatment was found to be ethically
acceptable for Slovenian intensivists, although they were not convinced S196 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 arm of the study, we examined the patient experience after critical
illness, their views of the exercise programme and the eff ects on their
HRQoL. Results and conclusion The research shows that the diffi culty of com-
muni cation is a factor that impacts negatively on the grieving process. Moreover, it stresses the importance for parents to rediscuss the
moment of their child’s death with health professionals. Methods A randomised controlled trial was undertaken in adult
survivors of ICU admission. They were allocated to receive an
8-week in-hospital supervised aerobic programme consisting of two
cycle ergometry and one unsupervised session per week (exercise
group) or no exercise (control group). Primary outcomes were the
anaerobic threshold (in ml O2/kg mass/minute), physical function
and mental health scores (SF-36 questionnaire), measured at weeks
9 and 26. Participants were then allocated to focus groups where the
interpretation of experiences was compared with outcomes from the
PIX study. 1. Meyer E, Ritholz M, Burns J, Truog R: Improving the quality of end-of-life
care in the pediatric intensive care unit: parents priorities and
recommendations. Pediatrics 2006, 3:469-457. 2. Meyer E: On speaking less and listening more during end-of-life
conferences. Feasibility of neuromuscular electrical stimulation in critically ill
patients The potential factors associated
with the feasibility were: gender, age, body mass index (BMI), diagnosis
of sepsis, Barthel index prior to admission to the hospital, APACHE II
score, Glasgow Coma Scale (GCS), fi ve questions for adequacy, stimulus
intensity and leg edema. A multiple regression analysis was performed
to identify the factors determining whether or not a contraction could
be expected in a patient. Safety of NMES was assessed through heart
rate, blood pressure, oxygen saturation and respiratory rate. 1. Lusardi P, et al.: The going home initiative: getting critical care patients
home with hospice. Crit Care Nurse 2011, 31:46-57. P534 Project PIX (Post Intensive care eXercise): impact on physical
fi tness and focus group analysis of quality of life following exercise
rehabilitation Parent’s perception on end-of-life care in Brazil
P Lago, J Piva, G Halal, M Halal
Hospital de Clinicas de Porto Alegre, Brazil
Critical Care 2013, 17(Suppl 2):P532 (doi: 10.1186/cc12470) Crit Care 2004, 32:1609-1611. 3. Piva, JP, Lago P, Othero J, Garcia PC: Evaluating end of life in ten Brazilian
pediatric and adults intensive care units. J Med Ethics 2010, 36:344-348. 3. Piva, JP, Lago P, Othero J, Garcia PC: Evaluating end of life in ten Brazilian
pediatric and adults intensive care units. J Med Ethics 2010, 36:344-348. 4. Meert, K et al.: Parents’ perspectives on physician-parent communication
near the time of a child’s death in the pediatric intensive care unit. Pediatr
Crit Care 2008, 9:2-7. 4. Meert, K et al.: Parents’ perspectives on physician-parent communication
near the time of a child’s death in the pediatric intensive care unit. Pediatr
Crit Care 2008, 9:2-7. y
Results Fifty-nine patients were recruited to the study. The anaerobic
threshold increased at week 9 in the exercise group by a clinically and
statistically signifi cant amount of 2 ml O2/kg mass/minute (90% CI, 1
to 3 ml/kg/minute). There was further improvement in fi tness levels in
both groups by week 26 (although no signifi cant diff erence between
groups). No signifi cant diff erence in HRQOL measures between
groups was demonstrated; however, the exercise group did show an
improvement in their mental health scores. The focus groups centred
on feelings of isolation, abandonment, vulnerability, dependency and
reduced physical activity post hospital discharge. Many reported a
lack of social inclusion as they did not have the energy or confi dence
to venture outside. However, those in the exercise group felt that
the rehabilitation programme was motivating, built up confi dence,
improved fi tness, helped social interaction and gave them a sense of
achievement. P533
G
i Getting critical care patients home for end-of-life care
LS Nielsen
Kolding Sygehus, Kolding, Denmark
Critical Care 2013, 17(Suppl 2):P533 (doi: 10.1186/cc12471) Implementation of an ICU follow-up clinic: outcomes and patient
satisfaction after 1 year Implementation of an ICU follow-up clinic: outcomes and patient
satisfaction after 1 year Implementation of an ICU follow-up clinic: outcomes and patient
satisfaction after 1 year y
Results Sixty-two patients matched inclusion criteria and were
contacted. Thirty-three patients (90.9% males; age 38.7 ± 14.3 years;
mean SAPS II 33.75 ± 11.79) were interviewed. Mean ICU stay was
25.7 ± 15.3 days. Fifteen patients (45.5%) had undergone temporary
tracheostomy. Twenty-six patients were employed at the moment
of trauma: of these, 11 (42.3%) stated to be jobless and one was
pensioned at follow-up. In the whole cohort, the unemployment
rate increased from 12.1 to 45.6%. Mean PCS was 43.59 ± 6.43, mean
MCS was 41.04 ± 7.69. At follow-up, the 14 employed patients had a
lower MCS versus the 19 jobless patients (37.94 ± 7.14 vs. 43.33 ± 7.44;
P = 0.04), while no diff erences concerning PCS were found. The three
patients that were living on their own when the trauma occurred
showed, at follow-up, a lower mean MCS score towards the rest of the
cohort (32.2 ± 0.47 vs. 41.93 ± 7.51; P <0.001). Introduction The aim was to analyse the outcomes and patient
satisfaction of a recently implemented ICU follow-up clinic. These
clinics are National Institute for Clinical Excellence recommended [1]. Introduction The aim was to analyse the outcomes and patient
satisfaction of a recently implemented ICU follow-up clinic. These
clinics are National Institute for Clinical Excellence recommended [1]. Methods A retrospective analysis of prospective collected data
from January to December 2012. The clinic is run monthly by an ICU
consultant and a critical care outreach sister. Criteria to be invited to the
clinic are mechanical ventilation ≥3 days. Patients fi lled an anonymous
satisfaction survey after the clinic. Methods A retrospective analysis of prospective collected data
from January to December 2012. The clinic is run monthly by an ICU
consultant and a critical care outreach sister. Criteria to be invited to the
clinic are mechanical ventilation ≥3 days. Patients fi lled an anonymous
satisfaction survey after the clinic. y
Results Our attendance rate is 50% (26 patients), which is similar to
other series reported in the literature. Those patients who attended the
clinic required a longer length of mechanical ventilation (5.3 days vs. 7.1) and a longer length of stay in the ICU (7.6 vs. 13) and in hospital
(14 vs. 28). Follow-up and counseling service in trauma patients: needs and
goals – a preliminary study
DL Grieco, M Biancone, SL Cutuli, G Pintaudi, MT Santantonio,
ES Tanzarella, F Toni, AG De Belvis, MG Bocci, C Sandroni, M Antonelli
Catholic University School of Medicine, Rome, Italy
Critical Care 2013, 17(Suppl 2):P536 (doi: 10.1186/cc12474) g
p
y
y
DL Grieco, M Biancone, SL Cutuli, G Pintaudi, MT Santantonio,
ES Tanzarella, F Toni, AG De Belvis, MG Bocci, C Sandroni, M Antonelli
Catholic University School of Medicine, Rome, Italy
Critical Care 2013, 17(Suppl 2):P536 (doi: 10.1186/cc12474) Conclusion This survey demonstrates that the provision of ICU clinics
in the UK is not well established, with only 48% currently providing
a regular service. Currently 67% of clinics are not funded and further
resources should be employed so this service becomes an integral
part of the ICU pathway. Despite recent evidence demonstrating that
diaries reduce new-onset PTSD, only 44% of ICUs in our study provide
this service. Ninety-one per cent of ICUs do not provide family support
groups; similarly, it appears that fi nancial constraints are the limiting
factor. Introduction Trauma is the most common cause of morbidity in young
people. It has a high social impact both because of the high cost of
the acute treatments and because of the physical and psychological
consequences that it may cause. A prospective, observational, single-
center study on quality of life 12 to 24 months after trauma was carried
out. The aim of the study is to evaluate life quality after trauma and to
identify the most important needs of the patients, in order to improve
the level of care after an ICU stay and to implement a faster and more
eff ective reintegration into the active and productive society. References 1. DH and Modernisation Agency: The National Outreach Report. London: NHS
Modernisation Agency; 2003. 2. Jones C, et al.: Crit Care Med 2010, 14:168. 3. Orcutt TA:. Nurs Crit Care 2010, 5:33-37. 1. DH and Modernisation Agency: The National Outreach Report. London: NHS
Modernisation Agency; 2003. f
Methods All patients admitted to the 18-bed ICU of our university
hospital between 5 May 2010 and 28 February 2011 because of a
trauma who stayed in the ICU for at least 48 hours were eligible for the
study. Patients were contacted and invited to attend a multidisciplinary
interview. Data concerning family, working and economic status, main
disabilities and medical needs were collected. The SF12 questionnaire
was administered to defi ne physical component summary (PCS) and
mental component summary (MCS) scores. SPSS 20.0 was used to
perform statistical analysis. Project PIX (Post Intensive care eXercise): impact on physical
fi tness and focus group analysis of quality of life following exercise
rehabilitation Critical Care 2013, 17(Suppl 2):P534 (doi: 10.1186/cc12472) Introduction This study assessed the eff ect of a hospital-based aerobic
exercise programme on physical fi tness and health-related quality of
life (HRQoL) for survivors of ICU admission. Including the qualitative Results In 48% of the patients we were able to achieve adequate muscle
contractions in more than 75% of the sessions. GCS (P = 0.047), edema S197 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 (P = 0.001) and sepsis (P = 0.010) were signifi cantly diff erent between
responders and nonresponders. Responders had a lower mean GCS
(7 ± 3 vs. 9 ± 3), lower amount of edema and were less likely to have
had sepsis. In a multiple regression analysis, sepsis, edema, BMI and age
explained 51% of the variance. As for safety, none of the parameters
changed signifi cantly. incidence of new-onset PTSD [2]. Family support groups may also play
a pivotal role [3]. We performed a national survey via the Intensive Care
Society (ICS) to determine the provision of follow-up clinics, patient
diary services and family support groups. Methods An online structured questionnaire was sent via the ICS to all
ICU linkmen at 124 hospitals in the UK. Responses were received from
77 ICUs, a 62% response rate. Conclusion In patients with a better neurological condition, sepsis and/
or leg edema it was more diffi cult to obtain an adequate quadriceps
contraction with NMES. NMES is safe to apply on the ICU. Results Out of 77 ICUs, 37 (48%) run a follow-up clinic. The majority
of clinics (51%) only invite patients who have been admitted for more
than 3 to 4 days. Only 10 clinics (30%) receive funding from the ICU
budget or PCT, with the majority (67%) receiving no funding at all. Only
44% (34) of ICUs use patient diaries, mostly as a nonfunded service
(68%). Additionally, 91% of ICUs do not run family support groups; from
the minority that do, these are mostly held quarterly and are largely not
funded (55%). Implementation of an ICU follow-up clinic: outcomes and patient
satisfaction after 1 year We identifi ed a wide range of physical and nonphysical
morbidities on these patients (Figure 1). We referred them to the
appropriate specialities. Patients were very satisfi ed with this new
service (Figure 2). Conclusion Trauma is associated with an increase of the unemployment
rate in young people. Patients that are employed at follow-up seemed
to have a worse mental health status towards the unemployed ones. Conversely, the employed patients did not show any trend to a
lower physical health status. Thus a counseling service that provides
psychological care would be useful to implement more eff ective work
reintegration for trauma patients. Figure 1 (abstract P538). Physical and nonphysical morbidity. Follow-up and counseling service in trauma patients: needs and
goals – a preliminary study Follow-up and counseling service in trauma patients: needs and
goals – a preliminary study
DL Grieco, M Biancone, SL Cutuli, G Pintaudi, MT Santantonio,
ES Tanzarella, F Toni, AG De Belvis, MG Bocci, C Sandroni, M Antonelli
Catholic University School of Medicine, Rome, Italy
Critical Care 2013, 17(Suppl 2):P536 (doi: 10.1186/cc12474) Introduction Optimal patient evaluations of ICU rehabilitation therapy
remain unclear. Introduction Optimal patient evaluations of ICU rehabilitation therapy
remain unclear. Methods One hundred ICU patients with acute respiratory failure
were randomized to receive early rehabilitation (ER) or usual-care (UC). Cohort 1 (n = 50) received ER as one physical therapy (PT) session/day
versus UC; Cohort 2 (n = 50) received ER as 2 PT/day with the second
session resistance training, versus UC. UC was without ER. Blood was
drawn for cytokines through day 7. Cohort 2 underwent strength and
physical functional assessments using the Short Physical Performance
Battery (SPPB), a valid and reliable measure of physical function
consisting of walking speed, balance, and repeated chair stands. It is
a well-studied composite measure in older persons, but has not been
used in ICU survivors. Small changes of 0.5 to 0.6 points in the SPPB
have been shown to be clinically meaningful. Methods This study is a prospective randomised controlled trial with
a repeated-measures crossover design. Eligible participants, requiring
mechanical ventilation for 4 or more days, completed two exercise
activities routinely used in early critical care rehabilitation, a passive
chair transfer (PCT) and active sitting on the edge of the bed (SOEOB). The oxygen consumption and cardiovascular parameters were
measured to quantify and compare the exercise response between the
two activities. Results Data are presented as the median (interquartile range). Data for
fi ve patients have been collected, aged 68 years (23), with an ITU stay of y
g
Results Baseline parameters were similar between groups. Median
days from enrollment to fi rst PT were 4 (IQR 1 to 7.25). Deaths occurred
in eight UC subjects and four in ER (P = 0.22). For both arms, ventilator
days, ICU days and hospital days were not statistically diff erent. ER had
ventilator-free days of 22, 95% CI = 19.9 to 24.6, where UC had 22.3 days,
19.9 to 24.6, P = 0.99. ICU-free days for ER was 21, 95% CI = 19.1 to 23.6,
and that for UC was 21.0, 18.8 to 23.2, P = 0.84. Similarly, hospital days
for ER was 16.7, 95% CI = 11.8 to 21.4, and for UC was 18.2, 13.8 to 22.7,
P = 0.45. TNF, IL-6 and IL-8 through day 7 were not diff erent between
groups. Despite similar baseline acuity and infl ammatory profi les,
Cohort 2 ER group strength scores were numerically but not statistically
higher. P537 Post-traumatic stress disorder – are we doing all we can? A UK
national-based survey
X Watson
Ashford and St Peter’s Hospital, Surrey, UK
Critical Care 2013, 17(Suppl 2):P537 (doi: 10.1186/cc12475) Introduction Post-traumatic stress disorder (PTSD) is a well-recognised
complication in patients discharged from the ICU. ICU clinics have
been recommended to treat physical and psychosocial problems post
discharge, with guidelines issued by the Department of Health UK [1]. Recent evidence has advocated the use of patient diaries to reduce the Figure 1 (abstract P538). Physical and nonphysical morbidity. Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 S198 Critical Care 2013, Volume 17 Suppl 2
http://ccforum com/supplements/17/S2 pp
http://ccforum.com/supplements/17/S2 Figure 2 (abstract P538). Patient satisfaction survey. Figure 2 (abstract P538). Patient satisfaction survey. Conclusion Our follow-up clinic has enabled us to identify a wide range
of complications related to ICU admission and coordinate their future
management. This clinic improved patient satisfaction. Reference 15 days (10.5) and duration of mechanical ventilation 8 days (12), at the
point of intervention. Exercise response results are reported (Table 1). Conclusion Intensive care patients with prolonged mechanical
ventilation demonstrate a higher rate of oxygen consumption when
actively sitting on the edge of the bed, compared with a passive chair
transfer. This may have important consequences for early mobilisation
of the critically ill. 1. Rehabilitation after Critical Illness [www.nice.org.uk/CG83] References References
1. Unroe et al.: Ann Intern Med 2010, 153:167-175. 2. Thomas: Phys Ther Rev 2011, 16:46-57. Acute exercise response in the critically ill
N Collings1, R Young2 g
g
1University Hospital Southampton, UK; 2Sheffi eld Hallam University, Sheffi eld,
UK Randomized, controlled pilot study of early rehabilitation strategies
in acute respiratory failure Critical Care 2013, 17(Suppl 2):P539 (doi: 10.1186/cc12477) Introduction This study aims to quantify the acute exercise response to
early passive and active activities in order to inform exercise prescription
when designing rehabilitation programmes for the critically ill. Critical
care survival is often associated with a poor functional outcome [1],
with recent investigations presenting the case for early rehabilitation
in order to optimise functional recovery [2]. There, remains, however,
a scarcity of research investigating the immediate response to exercise
and subsequent exercise prescription, in the acute phase following
critical illness. p
y
D Files, P Morris, S Shrestha, S Dhar, M Young, J Hauser, E Chmelo,
C Thompson, L Dixon, K Murphy, B Nicklas, M Berry
Wake Forest University School of Medicine, Winston Salem, NC, USA
Critical Care 2013, 17(Suppl 2):P540 (doi: 10.1186/cc12478) D Files, P Morris, S Shrestha, S Dhar, M Young, J Hauser, E Chmelo,
C Thompson, L Dixon, K Murphy, B Nicklas, M Berry
Wake Forest University School of Medicine, Winston Salem, NC, USA
Critical Care 2013, 17(Suppl 2):P540 (doi: 10.1186/cc12478) Introduction Optimal patient evaluations of ICU rehabilitation therapy
remain unclear. Grip strength, as a percentage of predicted for ER was 66 versus Table 1 (abstract P539)
Parameter
PCT (n = 5)
SOEOB (n = 5)
Oxygen consumption (ml/minute)
270 (46.5)
334 (121)
Carbon dioxide production (ml/minute)
142 (45.5)
185 (121)
Heart rate (bpm)
92 (24)
77.5 (29)
Mean arterial pressure (mmHg)
100.3 (20)
97.3 (13)
Minute ventilation (l/minute)
11.0 (3.5)
12.1 (3.4) S199 Critical Care 2013, Volume 17 Suppl 2
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http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 39 for UC, P = 0.06. Dynamometry for ER was 211 versus 181 lbs for UC,
P = 0.124. Although the diff erence in SPPB values for ER versus UC (5
vs. 3, P = 0.172) was not statistically diff erent, it was greater than the
minimal clinically signifi cant diff erence. There were no diff erences in
adverse events. or alleviate this stress [4]. To action this it is important to identify the
main stressors from the patient’s perspective. A systematic review
was performed to provide a list of what patients consider stressors
in intensive care. These were then ranked in order to provide an
identifi cation tool that can be used to shape appropriate care. Conclusion In this pilot study, early ICU rehabilitation was safe, and was
associated with numerically although not statistically shorter hospital
stay, greater strength and improved functional scores. Particularly, the
SPPB demonstrated discriminatory ability in groups of ICU survivors
with low physical function. Future early ICU rehabilitation studies
should consider ICU survivor assessments using the SPPB due to its
ease, reproducibility and discriminatory ability following ICU and
hospital discharge. i
Methods A systematic review was performed using MEDLINE, CINAHL,
Psych INFO and Academic Search Complete. Grey literature was
included and searches were not restricted to type of intensive care or
country. Criteria were used to fi lter those articles that identifi ed the
patients’ views of their stressor, not the patient experience. Eligible
articles were critiqued using the Critical Appraisal Skills Programme for
qualitative studies [5] and brought together using a narrative synthesis. All of the reviewed studies used a questionnaire as a means to identify
what elements on the intensive care patients found stressful. A list
of the top-10 stressors could then be expressed for each study and
compared. From this information, a set of guidelines for best practice
were devised. y
References y
p
References 1. Baker C, et al.: Physical rehabilitation following critical illness. J Intensive Care
Soc 2008, 9:166-169. 2. Jones C, et al.: Long term outcome from critical illness. Anaesth Intensive
Care Med 2006, 7:155-156. Results In six hospitals the CQI ‘R-ICU’ is sent to relatives after receiving
informed consent (n = 441), 55.1% of the respondents are the patient’s
partner. Respondents seem to be most satisfi ed with the presence
of a professional at fi rst entrance to the ICU. The highest need for
improvement scores relate to information about meals, parking
and other disciplines (for example, social worker, spiritual worker or
psychologist). Factor analysis shows that quality of care is determined
by four clusters of items: Support, Communication, General Information
and Organisation. The reliability of the CQI ‘R-ICU’ is suffi ciently high,
only Communication and Support are signifi cant predictors of total
quality judgement of relatives (adj. R2 = 0.74). In addition, there is a
signifi cant diff erence in mean total quality judgement between the six
hospitals as well as between the four wards within Erasmus MC. None 3. CG83 Critical Illness Rehabilitation: Guideline 2009 [www.nice.org.uk/CG83] P543
Quality of care in the ICU from the perspective of relatives
M Van Mol1, E Bakker2, A Rensen3, I Menheere3, L Verharen3
1Erasmus MC, Rotterdam, the Netherlands; 2OU, Heerlen, the Netherlands;
3HAN, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P543 (doi: 10.1186/cc12481) P543
Quality of care in the ICU from the perspective of relatives
M Van Mol1, E Bakker2, A Rensen3, I Menheere3, L Verharen3
1Erasmus MC, Rotterdam, the Netherlands; 2OU, Heerlen, the Netherlands;
3HAN, Nijmegen, the Netherlands
Critical Care 2013, 17(Suppl 2):P543 (doi: 10.1186/cc12481) Introduction This study describes the development and validation
of the Consumer Quality Index Relatives in ICUs (CQI ‘R-ICU’), which
aims to measure the satisfaction of relatives and to identify aspect
of care that need improvement in the ICU in a reliable and valid way. According to the quality standards of the Dutch Society of Intensive
Care, every ICU needs to record the satisfaction of relatives [1]. At this
moment there is insuffi cient insight into the quality of care off ered to
relatives on the ICU because an evidence-based Dutch measurement
instrument is missing. Results At baseline measurements, there were no statistical diff erences
in age, gender, length of stays or APACHE II scores between the two
groups. Results indicate that the exercise group (n = 10) had signifi cantly
greater improvements in cardiopulmonary fi tness (P <0.001) and
balance (P <0.05) compared with the control group (n = 10). Greater
improvements were also evident in anxiety, depression and grip
strength in the exercise group, although not statistically signifi cant. i
Conclusion This pilot study highlights that a 6-week supervised
exercise programme can signifi cantly improve cardiopulmonary fi tness
and balance in post-ICU patients. Further recruitment to the study and
6-month/1-year follow-up is needed. Methods The CQI ‘R-ICU’ has been developed based on a scientifi c and
standardised method [2]. A mixed design method is used, consisting of
qualitative and quantitative survey studies. Factor analyses are carried
out to determine the underlying structure of the newly developed
questionnaire. Multiple regression analysis is used to explore the
relationship between demographic variables and the perceived quality
of care. P541 P541
Early results of a 6-week exercise programme in post-ICU patients
C Battle1, K James1, P Temblett1, H Hutchings2
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK
Critical Care 2013, 17(Suppl 2):P541 (doi: 10.1186/cc12479) Early results of a 6-week exercise programme in post-ICU patients
C Battle1, K James1, P Temblett1, H Hutchings2
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK
Critical Care 2013, 17(Suppl 2):P541 (doi: 10.1186/cc12479) Results A total 1,424 articles were systematically assessed for suitability
and applicability. Of these, 14 articles remained eligible for review. The
stressors were ranked by their frequency in the individual studies’ top-
10 lists. In rank order: 1, Tubes. 2, Pain. 3, Sleep Diffi culties. 4, Thirst. 5, Lack of Patient Understanding. Overall, stressors were found to be
similar throughout all of the studies.i Early results of a 6-week exercise programme in post-ICU patients
C Battle1, K James1, P Temblett1, H Hutchings2 g
1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK 1Morriston Hospital, ABMU Health Board, Swansea, UK; 2Swansea University,
Swansea, UK Introduction The aim of this study was to investigate the eff ect of
a 6-week exercise programme on outcomes in post-ICU patients. With improvements in intensive care medicine, increasing numbers
of patients are surviving catastrophic illness [1]. Severe weakness
is common in patients with prolonged critical illness and results in
considerable morbidity, mortality and healthcare costs [2]. The NICE83
guidelines Rehabilitation in Critical Care recommend follow-up for
post-ICU patients and that further research is needed in this fi eld [3]. Conclusion The review identifi ed a list of the most pertinent common
stressors. Awareness of these and ranking in priority may enable plans
of care to be instigated to eff ectively alleviate patient stress. References References
1. Jones C, et al.: Clin Intensive Care 1998, 9:199-205. 2. Scragg P, et al.: Anaesthesia 2001, 56:9-14. 3. Perrins J, et al.: Intensive Crit Care Nurs 1998, 14:108-116. 4. Ballard KS: Issues Ment Health Nurs 1981, 3:89-108. 5. Public Health Resource Unit: Critical Appraisal Skills Programme. 10 Questions
to Help You Make Sense of Qualitative Research. [http://www.medev.ac.uk/
static/uploads/workshop_resources/166/166_Qualitative_Appraisal_Tool.pdf] References
1. Jones C, et al.: Clin Intensive Care 1998, 9:199-205. 2. Scragg P, et al.: Anaesthesia 2001, 56:9-14. 3. Perrins J, et al.: Intensive Crit Care Nurs 1998, 14:108-116. 4. Ballard KS: Issues Ment Health Nurs 1981, 3:89-108. 5. Public Health Resource Unit: Critical Appraisal Skills Programme. 10 Questions
to Help You Make Sense of Qualitative Research. [http://www.medev.ac.uk/
static/uploads/workshop_resources/166/166_Qualitative_Appraisal_Tool.pdf] 1. Jones C, et al.: Clin Intensive Care 1998, 9:199-205. i
Methods Patients who have been discharged home from hospital
following an ICU stay of 48 hours or more were recruited to the study. Patients were only excluded if they were not considered safe for
exercise. Baseline measurements were completed prior to stratifi ed
(age, gender, APACHE II score) random allocation to either the exercise
or control group. Outcome measures included cardiopulmonary
fi tness (6-minute walk test), balance (Berg Balance Scale), grip strength
(JAMAR grip dynamometer) and hospital anxiety and depression (HAD
score). The exercise group completed a 6-week supervised exercise
programme, twice a week for up to 1 hour. In the seventh week, all
patients repeated the baseline measurements. An unpaired Student’s
t test was used to compare any diff erences between the control and
exercise groups.f Introduction Optimal patient evaluations of ICU rehabilitation therapy
remain unclear. Acknowledgement Supported by NIH grants 1R011186-01 and P30
AG21332. Family satisfaction in the ICU: enhancing patient experience
E Kursumovic, J Bilinska, A Molokhia
University Hospital Lewisham, London, UK
Critical Care 2013, 17(Suppl 2):P544 (doi: 10.1186/cc12482) p
p
J Othero, AB Cavalcanti, JC Mouro, K Normilio-da-Silva, R Pusch, F Moreira,
AA Kodama, O Berwanger
Hospital do Coração – HCor, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P545 (doi: 10.1186/cc12483) Introduction Following recommendations made by the Scottish
Intensive Care Society [1], we have introduced an annual Family
Satisfaction Survey in the intensive care unit (FS-ICU), in anticipation
that this will become an integral part of how we measure, improve
and strive to advance patient care in the future. We aim to recognise
factors impacting patient care and highlight areas for improvement. Improving understanding of patients’ needs is currently a UK national
goal for CQUIN 2011/2012. An estimated £6.9 billion per year can be
saved by reaching these goals [2] and a positive patient experience
results in improved long-term outcomes and shorter hospital stays [2]. Methods We performed a study to assess Family Satisfaction over a 10-
week period in a 16-bed critical care unit (CCU). A modifi ed version of
an FS-ICU published in US and Canadian studies [3] was distributed to
up to two family members per patient. Five-point Likert scale responses
were linearly transformed to give percentage scores. Higher values
represented a greater degree of satisfaction. Introduction We are conducting a cluster randomized trial with two
parallel arms to evaluate strategies to improve family satisfaction with
the care that themselves and their critically ill relatives receive in the
ICUs of nonacademic Brazilian public hospitals. Here we report the
results of the baseline phase of this trial. Introduction We are conducting a cluster randomized trial with two
parallel arms to evaluate strategies to improve family satisfaction with
the care that themselves and their critically ill relatives receive in the
ICUs of nonacademic Brazilian public hospitals. Here we report the
results of the baseline phase of this trial. Methods In this baseline phase, we interviewed the family member
most closely involved with the care of critically ill patients who stayed
in the ICU for at least 72 hours. We applied a form with 24 questions
divided into four domains: overall ICU experience, communication,
decision-making, and questions related to end-of-life care for patients
who died in the ICU. Each question scored from 0 (very poor) to 100
(excellent). The form was adapted from the Family Satisfaction with
Care in the ICU (FS-ICU 24). Recognition of the primary stressors aff ecting intensive care
patients: a systematic review p
y
S Birch, S Elliot S Birch, S Elliot
Leeds Teaching Hospitals, Leeds University Teaching Hospitals Trust, Leeds, UK
Critical Care 2013, 17(Suppl 2):P542 (doi: 10.1186/cc12480) ,
Leeds Teaching Hospitals, Leeds University Teaching Hospitals Trust, Leeds, UK
Critical Care 2013, 17(Suppl 2):P542 (doi: 10.1186/cc12480) Introduction Intensive care patients suff er psychological and
physiological distress that may have debilitating and long-lasting
eff ects [1-3]. Healthcare professionals are in a position to help avoid S200 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 Critical Care 2013, Volume 17 Suppl 2
http://ccforum.com/supplements/17/S2 References 3. Wall et al.: Refi nement, scoring, and validation of the Family Satisfaction in
the Intensive Care Unit (FS-ICU) survey. Crit Care Med 2007, 35:271-279. 1. Vos et al.: Quality measurement at intensive care units: which indicators
should we use? J Crit Care 2007, 22:267-274. 2. Sixma et al.: Handboek CQI Ontwikkeling: richtlijnen en voorschriften voor de
ontwikkeling van een CQI meetinstrument. Utrecht: Nivel; 2008. 2. Sixma et al.: Handboek CQI Ontwikkeling: richtlijnen en voorschriften voor de
ontwikkeling van een CQI meetinstrument. Utrecht: Nivel; 2008. References of the demographic variables such as sex, age, education, race and
length of stay had an eff ect on perceived quality of care. 1. Quality Indicators for Critical Care in Scotland [http://www.sicsag.scot.nhs. uk/SICSQIG-report-2012-120209.pdf] 1. Quality Indicators for Critical Care in Scotland [http://www.sicsag.scot.nhs. uk/SICSQIG-report-2012-120209.pdf] 1. Quality Indicators for Critical Care in Scotland [http://www.sicsag.scot.nhs. uk/SICSQIG-report-2012-120209.pdf] f
Conclusion The CQI ‘R-ICU’ turned out to be a valid, reliable, sensitive
and feasible instrument. Large-scale implementation is recommended. Acknowledgements Thanks to researchers of Ziekenhuis Gelderse
Vallei, Kennemer Gasthuis, Catharinaziekenhuis, Ziekenhuis Rivieren-
land and Scheperziekenhuis. References 2. Using the Commissioning for Quality and Innovation (CQUIN) Payment
Framework – Guidance on National Goals for 2011/12 [http://www.dh.gov. uk/prod_consum_dh/groups/dh_digitalassets/@dh/@en/@ps/documents/
digitalasset/dh_123009.pdf] 2. Using the Commissioning for Quality and Innovation (CQUIN) Payment
Framework – Guidance on National Goals for 2011/12 [http://www.dh.gov. uk/prod_consum_dh/groups/dh_digitalassets/@dh/@en/@ps/documents/
digitalasset/dh_123009.pdf]i Family satisfaction with patient care in ICUs of nonacademic
Brazilian public hospitals Family satisfaction with patient care in ICUs of nonacademic
Brazilian public hospitals
J Othero, AB Cavalcanti, JC Mouro, K Normilio-da-Silva, R Pusch, F Moreira,
AA Kodama, O Berwanger
Hospital do Coração – HCor, São Paulo, Brazil
Critical Care 2013, 17(Suppl 2):P545 (doi: 10.1186/cc12483) Family satisfaction in the ICU: enhancing patient experience
E Kursumovic, J Bilinska, A Molokhia
University Hospital Lewisham, London, UK
Critical Care 2013, 17(Suppl 2):P544 (doi: 10.1186/cc12482) As many questions assessed the quality
of intensivist care or communication, the interview was applied by a
psychologist or a nurse. y
g
Results Families of 564 patients were interviewed. A total 45/564
(8.3%) died in the ICU. Most respondents were satisfi ed with overall ICU
experience (mean ± SD score 85.5 ± 11.9). However, family satisfaction
with communication (67.8 ± 18.0) and decision-making (69.5 ± 21.1)
resulted in somewhat lower scores. Most families of patients who
died in the ICU (38/45 (82.6%)) considered that their relative’s life
was neither extended nor shortened unnecessarily. Also, most of the
families believed that their relative did not suff er or suff ered little in the
ICU (37/46 (80.4%)) and felt supported by the healthcare team (40/46
(87.0%)).i Results We received and analysed 32 completed surveys. Seventy-
eight per cent of relatives reported that the treatment of the patient
was excellent, including symptom control such as pain, breathlessness
and agitation (72%, 69%, 100%, respectively). Ninety-one per cent
felt the care and frequency of communication provided by nurses
was excellent or very good, compared with 81% by doctors. Only 9%
of family members were off ered spiritual support during their CCU
experience. Forty-seven per cent felt they would have liked more
involvement in the decision-making process. Conclusion Most families were satisfi ed with the care themselves and
their critically ill relatives received in the ICU. Also, most relatives of
patients who died in the ICU felt that end-of-life care was adequate. Although we believe there is much room for improvement in
communication, decision-making and support critically ill patients and
their families, as their baseline satisfaction with patient care is quite
high, it may be hard to demonstrate substantial improvement after
interventions. Conclusion The vast majority of relatives rated their overall experience
on CCU as excellent or very good. The study highlighted two main areas
for improvement: provision of spiritual support as well as family and
patient involvement in the decision-making process. We will arrange
multi-disciplinary teaching sessions focusing on the positive impact,
and therefore importance of patient satisfaction. A poster providing
information about available spiritual support will be displayed around
our CCU. Further studies are required to evaluate these measures.
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Types of teacher-AI collaboration in K-12 classroom instruction: Chinese teachers’ perspective
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Education and information technologies
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* Jinhee Kim
jhkim@odu.edu Education and Information Technologies (2024) 29:17433–17465
https://doi.org/10.1007/s10639-024-12523-3 Education and Information Technologies (2024) 29:17433–17465
https://doi.org/10.1007/s10639-024-12523-3 Abstract The advancing power and capabilities of artificial intelligence (AI) have expanded
the roles of AI in education and have created the possibility for teachers to collabo-
rate with AI in classroom instruction. However, the potential types of teacher-AI
collaboration (TAC) in classroom instruction and the benefits and challenges of
implementing TAC are still elusive. This study, therefore, aimed to explore differ-
ent types of TAC and the potential benefits and obstacles of TAC through Focus
Group Interviews with 30 Chinese teachers. The study found that teachers antici-
pated six types of TAC, which are thematized as One Teach, One Observe; One
Teach, One Assist; Co-teaching in Stations; Parallel Teaching in Online and Offline
Classes; Differentiated Teaching; and Team Teaching. While teachers highlighted
that TAC could support them in instructional design, teaching delivery, teacher pro-
fessional development, and lowering grading load, they perceived a lack of explicit
and consistent curriculum guidance, the dominance of commercial AI in schools,
the absence of clear ethical guidelines, and teachers’ negative attitude toward AI as
obstacles to TAC. These findings enhance our understanding of how TAC could be
structured at school levels and direct the implications for future development and
practice to support TAC. Keywords AI in education · Teacher-AI collaboration · Teaching with AI · Teacher
in the loop · Focus group interviews Types of teacher‑AI collaboration in K‑12 classroom
instruction: Chinese teachers’ perspective Jinhee Kim1 Received: 14 August 2023 / Accepted: 24 January 2024 / Published online: 23 February 2024
© The Author(s) 2024 1
Department of STEM and Professional Studies, Old Dominion University, 4301 Hampton Blvd,
Room 2333, Norfolk, VA 23529, USA 1 Introduction Embedding artificial intelligence (AI) technology into the classroom as a
teaching assistant, a tutor, and an advisor has increased and brought opportu-
nities to realize the complementary synergy with human teachers in teaching 456789)
3 Education and Information Technologies (2024) 29:17433–17465 17434 activities (Kim, 2023; Kim et al., 2022). In fact, much prior work explored
and designed AI-based systems or tools to augment teachers’ ability and co-
orchestrate complex classroom learning situations between teachers and AI
to better support students, rather than AI directly offering personalized sup-
port (Holstein & Aleven, 2022; Molenaar, 2022). For instance, AI-embedded
dashboards support teachers to understand and monitor students’ learning pro-
cesses and offer teachers supplementary insights to adequately respond to the
needs of the students (Molenaar & Knoop-Van Campen, 2018). Further, AI
could possibly facilitate co-orchestration between human teachers to leverage
the complementary strengths of each. As an example, Holstein et al. (2018)
designed Lumilo, mixed-reality smart glasses, that aid the teacher in describ-
ing the actions happening in the classroom in real-time to implement appro-
priate pedagogical actions. It should be noted, though, that AI cannot effec-
tively address student learning on its own such as students’ gaming the system
behaviors, or hint avoidance. The interest and demand for teacher-AI collaboration (TAC) have been increas-
ing to the point of becoming a critical research and design challenge for AI in
Education (AIED) (Rodríguez-Triana et al., 2017), however, the implementation
of TAC is complex and challenging in K-12 schools. In practice, teachers are used
to being the only teacher responsible for the course and the change required to
work efficiently in a team of colleagues is challenging even in a human-human
collaborative teaching context. This change is profound as it is not just a method
or single skill, but it actually changes the whole culture of teaching (Vangrieken
et al., 2015). 1 3 2 Literature review While many research and government initiatives have led to high expectations that
AI will save teacher time that could be redirected toward student learning (Miao
et al., 2021), others have claimed that AI will make teachers redundant – or teach-
ers’ roles will be at least reconceptualized to be learning designers and classroom
orchestrators who manage learner behavior, ensuring smooth interactions between
technology and students (Celik et al., 2022; Seldon & Abidoye, 2018). Amidst the contrasting opinions and perspectives regarding the teacher and
AI relationship, a rich line of recent research has proposed the notion of TAC, or
human-AI co-orchestration that amplifies the complementary strengths of each
participant without simply automating or replacing valuable human-human inter-
actions and limiting the roles of teachers (Holstein et al., 2019; Cukurova et al.,
2019; Mavrikis et al., 2021). This shifts the attention that was previously focused on
learner-supporting AI, which offers personalized support directly to learners (i.e.,
AI-powered adaptive tutoring, Du Boulay, 2016), to the design of effective TAC
processes and systems whereby teachers and AI are considered equal team members
in orchestrating individual and collaborative instructional activities during the class
(Dillenbourg et al., 2018). These AI systems are designed to augment human teacher
ability to overcome teacher limitations and co-orchestrate classroom interactions
and activities (Holstein & Aleven, 2022; Ji et al., 2023). For instance, Kaliisa and
Dolonen (2023) designed the Canvas Discussion Analytics Dashboard (CADA) to
augment teachers’ awareness of relevant features of online learning situations (e.g.,
students’ participation and discourse patterns), and offer some hints regarding what
teachers need to do to support students. Teachers can take advantage of the hints
provided by the dashboard to facilitate relevant pedagogical decisions and make
relevant interventions. That is to say, the dashboard not only helps teachers sense
the status quo information about learning but also enhances and expands the range
of teachers’ instructional actions. Additionally, some research has explored design
approaches in which teachers help and train AI systems, via demonstration, to sense
relevant information about students’ broader contexts (e.g., students’ at-home dif-
ficulties that may negatively impact their academic performance, Bull & Kay, 2016),
recognize instructionally relevant features, and perceive learning situations and
information (Lee et al., 2015). 1 Introduction Without formal AIED training, teachers are expected to jointly
orchestrate classroom instruction in support of a fully-packed curriculum that
involves the planning and real-time management of diverse classroom activities
(Dillenbourg & Jermann, 2010) alongside AI without a clear model or best prac-
tices to serve as a guideline for that joint orchestration.f Considering that human teachers play critical roles in mediating the effective-
ness of AI in K-12 classrooms (Ritter et al., 2016), this study aims to explore the
types of TAC in teaching that could be adopted in classroom instruction and to
examine both the benefits and challenges of TAC perceived by K-12 teachers. The
questions guiding the study are as follows: at types of TAC can be implemented in the classroom instruction? 1. What types of TAC can be implemented in the classroom instruction? 2. What do teachers think are the main opportunities for TAC in classroom instruc
tion? 3. What are the obstacles to TAC perceived by teachers? The findings of this study can offer insights into how teachers leverage TAC at
the classroom levels and ensure teachers-in-the-loop in AI-supported instruction
(Ramos et al., 2020; Renz & Vladova, 2021) whereby AI empowers and not over-
powers teachers so that teachers’ expertise is respected and teachers are actively
involved and have control over the instructional process while balancing the pro-
cess of human and AI-driven decision-making and mutual monitoring. The study 1 3 Education and Information Technologies (2024) 29:17433–17465 17435 findings can also direct the implications for future developments and practice to
support TAC. findings can also direct the implications for future developments and practice to
support TAC. 2 Literature review In sum, the discourse around the utilization of AIED shifted from being narrowly
focused on automation of teaching tasks and AI-directed teaching to the augmen-
tation of human teacher capabilities linked to learning and teaching (Chatti et al.,
2020) and identifying the advantages, scope, and challenges in forming effective
TAC. Many studies anticipate and highlight TAC could support teachers in deliv-
ering more personalized learning experiences while promoting their motivation,
engagement, and immersion in learning, and enhancing self-efficacy, self-organiza-
tion, and ownership of their own learning (Kim, in press). For example, AI is widely 1 3 3 Education and Information Technologies (2024) 29:17433–17465 17436 used to create exploratory and gamified learning environments, generating and sug-
gesting personalized learning tasks to help teachers recognize, facilitate, and achieve
quality engagement of each individual learner (Miao et al., 2021). The Kid Space,
an interactive learning environment, incorporates extensive multimodal sensing
and sense-making technologies to engage learners both physically and socially with
play-based learning, guided by an animated teddy bear that analyzes and responds
to learners’ behaviors, emotions, and utterances in real time (Anderson et al.,
2018). On the other hand, AI could support teachers in preparing and developing
instructional materials by searching and scraping online educational resources (e.g.,
X5Learn, Perez-Ortiz et al., 2021), analyze and support teacher practices (Wise &
Jung, 2019), time management and course/lesson planning (e.g., Chounta et al.,
2022) . Despite these promising use cases, there is evidently widespread concern related
to the practice of effective TAC. First of all, it is important to acknowledge that
while AI has demonstrated a strong ability to solve a single task in a well-defined
scenario it has limitations in understanding various contexts, scenarios, and behav-
iors. These limitations restrict AI’s ability to address highly complex and open
problems in diverse teaching scenarios which serves as a key challenge in TAC
(Crescenzi-Lanna, 2023). Second, much research underlines the need to empower
teachers’ AI-related competencies. This encompasses not just general computational
thinking skills, but also the understanding and use of key concepts to create and
apply AI tools and AI-empowered pedagogical approaches to facilitate learning
while ensuring teachers’ agency and accountabilities (Kim et al., 2022; Miao et al.,
2021; Touretzky et al., 2019). 2 Literature review Particularly, the teachers’ lower levels of meta-knowl-
edge in recognizing and assessing complementarities and shortcomings of working
AI lead to poor delegation decisions (Fügener et al., 2022) and distrust toward AI
(e.g., algorithm aversion, Dietvorst et al., 2015). It remains unclear how a teacher and AI can form and apply TAC in the class-
room context. An active, collaborative relationship where these two heterogeneous
instructors are working purposefully, regularly, and collaboratively to deliver sub-
stantive instruction, and to meet the diverse learning needs of students has rarely
been demonstrated. This study, therefore, aims to explore the diverse forms of
TAC in classroom instruction perceived by teachers. Such foresight indicators can
enhance our understanding of what complementary strengths teachers and AI sys-
tems hold in a given context and can be used to guide the design of systems that
combine these strengths. 3.1 Participants This study adopted a purposeful sampling method to recruit 30 research participants. Each participant’s school level they are currently teaching, gender, education level,
teaching years, and subject as well as types of AI used for instruction were exam-
ined (see Table 1). 3.1 Participants Regarding the types of AI used for instruction, participants were 1 3 Education and Information Technologies (2024) 29:17433–17465 17437 Table 1 Participants’ characteristics
Participant
School level
Gender
Education level
Teaching years
Teaching subject
Types of AI used for instruction
Group A
1
Primary
F
Master’s degree
7 years
Art
ELE, AI-based dashboard
2
Middle
F
Master’s degree
12 years
English
AI-based dashboard, AWE, chatbot
3
Primary
M
Master’s degree
10 years
Social studies
ELE, AI-based dashboard
4
Middle
F
Master’s degree
4 years
Math
ITS, ELE, AI-based dashboard, AWE, chatbot
5
High
M
Master’s degree
8 years
Information Technology
ITS, AI-based dashboard, chatbot
Group B
6
Primary
M
Master’s degree
6 years
Physical education
AI-based dashboard
7
High
F
PhD’s degree
17 years
Math
ITS, ELE, AI-based dashboard, AWE, chatbot
8
Primary
M
Master’s degree
12 years
Ideology and morality
ELE, chatbot
9
Middle
M
Master’s degree
3 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
10
High
F
Master’s degree
7 years
Chinese
AWE, chatbot
Group C
11
High
M
Master’s degree
3 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
12
High
F
Master’s degree
9 years
Natural sciences
ITS, ELE, AI-based dashboard
13
Primary
F
Master’s degree
15 years
Music
AI-based dashboard, chatbot
14
Middle
M
Master’s degree
11 years
Physical education
AI-based dashboard, chatbot
15
Middle
F
Master’s degree
6 years
Political Science
AI-based dashboard, AWE, chatbot
Group D
16
Middle
F
PhD’s degree
20 years
Chinese
ELE, AI-based dashboard, AWE, chatbot
17
Primary
M
Master’s degree
10 years
Social studies
ELE, AI-based dashboard, chatbot
18
Middle
M
Master’s degree
4 years
Biology
ITS, ELE, AI-based dashboard, AWE, chatbot
19
Primary
F
Master’s degree
15 years
Chinese
ITS, AI-based dashboard, AWE, chatbot
20
High
M
Master’s degree
5 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
Group E
21
Middle
F
PhD’s degree
18 years
Geography
ITS, ELE, AI-based dashboard, AWE, chatbot
22
Middle
M
Master’s degree
10 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
23
High
F
Master’s degree
3 years
English
AI-based dashboard, AWE, chatbot
24
Primary
F
Master’s degree
5 years
Art
AI-based dashboard, chatbot 3 Education and Information Technologies (2024) 29:17433–17465 17438 1
(
)
Participant
School level
Gender
Education level
Teaching years
Teaching subject
Types of AI used for instruction
26
High
M
PhD’s degree
10 years
Natural sciences
ITS, ELE, AI-based dashboard
Group F
27
Primary
F
Master’s degree
12 years
English
AI-based dashboard, AWE, chatbot
27
Primary
M
Master’s degree
12 years
Physical education
ITS, ELE, AI-based dashboard, chatbot
28
High
M
PhD’s degree
7 years
Natural sciences
ELE, AI-based dashboard
29
Middle
F
Master’s degree
10 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
30
High
F
PhD’s degree
15 years
Social sciences
ELE, AI-based dashboard, chatbot 1 3
(
)
Participant
School level
Gender
Education level
Teaching years
Teaching subject
Types of AI used for instruction
26
High
M
PhD’s degree
10 years
Natural sciences
ITS, ELE, AI-based dashboard
Group F
27
Primary
F
Master’s degree
12 years
English
AI-based dashboard, AWE, chatbot
27
Primary
M
Master’s degree
12 years
Physical education
ITS, ELE, AI-based dashboard, chatbot
28
High
M
PhD’s degree
7 years
Natural sciences
ELE, AI-based dashboard
29
Middle
F
Master’s degree
10 years
Information Technology
ITS, ELE, AI-based dashboard, AWE, chatbot
30
High
F
PhD’s degree
15 years
Social sciences
ELE, AI-based dashboard, chatbot Education and Information Technologies (2024) 29:17433–17465 17439 asked to choose ones that apply to their own experience among five types: Intel-
ligent Tutoring System (ITS), Exploratory Learning Environments (ELE), AI-based
dashboard, Automatic writing evaluation (AWE), and chatbot (Homles at al., 2019a;
Lameras & Arnab, 2021). 3.1 Participants In addition, all participants were required to have at least
a year of teaching experience in AIED, either in teaching with AI or teaching about
AI (Holmes et al., 2019a, b). The former refers to the use of AI as direct teaching
tools such as AI-based mobile apps and adaptive or personalized learning manage-
ment systems, whereas the latter indicates actual teaching experience in AI skills
and understanding (i.e., AI concepts, AI applications, AI ethics, building AI). The
study did not require participants to have extensive knowledge or technical skills in
AI model development as the study aimed to examine their perspective on TAC and
its potential effects, not developing AI modeling techniques. f
This study obtained ethical approval from the author’s university’s Institutional
Review Board (NO. ER-AOFE-11000086620220922083531) and also received
informed consent from all participants. 3.2 Data collection The study conducted Focus Group Interviews (FGI) to allow the participants to
express opinions, describe experiences, and provide detailed information (Turner,
2010). Prior to conducting FGI, the study created scenarios to better facilitate the inter-
views as well as allow participants to shape their own perspectives and assess TAC
in a more critical and nuanced manner within a real-classroom context with their
peer group (Zimmerman & Forlizzi, 2017). To explore teachers’ diverse percep-
tions, the AI that collaborates with teachers in the study embraces all types of AI
being used in school instruction, both physical (Physical systems in which AI is
embedded such as Robot, AI speakers, etc.) and digital AI (e.g., model, algorithms,
generative AI, etc.)i To create scenarios, the study first conducted an online brain-writing activity
about TAC scenarios in K-12 classrooms (Linsey & Becker, 2011) among 6 teach-
ers (2 from primary, 2 from middle, 2 from high schools), with an average of 7.8
years (SD=3.5 years) of teaching experience to create technically feasible and rel-
evant TAC in a classroom instruction context. Each teacher wrote down their ideas
in response to TAC scenarios using Miro (https://miro.com/brainstorming/), a free
online collaborative whiteboard and passed them to another teacher for review and
refinement. Once every proposed scenario made a full round, the team shared all
the ideas together to review and refine each idea until all designers agreed that the
scenarios of TAC were technically feasible and contextualized in the real class-
room. These TAC scenarios were further validated through expert reviews. The
study conducted individual semi-structured interviews with a total of 5 experts (1
from instructional design and technology, 1 from primary education, 1 in secondary
education, and 2 from Human-Computer Interaction) with an average of 10.8 years
(SD=7.8 years) of research experience and 8 years (SD=6.2 years) of teaching expe-
rience, using a video conferencing platform (i.e., Zoom and Tencent platform) in 1 3 Education and Information Technologies (2024) 29:17433–17465 17440 60–70 minutes. The questions discussed include, but are not limited to: (1) Can you
improve this scenario to make it technically feasible?; (2) can you improve this sce-
nario to have a positive impact on classroom interaction based on your own teaching
experience?; (3) Do you have any research ideas that can be used as a new sce-
nario? 3.2 Data collection As a result, prior to conducting FGI with participants, a total of 15 scenarios
(Table 2) were developed by reflecting on the experts’ review of the teacher created
scenarios and AIED literature which were then grouped into six categories: (1) One
teach-One observe (one has primary responsibility while the other gathers specific
observational information on students or the instructing teacher); (2) One teach-One
assist (one has primary instructional responsibility, while the other assists students
with their work, monitor behaviors, or corrects assignments); (3) Station teaching
(the co-teaching pair divides the instructional content into parts and the students into
groups); (4) parallel teaching (each instructs half the students on the same lesson);
(5) alternative teaching (one teaches the main lesson to a larger group of students
while the other works with the small group of students on an entirely different les-
son); and (6) team teaching (both delivers the same instruction at the same time).f This study formed six focus groups consisting of teachers from different levels of
schools (i.e., primary, middle, and high schools), teaching subjects, years of teach-
ing experiences, schools, and gender (see Table 1). A single FGI was conducted per
week between 3 March 2023 to 7 April 2023 until all groups had been interviewed
with each session lasting approximately two hours. Each group of participants first
read each of the scenarios aloud and was asked to critically assess and discuss how
TAC could be implemented and contextualized in the actual classroom without
being too intrusive and, additionally, describe potential benefits for and obstacles
to TAC that could undermine or strengthen classroom teaching and learning with
guiding questions (see Table 3). Meanwhile, participants were also asked to choose
the forms of TAC that would work well and identify which did not sound desirable
to elicit their holistic point of view regarding perceived opportunities and drawbacks
of TAC. All interviews were conducted in Chinese, the participants’ native language, and
audio-recorded, transcribed, and later translated into English by research assistants
with research experience in translation in both Chinese and English. 1 3 3.3 Data analysis A reflective thematic analysis (Braun & Clarke, 2006) was undertaken to identify
and construct emergent themes for each RQ. After going through the data famil-
iarization stage by reading and re-reading the transcripts, three researchers indepen-
dently generated initial codes related to intriguing statements or phrases in the data. These were then combined into 20 potential overarching themes. The entire data-
set was once again thoroughly reviewed to uncover and develop any new emerging
codes and themes. These were compared and contrasted with each other until all
researchers reached an agreement on every theme. To ensure the reliability of the qualitative data analysis, the study employed two
strategies: (1) respondent validation and (2) employing moderators (Golafshani, 1 3 1 3 Education and Information Technologies (2024) 29:17433–17465 17441 1
No Category
Scenarios
Reference
1
One teach-One observe Teacher leads instruction and AI observes and gathers observational information on stu-
dents’ learning process/experience
Holmes et al., 2019a, b; Han et al., 2021
2
Teacher leads instruction and AI observes and gathers observational information on teach-
ers’ instructional practices
Sergis et al., 2019; Seo et al., 2021
3
Teacher leads instruction and AI observes and gathers observational information on
classroom climate including the social, emotional, and physical aspects of learning space
(e.g., moods, attitudes, and tone of instructor-student and student–student interactions,
student cohesiveness, etc.)
Amado & Jarque, 2022
4
AI leads instruction and teacher observes and gathers observational information on class-
room climate
Miao et al., 2021
5
One teach-One assist
Teacher leads instruction and AI assists teacher’s administrative tasks such as addressing
FAQ, students’ academic report, attendance checking
Goel & Polepeddi, 2016
6
Teacher leads instruction and AI assists teachers’ instruction by offering unobtrusive assis-
tance to students with their work, or correcting assignments, as needed
Kasepalu et al., 2022; Wise & Jung, 2019
7
Teacher leads instruction and AI assists in appropriate learning content and material sup-
ply for teachers and students as needed
Holmes et al., 2019a, b; Perez-Ortiz et al., 2021
8
Station teaching
The co-teaching pair of teacher and AI divides the students into groups and the instruc-
tional content into different learning activities. 3.3 Data analysis For instance, teacher leads a station where
students work on reading-related activities, while AI works with students on writing
tasks in English class
Miao et al., 2021
9
The co-teaching pair of teacher and AI divides the students into groups and the instruc-
tional content into different levels of learning tasks. For instance, teacher leads a station
for an advanced level, while AI runs introductory sessions
Miao et al., 2021
10
Parallel teaching
The co-teaching pair of teacher and AI splits the class into two groups and each teaches the
same information but that can be discussed from different views (e.g., debating) at the
same time
Schäffer & Lieder, 2023 Education and Information Technologies (2024) 29:17433–17465 17442 1
Table 2 (continued)
No Category
Scenarios
Reference
11
The co-teaching pair of teacher and AI splits the class into two groups and each teaches the
same information but in a different mode of learning space. For instance, teacher leads
instruction in the actual classroom, while AI leads instruction online
Anderson et al., 2018
12
Differentiated teaching
Teacher manages a large group of students, while AI takes a small group for a specific
instructional purpose (i.e., gifted students, students with learning difficulties)
Miao et al., 2021; Holmes et al., 2019a, b
13
AI manages a large group of students, while teachers takes a small group for a specific
instructional purpose (i.e., gifted students, students with learning difficulty)
Miao et al., 2021; Holmes et al., 2019a, b
14
Team teaching
Both teacher and AI are in the classroom at the same time and share the instruction, freely
interject information, assist students, and answer questions as playing different roles. For
instance, one explains the concept of knowledge, while the other generates thoughts-
awakening questions
Kim & Lee, 2023; Tuomi, 2019
15
Both teacher and AI are in the classroom at the same time and share the instruction yet
involving in different pedagogical instruction. For instance, one delivers instruction in
the form of experiential learning, while the other one leads research-based learning
Kim et al., 2022 Education and Information Technologies (2024) 29:17433–17465 17443 Table 3 Interview guiding questions
Questions
Forms of TAC
∙ Among the storyboards presented, which forms of TAC do you think
would work well in your classroom context and why? Also, which
one wouldn’t work well? ∙ What other forms of TAC can be developed? 3.3 Data analysis Opportunities for TAC
∙ How the implementation of TAC will support your teaching practice? ∙ Who could the TAC support and how could TAC support them? ∙ How can we turn the TAC into opportunities in the field of education? Obstacles to TAC
∙ Are there any obstacles that prevent the TAC to be formed and imple-
mented in the school and classroom environment? ∙ Who and/or what might cause problems in structuring and imple-
menting the TAC? ∙ Are there any standards, policies, and/or legislation that might nega-
tively impact the TAC? 2003). The interview transcripts and analysis results were confirmed with each
participant and were allowed to be revised when necessary. In addition, the study
conducted three iterative discussions with the team of instructional designers and
experts who were involved in the TAC scenario storyboard development to review,
organize, define, and name the themes. As a result, a total of 14 themes were identi-
fied which included 6 themes with 7 sub-themes under RQ1, 4 themes with 4 sub-
themes under RQ2, and 4 themes with 3 sub-themes under RQ3 (see Table 4). 4.1.1 One teach, one observe Teachers anticipated that they would serve as leading teachers while AI could play a
key role in learning analytics and teaching analytics. Although AI’s role in learning
analytics and teaching analytics share a similar end goal of effectively improving
teachers’ instructional processes, there are differences in the target audiences, objec-
tives, and educational data used. First, teachers suggest that teachers would serve
as the primary teachers, while AI gathers specific observational data on students’
learning progress, performance, and engagement with educational resources/tools
and tasks/activities as well as their peers in learning which otherwise is difficult to
gather and track by an individual teacher. In turn, teachers can better understand
how the learning process is carried out including challenges that exist within the
classroom experience, providing them the information necessary to maximize learn-
ing potential and reinforce behaviors while applying interventions that more closely
align with learners’ needs and preferences. Also, teachers perceive that AI’s support in student observation creates a start-
ing point for helpful conversations between both learners and teachers rather 1 3 3 7444
Education and Information Technologies (2024) 29:17433–174
1 3
Table 4 Summary of emergent themes
Category
Themes
Sub-themes
Exemplary quotes
Forms of TAC
(RQ1)
T1. One Teach, One Observe
Teacher as a lead teacher, AI as learning
analytics
Students are passive in presenting their opin-
ions during class, so I often conduct online
discussions and collaborative arguments. But
it’s simply impossible for one teacher can
fully recognize and monitor each individual
student in their learning process. AI in this
respect helps me a lot by monitoring students’
learning progress and presenting some of the
details and patterns that I wasn’t well aware
such as students’ misconceptions and their
social relationships like who reacts to who. (D16)
Teacher as a lead teacher, AI as Teaching
Analytics
Although the current AI is more directed
to monitor students’ data, AI can also be
applied to collect and analyze the data of
teachers’ teaching process such as teachers’
instructional behavior and resources for
preparing and delivering lessons in the TAC
context since AI will most closely work with
teachers. This can guide us to reflect on the
teaching process and outcomes and support
our diagnostic pedagogical ability to adjust
and improve the quality of teaching. (A1)
T2. 4.1.1 One teach, one observe One Teach, One Assist
Teacher as a lead teacher, AI as an adminis-
trative assistant
I developed a class chatbot to delegate my over-
burdened and exhausted tasks in answering
students’ frequently asked questions such as
questions related to key dates and deadlines,
the syllabus, school, and class events, class
preparation materials, etc. (A5) 17444 Education and Information Technologies (2024) 29:17433–17465 ble 4 Summary of emergent themes
ategory
Themes
Sub-themes
Exemplary quotes
orms of TAC
RQ1)
T1. One Teach, One Observe
Teacher as a lead teacher, AI as learning
analytics
Students are passive in presenting their opin-
ions during class, so I often conduct online
discussions and collaborative arguments. But
it’s simply impossible for one teacher can
fully recognize and monitor each individual
student in their learning process. AI in this
respect helps me a lot by monitoring students’
learning progress and presenting some of the
details and patterns that I wasn’t well aware
such as students’ misconceptions and their
social relationships like who reacts to who. (D16)
Teacher as a lead teacher, AI as Teaching
Analytics
Although the current AI is more directed
to monitor students’ data, AI can also be
applied to collect and analyze the data of
teachers’ teaching process such as teachers’
instructional behavior and resources for
preparing and delivering lessons in the TAC
context since AI will most closely work with
teachers. This can guide us to reflect on the
teaching process and outcomes and support
our diagnostic pedagogical ability to adjust
and improve the quality of teaching. (A1)
T2. One Teach, One Assist
Teacher as a lead teacher, AI as an adminis-
trative assistant
I developed a class chatbot to delegate my over-
burdened and exhausted tasks in answering
students’ frequently asked questions such as
questions related to key dates and deadlines,
the syllabus, school, and class events, class
preparation materials, etc. (A5) Education and Information Technologies (2024) 29:17433–17465 17445 ble 4 (continued)
tegory
Themes
Sub-themes
Exemplary quotes
Teacher as a lead teacher, AI as an assistant
tutor
Although current learning analytics tools avail-
able in the school only offer limited informa-
tion and visualization about students’ learning
processes and outcomes, I think AI has the
potential to further develop to support the
teachers’ pedagogical actions by proactively
interpreting its data analysis results, explain-
ing them to me and recommending feasible
intervention. 4.1.1 One teach, one observe (E22)
Teacher as a lead teacher, AI as an instruc-
tional materials suppliers
Though I don’t use every learning material
such as quizzes, worksheets, or videos recom-
mended by AI, any material is better than no
material at all. It really saves time in prepar-
ing lesson materials and also suggests neces-
sary content in keeping with the curriculum,
which I couldn’t think of. (D17)
T3. Co-teaching in stations
Teacher and AI co-teach for different learning
activity
The class usually breaks into small groups
at separate stations for different learning
activities; One group is with me and engages
in, for instance, physical interactive activities
with other peers such as role play to improve
speaking, while the other group works on
writing tasks with AI, the other is given with
vocabulary quizzes and puzzles from AI-
assisted programs. (E26) Education and Information Technologies (2024) 29:17433–17465 17446 (continued)
y
Themes
Sub-themes
Exemplary quotes
Teacher and AI co-teach for different levels of
learning tasks
I work with AI to deliver class activities in
a sequence of different levels of difficulty,
making tasks or activities easier and harder,
which shifts levels of understanding. For
example, AI works with students on simple
algebra through simple quizzes and gradually
engages them in difficult tasks by presenting
real-world examples to apply mathematic
equations, and I often work with students on
creative mathematical reasoning through
discussions. (B7)
T4. Parallel teaching in online and offline
classes
I and AI can divide the class in half; I lead
students in the actual classroom while AI
instructs those who participate online for sick
leave or other emergencies. (B9)
T5. Differentiated teaching
It is seldom that we have a class of students
who have the same learning levels. For this,
AI works with a small group of students,
either top-tier groups or lower-tier groups,
and offers them intensive instruction such
as working on front loading, re-teaching, or
offering an extension activity. Meanwhile, I
work with the rest of the class to complete the
planned lesson. (C13)
T6. Team teaching
Teacher and AI co-teach in different roles in
a lesson
In English class, I often teach grammatical
structure to students, while AI practices daily
conversations and pronunciation since I am
not a native speaker. (E23) T4. Parallel teaching in online and offline
l Education and Information Technologies (2024) 29:17433–17465 17447 ble 4 (continued)
tegory
Themes
Sub-themes
Exemplary quotes
nefits of SAC
Q2)
T7. 4.1.1 One teach, one observe Instructional design
AI systems gauge a student’s learning style,
pre-existing knowledge, and learning pro-
gress and this certainly helps me better tailor
lessons to an individual student. (F28)
T8. Teaching delivery
Creating opportunities to implement creative
and engaging activities
Instead of teacher-dominated delivery of knowl-
edge, teaching students how to solve problems
directly, students can try with and explore a
range of different ways using different digital
tools available on the AI system to solve frac-
tion problems guided by adaptive feedback
both by AI and me. This improves not only
students’ arithmetical computation but also
students’ overall math problem-solving. (F27)
flexible student grouping
The AI system analyzes data on individual
students’ academic performance and divides
them into groups that are well-balanced or
suggests a couple of choices so that I can
decide the optimal group makeup consists of
students of varying levels of academic perfor-
mance so that each brings their own unique
talents and interests to the group work. (D19) 3 Education and Information Technologies (2024) 29:17433–17465 17448 (continued)
y
Themes
Sub-themes
Exemplary quotes
T9. Teacher professional development
Reflection on instructional practice
AI could ask questions to guide and facilitate
my self-reflection during the recorded class-
room instruction video analysis. I found such
AI-guided reflection time is helpful to me as
a veteran teacher who is likely to recycle the
same instructional materials and is reluctant
to adjust to new pedagogies. And I don’t have
to feel stressed in scheduling the class obser-
vation with my colleagues nor pressured to be
observed by someone else. (E21)
Continuous study
For me to better make use of AI’s strength, I
also need to keep up with AI and ML trends
and test how to contextualize in my classroom
context. (C11)
T10. Lowering grading load
The AI grading system can scan students’
homework and speed up grading. And the
system can generate automatic reports. (B8)
s to TAC (RQ3) T11. Lack of explicit and consistent curricu-
lum guidance
An absence of informed visions and goals of
AIED
Isn’t the use of AI supposed to enable and map
personalized learning that responds to the
voice, respects the choices, and caters to the
needs of learners? 4.1.1 One teach, one observe Apparently, what we do
with AI for students mainly involves a drill
and practice approach in which students are
given the learning materials and contents
repeatedly by teachers and AI to better per-
form in the school exam. (C14) Table 4 (continued)
Category
Themes
Sub-themes
Exemplary quotes
T9. Teacher professional development
Reflection on instructional practice
AI could ask questions to guide and facilitate
my self-reflection during the recorded class-
room instruction video analysis. I found such
AI-guided reflection time is helpful to me as
a veteran teacher who is likely to recycle the
same instructional materials and is reluctant
to adjust to new pedagogies. And I don’t have
to feel stressed in scheduling the class obser-
vation with my colleagues nor pressured to be
observed by someone else. (E21)
Continuous study
For me to better make use of AI’s strength, I
also need to keep up with AI and ML trends
and test how to contextualize in my classroom
context. (C11)
T10. Lowering grading load
The AI grading system can scan students’
homework and speed up grading. And the
system can generate automatic reports. (B8)
Obstacles to TAC (RQ3) T11. Lack of explicit and consistent curricu-
lum guidance
An absence of informed visions and goals of
AIED
Isn’t the use of AI supposed to enable and map
personalized learning that responds to the
voice, respects the choices, and caters to the
needs of learners? Apparently, what we do
with AI for students mainly involves a drill
and practice approach in which students are
given the learning materials and contents
repeatedly by teachers and AI to better per-
form in the school exam. (C14) Education and Information Technologies (2024) 29:17433–17465 17449 Table 4 (continued)
Category
Themes
Sub-themes
Exemplary quotes
Lack of systematic learning content
What we are given is all about how to teach
programming language to build better AI,
which can easily be taught by a human
teacher. There are no contents and materials
that describe how teachers and AI can co-
deliver instruction effectively. (D20)
Limited TAC target outcomes and evaluation
mechanisms
It is not clearly communicated what outcomes
I need to make with AI and how they can
be evaluated. So, I am not sure how to plan
a series of activities that need to make the
best of TAC apart from making use of AI for
analyzing students’ data and recommending
appropriate learning content for students. 4.1.1 One teach, one observe (E24)
T12. Commerical-driven AI
Almost every educational AI offered to schools
is developed by big tech companies. They
defined what the AI systems should do for
education and how AI systems should affect
learners and teachers, and pushed us to use
their products. What do they know about
education? (C15) ble 4 (continued)
tegory
Themes
Sub-themes
Exemplary quotes
Lack of systematic learning content
What we are given is all about how to teach
programming language to build better AI,
which can easily be taught by a human
teacher. There are no contents and materials
that describe how teachers and AI can co-
deliver instruction effectively. (D20)
Limited TAC target outcomes and evaluation
mechanisms
It is not clearly communicated what outcomes
I need to make with AI and how they can
be evaluated. So, I am not sure how to plan
a series of activities that need to make the
best of TAC apart from making use of AI for
analyzing students’ data and recommending
appropriate learning content for students. (E24)
T12. Commerical-driven AI
Almost every educational AI offered to schools
is developed by big tech companies. They
defined what the AI systems should do for
education and how AI systems should affect
learners and teachers, and pushed us to use
their products. What do they know about
education? (C15) 3 Education and Information Technologies (2024) 29:17433–17465 17450 able 4 (continued)
ategory
Themes
Sub-themes
Exemplary quotes
T13. Absence of clear AI ethics
In contrast to those research and practices in
the medical sector where there are long-
established ethical principles and codes
of practice to comply with, we don’t have
clear-cut protocols and guidelines concern-
ing the biodata collected from AI-based
robots in the school for students. I am not
sure whom I need to ask about the nitty–gritty
details involved in the data maintenance and
analysis. (D18)
T14. Negative attitude toward AI
It’s quite a paradoxical situation whereby most
teachers are using AI for their classroom
instruction in some ways and enjoying the
benefits of having AI in the school, at least for
their attendance checking, but they think AI is
difficult to learn, interact with, or something
scary that will take away our jobs one day,
with exceptions for informatics teachers. (F30) T14. Negative attitude toward AI Education and Information Technologies (2024) 29:17433–17465 17451 than having a broad idea that is unable to target specific problems. 4.1.1 One teach, one observe Teachers can
take this opportunity presented by AI to help students feel connected with class-
room learning and to engage students as partners in improving their learning
environments. than having a broad idea that is unable to target specific problems. Teachers can
take this opportunity presented by AI to help students feel connected with class-
room learning and to engage students as partners in improving their learning
environments. AI’s observable data may not fully pinpoint students’ various learning chal-
lenges but I can take them as a point of reference to ask students questions
about their behavior and interpret and discuss data together and clarify and
plan for the next steps. (D18) By contrast, AI-powered teaching analytics supports the primary teacher in
reflecting on the effectiveness of their instructional design and delivery process
and in developing an approach for further action and refinement. Hence, AI is
directed to teachers and collects data that is relevant to support teachers in reflec-
tion on teaching approaches and their engagement with the activities, such as the
frequency of teachers’ participation and engagement in discussion activities, lev-
els of engagement in the teaching process, analysis of their teaching approach,
and any points of modification including the learning content delivery mode. 4.1.3 Co‑teaching in stations Another form of TAC perceived by teachers occurs in the station teaching approach;
the co-teaching pair of human teacher and AI divides the class into workstations or
learning centers where learners rotate in and out in a small group, which is either
managed by a teacher or AI. Teachers proposed that stations could be designed with
different learning activities/tasks and/or different concepts related to the same lesson
to utilize both teacher and AI lesson-planning abilities and content area strengths
as well as enhance students’ deeper understanding of the subject and applied
knowledge. On the other side, learning activities could be designed and conducted with the
same concepts and ideas, but in a sequence of different difficulty levels to move stu-
dents progressively toward stronger understanding and, ultimately, greater independ-
ence in the learning process. Students, for instance, can work with a teacher to solve
a set of five polynomial equations at station 1. At station 2, students will use laptops
to complete a Desmos activity on graphing polynomials with AI and so on (A4). 4.1.4 Parallel teaching in online and offline classes The TAC can also take place in the form of parallel teaching in which the class is
split in half, with each half being taught the same lesson by either a human teacher
or AI. Teachers are expected to collaborate with AI to functionalize an effective
Hybrid-flexible (Hyflex) model of instruction that could offer equivalent learning
experiences for both those attending online and in-person classes to ensure that
students present in the classroom and those studying remotely due to a variety of
unique situations (e.g. health, location, etc.) can learn together. However, imple-
menting a Hyflex mode of instruction puts increased pressure on teachers to plan
and design instruction that engages both in-person and online attendees, while also
considering potential tech issues in the online learning environment and fully sup-
porting interaction between in-person students, virtual students, and the teacher dur-
ing instruction. In this regard, teachers expected TAC could take the form of one
class being taught by a human teacher in a traditional classroom environment while
also independently having the AI support the online learning environment. TAC in
this context could contribute to the development of creative solutions to such chal-
lenges by reducing the student-teacher ratio and increasing instructional intensity for
an individual student. 4.1.2 One teach, one assist In line with the abovementioned themes, teachers expected TAC would consist of
a teacher serving in a lead role while AI served in a secondary role, in this case as
an administrative assistant. To be specific, teachers expected AI to carry out routine
administrative and clerical tasks that distract them from their core teaching respon-
sibilities. For instance, responding to students’ frequently asked questions (A4,
B7); analyzing attendance data (B9, C13); digital device troubleshooting such as
offering virtual technical support (D20, E22); processing exam results (A3, F26);
and filing, collating, and reporting records of their student’s academic progress and
constantly update this information in digital forms (B8, C15). With respect to exam-
related tasks, although teachers understood AI could automatically assess learner
assignments and exams which would save time, teachers argued that AI’s depth of
interpretation or accuracy of analysis cannot excel beyond what one human teacher
can offer (Holmes et al., 2019a, b). Also, teachers highlighted that teachers learn
about their learners while assessing outcomes that allow teachers to develop insights
on what and how to support individual students. Thereby, teachers envision TAC in
assessment in a way that AI supports the grading/assessment process (i.e., offering
prompts and shortcuts) while teachers do the actual grading. Furthermore, while teachers take primary responsibility for delivering whole-
class instruction, AI could support individual students within the whole group
as an assistant tutor by answering student questions (E21, F26), monitoring/cor-
recting students’ works (A1, B7), and suggesting a broad range of instructional
resources which may include but is not limited to multimedia resources, supple-
mentary readings and task sheets (C12, D17) to encourage, improve, and promote
teaching and learning activities during the process of instruction. 1 3 Education and Information Technologies (2024) 29:17433–17465 17452 4.1.6 Team teaching Teachers suggested that the teacher and AI could co-teach at the same time, but take
turns, sharing the responsibilities of lead instruction, with equally active, but pos-
sibly different roles in a lesson (e.g., the teacher makes a point, AI can jump in and
elaborate if needed or vice versa, D16; Teacher leads the class, AI leads the experi-
ment, E25).lf In line with this, it was interesting to capture that teachers reflected on the differ-
ent strengths of humans and AI teachers in a team teaching scenario. For instance,
AI is efficient in teaching facts, logic, and theories whereas human teachers can read
social cues and strong sensory feelings and emotions, to nurture students’ whole-
person development, also known as holistic learning, developing students both intel-
lectually (e.g., higher-order thinking like analytic abilities, technical skills — things
like C++ programming, data set analysis, or process mapping, and the acquisition of
subject-based knowledge) and emotionally (e.g., value, consciousness, self-/social
awareness, intrapersonal communication, adaptability, and conflict resolution). AI teaches with something that it is good at or even excels over humans such
as teaching programming since AI automatically detects errors in code that I
often miss and even takes so much time during the class. I then support stu-
dents’ soft skills such as defining genuine problems, collaborating with peers,
and presentation skills, which are unable to articulate and explain clearly for
AI to instruct them to students. (F29) 4.1.5 Differentiated teaching Another form of TAC is the differentiated teaching approach, as identified in TAC
scenarios of Table 2, in which one instructs most of the class while the other works
with a smaller group of students who need specialized attention. Teachers and AI
can work to analyze and review students’ data to figure out which students need sup-
port filling in gaps in background knowledge, which students need remediation, or
which students could benefit from accelerated learning because they already know
the content or have mastered the skills of the large group lesson and offer a specific 1 3 Education and Information Technologies (2024) 29:17433–17465 17453 instruction accordingly. Teachers anticipated that this approach, whether teachers
take a big group or a smaller group, allows teachers instructional flexibility in better
managing what students learn, how students learn, and how students are assessed
and tailoring instructional strategies to maximize individual growth in the teaching
content. 4.2.1 Instructional design Teachers described that TAC can help teachers better design classroom instruction
for engaging students in authentic problem-solving while deepening subject content
knowledge. In particular, teachers highlighted that working with learning analyt-
ics systems powered by AI improves an understanding of their students and better
informs them about students’ learning difficulties and dispositions that they may be
unable to identify during traditional classroom instruction. Further, learning ana-
lytics supports monitoring of the evolution of individual student learning progress
and can predict the learning time needed for mastering a subject. In turn, teachers
can plan for better customized pedagogical interventions and resources, improve
the structure of learned knowledge, and design differentiated and flexible learn-
ing opportunities for students in the classroom, hopefully leading to better learning 1 3 3 17454 Education and Information Technologies (2024) 29:17433–17465 gains (see Table 4). Teachers, however, argued that their efforts in working out the
details of individual student academic progress and performance and implement-
ing personalized instruction through TAC are impacted by schools and governments
that set the same fixed learning outcomes that are ultimately measured by stand-
ardized exams. In other words, the learning path design may be set in personalized
ways but not the destination (Holmes et al., 2018). Meanwhile, teachers highlighted
that education is not merely about supporting students’ cognitive development only. Thereby, they aspired to work with AI systems to develop personalized instruction
that can support learners’ broader areas of development including cognition, social
interaction with peers, emotional intelligence, well-being, and so on. 4.2.2 Teaching delivery Another benefit of TAC perceived by teachers was creating more opportunities to
implement creative and engaging activities. As F27 said in Table 4, teachers recog-
nized that working with AI supports technology-rich environments whereby teach-
ers and AI can both conduct hands-on activities, (re)create a realistic and authentic
real-life event or a set of learning situations to solve problems in the areas of the
instruction. Through such activities, teachers expected their teaching delivery of
subject-specific knowledge such as facts and definitions could be digested and con-
solidated by students through presentation, meaning-making, problem-solving, and
inquiry activities supported by both teachers and AI.ll TAC also allows for more flexible student grouping. Teachers reflected on data
analysis proposed by AI and put students into small groups for instruction consid-
ering their different readiness, interest, or learning styles. Students can be grouped
at the same skill level or with varying skill levels. Flexible grouping also allowed
smaller student-to-teacher ratios in which teachers felt more confident in treating
and supporting the unique needs of individual learners with learning and attention
issues. It’s simply impossible for one teacher to listen in on the conversations of mul-
tiple collaborative groups at once. The AI system can monitor multiple written
discussions simultaneously, providing timely alerts to me if a group’s conver-
sation repeatedly goes off-topic, or conversely if a group is progressing at a
faster rate than their peers. In turn, I feel less overwhelmed to address lapses in
effective collaboration and provide students with more individualized support
(C15) 4.2.4 Lowering grading load TAC could reduce teachers’ grading load, enabling teachers to provide greater
amounts of feedback on students’ learning as well as provide students with immedi-
ate automated feedback to help students revise and improve their work (Grimes &
Warschauer, 2010; Wilson & Roscoe, 2020), although most teachers highlight such
an automatic evaluation system works well only on multiple-choice or assignments
with well-defined answers such as math and coding. i
However, given that many types of assignments (i.e., daily exercises and quiz-
zes) still follow conventional paper-based practices, teachers highlight that there is
a need to establish and adopt a learning management system (LMS) that can auto-
mate or support data processing tasks such as inputting data, digitalizing records,
and tidying up the formats of diverse types of assignment. 4.2.3 Teacher professional development Teachers perceived that TAC supports their professional development. First, teach-
ers critically examine their own instructional practices, experiences, and assump-
tions with data collection and analysis conducted by AI which is largely engaged
in three stages: (1) the collection of data about what is happening in the classroom,
(2) the analysis and evaluation of this data, and (3) an exploration of how this data
relates to and can inform their practice and beliefs as a teacher (Wise & Jung, 2019). 1 3 Education and Information Technologies (2024) 29:17433–17465 17455 Teachers found reflection through TAC is more flexible, less time-consuming, and
self-directed compared to ones that involve their colleagues in the reflective pro-
cess through class observation and consultation (see Table 4). However, teachers
expressed that the process of reflection with AI requires high-quality data about
classroom practices, student experiences, and outcomes, followed by thoughtful
analysis, and contextualized interpretation in an instructional setting. Thereby, there
needs to be more support for teacher reflection on their practices through the col-
lection and analysis of sustainable, continuous, and robust collection, analysis, and
interpretation of multimodal data (Persico & Pozzi, 2015) while respecting qualita-
tive insights teachers have into their role as instructors and instructional designers
(Warr & Mishra, 2021) and promoting dynamic TAC interaction mechanisms nec-
essary for teacher reflection to meet changes in curriculum and the varied needs of
their students (Wise & Jung, 2019).i Teachers also identified that TAC requires them to engage in continual learning
to stay current in the most effective and efficient uses of AI, a landscape that is con-
stantly changing, to develop an understanding of how to combine and leverage con-
tent, pedagogy, and technology knowledge to better support students’ learning (e.g.,
acquisition of content knowledge, F28; engaging students in authentic problem-solv-
ing, D19) while benefiting teaching practices. 4.3.1 Lack of explicit and consistent curriculum guidance The lack of explicit and consistent curricular structure and guidance is a prominent
challenge identified by teachers. This deficiency impacts the four fundamental com-
ponents of classroom instruction, as shaped through TAC: the ’why’, the ’what’,
the ’how’ of learning, and ’how well’ the classroom instruction achieves its goals
(Tyler, 1949). First of all, teachers expressed that an absence of informed visions and goals of
AIED inhibits consistently defining the purpose of TAC for classroom instruction, 1 3 3 Education and Information Technologies (2024) 29:17433–17465 17456 diversifying the forms of TAC, and aligning TAC with diverse pedagogy. Although
personalization of learning has been the strong rationale for the integration of AIED,
most teachers described that the dominant goals for AIED tend to be two-fold: (1)
Cultivating high-level Talents in AI development and (2) Improving standardized
exam results. Reflecting on the former goal, TAC’s focus and outcomes prioritize
improving students’ technical skills such as programming languages, and data sci-
ence, and developing advanced AI. Meanwhile, the latter goals directed TAC to
support students to achieve a higher grade in a standardized examination, such as
Gaokao (The national college entrance examination), and prioritize rote memoriza-
tion over higher-order thinking (i.e., creativity, metacognition), and knowing facts
over a critical engagement (i.e., critical analysis and reflection). For instance, B10
expressed, I think we somehow misuse or underuse AI when it comes to the educational
domain since we can’t direct AI to play what significant tasks to be engaged in
and what contributions it should offer to education. We simply use such state-
of-art technologies just to prepare students for exams. Her views reflect critical questions discussed by previous literature: (1) ‘what is
the right kind of education that we need to pursue and deliver jointly with AI?’;
(2) ‘are the AI technologies being used in schools addressing the right educational
tasks?’; and ‘are they enhancing learning as an essentially human and social activ-
ity, or aiming to simply make learning ‘more efficient?’ (Holmes, 2020; Kim et al.,
2022; Miao & Holmes, 2021). In line with the disjointed goals of AIED, the lack of systematic and authoritative
guidance on teaching content and related materials was another significant challenge
to effective TAC. 1 3 4.3.1 Lack of explicit and consistent curriculum guidance Although teachers list numerous examples of teaching approaches
in AIED, including teaching AI concepts in ICT/STEM activities, the use of AI-
powered apps to support student subject knowledge learning, and interdisciplinary
learning under the AI+X framework, teachers argued that existing textbooks and
ready-to-use instruction packages are all directed towards the emphasis on technol-
ogy (e.g., the concepts and inner mechanisms of AI systems). In this regard, teach-
ers expressed concerns that such technology-centric content caters to the needs of
students who want to pursue further study or work in AI disciplines, thereby TAC
mainly occurs within the domain of STEM and coding education while underserv-
ing other domains and fails to accommodate students’ diverse characteristics (i.e.,
learning interests, motivation and understanding of AI). Their views allude that
insufficient understanding of AI and pedagogical content and materials could exac-
erbate the implementation of a broader coverage of TAC and thereby the teaching
areas delivered through TAC could also be narrowly defined (Long & Magerko,
2020; Zhou et al., 2020). Furthermore, limited TAC target outcomes and evaluation mechanisms led teach-
ers to remain unclear about the scope and scale of the TAC; a question of how dif-
ferent activities and relations of collaboration between teachers and AI fit together
within a wider educational practice and what tangible outcomes to pursue over
long-time periods to make the most sustainable TAC was not addressed well. As
E24 illustrated in Table 4, although teachers were informed about a wide range of 1 3 Education and Information Technologies (2024) 29:17433–17465 17457 benefits that TAC could offer, as noted earlier from the literature review, teachers
argue that learners’ high-level academic performance and achievements from school
exams were considered and counted as the key intended outcomes of TAC, while its
potential impacts on other areas such as teachers’ decision-making process, empow-
ering students’ cognition or supporting mental health, facilitating classroom prac-
tices are not taken into account. 4.3.2 Commercial‑driven AI Most AI tools currently in use in educational contexts have been a commercial ver-
sion of its AI model, which are not generally developed in consideration of edu-
cational usage nor reflective of proven pedagogical need. Although teachers
acknowledged the diverse roles of the private sector in AIED and support for AIED
in schools (e.g., offering training bases for teachers, D17; supporting students’ AI-
related extra-activities such as contests and boot camp, E22 and building AI innova-
tion space in the school, E21), teachers claimed that AI that they currently encounter
and work with for the purpose of instruction, a specific application domain, has been
designed, developed, and deployed dominated by computer-centric views without
sufficient understanding and considerations of underlying education rationale and
adequate pedagogical principles (Holmes et al., 2019a, b). Thereby, the educational
application of these models may not be aligned with classroom instruction. In turn,
teachers argued that such pedagogically insufficient and incompetent AI may scale
up poor pedagogic practice, rather than supporting students and teachers in develop-
ing innovative approaches to learning and instruction, and thereby its roles would
ultimately diminish and remain merely as mundane back-end AI-as-a-service plug-
ins, or a specific type of recommender system, although its potential goes far beyond
these uses (Williamson & Eynon, 2020). 4.3.3 Absence of clear AI ethics In contrast with the health sector, where there are long-established ethical principles
and codes of practice with regard to the treatment of human subjects, an absence of
a commonly accepted model of ethics or a robust ethical framework, approach, and
procedures exists for AIED and TAC research which was found to be another criti-
cal challenge to further understanding and developing TAC for classroom instruc-
tion. In particular, as processes and outcomes of TAC have an unknown and unin-
tended impact on individual learner’s learning experience, and their neurological,
cognitive, and socio-emotional development for life (Gottschalk, 2019). As TAC is
used at scale, teachers expressed a great need for clear and robust ethical guidelines
and interventions that demonstrate AI efficacy and safety in the educational context,
including foreseeable misuse, through the entire life cycle of TAC use in classroom
instruction. Meanwhile, teachers argued that the ethics of data (i.e., data bias, privacy,
transparency, and trust in data analysis) and computational power (i.e., accu-
racy in prediction modeling and AI recommendation) are crucial but not suf-
ficient alone. Given that TAC is set to take place in diverse areas and processes 1 3 3 17458 Education and Information Technologies (2024) 29:17433–17465 in classroom instruction, the ethics of AI also needs to account for the ethics
of education including, but not limited to, the ethics of teacher expectations, of
resources allocations (including teacher expertise), of gender and ethnic biases,
of behavior and discipline, of the accuracy and validity of assessments, of what
constitutes useful knowledge, of teacher roles, of power relations between teach-
ers and their students, and of particular approaches to pedagogy, such as behav-
iorism versus constructivism (Holmes et al., 2022b). 5 Conclusion The present study explored teachers’ perceptions of diverse forms of TAC as well
as the opportunities of and obstacles to TAC pertinent to implementation in a
K-12 context. Participating teachers in the study recognized the diverse forms
of TAC that could be adopted to contribute to classroom instruction. Also, they
described opportunities and obstacles to TAC implementation in the K-12 con-
text. Based on the findings, this study provides implications for the current and
future AIED. First, it must be executed with deliberate long-term educational vision and
goals with built-in curriculum systems/structure and pedagogical supports to
maximize the potential of TAC and minimize barriers to impactful TAC. Teachers
in the study echo the barriers found in other studies related to shallow discussions
regarding the long-term visions and goals of AIED (Abdelaziz, 2019; Kim et al.,
2022). Although much of the integration of AIED and rationale for TAC has
been supported for the sake of personalization of learning, teachers in the study
expressed that the current forms of TAC are likely set to improve students’ aca-
demic performance in a standardized examination by spoon-feeding pre-specified
content, which is driven by the overwhelming attention to performance-driven
curriculum design. In this regard, teachers directed us to ponder upon the mean-
ing and purpose of education and the right kind of education we need to pursue
TAC. In fact, teachers are expected to realize and implement students’ life-long
learning and personalized learning both in the learning process and through out-
comes. Additionally, they are expected to incorporate diverse teaching pedagogy
such as guided discovery, productive failure, project-based learning, active learn-
ing, and deep learning to support students’ higher-order thinking rather than rote
memorization, while also empowering students’ critical and active engagement in
learning rather than simply knowing facts in the textbooks. This resonates with
the prior research suggesting that we should aspire and focus on leveraging the
power of teachers and AI to address genuine education as a ‘wicked problem’,
such as promoting students’ engagement and inclusion alongside knowledge
delivery and formative assessment and feedback (Holmes et al., 2022a). 4.3.4 Negative attitude toward AI Teachers in the study indicated that teachers’ biased and negative attitudes
toward AI could be another barrier to effective TAC in the classroom. Many
teachers made comments about their colleagues’ attitude toward AI during the
interviews and confided that most of their colleagues naively assume that AI
would replace human teachers, or they simply consider AI as another kind of
sophisticated machine that has been put into the classroom instruction forced
by the government. To some extent, their concern was valid; teachers without
knowing what AI is, what technical levels are required for teachers to interact
with AI, and how AI could work with teachers and students in what ways, could
have been overwhelmed with concerns and anxiety. However, one interesting
point to capture is that teachers have, in fact, been using technology in classroom
instruction and many of those technology tools are powered by AI (Schmidt,
2016). Teachers appeared to have formed a generally positive attitude towards
ICT usage in contrast to their critical view AI. For instance, D17 expressed: Teaching with ICTs has been promoted among Chinese education circles
for many years. I am sure all the teachers nowadays use ICTs, more or less. We have reached the point of no return. But they are not fully aware that
AI has been embedded into those technologies so our teaching practice has
now become convenient. This quote resonates with previous studies highlighting that it is crucial to
understand how people feel about AI, how people are affected by AI presence,
and how people’s attitudes toward AI may affect or bias subsequent interac-
tion with AI in order for AI to be integrated and accepted in human societies
(Nomura et al., 2006). Accordingly, teachers’ attitude toward AI constitutes
an important element for their teaching experience and performance with AI
as well as persistence in teaching with technology (Tas, 2016; Wentzel et al.,
2010). In addition, existing literature has shown that humans’ incomplete and
primitive understanding/conceptualization of AI can influence their interactions
with technology, and determine their actions with technologies (Phillips et al.,
2011). Also, an incomplete understanding could potentially lead humans to mis-
understand AI’s abilities and thereby create a pitfall for under-utilization (e.g.,
misuse or disuse). 3 1 Education and Information Technologies (2024) 29:17433–17465 17459 5 Conclusion It is,
therefore, crucial to acknowledge the varied beliefs, values, and assumptions in
the nature of education and establish informed public discussions around why,
what, how, and how well TAC reflects a bigger picture/definition of education
that takes account of contemporary and future learner needs and context should
come into first, otherwise the roles of AI will passively remain ancillary to class-
room instruction and scale up the poor pedagogical practice (Abdelaziz, 2019;
Holmes et al., 2022a).i In line with this, the findings revealed that the AI technologies that are meant
to collaborate with teachers in the educational domain are mainly developed and
given by the commercial sector. Although teachers acknowledge the support from
and collaboration with the commercial sector, they bring with it fundamental
ethical questions over what constitutes appropriate use and rationale by a private
company of learner-AI, teacher-AI, and learner-teacher-AI interactions, and what 1 3 Education and Information Technologies (2024) 29:17433–17465 17460 information and interventions they suggest or do not suggest to teachers. These
further link to the question of AI loyalty- for whom does the AI system work (Hol-
mes et al., 2022a). In this regard, earlier research argued that it is crucial to set
underlying educational rationale for the design and development of AI applica-
tions, design and implement robust ethical guidelines, and avoid ethics washing-
a growing instrumentalization of ethical language by private tech companies that
have been often used to justify their self-interest in design, test, and implement
their AI technologies (Knox, 2020; Bietti, 2020). To increase the transparency and
trustworthiness of the process and effects of TAC, system developers should be
obliged to explicitly align the loyalty of their AI systems and governance struc-
tures with the best interests of the learners and teachers as well as others affected
by the system. This should include measures to involve multiple stakeholders that
fall outside of the commercial sector in the AI tool’s design, procurement, and
deployment including learners, teachers, parents, researchers, tech providers, and
policymakers. This can serve to ensure multivocality on ethical oversight and
clear communication of the expectations of what will be done with both the main
and incidental impact of TAC. 5 Conclusion For the same reasons, there is a critical need for
appropriate professional development for teachers as well as for administrators and
policymakers, so that they can make informed decisions about which AI systems
might be appropriate for their classroom, how those systems broadly work with
teachers and students, what they might achieve, their challenges and risks, and
what unintended consequences there might be. Furthermore, teachers in the study characterized and anticipated the diverse
possibilities and forms of TAC by taking into account mutual/shared interaction
in which teachers and AI systems can augment each other’s strengths, not a uni-
lateral adaptivity in AI systems or teachers separately (Holstein et al., 2020). In
this regard, emerging lines of research are beginning to explore the design oppor-
tunities of human-AI co-orchestration systems that optimize the complementary
strengths of human intelligence and AI, so that the two together behave more
intelligently than the two separately (Dellermann et al., 2019), and it envisions
collaboration between human and artificial intelligence in a social, technologi-
cal ensemble (Cukurova et al., 2019). For instance, while teachers work with
students, AI systems follow along with teachers’ instruction and adaptively pre-
sent education resources (e.g., relevant readings, videos, or practice materials)
that support their current goals. Alternatively, a system may respond during or
after the teacher by adaptively providing feedback on the quality of the instruc-
tion (e.g., the clarity of a particular explanation the teacher provides), to help the
teachers adjust and improve over time (Holstein et al., 2019; Walker et al., 2014). Meanwhile, teachers can augment the set of instructional moves available to an
AI system by either customizing or creating new actions for the system (Heffer-
nan & Heffernan, 2014; Holstein et al., 2019). Yet, there are numerous challenges
in conceptualizing and developing such systems. Four challenges are set out by
Akata et al. (2020): (1) how do we develop AI systems that work in synergy with
humans, (2) how do these systems learn from and adapt to their environment,
(3) how do we ensure ethical and responsible behavior, and (4) how can humans
and AI share and explain their awareness, goals, and strategies to each other? 1 3 Education and Information Technologies (2024) 29:17433–17465 17461 Developing collaborative, adaptive, responsible, and explainable hybrid intelli-
gent systems is important across many application domains but is especially use-
ful for our thinking about the role of AI in education. 5 Conclusion The focus on the interplay
between learners, teachers, and AI to optimize learning will be central to the suc-
cessful application of AI in this field. Future research thereby takes a step for-
ward and towards the development of richer instructional theory, practice, and
technology for TAC, rather than simply limiting the value of AI development and
applications in education to accessing, organizing, and managing data to support
teachers’ decision-making. More research needs to be conducted to understand
the nature of TAC and the interaction between teachers and AI based on learn-
ing and teaching theories, to develop feasible instructional models and strategies
to assist teachers in navigating and formulating more meaningful and impactful
TAC and to advance AI models and technology to support TAC.i Although the present study presented important findings and implications for
structuring and implementing TAC in the context of K-12 instruction, the study
recognizes some limitations that should be addressed in a future study. To begin
with, the current study was conducted by reviewing the storyboards that have been
developed by instructional experts, rather than directly interacting with AI systems,
although participating teachers mostly have extensive teaching experience in AIED. This might not have fully revealed participants’ perceptions about TAC. If partic-
ipants have direct and longitudinal collaborative interactions with the AI systems
in the real classroom environment, their perceptions may change over time. Future
studies can examine teachers’ experiences and views of actual collaboration with
AI systems on various teaching and administrative tasks. One possible way is to
conduct a virtual experiment/simulation using AI-embedded virtual and immersive
technologies or the Wizard of Oz method, a prototyping method that involves inter-
action with a mock interface manually controlled by operators (e.g., Kasepalu et al.,
2022) to explore how teachers build a sequence of actions TAC on a task given to
map a variety of types of TAC. Furthermore, a total number of 30 participating
teachers in the study may somewhat limit the generalization of the findings. Future
research thereby needs to increase the number of research participants and consider
quantitative data collection and analysis. Authors’ contributions Jinhee Kim was fully responsible for the research design, material preparation,
data collection and analysis. The entire draft of the manuscript was also written by Jinhee Kim. Funding The author has no relevant financial or non-financial interests to disclose. Data availability N/A. Code availability N/A. Code availability N/A. Authors’ contributions Jinhee Kim was fully responsible for the research design, material preparation,
data collection and analysis. The entire draft of the manuscript was also written by Jinhee Kim. Authors’ contributions Jinhee Kim was fully responsible for the research design, material preparation,
data collection and analysis. The entire draft of the manuscript was also written by Jinhee Kim.
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which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long
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Man-Specific Lectins from Plants, Fungi, Algae and Cyanobacteria, as Potential Blockers for SARS-CoV, MERS-CoV and SARS-CoV-2 (COVID-19) Coronaviruses: Biomedical Perspectives
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Cells
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Johansson
Received: 28 April 2021
Accepted: 25 June 2021
Published: 28 June 2021 Citation: Barre, A.; Van Damme,
E.J.M.; Simplicien, M.; Le Poder, S.;
Klonjkowski, B.; Benoist, H.; Peyrade,
D.; Rougé, P. Man-Specific Lectins
from Plants, Fungi, Algae and
Cyanobacteria, as Potential Blockers
for SARS-CoV, MERS-CoV and
SARS-CoV-2 (COVID-19)
Coronaviruses: Biomedical
Perspectives. Cells 2021, 10, 1619. https://doi.org/10.3390/cells10071619 Received: 28 April 2021
Accepted: 25 June 2021
Published: 28 June 2021 Received: 28 April 2021
Accepted: 25 June 2021
Published: 28 June 2021 Keywords: plant lectins; mannose-specific lectins; plant lectins; algae lectins; fungi lectins; cyanobac-
teria lectins; pea lectin; artocarpin; snowdrop lectin; biomedical applications; coronaviruses; SARS-
CoV; MERS-CoV; SARS-CoV-2; COVID-19 Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations. Man-Specific Lectins from Plants, Fungi, Algae and Cyanobacteria,
as Potential Blockers for SARS-CoV, MERS-CoV and
SARS-CoV-2 (COVID-19) Coronaviruses: Biomedical Perspectives Annick Barre 1
, Els J. M. Van Damme 2
, Mathias Simplicien 1
, Sophie Le Poder 3, Bernard Klonjkowski 3,
Hervé Benoist 1, David Peyrade 4 and Pierre Rougé 1,* 1
UMR 152 PharmaDev, Institut de Recherche et Développement, Faculté de Pharmacie, Université Paul
Sabatier, F-31062 Toulouse, France; annick.barre@univ-tlse3.fr (A.B.); simplicien.mathias@gmail.com (M.S.);
herve.benoist@ird.fr (H.B.) 2
Department of Biotechnology, Faculty of Bioscience Engineering, Ghent University, B-9000 Ghent, Belgium;
elsjm.vandamme@ugent.be 2
Department of Biotechnology, Faculty of Bioscience Engineering, Ghent University, B-9000 Ghent, Belgium;
elsjm.vandamme@ugent.be j
g
3
UMR Virologie, INRA, ANSES, Ecole Nationale Vétérinaire d’Alfort, F-94700 Maisons-Alfort, France;
sophie.lepoder@vet-alfort.fr (S.L.P.); bklonjkowski@vet-alfort.fr (B.K.)
4 3
UMR Virologie, INRA, ANSES, Ecole Nationale Vétérinaire d’Alfort, F-94700 Maisons-Alfort, France;
sophie.lepoder@vet-alfort.fr (S.L.P.); bklonjkowski@vet-alfort.fr (B.K.)
4
UMR5129 Laboratoire des Technologies de la Microélectonique, Université Grenoble Alpes, CNRS, CEA-Leti
Minatec, CEDEX, F-38054 Grenoble, France; david.peyrade@cea.fr
*
Correspondence: pierre.rouge.perso@gmail.com; Tel.: +33-069-552-0851 sophie.lepoder@vet alfort.fr (S.L.P.); bklonjkowski@vet alfort.fr (B.K.)
4
UMR5129 Laboratoire des Technologies de la Microélectonique, Université Grenoble Alpes, CNRS, CEA-Leti
Minatec, CEDEX, F-38054 Grenoble, France; david.peyrade@cea.fr
*
C
d
i
@
il
T l
33 069 552 0851 g
q
,
p
,
,
Minatec, CEDEX, F-38054 Grenoble, France; david.peyrade@cea.fr p y
*
Correspondence: pierre.rouge.perso@gmail.com; Tel.: +33-069-552-0851 cells cells
Abstract: Betacoronaviruses, responsible for the “Severe Acute Respiratory Syndrome” (SARS)
and the “Middle East Respiratory Syndrome” (MERS), use the spikes protruding from the virion
envelope to attach and subsequently infect the host cells. The coronavirus spike (S) proteins contain
receptor binding domains (RBD), allowing the specific recognition of either the dipeptidyl peptidase
CD23 (MERS-CoV) or the angiotensin-converting enzyme ACE2 (SARS-Cov, SARS-CoV-2) host
cell receptors. The heavily glycosylated S protein includes both complex and high-mannose type
N-glycans that are well exposed at the surface of the spikes. A detailed analysis of the carbohydrate-
binding specificity of mannose-binding lectins from plants, algae, fungi, and bacteria, revealed that,
depending on their origin, they preferentially recognize either complex type N-glycans, or high-
mannose type N-glycans. Since both complex and high-mannose glycans substantially decorate the S
proteins, mannose-specific lectins are potentially useful glycan probes for targeting the SARS-CoV,
MERS-CoV, and SARS-CoV-2 virions. Mannose-binding legume lectins, like pea lectin, and monocot
mannose-binding lectins, like snowdrop lectin or the algal lectin griffithsin, which specifically
recognize complex N-glycans and high-mannose glycans, respectively, are particularly adapted
for targeting coronaviruses. The biomedical prospects of targeting coronaviruses with mannose-
specific lectins are wide-ranging including detection, immobilization, prevention, and control of
coronavirus infection.
Citation: Barre, A.; Van Damme,
E.J.M.; Simplicien, M.; Le Poder, S.;
Klonjkowski, B.; Benoist, H.; Peyrade,
D.; Rougé, P. Man-Specific Lectins
from Plants, Fungi, Algae and
Cyanobacteria, as Potential Blockers
for SARS-CoV, MERS-CoV and
SARS-CoV-2 (COVID-19)
Coronaviruses: Biomedical
Perspectives. Cells 2021, 10, 1619. https://doi.org/10.3390/cells10071619
Academic Editor: Mats W. Johansson
Received: 28 April 2021
Accepted: 25 June 2021
Published: 28 June 2021 Citation: Barre, A.; Van Damme,
E.J.M.; Simplicien, M.; Le Poder, S.;
Klonjkowski, B.; Benoist, H.; Peyrade,
D.; Rougé, P. Man-Specific Lectins
from Plants, Fungi, Algae and
Cyanobacteria, as Potential Blockers
for SARS-CoV, MERS-CoV and
SARS-CoV-2 (COVID-19)
Coronaviruses: Biomedical
Perspectives. Cells 2021, 10, 1619. https://doi.org/10.3390/cells10071619
Academic Editor: Mats W. Johansson
Received: 28 April 2021
Accepted: 25 June 2021
Published: 28 June 2021 Citation: Barre, A.; Van Damme,
E.J.M.; Simplicien, M.; Le Poder, S.;
Klonjkowski, B.; Benoist, H.; Peyrade,
D.; Rougé, P. Man-Specific Lectins
from Plants, Fungi, Algae and
Cyanobacteria, as Potential Blockers
for SARS-CoV, MERS-CoV and
SARS-CoV-2 (COVID-19)
Coronaviruses: Biomedical
Perspectives. Cells 2021, 10, 1619. https://doi.org/10.3390/cells10071619
Academic Editor: Mats W. 1. Introduction The S1 subunit consists of a signal peptide (S), followed by the N-terminal
domain (NTD) containing the receptor-binding domain (RBD) interacting with the ACE2 receptor via the RBM motif (RBM). The S2 subunit comprises the fusion peptide (FP), two heptad repeats HR1 and HR2, the transmembrane spanning domain
(TM) that anchors the S protein into the virion membrane, and the cytoplasmic domain CP. The S proteins forming the coronavirus spikes, consist of heavily N-glycosylated pro-
teins containing a high number of potential N-glycosylation sites NXT/S, actually occupied
by complex-type glycans, high-mannose type glycans, and a few hybrid type glycans:
22 sites for SARS-CoV and SARS-CoV-2, and 23 sites for MERS-CoV, respectively [11,12]. A very limited number of potential O-glycosylation sites occupied by GalNAc and T/Tn-
antigens have been identified, especially in the SARS-CoV-2 S-glycoprotein [12]. According
to their heavily N-glycosylated character, S-glycoproteins of coronaviruses represent partic-
ularly relevant targets for mannose-specific (Man-specific) lectins, a heterogeneous group
of proteins involved in the recognition of Man, mannosides, and complex Man-containing
glycans, already known for their potential anti-viral and anti-cancer properties [13–18]. g y
y
p
p
p
The Man-specific lectins are widely distributed in all the living organisms since
they have been isolated and characterized from viruses, bacteria, fungi, algae, plants,
animals, and humans [19]. Even though they constitute a disparate group of proteins
poorly related by their phylogenetic relationship and structural features, they share a
very common property to specifically recognize Man and their derivatives, including
mannosides, oligomannosides, and high-mannose glycans [20]. This specific recognition
also concerns other Man-containing glycans like the complex N-glycans which are built
around a trimannosyl Manα1,3Manα1,6Man core, readily recognized by Man-specific
legume lectins [21,22]. As shown for plant lectins, the differences observed in the specific
recognition of Man-containing glycans, essentially depend on both the axial position of the
hydroxyl group at C2 in Man which is more or less strictly recognized by the carbohydrate-
binding sites (CBSs) of lectins, and the type of glycosidic bonds α1,2, α1,3 or α1,6, and
their internal or external position in the glycan chain, recognized by lectins [23]. According
to these discrepancies and with respect to the diversity of N-glycans occurring at the
surface of the coronavirus spikes, many distinct Man-specific lectins from plants, algae,
fungi, and bacteria, could serve as relevant glycan probes for SARS-CoV, MERS-CoV, and
SARS-CoV-2 viruses. 1. Introduction The human Betacoronaviruses SARS-CoV, MERS-CoV, and SARS-CoV-2 responsible
for the pandemic COVID 19 that still rages worldwide, share common structural char-
acteristics that account for their transmissibility and infectivity among sensitive people. The virions are covered by spikes, which protrude outside of the lipid bilayer forming the
envelope of coronaviruses [1–3]. Spikes are built up from the homotrimeric association
of the S protein), an acidic class I viral fusion glycoprotein of ~130 kDa which consists of
two S1 and S2 subunits [4–7]. The S protein is cleaved between the S1 and S2 subunits
(S1/S2 cleavage site) and at an additional site S2’ by host cell protease. The S1 subunit Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article
distributed
under
the
terms
and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/). https://www.mdpi.com/journal/cells Cells 2021, 10, 1619. https://doi.org/10.3390/cells10071619 Cells 2021, 10, 1619 2 of 36 contains a receptor-binding domain (RBD), which allows the virus to specifically recognize
host cell receptors via a receptor-binding motif (RBM). The dipeptidyl peptidase 4 (DPP4),
also known as CD26, was identified as the host receptor for MERS-CoV [8], whereas the
angiotensin-converting enzyme 2 (ACE2) serves as the host receptor for SARS-CoV and
SARS-CoV-2 [9]. The S2 subunit is subsequently involved in the fusion of both coron-
avirus and host cell membranes through the fusion peptide (FP) and the first heptad repeat
HR1 [10], allowing the entry of the viral RNA genome into the host cell, which will further
trigger the replication of the coronavirus at the detriment of the host cell (Figure 1). Figure 1. Molecular organization of the SARS-CoV-2 S-glycoprotein, made up of two linked S1 and S2 subunits that are
further separated at the furin cleavage site (FCS). The S1 subunit consists of a signal peptide (S), followed by the N-terminal
domain (NTD) containing the receptor-binding domain (RBD) interacting with the ACE2 receptor via the RBM motif (RBM). The S2 subunit comprises the fusion peptide (FP), two heptad repeats HR1 and HR2, the transmembrane spanning domain
(TM) that anchors the S protein into the virion membrane, and the cytoplasmic domain CP. Figure 1. Molecular organization of the SARS-CoV-2 S-glycoprotein, made up of two linked S1 and S2 subunits that are
further separated at the furin cleavage site (FCS). 2. Man-Specific Lectins from Higher Plants, Fungi, Algae, and Cyanobacteria Mannose-specific lectins consist of a group of lectins that are widely distributed
in all living organisms and, especially, in higher plants (monocot and dicot plants), in
fungi (Ascomycota and Basidiomycota), in the different groups of algae (essentially red
and green algae), and in bacteria, mainly the cyanobacteria (formerly known as blue
algae). Although they belong to quite different structural scaffolds [20], all of these lectins
contain CBSs that display a more or less strict specificity for mannose (Man) and Man-
containing glycoproteins. 1. Introduction In addition to their Man-binding capacity, Man-specific lectins from plants, algae,
fungi, and bacteria, have been largely studied with respect to their anti-viral properties
against different types of enveloped viruses, including HIV-1 [24], papilloma virus [25],
herpes virus, hepatitis C virus [26], and Ebola virus [27]. In this respect, the algal lectin grif-
fithsin [28], the cyanobacteria lectins cyanovirin [29], actinohivin [30], and microvirin [31], Cells 2021, 10, 1619 3 of 36 and various GNA-related lectins like NPA from daffodil (Narcissus pseudonarcissus) and
ASA from garlic (Allium sativum) [32–34], have been particularly well documented. Most
of these Man-specific lectins prevent the virus replication, at least under in vitro conditions,
by interfering with the Man-containing N-glycans present on the cell surface of the virion
envelope [35]. Depending on the virus, different Man-containing glycans serve as targets
for the Man-specific lectins, e.g., gp120 for HIV-1 [36] or hemagglutinin for influenza
virus [37]. Owing to their smaller size, compared to other plant Man-specific lectins, Man-
specific lectins from cyanobacteria such as CVN-N (cyanovirin), MVN (microvirin), and
AH (actinohivin) from actinobacteria, have been deeply investigated for their anti-viral
properties. In addition, only very few publications report the ability of Man-specific lectins
from higher plants to interact with glycans of the coronavirus envelope [13,38–40]. The present review aims at presenting a detailed analysis of the Man-specific lectins
from plants, algae, fungi, and bacteria that could serve as relevant probes for targeting the
N-glycans coating the spikes of SARS-CoV, MERS-CoV, and SARS-CoV-2 coronaviruses,
with a focus on their potential biomedical applications in controlling coronavirus infections,
notably the COVID-19 pandemic associated with SARS-CoV-2. 2.1. Man-Specific Lectins from Higher Plants Seeds and tubers of higher plants often contain Man-specific lectins which have been
associated with different structural scaffolds (Figure 2): -
The β-sandwich structure (jelly-roll structural scaffold), present in legume lectins. They are classically divided into two groups of single- and two-chain lectins, ac-
cording to the cleavage (two-chain) or the absence of cleavage (single-chain) of their
protomers. Single-chain, tetravalent lectins consist of the non-covalent association of
four identical protomers (α4), each containing a CBS that specifically recognizes Man
and its derivatives. In two-chain lectins, a proteolytic cleavage of the protomeric chain
results in the separation of a light (α-chain) and a heavy (β-chain), which remain
associated with non-covalent bonds. Two-chain, bivalent lectins (α2β2)) result from
the non-covalent association of two identical two-chain protomers, each possessing a
CBS specific for Man and its derivatives. -
The β-sandwich structure (jelly-roll structural scaffold), present in legume lectins. They are classically divided into two groups of single- and two-chain lectins, ac-
cording to the cleavage (two-chain) or the absence of cleavage (single-chain) of their
protomers. Single-chain, tetravalent lectins consist of the non-covalent association of
four identical protomers (α4), each containing a CBS that specifically recognizes Man
and its derivatives. In two-chain lectins, a proteolytic cleavage of the protomeric chain
results in the separation of a light (α-chain) and a heavy (β-chain), which remain
associated with non-covalent bonds. Two-chain, bivalent lectins (α2β2)) result from
the non-covalent association of two identical two-chain protomers, each possessing a
CBS specific for Man and its derivatives. Cells 2021, 10, 1619 4 of 36 Figure 2. Overview of lectins from representative genera/species belonging to higher plant familie
with their associated structural scaffolds, able to recognize the spike proteins from coronaviruse
(Coronavirus Credit: Maria Voigt/RCSB PDB). A-K correspond to lectins used as structural sca
folds ((A): Con A, (B): LcA, (C): BanLec, (D): Morniga-M, (E): PPL, (F): Heltuba, (G): Gastrodianin
(H): ASA, (I): GNA, (J): Nictaba, (K): IraB. References [41–98] correspond to the lectins presented i
the figure. Both Con A (concanavalin A) from jack bean seeds (Canavalia ensiformis) (Figure 2A Figure 2. Overview of lectins from representative genera/species belonging to higher plant families,
with their associated structural scaffolds, able to recognize the spike proteins from coronaviruses
(Coronavirus Credit: Maria Voigt/RCSB PDB). 2.1. Man-Specific Lectins from Higher Plants A-K correspond to lectins used as structural scaf-
folds ((A): Con A, (B): LcA, (C): BanLec, (D): Morniga-M, (E): PPL, (F): Heltuba, (G): Gastrodianin,
(H): ASA, (I): GNA, (J): Nictaba, (K): IraB. References [41–98] correspond to the lectins presented in
the figure. Both Con A (concanavalin A) from jack bean seeds (Canavalia ensiformis) (Figure 2A)
and LcA (lentil lectin) from Lens culinaris (Figure 2B) are classical examples of single-chain
and two-chain lectins, respectively. -
The β-prism I structure (β-barrel structural scaffold), present in the jacalin-related
lectin group. Their protomers are organized in three bundles of four antiparallel
β-strands arranged into a β-prism structure along a longitudinal axis. Depending on -
The β-prism I structure (β-barrel structural scaffold), present in the jacalin-related
lectin group. Their protomers are organized in three bundles of four antiparallel
β-strands arranged into a β-prism structure along a longitudinal axis. Depending on Cells 2021, 10, 1619 5 of 36 the number of identical protomers associated by non-covalent bonds, Man-specific
jacalin-related lectins consist of bivalent (two CBSs) lectins, e.g., Calsepa from hedge
bindweed Calystegia sepium [61] (Figure 2C); tetravalent (four CBSs) lectins, e.g.,
Artocarpin from black mulberry Artocarpus integer [65] (Figure 2D); hexavalent (six
CBSs) lectins, e.g., PPL from the African locust bean Parkia platycephala [70] (Figure 2E);
or octavalent (eight CBSs) lectins, e.g., Heltuba from the Jerusalem artichoke Helianthus
tuberosus [71] (Figure 2F). (
g
)
The β-prism II structure (β-trefoil structural scaffold), present in the GNA-related
lectin group. The protomer consists of three bundles of four antiparallel β-strands
arranged into a flattened β-trefoil structure around a central pseudoaxis. A CBS occurs
in each of the bundles of β-strands. Except for gastrodianin, which is composed of
a single protomer (Figure 2G), lectins belonging to the GNA-related group result
from the non-covalent association of two protomers, e.g., the hexavalent NPA from
daffodil (Narcissus pseudonarcissus) bulbs [80] (Figure 2H) or four protomers, e.g., the
dodecavalent GNA from snowdrop (Galanthus nivalis) bulbs [94] (Figure 2I). g
The β-prism II structure (β-trefoil structural scaffold), present in the GNA-related
lectin group. The protomer consists of three bundles of four antiparallel β-strands
arranged into a flattened β-trefoil structure around a central pseudoaxis. A CBS occurs
in each of the bundles of β-strands. 2.1. Man-Specific Lectins from Higher Plants Except for gastrodianin, which is composed of
a single protomer (Figure 2G), lectins belonging to the GNA-related group result
from the non-covalent association of two protomers, e.g., the hexavalent NPA from
daffodil (Narcissus pseudonarcissus) bulbs [80] (Figure 2H) or four protomers, e.g., the
dodecavalent GNA from snowdrop (Galanthus nivalis) bulbs [94] (Figure 2I). Two covalently linked β-trefoil-domains form the B-chain of the lectin from Iris
hollandica, a type II ribosome-inactivating protein (RIP-II) (Figure 2J). Each domain
contains a CBS which can accommodate Man and Gal/GalNAc. This Man-binding
property confers an unusual Man-specificity to the Iris RIP-II, which readily differs
from the Gal/GalNAc-specificity of other RIP-IIs [97]. -
Nictaba, the jasmonic acid-induced tobacco (Nicotiana tabacum) lectin displays speci-
ficity for chitin and GlcNAc oligomers but it can also bind to high mannose glycans
with a lower affinity, as shown from glycan array experiments. The molecular struc-
ture of Nictaba is still unknown but modeling experiments suggest a β-sandwich
structure for this tobacco lectin [98] (Figure 2K). All of these lectins readily interact with Man and oligomannosides but, more inter-
estingly, they display a higher affinity for more complex Man-containing glycan chains
that are usually associated with cell surfaces. In this respect, different Man-specific lectins
have been co-crystallized in the presence of Man (Figure 3A–C), di-mannoside (Figure 4A),
tri-mannoside core Manα1,3Manα1,6Man (Figure 4B,C), and tetra-mannoside (Figure 4D)
occurring in N-glycans of the complex and high-mannose type. Figure 3. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino acid
residues A30, E31, D81, G99, and N135, forming the CBS of LoLI from Lathyrus ochrus (PDB code
1LOB). Aromatic residue F123 participating in stacking interaction with the pyranose ring of Man
is colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the
amino acid residues G27, G149, V151, and D153, forming the CBS of Morniga-M from Morus nigra
(PDB code 1XXR). Aromatic residue F150 involved in stacking interaction with the pyranose ring of
Man is colored orange. (C). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to
the amino acid residues Q90, D92, and N94, forming the CBS of PCL from Polygonatum cyrtonema
(PDB code 3A0D). Aromatic residue Y98 participating in stacking interaction with the pyranose ring
of Man is colored orange. Figure 3. (A). 2.1. Man-Specific Lectins from Higher Plants Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino acid
residues A30, E31, D81, G99, and N135, forming the CBS of LoLI from Lathyrus ochrus (PDB code
1LOB). Aromatic residue F123 participating in stacking interaction with the pyranose ring of Man
is colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the
amino acid residues G27, G149, V151, and D153, forming the CBS of Morniga-M from Morus nigra
(PDB code 1XXR). Aromatic residue F150 involved in stacking interaction with the pyranose ring of
Man is colored orange. (C). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to
the amino acid residues Q90, D92, and N94, forming the CBS of PCL from Polygonatum cyrtonema
(PDB code 3A0D). Aromatic residue Y98 participating in stacking interaction with the pyranose ring
of Man is colored orange. Figure 3. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino acid
residues A30, E31, D81, G99, and N135, forming the CBS of LoLI from Lathyrus ochrus (PDB code
1LOB). Aromatic residue F123 participating in stacking interaction with the pyranose ring of Man
is colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the
amino acid residues G27, G149, V151, and D153, forming the CBS of Morniga-M from Morus nigra
(PDB code 1XXR). Aromatic residue F150 involved in stacking interaction with the pyranose ring of
Man is colored orange. (C). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to
the amino acid residues Q90, D92, and N94, forming the CBS of PCL from Polygonatum cyrtonema
(PDB code 3A0D). Aromatic residue Y98 participating in stacking interaction with the pyranose ring
of Man is colored orange. Figure 3. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino acid Cells 2021, 10, 1619 6 of 36 Figure 4. (A). Network of hydrogen bonds (black dashed lines) anchoring Manα1,2Man (M2) to
Heltuba from Helianthus tuberosus (PDB code 1C3N). Aromatic residues Y89 and Y90 participating in
stacking interactions with the pyranose rings of mannobiose are colored orange. (B). Network of
hydrogen bonds (black dashed lines) anchoring Manα1,6Manα1,3Man (M3) to GNA from Galanthus
nivalis (PDB code 1JPC). Aromatic residues Y89 and Y90 participating in stacking interactions with
the pyranose rings of mannobiose are colored orange. (C). 2.1. Man-Specific Lectins from Higher Plants Network of hydrogen bonds (black dashed
lines) anchoring Manα1,3Manα1,6Man (M3) to PAL from Pterocarpus angolensis (PDB code 1Q8V). Aromatic residues Y89 and Y90 participating in stacking interactions with the pyranose rings of
mannobiose are colored orange. (D). Network of hydrogen bonds (black dashed lines) anchoring
mannotetraose (M4) to artocarpin from Artocarpus integer (PDB code 1VBP). Figure 4. (A). Network of hydrogen bonds (black dashed lines) anchoring Manα1,2Man (M2) to g
( )
y
g
(
)
g
(
)
Heltuba from Helianthus tuberosus (PDB code 1C3N). Aromatic residues Y89 and Y90 participating in
stacking interactions with the pyranose rings of mannobiose are colored orange. (B). Network of
hydrogen bonds (black dashed lines) anchoring Manα1,6Manα1,3Man (M3) to GNA from Galanthus
nivalis (PDB code 1JPC). Aromatic residues Y89 and Y90 participating in stacking interactions with
the pyranose rings of mannobiose are colored orange. (C). Network of hydrogen bonds (black dashed
lines) anchoring Manα1,3Manα1,6Man (M3) to PAL from Pterocarpus angolensis (PDB code 1Q8V). Aromatic residues Y89 and Y90 participating in stacking interactions with the pyranose rings of
mannobiose are colored orange. (D). Network of hydrogen bonds (black dashed lines) anchoring
mannotetraose (M4) to artocarpin from Artocarpus integer (PDB code 1VBP). Heltuba from Helianthus tuberosus (PDB code 1C3N). Aromatic residues Y89 and Y90 participating in
stacking interactions with the pyranose rings of mannobiose are colored orange. (B). Network of
hydrogen bonds (black dashed lines) anchoring Manα1,6Manα1,3Man (M3) to GNA from Galanthus
nivalis (PDB code 1JPC). Aromatic residues Y89 and Y90 participating in stacking interactions with
the pyranose rings of mannobiose are colored orange. (C). Network of hydrogen bonds (black dashed
lines) anchoring Manα1,3Manα1,6Man (M3) to PAL from Pterocarpus angolensis (PDB code 1Q8V). Aromatic residues Y89 and Y90 participating in stacking interactions with the pyranose rings of
mannobiose are colored orange. (D). Network of hydrogen bonds (black dashed lines) anchoring
mannotetraose (M4) to artocarpin from Artocarpus integer (PDB code 1VBP). In addition, results from glycan arrays and binding assays, and X-ray crystallography
of lectins co-crystallized in the presence of complex glycan chains revealed that Man-
binding lectins from higher plants also accommodate more complex N-glycan chains,
through a complex network of hydrogen bonds and stacking interactions between the
pyranose ring of sugars and aromatic residues (F, W, Y) located in the vicinity of the CBS of
lectins (Figure 5A–C). Figure 5. (A). 2.1. Man-Specific Lectins from Higher Plants Network of hydrogen bonds (black dashed lines) anchoring Man9 (M9) to PAL, the
Pterocarpus angolensis lectin (PDB code 2PHN). Aromatic residues F132 and Y133 participating in
stacking interactions with the pyranose rings of the oligomannoside are colored orange. (B). Network
of hydrogen bonds (black dashed lines) anchoring N2 oligosaccharide to LoLII, isolectin II from
Lathyrus ochrus (PDB code 1LGB). Aromatic residues Y77, Y100, F123, Y124, and W128 involved
in stacking interactions with the pyranose rings of the oligosaccharide, are colored orange. The
α1,6-linked fucose (Fuc) participates in the H-bond network. (C). Network of hydrogen bonds (black
dashed lines) anchoring a biantennary glycan (GLY) to Calsepa from Calystegia sepium (PDB code
5XFI). Aromatic residue Y141 and Y142 participating in stacking interaction with the pyranose rings
of the glycan are colored orange. Figure 5. (A). Network of hydrogen bonds (black dashed lines) anchoring Man9 (M9) to PAL, the Figure 5. (A). Network of hydrogen bonds (black dashed lines) anchoring Man9 (M9) to PAL, the
Pterocarpus angolensis lectin (PDB code 2PHN). Aromatic residues F132 and Y133 participating in
stacking interactions with the pyranose rings of the oligomannoside are colored orange. (B). Network
of hydrogen bonds (black dashed lines) anchoring N2 oligosaccharide to LoLII, isolectin II from
Lathyrus ochrus (PDB code 1LGB). Aromatic residues Y77, Y100, F123, Y124, and W128 involved
in stacking interactions with the pyranose rings of the oligosaccharide, are colored orange. The
α1,6-linked fucose (Fuc) participates in the H-bond network. (C). Network of hydrogen bonds (black
dashed lines) anchoring a biantennary glycan (GLY) to Calsepa from Calystegia sepium (PDB code
5XFI). Aromatic residue Y141 and Y142 participating in stacking interaction with the pyranose rings
of the glycan are colored orange. Figure 5. (A). Network of hydrogen bonds (black dashed lines) anchoring Man9 (M9) to PAL, the
Pterocarpus angolensis lectin (PDB code 2PHN). Aromatic residues F132 and Y133 participating in
stacking interactions with the pyranose rings of the oligomannoside are colored orange. (B). Network
of hydrogen bonds (black dashed lines) anchoring N2 oligosaccharide to LoLII, isolectin II from
Lathyrus ochrus (PDB code 1LGB). Aromatic residues Y77, Y100, F123, Y124, and W128 involved
in stacking interactions with the pyranose rings of the oligosaccharide, are colored orange. The
α1,6-linked fucose (Fuc) participates in the H-bond network. (C). Network of hydrogen bonds (black
dashed lines) anchoring a biantennary glycan (GLY) to Calsepa from Calystegia sepium (PDB code
5XFI). 2.1. Man-Specific Lectins from Higher Plants Aromatic residue Y141 and Y142 participating in stacking interaction with the pyranose rings
of the glycan are colored orange. N-glycans are classified into three types, including oligomannose or high-mannose
glycans, complex glycans, and hybrid glycans, according to the extensions of their common
core sequence Manα1,3(Manα1,6)Manβ1,4GlcNAcβ1,4GlcNAcβ1-Asn (Figure 6A–C): 7 of 36 Cells 2021, 10, 1619 Figure 6. (A). High-mannose N-glycan, in which only Man (green circles) residues extend the core. (B). Complex N-glycans with two antennae initiated by a GlcNAc residue (blue square) extending the
core. (C). Hybrid N-glycan with Man extending the Man1,6 arm of the core and GlcNAc extending
the Man1,3 arm of the core. The core is delineated by a dashed red line. Figure 6. (A). High-mannose N-glycan, in which only Man (green circles) residues extend the core. (B). Complex N-glycans with two antennae initiated by a GlcNAc residue (blue square) extending the
core. (C). Hybrid N-glycan with Man extending the Man1,6 arm of the core and GlcNAc extending
the Man1,3 arm of the core. The core is delineated by a dashed red line. Glycan array analyses were performed by the Consortium for Functional Glycomics
(http://www.functionalglycomics.org, accessed on 14 March 2021) for Con A, a typical
member of the Man-specific single-chain lectins from the Fabaceae. Con A yielded the best
interaction with the following glycans arranged in decreasing order of affinity (Figure 7): Figure 7. Glycans recognized by Con A in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Glycan array analyses performed for PsA, a typical member of the Man-specific tw by Con A in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 7. Glycans recognized by Con A in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Glycan array analyses performed for PsA, a typical member of the Man-specific two-
chain lectins from the Vicieae (a tribe of Leguminosae/Fabaceae), yielded the best results
with the following glycans arranged in decreasing order of affinity (Figure 8): 8 of 36 Cells 2021, 10, 1619 Figure 8. Glycans recognized by PsA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 8. Glycans recognized by PsA in glycan array experiments, arranged from 1 to 10 in decreasin ans recognized by PsA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. 2.1. Man-Specific Lectins from Higher Plants Glycan array results obtained for GNA, the Man-specific lectin from snowdrop, the
prototype of monocot Man-binding lectins, are in decreasing order of affinity (Figure 9): Glycan array results obtained for GNA, the Man-specific lectin from snowdrop, the
prototype of monocot Man-binding lectins, are in decreasing order of affinity (Figure 9): Figure 9. Glycans recognized by GNA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 9. Glycans recognized by GNA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. gnized by GNA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 9. Glycans recognized by GNA in glycan array experiments, arranged from 1 to 10 in d Results obtained with Morniga-M, a typical representative of the jacalin-related Man-
binding lectins, reveal some reactivity towards bisected hybrid glycans (Figure 10): Figure 10. Glycans recognized by Morniga-M in glycan array experiments, arranged from 1 to 10 in decreasing order
of affinity. recognized by Morniga-M in glycan array experiments, arranged from 1 to 10 in decreasing order Figure 10. Glycans recognized by Morniga-M in glycan array experiments, arranged from 1 to 10 in decreasing order
f ffi Figure 10. Glycans recognized by Morniga-M in glycan array experiments, arranged from 1 to 10 in decreasing order
of affinity. From the glycan array results and X-ray crystallographic analysis of lectins in complex
with Man-containing glycans, it clearly appears that beyond a similar affinity for the
trimannosyl Manα1,6Manα1,3Man core occurring in both N-glycans of the complex, high-
mannose and hybrid types, Man-specific lectins from higher plants are not equivalent and
readily differ in their capacity to specifically recognize subtle chemical arrangements in the
complex glycan chains. In this respect, Man-specific legume lectins essentially recognize
complex glycans α1,6 fucosylated at the GlcNAc residue linked to asparagine, whereas
Man-specific lectins belonging to the GNA-related lectin group, essentially recognize more Cells 2021, 10, 1619 9 of 36 or less branched high-mannose N-glycans. The Man-specific lectins of the jacalin-related
group differ from the other lectins by their strong specificity for high-mannose glycans. Depending on the differences observed in the fine specificity towards complex glycans,
high-mannose glycans, and hybrid glycans, Man-specific lectins from diverse origins can
be used as glycan probes for targeting diverse types of N-glycans covering both host cell
and viral surfaces. 2.2. Man-Specific Lectins from Lower Plants and Fungi p
y
y
p
p
y
-
The β-prism II structure (β-trefoil structural scaffold), that builds the GNA-related
lectin protomer, also occurs in Marpola, the Man-specific lectin isolated from the
liverwort Marchantia polymorpha, and in mushrooms (Abmb from Agaricus bisporus,
MOA from Marasmius oreades). Special mention should be made to the apparent β-
trefoil structure that was found in the mini-lectin PhosL from the mushroom Pholiota
squarrosa. In fact, PhosL results from the trimeric association of three antiparallel
β-sheets, each possessing a CBS, that mimics a β-trefoil structure with three identical
CBSs. This newly identified lectin fold is involved in the specific recognition of the
core α1,6-fucosylation of fucosylated N-glycans [100] (Figure 12A,B). Figure 12. (A). Arrangement of the three antiparallel β-sheets (I, II, III) forming the PhosL ho-
motrimer (PDB code 5XZK) to mimic a pseudo-β-trefoil structure. (B). Network of hydrogen bonds
(black dashed lines) anchoring the fucose (Fuc, colored cyan) α1,6-linked for the first GlcNAc from
a fucosylated N-glycan to the CBS of PhosL (PDB code 6FX1). Aromatic residues Y89 and Y90
participating in stacking interactions with the pyranose rings of mannobiose are colored orange. Figure 12. (A). Arrangement of the three antiparallel β-sheets (I, II, III) forming the PhosL ho-
motrimer (PDB code 5XZK) to mimic a pseudo-β-trefoil structure. (B). Network of hydrogen bonds
(black dashed lines) anchoring the fucose (Fuc, colored cyan) α1,6-linked for the first GlcNAc from
a fucosylated N-glycan to the CBS of PhosL (PDB code 6FX1). Aromatic residues Y89 and Y90
participating in stacking interactions with the pyranose rings of mannobiose are colored orange. -
A rather unusual 6-bladded β-propeller organization was identified in tectonin 2, the
Man-specific lectin from the basidiomycete Laccaria bicolor [109]. As commonly ob-
served in other lectins exhibiting a β-propeller structure, tectonin 2 contains 6 identical β p
p
g
,
Man-specific lectin from the basidiomycete Laccaria bicolor [109]. As commonly ob-
served in other lectins exhibiting a β-propeller structure, tectonin 2 contains 6 identical served in other lectins exhibiting a β propeller structure, tectonin 2 contains 6 identical
Man-specific CBSs located at the top of each of the six blades forming the β-propeller. g
p
p
Man-specific CBSs located at the top of each of the six blades forming the β-propeller. 2.2. Man-Specific Lectins from Lower Plants and Fungi Although they are less documented compared to the lectins from higher plants, a
few Man-specific lectins have also been identified in lower plants including Bryophyta,
Pteridophyta, and Gymnospermae. Man-specific lectins also occur in both groups of
fungi, Ascomycota and Basidiomycota. They usually consist of structural scaffolds very
similar to those found in higher plant lectins but few of them also exhibit quite unusual
structural scaffolds like the β-propeller fold found in fungi or the α + β fold found in the
Gymnospermae (Figure 11A–L): Figure 11. Overview of lectins from the major representative genera/species of lower plants and fungi,
with their associated structural scaffolds, able to recognize the spike proteins from coronaviruses (Coron-
avirus Credit: Maria Voigt/RCSB PDB). A-L correspond to lectins used as structural scaffolds (A): Marpola,
(B): PAL, (C): CrCVNH, (D): RCLL-N, (E): Gnk2, (F): PeCL, (G): Flo1p, (H): TbCVNH, (I): Abmb, (J): HRL,
(K): Tectonin 2, (L): MOA. References [99–111] correspond to the lectins presented in the figure. Figure 11. Overview of lectins from the major representative genera/species of lower plants and fungi,
with their associated structural scaffolds, able to recognize the spike proteins from coronaviruses (Coron-
avirus Credit: Maria Voigt/RCSB PDB). A-L correspond to lectins used as structural scaffolds (A): Marpola,
(B): PAL, (C): CrCVNH, (D): RCLL-N, (E): Gnk2, (F): PeCL, (G): Flo1p, (H): TbCVNH, (I): Abmb, (J): HRL,
(K): Tectonin 2, (L): MOA. References [99–111] correspond to the lectins presented in the figure. Cells 2021, 10, 1619 10 of 36 10 of 36 -
The β-sandwich structure (jelly-roll structural scaffold) typical for legume lectins, also
occurs in the Man-specific lectin PeCL from Penicillium chrysogenum, and in floculins
Flo1p from the yeast Saccharomyces pasteurianum and Flo5p from Saccharomyces cerevisiae. p
y
y
p
p
y
-
The β-prism II structure (β-trefoil structural scaffold), that builds the GNA-related
lectin protomer, also occurs in Marpola, the Man-specific lectin isolated from the
liverwort Marchantia polymorpha, and in mushrooms (Abmb from Agaricus bisporus,
MOA from Marasmius oreades). Special mention should be made to the apparent β-
trefoil structure that was found in the mini-lectin PhosL from the mushroom Pholiota
squarrosa. In fact, PhosL results from the trimeric association of three antiparallel
β-sheets, each possessing a CBS, that mimics a β-trefoil structure with three identical
CBSs. This newly identified lectin fold is involved in the specific recognition of the
core α1,6-fucosylation of fucosylated N-glycans [100] (Figure 12A,B). 2.2. Man-Specific Lectins from Lower Plants and Fungi -
Another quite unusual structural scaffold, which consists of an antiparallel five-
stranded β-sheet associated to two α-helices (α + β fold), occurs in the monomeric
ginkbilobin 2, a Man-specific lectin from Ginkgo biloba [103]. Hydrophilic residues
occurring on the loops connecting the β-strands associated with charged residues of
two β-strands, to form a Man-binding pocket located at the top of the monomer [103]. The mannose-binding specificity of lectins from lower plants (Bryophyta, Pterido-
phyta, Gymnospermae), yeasts, and mushrooms (Ascomycota and Basidiomycota), is
directed against both Man, mannosides, and more complex high-mannose glycans, as
shown from the X-ray crystallographic studies for various Man-specific lectins complexed
to Man and oligomannosides (Figure 13 and Figure 14). 11 of 36 Cells 2021, 10, 1619 Figure 13. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino
acid residues N11, R93, and E104, forming the CBS of ginkbilobin from Ginkgo biloba (PDB code
4XRE). Aromatic residue Y103 and F108 participating in weak stacking interactions with the pyranose
ring of Man are colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring Man
(M) to the amino acid residues Q98, D160, D161, V226, and S227, forming the CBS of flocculin Flo5
from Saccharomyces cerevisiae (PDB code 2XJP). Aromatic residues Y54 and W228 involved in stacking
interactions with the pyranose ring of Man are colored orange. Figure 13. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino
acid residues N11, R93, and E104, forming the CBS of ginkbilobin from Ginkgo biloba (PDB code
4XRE). Aromatic residue Y103 and F108 participating in weak stacking interactions with the pyranose
ring of Man are colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring Man
(M) to the amino acid residues Q98, D160, D161, V226, and S227, forming the CBS of flocculin Flo5
from Saccharomyces cerevisiae (PDB code 2XJP). Aromatic residues Y54 and W228 involved in stacking
interactions with the pyranose ring of Man are colored orange. Figure 14. (A). Network of hydrogen bonds (black dashed lines) anchoring Mannobiose (M2) to
the amino acid residues forming the CBS of flocculin Flo5 from Saccharomyces cerevisiae (PDB code
2XJU). Aromatic residue Y54 and FW228 participating in stacking interactions with the pyranose
ring of the first Man residue are colored orange. (B). 2.2. Man-Specific Lectins from Lower Plants and Fungi Figure 16. Glycans recognized by ABA in glycan array experiments, arranged from 1 to 10 in decreasing o ans recognized by ABA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. ycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Glycan array analyses performed for the lectin MOA from the Marasmius oreades
mushroom, yielded the best results with the following glycans arranged in decreasing
order of affinity (Figure 17): Figure 17. Glycans recognized by MOA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. ns recognized by MOA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 17. Glycans recognized by MOA in glycan array experiments, arranged from 1 to 10 in decreasing MOA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Mannose-specific lectins from Ascomycota, such as the TbCVN and NcCVN cyanovirin-
related lectins from Tuber borchii (TbCVN) and Neurospora crassa (NcCVN), usually differ
from lectins of the Basidiomycota by a more pronounced affinity to high-mannose glycans
and a lesser affinity for complex type glycans, such as ABA from Agaricus bisporus and
MOA from Marasmius oreades. 2.2. Man-Specific Lectins from Lower Plants and Fungi Network of hydrogen bonds (black dashed
lines) anchoring Mannopentaose (M5) to the amino acid residues forming the CBS of flocculin Flo5
from Saccharomyces cerevisiae (PDB code 2XJT). Aromatic residue Y54 and W228 involved in stacking
interactions with the pyranose ring of the first Man of the oligomannoside are colored orange. Figure 14. (A). Network of hydrogen bonds (black dashed lines) anchoring Mannobiose (M2) to
the amino acid residues forming the CBS of flocculin Flo5 from Saccharomyces cerevisiae (PDB code
2XJU). Aromatic residue Y54 and FW228 participating in stacking interactions with the pyranose
ring of the first Man residue are colored orange. (B). Network of hydrogen bonds (black dashed
lines) anchoring Mannopentaose (M5) to the amino acid residues forming the CBS of flocculin Flo5
from Saccharomyces cerevisiae (PDB code 2XJT). Aromatic residue Y54 and W228 involved in stacking
interactions with the pyranose ring of the first Man of the oligomannoside are colored orange. Figure 14. (A). Network of hydrogen bonds (black dashed lines) anchoring Mannobiose (M2) to
the amino acid residues forming the CBS of flocculin Flo5 from Saccharomyces cerevisiae (PDB code
2XJU). Aromatic residue Y54 and FW228 participating in stacking interactions with the pyranose
ring of the first Man residue are colored orange. (B). Network of hydrogen bonds (black dashed
lines) anchoring Mannopentaose (M5) to the amino acid residues forming the CBS of flocculin Flo5
from Saccharomyces cerevisiae (PDB code 2XJT). Aromatic residue Y54 and W228 involved in stacking
interactions with the pyranose ring of the first Man of the oligomannoside are colored orange. In the glycan array analyses performed at Consortium for Functional Glycomics for
TbCVN, the Tuber borchii Man-specific lectin yielded the best results with the following
glycans arranged in decreasing order of affinity (Figure 15): Figure 15. Glycans recognized by TbCVN in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 15. Glycans recognized by TbCVN in glycan array experiments, arranged from 1 to 10 in decreasing lycans recognized by TbCVN in glycan array experiments, arranged from 1 to 10 in decreasing order of affinit 12 of 36 12 of 36 Cells 2021, 10, 1619 Glycan array studies performed for ABA, the Agaricus bisporus lectin, yielded the best
results with the following glycans arranged in decreasing order of affinity (Figure 16): Figure 16. Glycans recognized by ABA in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. 2.3. Man-Specific Lectins from Algae and Bacteria Seaweeds belonging to the different classes of algae, including red algae (Rhodophyta),
brown algae (Phaeophyta), green algae (Chlorophyta), and yellow-green algae (Chrys-
ophyta), are well-known sources of Man-specific lectins, together with the lectins from
Actinobacteria and Cyanobacteria, formerly classified as blue algae. Seaweed Man-specific lectins exhibit well-identified structural scaffolds but some of
them also exhibit unrelated structural scaffolds that still remain to be characterized. -
The β-prism I or β-barrel scaffold occurs predominantly in red algae (Rhodophyta)
but is less common in other classes of algae (Figure 18A–M). Algae of the genera Agard-
hiella, Eucheuma, Kappaphycus, Meristiella, Meristotheca, and Solieria exhibit this type of
molecular organization, in which the CBS is located at the top of the barrel structure. -
The β-prism I or β-barrel scaffold occurs predominantly in red algae (Rhodophyta)
but is less common in other classes of algae (Figure 18A–M). Algae of the genera Agard-
hiella, Eucheuma, Kappaphycus, Meristiella, Meristotheca, and Solieria exhibit this type of
molecular organization, in which the CBS is located at the top of the barrel structure. Cells 2021, 10, 1619 13 of 36 Figure 18. Overview of lectins from representative genera/species of algae and cyanob
with their associated structural scaffolds, able to recognize the spike proteins from corona
(Coronavirus Credit: Maria Voigt/RCSB PDB). A-M correspond to lectins used as structural sc
((A): Griffithsin, (B): ASL-1, (C): BU14, (D): HfL, (E): BCA, (F): OtL, (G): BPL-2, (H): N
Actinohivin, (J): CVN, (K): MVN, (L): OAA, (M): Scytovirin. References [28–30,112–129] corr
to the lectins presented in the figure. Figure 18. Overview of lectins from representative genera/species of algae and cyanobacteria,
with their associated structural scaffolds, able to recognize the spike proteins from coronaviruses
(Coronavirus Credit: Maria Voigt/RCSB PDB). A-M correspond to lectins used as structural scaffolds
((A): Griffithsin, (B): ASL-1, (C): BU14, (D): HfL, (E): BCA, (F): OtL, (G): BPL-2, (H): NgL, (I):
Actinohivin, (J): CVN, (K): MVN, (L): OAA, (M): Scytovirin. References [28–30,112–129] correspond
to the lectins presented in the figure. -
A less common β−prism II organization occurs in red algae (Grateloupia chiangii) and
green algae (Boodlea coacta), in which CBSs are located in the grooves delineated by
the bundles of β-strands. The MFP2-like structural scaffold, related to the β-prism II
organization, occurs in the green alga Bryopsis plumosa. -
A rather uncommon β-sandwich organization was found in the red alga Hydropuntia
(Gracilaria) fisheri, and in the green alga Ostreococcus tauri. 2.3. Man-Specific Lectins from Algae and Bacteria -
It is interesting to note that the structural scaffolds of many other Man-specific lectins,
identified in red algae (Carpopeltis flabellata, Gracilaria bursa-pastoris) and green algae Cells 2021, 10, 1619 14 of 36 14 of 36 (Enteromopha prolifera, Halimeda renschii), still remain unknown and are not apparently
related to any of the known structural scaffolds reported in algae. Man-specific lectins from the actinobacteria (actinohivin) and cyanobacteria/ex-blue
algae group (cyanovirin CVN, microvirin MVN, scytovirin SVN, and the Oscillatoria agard-
hii agglutinin OAA), exhibit a β-prism II (β-trefoil) structure and a β-prism I (β-barrel)
structure, respectively (Figure 18). The three-dimensional structures for different bacterial lectins in complex with Man,
mannosides, and more complex oligomannoside chains are available at the PDB (Figure 19
and Figure 20). Figure 19. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino
acid residues forming the CBS of griffithsin (PDB code 2RDK). Aromatic residue Y28 and Y110
participating in stacking interactions with the pyranose ring of Man are colored orange. (B). Network
of hydrogen bonds (black dashed lines) anchoring 6α-mannobiose (M2) to the amino acid residues
of the CBS of griffithsin (PDB code 2HYQ). Aromatic residue Y28 and Y110 involved in stacking
interactions with the pyranose ring of the first Man residue are colored orange. (C). Network of
hydrogen bonds (black dashed lines) anchoring a high-mannose branched glycan (M8) to the amino
acid residues of the CBS of griffithsin (PDB code 3LL2). Aromatic residue Y28, Y68, and Y110,
participating in stacking interactions with the pyranose rings of Man residues, are colored orange. Figure 19. (A). Network of hydrogen bonds (black dashed lines) anchoring Man (M) to the amino
acid residues forming the CBS of griffithsin (PDB code 2RDK). Aromatic residue Y28 and Y110
participating in stacking interactions with the pyranose ring of Man are colored orange. (B). Network
of hydrogen bonds (black dashed lines) anchoring 6α-mannobiose (M2) to the amino acid residues
of the CBS of griffithsin (PDB code 2HYQ). Aromatic residue Y28 and Y110 involved in stacking
interactions with the pyranose ring of the first Man residue are colored orange. (C). Network of
hydrogen bonds (black dashed lines) anchoring a high-mannose branched glycan (M8) to the amino
acid residues of the CBS of griffithsin (PDB code 3LL2). Aromatic residue Y28, Y68, and Y110,
participating in stacking interactions with the pyranose rings of Man residues, are colored orange. Figure 20. (A). 2.3. Man-Specific Lectins from Algae and Bacteria Network of hydrogen bonds (black dashed lines) anchoring mannobiose (M2) to the
amino acid residues forming the CBS of cyanovirin CVN (PDB code 2PYS). Aromatic residue F54
participating in a very weak stacking interaction with the pyranose ring of Man is colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring mannotetraose (M4) to cyanovirin
(PDB code 3GXZ). Amino acid residues of A and B protomers forming the swapped CVN dimer,
participate in the interaction with the oligomannoside. Figure 20. (A). Network of hydrogen bonds (black dashed lines) anchoring mannobiose (M2) to the
amino acid residues forming the CBS of cyanovirin CVN (PDB code 2PYS). Aromatic residue F54
participating in a very weak stacking interaction with the pyranose ring of Man is colored orange. (B). Network of hydrogen bonds (black dashed lines) anchoring mannotetraose (M4) to cyanovirin
(PDB code 3GXZ). Amino acid residues of A and B protomers forming the swapped CVN dimer,
participate in the interaction with the oligomannoside. High-mannose glycan-binding activity measured by Hori et al. [130] for ESA-2, the
Eucheuma serra lectin, yielded the best results with the following glycans arranged in
decreasing order of affinity (Figure 21): 15 of 36 Cells 2021, 10, 1619 Figure 21. Glycans recognized by ESA-2 in glycan-binding activity measurements, arranged from 1 to 10 in decreasing
order of affinity. d by ESA-2 in glycan-binding activity measurements, arranged from 1 to 10 in decreasing Figure 21. Glycans recognized by ESA-2 in glycan-binding activity measurements, arranged from 1 to 10 in decreasing
d
f ffiit Figure 21. Glycans recognized by ESA-2 in glycan-binding activity measurements, arranged from 1 to 10 in decreasing
order of affinity. Glycan array analyses performed at Consortium for Functional Glycomics for cyanovirin-
N CVN-N, a typical member of the cyanobacterial Man-specific lectins, yielded the best
results with the following glycans arranged in decreasing order of affinity (Figure 22): Figure 22. Glycans recognized by CVN-N in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 22. Glycans recognized by CVN-N in glycan array experiments, arranged from 1 to 10 in decreasing ns recognized by CVN-N in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Glycan array studies performed for actinohivin, a typical member of the actinobac-
teria Man-specific lectins, yielded the best results with the following glycans arranged in
decreasing order of affinity (Figure 23): Figure 23. 2.3. Man-Specific Lectins from Algae and Bacteria Glycans recognized by actinohivin in glycan array experiments, arranged from 1 to 10 in decreasing order
of affinity. Figure 23. Glycans recognized by actinohivin in glycan array experiments, arranged from 1 to 10 in decreasing order
f ffii Figure 23. Glycans recognized by actinohivin in glycan array experiments, arranged from 1 to 10 in decreasing order
of affinity. Man-specific lectins from algae, actinobacteria, and cyanobacteria, almost exclusively
interact with high-mannose glycans and, to a very little extent, with complex N-glycans. Most of the high-mannose glycans recognized by algal and bacterial lectins readily partici-
pate in the glycan equipment of the S proteins forming the spikes that decorate the surface
of SARS-CoV, MERS-CoV, and SARS-CoV-2 virions. Cells 2021, 10, 1619 16 of 36 3. Diversity of Glycans Decorating the Spike-Forming S Glycoproteins of Coronaviruses 3. Diversity of Glycans Decorating the Spike-Forming S Glycoproteins of Coronaviruses The S proteins are organized in homotrimers that form the spikes protruding from
the virion lipid bilayer. The S-glycoproteins of SARS-CoV, MERS-CoV, and SARS-CoV-
2, consist of heavily N-glycosylated proteins consisting of more than 1000 amino-acids,
exhibiting a high MW close to 130–140 kDa (133,568 Da for SARS-CoV, 146,594 Da for
MERS-CoV, and 141,178 Da for SARS-CoV-2), and an acidic pI (5.55 for SARS-CoV, 5.41 for
MERS-CoV, and 6.24 for SARS-CoV-2). These proteins contain a high number of potential
N-glycosylation sites NXT/NXS, that are actually glycosylated (22 for SARS-CoV and
SARS-CoV-2, 23 for MERS-CoV) [11,12]. These N-glycosylation sites are arrayed along the
amino acid sequence of the S-glycoproteins, predominantly at both the N- (S1 chain) and
C-termini (S2 chain) of the polypeptide chain (Figure 24). Figure 24. Amino acid sequences of S proteins from SARS-CoV, MERS-CoV, and SARS-CoV-2
showing the N-glycosylation sites occupied (almost) exclusively by complex type N-glycans (red
high-mannose type N-glycans (green), hybrid type N-glycans (purple), and predominantly occupied
by high-mannose glycans (pale green) or complex glycans (magenta). Figure 24. Amino acid sequences of S proteins from SARS-CoV, MERS-CoV, and
showing the N-glycosylation sites occupied (almost) exclusively by complex type N-g
high-mannose type N-glycans (green), hybrid type N-glycans (purple), and predominan
by high-mannose glycans (pale green) or complex glycans (magenta). Figure 24. Amino acid sequences of S proteins from SARS-CoV, MERS-CoV, and SARS-CoV-2,
showing the N-glycosylation sites occupied (almost) exclusively by complex type N-glycans (red),
high-mannose type N-glycans (green), hybrid type N-glycans (purple), and predominantly occupied
by high-mannose glycans (pale green) or complex glycans (magenta). 17 of 36 Cells 2021, 10, 1619 17 of 36 A detailed study of N-glycans present on the S-glycoprotein of coronaviruses revealed
the extreme diversity of N-glycans covering the coronavirus envelope [11,12]: -
High-mannose type glycans consist essentially of tri-antennary glycans GlcNAc2-
Man5–9 (Figure 25). -
High-mannose type glycans consist essentially of tri-antennary glycans GlcNAc2-
Man5–9 (Figure 25). f
f g
-
Complex type glycans are represented by essentially fucosylated and often sialylated,
bi-, tri-, and tetra-antennary glycans (Figure 25). Figure 25. (1–5). High-mannose-type glycans: GlcNAc2Man5 (1); GlcNAc2Man6 (2), GlcNAc2Man7 (3), GlcNAc2Man8
(4) and GlcNAc2Man9 (5). (6–15). Complex-type glycans: bi-antennary glycans (6,7), fucosylated bi-antennary glycan (8),
sialylated bi-antennary glycan (9), sialylated and fucosylated bi-antennary glycan (10), sialylated tri-antennary glycan (11),
sialylated and fucosylated tr-antennary glycan (12), sialylated tetra-antennary glycans (13,14) and sialylated and fucosylated
tetra-antennary glycan (15). Figure 25. (1–5). High-mannose-type glycans: GlcNAc2Man5 (1); GlcNAc2Man6 (2), GlcNAc2Man7 (3) Figure 25. (1–5). High-mannose-type glycans: GlcNAc2Man5 (1); GlcNAc2Man6 (2), GlcNAc2Man7 (3), GlcNAc2Man8
(4) and GlcNAc2Man9 (5). (6–15). Complex-type glycans: bi-antennary glycans (6,7), fucosylated bi-antennary glycan (8),
sialylated bi-antennary glycan (9), sialylated and fucosylated bi-antennary glycan (10), sialylated tri-antennary glycan (11),
sialylated and fucosylated tr-antennary glycan (12), sialylated tetra-antennary glycans (13,14) and sialylated and fucosylated
tetra-antennary glycan (15). Figure 25. (1–5). High-mannose-type glycans: GlcNAc2Man5 (1); GlcNAc2Man6 (2), GlcNAc2Man7 (3), GlcNAc2Man8
(4) and GlcNAc2Man9 (5). (6–15). Complex-type glycans: bi-antennary glycans (6,7), fucosylated bi-antennary glycan (8),
sialylated bi-antennary glycan (9), sialylated and fucosylated bi-antennary glycan (10), sialylated tri-antennary glycan (11),
sialylated and fucosylated tr-antennary glycan (12), sialylated tetra-antennary glycans (13,14) and sialylated and fucosylated
tetra-antennary glycan (15). y
g y
(
)
-
Scarce potential O-glycosylation sites T/S, actually occupied by O-glycans, have been
identified essentially in the S-glycoprotein of SARS-CoV-2 (T323, S325, and T678) [12] Lectin Family
Plant Species
Lectin
Structural
Scaffold
EC50
(µg/mL)
GNA-related
Allium porum
APA
β-trefoil
0.45 ± 0.08
Epipactis helleborine
EHA
β-trefoil
1.8 ± 0.3
Listera ovata
LOA
β-trefoil
2.2 ± 1.3
Hyppeastrum hybrid
HHA
β-trefoil
3.2 ± 2.8
Cymbidium hybrid
CA
β-trefoil
4.9 ± 0.8
Narcissus pseudonarcissus
NPA
β-trefoil
5.7 ± 4.4
Galanthus nivalis
GNA
β-trefoil
6.2 ± 0.6
Allium ursinum
AUA
β-trefoil
18 ± 4
Tulipa hybrid
TL MI
β-trefoil
22 ± 6
Lycoris radiata
LRA
β-trefoil
48
Colocasia esculenta
CEA
β-trefoil
>60
Allium sativum
ASA
β-trefoil
>100
JRL-related
Morus nigra
Morniga-M
β-barrel
1.6 ± 0.5
Nictaba-related
Nicotiana tabacum
Nictaba
β-sandwich
1.7 ± 0.3
Legume lectins
Cladrastis lutea
Cladrastis
β-sandwich
7.4 ±0.2
Figure 26. Site-specific N-linked glycosylation of SARS-CoV, MERS-CoV, and SARS-CoV-2. Sites
containing (almost) exclusively complex glycans (colored red), high-mannose glycans (colored green),
and hybrid glycans (colored magenta), are indicated. Sites harboring a mixture of complex glycans,
high-mannose glycans, and a few hybrid glycans are colored pale green or pink, depending on the
predominant high-mannose glycans (pale green) or complex glycans (orange). Table 1. Overview of Man-specific lectins from higher plants investigated in [13], for their anti-SARS-
CoV activity, classified in decreasing order of activity (EC50 (µg/mL)). Adapted from [13]. Lectin Family
Plant Species
Lectin
Structural
Scaffold
EC50
(µg/mL)
GNA-related
Allium porum
APA
β-trefoil
0.45 ± 0.08
Epipactis helleborine
EHA
β-trefoil
1.8 ± 0.3
Listera ovata
LOA
β-trefoil
2.2 ± 1.3
Hyppeastrum hybrid
HHA
β-trefoil
3.2 ± 2.8
Cymbidium hybrid
CA
β-trefoil
4.9 ± 0.8
Narcissus pseudonarcissus
NPA
β-trefoil
5.7 ± 4.4
Galanthus nivalis
GNA
β-trefoil
6.2 ± 0.6
Allium ursinum
AUA
β-trefoil
18 ± 4
Tulipa hybrid
TL MI
β-trefoil
22 ± 6
Lycoris radiata
LRA
β-trefoil
48
Colocasia esculenta
CEA
β-trefoil
>60
Allium sativum
ASA
β-trefoil
>100
JRL-related
Morus nigra
Morniga-M
β-barrel
1.6 ± 0.5
Nictaba-related
Nicotiana tabacum
Nictaba
β-sandwich
1.7 ± 0.3
Legume lectins
Cladrastis lutea
Cladrastis
β-sandwich
7.4 ±0.2 Table 1. Overview of Man-specific lectins from higher plants investigated in [13], for their anti-SARS-
CoV activity, classified in decreasing order of activity (EC50 (µg/mL)). Adapted from [13]. Table 1. Overview of Man-specific lectins from higher plants investigated in [13], for their anti-SARS-
CoV activity, classified in decreasing order of activity (EC50 (µg/mL)). Adapted from [13]. Table 1. Overview of Man-specific lectins from higher plants investigated in [13], for their anti-SARS-
CoV activity, classified in decreasing order of activity (EC50 (µg/mL)). Adapted from [13]. -
Hybrid type glycans, often bisected, have been detected at different N-glycosylation
sites but with the exception of the last C-terminal N-glycosylation site occurring in
S-glycoprotein of MERS-CoV which is exclusively occupied by hybrid-type glycans. All other N-glycosylation sites are occupied by a mixture of high-mannose, complex,
and hybrid glycans (Table 1). -
Hybrid type glycans, often bisected, have been detected at different N-glycosylation
sites but with the exception of the last C-terminal N-glycosylation site occurring in
S-glycoprotein of MERS-CoV which is exclusively occupied by hybrid-type glycans. All other N-glycosylation sites are occupied by a mixture of high-mannose, complex,
and hybrid glycans (Table 1). -
Hybrid type glycans, often bisected, have been detected at different N-glycosylation
sites but with the exception of the last C-terminal N-glycosylation site occurring in p
g y
y
g
S-glycoprotein of MERS-CoV which is exclusively occupied by hybrid-type glycans
All other N-glycosylation sites are occupied by a mixture of high-mannose, complex
and hybrid glycans (Table 1). y
g y
(
)
-
Scarce potential O-glycosylation sites T/S, actually occupied by O-glycans, have been
identified essentially in the S-glycoprotein of SARS-CoV-2 (T323, S325, and T678) [12]. y
g y
-
Scarce potential O-glycosylation sites T/S, actually occupied by O-glycans, have been
identified essentially in the S-glycoprotein of SARS-CoV-2 (T323, S325, and T678) [12]. The diversity of N-glycosylation occurring at the different N-glycosylation sites of
both SARS-CoV, MERS-CoV, and SARS-CoV-2, has been summarized in Figure 26. This
table suggests that SARS-CoV and SARS-CoV-2 share a rather similar N-glycosylation
pattern, which readily differs from that observed in MERS-CoV S-glycoprotein. In addition to the different N-glycosylation patterns found at the glycosylation sites
arrayed along the S-glycoprotein trimers forming the coronavirus spikes, spikes from
the different coronavirus readily differ by a distinct distribution of the complex and high-
mannose glycans at their surface and, especially at the top and the lateral faces of the spikes. Cells 2021, 10, 1619 18 of 36 Table 1. Overview of Man-specific lectins from higher plants investigated in [13], for their anti-SARS-
CoV activity, classified in decreasing order of activity (EC50 (µg/mL)). Adapted from [13]. Lectin Family
Plant Species
Lectin
Structural
Scaffold
EC50
(µg/mL)
GNA-related
Allium porum
APA
β-trefoil
0.45 ± 0.08
Epipactis helleborine
EHA
β-trefoil
1.8 ± 0.3
Listera ovata
LOA
β-trefoil
2.2 ± 1.3
Hyppeastrum hybrid
HHA
β-trefoil
3.2 ± 2.8
Cymbidium hybrid
CA
β-trefoil
4.9 ± 0.8
Narcissus pseudonarcissus
NPA
β-trefoil
5.7 ± 4.4
Galanthus nivalis
GNA
β-trefoil
6.2 ± 0.6
Allium ursinum
AUA
β-trefoil
18 ± 4
Tulipa hybrid
TL MI
β-trefoil
22 ± 6
Lycoris radiata
LRA
β-trefoil
48
Colocasia esculenta
CEA
β-trefoil
>60
Allium sativum
ASA
β-trefoil
>100
JRL-related
Morus nigra
Morniga-M
β-barrel
1.6 ± 0.5
Nictaba-related
Nicotiana tabacum
Nictaba
β-sandwich
1.7 ± 0.3
Legume lectins
Cladrastis lutea
Cladrastis
β-sandwich
7.4 ±0.2
Figure 26. Site-specific N-linked glycosylation of SARS-CoV, MERS-CoV, and SARS-CoV-2. Sites
containing (almost) exclusively complex glycans (colored red), high-mannose glycans (colored green),
and hybrid glycans (colored magenta), are indicated. Sites harboring a mixture of complex glycans,
high-mannose glycans, and a few hybrid glycans are colored pale green or pink, depending on the
predominant high-mannose glycans (pale green) or complex glycans (orange). Figure 26. Site-specific N-linked glycosylation of SARS-CoV, MERS-CoV, and SARS-CoV-2. Sites
containing (almost) exclusively complex glycans (colored red), high-mannose glycans (colored green),
and hybrid glycans (colored magenta), are indicated. Sites harboring a mixture of complex glycans,
high-mannose glycans, and a few hybrid glycans are colored pale green or pink, depending on the
d
h
h
l
(
l
)
l
l
(
) Figure 26. Site-specific N-linked glycosylation of SARS-CoV, MERS-CoV, and SARS-CoV-2. Sites
containing (almost) exclusively complex glycans (colored red), high-mannose glycans (colored green),
and hybrid glycans (colored magenta), are indicated. Sites harboring a mixture of complex glycans,
high-mannose glycans, and a few hybrid glycans are colored pale green or pink, depending on the
predominant high-mannose glycans (pale green) or complex glycans (orange). 19 of 36 Cells 2021, 10, 1619 19 of 36 4. Structural Organization and Glycosylation Pattern of S Glycoproteins Forming the
Spikes of Coronaviruses The distribution of the high-mannose N-glycans and complex N-glycans at the surface
of the S-glycoprotein strikingly differs depending on the coronaviruses SARS-CoV, MERS-
CoV, and SARS-CoV-2 (Figure 27): -
high-mannose glycans are predominantly distributed in the upper part of the S-
glycoprotein from MERS-CoV, whereas complex glycans mainly occur along the lower
part of the S-glycoprotein. -
complex glycans predominate in the upper part and lower part of S-glycoprotein from
SARS-CoV, while the less abundant high-mannose glycans are equally distributed
along with the glycoprotein. g
g y
p
-
in all three coronaviruses, the RBDs only contain complex N-glycans. g
g y
p
-
in all three coronaviruses, the RBDs only contain complex N-glycans. Figure 27. (A–C). Lateral face of the ribbon diagram representing the heavily glycosylated S glyco-
protein of SARS-CoV (PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2
(PDB code 6VXX) (C). High-mannose glycans and complex glycans are colored yellow and cyan,
respectively. The RBD is indicated by a red dashed circle. (D–F). Lateral face of the molecular
surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV (E), and SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. Figure 27. (A–C). Lateral face of the ribbon diagram representing the heavily glycosylated S glyco-
protein of SARS-CoV (PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2
(PDB code 6VXX) (C). High-mannose glycans and complex glycans are colored yellow and cyan,
respectively. The RBD is indicated by a red dashed circle. (D–F). Lateral face of the molecular
surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV (E), and SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. According to the different distribution of high-mannose and complex N-glycans along
the S-glycoprotein of SARS-CoV, MERS-CoV, and SARS-CoV-2, both types of glycans
are differentially distributed at the surface of the spikes of SARS-CoV, MERS-CoV, and
SARS-CoV-2, as shown from pictures of the lateral faces (Figure 28A–F) and front faces
(Figure 29A–F) of the spikes: 20 of 36 Cells 2021, 10, 1619 Figure 28. (A–C). Lateral face of the ribbon diagram representing the homotrimeric spike of SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-
mannose glycans and complex glycans are colored yellow and cyan, respectively. (D–F). 4. Structural Organization and Glycosylation Pattern of S Glycoproteins Forming the
Spikes of Coronaviruses Lateral face
of the molecular surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV (E), and
SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. Figure 28. (A–C). Lateral face of the ribbon diagram representing the homotrimeric spike of SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-
mannose glycans and complex glycans are colored yellow and cyan, respectively. (D–F). Lateral face
of the molecular surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV (E), and
SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. Figure 29. (A–C). Top face of the ribbon diagram representing the homotrimeric spike of SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. (D–F). Top
face of the molecular surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV
(E), and SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and
cyan, respectively. Figure 29. (A–C). Top face of the ribbon diagram representing the homotrimeric spike of SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-mannose glycans and complex glycans are colored yellow and cyan, respectively. (D–F). Top
face of the molecular surface (colored grey) of the S-glycoprotein of SARS-CoV (D), MERS-CoV
(E), and SARS-CoV-2 (F). High-mannose glycans and complex glycans are colored yellow and
cyan, respectively. Discrepancies observed in the distribution and exposure of both types of N-glycans,
especially at the top of the spikes, between SARS-CoV, MERS-CoV, and SARS-CoV-2
(Figure 29), allow for predicting the quite different accessibility of the high-mannose and Cells 2021, 10, 1619 21 of 36 21 of 36 complex glycan shield of the different coronaviruses to the Man-specific lectins from plants,
fungi, algae, and bacteria. Interestingly, none of the N-glycosylation sites was impacted by the point mutations
identified on the SARS-CoV-2 variants B.1.351 and B.1.1.7 that display increased resistance
to antibody neutralization [131]. The only asparagine residue impacted by the N501Y
mutation in the U.K., South African, and Brazilian variants of SARS-CoV-2, is not part of
an N-glycosylation site (Figure 30). Figure 30. Localization of point mutation N501Y in the amino acid sequence of the S-glycoprotein from SARS-CoV-2. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses Depending on the results obtained in glycan-binding assays and glycan array experi-
ments, one can predict the potential interaction with high-mannose glycans and complex
glycans identified in the glycan shield covering the spikes from SARS-CoV, MERS-CoV,
and SARS-CoV-2 viruses (Figures 31–33): -
Man-specific single- and two-chain lectins from the Fabaceae, which primarily rec-
ognize the (α3,α6)mannoside core of the branched GlcNAc2Man3 oligosaccharides,
readily interact with the complex glycans and, to a lesser extent, with high-mannose
glycans of the spikes from SARS-CoV (Figure 31), MERS-CoV (Figure 32) and SARS-
CoV-2 (Figure 33). In this respect, two-chain lectins from the Viciae, e.g., pea lectin PsA
and lentil lectin LcA, differ from single-chain lectins from other families of Fabaceae,
e.g., Con A from Canavalia ensiformis or DGL from Dioclea grandiflora, by a much more
pronounced affinity for α6-fucosylated complex glycans [21]. In addition, due to the
particularly well-exposed distribution of complex glycans at the top of the SARS-CoV
and SARS-CoV-2 spikes, both types of coronaviruses are predicted to better interact
with lectins from the Fabaceae, compared to MERS-CoV spikes (Figure 29). -
Man-specific single- and two-chain lectins from the Fabaceae, which primarily rec-
ognize the (α3,α6)mannoside core of the branched GlcNAc2Man3 oligosaccharides,
readily interact with the complex glycans and, to a lesser extent, with high-mannose
glycans of the spikes from SARS-CoV (Figure 31), MERS-CoV (Figure 32) and SARS-
CoV-2 (Figure 33). In this respect, two-chain lectins from the Viciae, e.g., pea lectin PsA
and lentil lectin LcA, differ from single-chain lectins from other families of Fabaceae,
e.g., Con A from Canavalia ensiformis or DGL from Dioclea grandiflora, by a much more
pronounced affinity for α6-fucosylated complex glycans [21]. In addition, due to the
particularly well-exposed distribution of complex glycans at the top of the SARS-CoV
and SARS-CoV-2 spikes, both types of coronaviruses are predicted to better interact
with lectins from the Fabaceae, compared to MERS-CoV spikes (Figure 29). -
The Man-specific jacalin-related lectins (JRL), such as Morniga-M from Morus nigra,
which also recognizes N-glycans of the hybrid type in addition to complex glycans and
high-mannose glycans, could serve as glycan probes for coronaviruses, particularly
for binding to MERS-CoV, which exhibits hybrid glycans at the N1288 glycosylation
site (Figure 26). 4. Structural Organization and Glycosylation Pattern of S Glycoproteins Forming the
Spikes of Coronaviruses The actually glycosylated N-glycosylation sites distributed along the amino acid sequence are indicated by red rods
and numbered. Figure 30. Localization of point mutation N501Y in the amino acid sequence of the S-glycoprotein from SARS-CoV-2. The actually glycosylated N-glycosylation sites distributed along the amino acid sequence are indicated by red rods
and numbered. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses -
The GNA-related lectins, which specifically interact with branched Man5-Man9 oligo-
mannosides, would predominantly recognize high-mannose glycans (Figures 31–33)
and, especially, the well-exposed high-mannose glycans protruding at the top of the
MERS-CoV spikes (Figure 29). The less exposed high-mannose glycans located at
the bottom of the SARS-CoV and SARS-CoV-2 spikes (Figure 28), should also be
recognized, but to a lesser extent. 22 of 36 Cells 2021, 10, 1619 Figure 31. SARS-CoV glycans that can be recognized (X) by Man-specific lectins from plants, mush-
rooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed with
purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. Figure 31. SARS-CoV glycans that can be recognized (X) by Man-specific lectins from plants, mush-
rooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed with
purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. Figure 32. MERS-CoV glycans that can be recognized (X) by Man-specific lectins from plants,
mushrooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed
with purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. Figure 32. -
Nictaba lectin from Nicotiana tabacum interacts with the following glycans arranged in
decreasing order of affinity according to glycan array experiments performed at the
Consortium for Functional Glycomics, and should therefore recognize the complex
glycans and high-mannose glycans forming the glycan shield of the spikes from
coronaviruses (Figure 34): 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses MERS-CoV glycans that can be recognized (X) by Man-specific lectins from plants,
mushrooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed
with purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. 23 of 36 Cells 2021, 10, 1619 Figure 33. ARS-CoV-2 glycans that can be recognized (X) by Man-specific lectins from plan
mushrooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performe
with purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legum
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictab
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomyco
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectin
from cyanobacteria. Figure 33. ARS-CoV-2 glycans that can be recognized (X) by Man-specific lectins from plants,
mushrooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed
with purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. Figure 33. ARS-CoV-2 glycans that can be recognized (X) by Man-specific lectins from plants,
mushrooms, algae, and cyanobacteria, as predicted from the results of glycan arrays performed
with purified lectins: Leg4 (Con A): single-chain legume lectins; Leg2 (PsA, LcA): two-chain legume
lectins; JRL (Morniga-M): jacalin-related lectins; GNA (GNA, ASA): GNA-like lectins; Nic: Nictaba
from Nicotiana tabacum; Ric: ricin-B from Ricinus communis; Asc (NcCVN): lectins from Ascomycota;
Bas (ABA): lectins from Basidiomycota; Rh (griffithsin, KAA-2): lectins from Rhodophyta; Chl (BCA):
lectins from Chlorophyta; Act (actinohivin): lectins from actinomycetes; Cya (cyanovirin): lectins
from cyanobacteria. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses -
Nictaba lectin from Nicotiana tabacum interacts with the following glycans arranged in
decreasing order of affinity according to glycan array experiments performed at the
Consortium for Functional Glycomics, and should therefore recognize the complex
glycans and high-mannose glycans forming the glycan shield of the spikes from
coronaviruses (Figure 34): -
Nictaba lectin from Nicotiana tabacum interacts with the following glycans arranged in
decreasing order of affinity according to glycan array experiments performed at the
Consortium for Functional Glycomics, and should therefore recognize the complex
glycans and high-mannose glycans forming the glycan shield of the spikes from
coronaviruses (Figure 34): 24 of 36 Cells 2021, 10, 1619 Figure 34. Glycans recognized by Nictaba in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. ns recognized by Nictaba in glycan array experiments, arranged from 1 to 10 in decreasing order of affinity. Figure 34. Glycans recognized by Nictaba in glycan array experiments, arranged from 1 to 10 in decreasing -
The ricin-B lectin from the Iris hollandica RIP-II was also shown to interact with complex
(α6-fucosylated) glycans, high-mannose glycans, and hybrid glycans in glycan arrays
experiments. Accordingly, it should readily interact with the spikes covering all types
of coronaviruses (Figures 31–33). g
-
Man-specific lectins from mushrooms belonging to the Ascomycota, essentially recog-
nize high-mannose glycans (Nc-CVN from Neurospora crassa, and Tb-CVN from Tuber
borchii, Flo1p from Saccharomyces pasteurianus, and Flo5A from S. cerevisiae), whereas
other Man-specific lectins from Basidiomycota, e.g Abmb from Agaricus bisporus, and
MOA from Marasmius oreades, preferentially recognize complex glycans. Mushroom
lectins should recognize the high-mannose glycans and the complex glycans from
coronaviruses. -
Man-specific lectins from red algae and green algae, both recognize exclusively high-
mannose glycans containing Man5-Man9 mannose residues. They should be nicely
adapted for the recognition of the high-mannose glycan shield exposed at the top
of the MERS-CoV spikes (Figures 28 and 29). Obviously, they should also recognize
the high-mannose glycans associated with the SARS-CoV and SARS-CoV-2 spikes
(Figures 31–33). g
-
Man-specific lectins from algae, actinobacteria (actinohivin) and cyanobacteria (cyanovirin,
microvirin, scytovirin), which predominantly possess a β−barrel structure and display
an exclusive specificity for branched Man5-Man9 oligomannosides, should recognize
the high-mannose glycan shield covering the coronavirus spikes and, especially, the
MERS-CoV virus (Figures 28 and 29). 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses The above predictions have already been corroborated by coronavirus-binding experi-
ments using several lectins from plants, algae, and bacteria, confirming the potential of
Man-specific lectins as relevant probes for the proper targeting of coronavirus spikes. p
p
p
p
g
g
p
The pioneering work of van der Meer et al. (2007) demonstrated the anti-viral prop-
erties of Man-specific GNA-related lectins such as CA from Cymbidium sp., HHA from
Hippeastrum hybrid, and GNA from Galanthus nivalis against SARS-CoV, and also high-
lighted the efficacy of the bacterial cyanovirin CVN [132]. These results were further
extended to a broader list of 33 lectins, including Morniga-M from Morus nigra and other
Man-specific lectins exhibiting a different structural scaffold [13]. The most efficient lectins
against SARS-CoV predominantly consist of GNA-related Man-specific lectins (Table 1). In
addition, lectins with different specificities, e.g., Gal-specific lectins (Morniga-G from Morus
nigra), GalNAc-specific lectins (ML II from Viscum album), and GlcNAc-specific lectins
(UDA from Urtica dioica), also displayed a significant anti-SARS-CoV activity. In addition,
two targets for HHA, the Man-specific lectin from Amaryllis (Hippeastrum hybrid), were
identified at the beginning and at the end of the infectious virus cycle. These target proteins
most probably are involved in the viral attachment to the host cell surface and the virus
release from the infected cells, respectively. The GNA-related Man-specific lectins interact
with the glycan shield from the SARS-CoV spikes and thus interfere with the attachment of
the virions to and the release from the host cells, with the exception of the garlic (Allium
sativum) lectin ASA, which is devoid of any anti-viral activity against SARS-CoV. Obviously,
the specific interaction of GNA-related lectins with the SARS-CoV virions depends on the Cells 2021, 10, 1619 25 of 36 preferential recognition by lectins of high-mannose glycans well exposed at the top of the
SARS-CoV spikes (Figure 29). Recently, another Man-specific lectin FRIL from the legume Lablab (Dolichos) purpureus
seeds, was shown to interfere with the SARS-CoV-2 entry in host cells, and its antiviral
activity is mediated by specific recognition of complex glycans occurring on the surface
of the S-glycoprotein [133]. In glycan array experiments, FRIL exhibited the best binding
to complex-type N-glycans with α1,3 or α1,4 fucosylated sub-terminal GlcNAc whereas
a slightly weaker binding was obtained with non-terminally fucosylated complex- and
hybrid-type N-glycans. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses In addition, GRFT was Cells 2021, 10, 1619 26 of 36 identified as a potent inhibitor of the MERS-CoV infection, inhibiting the entry of virions
into the cells [138]. Other griffithsin-related Man-specific lectins from Ascomycota, e.g.,
Nc-GRFT from Neurospora crassa and Tb-GRFT from Tuber borchii, should also interact with
the glycan shield covering the SARS-CoV spikes. Actinohivin, microvirin, and scytovirin
should also specifically interact with SARS-CoV but, to date, no information is available
yet on their anti-SARS-CoV properties. identified as a potent inhibitor of the MERS-CoV infection, inhibiting the entry of virions
into the cells [138]. Other griffithsin-related Man-specific lectins from Ascomycota, e.g.,
Nc-GRFT from Neurospora crassa and Tb-GRFT from Tuber borchii, should also interact with
the glycan shield covering the SARS-CoV spikes. Actinohivin, microvirin, and scytovirin
should also specifically interact with SARS-CoV but, to date, no information is available
yet on their anti-SARS-CoV properties. Similar to the Man-specific lectins from plants and algae, the so-called mannose-
binding lectins MBL from animals and especially, the calcium-dependent serum C-lectins,
which play a key role as opsonins in innate immunity towards a number of pathogens,
including viruses, bacteria, and protozoa, were identified as selective inhibitors for the
infection of host cells by SARS-CoV [139]. In addition, a single N-glycosylation site N330 of
the SARS-CoV S-glycoprotein, predominantly occupied by N-glycans of the complex type
(Figure 26), was identified as a critical target for the interaction from directed mutagenesis
experiments. In contrast, other membrane-associated Man-specific lectins such as DC-
SIGN and DC-SIGNR were assigned as receptors promoting infection and dissemination
of SARS-CoV through the specific recognition of the glycan shield covering the virion
spikes [140]. Recently, a detailed study of the interactions of the N-linked glycans in the
RBD from the SARS-CoV-2 spike protein has been performed with human galectins, using
NMR spectroscopy [141]. Moreover, the inhibition of Galectin-3, another animal lectin, was
proposed as a possible treatment capable of impeding the SARS-CoV-2 attachment to the
host cells and suppressing the host inflammatory response [142]. The mechanism by which lectins block viral replication once they have recognized the
glycoproteins covering the enveloped viruses, still remains unclear. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses This situation is reminiscent of the preferential affinity of Man-
specific Viciae lectins (LcA, LoL-I, PsA) for α1,6-fucosylated N-glycans of the complex
type [21,134,135], with the difference that Viciae lectins bind α1,6-Fuc on reducing ends of
complex-type N-glycans, which are therefore suspected to be less accessible to the lectins. However, Vicieae lectins also interact with the non-fucosylated tri-mannosyl core of high
mannose and complex-type glycans, which appear well exposed on the surface of the
coronavirus spikes (Figure 35A–F). This is consistent with the predicted preferential affinity
of Man-specific legume lectins for α1,6-fucosylated N-glycans of the complex type which
predominantly occur at the top of the SARS-CoV-2 spikes (Figure 29). These results obtained
with a legume lectin, compared to those obtained with GNA-related lectins towards SARS-
CoV, highlight the importance of a differential distribution of the high-mannose glycans
and complex glycans on the surface of the spikes, which, in fact, define the type of glycans
that will be accessible to the Man-specific lectins from the different groups. Figure 35. (A–C). Lateral views of the molecular surfaces (colored grey) of spikes from SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C),
showing the exposure of the fucosylated and non-fucosylated tri-mannosyl cores (colored magenta)
of high mannose glycans and complex glycans (colored cyan). (D–F). Front views of the molecular
surfaces (colored grey) from SARS-CoV (D), MERS-CoV (E), and SARS-CoV-2 (F), showing the
exposure of the fucosylated or non-fucosylated tri-mannosyl cores (colored magenta) of high mannose
glycans and complex glycans (colored cyan). Figure 35. (A–C). Lateral views of the molecular surfaces (colored grey) of spikes from SARS-CoV
(PDB code 6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C),
showing the exposure of the fucosylated and non-fucosylated tri-mannosyl cores (colored magenta)
of high mannose glycans and complex glycans (colored cyan). (D–F). Front views of the molecular
surfaces (colored grey) from SARS-CoV (D), MERS-CoV (E), and SARS-CoV-2 (F), showing the
exposure of the fucosylated or non-fucosylated tri-mannosyl cores (colored magenta) of high mannose
glycans and complex glycans (colored cyan). Man-specific lectins from red algae, and especially griffithsin GRFT, were recognized
very early as specific carbohydrate-binding agents for SARS-CoV, due to its strict specificity
for high-mannose glycans containing Man5-Man9 units [136,137]. 6. Biomedical Perspectives The extensive glycosylation at the surface of the S protein offers a shielding effect to
SARS-CoV, MERS-CoV, and SARS-CoV-2. In addition to this protective role, a possible
stabilization function has been postulated for some of the N-glycans covering the SARS-
CoV-2 S protein, that could be exploited in the future for targeting the glycan coat by
therapeutic drugs [144]. However, the distribution of the glycan shield at the surface of the
spikes and especially, at the top of the spikes, does not consistently support a protective
function susceptible to reduce the vulnerability of the coronaviruses towards antibody
epitopes and/or exogenous proteases. In this respect, the exposed area of the RBDs at the
top of the spikes is substantially free of glycans and thus, remains available for specific
recognition by the opened spike conformation of the corresponding ACE2 (SARS-CoV and
SARS-CoV-2) and DPP4 (MERS-CoV) host cell receptors (Figure 36A–C). Figure 36. (A–C). Top face of the ribbon diagram of the homotrimeric spike of SARS-CoV (PDB code
6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-mannose
glycans and complex glycans are colored cyan. Molecular surfaces corresponding to the RBDs are
colored gold and the remaining molecular surfaces are colored grey. Figure 36. (A–C). Top face of the ribbon diagram of the homotrimeric spike of SARS-CoV (PDB code
6ACD) (A), MERS-CoV (PDB code 5W9H) (B), and SARS-CoV-2 (PDB code 6VXX) (C). High-mannose
glycans and complex glycans are colored cyan. Molecular surfaces corresponding to the RBDs are
colored gold and the remaining molecular surfaces are colored grey. Nevertheless, the steric hindrance created by the interaction of Man-specific lectins
such as the dimeric PsA (50 kDa), the tetrameric GNA (50 kDa), and Morniga-M (60 kDa)
or the octameric Heltuba (120 kDa), with the high-mannose and complex N-glycans located
in the vicinity of the RBD area, should interfere with the receptor recognition process and
could disturb or prevent the entry of coronaviruses in host cells. In this respect, the di-,
tetra-, hexa- (Parkia platicephala lectin), and octameric Man-specific lectins from higher
plants, which exhibit high molecular weights in the range between 50–100 kDa, could serve
as blocking agents that can prevent the coronavirus infection. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses g y
p
g
p
Previous experiments performed to elucidate the inhibitory mode of action of Man-
specific lectins (GNA from snowdrop Galanthus nivalis, and HHA from Hippeastrum hybrid)
and UDA, the GlcNAc-binding lectin from stinging nettle Urtica dioica, on feline infectious
peritonitis virus (FIPV) and mouse hepatitis virus (MHV), showed that lectins do not inhibit
the attachment of coronavirus to the host cells but affect coronavirus entry at a post-binding
stage. The lectins could interfere with some spike protein rearrangements required for
driving the fusion between the viral envelope and the host cell membrane [143]. g
p
Griffithsin, the red algal Man-specific lectin, was identified as a potent inhibitor of
MERS-CoV infectivity and replication in vitro. The lectin was shown to interact with the
MERS-CoV spike protein during the early attachment step to the virions, suggesting that
griffithsin could prevent the attachment of the virions to the DPP4 receptors of the host
cells. Accordingly, the lectin cannot block MERS-CoV infection when added to the virions
at a post-binding step [138]. Investigating the mechanism by which FRIL neutralizes influenza viruses H1N1,
H3N2, H5N1, and H7N9, under in vitro and in vivo conditions, Liu et al. (2020) showed
that (1) FRIL neutralizes only influenza viruses possessing complex-type N-glycans, (2) due
to its tetravalent character, FRIL binds to the virions and creates virion aggregates before
the endocytosis into the host cell, (3) FRIL halts the host cell infection process in the late
endosome/lysosome and thus prevents the nuclear import of the virions [133]. Accord-
ing to these results, the authors proposed a model in which the lectin causes the virion
aggregation outside the host cell and, once endocytosed, prevents their nuclear import
by entrapping the virion-lectin aggregates in the late endosome/lysosome stage. In this
process, the multivalency of FRIL appears as a key factor for bridging the influenza virions
and create virion aggregates outside the host cell, prior to their endocytosis. Similarly, FRIL
also binds to the complex-type glycans of SARS-CoV-2 virions and displays a lower affinity
towards high-mannose glycans. These results confirm that FRIL exhibits a better affinity to
the complex-type glycans with α1,3 or α1,4 fucosylated sub-terminal GlcNAc, compared
to α1,3 and α1,6 linked Man of the α1,6-fucosylated or non-fucosylated tri-mannosyl core
GlcNAc2Man3. The affinity of FRIL for complex-type glycans could explain why the lectin
neutralizes both influenza and SARS-CoV-2 viruses but has almost no effect on HIV. 5. Man-Specific Lectins from Higher Plants, Fungi, Algae and Cyanobacteria,
Specifically Interact with the Highly Glycosylated S Glycoprotein from Coronaviruses Cells 2021, 10, 1619 27 of 36 Figure 37. Cartoon showing the relative size of different Man-binding lectins (dimeric GNA-like lectin
(A), bivalent legume lectin β2α2 (B), and Jacalin-related lectin (C)), complexed to the homotrimeric
spike (front face) from SARS-CoV-2. 6. Biomedical Perspectives Other Man-specific lectins
from algae, fungi, and cyanobacteria seem to be less adapted to act as a blocking agent
owing to their smaller size, e.g., 25 kDa for the griffithsin swapped homodimer and 20 kDa
for the cyanovirin swapped homodimer, respectively (Figure 37). Figure 37. Cartoon showing the relative size of different Man-binding lectins (dimeric GNA-like lectin
(A), bivalent legume lectin β2α2 (B), and Jacalin-related lectin (C)), complexed to the homotrimeric
spike (front face) from SARS-CoV-2. Figure 37. Cartoon showing the relative size of different Man-binding lectins (dimeric GNA-like lectin
(A), bivalent legume lectin β2α2 (B), and Jacalin-related lectin (C)), complexed to the homotrimeric
spike (front face) from SARS-CoV-2. Cells 2021, 10, 1619 28 of 36 28 of 36 As blocking agents for coronaviruses SARS-CoV, MERS-CoV, and SARS-CoV-2, Man-
specific lectins from higher plants, algae, fungi, and cyanobacteria could be used to entrap
and immobilize the virions using air-conditioned filters impregnated with lectins to prevent
the coronavirus dissemination by contaminated people. However, the covalent fixation
of lectins to air-conditioned filters necessitates large amounts of purified lectins, which
may be problematic in view of low yields usually obtained during purification of Man-
specific lectins from higher plants. Recent improvements performed in the large-scale
purification of the algal lectins from Eucheuma [114], and griffithsin from tobacco leaves,
could provide the bulk production of Man-specific lectins required for the manufacturing of
lectin-impregnated filters [145]. However, the blocking activity of dried lectins immobilized
on air-conditioned filters seems unlikely because these proteins need water to retain both
their conformation and carbohydrate-binding activity. At most, one can imagine a filtration
device equipped with a wet filter containing immobilized lectins, which would remove the
air from the virions before entering the air-conditioned filter. Another potential biomedical application for Man-specific lectins could consist of their
use as glycan probes for the detection of coronaviruses in the environment, e.g., on various
domestic surfaces such as doorknobs, home furnishings, handrails, mirrors, computers, etc. Virions deposited on these surfaces could be detected under UV illumination, e.g., using
specific antibodies labeled with a fluorochrome. It is possible to imagine further therapeutic issues for Man-specific lectins with respect
to the coronavirus infection. Recently, a risk of contamination by SARS-CoV-2 has been
identified via spoiled blood or blood products transfused to healthy people [146]. 6. Biomedical Perspectives Although
the risk of a transfusion transmission is essentially theoretical [147], a plasmapheresis
through a column of immobilized Man-specific lectin would be sufficient to eliminate the
eventual blood contaminants. A similar filtration of the suspected blood samples through
a column containing an immobilized Man-specific lectin could be performed in blood
centers or blood banks and would be a control measure sufficient to purge the blood from
coronavirus particles and thus prevent possible contamination for the transfused people. In
this respect, up to 80% of MERS-CoV and 70% of MARV (Marburg virus), were successfully
eliminated by lectin affinity plasmapheresis using GNA as a blocking agent immobilized
in the extra-capillary space of a standard plasma filter [148]. From the scarce in vitro experiments conducted to study the effects of Man-specific
lectins from higher plants [13,132,133], and griffithsin [137,138], towards MERS-CoV and
SARS-CoV-2, the anti-viral activity of lectins appeared to prevent both attachment and
entry of the virus in the host cell but does not interfere with the coronavirus replication
process within the cell. Consequently, Man-specific lectins cannot be considered as repli-
cation blockers for coronaviruses. A very different situation occurs for other enveloped
viruses such as HIV-1 [24], influenza virus [37], and Herpes virus [25], whose replication is
apparently blocked in vitro, in the presence of Man-specific lectins. In addition, some of the biomedical applications presented here could be adapted to
veterinary medicine by addressing animal coronavirus concerns [149] as part of the “one
health” approach. 8. Discussion Glycans of the S-glycoproteins forming the spikes of SARS-CoV, MERS-CoV and SARS-
CoV-2, consist of high-mannose glycans and often sialylated N-glycans that predominantly
occupy their N-glycosylation sites [11,12]. However, depending on the coronaviruses, some
discrepancies occur between the distribution of the two types of glycans on the surface of
the virion, which introduces some diversity in the glycan shield covering the coronavirus
spikes [11]. Accordingly, Man-specific lectins from plants, algae, fungi, and bacteria, which
differ slightly due to their fine sugar-binding specificities, offer a vast panel of glycan probes
more or less adapted to the specific recognition of the different coronaviruses. In this respect,
GNA-related lectins together with Man-specific lectins from algae and cyanobacteria,
appear as glycan probes nicely adapted to the recognition of the high-mannose shield
which predominates at the top of the MERS-CoV spike. Otherwise, legume lectins with
a higher affinity for N-glycans possessing the trimannoside Manα1,3Manα1,6Man core,
seem better adapted to the recognition of the N-glycans distributed predominantly at the
top of the glycan shield from SARS-CoV and SARS-CoV-2. p
g y
Besides Man-specific lectins, other lectins displaying different carbohydrate-binding
specificities have previously been identified as glycan probes for MERS-CoV [13,161]. In this
respect, lectins exhibiting Man/Glc-specificity (CLA from Cladrastris lutea), Gal/GalNAc-
specificity (Morniga-G from Morus nigra), GlcNAc-specificity (Nictaba from Nicotiana
tabacum), chitin-specificity (UDA from Urtica dioica), display an antiviral activity often
higher or similar to those displayed by genuine Man-specific lectins, essentially the GNA-
related lectins from monocot plants [39]. The occurrence of (sialylated)complex glycans and
hybrid glycans arrayed on the surface of coronavirus spikes, readily accounts for the differ-
ences observed among the carbohydrate-binding specificities towards the coronaviruses. In-
hibition of the SARS-CoV replication was observed in a lethal SARS-CoV-infected BALB/c
mouse model after treatment with the (GlcNAc)n-binding stinging nettle (Urtica dioica)
lectin [161]. Nictaba from Nicotiana tabacum and UDA, two GlcNAc-binding lectins, also
displayed antiviral properties against several families of enveloped viruses including
influenza A/B, Dengue virus type 2, herpes simplex virus, and HIV, respectively [162]. Accordingly, a mixture of lectins with different carbohydrate-binding specificities should
provide a glycan probe tool well adapted to all of the coronavirus types and strains. Author Contributions: A.B., M.S. and H.B., provided and analyzed the bibliographic information
for lectins. S.L.P. and B.K. provided and analyzed the bibliographic information dealing with coron-
aviruses and D.P. analyzed the technical aspects of the bibliographic information. P.R. 7. Bioinformatics The X-ray coordinates available for lectins, lectins in complex with simple sugars,
oligosaccharides, and more complex glycans, and S-glycoproteins from SARS-CoV, MERS-
CoV, and SARS-CoV-2, were taken from the PDB. Homology modeling of other lectins was performed with the YASARA Structure
program [150] using various structurally related proteins as templates from the PDB [151],
depending on the overall structural scaffold to which they belong. PROCHECK [152],
ANOLEA [153], and the calculated QMEAN scores [154,155] were used to assess the
geometric and thermodynamic qualities of the resulting three-dimensional models. Docking of simple sugars and oligosaccharides was performed with YASARA and
SwissDock [156,157]. The molecular surface of the lectins and S-glycoproteins from SARS- Cells 2021, 10, 1619 29 of 36 29 of 36 CoV, MERS-CoV, and SARS-CoV-2, were calculated and displayed with Chimera [158]. Molecular cartoons were drawn with Chimera and YASARA. CoV, MERS-CoV, and SARS-CoV-2, were calculated and displayed with Chimera [158]. Molecular cartoons were drawn with Chimera and YASARA. Assuming that putative N-glycosylation sites Asn-X-Thr/Ser of SARS-CoV, MERS-
CoV, and SARS-CoV2, are actually glycosylated, a classic bi-antennary high-mannose
glycan chain with a tri-mannoside core (Man)2-(Man)3-(GlcNAc)2 was modeled using
the GlyProt server (http://www.glycosciences.de/modeling/glyprot/php/main.php, ac-
cessed on 1 March 2021) [159], and represented in CPK on the molecular surface of the
spike S-glycoproteins. According to the known differential distribution of high-mannose
N-glycans and complex N-glycans on the surface of the S-glycoproteins, another run of
modeling was performed using standard high-mannose glycans and complex glycans
as models. Cartoons for high-mannose glycans, complex N-glycans, and hybrid glycans, were
built and represented with the DrawGlycan SNFG package for Mac [160]. Classical colored
symbols were used for representing Fuc (red triangle), Gal (yellow circle), GalNAc (yel-
low square), GlcNAc (blue square), Man (green circle), and sialic acid/Neu5Ac (purple
losange), respectively. 8. Discussion performed the
in silico molecular modelling and docking experiments, and provided the molecular cartoons and
the docking pictures. E.J.M.V.D. and P.R. wrote the review. All authors have read and agreed to the
published version of the manuscript. 30 of 36 Cells 2021, 10, 1619 Funding: This research received no external funding. Funding: This research received no external funding. Institutional Review Board Statement: Not applicable. Conflicts of Interest: The authors declare no conflict of interest. Conflicts of Interest: The authors declare no conflict of interest. 1.
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Ways of Viewing Pictorial Plasticity
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Keywords y
stereopsis, three-dimensional vision, synopter, depth perception, pictorial space, plastic effect y
stereopsis, three-dimensional vision, synopter, depth perception, pictorial space, plastic effect Article i-Perception
March-April 2017, 1–10
! The Author(s) 2017
DOI: 10.1177/2041669517699220
journals.sagepub.com/home/ipe i-Perception
March-April 2017, 1–10
! The Author(s) 2017
DOI: 10.1177/2041669517699220
journals.sagepub.com/home/ipe Ways of Viewing Pictorial
Plasticity Ways of Viewing Pictorial
Plasticity Maarten W. A. Wijntjes
Perceptual Intelligence Lab, Industrial Design Engineering,
Delft University of Technology, The Netherlands Abstract The plastic effect is historically used to denote various forms of stereopsis. The vivid impression of
depth often associated with binocular stereopsis can also be achieved in other ways, for example,
using a synopter. Accounts of this go back over a hundred years. These ways of viewing all aim to
diminish sensorial evidence that the picture is physically flat. Although various viewing modes have
been proposed in the literature, their effects have never been compared. In the current study, we
compared three viewing modes: monocular blur, synoptic viewing, and free viewing (using a
placebo synopter). By designing a physical embodiment that was indistinguishable for the three
experimental conditions, we kept observers naı¨ve with respect to the differences between them;
197 observers participated in an experiment where the three viewing modes were compared by
performing a rating task. Results indicate that synoptic viewing causes the largest plastic effect. Monocular blur scores lower than synoptic viewing but is still rated significantly higher than the
baseline conditions. The results strengthen the idea that synoptic viewing is not due to a placebo
effect. Furthermore, monocular blur has been verified for the first time as a way of experiencing
the plastic effect, although the effect is smaller than synoptic viewing. We discuss the results with
respect to the theoretical basis for the plastic effect. We show that current theories are not
described with sufficient details to explain the differences we found. p
g
Maarten W. A. Wijntjes, Perceptual Intelligence Lab, Industrial Design Engineering, Delft University of Technology,
Landbergstraat 15, 2628 CE Delft, The Netherlands.
Email: m.w.a.wijntjes@tudelft.nl Creative Commons CC-BY: This article is distributed under the terms of the Creative Commons Attribution 3.0 License
(http://www.creativecommons.org/licenses/by/3.0/) which permits any use, reproduction and distribution of the work without
further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sage-
pub.com/en-us/nam/open-access-at-sage). Corresponding author:
Maarten W. A. Wijntjes, Perceptual Intelligence Lab, Industrial Design Engineering, Delft University of Technology,
Landbergstraat 15, 2628 CE Delft, The Netherlands.
Email: m.w.a.wijntjes@tudelft.nl Introduction The term stereopsis is often associated with the perception of depth based on binocular
disparities. However, it has recently been argued (Vishwanath & Hibbard, 2013) that the
subjective impression evoked by stereo pictures very much resembles the depth impression of
monocular aperture viewing. This paradoxical (Clapare` de, 1904; Koenderink, van Doorn, &
Kappers, 1994) stereoscopic impression had been predicted before (Ames, 1925; Ebbinghaus,
1902) but was never directly and qualitatively compared with stereoscopic picture viewing. Monocular aperture viewing is a single case of a collection of viewing modes that achieve this Creative Commons CC-BY: This article is distributed under the terms of the Creative Commons Attribution 3.0 License
(http://www.creativecommons.org/licenses/by/3.0/) which permits any use, reproduction and distribution of the work without
further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sage-
pub.com/en-us/nam/open-access-at-sage). 2 i-Perception effect, also known as the plastic effect (Higashiyama & Shimono, 2012; Koenderink, Wijntjes,
& van Doorn, 2013). These ways of viewing include the following: 1. Monocular viewing (even better when using an aperture). 2. Optically decrease interocular distance (Javals’ iconoscope). 3. Increase the viewing distance. 4. Looking through an artificial pupil. 5. Change accommodation to infinity (using a lens). 6. Change convergence to parallel. 7. Introduce blur in one eye. 8. Looking through a mirror. 9. Looking through a synopter. These ways of viewing have all been discussed earlier (Ames, 1925; Blundell, 2015;
Schlosberg, 1941), including a concise overview of our own (Wijntjes, Fu¨ zy, Verheij,
Deetman, & Pont, 2016). What all these viewing modes have in common is that they change or eliminate the
nonpictorial depth cues of accommodation, vergence, and binocular disparity. When
viewing a picture normally, these cues will signal that the stimulus is flat. Although
pictures are indeed physically flat, they often contain pictorial depth cues that create an
impression of three-dimensional space, also known as pictorial space. In these cases (that
occur continuously in our daily lives), the visual system is confronted with a cue conflict
situation: Nonpictorial cues signal a flat stimulus, while pictorial cues signal a deep stimulus. A common explanation of the plastic effect is that perceived depth is the result of (possibly
weighted) averaging of physical flatness and pictorial depth (Ames, 1925). Thus, removing
the physical flatness would result in a relatively increased impression of depth, which is the
plastic effect. Introduction Aside from these theoretical considerations, we informally found that
looking through a very tiny aperture is rather cumbersome. The reader can easily verify
this him or herself by punching holes in an opaque sheet of cardboard (we used a hole
puncher that ships with a DIY music box kit). The other neglected viewing mode appears more promising: monocular blurring. Here, one
eye receives a blurred image while the other eye receives a sharp image. This binocular
sharpness difference disturbs the stereo correspondence problem, that is, the accurate
matching of retinal image features from which disparities are computed. The obvious
apparent downside is that the observer may become aware of the binocular conflict and
experience alternating rivalry between the sharp and blurred image. However, it is well
known (Levelt, 1966) that in this specific case of a sharp and blurred image, the sharpest
image generally dominates awareness. Therefore, the observer should theoretically not be too
much bothered by an alternating percept. In the study presented here, we wanted to test this aforementioned viewing mode and
verify whether the plastic effect can be induced by monocular blur. Initially, we planned to
use the synopter as a baseline condition for the plastic effect. The synopter is an optical device
invented by von Rohr (1907) that gives both eyes a similar viewpoint and thus eliminate
disparity information. Furthermore, it forces the eyes to have parallel binocular vergence. Previously, we investigated the contribution of pictorial cues to the plastic effect by using a
variety of paintings using a newly designed synopter (Wijntjes et al., 2016). While the synopter appears to be a good candidate to compare new, untested viewing
modes with, we realized that the synopter itself has never been thoroughly tested against a
baseline condition of normal binocular picture viewing. In our previous study (Wijntjes et al.,
2016), we let observers compare synoptic viewing with free viewing. Although we found
coherent results among observers for a large variety of paintings, we did not address the
potential contribution of a placebo effect: Maybe merely viewing through a box and thereby
introducing an artificial frame was evoking the sensation of stereoscopic depth, and not the
optical design itself. Thus, for a fair comparison between the synopter and monocular blur we
should also involve normal binocular viewing. It is not straightforward to make a prediction which viewing mode will result in the largest
plastic effect. Introduction The reason is that monocular blur and synoptic viewing perturb nonpictorial
depth cues in different ways. Monocular blur theoretically alters the sensitivity of both
vergence and disparity signals, while synoptic viewing introduces a bias in the vergence
and disparity signals. Theories explaining the plastic effect (e.g., Ames [1925] or
Vishwanath [2014]) do not make a distinction between these two ways of altering the
nonpictorial signals. Therefore, our experiment will not settle the debate which theory best
describes the plastic effect, neither are we able to predict which viewing mode may results in
the strongest plastic effect. However, the current study will provide empirical insights that
may help theory formation concerning the plastic effect. Furthermore, the study will also
reveal whether it is more efficient to alter the bias (synopter) or sensitivity (monocular blur) to
achieve the optimal plastic effect. Introduction This explanation predicts quantitative changes in depth as has been found by
Koenderink et al., (1994) but could not be verified by Vishwanath & Hibbard, (2013). An
alternative explanation of the plastic effect is the ‘‘absolute depth scaling hypothesis’’
(Vishwanath, 2014) which does not predict a quantitative depth change. According to this
hypothesis, perturbing the reliability of depth cues specifying the flat picture surface results in
ascribing distance information deriving from absolute depth cues (like accommodation,
vergence, and vertical disparities) to pictorial depth cues allowing them to be scaled in a
manner similar to the scaling of binocular disparities. The similar scaling mechanism is
according to Vishwanath (2014) responsible for the phenomenological similarity. Not all of the viewing modes have been empirically investigated, and some of these
(although mentioned separately in the literature) are largely similar. For example, Javals’
iconoscope uses four mirrors to decrease the virtual interocular distance, while the synopter
completely nullifies the virtual interocular distance due to a half translucent mirror. The
viewing modes that have attracted the least amount of attention in the literature seem to
be the artificial pupil (4) and monocular blurring (7). The concept of the artificial pupil
originates from Ames (1925) who recommends an aperture size of 2 mm or larger. Similar
to photography, the sharpness of the retinal image depends less on the accommodation for
small aperture sizes than for large apertures. The limiting case is the pinhole camera where no
lens is needed. Using a small artificial pupil will thus decrease the reliability of the
accommodation cue. Ames (1925) notes that the eye seems to automatically change
accommodation (because it cannot find a sharp image) resulting in apparent changes of
object size. Vishwanath and Hibbard (2013) used a much larger aperture (12–15 mm)
keeping potential depth information from accommodation reliable. This raises the Wijntjes Wijntjes 3 question of whether reducing the reliability even more would substantially improve the
plastic effect. Aside from these theoretical considerations, we informally found that
looking through a very tiny aperture is rather cumbersome. The reader can easily verify
this him or herself by punching holes in an opaque sheet of cardboard (we used a hole
puncher that ships with a DIY music box kit). question of whether reducing the reliability even more would substantially improve the
plastic effect. Venue and Context The experiment took place during a three-day festival called Campingflight to Lowlands,
in 2015. About 48,000 people visited this festival, which is primarily based on pop music
performances but also various other kinds of cultural activities. Our experiment was part i-Perception 4 of the Lowlands Science program and was located in the ArtTube tent, a venue where visitors
could take part in various art-related activities and experiments. The experiment was
conducted during 3 consecutive days at daytime between 12.00 and 18.00. of the Lowlands Science program and was located in the ArtTube tent, a venue where visitors
could take part in various art-related activities and experiments. The experiment was
conducted during 3 consecutive days at daytime between 12.00 and 18.00. Participants A total of 193 visitors participated in our experiment; 97 were male, 93 were female, and 3 did
not specify their gender. Average age was 30 years with the youngest participant being 15 and
the oldest being 63. Participants were asked whether they had used something ‘‘that could
influence their judgments,’’ 54 responded affirmative. Specifying their usage, 46 responded
that they had used alcohol (13 consumed one unit, 2 consumed two units, 1 consumed three
units, and 30 did not specify the total). A total of six had used cannabis (four in combination
with alcohol). Binocular stereo vision was assessed through self-report: Five responded to
have ill binocular stereo vision. Procedure Observers were first explained that we were investigating various devices that had been
hypothesized to increase pictorial depth. We asked the observers to assess the three devices
by rating them between 1 (no effect) and 10 (huge depth effect). They were informed that the
absolute values of the ratings were not very important to us, and that we were primarily
interested in the relative differences between the values. To assess the effect of the viewing
devices, the observers were asked to compare perception with and without device by
alternately looking through the device, and looking past it. y
g
g
g p
Observers were instructed to look at approximately 2 m distance, although in practice this
distance could vary somewhat. The devices were presented on a table, and the observers
could freely alternate between them. The order of viewing was random in the sense that the
devices were randomly presented on the table; the observers chose their own order. Most
observers were either looking at the Watermill or the Milkmaid. Only at Day 2 did we start
logging this independent variable, which amounts to about 100 cases. The experiment was performed in agreement with the Declaration of Helsinki and
approved by the TU Delft Human Research Ethics Committee. Apparatus and Stimuli We used three types of viewing devices. From the outside, all three looked similar, but the
interior was very different. The first device (see Figure 1) was a synopter close to described by
Wijntjes et al. (2016). We used a wooden (plywood) embodiment with a black spray painted
interior. The second device was similar to the synopter but we removed the full mirror in the
left eye. Therefore, the right eye was looking through a half mirror while the left eye had no
obstruction. With this design, stereovision is not impaired and should work normally: It is a
placebo synopter. We kept the half mirror on the right eye to introduce luminance differences
between the two eyes, which also occurs in our normal synopter due to the low beam splitting
quality of the acrylate half mirror. The third device had a similar embodiment as the other
two devices but did not have any mirrors. Instead, we glued a reading glass lens of þ3 diopter Synopter
Placebo
Blur
Figure 1. The three devices used in the experiment. Top row depicts them from the point of view of the
stimulus (the observers’ face should be behind the device). Bottom row shows the qualitative optics involved. Figure 1. The three devices used in the experiment. Top row depicts them from the point of view of the
stimulus (the observers’ face should be behind the device). Bottom row shows the qualitative optics involved. Wijntjes Wijntjes 5 behind the right eye aperture. This lens introduces blur when viewing at the distance we used
in our experiment (approximately 2 m). behind the right eye aperture. This lens introduces blur when viewing at the distance we used
in our experiment (approximately 2 m). As stimuli, we used matte prints of two paintings: The Watermill (c. 1664) by Meinder
Hobbema and The Milkmaid (c. 1660) by Johannes Vermeer, both paintings are in the
collection of the Rijksmuseum in Amsterdam. They were printed on A2 sized paper, resulting
in a size of 59.4 cm wide and 42 cm high for the Watermill, and 42 cm wide and 47.3 cm high for
the Milkmaid. The prints were attached to poster boards at approximately eye height (about
170 cm). Pictures documenting the experimental environment are shown in Figure 2. Data Analysis Responses were analyzed using a repeated measures analysis of variance with age as
covariate, and gender, stimulus, and reported drug use as between-visitor factors. Results The average ratings per viewing device are displayed in Figure 3. As can be seen, the real
synopter was rated highest, the placebo version the lowest and the blur device was rated in Figure 2. The experimental scene. On the left, an experimenter explains the experiment with the stimuli
visible in the background. Middle and right pictures show observer holding the devices to look at either or
the stimuli. Figure 2. The experimental scene. On the left, an experimenter explains the experiment with the stimuli
visible in the background. Middle and right pictures show observer holding the devices to look at either or
the stimuli. i-Perception 6 Real
Placebo
Blur
0
1
2
3
4
5
6
7
Grade
N=193
Figure 3. Average ratings and standard errors for the three viewing conditions. Figure 3. Average ratings and standard errors for the three viewing conditions. between. A statistical main effect was confirmed by the (Greenhouse-Geisser corrected)
analysis of variance: F(1.921, 312)¼11.831, p < .05. Furthermore, Bonferroni-corrected
pairwise comparisons revealed that all conditions were significantly different from each
other (p < .05). Neither the covariates age, nor the between-subject factors gender, stimulus
and reported drug use had a significant influence on the ratings. Because we had instructed the participants to primarily focus on the relative differences
between their ratings, and not so much on the absolute values, we also (descriptively)
analyzed the subjective ordering. In Figure 4, we plotted a histogram of the various
ordering combinations. The numbers on the y axis denote the order: 2 is highest, 1
intermediate, and 0 lowest. As can be seen, the synopter scored highest in almost 70% of
the cases (the sum of the lowest three bars). Furthermore, the configuration with the synopter
highest, placebo lowest, and monocular blur intermediate (lowest bar, 42%) is by far
occurring most often: It is three times more than the follow-up configuration (synopter
highest, no difference between placebo and monocular blur: 14%). 7 Wijntjes 42.%
14.%
13.5%
6.7%
6.2%
4.7%
3.6%
2.1%
2.1%
2.1%
1.6%
1.6%
0
10
20
30
40
0 2 1
0 2 2
0 0 0
0 0 2
2 0 2
0 2 0
1 2 0
1 0 2
0 1 2
2 1 0
2 0 0
2 0 1
Synopter
Placebo
Blur
Figure 4. Histogram of the various orderings that were present in the results. Results Highest is denoted by 2,
middle by 1, and lowest by 0. If a number is presented twice, these viewing modes were graded similarly. As
can be seen, synopter highest (2), monocular blur middle (1), and placebo lowest (0) dominate the data. 42.%
14.%
13.5%
6.7%
6.2%
4.7%
3.6%
2.1%
2.1%
2.1%
1.6%
1.6%
0
10
20
30
40
0 2 1
0 2 2
0 0 0
0 0 2
2 0 2
0 2 0
1 2 0
1 0 2
0 1 2
2 1 0
2 0 0
2 0 1
Synopter
Placebo
Blur Figure 4. Histogram of the various orderings that were present in the results. Highest is denoted by 2,
middle by 1, and lowest by 0. If a number is presented twice, these viewing modes were graded similarly. As
can be seen, synopter highest (2), monocular blur middle (1), and placebo lowest (0) dominate the data. Discussion Sometimes, observers responded that
they saw two images when looking through the synopter. This occurs when observers set their
vergence at the physical viewing distance. Possibly, accommodation or familiar size (the
images had been seen when entering the space and may have a typical size of an average
painting or poster) causes this vergence setting to occur. We heard from at least one
participant that he rated the placebo higher because it did not give him double images. As
can be inferred from Figure 3, there are four types of orderings in which the placebo was
higher rated than the synopter. These four amount to 11.4% of the observers. Although this
is not a large amount, it is important to consider that these cases may mitigate an actually
larger difference between the synopter and placebo condition. Although accounts referring to the plastic effect have existed in the literature for over a
hundred years, it has hardly been studied. The most salient experimental findings are changes
in pictorial relief using either a synopter (Koenderink et al., 1994) or a zograscope or
graphoscope (Koenderink et al., 2013) and the experiential similarity between binocular
stereo
and
monocular
aperture
viewing
(Vishwanath
&
Hibbard,
2013). The
aforementioned viewing devices are illustrated in Figure 5. Besides the empirical findings,
there is relatively little theoretical work. As outlined in the introduction, the two main
theories explaining the plastic effect (cue combination and absolute depth scaling) do not
seem to have been described in sufficient detail to quantitatively predict effect differences. One
difficulty is the distinction between sensitivity and bias of the physiologically altered cues. For
monocular blur, the sensitivities of the vergence and disparity cues are decreased. For
synoptic viewing, the vergence and disparity cues are biased. In the latter case, the biasing
of the cues toward infinity may result in a decrease of their reliability. Therefore, in both
viewing modes, the reliability of physiological depth cues is decreased but in very different
ways. This made it difficult to compare quantitatively in this study. Nevertheless, it is not
impossible to design future experiments that address this by systematically investigate
nonpictorial cues. For example, sensitivity studies can be conducted on the various ways
of viewing to quantify the size of variances introduced by the viewing modes. Also, it is Zograscope
Graphoscope
Hills’ Graphoscope
‘Graphoscopic’ optics
Figure 5. Illustration of the zograscope and graphoscope, including Hills’ variation. Discussion We found that both monocular blur and synoptic viewing induced the plastic effect in
comparison with the baseline condition of normal binocular viewing. Furthermore, the
synopter evoked a larger plastic effect than monocular blur. Therefore, we can conclude
that previous findings (e.g., Wijntjes et al., 2016) cannot be attributed to a placebo effect
and that the hitherto neglected viewing mode of monocular blur indeed enhances the
impression of pictorial depth as predicted (Ames, 1925; Schlosberg, 1941). Based on our (informal) experience with synopter demonstration events, the large
difference between the synopter and the placebo hardly comes as a surprise. Often,
observers express surprise when looking at a painting through a synopter. However, it had
never been experimentally verified whether these expressions could be attributed to some kind
of placebo effect. Since our placebo version has a similar embodiment as the synopter, we can
also safely argue that the synoptic effect cannot be attributed to looking through a box that
masks much of the periphery. It may certainly help with concentrating on an artwork while
wearing blinkers (used to focus the horse gaze), but it is not solely responsible for the plastic
effect. We also found that monocular blur increases the plastic effect, in comparison with the
baseline condition of the placebo. Perturbing binocular vergence and disparities through
blurring one of the images increased the impression of pictorial depth. This finding implies
the design of a very simple plastic effect device: A pair of reading glasses and with one of the
lenses removed. However, we found that the effect of monocular blur is substantially weaker
than the synopter. It may also be noteworthy that we never observed expressions of
spontaneous surprise associated with synoptic viewing. This may be reflected by the
absolute rating values. Although observers were instructed to primarily focus on the 8 i-Perception relative grades, it is likely that observers were inclined to use an absolute scale. Since the scale
used on the experiment was between 1 and 10, it is probable that observers referred to the
examination grades used in the Netherlands, that fall between 1 and 10. In this system, 5 and
lower is insufficient, 6 and higher is sufficient. Taking this into account, monocular blur
scores insufficient, while the synopter passes the exam. ffi
y
p
p
During the experiment, we picked up introspective comments indicating unforeseen
negative side effects of the synoptic parallel vergence. Discussion The principle relies at
least partly on the prismatic nature of the lens edges (Michaud, 1908) illustrated at the right. Figure 5. Illustration of the zograscope and graphoscope, including Hills’ variation. The principle relies at
least partly on the prismatic nature of the lens edges (Michaud, 1908) illustrated at the right. Wijntjes Wijntjes 9 possible to alter the vergence of the synopter toward the viewing distance without affecting
the disparities, which could reveal the contribution of binocular vergence. Another
interesting device that may help answering these questions is Hill’s (1898) graphoscope
which forces vergence to be parallel using prisms, without affecting disparities as shown in
Figure 5. possible to alter the vergence of the synopter toward the viewing distance without affecting
the disparities, which could reveal the contribution of binocular vergence. Another
interesting device that may help answering these questions is Hill’s (1898) graphoscope
which forces vergence to be parallel using prisms, without affecting disparities as shown in
Figure 5. Besides the fundamentally interesting underlying mechanisms of the plastic effect, studies
on various ways of viewing the plastic effect also have practical relevance. The consumer
interest in three-dimensional displays and virtual reality indicates that people are interested in
seeing pictures differently. However, these novel techniques are only applicable when a stereo
image pair is available. There are numerous examples of occasions where this is impossible. Our personal favorite application is looking at paintings in a museum, but also more
mundane activities such as viewing personal photos on a screen may become more
interesting when the plastic effect is present. Thus, studying and applying old techniques
and inventions concerning the plastic effect may be a valuable addition next to the high-
tech advances of virtual reality, as has recently been argued by Blundell (2015). The current
findings in combination with our previous study (Wijntjes et al., 2016) indicates that the
synopter version we used in these studies is a good candidate for viewing pictures at distances
of about 1.5 to 2 m. For presentations on small displays that are close to the eyes, other
devices may be useful, such as the Verant lens (also by von Rohr), or Hill’s graphoscope. And
possibly there is some space left for novel devices yet to be invented. Acknowledgements The author would like to thank Steffen de Jonge, Willemijn Elkhuizen, Jur Vleugel, Mitra Smit, Argun
Cencen, and Michiel Baay for their help in setting up and conducting the experiments and also thank
Noortje Jacobs and Rewan Jansen from BKB, who organized Lowlands Science and gave us the
opportunity to conduct this large-scale study on the plastic effect. Funding The author(s) received no financial support for the research, authorship, and/or publication of this
article. Declaration of Conflicting Interests The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or
publication of this article. Higashiyama, A., & Shimono, K. (2012). Apparent depth of pictures reflected by a mirror: The plastic
effect. Attention, Perception, & Psychophysics, 74, 1522–1532. Retrieved from http://doi.org/10.3758/
s13414-012-0346-4 Ebbinghaus, H. (1902). Grundzu¨ge der psychologie (Vol. 9). Leipzig, Germany: Verlag von Veit &
Comp. Retrieved from http://doi.org/10.1037/h0057450 References Ames, A. (1925). The illusion of depth from single pictures. Journal of the Optical Society of America,
10, 137–148. , B. G. (2015). On alternative approaches to 3D image perception: Monoscopic 3D technique
Research, 6, 1–17. ,
,
Clapare` de, E. (1904). Ste´ re´ oscopie monoculaire paradoxale. Annales d’Oculistique, 132, 465–466. Ebbinghaus, H. (1902). Grundzu¨ge der psychologie (Vol. 9). Leipzig, Germany: Verlag von Veit &
Comp. Retrieved from http://doi.org/10.1037/h0057450 Ebbinghaus, H. (1902). Grundzu¨ge der psychologie (Vol. 9). Leipzig, Germany: Verlag von Veit &
Comp. Retrieved from http://doi.org/10.1037/h0057450 Higashiyama, A., & Shimono, K. (2012). Apparent depth of pictures reflected by a mirror: The plastic
effect. Attention, Perception, & Psychophysics, 74, 1522–1532. Retrieved from http://doi.org/10.3758/
s13414-012-0346-4 Higashiyama, A., & Shimono, K. (2012). Apparent depth of pictures reflected by a mirror: The plastic
effect. Attention, Perception, & Psychophysics, 74, 1522–1532. Retrieved from http://doi.org/10.3758/
s13414-012-0346-4 10 i-Perception Hill, H. H. (1898). Improved graphoscope for obtaining stereoscopic effects, UK Patent 189815299–A. Michaud, G. (1908). A stereoscope for single photographs. Scientific American, 99, 320–320. Koenderink, J. J., van Doorn, A. J., & Kappers, A. M. L. (1994). On so-called paradoxical m
stereoscopy. Perception, 23, 583–594. Koenderink, J. J., Wijntjes, M. W. A., & van Doorn, A. J. (2013). Zograscopic viewing. I-Perception, 4,
192–206. Levelt, W. J. M. (1966). The alternation process in binocular rivalry. British Journal of Psychology, 57,
225–238. Schlosberg, H. (1941). Stereoscopic depth from single pictures. The American Journal of Psychology, 54,
601–605. Vishwanath, D. (2014). Toward a new theory of stereopsis. Psychological Review, 121, 151–178. Retrieved from http://doi.org/10.1037/a0035233 Vishwanath, D., & Hibbard, P. B. (2013). Seeing in 3-d with just one eye: Stereopsis without binocular
vision. Psychological
Science,
24,
1673–1685. Retrieved
from
http://doi.org/10.1177/
0956797613477867 von Rohr, M. (1907). Reflecting intruments annulling the perception of depth in binocular vision, UK
Patent 190700916-A. Wijntjes, M. W. A., Fu¨ zy, A., Verheij, M. E. S., Deetman, T., & Pont, S. C. (2016). The synoptic art
experience. Journal of Art and Perception, 4, 73–105. Author Biography Author Biography
Maarten W. A. Wijntjes received a PhD degree from Utrecht
University for work on haptic perception. He is currently an
assistant professor at the faculty of Industrial Design Engineering
(Delft University of Technology, Netherlands), where he teaches
Visual
Communication
Design. His
(research)
interests
are
pictorial space, material perception, visual design and (technical)
art history. Maarten W. A. References Wijntjes received a PhD degree from Utrecht
University for work on haptic perception. He is currently an
assistant professor at the faculty of Industrial Design Engineering
(Delft University of Technology, Netherlands), where he teaches
Visual
Communication
Design. His
(research)
interests
are
pictorial space, material perception, visual design and (technical)
art history. Maarten W. A. Wijntjes received a PhD degree from Utrecht
University for work on haptic perception. He is currently an
assistant professor at the faculty of Industrial Design Engineering
(Delft University of Technology, Netherlands), where he teaches
Visual
Communication
Design. His
(research)
interests
are
pictorial space, material perception, visual design and (technical)
art history.
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Realidade de serviços e da assistência em saúde após acometimento pela covid-19: uma revisão integrativa de literatura
|
Revista Interdisciplinar de Promoção da Saúde
| 2,022
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cc-by
| 7,516
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Palavras-chave:
Covid-19; Serviços de
reabilitação; Atenção à
saúde; Assistência Inte
gral à Saúde.. RESUMO Introdução: o surgimento da Covid-19 causada pelo Sars-COV-2, ao final do ano de 2019, produziu, nos
diversos âmbitos da gestão pública, dos serviços e na assistência em saúde, adaptações, melhorias e tam
bém algumas limitações ainda não bem dimensionadas. Objetivo: sistematizar dados da produção cientí
fica nacional e internacional acerca da realidade dos serviços e da assistência à saúde prestada à popula
ção, após o acometimento pela Covid-19. Método: estudo de revisão integrativa da literatura com busca
dos artigos publicados entre janeiro a setembro de 2021, na base de dados Portal de Periódicos da CAPES,
utilizando os Descritores em Ciências da Saúde (DeCS) “Covid-19”, “serviços de reabilitação”, “atenção
à saúde” e “assistência integral à saúde” e seus equivalentes em língua inglesa. Foram encontrados 234
artigos, sendo que artigos duplicados, os que não abordavam a temática, artigos de revisão e meta-análise
foram excluídos, resultando em três estudos selecionados. A análise dos dados foi realizada por meio da
Análise de Conteúdo, com a elaboração de três campos temáticos: (re) estruturação de fluxos e processos
de trabalho; saúde e segurança dos trabalhadores da saúde; modelos de assistência à saúde. Resultados:
os artigos selecionados abordavam questões relacionadas à necessidade de alterações e adaptações nos
serviços de saúde, acerca dos atendimentos assistenciais nos casos de Covid-19, como também medidas
de promoção da saúde, prevenção do risco de contágio da doença e proteção dos trabalhadores da saúde,
além de reformulações necessárias nos padrões de assistência à saúde dos pacientes. Considerações fi
nais: buscou-se dar visibilidade ao tema em questão, reforçando a necessidade de investigações científicas
a respeito, considerando a pequena quantidade de estudos encontrados no contexto de pós- Covid-19 nos
serviços e na assistência à saúde. Os resultados apontaram aspectos relacionados à necessidade de diver
sas modificações e adaptações dos serviços de saúde após o acometimento pela doença. Exceto onde especificado diferentemente, a matéria publicada neste periódico é licenciada
sob forma de uma liença Creative Commons - Atribuição 4.0 Internacional.
http://creativecommons.org/licenses/by/4.0/ ARTIGO DE REVISÃO
Realidade de serviços e da assistência em saúde após
acometimento pela covid-19: uma revisão integrativa
Reality of services and health care after acquiring covid-19:
an integrative review
Volume 4 - Número 2 - Abril/Junho 2021
https://doi.org/10.17058/rips.v4i2.17264
REVISTA INTERDISCIPLINAR DE PROMOÇÃO DA SAÚDE
INTERDISCIPLINARY JOURNAL OF HEALTH PROMOTION Suzane Beatriz Frantz Krug1, Henrique Ziembowicz1, Iagro Cesar de Almeida Bavaresco1, Laura
Schmidt Rizzi1, Irene Souza1, Morgana Pappen1, Guilherme Mocelin1 1 - Universidade de Santa Cruz do Sul – UNISC, Santa Cruz do Sul, RS, Brasil. Keywords:
COVID-19; Rehabilita
tion Services; Delivery of
Health Care; Comprehen
sive Health Care. SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 INTRODUÇÃO surgiram necessidades de reabilitação da população
quanto à saúde mental. Nesse contexto, percebeu-se a
exposição da sociedade a uma variedade de doenças
psicológicas, como distúrbios do sono, depressão,
ansiedade e esgotamento. Tornando-se essencial
promover apoio aos indivíduos que foram afetados
pelo isolamento social e pela incapacidade de acessar
cuidados de saúde para doenças pré-existentes ou
novas, não-Covid-19 durante a atual pandemia, ainda
em curso.5 Com o surgimento da Covid-19 causada
pelo Sars-COV-2 ao final do ano de 2019, produziu-
se, nos diferentes âmbitos da gestão pública, dos
serviços e na assistência em saúde, adaptações,
alterações, melhorias e alguns prejuízos ainda não
bem dimensionados. A saber, a doença apresenta
acometimento multissistêmico, onde os sintomas
mais comuns são inespecíficos, como febre, tosse,
fadiga e dispneia e, ainda, dentre as complicações
mais prevalentes estão a síndrome do desconforto
respiratório agudo, choque séptico, disfunção
miocárdica e lesão renal aguda.1 Envoltos nessa realidade da Covid-19,
que ainda persiste, é imperativo compreender as
exigências que o momento vivido trouxe, atentando
para a continuidade do cuidado quanto à infecção
e buscando maneiras de reorganizar os serviços
assistenciais à saúde. A sistematização de dados e a
produção científica auxiliam nesse processo, o que
pode beneficiar os sistemas de saúde, promover o bem-
estar da população e dos trabalhadores dessa área,
qualificar os atendimentos e proporcionar pesquisas de
qualidade. Desse modo, o estudo teve como objetivo:
sistematizar dados da produção científica nacional
e internacional acerca da realidade dos serviços e
da assistência à saúde prestada à população, após o
acometimento pela Covid-19. Nesse sentido, a literatura relata consequências
como pneumonia viral com infiltrados e hipóxia com
resolução prolongada, exigindo suporte de oxigênio
suplementar, além do estado inflamatório que pode
causar disfunção múltipla de órgãos. Diante dessa
situação, a reversão do quadro clínico frente à alta
transmissibilidade do vírus gerou acréscimo da
necessidade de leitos hospitalares e equipamentos de
terapia intensiva, de tal maneira que desafia a gestão
de serviços de saúde e a implementação de políticas
públicas para contemplar a demanda na reabilitação e
assistência de pacientes no período pós infecção.2 MÉTODO A pandemia da Covid-19 afetou diversos
aspectos da vida diária da população em todo o mundo
e, sendo uma crise de saúde, impactou severamente o
setor.3 Intensificada pelas estratégias que precisaram
ser tomadas, como o lockdown e o receio de parte da
população pelo grande número de casos da Covid-19,
a pandemia promoveu modificações no padrão de
oferta e procura em saúde. Evidenciou-se que não
apenas havia uma nova doença baseada em quadros
de síndrome respiratória aguda, como também
ocorreu uma alteração no seguimento dos pacientes
anteriormente diagnosticados por outras patologias e
uma descontinuidade do cuidado pelo medo da nova
infecção. Desse modo, o sistema de saúde como um
todo precisou adequar-se à situação vivida e passou
a utilizar algumas tecnologias e inovações em saúde
como a telemedicina, que se constitui como uma
ferramenta promovida e ampliada para reduzir o
risco de transmissão e ajudar a expandir o acesso aos
cuidados.4 O presente estudo trata-se de uma revisão
integrativa da literatura, caracterizada por sintetizar
resultados já alcançados em pesquisas sobre
determinado tema, de forma estruturada, sistemática
e ampla. Esse tipo de revisão baseia-se em seis etapas,
sendo elas: elaboração da pergunta norteadora, busca
ou amostragem na literatura, coleta de dados, análise
crítica dos estudos incluídos, discussão dos resultados
e apresentação da revisão integrativa.6,7 O presente estudo trata-se de uma revisão
integrativa da literatura, caracterizada por sintetizar
resultados já alcançados em pesquisas sobre
determinado tema, de forma estruturada, sistemática
e ampla. Esse tipo de revisão baseia-se em seis etapas,
sendo elas: elaboração da pergunta norteadora, busca
ou amostragem na literatura, coleta de dados, análise
crítica dos estudos incluídos, discussão dos resultados
e apresentação da revisão integrativa.6,7 A primeira etapa da revisão integrativa é a
definição da questão norteadora, a qual definiu-se:
como está constituída a produção científica nacional
e internacional acerca da realidade dos serviços e
da assistência à saúde prestada à população, após o
acometimento pela Covid-19? i A segunda etapa é a definição dos critérios
de inclusão e exclusão dos documentos a serem
analisados. Assim sendo, como critério de inclusão,
optou-se por artigos originais ou relatos de experiência,
disponíveis online, de acesso livre, em língua inglesa
ou portuguesa, com texto completo, publicados no
ano de 2021, revisados por pares. ABSTRACT Introduction: the onset of COVID-19, caused by the SARS-COV-2 virus at the end of 2019, produced,
in the various spheres of public management, services and health care, adaptations, improvements and
also some limitations that were not well-dimensioned at the moment. Objective: systematize data from
national and international scientific production about the reality of services and health care provided to
the population, after acquiring COVID-19. Method: integrative literature review study with search for
articles published between January and September 2021, in the CAPES Periodicals Portal database, using
the Health Sciences Descriptors (DeCS) “COVID-19”, “rehabilitation services”, “delivery of health care”
and “comprehensive health care” and their Portuguese equivalents. A total of 234 articles were found,
from which duplicated articles, those that did not address the topic, review articles and meta-analyses
were excluded, resulting in three selected studies. Data analysis was carried out through content analy
sis, with the elaboration of three thematic fields: (re)structuring of work flows and processes; health and
safety of health workers; healthcare models. Results: the selected articles addressed issues related to the
need for changes and adaptations in health services, about care assistance in cases of COVID-19, as well
as health promotion measures, prevention of the risk of contagion of the disease and protection of health
workers, in addition to reformulations in patient health care standards. Final considertions: visibility was
given to the issue in question, reinforcing the need for scientific investigations in this regard, considering
the low number of studies found in the post-COVID-19 context in health services and health care. The
results pointed to aspects related to the need for several changes and adaptations of health services after
the onset of the disease. Keywords:
COVID-19; Rehabilita
tion Services; Delivery of
Health Care; Comprehen
sive Health Care. ISSN: 2595-3664 MÉTODO Como critérios Ademais,
percebeu-se
que,
associada
à reabilitação dos pacientes da Covid-19 e à
importância da reorganização das consultas e do
acesso dos sistemas de saúde à população em geral, Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 76 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 de exclusão, considerou-se: artigos de meta-análise,
artigos de revisão, artigos publicados em outras
línguas e que não respondessem à questão norteadora. dos estudos incluídos, sendo que na busca com os
descritores na língua inglesa, foram encontrados
inicialmente 234 artigos, sendo que os artigos
duplicados, bem como, aqueles em outras línguas
foram excluídos, totalizando 114 artigos pré-
selecionados. A terceira etapa compreende a busca na base
de dados selecionada a partir dos critérios de inclusão
estabelecidos, dessa forma, utilizou-se os descritores
“Covid-19”, “serviços de reabilitação”, “atenção à
saúde” e “assistência integral à saúde”, bem como,
seus equivalentes em língua inglesa “Covid-19”,
“rehabilitation services”, “delivery of health care” e
“comprehensive health care”, retirados dos Descritores
em Ciências da Saúde (DeCS). Utilizou-se a base
de dados do Portal de Periódicos da CAPES para a
referida busca, sendo incluídos no estudo, artigos
publicados entre 01 de janeiro a 08 de setembro de
2021, que continham no título e/ou resumo e/ou no
corpo do texto e/ou nas palavras-chave os descritores
pesquisados. Após nova leitura criteriosa e detalhada
dos textos, foram excluídos 87 por não abordar a
temática, 18 por serem artigos de revisão e meta-
análise, e seis artigos pagos. Assim, três artigos foram
selecionados para a amostra final e foram exportados
para a plataforma Mendeley®, a fim de organizar e
realizar as demais etapas. Já com os descritores na
língua portuguesa, foi encontrado um artigo, porém,
o mesmo não apresentava revisão por pares e não
abordava a temática, sendo excluído do presente
estudo. Utilizou-se neste artigo a recomendação
“Principais Itens para Relatar Revisões sistemáticas e
Meta-análises” - PRISMA (Preferred Reporting Items
for Systematic reviews and Meta-Analyses) visando
a clareza em relação ao processo de coleta de dados. Essa recomendação segue, de forma sistematizada,
um fluxo ascendente de ideias, de modo a permitir,
didaticamente, colher e analisar informação por meio
de quatro fases: identificação, seleção, elegibilidade
e inclusão.8 Seguindo a recomendação PRISMA, um
fluxograma da etapa de coleta foi elaborado, para
sintetizar e apresentar o processo da busca dos artigos. RESULTADOS A busca dos artigos apontou a importante
predominância
de
artigos
internacionais
em
comparação a artigos nacionais, já que do total
de artigos encontrados, nenhum foi publicado em
periódico nacional. Outro aspecto a ser considerado
na busca dos estudos é o grande volume de artigos
duplicados, de artigos de revisão e de estudos que
não abordavam a temática específica, o que indica a
necessidade de mais investigações sobre a questão. Por
outro lado, apesar da pequena quantidade de artigos
selecionados para análise, é importante considerar
a existência de estudos sobre serviços e assistência
à saúde após acometimento pela Covid-19, já que a
pandemia ainda se encontra em curso. Como resultados do campo temático dos
modelos de assistência à saúde aponta-se: os
procedimentos eletivos passaram a ser (re)agendados
para momentos oportunos, em função da prioridade
de atendimento aos casos da Covid-19; assistência à
saúde no contexto de reabilitação cardiorrespiratória
(fator de maior prevalência e incidência nos casos
de pós Covid-19); reabilitação pós Covid-19 para
reinserção social. Os dados dos artigos selecionados foram
condensados e dispostos no Quadro 1 para descrição
das produções revisadas, as quais foram divididas por
autor, título, objetivos, delineamento metodológico e
principais resultados encontrados. Quanto ao delineamento metodológico dos
três artigos selecionados, dois tratavam-se de artigos
originais e um relato de experiência. Identificou-se
que, dois estudos utilizaram como método de pesquisa
a abordagem mista (qualitativo e quantitativo) e um
artigo a abordagem qualitativa descritiva. No que
se refere às técnicas de coleta dos dados, os artigos
originais fizeram uso de questionário com perguntas
abertas semiestruturadas. Com o objetivo de mapear e compreender as
inovações dos serviços especializados de cuidados
paliativos e as mudanças na prática dos serviços
em resposta a Covid-19, o artigo 1 se refere a uma
pesquisa online de prestadores de cuidados paliativos,
com dados coletados sobre as características do
serviço, suas inovações e mudanças em resposta à
pandemia. O estudo concluiu que a crise nos serviços
demandou rápidas mudanças que envolveram
principalmente a simplificação dos atendimentos
como forma de aumentar o alcance dos serviços. Tais
medidas ainda estão sendo avaliadas, mas relatou-
se a maior utilização das tecnologias para facilitar
a comunicação e implementar inovações no bem-
estar da equipe. SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Por fim, a quinta e sexta etapa referem-se,
respectivamente, à discussão e apresentação dos
dados. Para este estudo, utilizou-se a Análise de
Conteúdo proposta por Minayo e Bardin para a
análise e discussão dos dados. Os resultados foram
divididos em três campos temáticos de análise: (Re)
estruturação de fluxos e processos de trabalho; Saúde
e segurança dos trabalhadores da saúde; Modelos de
assistência à saúde. ao encontro da simplificação dos processos para
a agilidade nos atendimentos; inovação nos
atendimentos para suprir às demandas dos serviços, de
modo a facilitar a comunicação entre os profissionais
e entre os profissionais com os usuários; incentivo aos
modelos de trabalhos e fluxos que fomentem o bem-
estar da equipe. No que se refere ao campo temático da saúde
e segurança dos trabalhadores da saúde verificou-
se: ações de educação permanente para a equipe de
saúde no enfrentamento da Covid-19 por falta de
conhecimento e insegurança sobre a doença e seus
prognósticos; intensificação de atividades de incentivo
ao uso de Equipamentos de Proteção Individuais
(EPIs); altas e longas demandas laborais com impacto
na saúde mental e física dos profissionais. MÉTODO A terceira etapa diz respeito à coleta de dados
propriamente dita, para tanto, foram definidos dois
conjuntos de descritores, cada conjunto composto
por agrupamento de termos, sendo eles os descritores
acima referidos, tanto em língua portuguesa quanto
em inglesa, não havendo atravessamentos de ambas
as línguas para as buscas. Foi utilizado o navegador
Booleano “AND”, para somar as buscas e não excluir
nenhuma informação nela contida, garantindo a
abrangência e fidedignidade às buscas. A quarta etapa constitui-se da análise crítica Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664
Figura 1 - Fluxograma de busca, análise e seleção dos estudos para a revisão. Fonte: Banco de dados da pesquisa,
2021. Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 77 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 000Research
A
national
survey
of
community rehabilitation
service
provision
for
people with long COVID
in Scotland [version 2;
peer review: 2 approved]
O objetivo da pesquisa
foi descobrir o atual
fornecimento de serviços
para
reabilitação
de
pessoas que apresentam
sintomatologia
pós-
Covid na comunidade. Esta
pesquisa
nacional
teve como objetivo prover
uma descrição detalhada
do fornecimento atual de
reabilitação
comunitária
para pessoas com pós-Covid
na Escócia
Atualmente, quase todos os Conselhos de Saúde (13/14)
oferecem reabilitação para pessoas com Covid em serviços pré-
existentes. A maioria dos serviços de reabilitação comunitária
pós-Covid é fornecido por meio de uma combinação de contato
face a face e digital (13/14). RESULTADOS Os serviços de clínicas e de cuidados paliativos especializados
tiveram que implementar mudanças rapidamente para responder
ao impacto previsto e real da pandemia Covid-19. As mudanças
envolveram simplificar, ampliar e aumentar o alcance dos
serviços, bem como o uso de tecnologia de comunicação e
inovações para o bem-estar da equipe. Uma série de barreiras
e facilitadores para a mudança eram evidentes, como medo e
ansiedade do paciente, familiar cuidador e profissional de saúde,
duplicação de esforços e a combinação de recursos de pessoal
de saúde e trabalho colaborativo de equipe. Pesquisa
transversal,
multinacional, online, com
prestadores
de
cuidados
paliativos, clínicas, equipes
de
cuidados
paliativos
de
hospitais,
equipes
de
cuidados
paliativos
domiciliares e serviços de
enfermagem domiciliar. É preciso considerar um tempo de adaptação aos profissionais
alocados para novos ambientes, visto que pode lhe exigir trabalho
além de suas competências usuais na resposta à pandemia. Embora a modificação de ambiente possa interromper o
treinamento, o ensino e os testes na pós-graduação, o tratamento
intensivo é um rico ambiente de aprendizado para médicos
iniciantes. Profissionais mais experientes devem prestar atenção
especial ao bem-estar do profissional iniciante, auxiliando no
treinamento de simulação e na comunicação interprofissional. É preciso considerar um tempo de adaptação aos profissionais
alocados para novos ambientes, visto que pode lhe exigir trabalho
além de suas competências usuais na resposta à pandemia. Embora a modificação de ambiente possa interromper o
treinamento, o ensino e os testes na pós-graduação, o tratamento
intensivo é um rico ambiente de aprendizado para médicos
iniciantes. Profissionais mais experientes devem prestar atenção
especial ao bem-estar do profissional iniciante, auxiliando no
treinamento de simulação e na comunicação interprofissional. SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Quadro 1 - Características dos estudos incluídos na revisão integrativa adro 1 - Características dos estudos incluídos na revisão integrativa
AUTOR -
PERIÓDICO
TÍTULO
OBJETIVO
DELINEAMENTO
METODOLÓGICO
RESULTADOS
RTIGO
1-
Dunleavy
L,
eston N, Bajwah S, Bradshaw
Cripps R, Fraser LK,
addocks M, Hocaoglu M,
urtagh FEM, Oluyase AO,
eeman KE, Higginson IJ,
alshe C. Palliative Medicine
urnal
Necessity is the mother
of invention’: Specialist
palliative
care
service
innovation and practice
change in response to
COVID-19. Results from
a
multinational
survey
(CovPall)
Mapear e compreender
as inovações dos serviços
especializados
em
cuidados paliativos e as
mudanças na prática em
resposta ao Covid-19. Pesquisa
transversal,
multinacional, online, com
prestadores
de
cuidados
paliativos, clínicas, equipes
de
cuidados
paliativos
de
hospitais,
equipes
de
cuidados
paliativos
domiciliares e serviços de
enfermagem domiciliar. Os serviços de clínicas e de cuidados paliativos especializados
tiveram que implementar mudanças rapidamente para responder
ao impacto previsto e real da pandemia Covid-19. As mudanças
envolveram simplificar, ampliar e aumentar o alcance dos
serviços, bem como o uso de tecnologia de comunicação e
inovações para o bem-estar da equipe. Uma série de barreiras
e facilitadores para a mudança eram evidentes, como medo e
ansiedade do paciente, familiar cuidador e profissional de saúde,
duplicação de esforços e a combinação de recursos de pessoal
de saúde e trabalho colaborativo de equipe. RTIGO 2 - Coughlan C,
afde C, Khodatars S, Jeanes
L, Habib S, Donaldson E,
si C, Kooner GK. Postgrad
ed Journal
COVID-19: lessons for
junior doctors redeployed
to critical care
Descrever
princípios
relevantes para médicos
em formação designados
para
os
cuidados
intensivos
durante
a
pandemia Covid-19. Foram
estruturadas
seis
situações
por
meio
da
experiência de médicos que
trabalham em um hospital
universitário
urbano
de
um país de alta renda com
sistema de saúde universal. É preciso considerar um tempo de adaptação aos profissionais
alocados para novos ambientes, visto que pode lhe exigir trabalho
além de suas competências usuais na resposta à pandemia. Embora a modificação de ambiente possa interromper o
treinamento, o ensino e os testes na pós-graduação, o tratamento
intensivo é um rico ambiente de aprendizado para médicos
iniciantes. Profissionais mais experientes devem prestar atenção
especial ao bem-estar do profissional iniciante, auxiliando no
treinamento de simulação e na comunicação interprofissional. RTIGO 3 - Duncan E, Cooper
Cowie J, Alexander L,
orris J, Preston J. RESULTADOS Como barreiras, foi identificado que
houve aumento de medo e ansiedade nos profissionais
atuantes, onde a elevação do trabalho colaborativo e
a flexibilidade e liderança na equipe são importantes
para minimizar a duplicação de esforços e o uso de
recursos organizacionais. Os serviços onde os estudos foram realizados
se tratam de uma Unidade de Terapia Intensiva (UTI),
um serviço especializado em cuidados paliativos e
um serviço de reabilitação comunitária. Quanto aos
países/localidades de desenvolvimento dos estudos,
um foi realizado na Escócia, um no Reino Unido (UK)
e o outro de forma multicêntrica global, abarcando o
UK em sua maioria e diversos países europeus, sendo
que o país com maior prevalência de estudos foi o
UK. Quanto ao campo temático da (re)estruturação
de fluxos e processos de trabalho identificou-se os
seguintes resultados: reformulações e modificações
rápidas de fluxos e estruturas físicas dos serviços;
utilização de novas tecnologias que caminham 78 78 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 No artigo 2, estruturam-se recomendações
para médicos em formações com base em experiências
coletivas de profissionais em um hospital universitário
durante momentos críticos da pandemia no UK. Os principais postulados salientam o ambiente de
estresse gerado na área da saúde e que profissionais
com mais tempo de formação devem prestar apoio a
médicos iniciantes, auxiliando em seu treinamento
e na comunicação interprofissional. Embora tenha
ocorrido mudanças na dinâmica da assistência, a
atuação intensiva proporciona um ambiente rico de
aprendizado para profissionais iniciantes. a segurança da sociedade e a mitigação do risco de
contágio, como uma das repercussões apontadas nos
artigos selecionados.10 O emprego de ferramentas
digitais mostra-se como importante aliado diante
dessa realidade, na qual emergem diversas questões
psicossociais que condicionam a articular novos
modelos assistenciais, a exemplo do atendimento de
forma virtual ou presencial quando possível, ambos
visando manter a segurança e o bem-estar de todos.11 O uso da internet e suas variantes viabilizou
de forma fluida o monitoramento de pacientes e a
assistência à saúde, facilitando a comunicação. O
formato virtual também evidenciou as fragilidades
de um sistema de saúde, trazendo à tona inúmeras
mazelas em relação ao autocuidado e à credibilidade
científica, deixando explícito o complexo caminho a
ser estudado.12 O artigo 3 aborda uma pesquisa descritiva
no sistema de saúde escocês, realizada por meio de
perguntas relacionadas aos serviços de reabilitação. As perguntas foram direcionadas para os diretores
das 14 regiões correspondentes ao sistema de saúde
escocês. Treze conselheiros de saúde indicaram
oferecer serviços de reabilitação para pacientes pós
Covid-19 em serviços pré-existentes, uma região de
saúde criou um novo serviço e outra região está em
fase de elaboração de um novo serviço. Treze serviços
ofertam assistência combinada, presencial e por meio
digital. A reabilitação pós Covid-19 é uma realidade
emergente, sendo prestada predominantemente de
modo multidisciplinar, já que resultados apontam que
15 a 35% das pessoas infectadas podem apresentar
queixas após o acometimento pela doença e necessitar
desse tipo de assistência. Dessa forma, a emergência na continuidade das
práticas diante do fluxo aumentado de atendimentos
obrigou a rápida implementação e adaptação da rede
tecnológica disponível nos diferentes campos de
atuação. Nesse contexto, ações práticas não focadas
diretamente no manejo de pacientes graves podem
atrair menos atenção diante da crise, mas possuem
potencial semelhante para contribuir em melhorias que
continuarão mesmo após a pandemia. SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Devido a isso,
as circunstâncias que exigiram lidar com imprevistos
para manter os serviços durante padrões anormais de
atendimento são essenciais não somente na linha de
frente, onde a assistência foi duramente atingida pela
escassez de recursos para o manejo da Covid-19, mas
também na atenção social à comunidade.13 Assim, a
telesaúde, praticada previamente a pandemia, obteve
significativo destaque como uma ação que pôde
manter ativa a atuação de serviços na assistência à
saúde. Na mesma esteira, as consultas virtuais e o
suporte via telefone supriram parte da necessidade
em acompanhar pacientes com doenças crônicas
e até mesmo a triagem de quadros agudos, mas
notavelmente alargou ainda mais o distanciamento
gerado por medidas de isolamento social, sendo a
falta de proximidade e de contato humano um dos
principais amplificadores de patologías psíquicas
durante a pandemia.14 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 Foram
estruturadas
seis
situações
por
meio
da
experiência de médicos que
trabalham em um hospital
universitário
urbano
de
um país de alta renda com
sistema de saúde universal. Esta
pesquisa
nacional
teve como objetivo prover
uma descrição detalhada
do fornecimento atual de
reabilitação
comunitária
para pessoas com pós-Covid
na Escócia O objetivo da pesquisa
foi descobrir o atual
fornecimento de serviços
para
reabilitação
de
pessoas que apresentam
sintomatologia
pós-
Covid na comunidade. Atualmente, quase todos os Conselhos de Saúde (13/14)
oferecem reabilitação para pessoas com Covid em serviços pré-
existentes. A maioria dos serviços de reabilitação comunitária
pós-Covid é fornecido por meio de uma combinação de contato
face a face e digital (13/14). Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 79 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 DISCUSSÃO No que tange à necessidade de reformulações e
modificações rápidas de fluxos e estruturas físicas dos
serviços apontada nos artigos, a compreensão deste
novo contexto instaurado mundialmente exigiu dos
profissionais de saúde e gestão, novos planejamentos,
estratégias e condutas para propiciar modificações
de fluxos e estruturas dos serviços de atendimentos
à população. A fim de evitar o colapso dos sistemas
de saúde, os modelos de atenção foram (re)pensados
e (re)configurados em face das novas demandas,
de forma rápida. As capacitações profissionais e
revisitações de conceitos básicos de prevenção e
atenção à saúde como a correta lavagem de mãos e
uso de EPIs, fizeram-se importantes e indispensáveis
para a contenção de impactos negativos.9 Analogamente, ao passo que a tecnologia
foi utilizada para suprir a demanda populacional,
ferramentas de comunicação demonstraram grande
potencial em facilitar a dinâmica entre as equipes,
gerando maior agilidade na comunicação entre
os profissionais e, também, reduzindo o estresse A utilização de novas tecnologias caminha ao
encontro da simplificação dos processos e agilidade
nos atendimentos, os quais apresentam como objetivo 80 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 ocupacional, principalmente em jornadas intensas. Com efeito, constatou-se aumento significativo
de estresse, ansiedade e insônia de profissionais de
saúde frente à exposição e consequente maior risco
de infecção, assim como, o aumento da prevalência
de sofrimento psicológico entre trabalhadores fora
da linha de frente, com prováveis causas a falta de
acesso à informação e menor apoio psicológico.15
Isso demonstra que os mecanismos desenvolvidos
na pandemia podem trazer ao debate necessidades
anteriores que ainda não haviam sido evidenciadas,
afinal, medidas que impulsionam a comunicação entre
a equipe promovem maior bem-estar e apoio, tendo
em vista que os profissionais de saúde frequentemente
negligenciam os relacionamentos com familiares
e amigos devido as longas cargas de trabalho. Portanto, todo o contexto pode ser observado como
potencial inovador para recuperar a crise na saúde
e o impacto social, fortalecendo a resiliência dos
sistemas de trabalho, a assistência social e a gestão
de recursos, inclusive para imprevistos futuros. DISCUSSÃO Logo, a implementação de estratégias de trabalho
e o desenvolvimento de práticas inovadoras deve
manter-se como forma de avaliar limitações e adotar
melhorias.16 destaca-se a atuação das Estratégias Saúde da Família
(ESFs) na abordagem da saúde pública, de forma a
agirem diretamente com a população, promovendo
ações de prevenção e promoção da saúde, por meio de
princípios dispostos pelo SUS, como universalidade,
integralidade e equidade.19 Ademais, soma-se à essa atuação baseada em
processos de educação em saúde a percepção de que o
uso de EPIs se torna ainda mais essencial. A fragilidade
do setor de saúde foi colocada em evidência no decorrer
da pandemia, uma vez que a necessidade em promover
a segurança aos trabalhadores da saúde configurou-
se como um desafio, exigindo uma combinação de
abordagens administrativas nos serviços de saúde para
o cumprimento de medidas necessárias de proteção.20
Somado a lacunas prévias no quesito do conhecimento
e da aplicação das informações acerca do uso de EPIs,
notou-se evidente aumento das ações relacionadas ao
incentivo do seu uso. Ainda, destaca-se a importância
de órgãos nacionais e internacionais convergirem
com as recomendações preconizadas, salientando a
importância das informações disponibilizadas pela
Organização Mundial da Saúde.21 As altas e extensas demandas de serviços dos
trabalhadores da saúde influenciaram na saúde mental
e física durante a pandemia da Covid-19, considerando
o seu papel fundamental no atendimento aos casos
de pacientes infectados. Evidenciam-se horas-extras
no trabalho de enfermeiros na linha de frente da
assistência, impactando negativamente em sua saúde
e bem-estar pela maior pressão laboral envolvida
nessas circunstâncias.22 Nessa situação, entende-se
que há uma discrepância entre o esforço feito pelo
trabalhador e a recompensa pela sua tarefa laboral
realizada, levando-se em conta o maior desgaste físico
e mental ao longo da pandemia. DISCUSSÃO Em contrapartida,
estar engajado e satisfeito com o trabalho leva a uma
relação favorável na autoavaliação e autocuidado de
sua saúde.23i Nessa linha, a Educação Permanente em
Saúde (EPS) traz consigo a concepção de trabalho
ambientada no Sistema Único de Saúde, além da
aprendizagem cotidiana e o comprometimento com
o coletivo, é uma forma de promover uma atuação
satisfatória no que se refere à saúde pública no
Brasil.17 No âmbito da Política Nacional de Educação
Permanente em Saúde, a EPS objetiva nortear a
formação e a qualificação dos profissionais inseridos
nos serviços públicos de saúde, visando transformar
práticas profissionais e organizar o trabalho baseando-
se em demandas.18 Entende-se como uma alternativa
plausível a utilização da EPS para a capacitação dos
trabalhadores da saúde, o que se destaca de modo
mais ampliado no cenário atual mundial. A pandemia
acentuou a importância do cuidado em saúde e dessa
forma, a necessidade de profissionais com a habilidade
de reconhecer e tratar patologias.18 Nesse contexto, os profissionais de saúde
tiveram que lidar, em algumas situações, com o
improviso de recursos de saúde, uma escolha por
vezes difícil e complexa, envolvendo aspectos
bioéticos. Outra questão a ser ponderada é que esses
trabalhadores ainda precisa considerar suas próprias
necessidades holísticas, visto que estão expostos a
inúmeros estressores, sujeitos a agravos psicológicos
e danos morais a longo prazo.24i Acentua-se, de igual forma, a relevância
da educação em saúde no que se refere, também,
aos processos da EPS que podem ser ampliados a
diferentes níveis de gestão, vinculando os municípios
às referências nacionais para o seu processo de
formação e desenvolvimento.17 Nesse sentido, Identificou-se nos estudos que procedimentos Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 81 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 eletivos passaram a ser (re)agendados para momentos
oportunos, em função da prioridade de atendimento
aos casos de Covid-19. No Brasil, a Agência Nacional
de Saúde recomendou a avaliação criteriosa para que
apenas fossem agendados os atendimentos quando
o seu adiamento prejudicasse a saúde do paciente,
de modo a organizar os recursos assistenciais e
garantir a manutenção dos cuidados emergenciais. Especialmente no pico da pandemia, a priorização
dos recursos disponíveis foi feita através de critérios
clínicos, éticos e epidemiológicos locais. DISCUSSÃO A partir
disso, mostrou-se necessário informar ao paciente e
a sua família detalhadamente os riscos, benefícios e
necessidades advindas da recuperação domiciliar,
visto que há menor risco de contágio se a alta hospitalar
for realizada o mais precocemente possível.25 o trabalho multi e intersetorial deve continuar para
atender às necessidades de reabilitação causadas
pela mesma. A transformação da reabilitação deve
oferecer uma abordagem individualizada, com base
nas necessidades, para permitir que as pessoas vivam
com qualidade de vida, apesar do Covid-19.30 CONSIDERAÇÕES FINAIS Este estudo buscou dar visibilidade ao
tema em questão, reforçando a necessidade de
investigações científicas a respeito, considerando
a pequena quantidade de estudos encontrados
no contexto de pós-Covid-19 nos serviços e na
assistência à saúde. Por outro lado, o estudo realizado
considera a importância da existência dos três artigos
selecionados, reconhecendo que a pandemia ainda
está em curso e que publicações acerca do tema já
estejam sendo produzidas mundialmente. p
p
Os
artigos
selecionados
apontaram
a
predominância da assistência à saúde no contexto de
reabilitação cardiorrespiratória como fator de maior
prevalência e incidência nos casos de pós Covid-19. Sabe-se que a fisiopatologia da doença ainda é pouco
conhecida, podendo levar à síndrome do desconforto
respiratório agudo e também originar lesões
circulatórias. A doença provoca hiperviscosidade
sanguínea, determinando um fator de risco para a
obstrução de artérias por material trombótico de
forma generalizada, entretanto, o impacto futuro das
lesões é desconhecido.26 Os
artigos
selecionados
apontaram
a
predominância da assistência à saúde no contexto de
reabilitação cardiorrespiratória como fator de maior
prevalência e incidência nos casos de pós Covid-19. Sabe-se que a fisiopatologia da doença ainda é pouco
conhecida, podendo levar à síndrome do desconforto
respiratório agudo e também originar lesões
circulatórias. A doença provoca hiperviscosidade
sanguínea, determinando um fator de risco para a
obstrução de artérias por material trombótico de
forma generalizada, entretanto, o impacto futuro das
lesões é desconhecido.26 Os
artigos
selecionados
apontaram
a
predominância da assistência à saúde no contexto de
reabilitação cardiorrespiratória como fator de maior
prevalência e incidência nos casos de pós Covid-19. j
p
Os
resultados
apontaram
aspectos
relacionados à necessidade de diversas modificações
e adaptações dos serviços de saúde na realidade
de atendimentos após a infecção pela Covid-19. Essas situações se referem mais especificamente à
reestruturação de fluxos, protocolos e processos de
trabalho, com a utilização de tecnologias e novos
processos de comunicação. A adoção de medidas
de promoção da saúde, de prevenção da doença e
de proteção e segurança dos trabalhadores da saúde
foi muito enfatizada como resultado, já que se trata
de uma nova realidade de risco biológico presente
no trabalho em saúde. A reformulação de modelos
e padrões de assistência à saúde dos mesmos trouxe
como destaque a inserção de serviços de reabilitação
cardiorespiratória e também de reinserção social,
já que a pandemia afetou não somente a saúde das
pessoas, mas também aspectos econômicos, de renda
e trabalho da população. Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 REFERÊNCIAS 12. Coelho AL, de Araujo Morais I, da Silva Rosa WV. A
utilização de tecnologias da informação em saúde para o
enfrentamento da pandemia do Covid-19 no Brasil. Cadernos
Ibero-Americanos de Direito Sanitário. 2020 Sep 29;9(3):183-
99. https://www.cadernos.prodisa.fiocruz.br/index.php/
cadernos/article/download/709/754 1. Wang D, Hu B, Hu C, Zhu F, Liu X, Zhang J, Wang B, Xiang
H, Cheng Z, Xiong Y, Zhao Y, Li Y, Wang X, Peng Z. Clinical
Characteristics of 138 Hospitalized Patients with 2019 Novel
Coronavirus-Infected Pneumonia in Wuhan, China. JAMA
- J Am Med Assoc. 2020;323(11):1061–9. doi: http://dx.doi. org/10.1001/jama.2020.1585 1. Wang D, Hu B, Hu C, Zhu F, Liu X, Zhang J, Wang B, Xiang H, Cheng Z, Xiong Y, Zhao Y, Li Y, Wang X, Peng Z. Clinical Characteristics of 138 Hospitalized Patients with 2019 Novel
Coronavirus-Infected Pneumonia in Wuhan, China. JAMA Characteristics of 138 Hospitalized Patients with 2019 Novel 13. Wiedner R, Croft C, McGivern G. Improvisation during a
crisis: hidden innovation in healthcare systems. BMJ Leader. 2020 Jun 17:leader-2020. doi: http://dx.doi.org/10.1136/
leader-2020-000259 - J Am Med Assoc. 2020;323(11):1061–9. doi: http://dx.doi. org/10.1001/jama.2020.1585 2. Phua J, Weng L, Ling L, Egi M, Lim CM, Divatia JV, Shrestha
BR, Arabi YM, Ng J, Gomersall CD, Nishimura M, Hoh Y, Du
B. Intensive care management of coronavirus disease 2019
(COVID-19): challenges and recommendations. Lancet Respir
Med. 2020;8(5):506–17. doi: http://dx.doi.org/10.1016/S2213-
2600(20)30161-2 14. Dunleavy L, Preston N, Bajwah S, Bradshaw A, Cripps
R, Fraser LK, Maddocks M, Hocaoglu M, Murtagh FEM,
Oluyase AO, Sleeman KE, Higginson IJ, Walshe C. On behalf
of the CovPall study team‘Necessity is the mother of invention’:
Specialist palliative care service innovation and practice change
in response to COVID-19. Results from a multinational survey
(CovPall). Palliat Med. 2021 Mar 23;35(5):814–29. doi: http://
dx.doi.org/10.1177/02692163211000660 3. Organização Mundial da Saúde. Risk Communication
Strategy for Public Health Emergencies in the WHO South-East
Asia Region: 2019–2023. World Health Organization. Regional
Office for South-East Asia. 2019. doi: https://apps.who.int/iris/
handle/10665/326853 15. Tan BYQ, Chew NWS, Lee GKH, Jing M, Goh Y, Yeo LLL,
Zhang K, Chin HK, Ahmad A, Khan FA, Shanmugam GN, Chan
BPL, Sunny S, Chandra B, Ong JJY, Paliwal PR, Wong LYH,
Sagayanathan R, Chen JT, Ng AYY, Teoh HL, Ho CS, Ho RC,
Sharma VK. Psychological Impact of the COVID-19 Pandemic
on Health Care Workers in Singapore. Annals of Internal
Medicine. 2020 Ago 18;173(4):317–20. doi: http://dx.doi. org/10.7326/M20-1083 4. Ohannessian R, Duong TA, Odone A. SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 REFERÊNCIAS Global Telemedicine
Implementation and Integration Within Health Systems to
Fight the COVID-19 Pandemic: A Call to Action. JMIR Public
Health Surveill. 2020 abr 2;6(2):e18810. doi: http://dx.doi. org/10.2196/18810 5. Monroy-Fraustro D, Maldonado-Castellanos I, Aboites-
Molina M, Rodríguez S, Sueiras P, Altamirano-Bustamante NF,
Hoyos-Bermea A, Altamirano-Bustamante M. Bibliotherapy as
a Non-pharmaceutical Intervention to Enhance Mental Health
in Response to the COVID-19 Pandemic: A Mixed-Methods
Systematic Review and Bioethical Meta-Analysis. Front
Public Health. 2021 mar 15;9. doi: http://dx.doi.org/10.3389/
fpubh.2021.629872 16. Blake H, Bermingham F, Johnson G, Tabner A. Mitigating
the Psychological Impact of COVID-19 on Healthcare Workers:
A Digital Learning Package. IJERPH. 2020 abr 26;17(9):2997. doi: http://dx.doi.org/10.3390/ijerph17092997 17. Campos FE, Haddad AE, Roschke MA, Galvão ED. Política
nacional de educação permanente em saúde. In Política nacional
de educação permanente em saúde 2009 (pp. 65-65). https://
bvsms.saude.gov.br/bvs/publicacoes/pacto_saude_volume9.pdf
18. Ferreira L, Barbosa JS, Esposti CD, Cruz MM. Educação
Permanente em Saúde na atenção primária: uma revisão
integrativa da literatura. Saúde em Debate. 2019 May 6;43:223-
39. doi: https://doi.org/10.1590/0103-1104201912017i 6. França MS, Lopes MV, Frazão CM, Guedes TG, Linhares
FM, Pontes CM. Características da rede social de apoio ineficaz:
revisão integrativa. Revista Gaúcha de Enfermagem. 2018 Oct
22;39. doi: https://doi.org/10.1590/1983-1447.2018.20170303 7. Neta IS, Medeiros MS, Gonçalves MJ. Vigilância da saúde
orientada às condições de vida da população: uma revisão
integrativa da literatura. Saúde em Debate. 2018;42:307-17. doi:
https://doi.org/10.1590/0103-1104201811625 p
g
19. Silva RD, Rotondano PN, Sousa MM, Purificação ER, Santos
MP, Veras RP, Soares CD, Merces MC. Práticas interdisciplinares
no enfrentamento da COVID-19 na estratégia saúde da família. Enferm. foco (Brasília). 2020:246-53. http://revista.cofen.gov. br/index.php/enfermagem/article/download/4220/1014
20. Ribeiro AP, Oliveira GL, Silva LS, Souza ER. Saúde e
segurança de profissionais de saúde no atendimento a pacientes
no contexto da pandemia de Covid-19: revisão de literatura. Revista Brasileira de Saúde Ocupacional. 2020; 45. doi: https://
doi.org/10.1590/2317-6369000013920 19. Silva RD, Rotondano PN, Sousa MM, Purificação ER, Santos
MP, Veras RP, Soares CD, Merces MC. Práticas interdisciplinares
no enfrentamento da COVID-19 na estratégia saúde da família. Enferm. foco (Brasília). 2020:246-53. http://revista.cofen.gov. br/index.php/enfermagem/article/download/4220/1014 8. Moher D, Liberati A, Tetzlaff J, Altman DG, Prisma Group. Preferred reporting items for systematic reviews and meta-
analyses: the PRISMA statement. PLoS medicine. 2009
Jul 21;6(7):e1000097. doi: https://doi.org/10.1371/journal. pmed.1000097 20. Ribeiro AP, Oliveira GL, Silva LS, Souza ER. Saúde e
segurança de profissionais de saúde no atendimento a pacientes
no contexto da pandemia de Covid-19: revisão de literatura. Revista Brasileira de Saúde Ocupacional. 2020; 45. CONSIDERAÇÕES FINAIS No curso da pandemia,
enfatizou-se e priorizou-se o atendimento a casos
da Covid-19 em relação a situações eletivas de
atendimentos e procedimentos. Os artigos também apontaram serviços e
assistência em reabilitação pós Covid-19 voltados
à reinserção social, entre eles, às pessoas com
deficiência e os que têm necessidade de apoio social
por não poderem trabalhar.27 No entanto, sabe-se que
as pessoas com deficiência, independentemente da
infecção pelo Covid-19, sofrem com as restrições
impostas à prestação de serviços de reabilitação, que
corroboram com as questões de sua vulnerabilidade
social.28 Mundialmente, a circulação do vírus alterou
a mobilização de recursos necessários para o
desenvolvimento de estratégias voltadas a promover
a segurança coletiva, por meio de ações individuais. Sob essa perspectiva, a disponibilização de serviços
de reabilitação sofreu restrições impostas pelas
transformações causadas pela pandemia.29 Como lacuna do estudo, considera-se a não
publicação de investigações realizadas no Brasil,
destacando a necessidade de abordagem desse tema
em periódicos nacionais, já que a busca dos artigos
apontou somente estudos internacionais, mostrando
uma hegemonia predominante em relação aos estudos
brasileiros. A Covid-19 alterou a forma como a sociedade
vive, cuida e trabalha, sendo assim, os serviços de
reabilitação precisam se adaptar a esse novo contexto. À medida que os países saírem da onda pandêmica, Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 82 82 REFERÊNCIAS doi: https://
doi.org/10.1590/2317-6369000013920 9. Santos RD, Barreto CT, Lemos PF, Duarte CA, Moreira DD,
Reis AT, Silva FH, Nunes AS. Management of a university
ambulatory service: nursing in the coping of the pandemic of
COVID-19. Revista Brasileira de Enfermagem. 2021 Apr 14;74. doi: https://doi.org/10.1590/0034-7167-2020-0834 21.World Health Organization. Rational use of personal
protective equipment for coronavirus disease ( COVID-19)
and considerations during severe shortages: interim guidance,
6 April 2020. World Health Organization; 2020. https://apps. who.int/iris/bitstream/handle/10665/331695/WHO-2019-nCov-
IPC_PPE_use-2020.3-ara.pdf 10. Merhy EE, Feuerwerker LC, Santos ML, Bertussi DC, Baduy
RS. Basic Healthcare Network, field of forces and micropolitics:
implications for health management and care. Saúde em Debate. 2020 Jul 10;43:70-83. doi: https://doi.org/10.1590/0103-
11042019S606 22. Watanabe M, Yamauchi K. The effect of quality of overtime
work on nurses’ mental health and work engagement. Journal of
Nursing Management. 2018 Sep;26(6):679-88. doi: https://doi. org/10.1111/jonm.12595 11. Souza BA, Tritany ÉF. COVID-19: the importance of new
technologies for physical activity as a public health strategy. Cadernos de Saúde Pública. 2020 May 18;36. doi: https://doi. org/10.1590/0102-311X00054420 23. Ge J, He J, Liu Y, Zhang J, Pan J, Zhang X, Liu D. Effects of
effort-reward imbalance, job satisfaction, and work engagement Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 v. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 83 SERVIÇOS E ASSISTÊNCIA EM SAÚDE APÓS COVID-19 on self-rated health among healthcare workers. BMC Public
Health. 2021 Dec;21(1):1-0. doi: https://doi.org/10.1186/
s12889-021-10233-w 24. Ho CY, Kow CS, Chia CH, Low JY, Lai YH, Lauw SK, How
AE, Tan LH, Ngiam XL, Chan NP, Kuek TY. The impact of death
and dying on the personhood of medical students: a systematic
scoping review. BMC medical education. 2020 Dec;20(1):1-6. doi: https://doi.org/10.1186/s12909-020-02411-y 25. Di Martino M, Septiem JG, González RM, de Nova JL, de la
Hoz Rodríguez Á, Bonito AC, Martín-Pérez E. Cirugía electiva
durante la pandemia por SARS-CoV-2 (COVID-19): análisis
de morbimortalidad y recomendaciones sobre priorización
de los pacientes y medidas de seguridad. Cirugía Española. 2020 Nov 1;98(9):525-32. doi: https://doi.org/10.1016/j. ciresp.2020.04.029 26. José A, Muller MG, Malaguti C. Repercussões respiratórias
e funcionais após infecção por COVID-19. In: Hordonho AAC,
Napoleão AA, Lopes AC, Lopes CT, Kubiak CA, Martins JA,
Cipullo JP, Coppini LZ, Fidelix MSP, Karsten M, Corso SD,
Herdman TH, da Silva VM, organizadores. Especial Covid-19:
Ciclo 1. Porto Alegre: Artmed Panamericana; 2020. p. 9–29. (Sistema de Educação Continuada a Distância, v. 5). Recebido em: 15/10/2021
Aceito em:30/10/2021 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 content/uploads/2020/05/Journal_Infection_Control.pdf
30. De Biase S, Cook L, Skelton DA, Witham M, Ten Hove R.
The COVID-19 rehabilitation pandemic. Age and ageing. 2020
Aug 24;49(5):696-700. doi: https://doi.org/10.1093/ageing/
afaa118 REFERÊNCIAS https://
portal.secad.artmed.com.br/artigo/repercussoes-respiratorias-e-
funcionais-apos-infeccao-por-covid-19#_idParaDest-13 27. Gutenbrunner C, Stokes EK, Dreinhöfer K, Monsbakken J,
Clarke S, Côté P, Urseau I, Constantine D, Tardif C, Balakrishna
V, Nugraha B. Why rehabilitation must have priority during and
after the COVID-19 pandemic: A position statement of the global
rehabilitation alliance. Journal of rehabilitation medicine. 2020
l
( )
d i h
//d i
/
/ p
p
g
rehabilitation alliance. Journal of rehabilitation medicine. 2020
Jul 5;52(7):1-4. doi: https://doi.org/10.2340/16501977-2713
28. Andrenelli E, Negrini F, De Sire A, Arienti C, Patrini M,
Negrini S, Ceravolo MG. Systematic rapid living review on
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Jun 16. doi: https://doi.org/10.23736/S1973-9087.20.06435-7
29. Dias VM, Carneiro M, Vidal CF, Corradi MF, Brandão
D, Cunha CA, Chebabo A, Oliveira PD, Michelin L, Rocha
JL, Waib LF. Orientações sobre diagnóstico, tratamento e
isolamento de pacientes com COVID-19. J Infect Control. 2020
Apr 13;9(2):56-75. http://www.abennacional.org.br/site/wp-
content/uploads/2020/05/Journal_Infection_Control.pdf
30. De Biase S, Cook L, Skelton DA, Witham M, Ten Hove R. The COVID-19 rehabilitation pandemic. Age and ageing. 2020
Aug 24;49(5):696-700. doi: https://doi.org/10.1093/ageing/
afaa118 Jul 5;52(7):1 4. doi: https://doi.org/10.2340/16501977 2713
28. Andrenelli E, Negrini F, De Sire A, Arienti C, Patrini M,
Negrini S, Ceravolo MG. Systematic rapid living review on
rehabilitation needs due to Covid-19: update to May 31st 2020. European journal of physical and rehabilitation medicine. 2020 Jun 16. doi: https://doi.org/10.23736/S1973-9087.20.06435-7
29. Dias VM, Carneiro M, Vidal CF, Corradi MF, Brandão
D, Cunha CA, Chebabo A, Oliveira PD, Michelin L, Rocha
JL, Waib LF. Orientações sobre diagnóstico, tratamento e
isolamento de pacientes com COVID-19. J Infect Control. 2020
Apr 13;9(2):56-75. http://www.abennacional.org.br/site/wp- content/uploads/2020/05/Journal_Infection_Control.pdf
30. De Biase S, Cook L, Skelton DA, Witham M, Ten Hove R. The COVID-19 rehabilitation pandemic. Age and ageing. 2020
Aug 24;49(5):696-700. doi: https://doi.org/10.1093/ageing/
afaa118 Recebido em: 15/10/2021
Aceito em:30/10/2021 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 Rev. Interdisciplin. Promoç. Saúde - RIPS, Santa Cruz do Sul, 4(2):75-84, abr/jun. 2021 ISSN: 2595-3664 84 84
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https://openalex.org/W2967616910
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https://link.springer.com/content/pdf/10.1007/s00605-022-01807-w.pdf
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English
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On volume preserving almost Anosov flows
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Monatshefte für Mathematik
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cc-by
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Monatshefte für Mathematik (2023) 201:1003–1026
https://doi.org/10.1007/s00605-022-01807-w Monatshefte für Mathematik (2023) 201:1003–1026
https://doi.org/10.1007/s00605-022-01807-w We gratefully acknowledge the support of FWF grant P31950-N45. B Henk Bruin
henk.bruin@univie.ac.at Communicated by Adrian Constantin. Abstract The purpose of this paper is to establish limit laws for volume preserving almost
Anosov maps of T2 flows on 3-three manifolds having a transversally neutral periodic
orbit of cubic saddle type. In the process, we derive estimates for the Dulac maps for
neutral saddles in planar vector fields. Keywords Dulac map · Almost Anosov flows · Volume preserving flows · Stable
laws · Limit theorems Keywords Dulac map · Almost Anosov flows · Volume preserving flows · Stable
laws · Limit theorems Mathematics Subject Classification Primary 37A25; Secondary 28D05 · 37A40 ·
37A50 Henk Bruin1 Received: 15 May 2022 / Accepted: 25 November 2022 / Published online: 20 December 2022
© The Author(s) 2022 1
Faculty of Mathematics, University of Vienna, Oskar Morgensternplatz 1, 1090 Vienna, Austria On volume preserving almost Anosov flows Henk Bruin1 1 Introduction A flow φt : M × R →M on a (in our setting 3-dimensional) compact differentiable
manifold M is called Anosov if its tangent bundle has a continuous flow-invariant
mutually transversal splitting into a neutral flow direction Ec, a hyperbolically stable
direction Es and a hyperbolically unstable direction Eu. The uniform hyperbolicity
of such flows enables one to show various ergodic and statistical properties, such as
ergodicity (if the flow is topologically mixing) and the Central Limit Theorem (CLT)
for Hölder continuous observables. We obtain an almost Anosov flow (see Definition 1.1 below) by inserting a neutral
orbit ≃{(0, 0)} × S1 near which the flow has the following form in local Euclidean
coordinates: 1
Faculty of Mathematics, University of Vienna, Oskar Morgensternplatz 1, 1090 Vienna, Austria 3 1004 H. Bruin ⎛
⎝
˙x
˙y
˙z
⎞
⎠= X
⎛
⎝
x
y
z
⎞
⎠=
⎛
⎝
x(a0x2 + a1xy + a2y2)
−y(b0x2 + b1xy + b2y2)
1 + w(x, y)
⎞
⎠+ O(4)
(1) (1) where the parameters satisfy where the parameters satisfy a1, b1 ∈R, a0, a2, b0, b2 ≥0 with := a2b0 −a0b2 ̸= 0 and
a2
1 < 4a0a2, b2
1 < 4b0b2. (2) a1, b1 ∈R, a0, a2, b0, b2 ≥0 with := a2b0 −a0b2 ̸= 0 and
2
2 (2) The last two conditions of (2) imply that the first component in (1) is non-negative
and the second non-positive for x, y ≥0. The term O(4) indicates terms of order four
and higher, under the condition that they are of the form x2O(2) near the yz-plane
and of the form y2O(2) near the xz-plane, because otherwise these are not a small
perturbation compared to the leading terms of (1). These leading terms are cubic in
the direction transversal to , but this is the only source of non-hyperbolicity. Finally,
w is a linear combination of homogeneous functions in x and y, vanishing at (0, 0). Thus the period of is its length. The original motivation to study such systems was to find a class of natural examples
of non-uniformly hyperbolic invertible maps where operator renewal theory can be
applied to establish precise statistical laws. 1 Introduction The map in this context can be the Poincaré
map on a section ⊂R2 ×{0} or the time-1 map fhor = φ1
hor for the horizontal flow
where only the x- and y-coordinates are taken into account: ˙x
˙y
= Xhor
x
y
=
x(a0x2 + a1xy + a2y2)
−y(b0x2 + b1xy + b2y2)
+ O(4),
(3) (3) 123 with the restrictions (2). Initially, in [2] for the parameter range β2 := a2+b2
2b2
≤1 where fhor preserves an
infinite Sina˘ı-Bowen-Ruelle (SRB) measure, we gave mixing rates for C1 observables. Later [3], and more relevant to this paper, in the parameter range β2 > 1 where the
flow φt preserves a finite SRB-measure, we established limit laws (Stable Laws and
the CLT with standard or non-standard scaling, depending on whether β2 ∈(1, 2),
β = 2 or β2 > 2). All these results were obtained in the absence of mixed terms, i.e., a1 = b1 = 0 in
(1). This is of course not a natural assumption, and to our knowledge there is no change
of coordinates that allows one to remove the mixed terms. In fact, if a2
1 > 4a0a2 or
b2
1 > 4b0b2, then the dynamics near the saddle is not locally conjugate to the dynamics
under the current condition (2). The purpose of this paper is to perform the analysis when mixed terms are present,
and also the treatment of the perturbation terms (see Sect.3) is substantially different
and more straightforward than in [2, 3]. The crux of the analysis is the existence of
a local first integral (and its explicit form when O(4)-terms are absent in (9)), which
allows us to reduce the ODE to dimension one. We will show in Lemma 2.1 that the
first integral L can be found if 1005 On volume preserving almost Anosov flows Fig. 1 The Dulac map
D : (ξ(η0, T ), η0) →
(ζ0, ω(η0, T )) with
Dulac time T
b1
a1
= b2a0 + a2 + 2b0b2
b2a0 + a2 + 2a0a2
. (4)
Thi i
di
i
diti
i
t
H
if
l
ti
l t Fig. 1 The Dulac map
D : (ξ(η0, T ), η0) →
(ζ0, ω(η0, T )) with
Dulac time T (4) This is a co-dimension one condition in parameter space. However, if we also stipulate
that the flow φt is volume preserving, we must assume that div X = 0 in (1), which is
equivalent to div O(4) = 0 together with This is a co-dimension one condition in parameter space. 1 The precise values of ξ0(η) and ω0(η) are given in in the proof Proposition 2.1. with the restrictions (2). However, if we also stipulate
that the flow φt is volume preserving, we must assume that div X = 0 in (1), which is
equivalent to div O(4) = 0 together with 3a0 = b0, a2 = 3b2, a1 = b1. (5) (5) Condition (4) follows automatically from (5), and therefore (1) describes a generic
volume preserving almost Anosov flow with a single neutral periodic orbit of cubic
saddle type. We present the results on limit laws in the volume preserving setting, see
Theorem 1.2. Central to the proof of the main theorem of this paper (as well as precise mixing
rates) is the analysis of the Dulac map near the neutral equilibrium of (3). This means
that we take an incoming and an outgoing transversal to the flow, in our case an
unstable leaf W u(0, η), η ∈[η0, η1], and a stable leaf W s(ζ0, 0), see Fig.1. The Dulac
map D : W u(0, η) →W s(ζ0, 0) assigns to (ξ, η) the first intersection φT
hor(ξ, η)
of the integral curve through (ξ, η) with the outgoing transversal W s(ζ0, 0). The
corresponding flow-time is denoted by T . As main technical result of this paper we
obtain precise estimates of the Dulac map when (3) contains mixed terms, but using
the assumption (4). Dulac [5] introduced his map as an ingredient to prove that polynomial vector fields
in the plane have at most finitely many limit cycles, thus making a major contribution
to the solution of Hilbert’s 16th problem. Il’yashenko [7] corrected some weak parts in
Dulac’s arguments, see also the summary in Roussarie’s book [12, Sect.3.3]. Further
contributions are due to e.g. Dumortier and more recently Mardeši´c and collaborators
[4, 8–10, 13]. Specifically, polycycles (i.e., heteroclinic saddle connections) are not 12 3 1006 H. Bruin H. Bruin accumulated upon by limit cycles. Whereas our estimates only concern a single neutral
saddle, it is to our knowledge the first estimation of the Dulac times (and hence the
Dulac map, see (6)), at cubic saddle of this generality. accumulated upon by limit cycles. Whereas our estimates only concern a single neutral
saddle, it is to our knowledge the first estimation of the Dulac times (and hence the
Dulac map, see (6)), at cubic saddle of this generality. as T →∞. In particular, the functions ξ and ω are regularly varying of order −β2 and −β0
in T , respectively. That is: limT →∞
ξ(η,cT )
ξ(η,T ) = cβ2 for every c > 0 and analogous for
ω(η, T ). Moreover, the Dulac map D : W u(0, η) →W s(ζ0, 0) itself has the form (as
ξ →0) ω = D(ξ) = ω0(η)ξ0(η)−β0
β2 ξ
β0
β2
1 + O(ξ
1
2β2 )
. (6) (6) With assumptions (5) and c2
1 < 4c0c2 in place, we can use the change of coordinates
¯x = √a0x, ¯y = √b2y and ¯γ = a1/√a0b2 ∈(−4, 4) to transform (1) into the
one-parameter family ⎛
⎝
˙¯x
˙¯y
˙¯x
⎞
⎠=
⎛
⎝
¯x(¯x2 + ¯γ ¯x ¯y + 3 ¯y2)
−¯y(3¯x2 + ¯γ ¯x ¯y + ¯y2))
1 + ¯w(¯x, ¯y)
⎞
⎠+ O(4),
(7) (7) for some transformed function ¯w. 1.1 Main results The crucial estimates here are of the Dulac times, i.e., the times that orbits take to pass
from an “incoming” unstable transversal to an “outgoing” unstable transversal to the
flow, see Fig.1. Theorem 1.1 Consider a C3 vector field of local form (3) with parameters satisfying
(2) and (4). Define (2) and (4). Define β0 := a0 + b0
2a0
,
β2 := a2 + b2
2b2
. Then there constants1 ξ0(η), ω0(η) such that the following asymptotics hold: Then there constants1 ξ0(η), ω0(η) such that the following asymptotics hold: ξ(η, ˜T ) = ξ0(η) ˜T −β2(1 + O( ˜T −1
2 )) and and ω(η, ˜T ) = ω0(η) ˜T −β0(1 + O( ˜T −1
2 )). for some transformed function ¯w. Because of this genericity and reduced number of technicalities that Lebesgue
measure gives as opposed to the SRB-measure, we state our statistical result for volume 123 1007 On volume preserving almost Anosov flows Fig. 2 The first quadrant Q of the rectangle P0, with stable and unstable foliations drawn vertically and
horizontally, respectively Fig. 2 The first quadrant Q of the rectangle P0, with stable and unstable foliations drawn vertically and
horizontally, respectively preserving flows. Theorem 1.1 is used to estimate the measures of the strips {ϕ = n},
see Fig.2, which in turn, together with the spectral properties of an induced Poincaré
map ˆf , are crucial ingredients for the analysis required to establish the following
stochastic limit properties of the flow φt. preserving flows. Theorem 1.1 is used to estimate the measures of the strips {ϕ = n},
see Fig.2, which in turn, together with the spectral properties of an induced Poincaré
map ˆf , are crucial ingredients for the analysis required to establish the following
stochastic limit properties of the flow φt. Theorem 1.2 Consider a volume preserving almost Anosov flow (7) on M with ¯γ ∈
(−4, 4) and an observables v : M →R that is C1 on M \ and has the form
v = v0+o(ρ)where
τ
0 v0◦φt dt ishomogeneousoforderρ > −2inlocalcoordinates
(x, y) near p and o(ρ) stands for terms of order > ρ. 1. If ρ ∈(0, ∞), then v satisfies the Central Limit Theorem, i.e., 1.2 Set-up The set-up here is largely taken over from [3]. Our phase space will be the
3-dimensional compact manifold M. Definition 1.1 [6, Definition 1] A diffeomorphism f : T2 →T2 is called almost
Anosov if there exists two continuous families of non-trivial cones x →Cu
x , Cs
x such
that except for a finite set S, (i) DfxCu
x ⊆Cu
f (x) and DfxCs
x ⊇Cs
f (x);
(ii) |Df v| > |v| for any 0 ̸= v ∈Cu and |Df v| < |v| for any 0 ̸= v ∈Cs (i) DfxCu
x ⊆Cu
f (x) and DfxCs
x ⊇Cs
f (x); (i) DfxCu
x ⊆Cu
f (x) and DfxCs
x ⊇Cs
f (x); f ( )
f ( )
(ii) |Dfxv| > |v| for any 0 ̸= v ∈Cu
x and |Dfxv| < |v| for any 0 ̸= v ∈Cs
x. For x ∈S, Dfx is the identity. 3 A flow f t on 3-torus T3 is called almost Anosov flow if it has a finite set S of neutral
periodic orbits, but everywhere else observes the condition of an Anosov flow in that
there is a continuous splitting of the tangent bundle into a stable, an unstable and a
neutral (flow) direction. For x ∈S, the derivative at the return time τ is Df τ
x is the
identity. The time-1 map f of the flow φt of (1) has the form of a skew-product f
⎛
⎝
x
y
z
⎞
⎠=
⎛
⎝
x(1 + a0x2 + a1xy + a2y2)
y(1 −b0x2 −b1xy −b2y2)
z + O(|w(x, y)|)
⎞
⎠+ O(4),
(8) (8) (8) see [2, Sect.2.1]. Restricted to the (x, y)-coordinates, this restriction fhor of f to
the first two coordinates is a smooth almost Anosov map with a single neutral fixed
point p = (0, 0). Let {Pi}k
i=0 be the Markov partition for fhor (which we can assume
to exist since fhor is a local perturbation of a Anosov diffeomorphism on T2). We
assume that p belongs to the interior of P0. Clearly, the x- and y-axis are the unstable
and stable manifolds of p respectively. We assume that the Markov partition element P0 ⊂U is a small rectangle such that f −1
hor(P0) ∪P0 ∪fhor(P0) ⊂U.
0
3. If ρ ∈(−2, 0) then v satisfies a Stable Law of order
4
2−ρ ∈(1, 2). Theorem 1.1 also allows us to derive other limit theorems such as in the infinite
measure setting of [2], but with mixed terms. Since we restrict to flows preserving
Lebesgue measure (not just an SRB-measure), we don’t give any further details here. 1. If ρ ∈(0, ∞), then v satisfies the Central Limit Theorem, i.e., t
0 v ◦φs ds −t
v dV ol
σ√t
⇒dist N(0, 1)
as t →∞, where the variance σ 2 > 0 unless
τ
0 v ◦φt dt is a coboundary.
0
2. If ρ = 0, then v satisfies the Central Limit Theorem with non-standard scaling
√t log t, i.e., t
0 v ◦φs ds −t
v dV ol
σ√t log t
⇒dist N(0, 1)
as t →∞, 1008 H. Bruin H. Bruin and the variance σ 2 > 0 unless
τ
0 v ◦φt dt is a coboundary. 1.2 Set-up Due to the
symmetries (x, y) →(±x, ±y), it suffices to do the analysis only in the first quadrant
Q = [0, ζ0] × [0, η0] of P0, see Fig.2. Without loss of generality (see [2, Lemma
2.1]) we can think of [0, ζ0]×{η0} as a local unstable leaf and {ζ0}×[0, η0] as a local
stable leaf of the global diffeomorphism. f ϕ
hor : Y →Y for Y := T2 \ P0, where We consider an induced map Fhor = f ϕ
hor : Y →Y for Y := T2 \ P0, where We consider an induced map Fhor = f ϕ
hor : Y →Y for Y := T2 \ P0, where ϕ(z) = min{n ≥1 : f n
hor(z) /∈P0} is the first return time to Y. Note that Fhor is invertible because fhor is. In the first
quadrant of U \ P0, the sets {ϕ = n} := {z ∈f −1(Q) \ Q : ϕ(z) = n}, n ≥2, is the first return time to Y. Note that Fhor is invertible because fhor is. In the first
quadrant of U \ P0, the sets {ϕ = n} := {z ∈f −1(Q) \ Q : ϕ(z) = n}, n ≥2, 12 On volume preserving almost Anosov flows 1009 are vertical strips (see Fig.2) adjacent to the local unstable leaf [0, ζ0] × {η0}, and
converging to {0} × [η0, η1] as n →∞. The images Fhor({ϕ = n}) are horizontal
strips, adjacent to the local stable leaf {ζ0} × [0, η0], and converging to [ζ0, ζ1] × {0}
as n →∞. are vertical strips (see Fig.2) adjacent to the local unstable leaf [0, ζ0] × {η0}, and
converging to {0} × [η0, η1] as n →∞. The images Fhor({ϕ = n}) are horizontal
strips, adjacent to the local stable leaf {ζ0} × [0, η0], and converging to [ζ0, ζ1] × {0}
as n →∞. Incontrastto fhor,theinducedmap Fhor isuniformlyhyperbolic,butonlypiecewise
continuous. Indeed, continuity fails at the boundaries of the strips {ϕ = n}, n ≥2 (and
F is undefined on W s(p)), but these boundaries are local stable and unstable leaves,
and it is possible to create a countable Markov partition refining {Pi}k
i=1 of Y for F,
in which all the strips {ϕ = n} are partition elements. 2 Regular variation of (' > n) with mixed terms In this section, we allow quadratic mixed terms in (3), but for the moment leave out
the O(4)-terms. That is, we consider ˙x = x(a0x2 + a1xy + a2y2),
˙y = −y(b0x2 + b1xy + b2y2),
(9) (9) with the restrictions (2) and (4). The condition c2
1 < 4c0c2 avoids the formation of
invariant lines y = px, but in the below proofs it is used to guarantee that expressions
as c0 + c1M + c2M2 for M = y/x are positive. Our exposition closely follows [2],
but since the mixed terms require adjustments throughout the proof, we will give it in
full. Set ci := ai + bi, i = 0, 1, 2. The conditions a2
1 < 4a0a2, b2
1 < 4b0b2 imply
that a2
1b2
1 −16a0a2b0b2 ≤0 ≤4(a0b0 −a2b2)2, and thus a2
1b2
1 ≤4(a0b2 + a2b0)2. Since a0, b0, a2, b2 ≥0, we also have a1b1 ≤2(a0b2 + a2b0), which implies that
c2
1 ≤4c0c2. Let u, v ∈R be the solutions of the linear equations (u + 2)a0 = vb0
(v + 2)b2 = ua2
that is:
⎧
⎨
⎩
u = 2b2c0
,
v = 2a0c2
. (10) (10) Note that u, v and (recall ̸= 0) all have the same sign and (4) implies that
b1
a1 = u+1
v+1. Compute that Note that u, v and (recall ̸= 0) all have the same sign and (4) implies that
b1
a1 = u+1
v+1. Compute that β0 := a0 + b0
2a0
= u + v + 2
2v
,
β2 := a2 + b2
2b2
= u + v + 2
2u
,
β0
β2
= u
v = b2c0
a0c2
,
(11) (11) and note that β0, β2 > 1
2 (or = 1
2 if we allow b0 = 0 or a2 = 0 respectively). Under
the extra assumption (5) we obtain β0 = β2 = 2 and u = v = 1. Th fit
ti
t
i
b
t th D l
f (3) and note that β0, β2 > 1
2 (or = 1
2 if we allow b0 = 0 or a2 = 0 respectively). Under
the extra assumption (5) we obtain β0 = β2 = 2 and u = v = 1. The first estimates is about the Dulac map of (3). 2 Regular variation of (' > n) with mixed terms (13) (13) 123 123 On volume preserving almost Anosov flows 1011 In terms M = y/x, we have In terms M = y/x, we have = y/x, we have x(M)2 = ξ
2u
u+v+2 η
2v
u+v+2 M−
2v
u+v+2
c0ξ2 + c1ξη + c2η2
c0 + c1M + c2M2
2
u+v+2
= ξ
1
β2 η
1
β0 M−1
β0
c0ξ2 + c1ξη + c2η2
c0 + c1M + c2M2
1−1
2β0 −1
2β2
. (14) (14) Proof We carry out the proof for > 0, so L(x, y) = xu yv( a0
v x2 +
a1
v+1xy + b2
u y2)
as in Lemma 2.1. The case < 0 goes likewise. 2
b Proof We carry out the proof for > 0, so L(x, y) = xu yv( a0
v x2 +
a1
v+1xy + b2
u y2)
as in Lemma 2.1. The case < 0 goes likewise. 2
b g
We solve for x from L(x, y) = L(δ, δ) = δu+v+2( a0
v +
a1
v+1 + b2
u ): We solve for x from L(x, y) = L(δ, δ) = δu+v+2( a0
v +
a1
v+1 + b2
u ): x = x(y) = δ
u+v+2
u
y−v+2
u
1 +
a1u
b1(v + 1) + ua0
vb2
1
u
1 +
a1u
b1(v + 1)
x
y + c0
c2
x2
y2
−1
u
= U(y)δ1+ a2
b2 y−a2
b2 , giving the required expression with U(y) as in (13). Note that limy→δ U(y) = 1 (i.e.,
the limit as (x, y) approaches the diagonal), and U(y) is differentiable. giving the required expression with U(y) as in (13). Note that limy→δ U(y) = 1 (i.e.,
the limit as (x, y) approaches the diagonal), and U(y) is differentiable. For the other expression, we use that L(x, y) = L(ξ, η) = ξuηv( a0
v ξ2 +
a1
v+1ξη +
b2
u η2), and solve for x2: ξuηv
a0
v ξ2 +
a1
v + 1ξη + b2
u η2
= xu yv
a0
v x2 +
a1
v + 1xy + b2
u y2
= xu+v+2Mv
a0
v +
a1
v + 1 M + b2
u M2
. 2 Regular variation of (' > n) with mixed terms The first estimates is about the Dulac map of (3). 12 1010 H. Bruin Proposition 2.1 Consider a vector field on the 2-torus with local form (3) for
a0, a2, b0, b2 ≥0 and ̸= 0. There are functions ξ0(η), ω0(η), ξ1(η), ω1(η) > 0
independent of T (with exact expressions given in the proof) such that ξ(η, T ) = ξ0(η)T −β2
1 −ξ1(η)T −1 + O(T −2, T −2β2)
and and ω(η, T ) = ω0(η)T −β0
1 −ω1(η)T −1 + O(T −2, T −2β0)
. Lemma 2.1 The function L(x, y) =
⎧
⎨
⎩
xu yv( a0
v x2 +
a1
v+1xy + b2
u y2)
if > 0,
x−u y−v( a0
v x2 +
a1
v+1xy b2
u y2)−1
if < 0,
(12) (12) is a first integral of (9). is a first integral of (9). Proof of Lemma 2.1 First assume > 0, so u, v > 0 as well. By (10), we can write
L(x, y) as L(x, y) = xu yv
b0
u + 2 x2 +
a1
v + 1 xy + b2
u y2
= xu yv
a0
v x2 +
b1
u + 1 xy +
a2
v + 2 y2
. Using these two equivalent expressions we compute the Lie derivative directly Using these two equivalent expressions we compute the Lie derivative directly ˙L = ⟨∇L, X⟩
= xu−1yv
b0
u + 2(u + 2)x2 +
a1
v + 1xy + b2
u uxu−1y2
x(a0x2 + a1xy + a2y2)
−xu yv−1
a0
v x2v +
a1
v + 1xy +
a2
v + 2(v + 2)y2
y(b0x2 + b1xy + b2y2)
= 0.
= 0. = 0. Any function of a first integral is a first integral; in particular this holds for 1/L. Therefore the conclusion is immediate for < 0 too. Corollary 2.1 Every (x(y), y) on the integral curve from (ξ, η) to (δ, δ) satisfies x(y) = U(y)δ1+ a2
b2 η−a2
b2 , for U(y) :=
c2 +
a1uc2
b2(v + 1) + c0
1
u
c2 +
a1uc2
b2(v + 1)
x
y + c0
x2
y2
−1
u
. 2 Regular variation of (' > n) with mixed terms Here we used Here we used ξuηv
a0
v ξ2 +
a1
v + 1ξη + b2
u η2
= ξuηv
a0
2a0c2
ξ2 +
a1
2a0c2 + ξη + b2
2b2c0
η2
=
2c0c2
c0ξ2 +
2a1c0c2
2a0c2 + ξη + c2η2
=
2c0c2
c0ξ2 + c1ξη + c2η2
,
(15) (15) (where the last step follows from (4)) and a similar computation for the term with x, y. Use (10) and (11) to obtain ⎧
⎨
⎩
a0
v +
a1
v+1 M + b2
u M2 =
2c0c2 (c0 + c1M + c2M2),
a0ξ2
v
+
a1
v+1ξη + b2η2
u
=
2c0c2 (c0ξ2 + c1ξη + c2η2). 123 1012 H. Bruin H. Bruin This gives This gives x2 = ξ
2u
u+v+2 η
2v
u+v+2 M−
2v
u+v+2
c0ξ2 + c1ξη + c2η2
c0 + c1M + c2M2
2
u+v+2
= ξ
1
β2 η
1
β0 M−1
β0
c0ξ2 + c1ξη + c2η2
c0 + c1M + c2M2
1−1
2β0 −1
2β2
, where we recall that β0 =
u+v+2
2v
and β2 =
u+v+2
2u
from (11), which also gives
2
u+v+2 = 1 −
1
2β0 −
1
2β2 . ⊓⊔ Proof of Proposition 2.1 We carry out the proof for > 0, so L(x, y) = xu yv( a0
v x2+
a1
v+1xy + b2
u y2) as in Lemma 2.1. The case < 0 goes likewise. Fix η such that
(ξ(η, T ), η) ∈φ−1(Q) \ Q. We use the variable M = y/x, so y = Mx and differen-
tiating gives ˙y = ˙Mx + M ˙x. Recalling that ci = ai + bi and inserting the values for
˙x and ˙y from (9), we get ˙M = −M(c0 + c1M + c2M2)x2. (16) (16) Combined with (14), this gives Combined with (14), this gives ˙M = −G(ξ, η)M1−1
β0
c0 + c1M + c2M2 1
2β0 + 1
2β2
(17) (17) with with G(ξ, η) := ξ
1
β2 η
1
β0
c0ξ2 + c1ξη + c2η21−1
2β0 −1
2β2 . (18) (18) For the exit time T ≥0, recall that ξ(η, T ) and ω(η, T ) are such that the solution
of (3) satisfies (x(0), y(0)) = (ξ(η, T ), η) and (x(T ), y(T )) = (ζ0, ω(η, T )). 2 Regular variation of (' > n) with mixed terms Since G(ξ(η, T ), η)T = g(η, T )
1
β2 η
1
β0 (c0g(η, T )2T −2β2 + c1g(η, T )T −β2 + c2η2)1−1
2β0 −1
2β2 , and 1 −
1
2β0 −
1
2β2 > 0, we find that g(η, T ) converges2: and 1 −
1
2β0 −
1
2β2 > 0, we find that g(η, T ) converges2: ξ0(η) := lim
T →∞g(η, T ) = c
−1
u
2
η−a2
b2
⎛
⎝
∞
0
M
1
β0 −1 dM
c0 + c1M + c2M2 1
2β0 + 1
2β2
⎞
⎠
β2
,
(21) (21) here we have used −β2(1 −
1
2β0 −
1
2β2 ) = −
2β2
u+v+2 = −1
u for the exponent of c2,
nd 2
u + β2
β0 = v+2
u
= a2
b2 for the exponent of η. where we have used −β2(1 −
1
2β0 −
1
2β2 ) = −
2β2
u+v+2 = −1
u for the exponent of c2,
and 2
u + β2
β0 = v+2
u
= a2
b2 for the exponent of η. u
β0
u
b2
We continue the proof to get higher asymptotics. Differentiating (19) w.r.t. T gives −
η
1
β0 ξ
1
β2 −1ξ′
(c0ξ2 + c1ξη + c2η2)
1
2β0 + 1
2β2
−
ζ
1
β2
0 ω
1
β0 −1ω′
(c0ζ 2
0 + c1ζ0ω + c2ω2)
1
2β0 + 1
2β2
= ∂G(ξ, η)
∂ξ
T ξ′ + G(ξ, η),
(22 −
η
1
β0 ξ
1
β2 −1ξ′
(c0ξ2 + c1ξη + c2η2)
1
2β0 + 1
2β2
−
ζ
1
β2
0 ω
1
β0 −1ω′
(c0ζ 2
0 + c1ζ0ω + c2ω2)
1
2β0 + 1
2β2
∂G(ξ
) (22) where (by differentiating (18)) where (by differentiating (18)) ∂G(ξ, η)
∂ξ
=
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
ξ
1
β2 −1η
1
β0
c0ξ2 + c1ξη + c2η2−1
2β0 −1
2β2 . 2 Regular variation of (' > n) with mixed terms This
implies M(0) = η/ξ(η, T ) and M(T ) = ω(η, T )/ζ0. Inserting this in (17), separating
variables, and integrating we get η/ξ(η,T )
ω(η,T )/ζ0
M
1
β0 −1 dM
c0 + c1M + c2M2 1
2β0 + 1
2β2
= G(ξ(η, T ), η)T . (19) (19) In the rest of the proof, we will frequently suppress the dependence on η and T
in ξ(η, T ) and ω(η, T ). We know that L(ξ(η, T ), η) = ξuηv( a0
v ξ2 + b2
u η2) =
ζ u
0 ωv( a0
v ζ 2
0 + b2
u ω2) = L(η, ω(η, T )), which gives ξuηv(c0ξ2 + c1ξη + c2η2) = ζ u
0 ωv(c0ζ 2
0 + c1ζ0ω + c2ω2). (20) (20) From their definition, ξ(η, T ) and ω(η, T ) are clearly decreasing in T , so their T -
derivatives ξ′(η, T ), ω′(η, T ) ≤0. Since c0, c2 > 0 (otherwise = 0), the integrand 123 12 On volume preserving almost Anosov flows 1013 of (19) is O(M
1
β0 −1) as M →0 and O(M−1
β2 −1) as M →∞. Hence the integral is
increasing and bounded in T . But this means that G(ξ(η, T ), η)T is increasing in T
and bounded as well. Let g(η, T ) = ξ(η, T )T β2. Since of (19) is O(M
1
β0 −1) as M →0 and O(M−1
β2 −1) as M →∞. Hence the integral is
increasing and bounded in T . But this means that G(ξ(η, T ), η)T is increasing in T
and bounded as well. Let g(η, T ) = ξ(η, T )T β2. Since (
)
(
)
(
)
g
increasing and bounded in T . But this means that G(ξ(η, T ), η)T is increasing in
and bounded as well. Let g(η, T ) = ξ(η, T )T β2. 2 For the symmetric statement on ω(η, T ), define ˆg(η, T ) = ω(η, T )T β0. Then limT →∞ˆg(η, T )
limT →∞g(η, T )β0/β2η1+2/vζ −b0/a0
0
( c2
c0 )1/v. 2 Regular variation of (' > n) with mixed terms Combined with (18), (20) and (22), this gives Combined with (18), (20) and (22), this gives −η
1
β0 ξ′ −ζ
1
β2
0
ζ u
0 ωv
ξuηv
1
2β0 + 1
2β2 ω
1
β0 −1
ξ
1
β2 −1 ω′
=
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
T η
1
β0 ξ′ + η
1
β0 ξ
c0ξ2 + c1ξη + c2η2
. (23) (23) ause
1
2β0 +
1
2β2 −1 = −
2
u+v+2, using (11) and dividing by η
1
β0 , we can simplify
to Because
1
2β0 +
1
2β2 −1 = −
2
u+v+2, using (11) and dividing by η
1
β0 , we can simplify
(23) t Because 2 For the symmetric statement on ω(η, T ), define ˆg(η, T ) = ω(η, T )T β0. Then limT →∞ˆg(η, T ) =
limT →∞g(η, T )β0/β2η1+2/vζ −b0/a0
0
( c2
c0 )1/v. 123 1014 H. Bruin H. Bruin −ξ′ −ζ u
0
ηv
ωv−1
ξu−1 ω′ =
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
T ξ′
2
2 −ξ′ −ζ u
0
ηv
ωv−1
ξu−1 ω′ =
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
T ξ′
+ ξ(c0ξ2 + c1ξη + c2η2). + ξ(c0ξ2 + c1ξη + c2η2). (24) (24) Taking the derivative of (20) w.r.t. T and multiplying with /(c0c2) gives
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
ηvξu−1ξ′
=
2a0ζ 2
0 + +c1
b0
c0
+ a2
c2
ζ0ω + 2a2ω2
ζ u
0 ωv−1ω′. Hence, we can rewrite (24) as Hence, we can rewrite (24) as Hence, we can rewrite (24) as −
⎛
⎝1 +
2b0ξ2 + c1
b0
c0 + a2
c2
ξη + 2b2η2
2a0ζ 2
0 + c1
b0
c0 + a2
c2
ζ0ω + 2a2ω2
⎞
⎠ξ′
=
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + b2η2
T ξ′ + ξ(c0ξ2 + c1ξη + c2η2). 2 Regular variation of (' > n) with mixed terms We insert ξ′ = g′(T )T −β2 −β2g(T )T −(1+β2) and multiply with T β2, which leads to −
⎛
⎝1 +
2b0ξ2 + c1
b0
c0 + a2
c2
ξη + 2b2η2
2a0ζ 2
0 + c1
b0
c0 + a2
c2
ζ0ω + 2a2ω2
⎞
⎠(g′(T ) −β2g(T )T −1)
=
2b0ξ2 + c1
b0
c0
+ a2
c2
ξη + 2b2η2
g′(T ) T −
b2
g(T)3T −2β2. Since ξ = O(T −β2) and ω = O(T −β0), we can write this differential equation as g′
g = 1
T 2
β2
2
a0ζ 2
0 + b2η2 + O(T −2β2)
a0ζ 2
0 + O(T −2β0)
−
b2 g(T )2T −a2
b2
b2η2 + O(T −2β2) + O(T −1)
. Keeping the leading terms only (where we use that 2β2, 2β0 > 1), we get the
differential equation Keeping the leading terms only (where we use that 2β2, 2β0 > 1), we get the
differential equation g′
g =
ξ1(η) + O
max{T −1, T −a2
b2 }
1
T 2
for ξ1(η) := β2
2
1
a0ζ 2
0
+
1
b2η2
. Using the limit boundary value ξ0 = ξ0(η) = limT →∞g(η, T ), we find the solution g(η, T ) = ξ0e
−
ξ1+O
max{T −1,T
−a2
b2 }
T −1
= ξ0
1 −ξ1T −1 + O
max{T −2, T −2β2}
123 On volume preserving almost Anosov flows 1015 as required. The analogous asymptotics for ω and the constants ω0 and ω1 can be
derived by changing the time direction and the roles (a0, a2) ↔(b2, b0), and also by
the relation ξuηv+2c2 ∼ζ u+2
0
ωvc0 from (20): ω0(η) = c−1
v
0
ζ
−b0
a0
0
⎛
⎜⎝
∞
0
M
1
β2 −1 dM
c0M2 + c1M + c2
1
2β0 + 1
2β2
⎞
⎟⎠
β0
,
ω1(η) = β0
2
1
b2ζ 2
0
+
1
a0η2
. This concludes the proof. 3 Proof of Theorem 1.1 To prove that the regular variation established in Proposition 2.1 is robust under pertur-
bations of the vector field, we put the O(4) terms back into (3), but since we consider
it as a perturbation of (9), we write ˜X instead: ˜X =
˜X1
˜X2
=
x(a0x2 + a1xy + a2y2)
−y(b0x2 + b1xy + b2y2)
+ O(4),
(25) (25) so that | ˜X −X| = O(4). The quantities ξ(η, T ), ω(η, T ) will be written as
˜ξ(η, T ), ˜ω(η, T ) etc., and the goal is to show that ˜ξ(η, T ) is still regularly varying. As before, let ξ = ξ(η, T ) be such that for the unperturbed flow, φT (ξ, η) =
(ζ0, ω(η, T )). Proposition 2.1 gives the asymptotics of ξ(η, T ) as T →∞. At the
same time, under the perturbed flow associated to (25), φ ˜T (ξ, η) = (ζ0, ˜ω(η, ˜T ))
for some ˜T . Therefore we can write ξ(η, T ) = ˜ξ(η, ˜T ), and once we estimated ˜T
as function of T , we can express ˜ξ(η, ˜T ) explicitly as function of ˜T . We follow the
argument of the proof of Proposition 2.1, keeping track of the effect of the higher order
terms. The perturbed first integral To start, we construct a first integral ˜L on Q = [0, ζ0] ×
[0, η0] by defining ˜L( ˜φt(δ, δ)) = L(δ, δ) =
⎧
⎨
⎩
δu+v+2 ( a0
v +
a1
v+1 + b2
u )
if > 0,
δ−(u+v+2) ( a0
v +
a1
v+1 + b2
u )−1
if < 0, for 0 < δ ≤min{ζ0, η0} and t ∈R. (We continue the argument for the case > 0;
the other case goes analogously.)
˜
˜
˜
3 By construction, ˜L is constant on integral curves of ˙z = ˜X(z). Because ˜X is C3,
the integral curves are C3 curves, and form a C3 foliation of P0, see e.g. [14, Theorem
2.10]. Note that the coordinate axes consist of the stationary point (0, 0) and its stable
and unstable manifold; we put ˜L(x, 0) = ˜L(0, y) = 0. Then ˜L is continuous on Q
and C3 on the interior of Q. 12 3 H. Bruin 1016 Fig. 3 Proof of Theorem 1.1 3 Solutions of (27) and (28), starting from the same point z0 = (x0, y0) Fig. 3 Solutions of (27) and (28), starting from the same point z0 = (x0, y0) Now we compare ˜L with L on a small neighbourhood U of φ−1
hor(Q)∪Q∪φ1
hor(Q). Take y0 = η0 and x0 = x0(δ) such that the integral curve of ˙z = X(z) through
z0 := (x0, y0) intersects the diagonal at (δ, δ). Then the integral curve of ˙z = ˜X(z)
through z0 intersects the diagonal at (˜δ, ˜δ) for some ˜δ = ˜δ(δ), see Fig.3. For some z1 on this integral curve between z0 and (˜δ, ˜δ), the integral curve through
z1 of the unperturbed system intersects the diagonal in some point (δ′, δ′) between
(δ, δ) and (˜δ, ˜δ). Therefore ˜L(z1) = ˜L(˜δ, ˜δ) = L(˜δ, ˜δ) = L(δ′, δ′)
˜δ
δ′
u+v+2
= L(z1)
˜δ
δ′
u+v+2
. (26 (26) Estimating |˜δ −δ|: Parametrise the integral curve of X through z0 as (x(y), y) for
min{δ, ˜δ} ≤y ≤y0. (So x ≤y; the case y ≤x can be dealt with by switching the
roles of x and y.) Then by (9): Estimating |˜δ −δ|: Parametrise the integral curve of X through z0 as (x(y), y) for
min{δ, ˜δ} ≤y ≤y0. (So x ≤y; the case y ≤x can be dealt with by switching the
roles of x and y.) Then by (9): x′(y) = F(x, y) := −x(a0x2 + a1xy + a2y2)
y(b0x2 + b1xy + b2y2). (27) (27) For the perturbed vector field (25) we parametrise the integral curve through z0 as
(˜x(y), y) and we have the analogue of (27): ˜x′(y) = ˜F(˜x, y) := −
˜x(a0 ˜x2 + a1 ˜x y + a2y2 + 3
j=1 ˆa j ˜x j y3−j)
y(b0 ˜x2 + b1 ˜x y + b2y2 + 3
j=1 ˆb j ˜x j y3−j)
,
(28) (28) where ˆa j, ˆb j are bounded functions in ˜x and y. The absence of the terms ˆa0y3 and
ˆb0y3 is because of the assumption that O(4) = x2O(2) near the yz-plane. For the
region ˜x > y we need to leave out the terms ˆa3 ˜x3 and ˆb3 ˜x3 instead. where ˆa j, ˆb j are bounded functions in ˜x and y. 3 Proof of Theorem 1.1 The absence of the terms ˆa0y3 and
ˆb0y3 is because of the assumption that O(4) = x2O(2) near the yz-plane. For the
region ˜x > y we need to leave out the terms ˆa3 ˜x3 and ˆb3 ˜x3 instead. 123 volume preserving almost Anosov flows
1017 On volume preserving almost Anosov flows 1017 Combining (27) and (28) we have Combining (27) and (28) we have Combining (27) and (28) we have ˜x′(y) = −˜x(a0 ˜x2 + a1 ˜x y + a2y2)
y(b0 ˜x2 + b1 ˜x y + b2y2)(1 + ˜x q0(˜x, y))
(29) (29) for the bounded function q0 : [0, max{δ, ˜δ}] × [δ, y0] →R, given by q0(˜x, y) = 1
˜x
⎛
⎝
1 +
˜x 3
j=1 ˆa j ˜x j−1y3−j
a0 ˜x2 + a1 ˜x y + a2y2
1 +
˜x 3
j=1 ˆb j ˜x j−1y3−j
b0 ˜x2 + b1 ˜x y + b2y2
−1
−1
⎞
⎠. From Grönwall’s Lemma, we can bound the speed at which ˜x(y) and x(y) diverge
from each other, see [14, Theorem 2.8]: From Grönwall’s Lemma, we can bound the speed at which ˜x(y) and x(y) diverge
from each other, see [14, Theorem 2.8]: u(y) := |˜x(y) −x(y)| ≤|˜x(y0) −x(y0)|e(y0−y) Lip + V
Lip
e(y0−y) Lip −1
, (30) (30) for y ∈[δ, y0], where Lip is the Lipschitz constant of F(x, y) in x maximized over
y ∈[δ, y0] and for y ∈[δ, y0], where Lip is the Lipschitz constant of F(x, y) in x maximized over
y ∈[δ, y0] and V =
sup
y∈[δ,y0]
sup
x∈[0,max{δ,˜δ}]
|F(x, y)|q0(x, y)x. V =
sup
y∈[δ,y0]
sup
x∈[0,max{δ,˜δ}]
|F(x, y)|q0(x, y)x. The Lipschitz constant of F is supx,y
∂
∂x F(x, y), which is of order 1/δ. Therefore (30)
gives a very poor estimate if we apply it at once to the whole interval [δ, y0], but we
can improve it by partitioning [δ, y0] into smaller intervals. Proposition 3.1 Assume the equation ˜x′(y) = F(˜x, y)(1 + q0 ˜x) for some bounded
function q0 = q0(˜x, y) and initial value ˜x(y0) = x(y0) = x0. Then there is a constant
K for all y ∈[δ, y0] we have |˜x(y) −x(y)| ≤Kδ2. The estimate (30) applied to [y j+1, y j] gives the recursive formula The estimate (30) applied to [y j+1, y j] gives the recursive formula u(y j+1) ≤u(y j)e
(γ +ε)
y j −y j+1
y j+1
+ Cδ2(1+γ )y−(1+2γ )
j+1
(y j −y j+1). (31) u(y j+1) ≤u(y j)e
(γ +ε)
y j −y j+1
y j+1
+ Cδ2(1+γ )y−(1+2γ )
j+1
(y j −y j+1). (31)
Therefore, for 1 ≤M < N, (31) u(yM) ≤Cδ2(1+γ )
M
j=1
y−(1+2γ )
j
(y j−1 −y j)e(γ +ε) M
k= j
yk−1−yk
yk
. (32) (32) Then (32) gives for N = ⌈δ−1⌉and M < N, Then (32) gives for N = ⌈δ−1⌉and M < N, Then (32) gives for N = ⌈δ−1⌉and M < N, Then (32) gives for N = ⌈δ−1⌉and M < N, u(yM) ≤Cδ2(1+γ )y−2γ
0
N 2γ
M
j=1
e(γ +ε) M
k= j 1/(N−k)
(N −j)1+2γ
≤Cδ2y−2γ
0
1
(N −M)γ +ε
M
j=1
1
(N −j)γ −ε+1 ≤
C
γ −ε δ2y−2γ
0
. ≤Cδ2y−2γ
0
1
(N −M)γ +ε
M
j=1
1
(N −j)γ −ε+1 ≤
C
γ −ε δ2y−2γ
0
. Therefore |˜x(yM) −x(yM)| = u(yM) ≤Kδ2 as claimed. Therefore |˜x(yM) −x(yM)| = u(yM) ≤Kδ2 as claimed. ⊓⊔ Therefore |˜x(yM) −x(yM)| = u(yM) ≤Kδ2 as claimed. ⊓⊔ By (29) for y = δ = ˜x(δ) + O(δ2), the derivative ˜x′(δ) = a0+a1+a2
b0+b1+b2 + O(δ). Since
˜δ lies between δ and ˜x(δ) (see Fig.3), we have by a Taylor approximation |˜x(δ) −δ| = |˜x(δ) −˜δ| + |˜δ −δ| =
1 + a0 + a1 + a2
b0 + b1 + b2
+ O(δ)
|˜δ −δ|
= c0 + c1 + c2 + O(δ)
b0 + b1 + b2
|˜δ −δ|. (33) (33) and therefore |˜δ−δ| = O(|˜x(δ)−δ|) = O(δ2). Now that we have this relation between
˜δ and δ, we can estimate ˜T in terms of T , and effectively finish of Theorem 1.1. and therefore |˜δ−δ| = O(|˜x(δ)−δ|) = O(δ2). Now that we have this relation between
˜δ and δ, we can estimate ˜T in terms of T , and effectively finish of Theorem 1.1. 3 Proof of Theorem 1.1 Proof We divide the interval [δ, y0] into N interval [y j+1 −y j] of the same length,
so y j = δ + y0−δ
N (N −j) and set x j = x(y j). The Lipschitz constant [y j+1, y j] is
supy∈[y j+1,y j] supx∈[0,δ2]
∂
∂x F(x, y) ≤1
y (γ + ε) for γ = a2
b2 . Therefore Lip(y j)(y j −y j+1) ≤y0 −δ
N
γ + ε
δ + y0−δ
N
(N −j −1)
=
γ + ε
(N −( j + 1)) + δN/(y0 −δ)
Recall from (13) that x(y) = U(y)δ1+ a2
b2 y−a2
b2 . Let Lip(y j)(y j −y j+1) ≤y0 −δ
N
γ + ε
δ + y0−δ
N
(N −j −1)
=
γ + ε
(N −( j + 1)) + δN/(y0 −δ) Recall from (13) that x(y) = U(y)δ1+ a2
b2 y−a2
b2 . Let Recall from (13) that x(y) = U(y)δ1+ a2
b2 y−a2
b2 . Let C = 2
sup
y∈[y j+1,y j]
sup
x∈[0,2x(y)]
q0(x, y) a0x2 + a1xy + a2y2
b0x2 + b1xy + b2y2 U(y)2. Take K = 2C
γ y−2γ
0
and assume by induction that u(y j) ≤Kδ2 ≤1 (so we assume
that δ is sufficiently small). Clearly this induction hypothesis holds for j = 0 because
u(y0) = 0. Then H. Bruin 1018 H. Bruin V (y j) :=
sup
y∈[y j+1,y j]
sup
x∈[0,x(y)(1+Kδ2)]
q0(x, y) x F(x, y)
≤sup
x,y
a0x2 + a1xy + a2y2
b0x2 + b1xy + b2y2 q0(x, y)
sup
y∈[y j+1,y j]
x(y)2(1 + Kδ2)2
y
≤sup
x,y
a0x2 + a1xy + a2y2
b0x2 + b1xy + b2y2 q0(x, y) 2U(y)2
sup
y∈[y j+1,y j]
δ2(1+γ )y−(1+2γ )
= Cδ2(1+γ )y−(1+2γ )
j+1
. y [y j+1,y j] x∈[0,x(y)(1+Kδ )]
≤sup
x,y
a0x2 + a1xy + a2y2
b0x2 + b1xy + b2y2 q0(x, y)
sup
y∈[y j+1,y j]
x(y)2(1 + Kδ2)2
y ≤sup
x,y
y
y
b0x2 + b1xy + b2y2 q0(x, y)
sup
y∈[y j+1,y j]
y
y
≤sup
x,y
a0x2 + a1xy + a2y2
b0x2 + b1xy + b2y2 q0(x, y) 2U(y)2
sup
y∈[y j+1,y j]
δ2(1+γ )y−(1+2γ ) j
j
= Cδ2(1+γ )y−(1+2γ )
j+1
. The estimate (30) applied to [y j+1, y j] gives the recursive formula Proof of Theorem 1.1 Let z0 = (x0, y0) = (ξ(η, T ), η) = (˜ξ(η, ˜T ), η) as in Fig.3
be the point such that φT
hor(z0) = (ζ0, ω(η, T )) under the unperturbed flow and
˜φ ˜T
hor(z0) = (ζ0, ˜ω(η, ˜T )) under the perturbed flow. We estimate ˜T in terms of T . ˜φ ˜T
hor(z0) = (ζ0, ˜ω(η, ˜T )) under the perturbed flow. We estimate ˜T in terms of T . 123 123 On volume preserving almost Anosov flows 1019 Combining the estimate for ξ(η, T ) from Proposition 2.1 with L(δ, δ)
=
L(ξ(η, T ), η), we can find the relation between δ and T : δ = δ0T −1
2
1 −ξ1
2β2
T −1 + O(T −2, T −2β2)
,
(34) (34) for for δ0 = ξ
1
2β2
0
η1−1
2β2
⎛
⎝c0
ξ2
η2 + c1
ξ
η + c2
c0 + c1 + c2
⎞
⎠
1
u+v+2
. To estimate ˜T , we divide the trajectory ˜φt(z0) = (˜x(t), ˜y(t)) of z0 for the region
˜x ≤˜y into two parts delimited by points in time: To estimate ˜T , we divide the trajectory ˜φt(z0) = (˜x(t), ˜y(t)) of z0 for the region
˜x ≤˜y into two parts delimited by points in time: ˜T1 = min{t > 0 : 2˜x(t) = ˜y(t)},
˜T2 = min{t > 0 : ˜x( ˜T1 + t) = ˜y( ˜T1 + t)},
(35) (35) and symmetric quantities for the region ˜y ≤˜x. Let T1, T2 be the analogous quantities
for the unperturbed trajectory. For y1 := y(T1) and ˜y1 := y( ˜T1), we have | ˜y1 −y1| =
O(δ2). Assume that ˜y1 ≥y1 (the case ˜y1 < y1 goes likewise). We compute and symmetric quantities for the region ˜y ≤˜x. Let T1, T2 be the analogous quantities
for the unperturbed trajectory. For y1 := y(T1) and ˜y1 := y( ˜T1), we have | ˜y1 −y1| =
O(δ2). Assume that ˜y1 ≥y1 (the case ˜y1 < y1 goes likewise). We compute T1 =
y0
y1
dy
˙y =
y0
˜y1
dy
˙y +
˜y1
y1
dy
˙y
=
y0
˜y1
dy
−y(b0x(y)2 + b1x(y)y + b2y2) +
˜y1
y1
dy
˙y . The estimate (30) applied to [y j+1, y j] gives the recursive formula The first term is O(δ−2) as δ →0 and the second is O(1) (and therefore small
compared to the first term) because | ˜y1−y1| = O(δ2). Hence T1 = O(δ−2). Similarly, The first term is O(δ−2) as δ →0 and the second is O(1) (and therefore small
compared to the first term) because | ˜y1−y1| = O(δ2). Hence T1 = O(δ−2). Similarly, ˜T1 −T1 =
y0
˜y1
dy
˙˜y
−
y0
˜y1
dy
˙y −
˜y1
y1
dy
˙y =
y0
˜y1
1
˙˜y
−1
˙y dy + O(1). (36) (36) Using that ˜x(y) −x(y) ≤δ2 and abbreviating = 3
j=1 ˆb j ˜x j y3−j = O(y3) for
˜x ≤˜y, we compute the last integral of (36) as y0
˜y1
1
−y(b0 ˜x(y)2 + b1 ˜x(y)y + b2y2 + ) −
1
−y(b0x(y)2 + b1x(y)y + b2y2) dy
=
y0
˜y1
(˜x −x)(b0(˜x + x) + b1y) +
y(b0 ˜x(y)2 + b1 ˜x(y)y + b2y2 + )(b0x(y)2 + b1x(y)y + b2y2) dy
≤δ2
y0
˜y1
3b0 + 2b1
maxi bi y2 dy +
y0
˜y1
O(y−1) dy = O(δ−1). 123 1020 H. Bruin H. Bruin Together with (34) and the fact that
˜y1
y1
dy
˙y = O(1), this gives ˜T1 = T1(1 + O(δ)) = T1(1 + O(T −1
2 )). (37) (37) For ˜M = y/˜x, computations analogous to (16) show that ˙˜M = −˜M(c0 + c1 ˜M +
c2 ˜M2+ ˜x)˜x2 for ˆc j = ˆa j + ˆb j and := 2
j=0 ˆc3−j ˜M j. For every (˜x, y) = (˜x, ˜x ˜M)
on the ˜φ-trajectory of z0 (which is a level set of ˜L), we have ξuηv
a0
v ξ2 +
a1
v + 1ξη + b2
u η2
˜δ
δ′ = ˜xu+v+2 ˜Mv
a0
v +
a1
v + 1
˜M + b2
u
˜M2
, with δ′ as in (26). This gives the analogue of [2, formula (32)]: ˙˜M = −G(ξ, η) ˜M1−1
β0
c0 + c1 ˜M + c2 ˜M2 + ˜x(˜x, ˜M)
1
2β0 + 1
2β2
˜δ
δ′
1−1
2β0 −1
2β2
,
(38) (38) where G(ξ, η) is as in (18). The estimate (30) applied to [y j+1, y j] gives the recursive formula Because δ′ lies between δ and ˜δ, the conclusion |˜δ −δ| =
O(δ2) of (33) gives ˜δ
δ′ ≤1 + |˜δ −δ′|
δ′
≤1 + |˜δ −δ|
δ
= 1 + O(δ). Note also on the time interval [ ˜T1, ˜T1+ ˜T2], we have ˜x(y) ≤y ≤2˜x(y) ≤2 max{δ, ˜δ}
and therefore c0 + c1 ˜M + c2 ˜M2 + ˜x(˜x, ˜M) = c0(1 + O(˜x)) + c1 ˜M(1 + O(y)) + c2 ˜M2
= (c0 + c1 ˜M + c2 ˜M2)(1 + O(δ)). This combined with (38) we find This combined with (38) we find ˙˜M = −G(ξ, η) ˜M1−1
β0
c0 + c1 ˜M + c2 ˜M2 1
2β0 + 1
2β2 (1 + O(δ)). The choice of ˜T1 and T1 ensures that ˜M( ˜T1) = M(T1) = 2 and similarly ˜M( ˜T1+ ˜T2) =
M(T1 + T2) = 1. Therefore The choice of ˜T1 and T1 ensures that ˜M( ˜T1) = M(T1) = 2 and similarly ˜M( ˜T1+ ˜T2) =
M(T1 + T2) = 1. Therefore ˜T2 =
˜T1+ ˜T2
˜T1
dt =
˜M( ˜T1)
˜M( ˜T1+ ˜T2)
˙˜M−1 d ˜M
=
2
1
˜M
1
β0 −1(1 + O(δ))−1
G(ξ, η)(c0 + c1 ˜M + c2 ˜M2)
1
2β0 + 1
2β2
d ˜M
=
M(T1)
M(T1+T2)
M
1
β0 −1(1 + O(δ))−1
G(ξ, η)(c0 + c1M + c2M2)
1
2β0 + 1
2β2
dM 123 1021 On volume preserving almost Anosov flows = T2(1 + O(δ))−1 = T2(1 + O(T −1
2 )). = T2(1 + O(δ))−1 = T2(1 + O(T −1
2 )). = T2(1 + O(δ))−1 = T2(1 + O(T −1
2 )). Combining this with (37) gives ˜T = T (1+ O(T −1
2 )). The estimate of Proposition 2.1
now gives ˜ξ(η, ˜T ) = ξ0(η) ˜T −β2(1 + O( ˜T −1
2 )) as claimed. Combining this with (37) gives ˜T = T (1+ O(T −1
2 )). The estimate of Proposition 2.1
now gives ˜ξ(η, ˜T ) = ξ0(η) ˜T −β2(1 + O( ˜T −1
2 )) as claimed. g
ξ(η,
)
ξ0(η)
(
(
))
Reversing the roles (a0, a2) ↔(b2, b0) as in the end of the proof of Proposition 2.1
gives ˜ω(η, ˜T ) = ω0(η) ˜T −β0(1 + O( ˜T −1
2 )). The estimate (30) applied to [y j+1, y j] gives the recursive formula The formula (6) for the Dulac maps follows directly from Theorem 1.1 by inverting
T →ξ(η, T ) and inserting this in the formula for ω(η, T ). In the special case that
β0 = β2, formula (6) reduces to ω = D(ξ) =
b2
a0
1
β2−1 η
ζ0
2β2−1
ξ
1 + O(ξ
1
2β2 )
. Reducing further by assuming (5) (i.e., in the volume preserving setting), we get ω = D(ξ) = b2
a0
η
ζ0
3
ξ
1 + O(ξ
1
4 )
. This coefficient b2
a0
η
ζ0
3
=
∥Xhor(0,η)∥
∥Xhor(ζ0,0)∥agrees with the fact that for ω = D(ξ),
the flow-boxes ∪t∈[0,ε]φt
hor([0, ξ] × {η}) and ∪t∈[0,ε]φt
hor({ζ0} × [0, ω]) must have
the same volume. If the neutral saddle p is part of a heteroclinic cycle, then it is
accumulated by periodic solutions, but these are not limit cycles of course. 4 Time-1 map versus Poincaré map (40 (40) (40) For M →0, the leading term in the integrand is For M →0, the leading term in the integrand is For M →0, the leading term in the integrand is c
−1+(1−ρ
2 ) 1
β0
0
M
1
β0 −1−ρ
v
u+v+2 = c
−1+(1−ρ
2 ) 1
β0
0
M(1−ρ
2 ) 1
β0 −1, i.e., the exponent is > −1 for ρ < 2. For M →∞, the leading term in the integrand
is i.e., the exponent is > −1 for ρ < 2. For M →∞, the leading term in the integrand
is c
−1+(1−ρ
2 ) 1
β0
2
M−1
β2 −1−ρ(
v
u+v+2 +
2
u+v+2 −1) = c
−1+(1−ρ
2 ) 1
β0
0
M−(1−ρ
2 ) 1
β2 −1, i.e., the exponent is < −1 for ρ < 2. This means that the integral in (40) converges
to some constant C0 = C0(ρ) as T →∞, and ∼C0G(T )
ρ
2 −1 ∼CT 1−ρ
2 for
C = C0
c
1−1
2β0 −1
2β2
2
ξ
1
β0
0 η
1−1
β2
0
1−ρ
2
. This finishes the proof for ρ < 2. If ρ > 2, then the value of based on the leading terms of the integrand only, is i.e., the exponent is < −1 for ρ < 2. This means that the integral in (40) converges
to some constant C0 = C0(ρ) as T →∞, and ∼C0G(T )
ρ
2 −1 ∼CT 1−ρ
2 for
1
1
1
1 1−ρ C = C0
c
1−1
2β0 −1
2β2
2
ξ
1
β0
0 η
1−1
β2
0
1−ρ
2
. This fi C = C0
c
1−1
2β0 −1
2β2
2
ξ
1
β0
0 η
1−1
β2
0
1−ρ
2
. This finishes the proof for ρ <
If ρ > 2, then the value of based on the leading terms of the integrand only, is
If ρ > 2, then the value of based on the leading terms of the integrand only, is T )
ρ
2 −1 c
−1+(1−ρ
2 ) 1
β0
0
ρ
2 −1
β2
η0
ξ(T )
−(1−ρ
2 ) 1
β2 −β0
ω(T )
ζ0
(1−ρ
2 ) 1
β0
. 4 Time-1 map versus Poincaré map First we give an estimate of observables integrated over the flow-lines of Xhor of (9) Proposition 4.1 Let r =
x2 + y2, ρ > 0 and W(T ) be the integral curve for (9)
connecting (ξ(η0, T ), η0)) to (ζ0, ω(η0, T )), see Fig.1. Then there is a constant C =
C(ρ) > 0 such that Proposition 4.1 Let r =
x2 + y2, ρ > 0 and W(T ) be the integral curve for (9)
connecting (ξ(η0, T ), η0)) to (ζ0, ω(η0, T )), see Fig.1. Then there is a constant C =
C(ρ) > 0 such that = (T ) :=
W(T )
r(t)ρ dt =
⎧
⎪⎨
⎪⎩
CT 1−ρ
2 (1 + o(1))
if ρ < 2,
C log(T )(1 + o(1))
if ρ = 2,
C(1 + o(1))
if ρ > 2. (39) (39) Proof We build on the proof of Proposition 2.1 (or in fact Theorem 1.1), and in
the integral we change coordinates M = y/x. That is, rρ = (x2 + y2)ρ/2 =
xρ(1 + M2)ρ/2. Use (14) to get Proof We build on the proof of Proposition 2.1 (or in fact Theorem 1.1), and in
the integral we change coordinates M = y/x. That is, rρ = (x2 + y2)ρ/2 =
xρ(1 + M2)ρ/2. Use (14) to get x = G(T )
1
2 M−
v
u+v+2 (c0 + c1M + c2M2)
−1
u+v+2 , 123 123 1022 H. Bruin H. Bruin with G(T ) := G(ξ(η0, T )), η0) as in (18). Abbreviate ξ(η0, T ) = ξ(T ) and
ω(η0, T ) = ω(T ). Inserting the above in the integral of (19), we obtain with G(T ) := G(ξ(η0, T )), η0) as in (18). Abbreviate ξ(η0, T ) = ξ(T ) and
ω(η0, T ) = ω(T ). Inserting the above in the integral of (19), we obtain = G(T )
ρ
2 −1
η/ξ(T )
ω(T )/ζ0
M−
ρv
u+v+2 (c0 + c1M + c2M2)
−ρ
u+v+2 (1 + M2)
ρ
2
M1−1
β0
c0 + c1M + c2M2 1
2β0 + 1
2β2
dM. 4 Time-1 map versus Poincaré map Insert the values of ξ(T ) and ω(T ) from Proposition 2.1 as well as the leading term
of G(T ): =
c
1−1
2β0 −1
2β2
2
ξ
1
β0
0 η
1−1
β2
0
1−ρ
2
T 1−ρ
2 c
−1+(1−ρ
2 ) 1
β0
0
ρ
2 −1
β2η0
( ρ
2 −1) 1
β2 −β0ζ
( ρ
2 −1) 1
β0
0
T
ρ
2 −1. The powers of T cancel in this expression, proving the case ρ > 2. Finally, if ρ = 2,
then the factor G(T )
ρ
2 −1 in (40) disappears and the leading terms in the integrand
(both as M →0 and M →∞), are c−1
0 M−1. This gives, due to Proposition 2.1, ∼1
c0
log
η0
ξ(T ) −log ω(T )
ζ0
∼β2 + β0
c0
log T . ⊓⊔ ⊓⊔ 123 123 1023 On volume preserving almost Anosov flows The 3-dimensional time-1 map φ1 preserves no 2-dimensional submanifold of M. Yet in order to model φt as a suspension flow over a 2-dimensional map, we need a
genuine Poincaré map. For this we choose a section transversal to and containing
a neighbourhood U of p. As an example, could be T2 × {0}, and the Poincaré map
to T2 ×{0} could be (a local perturbation of) Arnol’d’s cat map; in this case (and most
cases) M is not homeomorphic to T3 because the homology is more complicated, see
[1, 11]. Let h : →R+, h(q) = min{t > 0 : φt(q) ∈} be the first return time. Assuming that sup |w(x, y)| < 1, the first return time h is bounded and bounded
away from zero, say 0 < inf h < sup h.
The Poincaré map f := φh : → has a neutral saddle point p at the origin. Its local stable/unstable manifolds are W s
loc(p) = {0} × (−ε, ε) and W u
loc(p) =
(−ε, ε) × {0}. Because the flow φt is a perturbation of an Anosov flow, and f is a
Poincaré map, it has a finite Markov partition {Pi}i≥0 and we can assume that p is in
the interior of P0. 4 Time-1 map versus Poincaré map In the sequel, let U be a neighbourhood of p that is small enough
that (1) is valid on U × [0, 1] but also that f (U) ⊃ˆP0 ∪P0. f
In order to regain the hyperbolicity lacking in f , let In order to regain the hyperbolicity lacking in f , let r(q) := min{n ≥1 : f n(q) ∈Y}
(41) (41) be the first return time to Y := \ P0. Then the Poincaré map F = f r = φτ of φt
to Y × {0} is hyperbolic, where τ(q) = min{t > 0 : φt((q, 0)) ∈Y × {0}} =
r−1
j=0
h ◦f j is the corresponding first return time. Consequently, the flow φt : M × R →M can be modeled as a suspension flow
on Y τ =
q∈Y {q} × [0, τ(q))
/(q, τ(q)) ∼(F(q), 0). Since the flow and section
Y × {0} are C1 smooth, τ is C1 on each piece {r = k}. Lemma 4.1 Let ˆτ(q) = min{t > 0 : φt
hor((q, η)) ∈W s(ζ0, 0)} be the time the
horizontal flow needs to reach ∂Q. Then τ(q) = ˆτ(q) + O(1) and r = ˆτ(q) +
(ˆτ(q)) + O(1), where is as in Proposition 4.1. Proof By the definition of ˆτ we have φ ˆτ
hor(q) ∈ˆW s. Therefore it takes a bounded
amount of time (positive or negative) for φ ˆτ(q, 0) to hit Y × {0}, so |τ(q) −ˆτ(q)| =
O(1). Proof By the definition of ˆτ we have φ ˆτ
hor(q) ∈ˆW s. Therefore it takes a bounded
amount of time (positive or negative) for φ ˆτ(q, 0) to hit Y × {0}, so |τ(q) −ˆτ(q)| =
O(1). The function w in the vertical component of the local vector field X of (1) is a linear
combination of homogeneous functions Ci ·(x2+y2)ρi/2. It suffices to take the leading
term, i.e., the one with the smallest ρi =: ρ. Using this term in (39), we obtain that
ˆτ(q)+(ˆτ(q)) indicates the vertical displacement by the flow φt. In particular, it gives
the number of times the flow-line intersects , and hence r = ˆτ(q)+(ˆτ(q))+O(1). ⊓⊔ 3 12 H. Bruin 1024 Fig. 4 The first quadrant of the rectangle P0, with stable and unstable foliations of Poincaré map f = φh
drawn vertically and horizontally, respectively. 4 Time-1 map versus Poincaré map Also one of the integral curves is drawn Fig. 4 The first quadrant of the rectangle P0, with stable and unstable foliations of Poincaré map f = φh
drawn vertically and horizontally, respectively. Also one of the integral curves is drawn Assume that φt and ˆf preserve Lebesgue measure. Assume that φt and ˆf preserve Lebesgue measure. Proposition 4.2 Recall that β2 = a2+b2
2b2
∈( 1
2, ∞). There exists C∗> 0 such that Proposition 4.2 Recall that β2 = a2+b2
2b2
∈( 1
2, ∞). There exists C∗> 0 such that Leb({τ > t}) = C∗t−β2(1 + o(1))
(42) (42) for the F-invariant SRB-measure μ ¯φ. Proof The function τ is defined on \P0 and τ ≥h2 = h + h ◦f on Y{r≥2} :=
f −1(P0)\P0. The set Y{r≥2} is a rectangle with boundaries consisting of two stable
and two unstable leaves of the Poincaré map f . Let W u(y) denote the unstable leaf of
f inside Y{r≥2} with (0, y) as (left) boundary point. Let y1 < y2 be such that W u(y1)
and W u(y2) are the unstable boundary leaves of Y{r≥2}. ˆ The unstable foliation of ˆf = φ1
hor does not entirely coincide with the unstable
foliation of f . Let ˆW u(y) denote the unstable leaf of ˆf with (0, y) as (left) boundary
point. Both ˆW u(y) and W u(y) are C1 curves emanating from (0, y); let γ (y) denote
the angle between them. Then the lengths Leb(W u(y) ∩{τ > t}) = | cos γ (y)| Leb( ˆW u(y) ∩{τ > t})(1 + o(1))
= | cos γ (y)| ξ0(y) t−β2(1 + o(1)) 123 123 On volume preserving almost Anosov flows 1025 as t →∞, where the last equality and the notation ξ0(y) and β2 = (a2 + b2)/(2b2)
come from Theorem 1.1 We decompose Lebesgue on Y{r≥2} as We decompose Lebesgue on Y{r≥2} as
Y{r≥2}
v dμ ¯φ =
y2
y1
W u(y)
v dμs
W u(y)
dνu(y). The conditional measures μW u(y) on W u(y) equals 1-dimensional Lebesgue mW u(y)
on W u(y) Therefore, as t →∞, The conditional measures μW u(y) on W u(y) equals 1-dimensional Lebesgue mW u(y)
on W u(y) Therefore, as t →∞, μ ¯φ(τ > t) =
y2
y1
μW u(y)(W u(y) ∩{τ > t}) dνu(y)
=
y2
y1
mW u(y)(W u(y) ∩{τ > t}) dνu(y)
=
y2
y1
| cos γ (y)| m ˆW u(y)( ˆW u(y) ∩{τ > t})(1 + o(1)) dνu(y)
=
y2
y1
| cos γ (y)| ξ0(y) t−β2(1 + o(1)) dνu(y) = C∗t−β2(1 + o(1)) for C∗=
y2
y1 | cos γ (y)| ξ0(y) dνu(y). This proves the result. ⊓⊔ for C∗=
y2
y1 | cos γ (y)| ξ0(y) dνu(y). This proves the result. ⊓⊔ Funding Open access funding provided by Austrian Science Fund (FWF). OpenAccess ThisarticleislicensedunderaCreativeCommonsAttribution4.0InternationalLicense,which
permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence,
and indicate if changes were made. The images or other third party material in this article are included
in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If
material is not included in the article’s Creative Commons licence and your intended use is not permitted
by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the
copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. 5 The proof of Theorem 1.2 Proof The proof of Theorem 1.2 is a direct application of Theorem 2.7 in [3], where
¯v =
τ
0 v ◦φt dt takes the role of ¯ψ in [3, Theorem 2.7], but the condition that
¯ψ = C −ψ0 for some positive ψ0 is only important for the results on the shape of the
pressure function in [3]. For us, only the tail of ¯v matters and since v is C1 on M\, ¯v
is C1 on each partition element {φ = n} of the Markov map F. Since Proposition 4.1
applies to v we get ¯v(x, y) ∼C pT 1−ρ
2 if the Dulac time of (x, y) is T . Since our
invariant measure is Lebesgue, and β2 = 2, Theorem 1.1 can be immediately used to
estimate Leb(¯v > t) ∼
η1
η0 ξ0(η) dη
Vol ( \ P0)
t
C p
−4
2−ρ
,
(43) (43) where is the Poincaré section and Vol ( \ P0) is the normalizing constant for
Lebesgue restricted to the domain \ P0 of F. If ρ ≥2, this asymptotic formula
should be interpreted as Leb(¯v > t) = 0 for t large, that is: ¯v is bounded. where is the Poincaré section and Vol ( \ P0) is the normalizing constant for
Lebesgue restricted to the domain \ P0 of F. If ρ ≥2, this asymptotic formula
should be interpreted as Leb(¯v > t) = 0 for t large, that is: ¯v is bounded. The exponent of the tail (43) is −2 if and only if ρ = 0, and in this case [3, Theorem
2.7(a)(ii)] gives the non-Gaussian CLT. g
If −2 < ρ < 0, [3, Theorem 2.7(a)(i)] gives a Stable Law of order 4/(2 −ρ) ∈
(1, 2). 123 12 1026 H. Bruin H. Bruin Finally, if 0 < ρ < 2 (or ρ ≥2 when ¯v is bounded), then we obtain the CLT
provided the variance σ 2 > 0, and this follows from ¯v not being a coboundary. In
other words, ¯v ̸= h −h ◦F for any h ∈B, the Banach space used in the proofs of [3],
and this we assumed explicitly. ⊓⊔ ⊓⊔ Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps
and institutional affiliations. References 1. Barbot, T., Fenley, S.: Pseudo-Anosov flows in toroidal manifolds. Geom. Topol. 17, 1877–1954 (201 1. Barbot, T., Fenley, S.: Pseudo-Anosov flows in toroidal manifolds. Geom. Topol. 17, 1877–1954 (2013)
2. Bruin, H., Terhesiu, D.: Regular variation and rates of mixing for infinite measure preserving almost
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and institutional affiliations. 123 12
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A Dynamic Reconfigurable Architecture for Hybrid Spiking and Convolutional FPGA-Based Neural Network Designs
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Journal of low power electronics and applications
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Irmak, Hasan; Corradi, Federico; Detterer, Paul; Alachiotis, Nikolaos;
Ziener, Daniel
A dynamic reconfigurable architecture for hybrid spiking and convolutional
FPGA-based neural network designs Irmak, Hasan; Corradi, Federico; Detterer, Paul; Alachiotis, Nikolaos;
Ziener, Daniel Irmak, Hasan; Corradi, Federico; Detterer, Paul; Alachiotis, Nikolaos;
Ziener, Daniel
Keywords: FPGA; NN; CNN; SNN; partial reconfiguration Keywords: FPGA; NN; CNN; SNN; partial reconfiguration Received: 10 July 2021
Accepted: 10 August 2021
Published: 17 August 2021 A dynamic reconfigurable architecture for hybrid spiking and convolutional
FPGA-based neural network designs Original published in:
Journal of Low Power Electronics and Applications. - Basel : MDPI. - 11
(2021), 3, art. 32, 25 pp. Original published:
2021-08-17
ISSN:
2079-9268
DOI:
10.3390/jlpea11030032
[Visited:
2022-03-03] TU Ilmenau | Universitätsbibliothek | ilmedia, 2022
http://www.tu-ilmenau.de/ilmedia
Original published in:
Journal of Low Power Electronics and Applications. - Basel : MDPI. - 11
(2021), 3, art. 32, 25 pp. Original published:
2021-08-17
ISSN:
2079-9268
DOI:
10.3390/jlpea11030032
[Visited:
2022-03-03]
This work is licensed under a Creative Commons Attribution 4.0
International license. To view a copy of this license, visit
https://creativecommons.org/licenses/by/4.0/ Original published in:
Journal of Low Power Electronics and Applications. - Basel : MDPI. - 11
(2021), 3, art. 32, 25 pp. Original published:
2021-08-17
ISSN:
2079-9268
DOI:
10.3390/jlpea11030032
[Visited:
2022-03-03] This work is licensed under a Creative Commons Attribution 4.0
International license. To view a copy of this license, visit
https://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0
International license. To view a copy of this license, visit
https://creativecommons.org/licenses/by/4.0/ TU Ilmenau | Universitätsbibliothek | ilmedia, 2022
http://www.tu-ilmenau.de/ilmedia Journal of
Low Power Electronics
and Applications Journal of
Low Power Electronics
and Applications Journal of
Low Power Electronics
and Applications Article
A Dynamic Reconfigurable Architecture for Hybrid Spiking
and Convolutional FPGA-Based Neural Network Designs
Hasan Irmak 1,*
Federico Corradi 2
Paul Detterer 2
Nikolaos Alachiotis 1
and Daniel Ziener 3 mak 1,*
, Federico Corradi 2
, Paul Detterer 2
, Nikolaos Alachiotis 1
and Daniel Ziener 3 Hasan Irmak 1,*
, Federico Corradi 2
, Paul Detterer 2
, Nikolaos Alachiotis 1
and D Hasan Irmak 1,* 1
Computer Architecture for Embedded Systems, Faculty of EEMCS, University of Twente,
7522 NB Enschede, The Netherlands; n.alachiotis@utwente.nl 1
Computer Architecture for Embedded Systems, Faculty of EEMCS, University of Twente,
7522 NB Enschede, The Netherlands; n.alachiotis@utwente.nl
2
Ultra-Low-Power Systems for IoT, Stichting IMEC Nederland, 5656 AE Eindhoven, The Netherlands;
federico.corradi@imec.nl (F.C.); paul.detterer@imec.nl (P.D.)
3
Computer Architecture and Embedded Systems, Department of Computer Science and Automation,
Technische Universität Ilmenau, 98693 Ilmenau, Germany; daniel.ziener@tu-ilmenau.de
*
Correspondence: h.irmak@utwente.nl 2
Ultra-Low-Power Systems for IoT, Stichting IMEC Nederland, 5656 AE Eindhoven, The Nether
federico.corradi@imec.nl (F.C.); paul.detterer@imec.nl (P.D.) Abstract: This work presents a dynamically reconfigurable architecture for Neural Network (NN)
accelerators implemented in Field-Programmable Gate Array (FPGA) that can be applied in a variety
of application scenarios. Although the concept of Dynamic Partial Reconfiguration (DPR) is increas-
ingly used in NN accelerators, the throughput is usually lower than pure static designs. This work
presents a dynamically reconfigurable energy-efficient accelerator architecture that does not sacrifice
throughput performance. The proposed accelerator comprises reconfigurable processing engines and
dynamically utilizes the device resources according to model parameters. Using the proposed archi-
tecture with DPR, different NN types and architectures can be realized on the same FPGA. Moreover,
the proposed architecture maximizes throughput performance with design optimizations while con-
sidering the available resources on the hardware platform. We evaluate our design with different NN
architectures for two different tasks. The first task is the image classification of two distinct datasets,
and this requires switching between Convolutional Neural Network (CNN) architectures having
different layer structures. The second task requires switching between NN architectures, namely a
CNN architecture with high accuracy and throughput and a hybrid architecture that combines
convolutional layers and an optimized Spiking Neural Network (SNN) architecture. We demonstrate
throughput results from quickly reprogramming only a tiny part of the FPGA hardware using DPR. Experimental results show that the implemented designs achieve a 7× faster frame rate than current
FPGA accelerators while being extremely flexible and using comparable resources. Citation: Irmak, H.; Corradi, F.;
Detterer, P.; Alachiotis, N.; Ziener, D.
A Dynamic Reconfigurable
Architecture for Hybrid Spiking and
Convolutional FPGA-Based Neural
Network Designs. J. Low Power
Electron. Appl. 2021, 11, 32. https://
doi.org/10.3390/jlpea11030032 Keywords: FPGA; NN; CNN; SNN; partial reconfiguration 1. Introduction Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations. The application of artificial intelligence models at the edge requires novel software
and hardware architectures capable of executing many tasks in an energy-efficient man-
ner. Many robotics applications are battery-limited, such as autonomous vehicles, drones,
and robotics. It is mandatory to execute many applications on the same hardware plat-
form in the most efficient way [1,2]. State-of-the-art Central Processing Unit (CPU) per-
forms 10–100 FLOP per second with typical power efficiency in the order of 1 GOP/J [3]. In contrast, GPUs offer up to 10 TOP/s of peak performance and are more suitable for
high-performance neural network (NN) applications at the expense of many hundreds
of watts of power consumption [4]. Recently, Field-Programmable Gate-Arrays (FPGA)
have been proposed for neural network processing [5]. They are ideal for highly parallel
workloads and can best exploit the properties of neural network computation. For this
reason, a promising approach is to build special optimized hardware blocks that can
accelerate several models of machine learning and deep neural network workloads [6]. Since NNs are increasingly becoming application specialized (such as language processing, Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article
distributed
under
the
terms
and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/). https://www.mdpi.com/journal/jlpea J. Low Power Electron. Appl. 2021, 11, 32. https://doi.org/10.3390/jlpea11030032 J. Low Power Electron. Appl. 2021, 11, 32 2 of 25 voice translation, object detection, and tracking), Dynamic Partial Reconfiguration (DPR)
on FPGAs can help alleviate the burden of executing various models onto a single hard-
ware platform. Among the most promising models of artificial NNs, Convolutional Neural
Networks (CNN) and Spiking Neural Networks (SNN) can be employed in almost all tasks. CNN and SNN are complementary; CNNs are mostly used in high-throughput applications
(object detection, image classification, etc.), while SNNs are more bio-inspired models that
require memory distributed with the processing [7] and offer energy-efficient temporally
distributed computation. The energy cost for these types of SNNs implemented in FPGA
hardware is in the order of hundreds of pico joules per synaptic operation [8,9], and this
makes them attractive for massively parallel implementations in FPGAs. Background This section introduces the basic system design for the typical FPGA-based NN accel-
erator design structure. In this work, we only focus on accelerated and efficient inference,
which means using a pre-trained and optimized model to perform prediction, regression,
or classification tasks. The highly parallel architecture of NNs requires massively parallel
computing capacity. In general, a NN model can be seen as a directed graph. Each vertex
of the graph indicates information flow between layers; data from a previous layer is used
as input to the next layer. In standard CNNs models, the parameters of each layer are
the connection weights, the neuron biases, and the input and output of each layer are
activations. CNN’s neurons do not store any state as their activations are computed at each
input cycle. In SNNs models, the parameters of each layer are also connection weights. The neurons are biologically inspired models that are stateful and require parameters such
as integration time constant, membrane decay, threshold, and refractory period. In contrast
to CNN models, in SNNs, inputs and outputs of each layer are binary spikes that indicate
when the neuron membrane potential has crossed a threshold, a spike is emitted. In SNNs,
it is required to store the membrane potential for each neuron. Communication is binary
(only spikes), and computation is event-based. Each neuron is immediately evaluated upon
the arrival of an input spike, and this makes SNNs models hugely parallel as no global
signals need to be shared among neurons. g
g
Partial reconfiguration is the ability to reconfigure part of the FPGA after initial
configuration. On the other hand, DPR allows reconfiguring part of the FPGA while the rest
of the device is still running. Thus, DPR feature offers to overcome the resource and power
limits of the design by enabling run-time reconfiguration of the selected regions of the
FPGA. Efficient utilization of the potential of the FPGA results in energy-efficient designs
compared to static designs. A DPR FPGA design can be divided into two parts. The static
part is the area of the FPGA that is not reconfigured again after the power-up configuration. On the other hand, partial reconfiguration region is the dynamic part of the FPGA, and it
can be updated at run time using partial bitstreams. This region is called Pblock which is a
collection of cells and one or more rectangular areas. 1. Introduction One of the major
differences among CNNs and SNNs is the fact that a CNN layer is a computational block,
and information needs to be fully computed before starting the computation of the next
layer (i.e., the matrix multiplication needs to be carried fully). In contrast, SNNs are truly
parallel, and spiking activity from each neuron is immediately sent to other neurons, which
are executed without the need of waiting for the computation from the previous layer to be
completed. Fully parallel SNNs implementation offers a trade-off between accuracy and
latency [10]. Background As shown in Figure 1, the same Pblock
can be used with different partial bitstreams to realize different implementations using
the same hardware resources. Thus, the functionality of the FPGA dramatically increases. For uploading the partial bitstream to the FPGA, Internal Configuration Access Port
(ICAP) and Processor Configuration Access Port (PCAP) are offered by Xilinx FPGAs [11]. The latter one is provided to reconfigure FPGA from the processor side in Zynq FPGAs. 3 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 1. The Concept of Partial Reconfiguration. Figure 1. The Concept of Partial Reconfiguration. Typical applications that benefit from DPR are network data processing, digital signal
processing, cognitive radio, and systems on a reconfigurable chip [12]. DPR capability is
also investigated in NN accelerators from different perspectives. In [13], it is proposed
an approach for realizing the deeper neural networks in low-resource FPGAs. In this
approach, a confidence level is calculated according to the output of the CNN. If it is under
a threshold, then the depth of the CNN is incremented using DPR. In other words, there is
an adaptive feedback mechanism for the decision of the classification, and if the image
is not classified, new layers are added to the NN architecture with DPR. Another similar
work [14] splits the NN into chunks consisting of a few layers and executes each chunk
separately. After execution of each chunk, the next chunk is uploaded to the FPGA using
DPR and then processed. Thus, resource usage of the accelerator is highly minimized. Similarly, in [15], the same hardware resources are used for each convolution layer using
DPR and they are executed in a sequential manner. However, in these studies, the classifier
throughput is significantly affected by the DPR overhead. In addition, some studies [16],
particularly for accelerating Binarized NNs, explores DPR on disaggregated FPGA plat-
forms to improve the throughput performance by encapsulating complex sequences of
operations as instructions of variable length. Moreover, DPR can also be used in low-power
applications. Youssef et al. [17] apply DPR to adjust the power consumption according
to the power level of the battery. If the battery level is low, low power and low accu-
rate design (i.e., smaller bit sizes) are loaded to decrease the power consumption of the
FPGA. There are also other studies exploiting DPR for classification problems of different
datasets [18]. Background To do so, the convolutional filters are selected according to the class datasets
using DPR. However, to change the kernel filter size at run time, DPR can be inefficient
since there are also some works that reconfiguration of the convolutional filters can be
achieved at run time without using DPR [19]. Moreover, there are also very low-power
NN accelerators [20] without using DPR but they lack flexibility and are designed for
specific CNN designs. Previously, we proposed a flexible DPR architecture specifically for
CNN architectures with only digit and letter recognition [21]. This work builds upon and
extends our previous work [20,21] by proposing a flexible hardware architecture for NN
accelerators, and different NN architectures and also combinations of them (i.e., hybrid
architectures) can be realized using DPR without sacrificing throughput or accuracy of
the accelerator. Because we employ DPR for switching between different NN accelerators. Thus, NN performance is not affected by the DPR overhead. p
y
The main contributions of the paper are the followings: The main contributions of the paper are the followings: •
It is proposed a flexible hardware design and computer architecture that allows to
easily modify the NN accelerators (i.e., insert/delete/update the structure) at run
time using DPR. g
•
The proposed architecture can adapt to different networks and, therefore, also to
different applications using DPR without any degradation in accuracy or throughput. •
The proposed architecture consists of processing elements (PE). These PEs exploit
pipelining and other optimizations for better performance. To exemplify, for the
convolutional PE, it is proposed a novel method to perform feature extraction that J. Low Power Electron. Appl. 2021, 11, 32 4 of 25 allows to hide the computations of a pooling layer behind the computations of a
convolutional layer. For other PEs, further optimizations are applied as well. The rest of the paper is organized as follows: Section 2 presents the proposed accel-
erator architecture, different NN designs such as CNN and SNN architectures. Section 3
provides implementation details and an experimental evaluation. Section 4 concludes
this work. 2.1. Accelerator Architecture As shown from the proposed hardware architecture in Figure 4, every PE can be
interconnected to any other PE, processor, or I/O port of the FPGA using internal bus
switches, and these interconnections can be configured at run time. This allows to dynami-
cally update the NN architecture by adding or removing layers, as well as by replacing the
structure of the NN with another structure to realize different types of networks. In the
updated structure, the weights and biases can be updated using the memory-mapped
interface of the PEs. As a result, different NN types can be implemented on the same FPGA
resources using the proposed hardware architecture. Figure 3. NN PE Types. Figure 3. NN PE Types. The proposed accelerator architecture consists of multiple PEs and configurable
switches that allow different connections for each PE interface (i.e., data, memory-mapped,
I/O). As shown from the proposed hardware architecture in Figure 4, every PE can be
interconnected to any other PE, processor, or I/O port of the FPGA using internal bus
switches, and these interconnections can be configured at run time. This allows to dynami-
cally update the NN architecture by adding or removing layers, as well as by replacing the
structure of the NN with another structure to realize different types of networks. In the
updated structure, the weights and biases can be updated using the memory-mapped
interface of the PEs. As a result, different NN types can be implemented on the same FPGA
resources using the proposed hardware architecture. The proposed accelerator architecture consists of multiple PEs and configurable
switches that allow different connections for each PE interface (i.e., data, memory-mapped,
I/O). As shown from the proposed hardware architecture in Figure 4, every PE can be
interconnected to any other PE, processor, or I/O port of the FPGA using internal bus
switches, and these interconnections can be configured at run time. This allows to dynami-
cally update the NN architecture by adding or removing layers, as well as by replacing the
structure of the NN with another structure to realize different types of networks. In the
updated structure, the weights and biases can be updated using the memory-mapped
interface of the PEs. As a result, different NN types can be implemented on the same FPGA
resources using the proposed hardware architecture. Figure 4. Proposed hardware architecture with multiple PEs. Figure 4. Proposed hardware architecture with multiple PEs. 2.1. Accelerator Architecture The essential processing element in the proposed accelerator architecture is referred
to as the PE. PE is a high-level generic block with predefined interfaces. Firstly, it has a
high-speed data interface (i.e., AXI4-Stream) used for the connection of different PEs to
exchange data. Secondly, an AXI4 interface is used as a memory-mapped interface to write
and read from the internal memory and registers of the PEs. Thus, the processor can update
the memory or registers of the PEs using this interface. Lastly, for I/O operations, a GPIO
interface is employed. The interfaces of the PE can be seen in Figure 2. In addition, each PE
has DPR capability to implement different functions using the same hardware resources. Figure 2. Interfaces of the Processing Element. Figure 2. Interfaces of the Processing Element. PEs are the building blocks of the hardware architecture. They implement the layers
of the NN accelerator. In this study, three different PE types are developed as shown in
Figure 3. Using these PEs, different NN accelerators can be realized. Firstly, a feedforward
PE is developed to implement Deep NNs (DNNs). In addition, to realize CNNs, a convo-
lution PE with optional integrated pooling is also designed. This is designed to optimize
performance by integrating the pooling layer with the convolution layer. CNN is a cascaded
connection of convolutional PEs and feedforward PEs. Lastly, for SNN implementations,
a spiking PE is developed. Some of the parameters of the PEs can be set before synthesis
and the others can be changed at run time. The data interface of the PEs is 64-bit wide. The optimum bit sizes for parameters of the PEs are determined based on the accuracy drop,
as compared with the accuracy achieved with single-precision floating-point arithmetic. Convolution and Feedforward PEs have 8-bit weights, and 16-bit activations and Spiking
PE has 6-bit weights, and 10-bit activations, with an accuracy drop of less than 0.1% in
comparison with the single-precision floating-point implementation. The hardware design
and optimizations of these PEs will be explained in the following sub-sections. 5 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 3. NN PE Types. The proposed accelerator architecture consists of multiple PEs and configurable
switches that allow different connections for each PE interface (i.e., data, memory-mapped,
I/O). 2.1. Accelerator Architecture In the proposed architecture, the hard/soft processor in the FPGA controls all the
connections of the PEs using the memory-mapped interface of the switches. In addition,
the stream switch supports the back pressure property and any PE can stop the input data
coming from another PE if it is not able to process the new data. In other words, it can
inform the previous PE to stop producing output by sending a busy signal. This means
that each PE can adjust the data transfer rate and it can also be run at different frequencies. Besides, if any PE needs more clock cycles for execution, there will be a chance to run it at
higher frequencies to preserve the overall throughput. 2.2. Convolution with Optional Integrated Pooling PE The convolution operation is the major operation for convolutional layers in CNN. In this operation, a two-dimensional filter (i.e., a kernel) is applied to the input to create a
feature map that shows the presence of detected features. The main idea of the convolution
operation in CNN is the ability to automatically learn a large number of filters in parallel
which are designed to detect specific features anywhere on the input images. A typical
convolution operation is shown in Figure 5. 6 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 5. Convolution Operation in CNNs. f
h
l
f
l d
h
f h Figure 5. Convolution Operation in CNNs. Figure 5. Convolution Operation in CNNs. After the convolution operation, an activation function is applied to the output of the
convolutional layer in order to learn the complex features in the image by adding non-
linearity into NN. Thus, a NN can easily solve non-linear complex classification problems. Moreover, the non-linearity of the activation functions allows back-propagation by having a
derivative function related to the inputs. Commonly used activation functions are Rectified
Linear Unit (ReLU), tanh, and sigmoid [22]. After the convolution operation, an activation function is applied to the output of the
convolutional layer in order to learn the complex features in the image by adding non-
linearity into NN. Thus, a NN can easily solve non-linear complex classification problems. Moreover, the non-linearity of the activation functions allows back-propagation by having a
derivative function related to the inputs. Commonly used activation functions are Rectified
Linear Unit (ReLU), tanh, and sigmoid [22]. g
In almost every CNN architecture, in order to decrease the data size and performance
sensitivity to the location of the features, pooling layers can be used after the convolutional
layer. These layers have no parameters to learn and only downsample the image according
to the pooling type. There are two common pooling types: average pooling and max-
pooling. Average pooling calculates the average of the patches in the feature map. Thus,
it smooths the image and it may not identify the sharp features. On the other hand, max-
pooling selects the highest value in the patch and helps to extract the edges and low-level
features in the feature maps. The most popular CNNs such as LeNet, Alexnet, and VGGNet use the pooling layer
after the convolution operation to decrease the data size [23–25]. 2.2. Convolution with Optional Integrated Pooling PE In this study, we are
proposing a less complex method for the computation of convolution layers followed by
max-pooling layers by hiding the computations of the pooling layers into the computa-
tion of convolution layers. Normally, the output pixel of a convolution layer with input
image in() and activation function f() followed by a max-pooling layer with 2 × 2 kernel,
and downsampling by 2 is given by Equation (1). out( i
2, j
2) = max( f (Conv(in(i, j)), f (Conv(in(i, j + 1)),
f (Conv(in(i + 1, j))), f (Conv(in(i + 1, j + 1))))
(1) (1) In Equation (1), if f() is a monotonically non-decreasing function, like ReLU, tanh,
or sigmoid, applying maximum pooling operation before the activation does not change
the result. Therefore, the activation and max pooling operations are interchangeable. Thus,
the order of applying the activation operation can be replaced with the pooling operation
and can be written as in Equation (2). J. Low Power Electron. Appl. 2021, 11, 32 7 of 25 out( i
2, j
2) = f (max(Conv(in(i, j)), Conv(in(i, j + 1)),
Conv(in(i + 1, j)), Conv(in(i + 1, j + 1))))
(2) out( i
2, j
2) = f (max(Conv(in(i, j)), Conv(in(i, j + 1)),
Conv(in(i + 1, j)), Conv(in(i + 1, j + 1))))
(2) Conv(in(i + 1, j)), Conv(in(i + 1, j + 1))))
(2) (2) Conv(in(i + 1, j)), Conv(in(i + 1, j + 1))))
(2) Conv(in(i + 1, j)), Conv(in(i + 1, j + 1)))) Using Equation (2) has two main advantages. Firstly, while computing the convolution,
the image data are already in the line buffers so the output of the convolution data can be
used without any write/read operations. In other words, the time for the computation of
the pooling layer is eliminated in the proposed method. Secondly, instead of the samples
of the convolution output, only downsampled samples are used to calculate the activation
functions. Thus, the complexity is less than the typical convolutional layer. A typical block diagram for proposed convolution using 5 × 5 kernels, 2 × 2 pool-
ing with stride 2 is depicted in Figure 6. If there are enough resources on the FPGA,
all processing can be performed in parallel using 100 DSP elements (i.e., 25 DSPs per con-
volution). Equation (2) can be processed in one clock cycle using the proposed convolution
in Figure 5. 2.2. Convolution with Optional Integrated Pooling PE In addition, according to the number of resources assigned to any convolution
layer, the number of clock cycles for this operation can be increased. For instance, if you
dedicate 50 DSPs for any convolution block, the execution time will become two clock
cycles for the computation in Figure 6. However, dedicating fewer DSPs results in more
memory and more logic usage to buffer the data temporarily and to realize the multiplexing
the data to feed the DSPs. Figure 6. Block Diagram of the Proposed Convolution. Lastly, in order to complete the proposed convolution in one clock cycle, the weights
have to be read in one clock cycle. Therefore, all the internal memories (i.e., BRAMs) storing
weights are 256 bits wide to read the kernel weights in one clock cycle. Each word of the
BRAM stores all kernel weights in a concatenated manner. Thus, switching from one kernel
to another has no time overhead in the convolution calculation. Figure 6. Block Diagram of the Proposed Convolution. Lastly, in order to complete the proposed convolution in one clock cycle, the weights
have to be read in one clock cycle. Therefore, all the internal memories (i.e., BRAMs) storing
weights are 256 bits wide to read the kernel weights in one clock cycle. Each word of the
BRAM stores all kernel weights in a concatenated manner. Thus, switching from one kernel
to another has no time overhead in the convolution calculation. 2.3. Feedforward PE Feedforward NNs are artificial NNs where the connections between nodes do not
form a cycle and information is transferred only in one direction. Feedforward NNs consist
of an input layer, an output layer, and hidden layers. A typical feedforward NN structure
can be seen in Figure 7. 8 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 7. Feedforward NN structure. Figure 7. Feedforward NN structure. Figure 7. Feedforward NN structure. From a computational point of view, the main computation of a feedforward NN is
matrix multiplication. The pseudo-code for the conventional computation of a feedforward
layer is shown in Algorithm 1. Algorithm 1 Conventional computation of feedforward layers Algorithm 1 Conventional computation of feedforward layers g
p
y
procedure FCLAYER(X, W, bias)
for i from 1 to outputNodeSize do
accumulator = 0
for j from 1 to inputNodeSize do
accumulator = accumulator + W[i][j]X[j]
end for
Y[i] = accumulator + bias[i]
end for
end procedure FPGAs are very suitable for pipeline processing and parallel matrix multiplication
which requires a substantial amount of DSP elements and memory resources. Thus, based
on the time budget and available resources, parallel multiplication operations result in
a significant reduction in processing time. In the feedforward PE, we define a 4-input
multiply-accumulators in order to multiply the 4 input data with the corresponding weights
and accumulates them in a single clock cycle. The parallelism of these 4-input multiply-
accumulators are achieved by unrolling the inner f or loop as shown in Algorithm 1 and
the degree of parallelism is limited by the available resources for the specific feedforward
layer. Besides, the bias summation can be omitted by initializing the accumulator with
the bias value. These optimizations reduce the execution times of the feedforward layers,
and the pseudo-code of the proposed computation is given in Algorithm 2. Algorithm 2 Proposed computation of feedforward layers
procedure FCLAYER(X, W, bias)
for i from 1 to outputNodeSize do
UNROLL_LOOP
accumulator = bias[i]
for j from 1 to inputNodeSize/4 do
accumulator = accumulator + MAC4DATA(&W[i][4 ∗j], &X[4 ∗j])
end for
Y[i] = accumulator
end for
end procedure
procedure MAC4DATA(∗a,∗b)
return a[0]b[0] + a[1]b[1] + a[2]b[2] + a[3]b[3]
end procedure Algorithm 2 Proposed computation of feedforward layers Algorithm 2 Proposed computation of feedforward layers
procedure FCLAYER(X, W, bias)
for i from 1 to outputNodeSize do
UNROLL_LOOP
accumulator = bias[i]
for j from 1 to inputNodeSize/4 do
accumulator = accumulator + MAC4DATA(&W[i][4 ∗j], &X[4 ∗j])
end for
Y[i] = accumulator
end for
end procedure
procedure MAC4DATA(∗a,∗b)
return a[0]b[0] + a[1]b[1] + a[2]b[2] + a[3]b[3]
end procedure Lastly, the output layer of a feedforward NN relies on a softmax function to convert
the vector of numbers to a vector of probabilities [26]. In a conventional NN, the output J. Low Power Electron. Appl. 2021, 11, 32 9 of 25 9 of 25 of the softmax function can be interpreted as the probability of membership of each class,
and the highest probability member is selected as the classification result. The softmax
function consists of complex exponentials and divisions which are highly compute- and
resource-demanding operations. Algorithm 1 Conventional computation of feedforward layers Using the softmax function only training of the NN offline
and instead of implementing a softmax function in hardware, the class having the highest
value in the output layer is selected as the classification result for simplification. Thus,
the same classification result is obtained in a computationally efficient way. In the design
of the feedforward PE, the data is processed in a pipelined manner so no additional clock
cycle is required to find the maximum. It is hidden in the computation of the output layer
of the feedforward PE. 2.4. Spiking PE The Spiking PE is based on a design presented in [8]. The architecture of the PE is
organized in layers of fully connected spiking neurons. The neuron model utilized in
the spiking PE is a leaky-integrate-and-fire neuron model. Each neuron in the network
receives spikes and emits spikes (i.e., binary pulses as in Figure 8a), and all of the neurons
in the network are executed in parallel and are physically implemented with digital logic. The connectivity among layers is fully connected, and the architecture supports forward
and recurrent connections, as shown in Figure 9. Input and output spikes can be routed in
and out of the PE using a high-speed (AXI4-Stream) data interface. Alternatively, spikes can
be generated by the input spike generators, and mean rate activity can be measured using
the Inter Spike Interval (ISI) calculator blocks. Once the input spikes are fed into the PE,
they enter a spike queue, and they are consumed by a weight controller that is in charge
of fetching the correct weight stored in on-chip memories (Block-RAM or Ultra-RAM). Once the weight has been fetched, the weight controller sends the weighted stimulus to
the neurons, which, in turn, perform the integration. If the neurons reach their threshold
value, then a spike is emitted to the next layer. The Spiking PE is completely event-driven,
meaning that computation happens upon the arrival of spikes only. On-chip memories are
used to store the synaptic weights, and this ensures a high memory bandwidth. The energy
performance of the PE varies depending on network size, mapping parameters, and re-
source availability but is in the order of ∼0.2 nano Joule per synaptic operation (nJ/SOP)
and can achieve throughput in the order of 10/40 Giga synaptic operations per second [8]. Figure 8. Leaky-Integrate-and-Fire (LIF) neuron model. (a) Behavior of the neuron. (b) Digital behav-
ior, neurons receive spikes in the form of a timestamp. It integrates them if it is not in the refractory
period. It would emit a spike if the membrane potential exceeded the threshold. The leakage block is
implemented with a bit shift for resource savings. Figure 8. Leaky-Integrate-and-Fire (LIF) neuron model. (a) Behavior of the neuron. (b) Digital behav-
ior, neurons receive spikes in the form of a timestamp. It integrates them if it is not in the refractory
period. It would emit a spike if the membrane potential exceeded the threshold. 2.4. Spiking PE The leakage block is
implemented with a bit shift for resource savings. Figure 8. Leaky-Integrate-and-Fire (LIF) neuron model. (a) Behavior of the neuron. (b) Digital behav-
ior, neurons receive spikes in the form of a timestamp. It integrates them if it is not in the refractory
period. It would emit a spike if the membrane potential exceeded the threshold. The leakage block is
implemented with a bit shift for resource savings. 10 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Layer
Layer
Weight controller
Layer
Weight controller
Neuron
wrapper
Weight controller
Weight controller
Neuron
wrapper
Timer
ISI
Buffer
Output
spikes
ISI
calculators
Mean
rates
Input
spikes
Weight
memory
Weight
memory
Weight
memory
Spike
generators
Spike queue
Spike queue
Spike queue
Neuron
wrapper
Figure 9. Spiking Neural Network PE. Spiking PE architecture is organized in layers of fully connected spiking neurons
that are executed in parallel. Timer Figure 9. Spiking Neural Network PE. Spiking PE architecture is organized in layers of fully connected spiking neurons
that are executed in parallel. In the following sections, we provide explanations for the different blocks of the
spiking PE, visible in Figure 9, and we give intuitions on their digital implementations. 2.4.1. Digital Leaky-Integrate-and-Fire Neurons 2.4.1. Digital Leaky-Integrate-and-Fire Neurons The neuron model, in its simplest form, is a cell membrane modeled by a leaky
capacitor. The neuron circuit can be seen as an exponentially decaying RC system with
constant decay time. In our discrete time-stepped model, the cell membrane voltage of
neuron i at step n + 1th is modeled as: Vi(n + 1) = Vi(n) · e
t
τm + ∑
j
Wi,jSj
(3) (3) in which t is the time step, Wi,j are the synaptic weights, and Sj are the input spikes. The input spikes are instantaneously integrated by the synapses that produce a step increase
(or decrease) of Wi,j to the membrane potential of neuron i when receiving spikes from
neuron j. The model also implements some features to more accurately model biological
neurons. These features are the implementation of a threshold Vthr that makes a neuron emit
one spike when Vmem > Vthr. After a spike is emitted, the neuron’s membrane potential
(Vmem) is reset to zero, see Figure 8a. Additionally, we have implemented a programmable
refractory period Tre during which a neuron is not sensitive to incoming spikes after the
emission of one spike, as shown in Figure 8. Importantly, our implementation of the
neuron model does not loop through time steps. Instead, our design is event-driven,
and the algorithm loops through spike events, indicating that computations only happen
upon spiking activity. Since calculating the exponential function for the neuron decay
is resource and time-expensive, we have approximated the exponential function with
bit-shifts operations. By shifting the membrane potential by n bits to the right, its value
is divided by 2n, and the least significant bits are eliminated. The function of spiking
PE digital logic is summarized in Algorithm 3. Since our architecture is based on fully
connected layers, the neurons in a layer receive a single previous spike time shared over
all neurons in a layer. Besides, an ‘else if’ statement is used to set the membrane potential
to zero if it becomes negative. While time-driven algorithm loops through all time steps,
our event-driven algorithm loops only through spike events, resulting in less computations
executed only upon spiking activity. in which t is the time step, Wi,j are the synaptic weights, and Sj are the input spikes. 2.4.2. The Spike Queue The spike queue is the purple module at the top of Figure 9. It receives two streams of
spikes, one for forward spikes generated by the previous layer and one for backward spikes
generated by the next layer. A spike is encoded by the address of the neuron that generated
the spike, including the forward or backward information. Each stream is separately
buffered using a FIFO. The FIFO outputs are arbitrated using a round-robin scheme. 2.4.1. Digital Leaky-Integrate-and-Fire Neurons M
Require: set of synaptic weights W ∈RN×M
t1...M
re
←0
t1...L
prev ←0
for t in St do
for s in Ssrc do
for l in ldst do
tδ ←t −tl
prev
tl
prev ←t
for d in Sl
dst do
Vi
m ←Vi
m · e−tδ/τm
if t > ti
re then
Vi
m ←Vi
m + WSksrc,i
end if
if Vi
m > Vthr then
Spike()
Vi
m ←Vi
reset
ti
re ←t + tre f rac
end if
if Vi
m < 0 then
Vi
m ←0
end if
end for
end for
end for
end for Require: set of sorted spike times St
Require: set of corresponding source neurons Ssrc ∈1 . . . N
Require: set of corresponding destination layers ldst ∈1 . . . L with neurons Sl
dst ∈1 . . . M
Require: set of synaptic weights W ∈RN×M
1
M
0 Require: set of synaptic w
t1...M
re
←0
t1...L
prev ←0
for t in St do
for s in Ssrc do
for l in ldst do
t ←t
tl 2.4.1. Digital Leaky-Integrate-and-Fire Neurons The input spikes are instantaneously integrated by the synapses that produce a step increase
(or decrease) of Wi,j to the membrane potential of neuron i when receiving spikes from
neuron j. The model also implements some features to more accurately model biological
neurons. These features are the implementation of a threshold Vthr that makes a neuron emit
one spike when Vmem > Vthr. After a spike is emitted, the neuron’s membrane potential
(Vmem) is reset to zero, see Figure 8a. Additionally, we have implemented a programmable
refractory period Tre during which a neuron is not sensitive to incoming spikes after the
emission of one spike, as shown in Figure 8. Importantly, our implementation of the
neuron model does not loop through time steps. Instead, our design is event-driven,
and the algorithm loops through spike events, indicating that computations only happen
upon spiking activity. Since calculating the exponential function for the neuron decay
is resource and time-expensive, we have approximated the exponential function with
bit-shifts operations. By shifting the membrane potential by n bits to the right, its value
is divided by 2n, and the least significant bits are eliminated. The function of spiking
PE digital logic is summarized in Algorithm 3. Since our architecture is based on fully
connected layers, the neurons in a layer receive a single previous spike time shared over
all neurons in a layer. Besides, an ‘else if’ statement is used to set the membrane potential
to zero if it becomes negative. While time-driven algorithm loops through all time steps,
our event-driven algorithm loops only through spike events, resulting in less computations
executed only upon spiking activity. 11 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Algorithm 3 Event-driven LIF neuron update Algorithm 3 Event-driven LIF neuron update
Require: set of sorted spike times St
Require: set of corresponding source neurons Ssrc ∈1 . . . N
Require: set of corresponding destination layers ldst ∈1 . . . L with neurons Sl
dst ∈1 . . . tδ ←t
tprev
tl
prev ←t
for d in Sl
dst do
Vi
m ←Vi
m · e−tδ/τm
if t > ti
re then
Vi
m ←Vi
m + WSksrc,i
end if
if Vi
m > Vthr then
Spike()
Vi
m ←Vi
reset
ti
re ←t + tre f rac
end if
if Vi
m < 0 then
Vi
m ←0
end if
end for
end for
end for
end for 2.4.3. The Neuron Wrapper The neuron wrapper module in Figure 9 consists of a set of neurons. All neurons exist
in parallel, but the layer’s output is a serial stream of spike sources addresses, identical to
the spike queue input streams. The output spikes from all neurons in a layer are serialized
using a round-robin arbiter to grant exclusive access to the output stream. However,
this solution introduces two issues. Firstly, when the number of neurons N in a layer is
large, the number of resources to implement the arbiter and the number of logic levels in
the arbiter increases. This leads to high resource utilization and a low maximum clock
frequency. Secondly, the serial stream processes at a maximum of one spike per clock
cycle, while the neurons generate many spikes in a single clock cycle. For these reasons,
the neuron wrapper module groups neurons in clusters, and a subset of neurons in each
cluster is activated in parallel. This choice enables to limit of the maximum number of
spikes generated in parallel. This, in turn, reduces the requirement only to have one buffer
for each neuron in a cluster. Depending on the cluster sizes, this can save a significant
amount of buffers and, therefore, hardware resources. Figure 10 shows the sharing of spike
buffers for three clusters of two neurons. The spike signals are fed through a logical or gate
to generate the buffer write signal. As spikes are represented using their source address, J. Low Power Electron. Appl. 2021, 11, 32 12 of 25 12 of 25 the information from the neuron addressing module is used. In this example, instead of
six buffers, one buffer for each neuron, only two buffers are required since, at maximum,
two neurons spike simultaneously. Lastly, the arbiter arbitrates between the buffers to
ensure fair and mutual access to the serial stream. the information from the neuron addressing module is used. In this example, instead of
six buffers, one buffer for each neuron, only two buffers are required since, at maximum,
two neurons spike simultaneously. Lastly, the arbiter arbitrates between the buffers to
ensure fair and mutual access to the serial stream. Figure 10. Spike buffer sharing mechanism. Figure 10. Spike buffer sharing mechanism. Figure 10. Spike buffer sharing mechanism. 2.4.4. The Weight Memory Block The weight memory block stores the weight matrix for the connections between two
layers. One of the major challenges when designing an efficient spiking PE is to map the
connection matrices to the on-chip memories efficiently. The way the weights are dis-
tributed over the memories determines the memory bandwidth and, ultimately, the system
throughput. Additionally, this mapping determines the implementation of the weight
controller. Our design exploits dual-port on-chip memories (BRAM or UltraRAM), and to
increase the number of weights per single memory access, we adopt several strategies. Firstly, we store quantized weights in memory (6-bits in this design). Secondly, we adopted
a generic and parametric description of the memory organization that can accommodate
on-chip memories of several bit-depth and bit-width. Lastly, we can explore the mapping
of the weights using a parametric search at design time. This ensures the selection of
optimal parameters for the available resources. 2.4.5. The Weight Controller The weight controller takes care of fetching the correct weight from memory, depending
on the source address. Since our design assumes fully connected layers, the only infor-
mation required to retrieve the correct weight is the neuron source address. The weight
controller receives the spike source addresses Ssrc and spike direction (recurrent or forward)
from the spike queue. The weight controller contains a state machine that decodes the
address received, fetches the correct weight, and generates valid weight signals for the
destination neurons. 2.5. DPR DPR is a powerful tool particularly for trying to reduce the resources and power
consumption needed to implement NNs with multiple functionalities. Moreover, instead
of training and implementing the NNs together, designing them separately, results in
more accurate results since separate designs are less complex and less deep than the
combined designs where different classes and datasets are trained together. Moreover,
power consumption is also substantially reduced with DPR, since some resources are
shared between different PEs and resource utilization is less than the combined designs. g
From an architectural point of view, DPR introduces considerable flexibility to the
proposed system architecture by allowing different NN realizations on the same FPGA
implementation. By only updating a small part of the FPGA (i.e., a PE) with a partial
bit stream, a different NN accelerator can be implemented. Besides, the depth of the
NN (i.e., number of layers) can be changed via programming the interconnections in
the proposed architecture. Without changing the placement and routing of the static
design, a different NN architecture can be realized only by implementing the new PE. Thus, implementation times are highly reduced while designing a new NN architecture by
changing some of the layers of the current NN. 2.4.6. Input and Output Interfaces The spiking PE communicates employing input and output spikes. In general, the spik-
ing PE is agnostic on the information coding scheme. In fact, it is compatible with temporal
coding and mean rate coding. To connect the spiking PE to traditional neural network
accelerators, we have implemented digital to spike conversion at the input of the spiking
PE that linearly converts M bits values into an input mean rate frequency. This is achieved
through a spike generator module, which is shown in Figure 11. This module uses the
digital input value as the probability of a neuron to spike within a fixed time window. Small stochasticity is implemented using a Linear-Feedback Shift Register (LFSR) triggered
once every time. A spike is generated if the pseudorandom number is greater than the
mean rate. The value of the mean rate is relative to its maximum value of 2M −1 for M J. Low Power Electron. Appl. 2021, 11, 32 13 of 25 13 of 25 bits. A mean rate of zero results in zero spikes, and a mean rate of 2M −1 results in one
spike every timer period. The probability that a neuron spike upon a timer trigger for a
mean rate of M bits is P(spike) =
MR
2M −1. (4) P(spike) =
MR
2M −1. (4) P(spike) =
M
2M −1. (4)
Figure 11. Spike generation with stochastic ISI. Figure 11. Spike generation with stochastic ISI. When a mean rate coding is used, the output of the spiking PE can be interpreted
by means of the Inter-Spike-Interval (ISI) measure. The ISI is measured for each output
neuron by computing the interval between two consecutive spikes. The resulting values
are fed into registers and can be read via the AXI-lite register interface. Additionally, input and output single spikes can be buffered via external (DDR)
memory and streamed to the spiking PE exploiting the AXI-Stream protocol. We used the
Xilinx DMA IP to outsource the communication to and from external memory. We have
only used AXI-stream for debugging as the use-cases and experimentation below are
carried using the mean rate mode of operation for the spiking neural network. 3. Results and Discussion The summary of both use cases is given in Figure 12. In this figure, intersection parts show
the common layers for the classifiers (i.e., static regions) whereas the other parts show the
layers specific to the related classifier (i.e., dynamic regions). Figure 12. Summary of Use Cases: (a) Same NN architecture with Different Classifiers, (b) Different
NN architectures for Digit Classifier. Figure 12. Summary of Use Cases: (a) Same NN architecture with Different Classifiers, (b) Different
NN architectures for Digit Classifier. 3. Results and Discussion For the proof of concept, two different use cases are implemented on the hardware. In the first use case, three different CNN classifiers trained by three different datasets
are designed and switched using DPR for two different scenarios. In the first scenario,
a digit classifier trained by the MNIST dataset [27] and a letter classifier trained by EMNIST
dataset [28] are switched only updating the feedforward layers since the number of classes
is different between digit and letter classifiers. In the second scenario, a digit classifier
trained by MNIST and an object classifier trained by CIFAR10 dataset [29] are switched
by updating the first convolution layer because the dataset for digit classifier is grayscale
whereas the object dataset is an RGB image. Thus, the number of input channels is different,
and the first convolutional layers are required to be different. In both scenarios, the weights J. Low Power Electron. Appl. 2021, 11, 32 14 of 25 14 of 25 and biases of the structurally identical layers are updated from the memory-mapped
interface of the PEs and the non-identical layers are dynamically partially reconfigured
with the corresponding PE. In the second use case, DPR is used to switch between a CNN
classifier and a hybrid NN (i.e., consisting of convolutional layers and spiking layers)
classifier. Both classifiers are digit classifiers and the convolutional layers of both classifiers
have the same structure and use the same weights and bias values. The switching between
CNN to hybrid NN is achieved by updating the feedforward layers with spiking layers. The summary of both use cases is given in Figure 12. In this figure, intersection parts show
the common layers for the classifiers (i.e., static regions) whereas the other parts show the
layers specific to the related classifier (i.e., dynamic regions). and biases of the structurally identical layers are updated from the memory-mapped
interface of the PEs and the non-identical layers are dynamically partially reconfigured
with the corresponding PE. In the second use case, DPR is used to switch between a CNN
classifier and a hybrid NN (i.e., consisting of convolutional layers and spiking layers)
classifier. Both classifiers are digit classifiers and the convolutional layers of both classifiers
have the same structure and use the same weights and bias values. The switching between
CNN to hybrid NN is achieved by updating the feedforward layers with spiking layers. 3.1. Use Case 1: CNN to CNN for Different Datasets The proposed architecture can be used to switch between the same type of NN
architectures used for the classification of different classes. Since different NN classifiers
can have different number of output classes or different number of input channels, an NN
architecture may not be used as-is for the recognition of another class set even if weights
and biases are changed for the new classification problem. In other words, at least the
input or the output layer should be changed if there is a change in the input or the output
parameters, respectively. Therefore, in this use case, we investigate both alternatives
(i.e., a change in the number of input channels and a change in the number of output
classes) using different scenarios. 3.1.1. Digit Classifier to Letter Classifier Scenario In the first scenario, we have two different CNN classifiers having a different number
of classes. The digit classifier is classifying the ten digits trained by the MNIST dataset. On the other hand, a letter classifier is used to classify the 26 letters in the English alphabet,
trained by the EMNIST dataset. The CNN structure of the digit and letter classifiers can be
seen in Figure 13. Except for the weights and biases, the only difference between the digit
and the letter classifier is the feedforward layers. Without implementing the whole design
and reloading the whole bitstream, only implementing a feedforward PE for the letter
classifier and updating only a small part of the FPGA with partial bitstream is enough to
switch the letter classifier. To put it another way, using the proposed architecture, it can be
switched between digit classifier to letter classifier in a very short period of time without
waiting for the reconfiguration of the whole FPGA. 15 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 13. Digit classifier and letter classifier, and the switching between them using DPR. Figure 13. Digit classifier and letter classifier, and the switching between them using DPR. As shown in Figure 13, the input of the CNNs is a 32 × 32 8-bit grayscale image,
and the output is the classification result. Each CNN accelerator consists of two convolu-
tional layers, two max-pooling layers, a hidden feedforward layer, and an output layer. ReLU activation function is used in the convolutional and hidden layers. In convolutional
layers, the convolutional kernel and pooling kernel are selected as 5 × 5 and 2 × 2 for
their better performance as compared to other size kernels. In the pooling layers, the
downsampling factor is selected as 2. After the convolutional layers, data is flattened and
fed to the feedforward layers. In the letter classifier, 48 nodes and ten nodes are used for
hidden and output layers, respectively. For the letter classifier, 96 nodes and 26 nodes are
used for the hidden and output layers, respectively. The switching between accelerators is
achieved by updating the partial bitstreams corresponding to the feedforward layers of the
related CNN. 3.1.2. Digit Classifier to Object Classifier Scenario The second scenario is based on the differences in the input format. Especially in
image classification applications, various datasets may have different image formats such
as grayscale, RGB, IR, and hyperspectral. Thus, the first convolutional layers are designed
according to the data format of the image. In this scenario, digit classifier and object
classifier are switched using DPR only by updating the first convolutional layer, since ob-
ject classifier is trained with CIFAR10 dataset consisting of three-channel colored image
(i.e., RGB) whereas digit classifier has single-channel grayscale image dataset for training. Both datasets have ten classes so the other layers are in the same structure (i.e., second
convolutional layers and feedforward layers). The CNN structure of the digit and object
classifiers can be seen in Figure 14. Note that, weights and biases also differ between the
two CNN accelerators but they are updated through the memory-mapped interface of
the PE. As shown in Figure 14, the digit classifier has the same structure as in Figure 13,
however, the object classifier RGB input format is 32 × 32 × 3. In other words, in the
first convolutional layer, the digit classifier has 1 input image and 3 output feature maps
whereas the object classifier has 3 input images and 3 output feature maps. Thus, to realize
the object classifier, the convolutional PE for layer 1 is updated from the digit classifier
implementation using DPR and in the order of milliseconds, both classifiers can be switched
between each other using partial bit files. 16 of 25 16 of 25 J. Low Power Electron. Appl. 2021, 11, 32 Figure 14. Digit classifier and object classifier, and the switching between them using DPR. Figure 14. Digit classifier and object classifier, and the switching between them using DPR. 3.2. Use Case 2: CNN to Hybrid for Different Architectures Each NN architecture has its own advantages that can be accuracy, throughput, power
consumption, or lower complexity. Thus, in some scenarios, it can be preferred to use
the same hardware resources to realize different NN designs, and switch between them
without reconfiguring the whole FPGA. Thus, in the second use case, we opt to switch
between different NN architectures using the proposed hardware architecture and DPR. To do so, a digit classifier trained by the MNIST dataset is implemented with two different
NN architecture. The first architecture is a CNN architecture which is explained above in
Use Case 1 whereas the second architecture is a hybrid NN which is a concatenation of
convolutional layers and spiking layers. In this architecture, feature maps are first extracted
using the convolutional layers, and then these feature maps are fed to the spiking layers
for the classification of the digits. The details of these NNs are given in Figure 15. Figure 15. CNN and Hybrid architectures and switching between each other using DPR. 3 3 I
l
t ti Figure 15. CNN and Hybrid architectures and switching between each other using DPR. Figure 15. CNN and Hybrid architectures and switching between each other using DPR. 3.3. Implementation The proposed hardware architecture in Figure 4 is implemented using Xilinx Vivado
2020.2. Using the proposed hardware architecture, two different use cases and three
different scenarios explained in the previous section are implemented. In total, 5 different
PEs are designed to realize those scenarios. Except for the spiking PE which is a pure
VHDL design, all PEs are designed using the Vitis HLS tool, which is used to transform
C, C++, and System C codes into the register transfer level implementations [30]. The PE
designs are first verified in the simulation environment. Then, they are implemented
and tested on a Zedboard FPGA board [31] which is equipped with a Xilinx Zynq 7020
FPGA. The summary of the resource usage of each PE can be seen in Table 1. As shown in
Table 1, the resource usage is dominated by the Convolution PEs in the NN architectures. J. Low Power Electron. Appl. 2021, 11, 32 17 of 25 17 of 25 In addition, Feedforward PEs use moderate DSP usage with the minimum deployment of
LUTs and FFs since the main computation is the matrix multiplication in Feedforward PEs. On the other hand, the Spiking PE does not use any DSPs because there is no multiplication
in the spiking layers. However, it consumes more logical resources, such as LUTs and FFs. In addition, Feedforward PEs use moderate DSP usage with the minimum deployment of
LUTs and FFs since the main computation is the matrix multiplication in Feedforward PEs. On the other hand, the Spiking PE does not use any DSPs because there is no multiplication
in the spiking layers. However, it consumes more logical resources, such as LUTs and FFs. Table 1. Resource usage of different PEs. Classifier
BRAM
DSP
LUT
FF
Convolutional PE
Digit/Letter Classifier Layer 1
17
34
4258
4604
Convolutional PE
Object Classifier Layer 1
21
47
7485
6382
Convolutional PE
Digit/Letter/Object Classifier Layer 2
15
113
13,894
11,893
Feedforward PE
Digit Classifier
8.5
20
810
740
Feedforward PE
Letter Classifier
20
20
601
616
Spiking PE
Digit Classifier
21
0
5313
8931 Table 1. Resource usage of different PEs. In each scenario, to switch one classifier to the other classifier, only a single PE is
updated using DPR. However, for another application, more than a single PE could be
updated as well. For the DPR, Vivado’s partial reconfiguration flow is used. 3.4. Evaluation To conduct a fair performance evaluation of the proposed architecture, we compare
the performance of our digit CNN classifier with three state-of-the-art digit CNN classifiers,
trained by the MNIST dataset and using the same CNN architecture (i.e., two convolu-
tional layers, two pooling layers, one hidden layer). The first accelerator is a static CNN
accelerator [32]. In that study, a ZCU102 evaluation board is used for the implementation. For each convolutional layer and pooling layer, separate accelerators are designed. The sec-
ond accelerator is a DPR design, and according to the energy level of the power source,
the processing system uploads the required partial bitstream at run time using ICAP [17]. The last work is using cascaded connections of processing engines designed to compute
the convolution [33]. Pipelining and tiling techniques are used to improve the performance
of that design. The performance comparison with these digit classifier accelerators is given
in Table 3. As can be seen from Table 3, our proposed digit classifier accelerator has the
shortest processing time to complete the classification of an image due to the hardware
optimizations in the PE designs that allow for a considerable throughput improvement
with using less number of DSPs in comparison with the other accelerators. However,
the LUT usage is slightly higher than the other designs because of the implementation of
the components for the flexibility of the proposed hardware architecture. Table 3. Performance Comparison of Different Digit Classifiers. BRAM
DSP
LUT
Processing Time
FPGA Model
Shi [32]
54
204
25,276
170 µs
Zynq UltraScale+ 9EG
Youssef [17]
9
N/A
19,141
270 µs
Zynq 7020
Li [33]
619
916
9071
490 µs
Virtex7 485t
This work
58.5
167
24,980
62 µs
Zynq 7020 Table 3. Performance Comparison of Different Digit Classifiers. In the proposed architecture, all data is processed in a pipelined manner. Although
the total processing time is 62 µs for the digit classifier; the accelerator can be fed with
a higher frame rate. Every PE can process new data after finishing its task, i.e., there is
no need to wait until the end of the overall processing of one image to proceed with the
next. As shown in Figure 17, the overall frame rate only depends on the processing time of
the PE with the largest delay which is 22 µs (processing time of Feedforward PE for the
digit classifier CNN). 3.3. Implementation 2021, 11, 32 18 of 25 18 of 25 the other CNNs as it has fewer nodes in the Feedforward PE for the output layer and
fewer channels in the Convolution PE for the input layer. Lastly, the Object classifier
has the highest DSP usages among them because three-channel classifiers require more
multiplications in the input layer in comparison with the one-channel classifiers. the other CNNs as it has fewer nodes in the Feedforward PE for the output layer and
fewer channels in the Convolution PE for the input layer. Lastly, the Object classifier
has the highest DSP usages among them because three-channel classifiers require more
multiplications in the input layer in comparison with the one-channel classifiers. Table 2. Resource usage of different classifier accelerators, The numbers in parenthesis show the
utilization on Zedboard. BRAM
DSP
LUT
FF
Digit Classifier (CNN)
58.5 (42%)
167 (76%)
24,980 (47%)
27,925 (26%)
Letter Classifier (CNN)
70 (50%)
167 (76%)
24,771 (47%)
27,801 (26%)
Object Classifier (CNN)
62.5 (45%)
180 (82%)
27,402 (52%)
29,496 (28%)
Digit Classifier (CNN + SNN)
71 (51%)
147 (67%)
31,093 (58%)
37,698 (35%) Table 2. Resource usage of different classifier accelerators, The numbers in parenthesis show the
utilization on Zedboard. 3.3. Implementation The size and
location of the Pblock of the corresponding PE is determined according to the resource
utilization of the related layer of the NN accelerator. In the use cases, two different
Pblocks are defined. The first Pblock is for the last layer of Digit CNN Classifier/Digit
Hybrid Classifier/Letter CNN Classifier. The second Pblock is for the first layer of Digit
CNN Classifier/Object CNN Classifier. Except for the Pblock, the remaining layout of the
implementation is static. Thus, only Pblock is placed and routed again for the new scenario
realizations. The layout of the implemented NN classifiers and the location of the related
Pblocks can be seen in Figure 16. Figure 16. The layout of the implemented classifier accelerators. The red rectangles correspond to
the partial reconfigurable areas (i.e., Pblocks), yellow color shows the dynamic part, and cyan color
shows the static part of the designs: (a) Digit CNN Classifier/Digit Hybrid Classifier/Letter CNN
Classifier (b) Digit CNN Classifier/Object CNN Classifier. Figure 16. The layout of the implemented classifier accelerators. The red rectangles correspond to
the partial reconfigurable areas (i.e., Pblocks), yellow color shows the dynamic part, and cyan color
shows the static part of the designs: (a) Digit CNN Classifier/Digit Hybrid Classifier/Letter CNN
Classifier (b) Digit CNN Classifier/Object CNN Classifier. In the hardware tests of the scenarios, the input images are loaded from the serial
port of the Zedboard and the results are written to the registers of the final PE in the chain. The registers are read from the memory-mapped interface of the PE. In addition, the internal
data transfers between PEs are tested on the system using Vivado Hardware Manager. All the implementations operate at 100-MHz clock frequency. Table 2 provides resource
usages of the classifier accelerator implementations: Digit Classifier (CNN), Letter Classifier
(CNN), Object Classifier (CNN), and Digit Classifier (CNN + SNN). In this table, the hybrid
classifier DSP usage is the lowest since spiking layers do not use any DSP because there is
no multiplication operation in the computation of the spiking layers. However, it consumes
more LUTs and FFs since some of the LUTs and FFs are used to convert the feature maps
to the spikes in Spiking PE. Moreover, the Digit Classifier CNN uses fewer BRAMs than J. Low Power Electron. Appl. 3.4. Evaluation For the power consumption of the processor side, since the pro-
with the same frequency with the same software architecture, the power
528 mW and is the same for all classifiers. These consumptions are taken
er report of the implemented (i.e., post-routed) design. Figure 17. Pipelined processing of the Digit Classifier Accelerator. Figure 17. Pipelined processing of the Digit Classifier Accelerator. Using the DPR concept in the proposed architecture has two main advantages. Firstly,
instead of training different datasets together, realizing specialized classifiers as different
NN greatly increases the accuracy in each task. Table 4 shows the accuracy of three different
implementations. In this table, the Character Classifier-1 is a mixed static CNN accelerator
trained with both letter and digit datasets together. On the other hand, Digit Classifier
and Letter Classifier in Figure 13 are two distinct CNNs. The switching between the digit
classifier and the letter classifier is achieved using DPR, reconfiguring only some network
layers. The accuracy of the Character Classifier-1 drops by 5% and 10% compared to the
specialized letter and the digit neural network models. The increased difficulty causes this
drop in performance for a single neural network that needs to learn multiple tasks. Table 4. Comparison of Digit Classifier, and Letter Classifier with Character Classifier-1 in terms of
resource usage and accuracy on Zedboard (Zynq7020). BRAM
DSP
LUT
Accuracy
Execution Time
Character Classifier 1 (Static)
70
167
25,117
87.53
87.55 µs
Letter Classifier (DPR)
70
167
24,771
92.45
84.92 µs
Digit Classifier (DPR)
58.5
167
24,980
98.88
62.19 µs Table 4. Comparison of Digit Classifier, and Letter Classifier with Character Classifier-1 in terms of
resource usage and accuracy on Zedboard (Zynq7020). The second advantage of using DPR is the reduction in power consumption. Without
sacrificing the accuracy and throughput, a NN design can be realized by using separate
Feedforward PEs for digit classifier and letter classifier respectively. Since the proposed
architecture allows to change the connection at run time, the classifier can be switched to
the related Feedforward PE by updating the connections in the AXI stream switch in the
proposed architecture. Thus, the accuracy and execution times remain the same as the digit
and letter classifiers given in Figure 13. However, instantiating both Feedforward PEs in
the same implementation results in an increase in resource usage and power consumption
compared with the DPR implementations as shown in Table 5. 3.4. Evaluation Therefore, using the pipelining in Figure 17, the proposed accelerator
architecture achieves frame rates of up to 45K images/sec in digit classification which is
7× higher than the state-of-the-art digit classifier accelerators given in Table 3. 19 of 25 19 of 25 J. Low Power Electron. Appl. 2021, 11, 32 d processing of the Digit Classifier Accelerator. PR concept in the proposed architecture has two main advantages. Firstly,
g different datasets together, realizing specialized classifiers as different
ses the accuracy in each task. Table 4 shows the accuracy of three different
In this table, the Character Classifier-1 is a mixed static CNN accelerator
h letter and digit datasets together. On the other hand, Digit Classifier
fier in Figure 13 are two distinct CNNs. The switching between the digit
letter classifier is achieved using DPR, reconfiguring only some network
acy of the Character Classifier-1 drops by 5% and 10% compared to the
and the digit neural network models. The increased difficulty causes this
nce for a single neural network that needs to learn multiple tasks. on of Digit Classifier, and Letter Classifier with Character Classifier-1 in terms of
accuracy on Zedboard (Zynq7020). BRAM
DSP
LUT
Accuracy
Execution Time
ifier 1 (Static)
70
167
25,117
87.53
87.55 µs
fier (DPR)
70
167
24,771
92.45
84.92 µs
fier (DPR)
58.5
167
24,980
98.88
62.19 µs
advantage of using DPR is the reduction in power consumption. Without
curacy and throughput, a NN design can be realized by using separate
for digit classifier and letter classifier respectively. Since the proposed
ws to change the connection at run time, the classifier can be switched to
orward PE by updating the connections in the AXI stream switch in the
ture. Thus, the accuracy and execution times remain the same as the digit
ers given in Figure 13. However, instantiating both Feedforward PEs in
entation results in an increase in resource usage and power consumption
he DPR implementations as shown in Table 5. In the table, Character
assifier design instantiating the Feedforward PEs of the digit classifier and
the same implementation. Thus, as clearly seen from Table 5, using DPR
n more energy- and resource-efficient compared with the design having
d PEs. There is a 7% and a 10% decrease in the power consumption of
ogic in DPR designs as compared to the static design having digit and
ogether. 3.4. Evaluation In the table, Character
Classifier-2 is a classifier design instantiating the Feedforward PEs of the digit classifier and
letter classifier in the same implementation. Thus, as clearly seen from Table 5, using DPR
makes the design more energy- and resource-efficient compared with the design having
both Feedforward PEs. There is a 7% and a 10% decrease in the power consumption of
programmable logic in DPR designs as compared to the static design having digit and
letter classifiers together. For the power consumption of the processor side, since the pro-
cessor is running with the same frequency with the same software architecture, the power
consumption is 1528 mW and is the same for all classifiers. These consumptions are taken
by Vivado’s power report of the implemented (i.e., post-routed) design. J. Low Power Electron. Appl. 2021, 11, 32 20 of 25 20 of 25 Table 5. Comparison of Digit Classifier, and Letter Classifier with Character Classifier-2 in terms of
resource usage and power consumption. BRAM
DSP
LUT
Power Consumption (Programmable Logic)
Character Classifier-2
79
187
26,525
1231 mW
Letter Classifier
70
167
24,771
1123 mW
Digit Classifier
58.5
167
24,980
1146 mW There are also some disadvantages of using DPR. First of all, using DPR increases the
difficulty of the design process. The size of the Pblock is to be defined well considering
the resource usage of the related PEs. Moreover, defining multiple Pblocks requires a
careful placement that should allow easy routing and prevent congestion. However,
these design difficulties are coped with only once and the location and size of the Pblocks
will probably not be updated unless there are major changes in the design. Besides,
another disadvantage of the DPR can be the power overhead during partial reconfiguration. However, as compared with the total reconfiguration of the FPGA, DPR has negligible
overhead in terms of power consumption [34]. Similarly, in [35], different experiments
were conducted to evaluate the power consumption overhead of DPR for Zynq devices
and measurements were done for the processor and the hardware logic fabrics in real-time. In that paper, it is noticed that the power consumed by the FPGA is almost unaffected
when applying DPR. Thus, it can be concluded that the proposed method has no significant
power or energy overhead while realizing different NN accelerators using DPR. 3.4. Evaluation p
gy
g
g
Lastly, in the proposed architecture, the switch time between digit and letter classifiers
(i.e., DPR time) is 9.4 ms, and the switch time between digit and object classifiers is
17.1 ms since the PCAP throughput is 145 MB/s [36] and the partial bit file sizes are
1.37 MB and 2.48 MB for the Pblocks given in Figure 16, respectively. However, for full
configuration time of Zedboard is around 250 ms [31]. Therefore, the proposed method
can be effectively used, especially in time-critical applications, without wasting the time
for updating the whole bitstream. The system can be switched in a very short period of
time between digit and letter classifiers only updating the partial bitstream. To do so, in a
short response time, the characters can be identified with higher accuracy as compared
to the static character classifier designs. However, when the switching between different
classifiers is very frequent, the throughput performance may be degraded by the DPR
overhead. Since the size of the partial bistream is directly proportional to the size of
the region it is reconfiguring, using smaller Pblock may help to decrease the DPR time. In addition, DPR overhead can also be decreased using compressed partial bit files. Yet,
these optimizations have limited improvement on DPR overhead and in order to preserve
higher overall throughput, the number of switchings between different classifiers should
be kept minimal. 3.5. Evaluation of the Spiking PE This section compares the spiking PE alone when solving MNIST digit recognition
with other spike-based neural network implementations in FPGA. We have implemented
a feed-forward network of four layers connected to the 784 input pixels from the MNIST
images. The layer size is 784-720-720-720-10. Input has been provided with the mean-rate
generators on board, and the outputs have been evaluated by means of the ISI calculators
blocks. The spiking PE achieves an accuracy of 99.3% and a peak-throughput of 40.71 Giga
Synaptic Operation Per second (GSOPS) with an efficiency of 0.050 nJ/SO. For compar-
ison, Table 6 shows a list of recently reported spiking neural networks implemented in
FPGA. The Bluehive project [37] from Cambridge University implemented 256,000 Izhike-
vich neurons with 1000 synapses each by interconnecting four FPGAs that each contains
64,000 neurons. Minitaur [9] and its improved version n-Minitaur [38] build upon [39]
and operate event-driven. They both time-multiplex 32 physical Leaky-Integrate-and-Fire
(LIF) neurons to emulate at a maximum of 65,536 neurons. Note that this is the only purely J. Low Power Electron. Appl. 2021, 11, 32 21 of 25 21 of 25 event-driven implementation in the reported work. In [40] the Efficient Neuron Archi-
tecture (ENA) is proposed, which consists of layers of neurons that communicate using
packets. Using the LIF model and 32-bit precision, it promises to emulate 3982 neurons and
400 k synapses. The authors implemented only three neurons, though. In [41] an FPGA
Design Framework (FDF) is proposed that time-multiplexes up to 200,000 neurons with
one physical conductance-based neuron. Furthermore, a network is presented that consists
of 1.5 M neurons and 92 G synapses on an FPGA. Only the most significant bits of the
exponential decay are stored to reduce memory usage, and the least significant bits are
stochastically generated. The same authors emulate 20 M to 2.6 B neurons in a so-called
Neuromorphic Cortex Simulator (NCS) [42] using only one FPGA. There is a significant
drop in performance, which is likely a result of using off-chip memory, but that increases
the supported network size. Table 6. FPGA-based spiking neural network implementations in chronological order. 3.5. Evaluation of the Spiking PE Name Bluehive
FDF
n-Minitaur
Pani
NCS
Tsinghua
This Work
Reference [37]
[41]
[38]
[43]
[42]
[44]
N/A
Year 2012
2014
2016
2017
2018
2020
2021
FPGA Stratix IV
Virtex 6
Spartan 6
Virtex 6
Stratix V
Zynq 7000
Zynq Ultrascale+
Clock (MHz) 200
266
105
100
200
200
250
Neuron model Izhikevich
Conductance LIF
Izhikevich
LIF
LIF
LIF
Network Unknown
Unknown
Feed-forward Recurrent
Unknown
Feed-forward Recurrent
Driven Time-driven Time-driven
Event-driven
Time-driven Time-driven Hybrid
Event-driven
Weight storage Off-chip
On-chip
Off-chip
On-chip
Off-chip
Off-chip
On-chip
Weight bit-width 12 bits
12 bits
16 bits
7 bits
4 bits
16 bits
6 bits
Cores 16
23
32
8
200 k
Unknown
Same as neurons
Number of neurons 256 k
1.5 M
1794
1440
100 M
2842
2954
Time resolution 1 ms
0.32 ms
N/A
0.1 ms
1 ms
N/A
N/A
Peak throughput (GSOPS) 0.256
1200
0.0535
0.0144
20
0.67
40.71
Power dissipation (W) Unknown
Unknown
1.5
8.5
32.4
0.5
2.039
Energy efficiency (nJ/SO) Unknown
Unknown
28
590
1.62
712
0.050
MNIST accuracy Unknown
Unknown
94.1%
Unknown
Unknown
97.1%
99.3% Table 6. FPGA-based spiking neural network implementations in chronological order. Digital ASIC implementations supporting spiking networks and can be extremely
power- and energy-efficient [45–47]. In ODIN [45], an open-source digital implementation
of a spiking neural network core is presented. ODIN’s design only supports one physi-
cal neuron core, but many (virtual) neurons can be simulated. Neuron states, as well as
synaptic weights, are stored in an on-chip SRAM memory. A state machine takes care
of time-multiplexing multiple virtual neurons and executes them in one physical core
sequentially. This choice results in a low-area and low power design with the number
of events per Synaptic Operation (SOP), which is in the order of ∼10 pJ/SOP at about
100 Mhz. ODIN has demonstrated an accuracy of 91.9% on MNIST using 4-bit synapses and
one physical neuron simulating 256 virtual neurons. In contrast, our spiking PE achieved
99.3% accuracy on MNIST—a significantly better result achieved through the fully param-
eterized digital implementation tailored to high-level abstraction designs. The behavior
agreement on application and RTL level allows the application specialist to optimize the
number of bits per synaptic contact, threshold parameters, and network structure (number
of neurons and layers) for better accuracy with fewer resources. Special attention is paid
to the memory allocation of synaptic weights to provide the required throughput with
minimal resources. 3.5. Evaluation of the Spiking PE Another recent digital ASIC has been presented by Intel [46], and it
is named Loihi neuromorphic processor. The Loihi processor exploits a similar concept
in which a neuron core emulates many (virtual) up to 1024 neurons and many (virtual)
synaptic connections. Time-multiplexing methods exploit timing slack provided by digital
silicon speed and constantly shuffle neuron’s membrane potential and synaptic weight
memory from/to a central memory. The Loihi architecture hosts multi-neuron cores whose
role is the emulation of a part of the network, e.g., a layer, which can exchange spikes J. Low Power Electron. Appl. 2021, 11, 32 22 of 25 22 of 25 asynchronously in a packet-switched form through a network-on-chip (NoC). The advan-
tage of the time-multiplexing approach is a higher neuron and synapse density because
the computational core is implemented only once, and the neuron states and synaptic
weights are stored in very dense memories. However, in contrast to distributed neuron
networks, time-multiplexed networks’ performance and energy efficiency are limited by
memory bandwidth and lack of parallelism. The distributed neurons offer a high degree of
parallelism and reduce the overhead of data movement caused by shuffling of neuron’s
membrane potential in exchange for manageable area overhead. Furthermore, the low-
power functionality can also be achieved with distributed neurons through voltage scaling,
power gating, and clock gating. For these reasons, our spiking PE architecture does not
time-multiplex neurons and exploits a layered organization. Another recent digital spiking
neural network architecture called µBrain has been presented in [47]. The µBrain also phys-
ically implements each neuron and does not use time-multiplexing, achieving microwatt
power consumption for an always-on system of 336 neurons with 19,878 synapses while
performing MNIST or radar-based gesture classification tasks. 4. Conclusions In this work, high-performance and flexible NN accelerator architecture is proposed
and implemented on an FPGA platform. Moreover, DPR is deployed to realize different
NN accelerators with updating only a part of the FPGA implementation. To compute
the layers in different NN architectures, different PEs with having the same interfaces
are designed. Thus, using these PEs with DPR concept makes the design very efficient
particularly for scenarios switching between different NN accelerators in a short period
of time and without implementing the whole project again. In addition, the proposed
architecture and DPR methodology are illustrated with different use cases to explain the
concept in detail. Lastly, the proposed architecture is evaluated by using one of the use
cases (i.e., digit classifier) with comparing other state-of-the-art methods. The results
showed that the proposed digit classifier achieves higher throughput with moderate DSP
and BRAM usage in comparison with previous implementations on FPGAs. Author Contributions: Conceptualization, H.I., N.A. and D.Z.; methodology, H.I., N.A. and D.Z.;
software, H.I. and P.D.; validation, H.I., F.C. and P.D.; formal analysis, H.I. and F.C.; investigation,
H.I.; data curation, H.I. and F.C.; writing—original draft preparation, H.I. and F.C.; writing—review
and editing, H.I., F.C., P.D., N.A. and D.Z.; visualization, H.I. and F.C.; supervision, N.A. and D.Z.;
project administration, H.I.; funding acquisition, H.I. and F.C. All authors have read and agreed to
the published version of the manuscript. Funding: This research was supported in part by The Scientific and Technological Research Council
of Turkey (TUBITAK). This research was also supported in part by the ECSEL Joint Undertaking (JU)
under grant agreement No 826610. The JU receives support from the European Union’s Horizon
2020 research and innovation programme and Spain, Austria, Belgium, Czech Republic, France, Italy,
Latvia, The Netherlands. Data Availability Statement: No new data were created or analyzed in this study. Data sharing is
not applicable to this article. Conflicts of Interest: The authors declare no conflict of interest. References [CrossRef] S.M.; El-Maleh, A. FPGA-based accelerators of deep learning networks for learning and classification
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12. Abbreviations The following abbreviations are used in this manus
ASIC
Application Specific Integrated Circuit
CNN
Convolutional Neural Network
CPU
Central Processing Unit
DMA
Direct Memory Access
DNN
Deep Neural Network
DPR
Dynamic Partial Reconfiguration
ENA
Efficient Neuron Architecture 23 of 25 J. Low Power Electron. Appl. 2021, 11, 32 FDF
FPGA Design Framework
FIFO
First In First Out
FPGA
Field-Programmable Gate Array
GSOPS
Giga Synaptic Operation per Second
ICAP
Internal Configuration Access Port
ISI
Inter Spike Interval
LFSR
Linear-Feedback Shift Register
LIF
Leaky-Integrate-and-Fire Neuron
NN
Neural Network
PCAP
Processor Configuration Access Port
PE
Processing Element
ReLU
Rectified Linear Unit
SNN
Spiking Neural Network
SOP
Synaptic Operation
SRAM
Static Random Access Memory 17.
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English
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New mHealth application software based on myofunctional therapy applied to sleep-disordered breathing in non-compliant subjects
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Sleep science and practice
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RESEARCH Open Access (2020) 4:3 (2020) 4:3 Sleep Science and Practice O’Connor Reina et al. Sleep Science and Practice
https://doi.org/10.1186/s41606-019-0040-8 © The Author(s). 2020 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. New mHealth application software based
on myofunctional therapy applied to sleep-
disordered breathing in non-compliant
subjects Carlos O’Connor Reina1,2*
, Guillermo Plaza3,4, Jose Maria Ignacio-Garcia5,6, Peter Baptista Jardin7,
Maria Teresa Garcia-Iriarte8, Juan Carlos Casado-Morente1,2, Eugenio De Vicente Gonzalez9 and
Agustin Rodriguez-Reina10 Abstract Background: Myofunctional therapy (MT) is a therapeutic option with demonstrated efficacy for treating sleep-
disordered breathing. We assessed the clinical application of a newly developed m-Health App, which aimed to
allow patients with OSAHS (obstructive sleep apnea/hypopnea syndrome) to perform oropharyngeal exercises only
by interacting with their smartphone without no other devices. Methods: We offered to treat the OSAHS of 20 patients with the App for 90 sessions. Inclusion criteria were adult
patients diagnosed with OSAHS who rejected, or could not tolerate or afford treatment for their OSAHS with other
modalities. The App was used by 15 patients, while 5 were noncompliant and then were used as a control group. Polygraphy was performed at the beginning of the study, and after 90 sessions in the App group or after 3 months
in the control group. Data for adherence to treatment, sex, body mass index (BMI), age, O2MIN (oxygen minimal
saturation) and apnea–hypopnea index (AHI) were collected for both groups. Results: 15 (75%) patients showed adherence to the treatment as measured as performing the exercises daily 5
days a week. In patients who performed the exercises, AHI decreased from 25.78 ± 12.6 to 14.1 ± 7,7 (p = 0.002),
Epworth changed from 18,2 ± 1,98 to 14,2 ± 7,7(p = 0,002) and (O2MIN) changed from 84,87 ± 7.02 to 89,27 ± 3,77
(p = 0.0189). In the control group, there was no significant change. Conclusions: To our knowledge, this is the first study reported where a mHealth App based on MT could be by
itself a therapeutic option in selected patients with OSAHS. Keywords: Myofunctional therapy, mHealth, Sleep apnea, Sleep-disordered breathing, Smartpho Background
b
l 4) can evaluate the perform-
ance of the exercises and achievements. A chat
function is available where the patient can contact
the therapist directly. There is a reminder every 5
days if the patient “forgets” to perform the exer-
cises. The English-language version of this App is
now available on Android and the iOS platform via
Google Play and the App Store, respectively. Full in-
formation about the App is provided in a Web Page
https://airwaygym.App/. Therapists can use it to en-
roll and follow up their patients. indications for all treatments depend on the adher-
ence to the treatment and severity of the disease.(-
Mendes et al. 2014) Myofunctional therapy (MT) has become one of the
newest treatments for sleep-disordered breathing.(Ca-
macho et al. 2015a) MT is based on daily exercises
using
oropharyngeal
muscles
in
an
attempt
to
strength them and facilitate opening of the airway. OSAHS originates from the lack of an optimal func-
tion of the dilator muscles of the airway. Therefore,
MT is a therapy designed theoretically to deal with
the
underlying
mechanism
of
this
disease.(Korhan
et al. 2015) MT is based on oropharyngeal exercises
described by diagrams, videos, and instructions from
a myofunctional therapist on a weekly basis. The pa-
tient is instructed to perform the exercises regularly,
for at least 3 months, between 20 to 40 min daily. In
some cases, patients perform exercises by themselves
at home without substantial feedback and without
giving an exact information to the therapist about
their performance of the exercises.(O’Connor Reina
et al. 2018) Most existing mHealth Applications for OSAHS
patient focus on diagnosis of snoring or OSAHS(I-
setta
et
al. 2017),
while
a
few
are
designed
to
promote adherence to treatment to CPAP(Camacho
et al. 2015b). So far, to our knowledge, there are
none focused in the treatment of OSAHS. However
mobile technology could be especially valuable in
treating OSAHS patient because of its potential to
promote
patient
empowerment
and
self-
management(Iftikhar et al. 2017). This App was developed with the most cutting-edge
technologies (e.g., Ionic, Angular provided by Google
and TypeScript by Microsoft) and the most consolidated
software languages (such as HTML5, CSS3, and PHP). The App takes advantage of 3D Touch technology, a
capacity available in the latest Apple devices to accur-
ately measure the pressure that is produced on the mo-
bile screen. Background
b
l morbidities such as cardiovascular and cerebrovascu-
lar diseases. The classic treatment of this disease is
based on dietary measures, losing weight, and exer-
cise, and the use of a continuous positive airway pres-
sure
(CPAP)
machine,
a
device
that
reduces
collapsibility of the upper airway by emitting a flow
of air. Other options are upper airway surgery, treat-
ing the obstacle of the airway or correcting the mus-
cles that do not perform their function properly, and
mandibular advancement devices (MAD), which pro-
trudes the tongue forward to avoid it falling backward
and collapsing the airway. The success rates and Obstructive sleep apnea/hypopnea syndrome (OSAHS)
has become one of the more serious health problems
worldwide.(Mendes et al. 2014) Because of its rela-
tionship with obesity, its prevalence and incidence
have been increasing and it is associated with severe * Correspondence: coconnor@us.es
1Cohead Department Otorhinolaryngology, Hospital Quiron Salud Marbella,
Avenida Severo Ochoa 22, 29603 Marbella, Spain
2Cohead Department of Otorhinolaryngology, Hospital Quiron Salud Campo
de Gibraltar, Cádiz, Spain
Full list of author information is available at the end of the article * Correspondence: coconnor@us.es
1Cohead Department Otorhinolaryngology, Hospital Quiron Salud Marbella,
Avenida Severo Ochoa 22, 29603 Marbella, Spain
2Cohead Department of Otorhinolaryngology, Hospital Quiron Salud Campo
de Gibraltar, Cádiz, Spain
Full list of author information is available at the end of the article O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 O’Connor Reina et al. Sleep Science and Practice Page 2 of 10 Page 2 of 10 therapists. The novelty of this App is that it is the
first in the health-care market where the patient can
interact directly with the smartphone without any
other device, which provides feedback about the ef-
ficacy of the exercises performed and focused on
sleep apnea disease. There are nine exercises based
on MT that attempt to enhance the tonicity of the
various
muscles
involved
in
the
pathogenesis
of
OSAHS. Before every exercise, there is an animated
gif demonstration that shows the patient how to
perform the exercise (Figs. 1a–c). App’s users can
follow the development of their activity daily over
time (Figs. 2a–c). At the conclusion of each exer-
cise, the patient receives feedback about the success
of their performance with a point score (Fig. 3). When the patient finishes the exercises, they are
saved on networked online storage (in the Cloud)
and a therapist (Fig. Background
b
l This App complies with regulation 2002/58/
CE and (UE) 2016/679 about Data protection. One of the best treatments for OSAHS is perform-
ing exercises and reducing weight.(KCC 2009) Be-
cause time is limited, we consider that there will be a
greater probability of patients performing the exer-
cises if they are able to do so while sitting comfort-
ably and watching television. Therefore, we designed
and developed a novel mHealth Application software
(App) to promote oropharyngeal exercises while inter-
acting with a smartphone. In this article, we intro-
duce
this
APP
and
we
present
results
regarding
adherence to the App and its effectiveness in a group
of patients with OSAHS. The main objective of these exercises was to increase
the tone of the extrinsic muscles of the tongue (genio-
glossus,hyoglossus,styloglossus and palatoglossus). The exercises are based on those described by Gui-
maraes in 2009(KCC 2009), adapted to obtain a feedback
with the phone. Due to hygienical reasons we recommend to cover the
screen with cling film or a hypoallergenic plastic wrap in
all exercises. Application function, and availability Exercise 1. Snake With your elbows touching your
body, hold the phone with both hands as you do when
holding a mirror in front of your face. This App is a smartphone App created in a collaboration
between the Sleep Units of Hospital Quirónsalud Mar-
bella and Campo de Gibraltar and their Engineering In-
formatics Departments and developed by Apnea Bye
company. It was presented last November 2018. Stick your tongue out and press the screen with it for
5 s and release. The tongue should not touch your teeth
or lips. We can think of this App as a portable gym with
athletes instead of patients and trainers instead of Repeat the exercise 15 times. O’Connor Reina et al. Sleep Science and Practice (2020) 4:3
Page 3 of 10 O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 O’Connor Reina et al. Sleep Science and Practice Page 3 of 10 (2020) 4:3 Fig. 1 a–c Exercise snake where patient should touch with his tongue the screen press it with suitable rhythm and strength and receive a
positive sound of feedback. Fig. 1 a–c Exercise snake where patient should touch with his tongue the screen press it with suitable rhythm and strength and receive a
positive sound of feedback. Fig. 1 a–c Exercise snake where patient should touch with his tongue the screen press it with suitable rhythm and strength and receive a
positive sound of feedback. Exercise 2. Pressure with your chin Fit your jaw 2 fin-
ger above the center of your phone. If you’ve got a beard
place your finger between your phone and your jaw. Exercise 2. Pressure with your chin Fit your jaw 2 fin-
ger above the center of your phone. If you’ve got a beard
place your finger between your phone and your jaw. Stick out your tongue and press on the square at the
bottom. The tongue must not touch your teeth or lips. Stick out your tongue and press on the square at the
bottom. The tongue must not touch your teeth or lips. Put your tongue down and keep pressing the screen
for 5 s, close your mouth and rest. Open your mouth and keep pushing the screen for 5 s,
close your mouth and rest. While doing the exercise say
/a/. Repeat the exercise 15 times. Repeat the exercise 15 times. Exercise 5. Application function, and availability Tongue left cheek Press with the tip of
your tongue the inside of the left cheek. Allow the
screen of your phone to touch gently your cheek. Exercise 3. Chameleon up Place the phone at a certain
distance in front of you, as if it were a mirror. Exercise 3. Chameleon up Place the phone at a certain
distance in front of you, as if it were a mirror. Contract the muscles of your cheek while pushing with
the tongue towards the cheek. Keep the pressure for 5 s. Stick out your tongue and press on the square at the
bottom. The tongue must not touch your teeth or lips. Repeat the exercise 15 times. Put your tongue up and keep the pressure on the
screen 5 s, close your mouth and rest. Repeat the exercise 15 times. Exercise 6. Tongue right cheek Press with the tip of
your tongue the inside of the right cheek. Allow the
screen of your phone to touch gently your cheek. Exercise 4. Chameleon down Due to hygienical reasons
we recommend to cover the screen with cling film or a
cleaning wipe. Contract the muscles of your cheek while pushing with
the tongue towards the cheek. Keep the pressure for 5 s. Place the phone at a certain distance in front of you,
as if it were a mirror. Repeat the exercise 15 times. O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 Page 4 of 10 O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 Fig. 2 a–c Evolution of the performing of the exercises during the week with proper information to the patient with the degree of accuracy Exercise 7. Pressure under chin Hold the phone with
both hands with the screen facing up. If you’ve got a
beard place your finger between your phone and your
jaw. were talking on the phone but with the screen to the
outside. Turn your jaw towards the phone and keep the pres-
sure with your index finger on the screen for 5 s. Say /i/
while doing it. Sitting upright put the phone under your chin so there
is no contact with the chin. Release the pressure and rest. Repeat the process 15
times. Bend your head forward like you are going to drink
from a cup and contact your chin on the phone. Keep the contact and move your head from side to
side during 10 s with your mouth closed. Lift your head
and rest. Exercise 9. Right mandibular pressure
Bring the
phone closer to the origin of the jaw on the right side. Fig. 3 Score achieved by patient after performing exercise American
Academy
of
Sleep
Medicine
(AASM)
Manual for the Scoring of Sleep and Associated
Events and were reviewed by certified physicians
(Adams et al., n.d.). Apnea was defined as an ab-
sence of airflow for a period lasting at least 10 s
and hypopnea was defined as at least a 30% reduc-
tion in airflow associated with a 4% or greater de-
crease in oxygen saturation. The AHI was defined
as the number of episodes of apnea and hypopnea
per hour of Total Sleep Time (American Academy
of Sleep Medicine 2007). Finally, the adherence to
treatment (those who performed the exercise at
least 5 days a week) were collected. Then, patients
were reevaluated after 90 sessions with the App
with the same tests, or after 3 months in the con-
trol group by the same physician who blindly evalu-
ated
the
results
achieved
from
each
group. All
patients
were
instructed
by
the
same
therapist
about the use of the App prior to enroll in the
study. Release the pressure and rest. Repeat the process 15
times. American
Academy
of
Sleep
Medicine
(AASM)
Manual for the Scoring of Sleep and Associated
Events and were reviewed by certified physicians
(Adams et al., n.d.). Apnea was defined as an ab-
sence of airflow for a period lasting at least 10 s
and hypopnea was defined as at least a 30% reduc-
tion in airflow associated with a 4% or greater de-
crease in oxygen saturation. The AHI was defined
as the number of episodes of apnea and hypopnea
per hour of Total Sleep Time (American Academy
of Sleep Medicine 2007). Finally, the adherence to
treatment (those who performed the exercise at
least 5 days a week) were collected. Then, patients
were reevaluated after 90 sessions with the App
with the same tests, or after 3 months in the con-
trol group by the same physician who blindly evalu-
ated
the
results
achieved
from
each
group. All
patients
were
instructed
by
the
same
therapist
about the use of the App prior to enroll in the
study. Release the pressure and rest. Repeat the process 15
times. Exercise 3. Chameleon up Place the phone at a certain
distance in front of you, as if it were a mirror. As if you were talking on the phone but with the screen
to the outside. Repeat the exercise 15 times. Turn your jaw towards the phone and keep the pres-
sure with your index finger on the screen for 5 s. Say /i/
while doing it. Exercise 8. Left mandibular pressure Bring the phone
closer to the origin of the jaw on the left side. As if you Page 5 of 10 O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 Fig. 3 Score achieved by patient after performing exercise Patients and methods This preliminary cross-sectional study Approved by
the local Ethics Committee AWGAPN-2019-01. Study
started January 2019 and all patients were studied in
a comparable frame of time. The inclusion criteria
were adult patients diagnosed with OSAHS by poli-
graphy and were non-compliant with any therapy and
had a mobile. The exclusion criteria were patients
with previous surgery in the UA, UA malformation
(i.e., significant tongue tie). All patients gave informed
consent. Data including sex, age, body mass index (BMI),
Epworth Sleepiness Scale (ESS), Minimal Oxigen Satur-
ation (O2MIN) and apnea–hypopnea index (AHI) ob-
tained by poligraphy (Sleep & Go, Sibelmed 2018) . All sleep studies were manually interpreted by a sleep
technician according to the standard criteria of the O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 Page 6 of 10 O’Connor Reina et al. Sleep Science and Practice Fig. 4 Information obtained by Therapist about the accuracy and adherence of the performance of the exercises performed by patient Statistical analysis included problems with the temporomandibular joint (1
case), irritation of the tongue (1 case), and fatigue (3 cases). included problems with the temporomandibular joint (1
case), irritation of the tongue (1 case), and fatigue (3 cases). The study population was mostly male, middle-aged,
and overweight. Twelve male (80%) and three women
had
adherence
to
the
treatment,
meanwhile
three
women (60%) and two men (40%) were non-compliant. The average aged of the compliant group were 36 ± 13,5,
on the other hand, 53,2 ± 7 years had the non-adherence
group. There were no significant differences in the IMC
of both groups (24,8 ± 2,8 to 24,7 p = 0,42, 25,3 ± 2,7 to
26,3 ± 3,3 p = 0,07 respectively). Quantitative variables were assessed by calculating the
arithmetic mean and standard deviation and group of
differences were assessed using a two-sample paired t
test, or using a Mann–Whitney rank sum test if the vari-
able was not normally distributed. P < 0.05 was consid-
ered
significant. IBM
SPSS
Statistics
for Windows
software (version 20; IBM Corp, Armonk, NY, USA) was
used for statistical analysis. The study population was mostly male, middle-aged,
and overweight. Twelve male (80%) and three women
had
adherence
to
the
treatment,
meanwhile
three
women (60%) and two men (40%) were non-compliant. The average aged of the compliant group were 36 ± 13,5,
on the other hand, 53,2 ± 7 years had the non-adherence
group. There were no significant differences in the IMC
of both groups (24,8 ± 2,8 to 24,7 p = 0,42, 25,3 ± 2,7 to
26,3 ± 3,3 p = 0,07 respectively). Discussion As the prevalence of obesity increases worldwide,
OSAHS
increases
proportionally. We
live
in
a
world where effort and involvement are being for-
gotten. OSAHS treatments that aim for the com-
fort of the patient overlook that this disease is
associated with sedentarism and passivity. Exercise
is one of the best treatments for this disease(Ifti-
khar et al. 2017). Time is limited; therefore, this
App was provided in an attempt to employ time
effectively to resolve this disease. People spend a
significant amount of time using their smartphones
(Andrews et al. 2015) as they are an excellent
companion and allow communication with social
media but also the use of Apps may allow better
use of a person limited time, in this case by the
performance of MT exercise. The results of the present study can be compared with
other therapeutic options currently available.(Iftikhar et al. 2017) This App has the advantage of being affordable
worldwide as a smartphone App and treats the etiology of
the disease. For patients who cannot afford other treat-
ment for their disease, this App presents an affordable op-
tion. While
it
may
not
substitute
for
established
treatments, in those patients where there is no other Understanding the reasons why some people with
OSAHS have poor muscle effectiveness are incomplete. The data are summarized in Tables 1 and 2 and
Fig. 5. contributing factors. In general, people with OSAHS
have poorer muscle effectiveness than non-OSAHS con-
trols (Dotan et al. 2015). Apps for smartphones and tablets have become indis-
pensable, especially for young generations; however, in-
creasing use of Apps in the middle-aged and elderly
population is observed, thus arguing for a common use
across generation borders. The estimated number of
mobile devices will exceed 2.16 billion in 2016. Over 2.2
million Apps are available in the Google Play store®, and
about 1.8 million Apps are available in the Apple App
Store®. Google and Apple distribute nearly 70,000 Apps
each in the category Health and Fitness, and about 33,
000 and 46,000 each in medical Apps. The WHO defines
these tools under the label “mHealth” or “eHealth” as
“medical and public health practice supported by mobile
devices, such as mobile phones, patient monitoring de-
vices, personal digital assistants, and other wireless devi-
ces”(Vogel et al. 2017). Results We enrolled 20 consecutive patients diagnosed with
OSAHS by physicians in our Sleep Unit. These patients
had no treatment for their disease because 10 patients
could not tolerate CPAP, 5 patients could not afford a
MAD, and 5 patients could not tolerate CPAP or MAD,
and rejected surgery. None had the option of a weekly visit
to a myofunctional therapist because of geographical re-
moteness. The App was used by 15 patients who complied
with its functions. Reasons for rejection of the App In the patients who performed the exercises the
AHI decreased from 25.78 ± 12.6 to 14.1 ± 7,7 (p =
0.002) t = 0,83, Epworth changed from 18,2 ± 1,98 to
14,2 ± 2,7(p = 0,002) t = 4,9 and (O2MIN) changed from
84,87 ± 7.02 to 89,27 ± 3,77 (p = 0.0189) t = −2,7. In
the control group, there was no significant change in
the AHI (31.460 ± 12.6 to 32.90 ± 12.8, p = 0.4065) t =
0,92 or (O2MIN) 83,9 ± 9 to 82 ± 9.4, (p = 0.29)t = 1 or
Epworth 18,8 ± 1,1 to 19,4 ± 1,95, (p = 0.208) t = −1,5. Page 7 of 10 O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 O’Connor Reina et al. Sleep Science and Practice The data are summarized in Tables 1 and 2 and
Fig. 5. Discussion However, poor coordination of the neural drive to the
various upper airway muscles during sleep, mechanically
inefficient muscle fibre orientation (due to excess fat or
muscle hypertrophy), or muscle fiber type changes
resulting in greater propensity for fatigue could all be Table 1 All results obtained in both groups pre and post use the app
Patient
Age
Sex
BMI (pre)
BMI (post)
AHI (pre)
AHI (post)
SatO2 Min
Sat O2Min (post)
ESS
ESS (POST)
ADH
1
23
M
24.2
25.1
17.8
14.2
83%
85%
16
14
Y
2
19
F
23.1
22.9
16.2
13.5
93%
94%
18
13
Y
3
34
M
26.2
26
21.9
21
85%
84%
19
21
Y
4
37
M
31.8
32
14.5
10.4
92%
93%
18
14
Y
5
17
M
23.2
21.9
12.6
9.3
89%
91%
16
13
Y
6
41
F
21.3
20.9
13.2
12.1
92%
92%
17
16
N
7
28
M
26
26.2
15.7
9.1
91%
90%
17
12
Y
8
32
M
24.3
24.1
28.8
17.7
86%
90%
19
13
Y
9
29
M
21.9
21.5
15.4
12.3
88%
88%
18
15
Y
10
25
M
22.1
22
40.2
15,9
74%
92%
21
15
Y
11
46
M
25,5
24,5
42,2
24,5
69%
86%
20
14
Y
12
50
M
22,1
22
26,6
18,3
88%
89%
15
14
Y
13
62
F
25,5
24,5
17,8
3,1
91%
93%
16
16
Y
14
55
M
24
24.2
24.1
4.5
87%
92%
19
10
Y
15
43
M
23.2
24.5
55
7
78%
91%
21
11
Y
16
47
F
30
29.5
38
32
79%
81%
21
18
Y
17
52
M
28
29.9
24
31
87%
86%
19
20
N
18
56
M
24.3
26.2
37
35.2
84%
85%
20
21
N
19
62
F
25.3
26.7
39
41
81%
80%
19
20
N
20
55
F
27.6
28.2
44.1
45.2
68%
67%
19
20
N O’Connor Reina et al. Discussion Sleep Science and Practice (2020) 4:3 Page 8 of 10 Table 2 Main grouped results
PATIENT
SEX
AGE
BMI PRE
BMI POST
IAH PRE
IAH POST
SAT O2 MIN
PRE
SAT O2 MIN
POST
ESS PRE
ESS POST
AD (15)
3 F,12 M
(80%)
36 ± 13,
5
24,8 ± 2,
83
24,7 ± 2891
P = 0,42 T = 0,
831
25.78 ± 12.6
14.1 ± 7,7
(p = 0.002)
T = 0,83
84,87 ± 7.02
9,27 ± 3,77
(p = 0.0189)
T = -2,7
18,2 ± 1,
98
14,2 ± 2,7
(p = 0,002)
T = 4,92
NON AD
(5)
3F,2M
(40%)
53,
2 ± 7,7
25,3 ± 2,
7
26,3 ± 3,3
P = 0,07
T = -2,45
31.460 ± 12.6 32.90 ± 12.8,
(p = 0.4065)
T = -0,927
83,9 ± 9
82 ± 9.4,
(p = 0.29)
T = 1
18,8 ± 1,
1
19,4 ± 1,95,
(p = 0.208). T = -1,5
Age years, M male, F female, BMI BODY MASS INDEX, AHI APNEA HIPOAPNEA INDEX, O2 Min Minimal o2 adherence, ESS Epworth Sleepiness scale,
ADH ADHERENCE choice, it presents a reasonable alternative. For patients
who are unable to attend weekly sessions with a myofunc-
tional therapist, the App provides a practical substitute. et al. 2018). In our most recent manuscript, we showed
a correlation between low IOPI tongue scores and the
presence of tongue collapse showed in the DISE of se-
vere OSA patients (O’Connor Reina n.f.). We believe
that patient with lowest IOPI scores are the best candi-
dates for our App. Under our experience, improving
IOPI scores obtained using our App is the best guideline
that exercises are being done correctly. We are perform-
ing a clinical trial to assure this matter. We have to point out that some patients have a small
variation in their results and that could be considered as a
night-to-night intra-individual variability due the small
sample of this study. This could generate a bias that should
be corrected in future investigations with larger samples. In our series, patients 10, 11, 13,14 and 15 showed a
significant improvement in their IAH. We are working
with IOPI(Adams et al., n.d.) as a tool to measure the
tongue strength prior to use the App. These patients
present
the
lowest
values
of
all
the
group. Discussion We
hypothesize that hypotonia is the main phenotype of
their OSAHS as suggested by Carberry et al.(Carberry Despite our small sample it is significant the age differ-
ence between the two groups, we think it could be asso-
ciated with the prevalence of the use of smartphones
between young people. Adherence to the described treatment is reasonable
and
can
be
compared
to
another
reported Fig. 5 Box plot showing statistical significant differences between study and control groups Fig. 5 Box plot showing statistical significant differences between study and control groups O’Connor Reina et al. Sleep Science and Practice (2020) 4:3 O’Connor Reina et al. Sleep Science and Practice Page 9 of 10 Page 9 of 10 noncompliance with standard treatments,(Burks et al. 2016) we consider this report worthwhile. series.(Hevener and Hevener 2016) There may be se-
lection bias because other therapeutic options have
failed for these patients. We are conducting now a
study with general population of patients with severe
OSAHS. This treatment also may increase compliance
with other established treatments as has been re-
ported previously for MT.(Diaféria et al. 2017) A limi-
tation of this study is that the patient must have a
smartphone. Reasons for rejection of the App included problems
with the temporomandibular joint (1 case), irritation of
the tongue (1 case), and fatigue (3 cases). We do not rec-
ommend this App in patients with temporomandibular
joint disorders because they can worsen with the exer-
cises. On the other side, we always recommend to use
an hypoallergenic plastic wrap to cover the screen before
contacting with the tongue in order to avoid adverse
reactions. The future treatment and diagnosis of this disease will
be based on the use of Apps(Lim and Pack 2017), as
shown by Kim et al. (2019) We currently recommend
that our patients use the App simultaneously with other
daily diagnosis tools(Camacho et al. 2015b) to provide
objective feedback of their progress in their activity. One
of the great advantages of this App is the possibility of
contacting a therapist who can observe the daily evolu-
tion of the patients’ exercises. This provides substantial
information for therapeutic decisions based on the evo-
lution of their disease. These exercises are based in those
presented in the literature and adapted to interact with
the phone(KCC 2009). Abbreviation ADH: adherence; AHÍ: apnea–hypopnea index; App: Application; BMI: Body
mass index; CE: European Community; CPAP: Continuous Airway pressure;
ESS: Epworth Sleepiness Scale; IOPI: Iowa Oral Performance Instrument;
MAD: Mandibular Advanced Device; MT: myofunctional therapy;
O2MIN: oxygen minimal saturation; OSAHS: obstructive sleep apnea/hypopnea
syndrome; UE: European Union Discussion The duration of the study and
the frequency of exercises are based in other myofunc-
tional studies (Guimarães et al. 2009; Ieto et al. 2015;
Nemati et al. 2015). We understand there is not enough
evidence to validate any exercise due the reduced litera-
ture about this matter(Camacho et al. 2015a). Goswami et al.(Goswami et al. 2019) reported recently
a randomized controlled trial with snorer patients using
a smartphone-based delivery with oropharyngeal exer-
cises created during vocalization. They excluded patients
with OSAHS. They present significant reduction in snor-
ing and ESS after 8 weeks of using the device. g
g
We have recently reported(O’Connor Reina et al. 2019) the first case of a OSAHS patient with a floppy
trap door epiglottis cured using this App. His initial ESS
was 22, AHI was 31.2 and minimal O2 saturation was
81.3%. After 90 sessions with the App, AHI was reduced
to 17.2, minimal O2 saturation improved to 85.1%, and
his Epworth sleepiness scale improved to 15. The floppy
epiglottis disappeared as was confirmed by final DISE. Being the first case reported of a floppy epiglottis Appar-
ently cured using an MT-based smartphone App, we do
not have a clear scientific reason to explain our finding. We understand that MT increases tone in the muscles
of the tongue, also achieving a reduction in its fat
content. One of the negative points of using MT to treat sleep-
disordered breathing is that patients need to perform the
exercises beyond a 3-month period and if they stop the
exercises, AHI might return to previous levels.(Guimar-
ães et al. 2009) Daily CPAP or MAD treatments do not
alter the AHI of patients with OSAHS if the patient is
not adherent to the treatment, and similarly AHI might
return to previous levels if therapy is stopped. In our
present series, patients did not show any difficulty chan-
ging their entire nights’ treatment for 20 min a day of
oropharyngeal exercises. Their feedback is that they slept
much better when they use the App. We have seen in
these patients that improvement in their AHI increased
if they maintain exercises beyond 3 months. Further in-
vestigations are needed to support this point. These changes are supposed to “tense” the glossoepi-
glottic folds, and this tension might act like suspenders
that prevent the collapse of the epiglottis. Further stud-
ies are needed to support this suggestion. Conclusion To our knowledge, this is the first study reported where
an mHealth App based on MT could be by itself a thera-
peutic
option
for
selected
patients
with
OSAHS. Randomized-controlled trials and cost-effectiveness in-
vestigations are needed to support this concept. There are serious concerns for ethical, security, priv-
acy, and connectivity issues surrounding Apps that
promise to cure diseases(De La Vega and Miró 2014) in
the mHealth realm. This App is focused on helping pa-
tients to perform exercises with their oropharyngeal
muscles with the aim of increasing their tone. This study
is anecdotal compared with the evidence existing for
other therapeutic options. The present study is prelimin-
ary, but quantitative improvement of OSAHS may be
supported with further reports on the use of this App. It
is difficult to present an App as a new therapeutic option
for
OSAHS,
but
considering
the
prevalence
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declaration and its later amendments or comparable ethical standards. Informed consent: Informed consent was obtained from all individual
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adherence to continuous positive airway pressure treatment. Sleep Breath. 2017;21(2):387–95. https://doi.org/10.1007/s11325-016-1429-6. Funding This study has no funds. Received: 1 July 2019 Accepted: 9 December 2019 O’Connor Reina C, García Iriarte MT, Casado-Morente JC, et al. New app “Apnea
Bye” increases adherence in myofunctional therapy to treat sleep disorder
breathing. Otolaryngol Neck Surg. 2018;159(1_suppl):P326–7. https://doi.org/
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MADs, exercise-training, and dietary weight loss for sleep apnea: a network
meta-analysis. Sleep Med. 2017 Feb;30:7–14. https://doi.org/10.1016/j.sleep. 2016.06.001. Dr. C O’Connor-Reina and Dra M Garcia-Iriarte are the otorhinolaryngologists
designers of this App and Mr. A. Rodriguez-Reina is the informatic engineer
who develop the App. G. Plaza declares no conflict of interest. Isetta V, Torres M, González K, et al. A new mHealth application to support
treatment of sleep apnoea patients. J Telemed Telecare. 2017 Jan;23(1):14–8. https://doi.org/10.1177/1357633X15621848. J. Ignacio-Garcia declares no conflict of interest. P. Baptista Jardin declares no conflicto of interest. JC Casado-Morente declares no conflict of interest. Guimaraes KCC. Apneia e Ronco. Tratamiento Miofuncional Orofacial:tratamento
miofuncional orofacial. In: Pulso, ed. San Jose dos Campos; 2009, pp 53–63. Kim JW, Ryu B, Cho S, et al. Impact of personal health records and wearables on
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miofuncional orofacial. In: Pulso, ed. San Jose dos Campos; 2009, pp 53–63. E. De Vicente Gonzalez declares no conflict of interest. miofuncional orofacial. In: Pulso, ed. San Jose dos Campos; 2009, pp 53–63. Kim JW, Ryu B, Cho S, et al. Impact of personal health records and wearables on
health outcomes and patient response: three-arm randomized controlled
trial. JMIR Mhealth Uhealth. 2019;7(1):e12070. https://doi.org/10.2196/12070. Authors’ contributions Dr. Carlos O’Connor Reina, Dr. Peter Baptista Jardin and Dr. Guillermo Plaza
had full access to all the data in the study and takes responsibility for the
integrity of the data and the accuracy of the data analysis. Dr. Carlos O’Connor Reina, Dr. Peter Baptista Jardin and Dr. Guillermo Plaza
had full access to all the data in the study and takes responsibility for the
integrity of the data and the accuracy of the data analysis. Dra Garcia Iriarte, Dr. De Vicente and Dr. Casado special contribution in data
collection and writing and editing assistance. Dr. Ignacio-Garcia assisted in the translation and statistics. Mr. Agustin Rodriguez assessed in the technical issues of the App, Dotan Y, Pillar G, Schwartz AR, Oliven A. Asynchrony of lingual muscle
recruitment during sleep in obstructive sleep apnea. J Appl Physiol. 2015;
118(12):1516–24. https://doi.org/10.1152/japplphysiol.00937.2014. Dra Garcia Iriarte, Dr. De Vicente and Dr. Casado special contribution in data
collection and writing and editing assistance. Goswami U, Black A, Krohn B, et al. Smartphone-based delivery of oropharyngeal
exercises for treatment of snoring: a randomized controlled trial. Sleep
Breath. 2019;23:243. https://doi.org/10.1007/s11325-018-1690-y. Dr. Ignacio-Garcia assisted in the translation and statistics. Mr. Agustin Rodriguez assessed in the technical issues of the App, Mr. Agustin Rodriguez assessed in the technical issues of the App, reath. 2019;23:243. https://doi.org/10.1007/s11325-018-1690-y. Guimarães KC, Drager LF, Genta PR, Marcondes BF, Lorenzi-Filhoy G. Effects of
oropharyngeal exercises on patients with moderate obstructive sleep apnea
syndrome. Am J Respir Crit Care Med. 2009;179(10):962–6. https://doi.org/10. 1164/rccm.200806-981OC. Author details
1 1Cohead Department Otorhinolaryngology, Hospital Quiron Salud Marbella,
Avenida Severo Ochoa 22, 29603 Marbella, Spain. 2Cohead Department of
Otorhinolaryngology, Hospital Quiron Salud Campo de Gibraltar, Cádiz, Spain. 3Head Otorhinolaryngology Department, Hospital Sanitas la Zarzuela, Madrid,
Spain. 4Head Department Otorhinolaryngology, Hospital Universitario
Fuenlabrada, Fuenlabrada, Spain. 5Head Neumology Department, Hospital
Quiron Salud Marbella, Marbella, Spain. 6Head Neumology Department,
Hospital Quiron Salud Campo de Gibraltar, Cádiz, Spain. 7Otorhinolaryngology Department, Clinica Universitaria de Navarra,
Pamplona, Spain. 8Otorhinolaryngology Department, Hospital la Merced
Osuna Sevilla, Sevilla, Spain. 9Department Otorhinolaryngology, Hospital
Universitario Miguel Servet, Zaragoza, Spain. 10Department Informatic &
Engineering, Apnea Bye S.L, Sevilla, Spain. Korhan I, Gode S, Midilli R, Basoglu OK. The influence of the lateral pharyngeal
wall anatomy on snoring and sleep apnoea. J Pak Med Assoc. 2015;65(2):
125–30. Lim DC, Pack AI. Obstructive sleep apnea: update and future. Annu Rev
MedAnnu Rev Med. 2017;68:99–112. https://doi.org/10.1146/annurev-med-
042915-102623. Mendes F, Marone S, Duarte B, Arenas A. Epidemiologic profile of patients with
snoring and obstructive sleep apnea in a university hospital. Int Arch Mendes F, Marone S, Duarte B, Arenas A. Epidemiologic profile of patients with
snoring and obstructive sleep apnea in a university hospital. Int Arch
Otorhinolaryngol. 2014;18(02):142–5. https://doi.org/10.1055/s-0033-1359309. Nemati S, Gerami H, Soltanipour S, Saberi A, Moghadam SK, Setva F. The effects
of oropharyngeal–lingual exercises in patients with primary snoring. Eur Arch
Otorhinolaryngol. 2015;272(4):1027–31. https://doi.org/10.1007/s00405-014-
3382-y. Received: 1 July 2019 Accepted: 9 December 2019 Availability of data and materials Hevener B, Hevener W. Continuous positive airway pressure therapy for
obstructive sleep apnea: maximizing adherence including using novel
information technology–based systems. Sleep Med Clin. 2016;11(3):323–9. https://doi.org/10.1016/j.jsmc.2016.04.004. The raw data on which the results in the current study are based may be
requested from the corresponding author upon reasonable request. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations. Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations. Camacho M, Robertson M, Abdullatif J, et al. Smartphone apps for snoring. J
Laryngol Otol. 2015b;129(10):974–9. https://doi.org/10.1017/
S0022215115001978. Camacho M, Robertson M, Abdullatif J, et al. Smartphone apps for snoring. J
Laryngol Otol. 2015b;129(10):974–9. https://doi.org/10.1017/
S0022215115001978.
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Review of ‘Audubon’s Birds of America’ at the National Museum of Scotland
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Peter Adkins, Review of ‘Audubon’s Birds of America’ at the National
Museum of Scotland. 19: Interdisciplinary Studies in the Long Nineteenth
Century, 19 Live (2022) <https://doi.org/10.16995/ntn.8974> Peter Adkins, Review of ‘Audubon’s Birds of America’ at the National
Museum of Scotland. 19: Interdisciplinary Studies in the Long Nineteenth
Century, 19 Live (2022) <https://doi.org/10.16995/ntn.8974> Peter Adkins, Review of ‘Audubon’s Birds of America’ at the National
Museum of Scotland. 19: Interdisciplinary Studies in the Long Nineteenth
Century, 19 Live (2022) <https://doi.org/10.16995/ntn.8974> 1 At the time of its publication it was the largest paper commercially available and only two paper mills in Britain
produced it.
2 Antoine Traisnel, Capture: American Pursuits and the Making of a New Animal Condition (Minneapolis: University of
Minnesota Press, 2020), p. 39.
3 Duncan Macmillan, ‘Art Reviews: Audubon’s Birds of America | Andrew Mackenzie | GPS Showcase’, Scotsman, 24
February 2022 <https://www.scotsman.com/arts-and-culture/art/art-reviews-audubons-birds-of-america-andrew-
mackenzie-gps-showcase-3585318> [accessed 13 June 2022]. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland Peter Adkins This is a review of the exhibition ‘Audubon’s Birds of America’, held at the National Museum of
Scotland, Edinburgh from 12 February to 8 May 2022. 19: Interdisciplinary Studies in the Long Nineteenth Century is a peer-reviewed open access journal published by the Open
Library of Humanities. © 2022 The Author(s). This is an open-access article distributed under the terms of the Creative
Commons Attribution 4.0 International License (CC-BY 4.0), which permits unrestricted use, distribution, and reproduction
in any medium, provided the original author and source are credited. See http://creativecommons.org/licenses/by/4.0/. OPEN ACCESS 19: Interdisciplinary Studies in the Long Nineteenth Century is a peer-reviewed open access journal published by the Open
Library of Humanities. © 2022 The Author(s). This is an open-access article distributed under the terms of the Creative
Commons Attribution 4.0 International License (CC-BY 4.0), which permits unrestricted use, distribution, and reproduction
in any medium, provided the original author and source are credited. See http://creativecommons.org/licenses/by/4.0/. OPEN ACCESS 19: Interdisciplinary Studies in the Long Nineteenth Century is a peer-reviewed open access journal published by the Open
Library of Humanities. © 2022 The Author(s). This is an open-access article distributed under the terms of the Creative
Commons Attribution 4.0 International License (CC-BY 4.0), which permits unrestricted use, distribution, and reproduction
in any medium, provided the original author and source are credited. See http://creativecommons.org/licenses/by/4.0/. OPEN ACCESS 2 Sent to subscribers between 1827 and 1838, the four-volume Birds of America is widely
considered one of the landmark works of American ornithological art. Even today, it
remains an imposing proposition: printed on paper that measures 99 by 66 centimetres
(a size fittingly called ‘double elephant’), the work presents 435 hand-coloured prints
that look to capture their subjects in life-size scale and often in situ, whether pouncing
on a rabbit or stripping corn ears.1 A kind of giant index of America’s avian life, the book
aimed to display every bird species in North America. Sent to subscribers between 1827 and 1838, the four-volume Birds of America is widely
considered one of the landmark works of American ornithological art. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland Even today, it
remains an imposing proposition: printed on paper that measures 99 by 66 centimetres
(a size fittingly called ‘double elephant’), the work presents 435 hand-coloured prints
that look to capture their subjects in life-size scale and often in situ, whether pouncing
on a rabbit or stripping corn ears.1 A kind of giant index of America’s avian life, the book
aimed to display every bird species in North America. Not shy of publicizing the ambition of the project, its author, John James Audubon
(1781–1851), was something of a raconteur. Presenting himself as an American
woodsman when looking to authenticate his knowledge of the continent (and raise
money for his projects), Audubon was French Haitian by birth, growing up in France
and emigrating to the USA at the age of eighteen to escape conscription in Napoleon’s
army. Forging his passport on entering the country, purporting to have been born in
Louisiana, Audubon’s precariously financed career as an artist and ornithologist was
at least partially built on performing a certain idea of American identity.2 A self-taught
artist, he also taught himself how to outwardly embody the rustic ideals of his adopted
country, especially when around those to whom such an identity was exotic and alluring. Indeed, key to Audubon’s ability to produce Birds of America was winning backers
and collaborators in Britain, where he found a warmer reception than in the USA due, in
part, to his willingness to play up to the imagined idea of the frontiersman. Edinburgh,
in particular, played a vital role in getting Birds of America off the ground. Finding
himself welcomed by the city’s cognoscenti after exhibiting his paintings in the Royal
Institution, Audubon was approached by local engraver William Lizars, who proposed
he might engrave and publish the images in a book.3 It is this connection with Scotland’s
capital that seems to have provided the impetus for his recent major exhibition at the
National Museum of Scotland. Presenting visitors with the chance to see Audubon’s work up close and in person,
‘Audubon’s Birds of America’ ran from 12 February to 8 May 2022, showcasing ‘46
unbound prints from National Museums Scotland’s collection, most of which have
never been on display before, as well as a rare bound volume of the book’. The latter, 3 3 incidentally, was not taken from the museum’s extensive Audubon collection but was
on loan from the Mitchell Library in Glasgow. 4 ‘Audubon’s Birds of America’, National Museums Scotland Press Release, 20 October 2021 <https://media.nms.ac.uk/
news/audubons-birds-of-america> [accessed 13 June 2022].
5 As quoted on the accompanying label. The exhibition sign also pointed out that kinder critics have suggested Audubon
mistook a juvenile bald eagle for a new species.
6 In December 2019 a complete four-volume edition, first purchased via subscription by the Yorkshire Philosophical
Society, was sold by Sotheby’s New York to a private buyer for $6.6 million. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland The Golden Eagle is presented clawing out
the eye of a white rabbit, a jet of viscera
shooting from the wound (Fig. 2), while
his plate framing the Red-Tailed Hawk
shows two of the species battling over
an ensnared and bloodied young hare
defecating in fear. Viewed at their intended
scale, the images are vivid and thrilling
(apparently the colours were unusually
bright due to the fact that the prints on
show had been largely kept in storage
since publication). Even the image of the
Snowy Owl, one of only a small number
given a nocturnal backdrop, clearly looks
to evoke moodiness; dark clouds break
in the sky as two of the birds cast a side-
eye at the viewer from their perch atop
a dead tree. It is a quality that speaks to
the success of Audubon’s ambitions to
capture not just the birds but the ‘spirit of
the moment’, as he described it in a letter
to Sir William Jardine included in the exhibition, and which in person lands som
between a proto-ecological sensibility and an eye for the drama of the natural
Fig. 2: John James Audubon, print o
Eagle, Birds of America. © National M
Scotland. Yet, to insist too much on his work’s document
made clear, to overlook the romanticism that so
captures many of his avian subjects at
their most intense, and often most violent. The Golden Eagle is presented clawing out
the eye of a white rabbit, a jet of viscera
shooting from the wound (Fig. 2), while
his plate framing the Red-Tailed Hawk
shows two of the species battling over
an ensnared and bloodied young hare
defecating in fear. Viewed at their intended
scale, the images are vivid and thrilling
(apparently the colours were unusually
bright due to the fact that the prints on
show had been largely kept in storage
since publication). Even the image of the
Snowy Owl, one of only a small number
given a nocturnal backdrop, clearly looks
to evoke moodiness; dark clouds break
in the sky as two of the birds cast a side-
eye at the viewer from their perch atop
a dead tree. 7 Charles Darwin attended a lecture delivered by Audubon on turkey buzzards at the Natural History Association in
Edinburgh, and Audubon is cited in both Origin of Species (1859) and The Descent of Man (1871). See Richard Rhodes,
John James Audubon: The Making of an American (London: Vintage, 2004), pp. 305–06. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland The fact that the museum holds only
an incomplete edition of the book, having let its subscription lapse during the original
publication run, speaks to one of the many practical and economic problems that
Audubon appears to have faced in keeping the project going.4 Spread
over
three
rooms,
the
exhibition offered an opportunity to take
in the breadth of the book’s ambitions and
scope, from a print of the Great American
Cock (more commonly known as the wild
turkey), which was the first painting to
be engraved, to his Bird of Washington,
an eagle species yet to be found and
which has led some critics to charge him
with invention (Fig. 1). In many respects,
the Bird of Washington highlights the
spirit of Audubon’s drive not just to
document but to innovatively capture
the essential character of his subjects. With its imposing talons and powerful
wings, it was to Audubon ‘indisputably
the noblest bird of its genus that has yet
been discovered in the United States’.5
The fact that it probably does not exist
has not detracted from its appeal over the
centuries.6 Fig. 1: John James Audubon, print of Bird
of Washington, Birds of America. Wikimedia
Commons. Fig. 1: John James Audubon, print of Bird
of Washington, Birds of America. Wikimedia
Commons. The critics and biographers of Audubon who appeared as talking heads on the videos
that accompanied the images seemed to agree that, on the whole, his work serves as an
admirable compromise between aesthetics and documentation. And there seems little
disagreement that Audubon’s contributions to the natural sciences were significant:
he identified over twenty new species of birds through his efforts to find, observe, 4 and paint the continent’s avian life. And while other birds he identified as being newly
discovered to science are now accepted as subspecies, the exhibition insisted on his
place within the complicated legacies of nineteenth-century science, as well as art.7 and paint the continent’s avian life. And while other birds he identified as being newl
discovered to science are now accepted as subspecies, the exhibition insisted on h Yet, to insist too much on his work’s documentarian dimensions is, as the ex
made clear, to overlook the romanticism that so clearly guided his vision. A
captures many of his avian subjects at
their most intense, and often most violent. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland It is a quality that speaks to
the success of Audubon’s ambitions to
capture not just the birds but the ‘spirit of
the moment’, as he described it in a letter
to Sir William Jardine included in the exhibition, a
between a proto-ecological sensibility and an eye
Fig. Eag Fig. 2: John James Audubon, print of Golden
Eagle, Birds of America. © National Museums
Scotland. Fig. 2: John James Audubon, print of Golden
Eagle, Birds of America. © National Museums
Scotland. Reading Antoine Traisnel’s chapter on Audubon in Capture: American Pursuits and
the Making of a New Animal Condition (2020) at home afterwards, I realized that I had
missed a tiny self-portrait in the print of the Golden Eagle, which apparently presents 5 Fig. 3: John James Audubon, Golden Eagle
with miniature self-portrait. Wikimedia
Commons. the artist astride a fallen tree trunk lodged
between a mountain precipice, a gun and
a dead eagle strung over his back (Fig. 3). Continuing to read, however, I discovered
that in fact I had not missed it, since it
was not there. When Audubon returned to
his painting and readied it for inclusion
in Birds of America, he kept the tree
trunk but removed the miniature hunter
(Traisnel, p. 55). This self-conscious
erasure of both artist and hunter from the
canvas (not mentioned in the exhibition
signage) speaks to the way in which Birds
of America both does and does not want to
acknowledge its origins in acts of violence
against its subjects. Perhaps too easily absolved by the
argument that all ornithology of the
period was couched in a culture of
hunting, Audubon used fresh corpses
for his models. His process was to spend
an extensive amount of time finding
and observing his subjects, taking in
movement and behaviour, before then shooting the targeted bird (he was, apparently,
an exceptional shot). Not shying away from this part of Audubon’s process, the
exhibition stressed the importance of his ‘position board’ for the modelling of the dead
birds. It was on this instrument that he was able to pose the corpse according to what
he had observed in the field (as the exhibition pointed out, it had been convention to
rely on taxidermy). Fig. 3: John James Audubon, Golden Eagle
with miniature self-portrait. Wikimedia
Commons. Fig. 3: John James Audubon, Golden Eagle
with miniature self-portrait. Wikimedia
Commons. 8 ‘Audubon’s Birds of America’, National Museums Scotland Press Release. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland The liveliness of the prints then, in some respects, derives from this process of
‘pinning birds into realistic poses he had observed in life and painting on the spot’.8
Yet, the position board also seemed to have invited artifice and manipulation, especially
when the larger birds would not fit onto even the largest paper available. The Golden
Eagle, for instance, is painted with its wings positioned in such a way that its pose is 6 aerodynamically impossible (possibly the most egregious example and certainly the
most commented upon). While the talking heads recorded for the exhibition are keen to
defend Audubon from accusations of anthropomorphism, it is, in fact, this aspect of his
work that makes him so interesting. The tension between a documentary naturalism and
a seductive romanticism is strangely productive. Even produced at life size, the sense of
natural scale is allusive — the birds seem often larger than life, perhaps a correlative to
the fact that it is impossible to lose sight of the fact that what we are being confronted
with is a huge book, imposing in both size and weight. The book cannot quite not point
to its own artifice, both in terms of what it is representing and as an artefact in itself. All of this constantly draws attention to the way in which, on encountering Audubon’s
work, we are always firmly in the world of representation and viewership at several
removes. The works on display register a remarkable veracity while also offering a way
into thinking about human-avian relationships, in which (despite Audubon’s desire
otherwise) we cannot ever quite remove ourselves from the frame. The result is not a lack of realism: the taxidermy displayed in the exhibition looked
surprisingly less real than the two-dimensional images. Partially, this is down to what
Audubon is able to include on the page. His attention to habitat situates the birds within
a specific context in contrast to the
violent decontextualization of the glass-
encased taxidermy. His insistence on the
environments
that
sustain
America’s
birds finds him having intuited the idea
of ecological interdependence. The Blue-
Winged Yellow Warbler plate positions
two of the species creeping around the
encompassing bloom of a large-flowered
hibiscus, presenting an image of fragile
dependence, while the Purple Grackle
plate draws the eye to the flesh-coloured
corn husk being torn open for sustenance
as much as to the less colourful birds
(Fig. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland 4). Elsewhere, ecological harmony is
suggested through aesthetic sympathy. The
yellows and blues of the Carolina Pigeon
are mirrored in the matching colours of
lichen covering the branch on which they
Fig. 4: John James Audubon, print of Purple
Grackle, Birds of America. © National
Museums Scotland. The result is not a lack of realism: the t
surprisingly less real than the two-dimens
Audubon is able to include on the page. His
a specific context in contrast to the
violent decontextualization of the glass-
encased taxidermy. His insistence on the
environments
that
sustain
America’s
birds finds him having intuited the idea
of ecological interdependence. The Blue-
Winged Yellow Warbler plate positions
two of the species creeping around the
encompassing bloom of a large-flowered
hibiscus, presenting an image of fragile
dependence, while the Purple Grackle
plate draws the eye to the flesh-coloured
corn husk being torn open for sustenance
as much as to the less colourful birds
(Fig. 4). Elsewhere, ecological harmony is
suggested through aesthetic sympathy. The
yellows and blues of the Carolina Pigeon
are mirrored in the matching colours of
lichen covering the branch on which they Fig. 4: John James Audubon, print of Purple
Grackle, Birds of America. © National
Museums Scotland. Fig. 4: John James Audubon, print of Purple
Grackle, Birds of America. © National
Museums Scotland. Fig. 5: John James Audubon, print of Carolina
Pigeons or Turtle Doves, Birds of America. © National Museums Scotland. are perched (Fig. 5). Yet, at this juncture,
the exhibition again invited reflection on
the tension between artistic arrangement
and verisimilitude. It is precisely because
of the plate’s formal harmony that it is able
to draw the viewer to ecological realities. Its willingness not to shy away from
Audubon’s many contradictions was one
of the strongest aspects of the exhibition. Aged in his forties by the time he began the
project, he was, as the signage and videos
pointed out, an enthusiastic beneficiary
of slavery and an unapologetic white
supremacist. At the same time, he was a
man less of principle than of pragmatism,
willing to name one of the birds he
‘discovered’, the Roscoe’s Yellow Throat,
after a prominent abolitionist who also
happened to be one of the project’s
backers, William Roscoe.9 Fig. 5: John James Audubon, print of Carolina
Pigeons or Turtle Doves, Birds of America. © National Museums Scotland. 9 This was one of Audubon’s misidentifications, now believed to be an example of the Common Yellowthroat. 10 See Matthew Whittle, ‘Lost Trophies: Hunting Animals and the Imperial Souvenir in Walton Ford’s Pancha Tantra’, Journal
of Commonwealth Literature, 51 (2016), 196–210. Review of ‘Audubon’s Birds of America’ at the
National Museum of Scotland One of the missed opportunities, then, is to have not capitalized on how Audubon’s
compromised positionality might offer the exhibition’s visitors a chance to reflect on
the contradictions that accompany ecological self-consciousness in the twenty-first
century. The last room in the exhibition was dedicated to prints of Audubon’s birds that
are now in crisis (including the Blue-Headed Vireo, the Eastern Towhee, the Dark-Eye
Junco, and the Snowy Owl), as well as those already extinct (the Carolina Parakeet and
Passenger Pigeon). Although habitat destruction and industrial agriculture are singled
out as impacting on ecosystems, the more ethically complicated question of the role
of institutions, organizations, and government policy in hastening such violence was
unaddressed. Also ignored were some of the trickier questions around the way in which
Audubon was contributing to a colonial settler project that was erasing as much as it
was documenting a world in the process of being ‘discovered’. Including contemporary
responses to Audubon, such as the self-reflexive work of Walton Ford (1960–), or
having included indigenous ways of knowing and thinking about the natural world that 8 8 were in the process of being actively displaced in the nineteenth century, would have
offered one way of situating Birds of America both within a postcolonial frame, as well
as having pointed to the challenges of finding an aesthetic expression adequate to our
own moment of ecological crisis.10 This would have required an exhibition with a far more radical eco-politics than
the National Museum of Scotland was willing, or able, to have delivered. Instead, the
exhibition focused on the legacy of a troubling and troubled man, the ongoing ability of
Birds of America to impress and beguile, and the degree to which, more often than not,
art, science, and ecology come as a package rather than as discrete modes of knowing.
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Microbial invasion of a toxic medium is facilitated by a resident community but inhibited as the community co-evolves
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INTRODUCTION occurs when new communities assemble and groups of species
follow one another in so-called “successions”, for example in the
formation of dental plaque [36, 37] or marine particle commu-
nities [32, 33]. Facilitation likely occurs in newly assembling
communities, as sterile environments are typically difficult to
colonize, for example if they have an extreme pH, contain toxic
compounds, or are lacking in easily accessible nutrients or water. Pioneer species may alter the environment in ways that facilitate
invasion by new species that would otherwise not survive
[24, 38–41]. This is in line with the Stress Gradient Hypothesis
(SGH), which predicts that species are more likely to interact
positively in stressful environments [42–48]. The link between the
SGH and microbial invasion has, however, not yet been explored
experimentally. Successful colonization of invader microorganisms into sterile
environments or existing microbial communities are common and
can impact ecosystem diversity and function, potentially with
significant consequences [1–3]. A better understanding of the
factors driving microbial invasions may help to prevent the spread
and establishment of invasive species, or to aid the intentional
introduction of a new species for a desired purpose. For example,
it might be desirable to prevent the invasion of a species that
reduces the efficiency of a bioremediation system [4], or to
promote the colonization of probiotic species in the intestinal
microbiome of a patient [5, 6]. What determines the ability of an invasive species to colonize
an existing ecosystem depends on the characteristics of both the
invading species and the resident community [7, 8]. Many
theoretical and empirical studies have established factors that
influence invasion outcome, such as propagule pressure [9–14],
resident community productivity [15], genotypic richness of
invaders [13, 16] or the resident community [3, 13, 17–20],
community niche coverage [3, 21, 22], and abiotic conditions (e.g.,
the presence of antibiotics [23]). As more species colonize the environment and species diversity
increases, previously available niches begin to fill up, such that
competition is expected to increase and invasion success to drop. The negative relationship between invasion success and species
richness and diversity have been well-established [3, 13, 17–20]. As time passes, resident species may co-evolve to reduce niche
overlap and availability in a way that would prevent further
invasion. www.nature.com/ismej Received: 3 March 2022 Revised: 27 July 2022 Accepted: 24 August 2022
Published online: 14 September 2022
1Département de Microbiologie Fondamentale, Université de Lausanne, Lausanne, Switzerland. 2Department of Environmental Systems Science, ETH Zurich, Zürich, Switzerland.
3Swiss Institute of Bioinformatics, Lausanne, Switzerland. ✉email: sara.mitri@unil.ch Philippe Piccardi1, Géraldine Alberti1, Jake M. Alexander2 and Sara Mitri
1,3✉ © The Author(s) 2022 © The Author(s) 2022 Predicting whether microbial invaders will colonize an environment is critical for managing natural and engineered ecosystems,
and controlling infectious disease. Invaders often face competition by resident microbes. But how invasions play out in
communities dominated by facilitative interactions is less clear. We previously showed that growth medium toxicity can promote
facilitation between four bacterial species, as species that cannot grow alone rely on others to survive. Following the same logic,
here we allowed other bacterial species to invade the four-species community and found that invaders could more easily colonize a
toxic medium when the community was present. In a more benign environment instead, invasive species that could survive alone
colonized more successfully when the residents were absent. Next, we asked whether early colonists could exclude future ones
through a priority effect, by inoculating the invaders into the resident community only after its members had co-evolved for
44 weeks. Compared to the ancestral community, the co-evolved resident community was more competitive toward invaders and
less affected by them. Our experiments show how communities may assemble by facilitating one another in harsh, sterile
environments, but that arriving after community members have co-evolved can limit invasion success. The ISME Journal (2022) 16:2644–2652; https://doi.org/10.1038/s41396-022-01314-8 The ISME Journal (2022) 16:2644–2652; https://doi.org/10.1038/s41396-022-01314-8 Received: 3 March 2022 Revised: 27 July 2022 Accepted: 24 August 2022
Published online: 14 September 2022 ARTICLE
OPEN
Microbial invasion of a toxic medium is facilitated by a resident
community but inhibited as the community co-evolves Philippe Piccardi1, Géraldine Alberti1, Jake M. Alexander2 and Sara Mitri
1,3✉ Philippe Piccardi1, Géraldine Alberti1, Jake M. Alexander2 and Sara Mitri
1,3✉ Pseudmonas fulva
CA (Centrimide agar, Sigma-Aldrich 22470, 4.67 g/
100 ml) + carbenicillin (50 μg/ml)
37
24
White colonies
hed each of the resident and invader species from one another according to their preferences in growth media and temperature, the time at which colonies became visible by eye and the colony does not manage to establish [56]. One prediction is that co-
evolved resident species would be less perturbed by species
invasion, presumably due to increased niche coverage [49]. Experimentally disentangling the role of the different factors
discussed above on invasion success and robustness against
invasion can be challenging. Table 1. Selective media composition. Species type
Species
Selective plate
Incubation
temperature (°C)
Time of colony
appearance (hours)
Colony morphology
Resident
Agrobacterium
tumefaciens
LBA (Luria Bertani agar, 4 g/100 ml) + sulfamethoxazole
(14.25 μg/ml) + trimethoprim (0.75 μg/ml)
28
48–72
Green fluorescent colonies
(GFP-labeled strain)
Comamonas testosteroni
LBA (4 g/100 ml)
28
24
White colonies, the first
to appear
Microbacterium
saperdae
LBA (4 g/100 ml) + colistin (10 μg/ml)
28
48–72
Green/yellow colonies
Ochrobactrum anthropi
LBA (4 g/100 ml) + colistin (10 μg/ml)
28
72–96
Red fluorescent colonies
(mCherry-labeled strain)
Invader
Aeromonas caviae
MMC (MM ChromoSelect agar, Sigma-Aldrich 00563,
4.91 g/100 ml) + nalidixic acid (15 μg/ml)
37
24
Dark blue colonies
Klebsiella pneumoniae
KIA (Klebsielle Isolation agar, Sigma-Aldrich 90925, 4.08 g/
100 ml) + carbenicillin (50 μg/ml)
37
24
Purple colonies
Providencia rettgeri
PIA (Pseudomonas Isolation agar, 4.5 g/
100 ml) + tetracycline (30 μg/ml) + 2% glycerol
28
48–72
White colonies
Pseudmonas fulva
CA (Centrimide agar, Sigma-Aldrich 22470, 4.67 g/
100 ml) + carbenicillin (50 μg/ml)
37
24
White colonies
We distinguished each of the resident and invader species from one another according to their preferences in growth media and temperature, the time at which colonies became visible by eye and the colony
morphology g g
Here we aim to test the effect of the two less-well understood
factors (the SGH and priority effects) on bacterial invasion success
and resistance by studying invasion into a synthetic bacterial
community whose composition is fixed at four species: Agrobacter-
ium tumefaciens, Comamonas testosteroni, Microbacterium saperdae,
and Ochrobactrum anthropi. These four species can grow and
bioremediate metal working fluids (MWF) [47, 57], an industrial
fluid used in metal manufacturing. MWFs contain mineral oils,
emulsifiers, and biocides, some of which are toxic to bacteria. In
previous work [47], we showed that when the four species were
grown together in this toxic environment, they facilitated each
other’s survival compared to when they were alone. Instead, when
we added amino acids to make the environment more permissive,
competition between species increased. Study system y y
The four bacterial species used to assemble the resident community were
isolated from MWF [57] and are referred to as: Agrobacterium tumefaciens
str. MWF001,
Comamonas
testosteroni
str. MWF001,
Microbacterium
saperdae str. MWF001, and Ochrobactrum anthropi str. MWF001 (as in ref. [47]). The additional four bacterial species, used to invade the resident
community, were kindly donated by Peter Küenzi from Blaser Swisslube
AG, Hasle-Rüegsau and we identified them using MALDI-TOF MS
performed at Mabritec AG, Switzerland as: Aeromonas caviae, Klebsiella
pneumoniae, Providencia rettgeri, and Pseudomonas fulva. We name these
four strains str. Blaser001. As mentioned in the main text, these species
were chosen from a set of 20 isolates, based on our ability to design
selective plates on which the invader but not the resident species would
grow (see Table 1). The choice of invader species might favor species that
differ metabolically from the residents, which could potentially increase
invasion ability, but we expect this effect to be small, given how different
the selective media were from MWF. The MWF (Castrol Hysol XF, acquired
in 2016) was prepared at a concentration of 0.5% (v/v), diluted in water
with the addition of selected salts and metal traces to support bacterial
growth. We also used MWF medium supplemented with 1% casamino
acids (Difco, UK) (MWF + AA). These media were prepared as in ref. [47]. Pseudmonas fulva
CA (Centrimide agar, Sigma-Aldrich 22470, 4.67 g/
100 ml) + carbenicillin (50 μg/ml)
37
24
White colonies
hed each of the resident and invader species from one another according to their preferences in growth media and temperature, the time at which colonies became visible by eye and the colony This system allows us to
study
biological
invasion
while
experimentally
manipulating
environmental conditions to control interactions between com-
munity members and holding all other factors constant. Another
advantage of this system is that the four species can coexist over
evolutionary time-scales, allowing us to explore the effect of
community co-evolution on microbial invasion. y
Using
four
invader
species,
Aeromonas
caviae,
Klebsiella
pneumoniae, Providencia rettgeri, and Pseudomonas fulva that
were isolated from waste MWF (chosen from a set of 20 based on
our ability to distinguish them from the resident species), we first
show that the resident community facilitates invasion of species
that cannot grow alone, but inhibits those that can. Whether or
not species could grow alone was modulated by changes in the
growth medium. Second, after co-evolving the four resident
species for 44 weeks, we found that invasions were still possible in
MWF, but the growth of the invaders was inhibited relative to the
ancestral community and the co-evolved resident species were
less affected by invasions. Together, our results show that
facilitative communities are easier to invade than competitive
ones, but that a co-evolved community is more robust to invasion
compared to an ancestral one. INTRODUCTION The Community Monopolization Hypothesis predicts
that early colonizers adapt to use available resources efficiently,
yielding a competitive advantage against later-arriving species
[31, 49–53], also known as a “priority effect” [31, 53]. Invasion success may also depend on the sign and strength of
interactions between resident community members, and between
residents and invaders [24]. Previous studies tend to find that
invaders compete with resident species [3, 13, 17, 20, 23, 25, 26],
which is consistent with competition being prevalent in microbial
communities [27–29]. However, in sterile environments, early
colonizers can facilitate the arrival of other species [30–35]. This Besides invasion success, it is important to consider whether
invaders perturb the resident community, possibly changing
community structure and function [24, 54, 55], even if the invader P. Piccardi et al. 2645 g
g
g
100 ml) + carbenicillin (50 μg/ml)
esident and invader species from one another according to their preferences in growth media and temperature, the time at which colonies became visible by eye and the colony Invasion assays g
g
P. fulva was the only one of the four invader species that could
colonize the MWF when alone (Fig. 1A (red dotted line), C). Instead, when the community was present, the number of
successful invasions increased: A. caviae, K. pneumoniae, and P. fulva colonized the MWF containing the resident community,
while P. rettgeri still did not (Fig. 1A (red solid line), C). These
results are in line with our previous findings [47] that species that
cannot grow alone in MWF are likely to be facilitated by other
species, explaining why in some cases invasion is only successful
when the community is present. Invasion was performed after 2 days of the first transfer of the resident
community. One single colony of each invader species was selected and
inoculated in 10 ml of TSB in Erlenmeyer flasks (50 ml) and incubated
overnight at 28 °C, shaken at 200 rpm. To achieve exponentially growing
bacteria, with a final concentration of ∼106–107 CFU/ml, each invader
strain was inoculated at an OD600 of 0.05 measured by spectrophotometry
(Ultrospec 10, Amersham Biosciences), in 20 ml of TSB in Erlenmeyer flasks
(100 ml) and cultivated at 28 °C (200 rpm). After 3 h, 200 μl of the invader
species were centrifuged (5 min, 10,000 rcf). The bacterial pellet was
resuspended in the same medium of the resident community. In total,
200 μl of this suspension were then added to the culture tubes, with or
without the resident communities. For the experiments where propagule
pressure was changed (Figs. S2 and S3), we took either 2 ml or 200 μl for
propagule size 107 or 106, respectively, centrifuged, and resuspended
them. For propagule size 105 or 104, we instead aliquoted 20 μl of cell
suspension into 180 μl of PBS and diluted it once more in PBS for 104,
before centrifuging and re-suspending in the growth medium of the
resident community. Quantifying bacterial abundance The abundance of each resident or invader species was quantified before
the inoculation in the MWF or MWF + AA (before combining resident
species) and before each transfer using serial dilution and selective plating
(Table 1). We also quantified population sizes of the resident species at the
time of invasion (Fig. S4C) and the same approach was used to quantify
growth curves shown in Figs. S5–S8. To define invasion outcomes we used
an invasion threshold representing the dynamics of an invader species
with a growth rate of 0 (its abundance changes only due to dilution, i.e.,
100-fold decrease every transfer from the initial population size). By
subtracting this threshold value from the abundance of the invader species
at transfer four, the invasion is defined as successful if >0 (the growth rate
is positive) or failed if ≤0 (the growth rate is 0 or negative). We used a
Kruskal–Wallis test to assess whether effects were significant. Raw CFU/ml
data and the results of all statistical tests are listed in Dataset 1. Experimental setup To assemble the resident community, a single isolated colony of each
species was selected and inoculated in 10 ml of Tryptic Soy Broth (TSB) in
Erlenmeyer flasks (50 ml), then incubated overnight at 28 °C (200 rpm). To The ISME Journal (2022) 16:2644 – 2652 P. Piccardi et al. 2646 2646 Quantifying pairwise interactions achieve exponentially growing bacteria, with a final concentration of
∼106–107 CFU/ml, each bacterial species was inoculated at an OD600 of
0.05 measured by spectrophotometry (Ultrospec 10, Amersham Bios-
ciences), in 20 ml of TSB in Erlenmeyer flasks (100 ml) and cultivated at
28 °C, shaken at 200 rpm. After 3 h, 200 μl of each of the four resident
species were combined and centrifuged (5 min, 10,000 rcf). The bacterial
pellet was resuspended in 30 ml of MWF or MWF + AA into borosilicate
glass tubes (16 × 125 mm, 30 ml). achieve exponentially growing bacteria, with a final concentration of
∼106–107 CFU/ml, each bacterial species was inoculated at an OD600 of
0.05 measured by spectrophotometry (Ultrospec 10, Amersham Bios-
ciences), in 20 ml of TSB in Erlenmeyer flasks (100 ml) and cultivated at
28 °C, shaken at 200 rpm. After 3 h, 200 μl of each of the four resident
species were combined and centrifuged (5 min, 10,000 rcf). The bacterial
pellet was resuspended in 30 ml of MWF or MWF + AA into borosilicate
glass tubes (16 × 125 mm, 30 ml). Quantifying pairwise interactions
Pairwise interactions between species (Figs. S6G, S7G, and S8F) were
quantified as in ref. [47]. Briefly, arrow thickness indicates the interaction
strength measured as the ten-fold change in area under the growth curve
(AUC, plotted in Figs. S6F, S7F, and S8E), with the color showing the sign of
fold-change and the p values resulting from a Kruskal–Wallis test
comparing each species alone and with a given partner species. We use
the AUC to represent species growth, as we have found it to adequately
summarize species’ effects on one another, combining growth rate, yield,
and lag phase length [47]. Residents inhibit invaders that can grow alone in a more
permissive medium p
MWFs are designed to prevent bacterial contamination and
include biocides [58], which make them quite toxic. This explains
why only one of the invader species was able to grow alone in the
MWF medium. To explore invasion in a less harsh environment, we
enriched the medium by adding 1% casamino acids (MWF + AA). Casamino acids are a nutrient source for three out of the four
resident community members and, according to previous work
[47, 59], we expect more negative interactions in a more
permissive medium. We found that K. pneumoniae, P. fulva, and
P. rettgeri could colonize MWF + AA alone, while A. caviae still
suffered from the environmental toxicity (invaders in Fig. 1B, C and
residents in Fig. S10A, right). The three species that were able to
colonize alone were still able to invade the community, but
significantly less well compared to when the community was
absent (Kruskal–Wallis, all p values < 0.05, Fig. 1C). Consistent with
previous work [47], our results suggest that in this more permissive
environment, the community competes with the invaders. The resident community facilitates the invasion of species
that cannot grow alone g
We first ask whether each of the four invader species (A. caviae, K. pneumoniae, P. rettgeri, and P. fulva), could colonize MWF and to
what extent the resident community promotes or inhibits
invasion. The resident community was cultured in MWF for
1 week, with 1% of the population transferred into fresh media
once a week for a total of 4 weeks (see Methods). Each invader
species was inoculated individually into three replicate micro-
cosms of the resident community 48 h after the first transfer,
presumably during the community’s exponential growth phase
(Fig. S9A). As a control treatment, we inoculated each of the
invader species into sterile MWF and performed transfers in
parallel (Fig. S9B). We quantified the abundance of all species at
inoculation and before each transfer (see Methods; invaders in
Fig. 1A and residents in Fig. S10A, left). Transfers All communities (the four-species or the three-species resident commu-
nities) were incubated at 28 °C and shaken at 200 rpm for seven days in
either MWF or MWF + AA medium. Every week, 300 μl (1%) of the week-
old culture was transferred into fresh medium and the growth cycle
repeated. Each week, we also harvested 1 ml of each culture, spun it down
at 10,000 rcf for 5 min, resuspended it in glycerol 25% (diluted in PBS) and
stocked it at −80 °C for future analyses. This was repeated for 44 transfers
(weeks) to co-evolve the resident communities or for four transfers in the
invasion assays. The evolutionary experiment was conducted in five
replicate
culture
tubes
for
each
condition
(three-
or
four-species
community), of which we show only one here (Fig. S1). After the 44 weeks,
we isolated one colony of each species, which we refer to as A. tumefaciens
str. MWF431, C. testosteroni str. MWF431, M. saperdae str. MWF431, and O. anthropi str. MWF431 for the four-species co-evolved community; and A. tumefaciens str. MWF351, C. testosteroni str. MWF351, M. saperdae str. MWF351, and O. anthropi str. MWF351 for the three-species co-evolved
community. This design choice simplified our experiments, but means that
we cannot test how intraspecies diversity affects invasion success. Quantifying growth rates The capacity to colonize a resident community might depend on
community history: resident species that have adapted to one
another in a given environment may be more likely to exclude
future colonists through a priority effect [49–51]. To test this
hypothesis, we extend the pre-invasion phase to 44 weeks,
allowing the four resident species to adapt to MWF and to each We quantified bacterial growth rates in Fig. S5 in two ways: first, we took
the difference in CFU/ml between all the measurements on consecutive
days that we had and took the maximum value (panels B and E). We also
computed the fold change between all consecutive CFU/mL measure-
ments and divided that by the number of days between measurements
(panels C and F). The ISME Journal (2022) 16:2644 – 2652 P. Piccardi et al. 2647 A
CFU/ml
10
10
10
6
10
2
1
2
3
4 1
3
4 1
2
3
4 1
2
3
4
Ac
Kp
r
P
f
P
Transfer
B
C
Transfer
10
10
10
6
10
2
1
2
3
4 1
2
3
4 1
2
3
4 1
2
3
4
CFU/ml
Ac
Kp
r
P
f
P
alone (MWF+AA)
into community (MWF+AA)
invasion threshold
alone (MWF)
into community (MWF)
invasion threshold
(alone)
invasion threshold
(into community)
0
102
104
106
108
1010
-102
-104
NS
*
*
*
*
*
*
Ac
Kp
Pr
Pf
A
C
A
A
A
C
C
C
invasion success
MWF
MWF+AA
invasion failure
A = alone
C = into community
Invasion magnitude at transfer 4
2
Fig. 1
Outcome of invasion in MWF or MWF + AA. A Red lines: abundances of the four invader species in MWF at different transfers,
quantified at first inoculation and before each transfer. The four invader species were grown alone or inoculated into the growing resident
community after it had been transferred once and until transfer four (one transfer every 7 days). At each transfer, the culture is diluted 100-
fold. The experiments were done in parallel except for the invader alone, which was done separately. Black lines: theoretical expectation of
what would happen if the invader did not grow at all and was just diluted, which we call the “invasion threshold”, whether invading alone
(dotted) and into the community (solid). We count an invasion as successful if the abundance of the invader at the end of the transfers is
higher than the invasion threshold. B Blue lines: abundances of the four invader species in MWF + AA at different transfers when alone
(dashed) and into the community (solid). Black lines: the theoretical invasion threshold simulating a non-growing invader. There is only one
invasion threshold as experiments were all conducted in parallel and had the same starting population sizes (compared to A). C To exclude
the effect of the choice of dilution rate on invasion success, we calculate the “invasion magnitude”: the abundance of each invader at transfer
four minus its corresponding invasion threshold. If this number is higher or lower than zero, then the invasion is successful (+) or failed (−),
respectively. From left to right: A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. If this number is higher or lower than zero, then the invasion is successful (+) or failed (−),
respectively. From left to right: A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. fulva (Pf). Statistical significance is marked above the data
points (p values: *<0.05, NS not significant). For abundance of resident species see Fig. S2. evolutionary process and allow it to invade at a later stage. We did
this by co-evolving three of the resident species, A. tumefaciens, C. testosteroni, and M. saperdae together in MWF for 44 weeks,
excluding O. anthropi. Next, we combined single isolates of the
three co-evolved species and invaded the wild-type O. anthropi
into this co-evolved three-species community (Fig. S9D). As
before, O. anthropi could not colonize the MWF when alone (as
in ref. [47], Fig. 2C), but invaded successfully when inoculated into
the ancestral or the co-evolved community of three. Consistent
with the previous invasion assays (Fig. 2A, B) and our hypothesis
that a co-evolved community is more difficult to invade, O. anthropi grew significantly worse when it was inoculated into the
co-evolved three-species community compared to the corre-
sponding ancestral one (invader O. anthropi in Fig. 2C, D and
residents in Fig. S12). other (see Methods). Next, we mixed one co-evolved isolate of
each species and call this the “co-evolved community” (Fig. S9C). We now ask to what extent the invader species can colonize
the co-evolved resident community compared to the ancestral
one. We found that while P. rettgeri could colonize neither, A. caviae, K. pneumoniae, and P. fulva colonized both the ancestral
and co- evolved communities (invaders in Fig. 2A, B and
residents in Fig. S10B). However, all three invader species had a
smaller invasion magnitude in the co-evolved compared to the
ancestral community (Kurskal–Wallis, A. caviae p value < 0.0005,
K. pneumoniae p value < 0.0005, P. fulva p value < 0.05, Fig. 2B). The invasion outcome for A. caviae was initially inconclusive,
where in one out of two biological replicates the invader went
extinct when inoculated into the ancestral community (Fig. S2A,
B). We tested whether this was due to variability in propagule
pressure [13], but found no evidence for this, as different
invasion population sizes of A. caviae all converged to a similar
population size at transfer four (invaders in Fig. S3 and residents
in Fig. S11). We therefore concluded that the death of A. fulva (Pf). Statistical significance is marked above the data
points (p values: *<0.05, NS not significant). For abundance of resident species see Fig. S2. 2 A
CFU/ml
10
10
10
6
10
2
1
2
3
4 1
3
4 1
2
3
4 1
2
3
4
Ac
Kp
r
P
f
P
Transfer
B
C
Transfer
10
10
10
6
10
2
1
2
3
4 1
2
3
4 1
2
3
4 1
2
3
4
CFU/ml
Ac
Kp
r
P
f
P
alone (MWF+AA)
into community (MWF+AA)
invasion threshold
alone (MWF)
into community (MWF)
invasion threshold
(alone)
invasion threshold
(into community)
0
102
104
106
108
1010
-102
-104
NS
*
*
*
*
*
*
Ac
Kp
Pr
Pf
A
C
A
A
A
C
C
C
invasion success
MWF
MWF+AA
invasion failure
A = alone
C = into community
Invasion magnitude at transfer 4
2 A Fig. 1
Outcome of invasion in MWF or MWF + AA. A Red lines: abundances of the four invader species in MWF at different transfers,
quantified at first inoculation and before each transfer. The four invader species were grown alone or inoculated into the growing resident
community after it had been transferred once and until transfer four (one transfer every 7 days). At each transfer, the culture is diluted 100-
fold. The experiments were done in parallel except for the invader alone, which was done separately. Black lines: theoretical expectation of
what would happen if the invader did not grow at all and was just diluted, which we call the “invasion threshold”, whether invading alone
(dotted) and into the community (solid). We count an invasion as successful if the abundance of the invader at the end of the transfers is
higher than the invasion threshold. B Blue lines: abundances of the four invader species in MWF + AA at different transfers when alone
(dashed) and into the community (solid). Black lines: the theoretical invasion threshold simulating a non-growing invader. There is only one
invasion threshold as experiments were all conducted in parallel and had the same starting population sizes (compared to A). C To exclude
the effect of the choice of dilution rate on invasion success, we calculate the “invasion magnitude”: the abundance of each invader at transfer
four minus its corresponding invasion threshold. caviae
in one biological replicate might have been due to a technical
error (Fig. S2). In summary, while invasions into a community co-evolved in
MWF are still possible, co-evolved community members inhibit
invading species more than their ancestors. The ISME Journal (2022) 16:2644 – 2652 Co-evolved communities are less affected by invasion
compared to their ancestors Positive or negative invasion magnitudes indicate successful (+) or failed (−) invasions, respectively. From left to
right: A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. fulva (Pf), O. anthropi (Oa). Statistical significances are marked above the data points
(p values: *<0.05, NS not significant, N/A not applicable). See Fig. 1 caption for more details. In A and B, the resident community consists of a
co-culture of four ancestral or evolved clonal populations, while in C and D, the community contained only three clonal populations (four
residents, without O. anthropi), either ancestral or evolved. For abundance of the four-species or three-species resident communities see
Figs. S2B or S6, respectively. 48 A
CFU/ml
10
10
10
6
10
2
1
2
3
4
1
2
3
4
1
2
3
4
Ac
Kp
Pr
Transfer
1
2
3
4
Pf
1
2
Oa
C
CFU/ml
10
10
10
6
10
2
T C 1
2
3
4
Pf B
0
102
104
106
108
1010
A
C
E
A
C
E
A
C
E
A
C
E
-102
-104
Invasion magnitude at transfer 4
Ac
Kp
Pr
Pf
*
*
*
*
*
*
*
*
N/A
N/A
N/A
NS B Oa Fig. 2
Invasion into ancestral or co-evolved communities. A, C Red/gray lines: invader species were grown alone (red dashed), or inoculated
into the ancestral (red solid) or the co-evolved community (gray solid) in MWF. Cultures were diluted 100-fold in fresh MWF every 7 days for a
total of four transfers. The experiments were all conducted in parallel. Black lines: the theoretical invasion threshold simulating a non-growing
invader. B, D Invasion magnitude (abundance at transfer four minus the invasion threshold) alone “A”, into the ancestral community “C”, or into
the evolved community “E”. Positive or negative invasion magnitudes indicate successful (+) or failed (−) invasions, respectively. From left to
right: A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. fulva (Pf), O. anthropi (Oa). Statistical significances are marked above the data points
(p values: *<0.05, NS not significant, N/A not applicable). See Fig. 1 caption for more details. In A and B, the resident community consists of a
co-culture of four ancestral or evolved clonal populations, while in C and D, the community contained only three clonal populations (four
residents, without O. anthropi), either ancestral or evolved. For abundance of the four-species or three-species resident communities see
Figs. Co-evolved communities are less affected by invasion
compared to their ancestors S2B or S6, respectively. By increasing the permissiveness of a harsh medium (MWF)
through the addition of amino acids (MWF + AA), the number of
invader species able to grow alone increased from a single one to
three out of four species. In almost all cases where invaders died
alone, the resident community facilitated their survival and
growth. Unfortunately, we do not yet know why more nutrients
or the presence of the resident community increased species
survival. The resident species might be producing additional
resources or removing toxic compounds [60]. Regardless of the
mechanism, though, if invaders could survive alone in the more
permissive environment, they experienced a net negative effect if
the community was present. cumulative effects), the abundance of two of the ancestral
residents, A. tumefaciens and O. anthropi, was significantly lower
when invaded by P. fulva (t-test, both p value < 0.005, Figs. 3A
and S13A). Otherwise, we detected no significant changes in their
final abundance following invasion by other species. This lack of
perturbation was also observed for C. testosteroni. M. saperdae
instead had a greater final population size in the presence of most
invaders (t-test, A. caviae, P. rettgeri, and P. fulva, all p values <
0.005, Fig. 3A and Fig. S13A). This is not unexpected, as we know
that M. saperdae strongly depends on other species to grow in
MWF [47]. cumulative effects), the abundance of two of the ancestral
residents, A. tumefaciens and O. anthropi, was significantly lower
when invaded by P. fulva (t-test, both p value < 0.005, Figs. 3A
and S13A). Otherwise, we detected no significant changes in their
final abundance following invasion by other species. This lack of
perturbation was also observed for C. testosteroni. M. saperdae
instead had a greater final population size in the presence of most
invaders (t-test, A. caviae, P. rettgeri, and P. fulva, all p values <
0.005, Fig. 3A and Fig. S13A). This is not unexpected, as we know
that M. saperdae strongly depends on other species to grow in
MWF [47]. Once the community had co-evolved, the abundances of A. tumefaciens and M. saperdae were no longer significantly affected
by the invasion of P. fulva (Fig. 3B). The abundance of O. anthropi
was still lower following invasion by P. fulva, but significantly less
compared to the ancestor (ancestral vs. co-evolved, t-test, p
value = 0.0167, Fig. 3A, B, last column). In addition, M. Co-evolved communities are less affected by invasion
compared to their ancestors So far, we have focused on the effect of the resident community
on the invading species. Next, we consider how robust the
resident community is to these invasion events. We next wondered whether the pattern observed for A. caviae,
K. pneumoniae, and P. fulva was specific to these invader species
colonizing our community of four resident species, or whether
there was anything particular about the four resident species. One
way to explore this is to exclude one species from the co- In all our treatments, the resident species were maintained over
the four transfers (Figs. S10 and S12). At transfer four (representing The ISME Journal (2022) 16:2644 – 2652 P. Piccardi et al. 2648 alone
into ancestral
community
invasion
threshold
into co-evolved
community
1
2
3
4
Oa
C
CFU/ml
10
10
10
6
10
2
0
102
104
106
108
1010
-102
-104
Invasion magnitude at transfer 4
D
*
*
*
A
C
E
Oa
Transfer
success
failure
A=
C=
E=
into ancestral
community
into co-evolved
community
alone
Invasion: A
CFU/ml
10
10
10
6
10
2
1
2
3
4
1
2
3
4
1
2
3
4
Ac
Kp
Pr
Transfer
1
2
3
4
Pf
B
alone
into ancestral
community
invasion
threshold
into co-evolved
community
1
2
3
4
Oa
0
102
104
106
108
1010
A
C
E
A
C
E
A
C
E
A
C
E
-102
-104
Invasion magnitude at transfer 4
Ac
Kp
Pr
Pf
C
CFU/ml
10
10
10
6
10
2
0
102
104
106
108
1010
-102
-104
Invasion magnitude at transfer 4
D
*
*
*
A
C
E
Oa
Transfer
success
failure
A=
C=
E=
into ancestral
community
into co-evolved
community
alone
Invasion:
*
*
*
*
*
*
*
*
N/A
N/A
N/A
NS
Fig. 2
Invasion into ancestral or co-evolved communities. A, C Red/gray lines: invader species were grown alone (red dashed), or inoculated
into the ancestral (red solid) or the co-evolved community (gray solid) in MWF. Cultures were diluted 100-fold in fresh MWF every 7 days for a
total of four transfers. The experiments were all conducted in parallel. Black lines: the theoretical invasion threshold simulating a non-growing
invader. B, D Invasion magnitude (abundance at transfer four minus the invasion threshold) alone “A”, into the ancestral community “C”, or into
the evolved community “E”. Co-evolved communities are less affected by invasion
compared to their ancestors saperdae
was no longer significantly positively affected by any of the
invaders. This may be because the co-evolved M. saperdae grows
significantly better within the resident community (Fig. 3A, B). The
co-evolved three-species community behaved similarly: while the
abundance of ancestral A. tumefaciens was significantly lower
following the invasion of O. anthropi (t-test, p value < 0.05), its co-
evolved counterpart was not (Fig. 3C, D). Altogether, co-evolved
resident communities were more robust to invasion compared to
ancestral ones. y
p
The observation that species that couldn’t survive alone
benefitted from those that could is consistent with our previous
research [47] and more generally with the SGH, which can now be
linked to invasion ecology due to our ability to modulate
environmental harshness and inter-species interactions: in a harsh
environment colonized by few species, invasion success may be
high, as niches are still available and invaders can rely on the
presence of the residents to survive. This is expected if early-
arriving species improve the environment, facilitating the growth
of others that are less well adapted to it [38–41]. Although MWF is
particularly toxic, we expect many sterile environments to be
“harsh” for many species, as every environment poses challenges
to species that are not adapted to it. Nevertheless, facilitation may
not always dominate, as it is possible that first colonizers alter the
environment in a way that inhibits future invaders [31], or that late
colonizers out-compete earlier ones and replace them [61], but
this is not what we observe here. DISCUSSION Our new intuition might then help to explain why the assembly
of many natural microbial communities is often highly predictable
[32, 33, 36, 37, 62]. Assuming that only few species can act as
pioneers and improve the environment in similar ways, the
following colonizers may come from a predictable set of species. For example, microbial colonization of the healthy mammalian gut
displays specific patterns of species arrival [62, 63]. But is a sterile Studies on microbial invasion often focus on how resident
community composition and species richness affect invasion
outcomes. Here, we chose instead to work with a resident
community whose composition was fixed at the same four species
and ask
how their
environment—specifically
environmental
harshness—and their common evolutionary history affect invasion
success and resistance. The ISME Journal (2022) 16:2644 – 2652 26 2649 104
106
108
1010
At
At(Ac)
At(Kp)
At(Pr)
At(Pf)
Ct
Ms
Oa
Ct(Ac)
Ct(Kp)
Ct(Pr)
Ct(Pf)
Ms(Ac)
Ms(Kp)
Ms(Pr)
Ms(Pf)
Oa(Ac)
Oa(Kp)
Oa(Pr)
Oa(Pf)
104
106
108
1010
Co-evolved community
Ancestral community
CFU/ml at transfer 4
CFU/ml at transfer 4
A
B
*
**
**
*
**
*
*
S
N
S
N
S
N
NS
NS
NS NS
NS
NS
NS
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
NS
At
At(Ac)
At(Kp)
At(Pr)
At(Pf)
Ct
Ms
Oa
Ct(Ac)
Ct(Kp)
Ct(Pr)
Ct(Pf)
Ms(Ac)
Ms(Kp)
Ms(Pr)
Ms(Pf)
Oa(Ac)
Oa(Kp)
Oa(Pr)
Oa(Pf) A B D 104
106
108
1010
At
At(Oa)
Ct
Ct(Oa)
Ms
Ms(Oa)
Ancestral 3-species community
CFU/ml at transfer 4
C
*
NS
NS 104
106
108
1010
At
At(Oa)
Ct
Ct(Oa)
Ms
Ms(Oa)
Co-evolved 3-species community
CFU/ml at transfer 4
D
S
N
S
N
S
N Fig. 3
Bacterial abundance of ancestral or co-evolved community members with or without invader. Each panel represents the total
population size (CFU/ml) at transfer four of a resident member in the ancestral community (A), the co-evolved community (B), the ancestral
three-species community (C) and the co-evolved three-species community (D). The full datasets are in Figs. S2 and S6. All experiments were
performed in MWF. The bacterial abundance of community members without any invader species is represented by colored dots and once
invaded by black dots (invader species indicated in brackets). From left to right: A. tumefaciens (At), C. testosteroni (Ct), M. saperdae (Ms), O. anthropi (Oa), A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. fulva (Pf). We compared the data points of each species when invaded to the
corresponding data points when co-cultured with other community members but without invasion. Statistical significance is marked above
the data points (p values: *<0.05, **<0.01, NS not significant). provide rare experimental support to it. To our knowledge, this
hypothesis has so far only been experimentally tested with single
microbial species invading an ancestral or evolved second species
[53, 68]. gut a harsh environment? Microorganisms colonizing a newborn
gut must survive the acidic conditions of the stomach, the host’s
immune system and bile acids, and cholesterol produced by the
host that are toxic for most microbial species [64]. A few
specialized Lactobacillus and Bifidobacterium species produce bile
resistance proteins [65], which allow them to colonize the gut and
may facilitate the arrival of other species [66]. Similar dynamics
may occur in other systems where strong ecosystem perturbations
clear the ground for new communities to assemble, such as
following antibiotic treatments, or the heavy pollution of soils. However, it remains to be seen whether first colonizers facilitate
future arrivals as would be predicted by the SGH [42, 47, 67], or
whether it is more of a race to fill available niches. [
,
]
But what makes the co-evolved MWF community more resistant
and robust to invasion? Possible explanations are summarized in
Fig. 4. First, more productive communities (greater population
size) are expected to be harder to invade (Fig. 4A) [15]. Although
we observed no significant differences in productivity between
ancestral and co-evolved communities at the time of transfer
(Fig. S4A, B), co-evolved A. tumefaciens and M. saperdae (but not C. testosteroni and O. anthropi) grew faster during the first days
(Fig. S5) and population sizes at the time of invasion were
significantly greater in the co-evolved communities (Fig. S4C). While productivity is a plausible explanation for reduced
invasion success in co-evolved communities, it need not be the
only one [15]. Another explanation could be that communities
with higher species- and strain-level diversity tend to be more
robust against invasions (Fig. 4B) [3, 13, 69]. However, diversity
cannot explain invasion outcomes in our system: by using single But what makes the co-evolved MWF community more resistant
and robust to invasion? Possible explanations are summarized in
Fig. 4. First, more productive communities (greater population
size) are expected to be harder to invade (Fig. 4A) [15]. Although
we observed no significant differences in productivity between
ancestral and co-evolved communities at the time of transfer
(Fig. S4A, B), co-evolved A. tumefaciens and M. saperdae (but not C. testosteroni and O. anthropi) grew faster during the first days
(Fig. S5) and population sizes at the time of invasion were
significantly greater in the co-evolved communities (Fig. S4C). Once a community has assembled despite the challenging
environment, we next asked whether the timing of invasion
matters. In our experiments, early invaders fared better than those
colonizing a community whose species had co-evolved, and co-
evolved species were less perturbed by (more robust against) the
invaders. P. Piccardi et al. 104
106
108
1010
At
At(Ac)
At(Kp)
At(Pr)
At(Pf)
Ct
Ms
Oa
Ct(Ac)
Ct(Kp)
Ct(Pr)
Ct(Pf)
Ms(Ac)
Ms(Kp)
Ms(Pr)
Ms(Pf)
Oa(Ac)
Oa(Kp)
Oa(Pr)
Oa(Pf)
104
106
108
1010
Co-evolved community
Ancestral community
CFU/ml at transfer 4
CFU/ml at transfer 4
A
B
104
106
108
1010
At
At(Oa)
Ct
Ct(Oa)
Ms
Ms(Oa)
Co-evolved 3-species community
CFU/ml at transfer 4
104
106
108
1010
At
At(Oa)
Ct
Ct(Oa)
Ms
Ms(Oa)
Ancestral 3-species community
CFU/ml at transfer 4
C
D
*
**
**
*
**
*
*
S
N
S
N
S
N
NS
NS
NS NS
NS
NS
NS
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
S
N
NS
*
NS
NS
S
N
S
N
S
N
At
At(Ac)
At(Kp)
At(Pr)
At(Pf)
Ct
Ms
Oa
Ct(Ac)
Ct(Kp)
Ct(Pr)
Ct(Pf)
Ms(Ac)
Ms(Kp)
Ms(Pr)
Ms(Pf)
Oa(Ac)
Oa(Kp)
Oa(Pr)
Oa(Pf)
Fig. 3
Bacterial abundance of ancestral or co-evolved community members with or without invader. Each panel represents the total
population size (CFU/ml) at transfer four of a resident member in the ancestral community (A), the co-evolved community (B), the ancestral
three-species community (C) and the co-evolved three-species community (D). The full datasets are in Figs. S2 and S6. All experiments were
performed in MWF. The bacterial abundance of community members without any invader species is represented by colored dots and once
invaded by black dots (invader species indicated in brackets). From left to right: A. tumefaciens (At), C. testosteroni (Ct), M. saperdae (Ms), O. anthropi (Oa), A. caviae (Ac), K. pneumoniae (Kp), P. rettgeri (Pr), P. fulva (Pf). We compared the data points of each species when invaded to the
corresponding data points when co-cultured with other community members but without invasion. Statistical significance is marked above
the data points (p values: *<0.05, **<0.01, NS not significant). On
measuring pairwise interactions between ancestral (Fig. S6G) and
co-evolved species (Figs. S7G and S8F, see Methods), however, we
only observed that positive interactions weakened between A. tumefaciens and C. testosteroni and increased in the other two
species (Figs. S7G and S8G), making this scenario less plausible. Another weakness of our study is the arbitrary choice to
perform four transfers at a 1% dilution rate. To compensate, we
were careful to define our measures independently of these
choices, such that we could compare between treatments rather
than considering absolute measures of invasion. We quantified
“invasion success”, representing absolute population increase or
decrease and “invasion magnitude”, which compares population
sizes between treatments at the end of the experiment. Another
possibility would have been to extend the length of the
experiment to observe whether invaders eventually went extinct
or established themselves. However, as we were interested in the
ecological dynamics of invasion separately from the evolutionary
dynamics of the resident community, we decided to keep the
invasion time-scale short and assume that species’ genetic
adaptation to the environment and each other was negligible. In reality, of course, invaders might acquire mutations that
increase invasion success. A final reason for reduced invasion success in co-evolved
communities
is
that
the
co-evolving
residents
may
have
partitioned the available niches among themselves, leaving little
“space” for new arrivals (Fig. 4D) [31]. Further investigation,
possibly using metabolomics analyses, would be needed to clarify
whether this is the case and to more mechanistically understand
resistance against invasion in our system. Taken together,
although our experimental data cannot conclusively test all the
different explanations (Fig. 4), we find support for the role of
community productivity [15], but cannot exclude increased
interference competitive or niche coverage as additional factors
reducing invasion success. In conclusion, we used a model system to disentangle
interactions between species and measure their effect on
microbial invasion. This revealed that a small, facilitative resident
community can improve the environment for species that would
otherwise be unable to colonize. However, a community whose
residents have adapted to the environment and each other is
more difficult to invade. Our findings corroborate several theoretical studies on
the Community Monopolization Hypothesis [49, 50, 52] and While productivity is a plausible explanation for reduced
invasion success in co-evolved communities, it need not be the
only one [15]. Another explanation could be that communities
with higher species- and strain-level diversity tend to be more
robust against invasions (Fig. 4B) [3, 13, 69]. However, diversity
cannot explain invasion outcomes in our system: by using single The ISME Journal (2022) 16:2644 – 2652 P. Piccardi et al. 2650 Fig. 4
Illustration of our hypotheses as to why invasion into an evolved community may be less successful than into the ancestral
community, leaving resident communities less perturbed. A Because the community might have evolved greater productivity (population
size); B because within-species diversity may be higher with additional strains occupying available niches; C because some evolved strains
may have evolved antagonistic phenotypes to compete with other community members; or D because evolved species better cover available
niches. More than one of these mechanisms could hold simultaneously. Created with BioRender.com. 0 Fig. 4
Illustration of our hypotheses as to why invasion into an evolved community may be less successful than into the ancestral
community, leaving resident communities less perturbed. A Because the community might have evolved greater productivity (population
size); B because within-species diversity may be higher with additional strains occupying available niches; C because some evolved strains
may have evolved antagonistic phenotypes to compete with other community members; or D because evolved species better cover available
niches. More than one of these mechanisms could hold simultaneously. Created with BioRender.com. isolates from the co-evolved communities, our experiments had
the same species- and strain-level diversity in all treatments
(Fig. S9A, C). successful in this context. This would also align with experiments
involving larger communities that presumably occupy more niches
and leave fewer resources for the invader [13, 17–20, 25]. Never-
theless, our community could help to understand the first phases
of community assembly, when only few species have colonized. It could also be that co-evolved communities are more resistant
to invasion because co-evolved residents actively inhibit other
species through interference competition (Fig. 4C) [70]. This is
difficult to test here, as spent media experiments are challenging
with MWF. But if this were true and species had evolved to
produce broad spectrum inhibitory molecules, we might expect
the resident species to interact negatively with one another. DATA AVAILABILITY
All data used in this manuscript are provided in Dataset 1. All data used in this manuscript are provided in Dataset One could also question whether a small synthetic community is
representative of natural communities and their diversity. A
mathematical
model
in
our
previous
study
indicated
that
competition would increase with a higher number of species in
MWF [47] and perhaps we would expect invasions to be less Our work provides new experimental
support
for
the
Community
Monopolization
Hypothesis
[42, 49, 71] and by linking invasion ecology with the SGH,
provides a fresh perspective on community assembly as a
sequence of invasion events into a harsh environment, where
facilitation may be dominant at first as species complement each
other, but decreases as niches are occupied through co-evolution
or through invasion. Our fixed-species experimental design revealed some interest-
ing patterns of invasion success, but also has its limitations. One
confounding factor is that adding amino acids to the growth
medium allowed more species to grow alone, but also provided
new or larger niches for invader species to occupy. This is reflected
in the higher overall invasion magnitude of species in MWF + AA
compared to MWF (Fig. 1C). But despite these additional niches,
invasion magnitude was still lower when the community was
present compared to its absence (Fig. 1C). This made it difficult to
interpret how invaders affected the resident community grown in
MWF + AA: the effects varied depending on the invader species
and the resident species with no clear pattern (Fig. S13). Further
exploring the mechanisms behind the interactions in our system
and developing a theoretical basis for what to expect may help to
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All data used in this manuscript are provided in Dataset 1. DATA AVAILABILITY
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common among culturable bacteria. Sci Adv. 2021;7:7159. 61. Enke TN, Leventhal GE, Metzger M, Saavedra JT, Cordero OX. Microscale ecology
regulates particulate organic matter turnover in model marine microbial com-
munities. Nat Commun. 2018;9:2743. 30. Connell JH, Slatyer RO. Mechanisms of succession in natural communities and
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effects in microbiome assembly. Nat Rev Microbiol. 2022;20:109–21. 32. Datta MS, Sliwerska E, Gore J, Polz MF, Cordero OX. Microbial interactions lead to
rapid micro-scale successions on model marine particles. Nat Commun. 2016;7:11965. 63. Coyte KZ, Rao C, Rakoff-Nahoum S, Foster KR. Ecological rules for the assembly of
microbiome communities. PLoS Biol. 2021;19:e3001116. The ISME Journal (2022) 16:2644 – 2652 P. Piccardi et al. 2652 ACKNOWLEDGEMENTS We thank Oliver Meacock, Shota Shibasaki, and three anonymous reviewers for useful
and constructive feedback on the manuscript. We thank Christopher van der Gast
and Ian Thompson for providing the four species used to assemble the resident
community and Peter Küenzi for providing the four additional species used as
invaders. We thank Flor-Inés Arias-Sánchez for designing the selective media. PP and
GA were funded by the University of Lausanne, SM by European Research Council
Starting Grant 715097. JMA received funding from the European Union’s Horizon
2020 research and innovation programme under grant agreement No. 678841. Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative
Commons license, and indicate if changes were made. The images or other third party
material in this article are included in the article’s Creative Commons license, unless
indicated otherwise in a credit line to the material. If material is not included in the
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creativecommons.org/licenses/by/4.0/. COMPETING INTERESTS 66. Gérard P. Metabolism of cholesterol and bile acids by the gut microbiota. Pathogens. 2013;3:14–24. The authors declare no competing interests. 67. Amarnath K, Narla AV, Pontrelli S, Dong J, Caglar T, Taylor BR, et al. Stress-induced
cross-feeding of internal metabolites provides a dynamic mechanism of micro-
bial cooperation. bioRxiv. 2021. https://doi.org/10.1101/2021.06.24.449802. AUTHOR CONTRIBUTIONS PP, GA, JMA and SM designed research. GA and PP performed the experiments. PP,
GA, JMA and SM analyzed the data. PP and SM wrote the manuscript with input from
GA and JMA. © The Author(s) 2022 © The Author(s) 2022 © The Author(s) 2022 The ISME Journal (2022) 16:2644 – 2652
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Tropical Paradise: Its Light and Its Shadows
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Citation Myer, Angela. “Tropical Paradise: Its Light and Its Shadows.”
Powerpoint presentation. Refereed Proceedings of the Inaugural
Tropics of the Imagination Conference, 2-3 November 2009. etropic:
electronic journal of studies in the tropics, Vol 9 (2010), <http:// Myer, Angela. “Tropical Paradise: Its Light and Its Shadows.”
Powerpoint presentation. Refereed Proceedings of the Inaugural
Tropics of the Imagination Conference, 2-3 November 2009. etropic:
electronic journal of studies in the tropics, Vol 9 (2010), <http:// Myer, Angela. “Tropical Paradise: Its Light and Its Shadows.”
Powerpoint presentation. Refereed Proceedings of the Inaugural
Tropics of the Imagination Conference, 2-3 November 2009. etropic:
electronic journal of studies in the tropics, Vol 9 (2010), <http:// www.jcu.edu.au/etropic> www.jcu.edu.au/etropic> Angela Myer Angela Myer Tropical Paradise:
Its Light and Its Shadows Angela Myer To define the inherent quality found in the
tropical paintings of Far North Queensland
and locate this research in the context of
Australian paintings - considering whether
this is a unique chapter in Australian Art. This notion of tropical light is compared to the
aesthetics of tropical paintings by artists such
as Gauguin and Matisse. Specifically North
Queensland occupies a wet tropic zone, and
this contributes to a particular visual stimulus. This notion of tropical light is compared to the
aesthetics of tropical paintings by artists such
as Gauguin and Matisse. Specifically North
Queensland occupies a wet tropic zone, and
this contributes to a particular visual stimulus. The research builds on European
traditions of paintings of the exotic and
Orient, via largely French influences. The research builds on European
traditions of paintings of the exotic and
Orient, via largely French influences. The research builds on European
traditions of paintings of the exotic and
Orient, via largely French influences. Extending into the Pacific Ocean Ray Crooke
empathised with the island lifestyle and the
casual attitudes which were advantageous for
an artist looking for models to draw. Extending into the Pacific Ocean Ray Crooke
empathised with the island lifestyle and the
casual attitudes which were advantageous for
an artist looking for models to draw. “When I go into the glass houses and I see
the strange plants of exotic lands, it seems to
me that I enter into a dream." Living in tropical
North Queensland may inhabit a Rousseau
dream. “When I go into the glass houses and I see
the strange plants of exotic lands, it seems to
me that I enter into a dream." Living in tropical
North Queensland may inhabit a Rousseau
dream. With a fascination for floral motifs Georgia O’Keeffe
participated in a changing aesthetic of modernism. Her
iconography is rooted in shells, rocks, bones and
flowers as well as spacious landscapes which morph
into luscious abstractions through her delicate
watercolours. With a fascination for floral motifs Georgia O’Keeffe
participated in a changing aesthetic of modernism. Her
iconography is rooted in shells, rocks, bones and
flowers as well as spacious landscapes which morph
into luscious abstractions through her delicate
watercolours. Italo Calvino suggests that “our ability to
desire a different kind of life, and our very
way of looking at the world: as a total way of
looking that sets us inwardly free to free
ourselves outwardly.” (Calvino: 1987, p. 222) Italo Calvino suggests that “our ability to
desire a different kind of life, and our very
way of looking at the world: as a total way of
looking that sets us inwardly free to free
ourselves outwardly.” (Calvino: 1987, p. 222) The unquantifiable realm of paradise is
attractive as it allows our imagination to run
wild, and conjure up our most desirable
aspirations. Certainly Michelangelo accepted
this cue with relish. The unquantifiable realm of paradise is
attractive as it allows our imagination to run
wild, and conjure up our most desirable
aspirations. Certainly Michelangelo accepted
this cue with relish. The research builds on European
traditions of paintings of the exotic and
Orient, via largely French influences. John Olsen likened North Queensland to a
Mahler symphony, which is harsh, discordant
and frenetic, and may suggest the gentle
breezes, mosquitoes and melting heat of
Cairns John Olsen likened North Queensland to a
Mahler symphony, which is harsh, discordant
and frenetic, and may suggest the gentle
breezes, mosquitoes and melting heat of
Cairns “My themes are influenced by my
spiritual beliefs. Currently, I go back
and forth between surrealism and
realism, like a bird with two nests.” “My themes are influenced by my
spiritual beliefs. Currently, I go back
and forth between surrealism and
realism, like a bird with two nests.” Perhaps Gauguin needed more of
everything, more colour, eroticism,
abandonment, freedom and moral
licence to discover and devour Perhaps Gauguin needed more of
everything, more colour, eroticism,
abandonment, freedom and moral
licence to discover and devour Rothenberg flies in the face of the
doomed plight of women
suggested by De Beauvoir. The theatre of expression taps into
highly emotive realms of the feminine
condition as in the case of Frida Khalo. The theatre of expression taps into
highly emotive realms of the feminine
condition as in the case of Frida Khalo. The theatre of expression taps into
highly emotive realms of the feminine
condition as in the case of Frida Khalo. Men are from Mars and Women are from Venus
comments about marriage, “Through understanding
the different languages we speak it is much easier to
recognize that our partners do love us and that they, in
their own way, are doing their best.” (Gray: 1994, p. 159) Men are from Mars and Women are from Venus
comments about marriage, “Through understanding
the different languages we speak it is much easier to
recognize that our partners do love us and that they, in
their own way, are doing their best.” (Gray: 1994, p. 159) When Kant articulates his notion of the
sublime and the beautiful, he is referring to
truly virtuous essences of feeling, very
primary reactions to things. When Kant articulates his notion of the
sublime and the beautiful, he is referring to
truly virtuous essences of feeling, very
primary reactions to things. When Kant articulates his notion of the
sublime and the beautiful, he is referring to
truly virtuous essences of feeling, very
primary reactions to things. Whether the art of the Tropics will achieve Kant’s
proposition that true virtue can be raised by an
appreciation and understanding of the beautiful and the
sublime, and thus enact the principle s of humanity’s
beauty and dignity, is a question I am willing to ask. Whether the art of the Tropics will achieve Kant’s
proposition that true virtue can be raised by an
appreciation and understanding of the beautiful and the
sublime, and thus enact the principle s of humanity’s
beauty and dignity, is a question I am willing to ask. The tropics are not just an escape, or
inspiration, but as necessary as the air
we breathe to get some clarity and
focus in our minds. The tropics are not just an escape, or
inspiration, but as necessary as the air
we breathe to get some clarity and
focus in our minds. It is as though exoticism from the
tropics allows a new conversation
within modernism and indeed culture. It is as though exoticism from the
tropics allows a new conversation
within modernism and indeed culture. It is as though exoticism from the
tropics allows a new conversation
within modernism and indeed culture.
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The study of n-3PUFAs protecting the intestinal barrier in rat HS/R model
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Lipids in health and disease
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Abstract Background: N-3 PUFAs have been demonstrated in vitro it could prevent the intestinal tight junctions (TJs) from
the ischemia/re-perfusion injury and the inflammatory reaction injury. The purpose of this study was to evaluate the
protection of n-3 PUFAs on the intestinal TJs in the rat model of hemorrhagic shock followed by resuscitation. Methods: Male SD rats (n = 72; 250 ~ 300 g) were randomly divided into 6 groups: SHAM, hemorrhagic shock (HS),
hemorrhagic shock/resuscitation (HS/R), ω-6 group, ω-3 group and ω-3 treatment group. Shock was induced, and
a mean arterial pressure was maintained at 35 to 40 mmHg for 60 minutes. Resuscitation was carried out by
returning half of the shed blood and Ringer’s lactate solution. In ω-6 and ω-3 group, Intralipid or fish oil (0.2 g/Kg),
respectively, was infused 30 minutes after shock. And fish oil was infused with resuscitation in ω-3 treatment group. Half of each group was killed at 30 minutes and 4 hours after resuscitation, respectively. The serum samples and
the intestinal sample was collected for further examination. Background: N-3 PUFAs have been demonstrated in vitro it could prevent the intestinal tight junctions (TJs) from
the ischemia/re-perfusion injury and the inflammatory reaction injury. The purpose of this study was to evaluate the
protection of n-3 PUFAs on the intestinal TJs in the rat model of hemorrhagic shock followed by resuscitation. Result: There is no difference between ω-3, ω-3 treatment and sham group in Chiu’s score, but the other three
groups have higher scores than they did. Compared with HS, HSR and ω-6 group, ω-3 and ω-3 treatment group
showed most intact in intestinal mucoscal villi and TJs through HE, SEM and LSCM. The levels of IL-6 and TNF-α of
bowel tissue in ω-3 and ω-3 treatment group were significantly lower than HS and HSR groups’. At the time point
of 30 min, the levels of serum endotoxin were dramatically higher in HS、HSR and ω-6 groups when compared
with ω-3, ω-3 treatment and sham group. However, it in ω-3 group was greater than sham and HS group until 4
hours. Conclusion: Fish oil pretreatment before resuscitation showed a beneficial effect to the intestinal TJs and
atteunated inflammation after H/R in HS/R rat model and is better than ω-6 PUFAs did. Keywords: Hemorrhagic shock, Resuscitation, Tight junction, Occludin protein, I-FABP, n-3 PUFA © 2014 Li et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative
Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain
Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article,
unless otherwise stated. The study of n-3PUFAs protecting the intestinal
barrier in rat HS/R model Yang Li†, Xinying Wang†, Ning Li and Jieshou Li* Yang Li†, Xinying Wang†, Ning Li and Jieshou Li* * Correspondence: xiaoran8210@gmail.com
†Equal contributors
Research Institute of General Surgery, Jinling Hospital, School of Medicine,
Nanjing University, 305 Zhongshan East Road, Nanjing 210002, China RESEARCH
Open Access
The study of n-3PUFAs protecting the intestinal
barrier in rat HS/R model
Yang Li†, Xinying Wang†, Ning Li and Jieshou Li*
Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 RESEARCH
Open Access
The study of n-3PUFAs protecting the intestinal
barrier in rat HS/R model
Yang Li†, Xinying Wang†, Ning Li and Jieshou Li*
Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Characteristics and shock preparation of rats All animals survived from shock and completed resuscita-
tion in the four groups. Among the groups, there was no
difference in the change of blood pressure into baseline
following shock and resuscitation (Table 1). Except for the
sham group, all animals were maintained at 35 to 40 mm
Hg for 60 minutes by periodic withdrawal or return of
blood. There were no detectable differences between the
four groups in terms of body weight, cannulation time, or
amount of blood withdrawn. Introduction (Tjs), serve as a fence between two cells, play a major role
in this mechanism. Hypoperfusion or ischemia can cause
disruption of the Tjs, with subsequent barrier failure. The
loss of intestinal cell integrity leads to paracellular leakage
of microbial products [3-6]. Hemorrhagic shock, an independent risk factor for
multiple-organ dysfunction syndrome (MODS) [1], is the
leading cause of death in traumatic patients. It followed by
resuscitation is considered as an insult frequently induces
a systemic inflammatory response syndrome (SIRS) that
results in MODS, which is a major clinical problem. The
concept has developed that the intestinal injury/resuscita-
tion (I/R) injury associated with hemorrhagic shock (HS)
followed by fluid resuscitation is responsible for the failure
of the intestinal barrier, bacterial translocation, systemic
sepsis, and even MODS [2]. Meanwhile, tight junctions The influence of the n-3 polyunsaturated fatty acids
(n-3 PUFAs) on inflammatory bowel disease indicated
50% reduction in steroid use and improved histology [7],
delayed relapse [8], reduced disease activity and drugs
usage [4]. Also, the influence of the n-3 fatty acids on
patients under surgical stress indicated reduced wound
and major infections, reduced intra-abdominal abscess
and multiple organ failure, and shortened hospital stay
in several prospective randomized studies in burn,
trauma and major surgery patients. The benefit of n- * Correspondence: xiaoran8210@gmail.com
†Equal contributors
Research Institute of General Surgery, Jinling Hospital, School of Medicine,
Nanjing University, 305 Zhongshan East Road, Nanjing 210002, China Page 2 of 9 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 response to hemorrhagic shock. TNF-α was markedly el-
evated in HS, HS/R, ω-6 group and ω-3 treatment group
(Table 2). However, there was no significant difference
between the control and fish oil groups. 3PUFAs on barrier barrier also was found in sever pan-
creatitis, trauma and septic shock. However, the effect of
the n-3PUFAs on intestinal permeability in hemorrhagic
shock/resuscitation (HS/R) is rarely reported [9]. So we make a hypothesis that intravenous n-3 PUFAs
would improve intestinal barrier function in a rat model
of hemorrhagic shock with resuscitation. Histology Hemorrhagic shock induced the slight intestinal inflam-
mation of the jejunum in the rats (Figure 2B), and the
intestine mucosa was destroyed after resuscitation or in-
fused by ω-6 PUFAs (Figure 2C,D). In rats, no matter
when injected fish oil, no distinguishable changes of in-
testinal histopathology were found compared with sham
rats (Figure 2E,F). On the basis of a mucosal damage
score, infusion of ω-3PUFAs was found to prevent intes-
tinal mucosal damage induced by ischemia/re-perfusion. However, treatment of ω-6PUFAs cannot reverse the ef-
fect of ischemia/re-perfusion injury on intestinal mucosa
(Figure 2G). Tight junctions ultrastructure In sham animals, the tight junctions appeared as typical
membrane fusions with intact TJs structure (Figure 3A). In contrast, TJs ultrastructure were altered after shock,
resuscitation or ω-6PUFAs infusion. Tight junctions
were discontinuous with few membrane fusions appar-
ently in colon tissues, which indicated disruption of TJs
morphology (Figure 3B-D). There were no morpho-
logical differences with the tight junctions in ω-3 group
and ω-3 treatment group compared with sham group
(Figure 3E-F). Lipid analysis Levels of docosahexenoic acid (DHA) and eicosapente-
noic acid (EPA) were found to be significantly elevated
in plasma in ω-3 group and ω-3 treatment group as
compared with other groups, whereas levels of arachi-
donic acid (AA) were correspondingly reduced. The
levels of DHA and EPA in intestinal epithelial cells had
the same change trend, although there were no signifi-
cant differences in among all groups (Tables 3 and 4). IL-6 and TNF-α in tissue Interleukin-6 (IL-6) was measured 4 hours postresuscita-
tion because it characteristically has a later response. Interestingly, the level of IL-6 were elevated in ω-3
group and ω-3 treatment group animals without signifi-
cant difference compared with the sham group. But the
levels of IL-6 in the other groups were elevated dramat-
ically (Table 2). I-FABP and endotoxin in plasma HS/R group has the highest concentration of intestinal
fatty acid binding protein (I-FABP) in all the groups. 30 min and 4 hours after resuscitation, the levels of I-
FABP in ω-3 group and ω-3 treatment group had little
change compared with sham group, but they were sig-
nificantly lower than HS, HS/R and ω-6 group (p < 0.05). (Figure 1A, B) 30 min after resuscitation, the levels of endotoxin in
ω-3 group and ω-3 treatment group were remarkablely
lower than HS, HS/R and ω-6 group (p < 0.05), however,
they increased 4 hours although there were no differ-
ences compared with HS/R and ω-6 group, respectively. (Figure 1C, D) Redistribution of occludin in epithelium Occludin was precisely localized in the villus surface
of epithelial cells in jejunum tissues from sham rats
(Figure 4A). After shock, resuscitation or intralipid infu-
sion, the distribution of occludin was altered and the The concentration of tumor necrosis factor-α (TNF-α)
was measured 30 minutes after shock and resuscitation
because it is known to be characteristic of an early Table 1 Hemorrhagic shock parametersa
sham
HS
HS/R
ω-6 group
ω-3 group
ω-3 Tre
P
Weight,(g)
232.4 ± 14.7
240.2 ± 16.4
223.3 ± 9.43
230.67 ± 11.0
230.8 ± 8.87
240.0 ± 10.8
NS
Cannulation time,(min)
20.13 ± 1.69
20.78 ± 1.40
20.60 ± 2.07
21.63 ± 1.25
19.12 ± 1.43
22.11 ± 3.61
NS
Blood removed, (ml)
5.34 ± 0.27
5.82 ± 0.23
6.15 ± 0.26
5.86 ± 0.64
6.03 ± 0.52
5.97 ± 0.49
NS
NS, not significant. aData are given as mean ± standard error of the mean. Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Page 3 of 9 Figure 1 The levels of I-FABP and endotoxin of plasma. I-FABP in Sham group, ω-3 group and ω-3 treatment group were significantly lower
than HS,HS/R and ω-6 group (p < 0.05). The datas in 30 min and 4 hours have the same trend (A, B). The levels of endotoxin in ω-3 group and
ω-3 treatment group were dramatically increased in 30 min when compared with 4 hours (C, D). Figure 1 The levels of I-FABP and endotoxin of plasma. I-FABP in Sham group, ω-3 group and ω-3 treatment group were significantly lower
than HS,HS/R and ω-6 group (p < 0.05). The datas in 30 min and 4 hours have the same trend (A, B). The levels of endotoxin in ω-3 group and
ω-3 treatment group were dramatically increased in 30 min when compared with 4 hours (C, D). epithelium was disrupted (Figure 4B,C,D). Occludin
from ω-3 group and ω-3 treatment group had no differ-
ence compared with that of sham group. certain acute systemic inflammatory diseases such as
asthma, chronic obstructive pulmonary disease, and in-
flammatory bowel disease [10]. And there have been
numerous researches about ω-3 PUFAs in critically ill
patients [11,12]. However, clinical use of ω-3 PUFAs in
the systemic inflammatory response syndrome and intes-
tinal barrier function in hemorrhagic shock remains
lacking in evidences. Discussion Hemorrhagic shock followed by resuscitation is known
to stimulate a systemic inflammatory response and intes-
tinal barrier dysfunction. Preventing the catastrophic
consequences of hemorrhagic shock remains an un-
solved
problem in clinical treatment of trauma. Daily
dietary supplementation with ω-3 PUFAs has been well
established as a method to achieve an alleviation of In our study, mucosal Damage Grading Score and H&E-
stained sections of jejunum indicated that ω-3 PUFAs alle-
viates morphological changes induced by ischemia and
re-perfusion. The transmission electron microscopy (TME) Table 2 Results of cytokine protein measurements (pg/mL tissue)
sham
HS
HS/R
ω-6 group
ω-3 group
ω-3 Tre
IL-6 (4 h)
35.23 ± 6.16
91.50 ± 33.62*
122.22 ± 41.28**
161.32 ± 19.31**
53.47 ± 25.81
71.44 ± 32.21
TNF-α (30 min)
30.84 ± 9.85
90.39 ± 33.76*
141.29 ± 75.43**
126.99 ± 19.89**
56.85 ± 17.46
88.12 ± 8.09
*P < 0.05. Data are given as mean ± standard error of the mean. Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Page 4 of 9 Table 3 Fatty acids in plasma (g/L)
sham
HS
HS/R
ω-6 group
ω-3 group
ω-3 Tre
P
EPA (c 20:5ω-3)
0.14 ± 0.00
0.11 ± 0.05
0.10 ± 0.02
0.15 ± 0.01
18.8 ± 5.7
19.1 ± 6.5
0.01
DHA (c 22:6ω-3)
3.5 ± 1.69
2.1 ± 0.74
2.0 ± 0.89
4.5 ± 0.25
16.4 ± 4.5
20.11 ± 3.61
0.01
AA (C20:4ω-6)
10.5 ± 3.17
11.2 ± 3.23
9.1 ± 4.01
21 ± 4.61
9.20 ± 3.42
10.97 ± 5.39
0.01
Data are given as mean ± standard error of the mean. AA, arachidonic acid; DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid. Table 3 Fatty acids in plasma (g/L) associated with the change of fatty acids in plasma and
epithelial cells (Tables 3 and 4). It is the administration
of ω-3 PUFAs that promotes the formation of eicosanoids,
which are anti-inflammatory rather than pro-inflammatory
mediators. In addition, intravenous infusion of ω-3 PUFAs
leads to decreases of prostaglandin E2, thromboxane
A2, and leukotriene B4, accompanied with increases of
thromboxane A3, prostacyclin PGI3, and leukotriene
B5. Generally, the leukotrienes, thromboxanes, and
prostaglandins derived from ω-3 PUFAs are less pro-
inflammatory than the homologus compounds derived
from ω-6 PUFAs. Discussion So, the levels of DHA and EPA signifi-
cantly elevated in plasma in ω-3 group and ω-3 treat-
ment group were the primary cause for less systemic
inflammation in the two groups. There are, however,
powerful anti-inflammatory metabolites from the ω-6
PUFAs AA, and one cannot say with validity that “ω-6
are pro-inflammatory and ω-3 are not” [19]. So we did
not find the inflammatory response becomes worse in
the rats of ω-6 group after ω-6 PUFAs infusion. assay revealed that ω-3 PUFAs extenuated the discontinity
and disruption of TJs in the mucosa. By use of immuno-
fluorescence, we demonstrated that ω-3 PUFAs could par-
tially prevent the decreased expression of occludin in
jejunum mucosa during hemorrhagic shock followed by re-
suscitation. These results are similar with other experimen-
tal results and the protective effect of fish oil attributed to
inflammation reaction alleviating and inflammation factors
reducing. Preclinical and clinical research is depicting a
therapeutic effect for consumption of the fish-oil derived in
the management of inflammatory disease in general [13]. The
small
intestine
is
highly
sensitive
to
hypo-
perfusion injury because of higher mucosal cell oxygen
demand [14,15]. Thus, subjected to hemorrhagic shock
and resuscitation, trauma, and surgery often develop
intestinal ischemia and tissue damage, which has been
documented by both experimental [16] and clinical stud-
ies [17]. In current research, we demonstrated that the
administration of ω-3 PUFAs could partly attenuate
intestinal epithelial barrier dysfunction in hemorrhagic
shock and resuscitation rats. The levels of I-FABP in
plasma were significantly decreased after ω-3 PUFAs
administration. Our preliminary study showed the intes-
tinal barrier is severely damaged after I/R, which is
related to the redistribution of I-FABP and indicated
that n-3 PUFAs protect the intestinal barrier by modify-
ing
intracellular
I-FABP,
activating
the
peroxisome
proliferator-activated receptor-γ (PPARγ) pathway, and
then upregulating TJs protein expression [18]. However, the levels of endotoxin in ω-3 group and ω-3
treatment group were dramatically escalated at 4 hours
after resuscitation when compared with them in 30 min. What makes the serum endotoxin levels grow up, further-
more why the elevated endotoxin concentration did not
induce more inflammatory response and gut barrier dam-
age? Pluess et al. Discussion have domesticated that healthy volun-
teers received twice 0.5 g/kg fish oil at 48 and 24 h before
lipopolysaccharide
(LPS)
injected
would
modify
the
phospholipid composition of platelets and blunted fever
and increased the neuroendocrine and the inflammatory
responses to LPS [20]. Meanwhile, in their previous study
in healthy subjects they showed that prolonged supple-
mentation with oral fish oil blunts the neuroendocrine
and inflammatory responses to LPS challenge [21]. Simi-
larly, Pittet et al. have divided male subjects into three
groups which received three different fish oil doses before In the present study, we found that administration of ω-
3 PUFAs regardless of being before or during resuscitation
would reduce the systemic inflammation. The levels of IL-
6 and TNF-α in intestinal tissue of ω-3 group and ω-3
treatment group were dramatically lower than HS, HSR
and ω-6 groups’ (Table 2). This phenomenon may be
explained as the effects of ω-3 PUFAs on regulation of
host inflammatory responses. And this phenomenon is Table 4 aFatty acids in intestinal epithelial cells
sham
HS
HS/R
ω-6 group
ω-3 group
ω-3 Tre
P
EPA (c 20:5ω-3)
2.01 ± 0.45
1.98 ± 0.12
2.10 ± 0.25
2.41 ± 0.51
5.47 ± 1.21
5.23 ± 1.5
NS
DHA (c 22:6ω-3)
2.16 ± 0.78
1.59 ± 0.44
2.50 ± 0.49
3.12 ± 0.75
6.45 ± 0.23
5.10 ± 1.45
NS
AA (C20:4ω-6)
15.3 ± 4.15
16.2 ± 3.63
17.1 ± 4.54
25 ± 5.46
17.5 ± 4.44
16.7 ± 3.72
NS
NS, not significant. aData are given as mean ± standard error of the mean. AA, arachidonic acid; DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid. Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Page 5 of 9 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Figure 2 Histological findings of the jejunum from rats (original magnification, 20 × 20). (A) Sham animals. (B) Hemorrhagic shock. (C) Hemorrhagic shock and resuscitation. (D) Intralipid infused before resuscitation. (E) Fish oil infused before resuscitation. (F) Fish oil infused
during resuscitation. (G) Mucosal damage grading was assessed. The mean histological score of animals in HS group, HSR group and omega-6
group are significantly higher than that of sham mice, and subsequent treatment of ω-3 PUFAs could not attenuate this damage. Figure 2 Histological findings of the jejunum from rats (original magnification, 20 × 20). (A) Sham animals. (B) Hemorrhagic shock. Materials and methods
Animals LPS. And the 0.2 g/kg per-fusion immediately (2 hours)
before LPS was the most efficient in blunting the re-
sponses, suggesting LPS capture in addition to the sys-
temic and membrane effects. These studies may explain
the phenomenon found in our experiments, although the
concentration of endotoxin magnified, the infusion of fish
oil will blunt the inflammatory and neuroendocrine re-
sponses to alleviate the damage of gut mucosa. Seventy-two male SD rats of a mean (±SD) weight of 269.26 ±
12.68 g were used and the protocol of this study was ap-
proved by the institutional animal care committee. Animals,
supplied by Medical Experiment Animal Center of Jinling
Hospital (Nanjing, China), were kept in a chamber with a
light cycle from 9 am to 9 pm and controlled temperature
of 25°C–28°C with food and water available ad libitum. Characteristics of the study groups are presented in Table 5. Conclusion
l In conclusion, our data indicate—to our knowledge—
that ω-3 PUFAs can ameliorate intestinal epithelium TJ
disruption in a rat model of hemorrhagic shock followed
by resuscitation. We speculate that ω-3 PUFAs, individu-
ally or in combination with other therapy, may serve as
a potential new therapeutic agent that restores the nor-
mal intestinal barrier function in hemorrhagic shock. Discussion (C) Hemorrhagic shock and resuscitation. (D) Intralipid infused before resuscitation. (E) Fish oil infused before resuscitation. (F) Fish oil infused
during resuscitation. (G) Mucosal damage grading was assessed. The mean histological score of animals in HS group, HSR group and omega-6
group are significantly higher than that of sham mice, and subsequent treatment of ω-3 PUFAs could not attenuate this damage. Rat model of hemorrhagic shock and resuscitation
ll
h d
d
h
d
h g
All rats were weighed and anesthetized with 60 mg/kg
ketamine intramuscular injection and then placed on a
temperature-controlled heating pads, which was heated
while shock was induced and temperature was sustained
37°C. Using aseptic techniques, the left femoral artery and Page 6 of 9 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Figure 3 Tight junction morphology was altered at 4 hours after resuscitation. The intact TJ structure was observed in sham rats
(A). TJ ultrastructure was altered characterized by decreased electron dense materials in the TJ (B–D). TJ ultrastructure was integrity as sham
group (E–F). Arrows, tight junctions. Bars = 0.5 μm. Figure 3 Tight junction morphology was altered at 4 hours after resuscitation. The intact TJ structure was observed in sham rats
(A). TJ ultrastructure was altered characterized by decreased electron dense materials in the TJ (B–D). TJ ultrastructure was integrity as sham
group (E–F). Arrows, tight junctions. Bars = 0.5 μm. vein were isolated and cannulated with a short incision
containing 0.1 ml heparinized saline (10 units/ml) [22]. Heparin (200 U/kg) was infused immediately through
femoral vein following cannulation. The artery catheter
was connected to a multi-channel recorder (Electronics
for Medi- cine, TX) for the monitoring of systolic, dia-
stolic, and mean arterial pressures (MAP) per 3 minutes. The target MAP (35 ~ 40 mm Hg) was induced by re-
moving blood slowly through the femoral vein cannula
during a total time of 15 minutes and maintained at 35
to 40 mm Hg with further withdrawals of blood as
required for 60 minutes. At the point of 75 minutes, re-
suscitation which would take 30 minutes was imple-
mented by returning half of the shed blood and Ringer’s
solution to the animal at the speed of 0.5 mL/min. And
Infusion was withheld if MAP exceeded 90% of baseline
and re-started if declined. At the 105th minute, resusci-
tation was finished and no interventions were carried
out. The rats in ω-6 group and ω-3 group were injected
by intralipid and fish oil (Omegaven, Huarui Pharma-
ceutical Co., Ltd. 0.2 g/Kg) respectively through the
caudal vena after shock 30 min. And the rats in ω-3
treatment group were injected by fish oil (0.2 g/Kg) during resuscitation. Rat model of hemorrhagic shock and resuscitation
ll
h d
d
h
d
h At the end of 30 minutes of resus-
citation, half of the animals of these three groups were
randomly sacrificed to collect blood samples, and the
others were killed at the end of 4 hours. The 2–3 ml
sample was centrifuged at 4°C. After centrifugation,
serum was kept refrigerated at -80°C until assayed. Measurement of I-FABP and endotoxin in plasma
The blood sample was centrifuged at 10 000 × g for
15 min at 4°C. The supernatant was assayed for I-FABP
and endotoxin levels by enzyme-linked immunosorbent
assay kits (R&D Systems, Minneapolis, MN, USA) in ac-
cordance with the manufacturer’s instructions. The ab-
sorbance at 450 nm was determined using a microplate
reader (Tecan’s, Sunrise, Unterberg strasse IA, Austria). Measurement of IL-6 and TNF-α in tissue The jejunum at 2 cm beyond Treitz ligament was
weighed and homogenized by adding 9 times gravimetric
saline. The 10% homogenate was centrifuged for 10 mi-
nutes (2500 r/min) and the supernatant was diluted 10
times with normal saline to 1% concentration. The 1%
supernatant was assayed for IL-6 and TNF-a levels by Page 7 of 9 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Figure 4 Redistribution of occludin in hemorrhagic shock rats. Occludin (A, E, F) was precisely localized to the tight junction in the jejunum
from sham group, ω-3 group and ω-3 treatment group. In contrast, the jejunum from shock, resuscitation and ω-6 group showed redistribution
of occludin (B, C, D). Bars = 25 mm. Figure 4 Redistribution of occludin in hemorrhagic shock rats. Occludin (A, E, F) was precisely localized to the tight junction in the jejunum
from sham group, ω-3 group and ω-3 treatment group. In contrast, the jejunum from shock, resuscitation and ω-6 group showed redistribution
of occludin (B, C, D). Bars = 25 mm. enzyme-linked immunosorbent assay kits (R&D Systems,
Minneapolis, MN, USA) in accordance with the manu-
facturer’s instructions. intestine samples were first homogenized and the cell pel-
let isolated by centrifugation and frozen until analysis. Total small intestine lipids were extracted with methanol
and methylene (both containing 50 mg/L BHT) as de-
scribed [23]. The organic phase was collected and the
solvent evaporated under nitrogen in a 45°C water bath. The samples were methylated as described [24]. The sam-
ples were analyzed by gas chromatography (GC-14A; Fatty acid analysised To verify that the treatments change resulted in the in-
corporation of significant amounts of DHA and EPA into
membrane lipids and plasma, fatty acid analysised was car-
ried out in intestinal epithelial cells and plasma. Small Table 6 Intestinal mucosal damage grading score Table 6 Intestinal mucosal damage grading score Table 6 Intestinal mucosal damage grading score
Grade
Histological characteristic(s)
Grade 0
Normal mucosal villi
Grade 1
Subepithelial Gruenhagen’s space (oedema),
usually at the apex of the villus
Grade 2
Extension of the subepithelial space with moderate
lifting of epithelial layer from the lamina propria
Grade 3
Massive epithelial lifting down the sides of villi;
a few tips may be denuded
Grade 4
Denuded villi with lamina propria and dilated
capillaries exposed
Grade 5
Digestion and disintegration of lamina propria;
haemorrhage and ulceration Table 6 Intestinal mucosal damage grading score
Grade
Histological characteristic(s)
Grade 0
Normal mucosal villi
Grade 1
Subepithelial Gruenhagen’s space (oedema),
usually at the apex of the villus
Grade 2
Extension of the subepithelial space with moderate
lifting of epithelial layer from the lamina propria
Grade 3
Massive epithelial lifting down the sides of villi;
a few tips may be denuded
Grade 4
Denuded villi with lamina propria and dilated
capillaries exposed
Grade 5
Digestion and disintegration of lamina propria;
haemorrhage and ulceration Competing interests
Th
h
d
l
h The authors declare that they have no competing interests. The authors declare that they have no competing interests. Histopathological evaluation
h
l n-3 PUFAs: n-3 polyunsaturated fatty acids; TJ: Tight junction;
HS: Hemorrhagic shock; HS/R: Hemorrhagic shock/resuscitation; MAP: Mean
arterial pressure; HE: Histologic examination; SEM: Scanning electron
microscopy; LSCM: Laser scanning confocal microscope; MODS: Multiple-
organ dysfunction syndrome; SIRS: Systemic inflammatory response
syndrome; I-FABP: Intestinal fatty acid binding protein;
LPS: Lipopolysaccharide. The proximal jejunum specimens were harvested imme-
diately after the animals were sacrificed, and rinsed in
cold saline with subsequent immersion in 10% buffered
formalin over- night at room temperature. Sections
measuring 5-μm in thickness were cut down, placed on
glass slides, and stained with hematoxylin and eosin
(H&E). Images were obtained using a Zeiss Image A1
light microscope at × 20 magnification with AxioVision
V4.5 software. Three pathologists, blinded to the source
of the slides, analyzed and made a report about every
slide. The degree of histopathologic changes was graded
semiquantitatively by using the histologic injury score
described by Chiu et al. [28] (Table 6). Table 5 Characteristics of study groups Table 5 Characteristics of study groups Table 5 Characteristics of study groups
SHAM (n = 12)
control (cannulation and Heparinized)
HS (n = 12)
control + shock
HS/R (n = 12)
HS + fluid resuscitation
ω-6 group (n = 12)
HS/R + intralipid (0.2 g/kg) intravenous
after shock 30 min
ω-3 group (n = 12)
HS/R + fish oil (0.2 g/kg) intravenous
after shock 30 min
ω-3 treatment group (n = 12)
HS/R + fish oil (0.2 g/kg) intravenous
during resuscitation
HS, hemorrhagic shock; HS/R, hemorrhagic shock and resuscitation. Table 5 Characteristics of study groups
SHAM (n = 12)
control (cannulation and Heparinized)
HS (n = 12)
control + shock
HS/R (n = 12)
HS + fluid resuscitation
ω-6 group (n = 12)
HS/R + intralipid (0.2 g/kg) intravenous
after shock 30 min
ω-3 group (n = 12)
HS/R + fish oil (0.2 g/kg) intravenous
after shock 30 min
ω-3 treatment group (n = 12)
HS/R + fish oil (0.2 g/kg) intravenous
during resuscitation
HS, hemorrhagic shock; HS/R, hemorrhagic shock and resuscitation. Page 8 of 9 Li et al. Lipids in Health and Disease 2014, 13:146
http://www.lipidworld.com/content/13/1/146 Shimadzu, Columbia, MD) using a fused silica capillary
column (SP-2560, 100 m; Supelco, Bellefonte, PA). Fatty
acid methyl esters are reported as percent of total and
were identified by comparison with known standards. were incubated with Alexa 488-conjugated secondary
antibodies for 60 min. DAPI was used for staining DNA
of cells. Images were captured using a Leica TCS SP2
laser confocal scanning microscope (Leica Micro- sys-
tems, Heidelberg GmbH, Mannheim, Germany). DHA and EPA in plasma were quantified by one-step
rapid extractive methylation for gas chromatographic
analysis [25], as described previously [26,27]. First, cit-
rate plasma was spiked with heptadecanoic acid as in-
ternal standard. Next, free fatty acids were converted to
methyl esters by mixing with ethereal diazomethane. Lastly,
the ethereal layer was dried, redissolved in chloroform, and
transferred to the gas chromatograph. The fatty acid me-
thyl esters were detected by use of a flame ionization de-
tector, and peak area integration was performed. Authors’ contributions Study conception and design: YL, XYW and JSL. Manuscript draft and data
analysis: YL. Interpretation and critical review of the manuscript: XYW, NL and
JSL. All authors read and approved the final manuscript. Statistical analysis
l Data results are presented as mean ± standard error. Comparisons of various hemorrhagic shock parameters,
I-FABP, endotoxin, oxidative and inflammatory mole-
cules, blood, and histological score between the four
groups were performed by ANOVA test. A significance
level of .05 was used for all analyses. Transmission electron microscopy (TEM) Transmission electron microscopy (TEM)
Two-millimeter sections of colon were fixed for 2 h in
4% buffered glutaraldehyde. The sections were cut into
smaller pieces, then fixed by 1% OsO4, sequentially
dehydrated through graded alcohols, infiltrated through
Epon 812 and then embedded in resin. Thin sections
were cut and stained with uranyl acetate and lead citrate,
and examined with a Hitachi H-600 (Tokyo, Japan)
transmission electron microscope operated at 75 kV. Received: 14 June 2014 Accepted: 5 August 2014
Published: 8 September 2014 Received: 14 June 2014 Accepted: 5 August 2014
Published: 8 September 2014 Acknowledgments
W
f l
i We are grateful to investigators and staff at laboratory of department of
general surgery in Jinling hospital for facilitating this study. We are grateful to investigators and staff at laboratory of department of
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and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit Submit your next manuscript to BioMed Central
and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit Submit your next manuscript to BioMed Central
and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit Submit your next manuscript to BioMed Central
and take full advantage of: Submit your next manuscript to BioMed Central
and take full advantage of: Submit your next manuscript to BioMed Central
and take full advantage of: 29. Laser scanning confocal microscopy (LSCM) Laser scanning confocal microscopy (LSCM)
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Cite this article as: Li et al.: The study of n-3PUFAs protecting the intestinal
barrier in rat HS/R model. Lipids in Health and Disease 2014 13:146.
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https://openalex.org/W2077123976
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http://rcin.org.pl/Content/55281/PDF/WA058_50657_P256-T17_Frag-Faun-Nr-21.pdf
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Ichtiofauna dorzecza górnego i środkowego Sanu
|
Fragmenta Faunistica
| 1,971
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cc-by
| 14,425
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[Z 2 rysunkam i i 4 tabelam i] [Z 2 rysunkam i i 4 tabelam i] W niniejszej pracy omawiam ryby stwierdzone w dorzeczu górnego i środkowego
biegu Sanu, analizę liczebności i rozmieszczenie poszczególnych gatunków, skład ga
tunkowy zespołów ichtiocenologicznych najbardziej charakterystycznych dla górnego
i środkowego biegu Sanu i jego dopływów oraz charakterystykę ekologiczną tych zes
połów. Dane dotyczące charakterystyki morfologicznej, biologicznej i stanowiska sys
tematycznego poszczególnych gatunków są częściowo już opublikowane ( R o l i k 1959,
1960, 1962, 1965a, 1965b, 1967a, 1970, 1971), inne ukażą się w najbliższym czasie. Wykaz gatunków z górnego i środkowego dorzecza Sanu podałam w referacie na IX
Zjeździe PTZool. w Lublinie, 22-25 września 1969 r. N ajstarsze dane bibliograficzne dotyczące w ystępow ania niektórych gatunków ryb w S anie pochodzą
od R z ą c z y ń s k ie g o (1721, za F e d o r o w ic z e m
1966). W X IX w. ichtiofaunę G alicji badali Z a w a d z k i
(1840) oraz H e c k e l i K n e r (1858). Spisy gatunków ry b , w k tó ry ch uw zględniono także dorzecze Sanu,
w raz z uw agam i dotyczącym i n iektórych gatunków opracow ali W a ł e c k i (1864) i N o w i c k i (1880a,
1880 b, 1882, 1889). K ilka publikacji, które ukazały się w ostatn im okresie (B o n t e m p s 1960, 1969, G ą -
s o w s k a 1960, M o r a w s k a 1964, P l is z k a 1951, 1953, S o l e w s k i 1962, 1964, S c h r a m m 1957, S z y p u ł a
1966, W a j d o w ic z 1966), pośw ięcono poszczególnym zagadnieniom dotyczącym ichtiofauny S anu, lecz
prace te nie odzw ierciedlają obecnego stanu ichtiofauny tej rzeki w całości. B adania w łasne zapoczątkow ałam w 1956 r., jed n ak w iększość m ateriału zebrałam w latach 1964-
-1968 podczas reg ularnych badań prow adzonych przy zastosow aniu agregatu prądotw órczego do p o
łow u ryb. B adania dorzecza górnego S anu ukończyłem w 1966 r., tj. przed pow staniem zbiornika zapo
row ego w Solinie, badania środkow ego biegu i jego dopływ ów w 1968 r. [Z 2 rysunkam i i 4 tabelam i] San został zbadany na obszarze
od Sm olnika (pow. U strzy k i D olne) do P rzem yśla (do daw nego jazu poniżej brow aru), na długości 220 km . O dcinek S anu długości 40 km położony m iędzy Soliną a L eskiem w yłączono z badań ze w zględu na opraco
w anie tego teren u przez W a jd o w ic z a (1966). Z badano także następujące dopływ y S anu: W ołosaty, N a-
siczniański, Solinka z W etlinką, H oczew ka, dolna O slaw a z K alniczką, Sanoczek, S tara R zeka, Szklary, F R A G M E N T A
F A U N I S T I C A
Tom XVII
Warszawa, 15 XI 1971
Nr 21 Nr 21 Halina R o l i k Halina R o l i k N ajstarsze dane bibliograficzne dotyczące w ystępow ania niektórych gatunków ryb w S anie pochodzą
od R z ą c z y ń s k ie g o (1721, za F e d o r o w ic z e m
1966).
W X IX w. ichtiofaunę G alicji badali Z a w a d z k i
(1840) oraz H e c k e l i K n e r (1858). Spisy gatunków ry b , w k tó ry ch uw zględniono także dorzecze Sanu,
w raz z uw agam i dotyczącym i n iektórych gatunków opracow ali W a ł e c k i (1864) i N o w i c k i (1880a,
1880 b, 1882, 1889). K ilka publikacji, które ukazały się w ostatn im okresie (B o n t e m p s 1960, 1969, G ą -
s o w s k a 1960, M o r a w s k a 1964, P l is z k a 1951, 1953, S o l e w s k i 1962, 1964, S c h r a m m 1957, S z y p u ł a
1966, W a j d o w ic z 1966), pośw ięcono poszczególnym zagadnieniom dotyczącym ichtiofauny S anu, lecz
prace te nie odzw ierciedlają obecnego stanu ichtiofauny tej rzeki w całości.
B adania w łasne zapoczątkow ałam w 1956 r., jed n ak w iększość m ateriału zebrałam w latach 1964-
-1968 podczas reg ularnych badań prow adzonych przy zastosow aniu agregatu prądotw órczego do p o
łow u ryb. B adania dorzecza górnego S anu ukończyłem w 1966 r., tj. przed pow staniem zbiornika zapo
row ego w Solinie, badania środkow ego biegu i jego dopływ ów w 1968 r. San został zbadany na obszarze
od Sm olnika (pow. U strzy k i D olne) do P rzem yśla (do daw nego jazu poniżej brow aru), na długości 220 km .
O dcinek S anu długości 40 km położony m iędzy Soliną a L eskiem w yłączono z badań ze w zględu na opraco
w anie tego teren u przez W a jd o w ic z a (1966). Z badano także następujące dopływ y S anu: W ołosaty, N a-
siczniański, Solinka z W etlinką, H oczew ka, dolna O slaw a z K alniczką, Sanoczek, S tara R zeka, Szklary, Rys. 1. Stanow iska połowów. San: 1 — Sm olnik, 2 — Sękowiec, 3 — Rajskie, 4 — Solina, 5 — P osto-
łów, 6 — Ł ukaw ica, 7 — Zasław , 8 — Sanok, 9 — T repcza, 10 — M rzygłód, 11 — T em eszów I, 12 —
T em eszów II, 13 — O barzym , 14 — D ynów , 15 — D ubiecko I, 16 — D ubiecko II, 17 — Iskań, 18 —
Babice. D opływ y S anu: 19 — W ołosaty, Pszczeliny, 20 — T erebow iec, U strzyki G órne, 21 — N asiczniań-
ski, B erehy G ó rn e, 22 — N asiczniański, D w ernik, 23 — Solinka, Ż ubracze, 24 — Solinka, Buk, 25 — So-
linka, W ołkow yja, 26 — Solinka, S olina, 27 — W etlinka, W etlina, 28 — W etlinka, Jaw orzec, 29 —
H oczew ka, Baligród, 30 — H oczew ka, N ow osiółki, 31 — H oczew ka, H oczew , 32 — Osława, D u szaty n ,
33 — O sław a, P rełuki, 34 -— Osława, Zagórz I, 35 — Osława, Zagórz II, 36 — K alniczka, T arnaw a G ó rn a, http://rcin.org.pl H. Rolik 560 D rohobyczka, górny W isłok do H aczow a, C zarny z G łuchym , O lszanica, D yrbek, S tupnica,
W iar,
a także starorzecza S anu w okolicy Przem yśla. Łącznie przeprow adzono 63 połow y w 57 stanow iskach
(56 w Sanie i dopływ ach, jed en w starorzeczu w H urko koło M edyki) (rys. 1). W niektórych stanow iskach CSRS http://rcin.org.pl http://rcin.org.pl 561 Ichtiofauna dorzecza Sanu połow y pow tórzono w kolejnych latach bądź tego sam ego roku. Połow ów dokonyw ano agregatem prądu
stałego (typ M L O V , p rodukcji Ć S S R ) o napięciu do 300V i n atężeniu 2-3A . D ługość odław ianego o d
cinka w ynosiła 250-350 m . Połow y agregatem prąd o tw ó rczy m przeprow adzono w Sanie po Babice (pow. P rzem yśl). M ateriały z K rasiczyna koło P rzem yśla i z P rzem yśla pochodzą w yłącznie z połow ów siecio
w ych lu b w ędkow ych. połow y pow tórzono w kolejnych latach bądź tego sam ego roku. Połow ów dokonyw ano agregatem prądu
stałego (typ M L O V , p rodukcji Ć S S R ) o napięciu do 300V i n atężeniu 2-3A . D ługość odław ianego o d
cinka w ynosiła 250-350 m . Połow y agregatem prąd o tw ó rczy m przeprow adzono w Sanie po Babice (pow. P rzem yśl). M ateriały z K rasiczyna koło P rzem yśla i z P rzem yśla pochodzą w yłącznie z połow ów siecio
w ych lu b w ędkow ych. Ł ącznie zebrano lub zidentyfikow ano 8500 okazów . Z tej liczby 1200 okazów pochodzi z połowów
sieciow ych. O kazów tych nie w łączono do analizy liczebności. Z różnych pow odów nie w ykorzystano
do obliczenia liczebności także m ateriałów z n astęp u jący ch stanow isk: 10, 16, 26, 32, 33, 36, 45 (nie p o
liczono w szystkich okazów podczas połow u z p o w odu złych w arunków hydrologicznych lub innych
przyczyn technicznych), 5, 35, 51 (pow tórzenie połow ów ), 47 (zm ienione środow isko). N ie analizow ano
liczebności pró b y ze starorzecza w H urko, gdyż składała się ona z pojedynczych okazów. 37 — Saneczek, Sanok, 38 — S tara R zeka, N iebocko, 39 — Baryczka, N ozdrzec, 40 — Szklary, Bachórz,
41 — D rohobyczka, D ubiecko, 42 — W isłok, Sieniaw a, 43 — W isłok, Besko, 44 — W isłok, Haczów,
45 — M oraw a, H aczów , 46 — C zarny, P olana, 47 — C zarn y , C hrew t, 48 — G łuchy, Polana, 49 — O lsza
nica, O lszanica, 50 — D yrbek, L ukaw ica, 51 — S tupnica, Ł azy Bachów skie, 52 — S tupnica, Bachów,
53 — W iar, W ojtkow a, 54 — W iar, Posada R ybotycka, 55 — W iar, K alw aria Pacław ska, 56 — W iar,
K rów niki, 57 — starorzecze S anu w H urko kolo M edyki. U w a g a : przy w ym ienianiu dopływ ów S anu na pierw szym m iejscu podano nazw ę potoku, na
d ru g im — m iejscow ość. http://rcin.org.pl D o badań nad
liczebnością w ykorzystano w yłącznie p ró b y zbierane w analogicznych okresach, tj. od końca m aja do
połow y czerw ca w latach 1966-1968. Zebrane materiały odzwierciedlają w dużej mierze stan pierwotny ichtiofauny dorze
cza Sanu ze względu na to, że w badanych wodach nie występowały stałe źródła zanie
czyszczenia rzeki, koryto Sanu uregulowane jest w nieznacznym stopniu, badane odcinki
dopływów nie są objęte regulacją w ogóle. Wpływ zapory wodnej w badanych wo
dach do 1968 r. był nieznaczny. W środkowym biegu Sanu przejawiał się on w postaci
regularnych wahań poziomu wody, doprowadzających do regularnego wynurzania
się z pod wody całych połaci dna rzeki oraz stałej obecności w wodzie mulistych zawie
sin. Czynniki te z pewnością wywierają ujemny wpływ na warunki rozrodcze gatunków
litofilnych oraz skład pogłowia poszczególnych gatunków w środkowym biegu Sanu. Zagadnienia te wymagają jednak specjalnych badań. Szczególnie obszar dorzecza gór
nego Sanu, ze względu na słabe zaludnienie i zagospodarowanie oraz naturalny cha
rakter cieków, przedstawiał doskonałą możliwość do przestudiowania naturalnych
zespołów ichtiocenologicznych, charakterystycznych dla tego obszaru Karpat. Zestawiając materiały własne, dotyczące występowania poszczególnych gatunków
w dorzeczu Sanu, uwzględniłam następujący podział dorzecza Sanu: bieg górny Sanu
po Rajskie, bieg środkowy po Przemyśl, dopływy biegu górnego i dopływy biegu środ
kowego. A nalizując w ystępow anie poszczególnych gatunków porów nuję sw oje obserw acje głów nie z danym i
N o w i c k i e g o (1880a i b, 1882, 1883, 1889), które m im o iż nie kom pletne zaw ierają bardzo szczegółowe
inform acje o rybach. N ie naw iązuję natom iast do skąpych danych zaw artych w pracy S oi.E W S K iE G O
(1964), ani do p racy W a j d o w i c z a (1966) om aw iającej zm iany w ichtiofaunie S anu w rejonie M yczkow iec. U w a g a : przy w ym ienianiu dopływ ów S anu na pierw szym m iejscu podano nazw ę potoku, na
d ru g im — m iejscow ość. http://rcin.org.pl C H A R A K T E R Y S T Y K A F IZ JO G R A F IC Z N A B A D A N E G O T E R E N U S an jest najw iększym i o statn im karpackim dopływ em W isły. W ypływ a na w ysokości 8 8 9 m ( C h l e
b o w s k i 1 8 8 9 ), w edług innych źródeł na w ysokości 843 m ( K l i m a s z e w s k i 1937) w okolicy Przełęczy
U żockiej. D ługość S an u w ynosi 444 km , z tego na bieg górski i podgórski przypada 279 km . Pow ierzchnia
zlew ni całego dorzecza obejm uje 16 732,1 km 2 ( M i k u l s k i 1963). Poniew aż badaniam i został objęty tylko
bieg górski i podgórski oraz dopływ y położone na tym obszarze, rozpatruję tylko ten teren. D olna granica g ó rn eg o biegu S anu określana je st dosyć różnie, najczęściej do ujścia O sław y. W o p ar
ciu o podział rzek karpackich na krain y rybne opracow any przez S t a r m a c h a (1956) jako górny bieg S anu
należy traktow ać tylko jego odcinek po Rajskie. Przem aw iają za tym nie tylko wysokości spadków jed n o
stkow ych i budow a koryta rzeki, lecz także w ystępow anie i liczebność określonych gatunków ryb. O d
R ajskiego pojaw ia się n p . Gobio kessleri, k tóry w okolicy Leska staje się ju ż liczny. S pada na tym o b
sz a r z e liczebność Phoxinus phoxinus, pojaw iają się pojedyncze okazy Perea fluviatilis. W górnym biegu S anu spadki jednostkow e w ynoszą od 1 6,1 do 3,4%o> a n a obszarze zbadanym ,
tj. od Sm olnika do R ajskiego — 3,9-3,4% o- K oryto rzeki szerokie, płytkie i skaliste. Bieg rzeki w edług
P a s t e r n a k a (1 9 6 4 ) zgodny je st z biegiem w arstw skalnych. T akie ukształtow anie się podłoża nie stw a r za
z b y t dogodnych w arunków do bytow ania ryb, gdyż brak tu głęboczków i kryjówek. C H A R A K T E R Y S T Y K A F IZ JO G R A F IC Z N A B A D A N E G O T E R E N U O d Rajskiego po P rzem yśl (z w yjątkiem jezior zaporow ych) rozpościera się obszar podgórski Sanu,
charakteryzujący się obecnością kolejno następujących po sobie odcinków przełom ow ych o zw ężonym
korycie rzeki, dużym spadku i dnie kam ienisto-skalistym oraz położonym i m iędzy nim i kilku- lub kilku
nastokilom etrow ym i odcinkam i o szerokim korycie, w odzie płynącej wolniej i dnie kam ienisto-żw iro-
w atym , które K l i m a s z e w s k i (1937) określa jako nieckow ate. Przeciętne spadki w biegu środkow ym w y
noszą 2,2-0 ,5 °/00. W środkow ym biegu Sanu bieg rzeki w edług K l i m a s z e w s k i e g o (1937) zasadniczo
nie je st zgodny z biegiem w arstw skalnych. M iejsc piaszczystych brak zarów no w górnym jak i śro d
kow ym biegu. M iejsc zam ulonych, z w yjątkiem basenów pow stałych w niektórych m iejscach w środko
w ym biegu w skutek regulacji rzeki (,,staw iki” ), w w arunkach naturalnych praw ie się nie spotyka. W szystkie zbadane dopływ y m ożna określić jako górskie i podgórskie. W szystkie charakteryzują
się w ysokim i spadkam i jednostkow ym i, w ynoszącym i 2 9 ,l-5 ,3 °/00. D no skaliste lub kam ienisto-żw iro-
w ate. M iejsc piaszczystych brak. Z am ulone odnogi spotyka się tylko w rozlew iskach W iaru w K alw arii
Pacławskiej i w O sław ię w Z agórzu. http://rcin.org.pl 562 A N A L IZ A C E N O L O G IC Z N A L iczebność gatunków w zespołach ichtiocenologicznych z uw zględnieniem poszczególnych stanow isk w procentach: I - I I — część górna d o
yw ów górnego biegu S anu, I I I - I V — część środkow a dopływ ów górnego biegu S anu, V — część dolna dopływ ów górnego biegu S anu, V I — dopły
y środkow ego biegu S anu, V II — dopływ y dolnego biegu S anu, A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e,
C — odcinki nieckow ate). h
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C ołius poecilopus
Salmo brutta m. fario
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Nemachilus barbatulus
Barbus petenyi
Coitus go bio
Leuciscus cephalus
Leuciscus leuciscus
Gobio gobio
Alburnus alburnus
Album oides bipunctatus
Barbus barbus
Barbus cyclolepis waleckii
Chondrostoma nasus
Gobio kessleri
Rhodeus sericeus amarus
Rutilus rut Hus
Carassius auratus gibelio
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Rvs. 2. L iczebność gatunków w zespołach ichtiocenologicznych z uw zględnieniem poszczególnych stanow isk w procentach: I - I I — część górna d o
pływ ów górnego biegu S anu, I I I - I V — część środkow a dopływ ów górnego biegu S anu, V — część dolna dopływ ów górnego biegu S anu, V I — dopły
wy środkow ego biegu S anu, V II — dopływ y dolnego biegu S anu, A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e,
C — odcinki nieckow ate). http://rcin org pl Rvs. 2. A N A L IZ A C E N O L O G IC Z N A Badania nad składem gatunkowym ichtiofauny i liczebnością poszczególnych
gatunków wykazały znaczne zróżnicowanie jakościowe i ilościowe w poszczególnych
odcinkach Sanu i większych jego dopływów, różnice między dopływami górnymi
i dopływami biegu środkowego, jak również między dopływami a górnym i środkowym,
biegiem Sanu. Strefowość w występowaniu prawie wszystkich gatunków, a także zes
połów gatunków zaznaczyła się bardzo wyraźnie. Strefy występowania poszczególnych
zespołów ustalam na podstawie spadków jednostkowych i dominacji poszczególnych
gatunków, podobnie jak to uczynił B
a
c
k
i e l (1964) dla Drwęcy i H
o
l C i' k (1966a i b)
w badaniach nad ichtiofauną Orawy i Hronu. Analizowane niżej dane, mimo szeregu niedociągnięć (jak np. różna efektywność
połowu w poszczególnych ciekach badanego obszaru), pozwalają zorientować się w ogól
nych prawidłowościach kształtowania się zespołów cenologicznych w poszczególnych
strefach i w stosunkach cenologicznych omawianych gatunków (tabela 1, rys. 2). http://rcin.org.pl http://rcin.org.pl ic h tio c e n o lo g ic z n y
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C ołius poecilopus
Salmo brutta m. fario
Phoxinus phoxinus
Nemachilus barbatulus
Barbus petenyi
Coitus go bio
Leuciscus cephalus
Leuciscus leuciscus
Gobio gobio
Alburnus alburnus
Album oides bipunctatus
Barbus barbus
Barbus cyclolepis waleckii
Chondrostoma nasus
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vs. 2. A N A L IZ A C E N O L O G IC Z N A L iczebność gatunków w zespołach ichtiocenologicznych z uw zględnieniem poszczególnych stanow isk w procentach: I - I I — część górna d o
pływ ów górnego biegu S anu, I I I - I V — część środkow a dopływ ów górnego biegu S anu, V — część dolna dopływ ów górnego biegu S anu, V I — dopły
wy środkow ego biegu S anu, V II — dopływ y dolnego biegu S anu, A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e,
C — odcinki nieckow ate). Rvs. 2. L iczebność gatunków w zespołach ichtiocenologicznych z uw zględnieniem poszczególnych stanow isk w procentach: I - I I — część górna d o
pływ ów górnego biegu S anu, I I I - I V — część środkow a dopływ ów górnego biegu S anu, V — część dolna dopływ ów górnego biegu S anu, V I — dopły
wy środkow ego biegu S anu, V II — dopływ y dolnego biegu S anu, A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e,
C — odcinki nieckow ate). http://rcin.org.pl H. Rolik 564 Tabela 1. Liczebność gatunków w poszczególnych zespołach ichtiocenologicznych w dorzeczu Sanu:
A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e, C — nieckow ate). Z espół ichtiocenolo-
giczny i liczeb-
ność
G atunek
I
[ i i
III
IV
0//o
n
°//o
n
0//o
n
°//o
n
Cottus poecilopus
60,00
39
21,29
145
5,88
98
Salmo trutta m . fario
29,23
19
12,33
84
5,52
92
1,63
12
Phoxinus phoxinus
10,77
7
46,85
319
22,76
379
19,64
145
Nemachilus barbatulus
14,11
96
10,38
173
13,96
103
Barbus petenyi
1,62
11
22,04
367
23,98
177
Cottus gobio
1,32
9
5,04
84
2,03
15
Leuciscus cephalus
0,73
5
8,94
149
10,30
76
Leuciscus leuciscus
1,17
8
5,88
98
1,90
14
Gobio gobio
0,29
2
2,10
35
3,79
28
Alburnus alburnus
0,29
2
5,88
98
1,36
10
Alburnoides bipunctatus
4,98
83
12,06
89
Barbus barbus
0,30
5
3,79
28
Barbus cyclolepis waleckii
0,30
5
1,22
9
Chondrostoma nasus
4,20
31
Gobio kessleri
0,14
1
Rhodeus sericeus arnarus
Rutilus rutilus
Carassius auratus gibelio
Acerina cernua
Tinea tinca
Perea fluviatilis
Blicca bjoerkna
Lota lota Tabela 1. A N A L IZ A C E N O L O G IC Z N A Liczebność gatunków w poszczególnych zespołach ichtiocenologicznych w dorzeczu Sanu:
A — górny bieg Sanu, B i C — środkow y bieg S anu (B — odcinki przełom ow e, C — nieckow ate). Dla górnych dopływów Sanu, do których zaliczyłam na podstawie ich charakte
rystyki hydrologicznej wszystkie dopływy Sanu w Bieszczadach oraz Osławę, górny
Wisłok po Haczów i Wiar po Kalwarię' Pacławską, można wyodrębnić kilka chara
kterystycznych zespołów ichtiocenologicznych, które nazwałam umownie zespołami
I, II, III, IV i V. Strefy występowania tych zespołów odpowiadają: I i II — górnym
biegom dopływów, III i częściowo IV — biegom środkowym i V — dolnym biegom. W zespołach wyodrębniłam oprócz gatunków dominujących gatunki towarzyszące oraz
uzupełniające. Ponieważ zarówno dopływy Sanu jak i poszczególne stanowiska w obrę
bie biegu górnego i środkowego cechuje podobny charakter pod względem spadków
i budowy geomorfologicznej, a co za tym idzie bardzo zbliżony skład ichtiofauny,
można było analizować poszczególne stanowiska łącznie w ramach odnośnych zespołów. Omawiając zespoły podaję w nawiasach badane stanowiska. Z e s p ó ł I (stan. 20, 21) obejmuje 2 lub 3 gatunki, występujące w górskich poto
kach. Są to: C. poecilopus, S’, trutta m. fario i Ph. phoxinus. A N A L IZ A C E N O L O G IC Z N A Pierwszy z nich jestgatun- http://rcin.org.pl Ichtiofauna dorzecza Sanu 565 I-V — dopływ y biegu górnego, V I — dopływ y biegu środkow ego, V II — dopływ y biegu dolnego,
V
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0,16
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27
0,85
6
241
5,60
35
4,59
20
17,63
125
0,76
3
1033
8,16
5 i
13,76
60
9,17
65
2,30
9
3,42
8
565
16,16
101
3,44
15
7,53
7
36,38
258
5,87
23
4,28
10
969
1,76
11
1,15
5
2,82
20
3,32
13
157
10,40
65
33,48
146
21,50
20
7,05
50
4,59
18
32,50
76
605
0,16
1
9,40
41
0,42
3
1,02
4
1,71
4
173
10,08
63
18,81
82
13,97
13
7,62
54
4,85
19
19,68
46
342
0,32
2
1,15
5
3,23
3
5,50
39
6,89
27
24,38
57
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11,36
71
5,73
25
12,90
12
2,54
18
3,32
13
2,99
7
318
14,88
93
0,92
4
4,30
4
4,80
34
20,15
79
1,71
4
251
9,76
61
0,23
1
2,15
2
2,11
15
1,28
5
98
8,80
55
0,92
4
33,33
31
2,82
20
40,81
160
6,00
14
315
0,96
6
0,14
1
4,59
18
2,14
5
31
0,80
5
5
0,16
1
0,26
1
0,43
1
3
0,16
1
0,23
1
2
0,16
1
1
0,16
1
1
0,14
1
0,43
1
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! 0,43
1
1 I-V — dopływ y biegu górnego, V I — dopływ y biegu środkow ego, V II — dopływ y biegu dolnego, I-V — dopływ y biegu górnego, V I — dopływ y biegu środkow ego, V II — dopływ y kiem dominującym, drugi towarzyszącym. Ph. phoxinus, który można uważać za gatu
nek uzupełniający, występował tylko w stan. 20. Wysokość spadków jednostkowych
w strefie zespołu I wynosi do 30°/00. kiem dominującym, drugi towarzyszącym. Ph. phoxinus, który można uważać za gatu
nek uzupełniający, występował tylko w stan. 20. Wysokość spadków jednostkowych
w strefie zespołu I wynosi do 30°/00. Z e s p ó ł II (stan. 22, 23, 27, 48, 53) liczący 10 gatunków można określić jako przej
ściowy od zespołu I do zespołu III. Skład gatunkowy powiększa się o następujące ga
tunki: N. barbatulus, C. gobio, B. petenyi, L. leuciscus, L. cephalus, G. gobio, A. a lb u r-
nus. A N A L IZ A C E N O L O G IC Z N A Gatunkiem dominującym staje się Ph. phoxinus. Gatunki towarzyszące to: C. poecilopus, N. barbatulus, S. trutta m. fario i C. gobio. B. petenyi, L. leuciscus, L. ce
phalus, G. gobio, A. alburnus, które są gatunkami uzupełniającymi, spotyka się poje
dynczo i nie we wszystkich stanowiskach. Liczebność C. poecilopus w tym zespole
znacznie się zmniejsza w porównaniu z zespołem I. Wysokość spadków jednostkowych
w strefie zespołu II wynosi około 16°/^. Z e s p ó ł III (stan. 19, 24, 28, 29, 42, 46). Liczba gatunków zwiększa się do 13-
Gatunkami dominującymi są Ph. phoxinus lub B. petenyi. Towarzyszącymi: N. bar- http://rcin.org.pl H. Rolik 566 batulus, L. cephalus, S. trutta m. fario, C. poecilopus, C. gobio, A. alburnus, L. leuci
scus, A bipunctatus, G. gobio. Gatunki uzupełniające to B. barbus i B. cyclolepis
waleckii, spotykane sporadycznie. Spadki jednostkowe w strefie zespołu III wynoszą
8,4-5,4 °/00. Z e s p ó ł IV (stan. 25, 30, 54) można określić jako przejściowy od zespołu III
do zespołu V. Spotyka się tu łącznie 14 gatunków. Gatunkiem dominującym jest B. petenyi. Gatunki towarzyszące to: Ph. phoxinus, N. barbatulus, A. bipunctatus, L. cephalus, G. gobio, CA. nasus, B. barbus, C. gobio, L. leuciscus, ć>. trutta m. /ano, A
alburnus, B. cyclolepis waleckii. Jako gatunek uzupełniający sporadycznie spotyka
się G. kessleri. Ph. phoxinus i N. barbatulus występują w tym zespole nadal licznie,
lecz brak już C. poecilopus, a liczebność S’, rrwrfa m. fario spada do pojedynczych oka
zów. Spadki jednostkowe w strefie zespołu IV wynoszą 8,6-6,3°/00. Mimo że wysokość
spadków jednostkowych jest podobna jak w strefie zespołu III jednak koryto rzeki
staje się szersze i głębsze, co powoduje zmniejszenie się liczebności Ph. phoxinus, zwię
kszenie się liczebności L. cephalus i B. barbus oraz pojawienie się Ch. nasus. Z e s p ó ł V (stan. 31, 34, 43, 44) liczy do 19 gatunków (wraz z nie stwierdzonym
w połowach agregatem, lecz występującym tu P. jluviatilis — 20 gatunków). Gatunkami
dominującymi są B. petenyi lub B. barbus. Towarzyszącymi: L. cephalus, A. bipuncta
tus, G. gobio, Ch. nasus, B. cyclolepis waleckii, N. barbatulus, PA. phoxinus, C. gobio. Gatunkami uzupełniającymi są G. kessleri, L. leuciscus, A alburnus i <S. trutta m. fario,
występujący sporadycznie i pojedynczo. Przy obecności odpowiednich mikro-nisz
ekologicznych sporadycznie występują także P. rutilus, PA. sericeus amarus, C. awra-
tas gibelio, P. tinea, P. jluviatilis, A cernua (stan. 34 i 35). Liczebność PA. phoxinus
w tym zespole znacznie spada, wzrasta liczebność CA. i P. cyclolepis waleckii. Spadki jednostkowe w strefie zespołu V wynoszą 7,4-2,5%0. Nieco inny skład gatunkowy, a przede wszystkim inny wzajemny stosunek procen
towy poszczególnych gatunków obserwowano w dopływach biegu środkowego, do
których zaliczono następujące zbadane cieki: Sanoczek, Stara Rzeczka, Baryczka,
Szklary, Drohobyczka, Olszanica, Dyrbek, Stupnica. Dopływy te, w odróżnieniu
od dopływów górnych, są niewielkie, w większości przypadków są to kilkumetrowej
szerokości potoki. Pobierano z nich zazwyczaj po jednej próbie. Zespół ichtioceno-
logiczny charakterystyczny dla tych dopływów wyodrębniłam jako zespół VI. Z e s p ó ł VI (stan. http://rcin.org.pl 39, 40,41, 50, 52) liczy łącznie 14 gatunków ryb. W odróżnieniu
od poszczególnych zespołów dopływów biegu górnego gatunkiem dominującym był tu
zawsze L. cephalus. Towarzyszącymi: G. gobio, N. barbatulus, L. leuciscus, A. bipunctatus,
S. trutta m. fario, B. petenyi, PA. phoxinus, C. Nielicznie i sporadycznie wystę
powały B. cyclolepis waleckii, CA. nasus, A. alburnus, C. auratus gibelio i C. carassius;
gatunki te należy uznać za uzupełniające. Do tego zespołu powinne być zaliczone także
stanowiska 37, 38, 49, lecz nietypowy skład jakościowy i ilościowy gatunków ryb w tych
stanowiskach nasuwa przypuszczenie, że jest on wynikiem zmian związanych z inge http://rcin.org.pl Ichtiofauna dorzecza Sanu 567 rencją ludzką. W Olszanicy (stan. 49) dominował L. leuciscus, co stanowiło jedyny tego
rodzaju przypadek podczas badań prowadzonych w dorzeczu Sanu. W Sanoczku
(stan. 37) gatunkiem dominującym był Ch. nasus. Stara Rzeka (stan. 38) miała bardzo
ubogi skład ichtiofauny. Ten dopływ jest częściowo regulowany i prawdopodobnie za
nieczyszczony. y
y
Dolny bieg Wiaru (stan. 56) różni się pod względem jakościowym i ilościowym
składu ichtiofauny od wszystkich innych omówionych dopływów. Wiar wpada do Sanu
poniżej Przemyśla i powinien być prawdopodobnie w swojej dolnej części zaliczony do
dopływów biegu dolnego. Gatunkiem dominującym jest tu Ch. nasus, towarzyszącymi
L. cephalus, G. gobio, A. bipunctatus, B. petenyi, B. barbus, A. alburnus, B. cyclolepis
waleckii. Pojedynczo występuje B. bjoerkna. Nie odnotowałam natomiast L. leuciscus,
Ph. phoxinus, N. barbatulus, tj. gatunków występujących licznie w dopływach biegu
górnego i biegu środkowego. g
g
g
g
Przy analizie ichtiofauny Sanu można było stwierdzić różnice jakościowe i ilościowe
w składzie ichtiofauny między biegiem górnym (zespół A) i biegiem środkowym
a w obrębie tego ostatniego między odcinkami przełomowymi (zespół B) i nieckowatvmi
(zespół C). Z e s p ó ł A (stan. 1, 2, 3). W górnym biegu Sanu stwierdziłam 15 gatunków ryb;
dominującym jest B. petenyi, towarzyszącymi: Ph. phoxinus, N. birbatulus, G. gobio,
L. cephalus, A. alburnus, B. barbus, C. gobio, Ch. nasus, A. bipunctatus, B. cyclolepis
waleckii, L. leuciscus. Jako gatunki uzupełniające nielicznie występują S. trutta m. fario, a w dolnej części także G. kessleri i P. fluviatilis. Spadki jednostkowe w strefie
zespołu A wynoszą od 3,9 do 3,4°/00. Z e s p ó ł A (stan. 1, 2, 3). W górnym biegu Sanu stwierdziłam 15 gatunków ryb;
dominującym jest B. petenyi, towarzyszącymi: Ph. phoxinus, N. birbatulus, G. gobio,
L. cephalus, A. alburnus, B. barbus, C. gobio, Ch. nasus, A. bipunctatus, B. cyclolepis
waleckii, L. leuciscus. Jako gatunki uzupełniające nielicznie występują S. trutta m. fario, a w dolnej części także G. kessleri i P. fluviatilis. Spadki jednostkowe w strefie
zespołu A wynoszą od 3,9 do 3,4°/00. Z e s p ó ł B (stan. 6, 7, 8, 9,11, 18). W środkowym biegu Sanu, w odcinkach prze
łomowych i na przemiałach wyniki połowów były dosyć niskie, ponieważ znaczną
część ryb znosi szybki prąd wody nim zostaną one policzone, a czasem nawet zidenty
fikowane. W połowach agregatem elektrycznym odnotowałam 14 gatunków. Gatunkiem
dominującym jest Ch. nasus, towarzyszącymi: B. barbus, B. petenyi, G. gobio, G. http://rcin.org.pl kes
sleri, A. alburnus, A. bipunctatus, L. cephalus, B. cyclolepis waleckii, C. gobio, N. bar-
batulus, L. leuciscus. Jako gatunek uzupełniający w stanowiskach położonych wyżej wystę
puje Ph. phoxinus, w stanowiskach położonych niżej — R. rutilus. Z e s p ó ł C (stan. 4, 12, 13, 14, 15, 17). W odcinkach nieckowatych wyniki po
łowów były nikłe. Na otwartej przestrzeni, pozbawionej kryjówek, przy bardziej spo
kojnym prądzie spłoszone ryby trzymają się poza zasięgiem działania pola elektrycznego. W połowach agregatem wystąpiło 13 gatunków. Można było zaobserwować, że w zes
pole tym gatunkiem dominującym jest L. cephalus. Jako gatunki towarzyszące na
pierwszym miejscu stoją A. alburnus i G. gobio, następnie Ch. nasus, A. bipunctatus,
G. kessleri, B. petenyi i L. leuciscus. Jako uzupełniające: B. barbus, R. rutilus, L. lota,
P. fluviatilis, N. barbatulus. Z materiałów uzyskanych drogą połowów siecią wynika,
że zespół ten w dolnym odcinku biegu środkowego wzbogaca się o szereg innych ga Salmo trutta L in n a e u s m. fario L in n a e u s — pstrąg Najliczniej występuje przy spadkach jednostkowych 30-14°/00 w górnych odcinkach
dopływów Sanu w Bieszczadach. Stanowi tam od 44,7 do 7,4% złowionych ryb, znaj
dując się na II-IV miejscu w połowach. W środkowych odcinkach dopływów stanowi
od 8,7 do 0,8% i zajmuje IV -X I miejsce. W dolnych odcinkach dopływów spotyka
się sporadycznie. Występuje w dopływach biegu środkowego, stanowiąc w Szklarach
(stan. 40) 25,4%, w Dyrbku (stan. 50) 4,5%, w Baryczce (stan. 39) 1,5%. W Sanie stwier
dziłam tylko pojedyncze okazy po Rajskie gdzie liczebność pstrąga wynosi mniej
niż 2,0%. http://rcin.org.pl H. Rolik 568 tunków, które nie wystąpiły w połowach agregatem. Są to: B. bjoerkna, A. brama, T-
tinea, A . aspius, F. vimba, /. nebulosus, S. glanis, F. lucius, A cernua. tunków, które nie wystąpiły w połowach agregatem. Są to: B. bjoerkna, A. brama, T-
tinea, A . aspius, F. vimba, /. nebulosus, S. glanis, F. lucius, A cernua. Ze starorzeczy Sanu w okolicy Przemyśla największe jest starorzecze w Hurko
koło Medyki, które jest stosunkowo mało zarośnięte i ma duże lustro otwartej wody. Z gatunków, których nie wykryłam w Sanie, występuje tam S’, erythrophthalmus i M. fossilis. Poza tym w starorzeczu żyje E. lucius, 7\ rmca, F. sericeus amarus, C. caras-
sius, P. fluviatilis. PRZEGLĄD GATUNKÓW Do opracowania tego rozdziału wykorzystałam wszystkie materiały, zarówno z połowów agregatem
elektrycznym, jak z połowów siecią i próbek narybkowych. Rutilus rutilus ( L in n a e u s) — płoć Nie wykryłam jej ani w górnym biegu Sanu ani w dopływach biegu górnego w Biesz
czadach. Sporadycznie i pojedynczo występuje w środkowym biegu Sanu, nieco licz
niejsza w głębszych i spokojniejszych wodach dolnego odcinka biegu środkowego. Z dopływów Sanu zasiedla pojedynczo tylko Olszanicę (stan. 49) i Wisłok (stan. 44). Esox lucius L in n a e u s — szczupak W połowach agregatem nie spotykany. Występuje w środkowym biegu Sanu,
głównie w głębszych i spokojniejszych partiach dolnego odcinka oraz w stawikach. Najwyżej stwierdzonym stanowiskiem tego gatunku przed spiętrzeniem wód Sanu
było Lesko. Występuje w starorzeczu w Hurko. Rutilus rutilus ( L in n a e u s) — płoć Leuciscus cephalus (L in n a eu s) — kleń Jeden z najliczniejszych i najczęściej spotykanych gatunków. W górnych odcinkach
dopływów górnego biegu Sanu występuje sporadycznie, bardziej liczny staje się w od
cinkach środkowych, stanowiąc od 5,2 do 25,8% i zajmując przeważnie IV-V pozycję
w połowach. Na tym samym poziomie utrzymuje się liczebność klenia w dolnych odcin
kach górnych dopływów. W dopływach biegu środkowego jest gatunkiem dominu
jącym, stanowiąc od 15,2 do 59,3%. W górnym biegu Sanu liczebność klenia utrzymu
je się na poziomie środkowych i dolnych odcinków górnych dopływów. W środkowym
biegu Sanu jest wyraźnie liczniejszy w odcinkach nieckowatych, niż na przełomach. W odcinkach nieckowatych jest gatunkiem dominującym, a liczebność jego wynosi
32,1-62,5%. W odcinkach przełomowych zajmuje przeważnie III pozycję po śwince
i brzanie, stanowiąc 1,6-16,0%. Lenciscus leuciscus (L in n a eu s) — jelec W dorzeczu Sanu jest wyraźnie liczniejszy w dopływach aniżeli w samym Sanie,
przy czym najobficiej występuje w środkowych odcinkach dopływów biegu górnego,
przy spadkach jednostkowych 8,6-5,4°/00. Liczebność jego waha się tam w granicach
0,5-8,4% i tylko w jednym stanowisku (24, Solinka — Buk) stanowił 14,2%. W do
pływach biegu środkowego — 7,4-12,8%, jedynie w Olszanicy (stan. 49) stanowił on
26,6% ; jednocześnie, jak już wspomniałam, skład gatunkowy oraz liczebność pozo
stałych gatunków były w Olszanicy nietypowe. W dolnych partiach dopływów gór
nych oraz w górnym i środkowym biegu Sanu jelec występuje pojedynczo. http://rcin.org.pl Ichtiofauna dorzecza Sanu 569 Lenciscus leuciscus (L in n a eu s) — jelec Phoxinus phoxinus ( L in n a e u s ) — strzebla potokowa Jeden z najliczniejszych i najczęściej spotykanych gatunków w dorzeczu górnego
Sanu. W najwyższych partiach potoków o dużym spadku, w których dominuje głowacz
pręgopłetwy, strzebla nie występuje lub spotyka się pojedynczo. Zasiedla natomiast
najmniejsze nawet potoczki oraz rowki o niższym spadku, w których brak wszelkich
innych ryb. Tym samym strzebla wykazuje znacznie większe zdolności przystosowaw
cze niż inne gatunki towarzyszące jej w dorzeczu górnego Sanu. Jest gatunkiem do
minującym w strefie II, jak również w niektórych stanowiskach strefy III (24 Solinka —
Buk; 29, Hoczewka — Baligród; 46, Czarny — Polana). Licznie występuje także w gór
nym biegu Sanu i w dopływach biegu środkowego. W środkowym biegu Sanu występuje
pojedynczo do Przemyśla. Tinea tinea (L in n a eu s) — lin W dorzeczu górnego i środkowego biegu Sanu występuje sporadycznie z powodu
braku odpowiednich nisz ekologicznych. Najwyżej stwierdzonym stanowiskiem tego
gatunku była Osława w Zagórzu (stan. 34). W środkowym biegu Sanu żyje w stawikach. Nieco częściej spotyka się w spokojniejszych wodach dolnego odcinka biegu środkowe
go. Występuje także w starorzeczach w Hurko i w Wy szaty cach. Chondrostoma nasus (L inn aeus) — św inka Jeden z najliczniejszych gatunków w dorzeczu Sanu. Liczniej występuje w środ
kowym biegu Sanu, gdzie na odcinkach przełomowych, poczynając od Sanoka w dół
rzeki, jest gatunkiem dominującym, stanowiąc od 33,3 do 78,1% wszystkich ryb ło
wionych na poszczególnych stanowiskach. W górnym biegu Sanu udział świnki w po
łowach wynosił tylko 1,2-3,5%. W dopływach Sanu także nie jest liczna. Zamieszkuje
tylko dolne i częściowo środkowe odcinki dopływów dużych, jak Hoczewka, Solinka,
Osława, Wisłok, Wiar, gdzie liczebność jej waha się w granicach 0,7-11,0% w partiach
środkowych i 1,4-23,6% w dolnych. W Sanoczku (stan. 37) była gatunkiem dominu
jącym, stanowiąc 24,5%. W innych dopływach biegu środkowego występowała poje
dynczo i tylko w niektórych (stan. 39, Baryczka; 52, Stupnica). Leucaspius delineatus (H eckel) — słonecznica W połowach agregatem nie była spotykana. Mam okazy słolecznicy tylko z Sanu
pod Przemyślem. Gobio gobio (L inn aeus) — kiełb Jest jednym z najczęściej i najliczniej spotykanych gatunków. Pojedyncze okazy
pojawiają się w strefie II górnych dopływów. Szczególnie liczny jest w dolnych odcin
kach górnych dopływów, gdzie stanowi od 7,3 do 23,3% i zajmuje IV -V I miejsce
w połowach; w dopływach biegu środkowego stanowi około 20% i zajmuje II -III
miejsce, w nieckowatych odcinkach środkowego biegu Sanu — od 6,3 do 18,9%. Aspius aspius (L in n a eu s) — b oleń Aspius aspius (L in n a eu s) — b oleń Aspius aspius (L in n a eu s) — b oleń W połowach agregatem nie spotykany. Mam okazy bolenia tylko z Sanu pod Kra
siczynem. Gatunek rzadki. Leucaspius delineatus (H eckel) — słonecznica Scardinius erythrophthalmus (L in n a e u s) — w zdręga Wzdręgę stwierdziłam tylko w starorzeczu w Hurko. http://rcin.org.pl http://rcin.org.pl H. Rolik 570 Gobio kessleri D ybow ski — kiełb Kesslera Gobio kessleri D ybow ski — kiełb Kesslera Górna granica występowania tego gatunku sięga w dorzeczu Sanu Rajskiego (stan. 3), Wołkowyi (stan. 25) i Beska (stan. 43). Z dopływów biegu górnego został stwier
dzony tylko w Solince i w Wisłoku. Zdecydowanie unika dopływów mniejszych. W dorzeczu górnego Sanu spotyka się pojedynczo. Od Leska do Przemyśla jest znacz
nie liczniejszy, niż to wynika z materiałów łowionych agregatem. Przyczyną tego jest
fakt, że rażony prądem rzadko wpada do kasarka, lecz zostaje znoszony przez prąd. Najliczniej przebywa w miejscach o szybkim prądzie, szczególnie na przemiałach. W dopływach biegu środkowego nie notowany. Barbus barbus (L inn aeus) — brzana Pojedyncze okazy brzany pojawiają się w strefie III. W strefie IV stanowi 3,5-8,2%
wszystkich ryb. Liczniejsza jest w dolnych partiach dopływów, osiągając 12,6-23,6%. W górnym biegu Sanu liczebność brzany jest wyższa niż liczebność świnki i wynosi
3,0-8,5% . W środkowym biegu Sanu znacznie liczniej występuje w odcinkach prze
łomowych, gdzie zajmuje II miejsce pod względem liczebności po śwince, a liczebność
jej wynosi 4,7-37,2%. Tylko w Łukawicy (stan. 6), tj. w górnej części biegu środkowego
brzana była gatunkiem dominującym. W odcinkach nieckowatych liczebność brzany
wynosi 1,5-6,3%. Z dopływów biegu środkowego występowała tylko w Stupnicy przy
ujściu do Sanu (stan. 52). Barbus petenyi H eckel — brzanka Należy do gatunków najczęściej i najliczniej spotykanych w dorzeczu Sanu. Szcze
gólnie liczna w dorzeczu górnego Sanu. W środkowych odcinkach dopływów górnych
a często i dolnych oraz w górnym biegu Sanu jest gatunkiem dominującym, a udział
jej w połowach stanowi od 18,4 do 39,8%. W dopływach biegu środkowego i w środko
wym biegu Sanu znacznie mniej liczna. W środkowym biegu Sanu występuje głównie
w odcinkach przełomowych. Pojedyncze okazy brzanki spotyka się aż do Przemyśla. Barbus cyclolepis waleckii R o lik — brzana karpack Ichtiofauna dorzecza Sanu Ichtiofauna dorzecza Sanu 571 Gobio kessleri D ybow ski — kiełb Kesslera Barbus cyclolepis waleckii R o lik — brzana karpacka Występuje w dorzeczu zarówno górnego jak i środkowego Sanu, lecz wszędzie
jest nieliczna. Pojawia się w środkowych partiach dopływów górnych, gdzie stanowi
mniej niż 1% wszystkich ryb w strefie III i około 2% w strefie IV. Nieco liczniej wy
stępuje w dolnych odcinkach dopływów górnych, gdzie stanowi od 3,3 do 4,6% , wy
jątkowo 9,7% (stan. 44, Wisłok — Haczów) i 12,9% (stan. 34, Osława — Zagórz). Ze
stanowisk górnego biegu Sanu najliczniej występowała w Rajskiem (stan. 3) — 3,5%. W środkowym biegu Sanu występuje pojedynczo tylko na przełomach, poniżej D u
biecka (stan. 16) nie notowana. Z dopływów biegu środkowego pojedyncze okazy zło http://rcin.org.pl Alburnus alburnus (L inn aeus) — ukleja Alburnus alburnus (L inn aeus) — ukleja Jest gatunkiem często i dosyć licznie spotykanym zarówno w dorzeczu górnego,
jak i środkowego biegu Sanu. Pojedyncze okazy pojawiają się w strefie II górnych
dopływów. W odcinkach środkowych udział uklei w połowach wynosi 0,5-15,3%. Ciekawe, że mniej licznie występowała ukleja w strefie dolnej tych dopływów a zwykle
w ogóle nie spotykano jej tam w połowach. W górnym biegu Sanu liczebność uklei
wynosiła 0,6-3,8% , a w Sękowcu (stan. 2) nawet 12,4%. Podobny poziom liczebności
tego gatunku notowano w materiałach zebranych w czerwcu 1958 r. Najliczniej żyje
w środkowym biegu Sanu w odcinkach nieckowatych (14,7-26,4%), gdzie w jednym
przypadku (stan. 12, San — Temeszów II) była gatunkiem dominującym, stanowiąc
80,0%. W dopływach biegu środkowego prawie nieobecna. Alburnoides bipunctatuś (B loch) — piekielnica Licznie występuje w środkowej i dolnej strefie dopływów górnych, przewyższając
liczebnością ukleję (0,34-27,6%). W górnym biegu Sanu liczebność piekielnicy była
prawie identyczna z liczebnością uklei w Smolniku (stan. 1); podobny stosunek ob
serwowano w 1958 r. W Rajskiem (stan. 3) liczniejsza była piekielnica. W środkowym
biegu Sanu spotyka się pojedynczo aż do Przemyśla, częściej występuje w odcinkach
przełomowych. W dopływach biegu środkowego piekielnica jest znacznie częstsza
i liczniejsza niż ukleja. Abramis brama (L in n a e u s) — leszcz Abramis brama (L in n a e u s) — leszcz W połowach agregatem nie stwierdzony. Występuje w tych samych miejscach
co krąp, lecz znacznie mniej
licznie. Vimba vimba (L in n a eu s) — certa Vimba vimba (L in n a eu s) — certa Vimba vimba (L in n a eu s) — certa Podczas badań przeprowadzonych w środkowym biegu Sanu w 1968 r. złowiłam
tylko jeden okaz certy koło Dubiecka (stan. 16). Jednoroczne okazy zebrałam pod Les
kiem 27 V 1956 i pod Sanokiem 9 VI 1956. Trudno powiedzieć czy brak certy w po http://rcin.org.pl http://rcin.org.pl H. Rolik 572 wiono w Baryczce (stan. 39) i w Stupnicy (stan. 52). Występuje także w dolnym biegu
Wiar u (stan. 56). wiono w Baryczce (stan. 39) i w Stupnicy (stan. 52). Występuje także w dolnym biegu
Wiar u (stan. 56). Blicca bjoerkna (L inn aeus) — krąp W połowach agregatem wystąpił tylko w Wiarze w Krównikach (stan. 56). Na pod
stawie materiałów łowionych siecią można stwierdzić, że krąpia spotyka się w głębszych
i spokojniejszych odcinkach dolnej części biegu środkowego. Abramis brama (L in n a e u s) — leszcz Carassius auratus gibelio (B locii) — karaś srebrzysty Jeden okaz tego gatunku złowiłam w Osławię pod Zagórzem (stan. 34) i jeden okaz
w Baryczce (stan. 41). Pelecus cultratus (L inn aeus) — ciosa W zbiorach mam jeden okaz złowiony na wędkę w Sanie koło Przemyśla. Jest to
prawdopodobnie w ogóle pierwsze stwierdzenie tego gatunku w karpackim dopływie
Wisły. Rhodeus sericeus amarus (B locii) — różanka W dorzeczu górnego Sanu stwierdziłam ją w Osławię pod Zagórzem (stan. 34). W środkowym biegu Sanu występuje sporadycznie, ze względu na brak miejsc mu-
listych. Liczna w rozlewiskach W iaru w Kalwarii Pacławskiej (stan. 55). Występuje
także w starorzeczu w Hurko. Noemacheilus barbatulus (L in n a eu s) — śliz Jest gatunkiem bardzo licznym w dorzeczu górnego Sanu. W strefie II stanowi
5,2-25,4% ogółu ryb. W odcinkach środkowych i dolnych jest również bardzo liczny
i występuje powszechnie. Jego udział w połowach w strefach III, IV i V jest podobny
i wynosi 0,8-21,8% . Bardzo liczny w górnym biegu Sanu (0,5-23,5%). W środkowym
biegu Sanu pojedyncze okazy spotykałam do Mrzygłodu (stan. 10). Z dopływów biegu
środkowego najliczniej występuje w Dyrbku (stan. 50) — 23,7%, w pozostałych dopły
wach
1,3-9,4%. Carassius carassius (L inn aeus) — karaś Dwa okazy karasia złowiłam w Olszanicy (stan. 49), gdzie jest on najprawdopo
dobniej uciekinierem z pobliskich stawów. Występuje w starorzeczach Sanu w Hurko
i w Wyszatycach. http://rcin.org.pl 573 Ichtiofauna dorzecza Sanu łowach w 1968 r. był przypadkowy, czy jest wynikiem zmian, jakie zaszły w wędrów
kach tego gatunku do dorzecza Sanu. Pelecus cultratus (L inn aeus) — ciosa Pelecus cultratus (L inn aeus) — ciosa Silurus glanis
L inn aeus
—
sum W połowach agregatem nie stwierdzony. Spotyka się w spokojniejszych i głębszych
wodach dolnego odcinka biegu środkowego. W połowach agregatem nie stwierdzony. Zajmuje w środkowym biegu Sanu po
dobne nisze ekologiczne co sum. Piskorza stwierdziłam tylko w starorzeczu w Hurko. Silurus glanis
L inn aeus
—
sum Misgurnus fossilis (L inn aeus) — piskorz Misgurnus fossilis (L inn aeus) — piskorz Lota lota
(L in n a e u s) —
m iętu s W połowach agregatem trafił się tylko jeden okaz w Sanie w Dubiecku (stan. 15)- Perea jiuviatilis L inn aeus — okoń Znaleziony pojedynczo w kilku stanowiskach dorzecza górnego Sanu: S an — Sę-
kowiec (stan. 2), Solina (stan. 4), Czarny — Chrewt (stan. 47), Osława — Prełuki
(stan. 33). Liczniejszy jest w dolnym odcinku środkowego biegu Sanu. Ze wszystkich
dopływów biegu środkowego stwierdzony tylko w Olszanicy (stan. 49). Występuje
także w starorzeczu w Hurko. Znaleziony pojedynczo w kilku stanowiskach dorzecza górnego Sanu: S an — Sę-
kowiec (stan. 2), Solina (stan. 4), Czarny — Chrewt (stan. 47), Osława — Prełuki
(stan. 33). Liczniejszy jest w dolnym odcinku środkowego biegu Sanu. Ze wszystkich
dopływów biegu środkowego stwierdzony tylko w Olszanicy (stan. 49). Występuje
także w starorzeczu w Hurko. http://rcin.org.pl H. Rolik 574 Cobitis (Sabanejewia) aurata (F i l i p p i ) — koza złotawa Cobitis (Sabanejewia) aurata (F i l i p p i ) — koza złotawa Dotychczas udało się wykryć ten gatunek tylko w Sanie pod Przemyślem (okazy
dorosłe i narybek). Na brak kozy złotawej w środkowym biegu Sanu wpływa prawdo
podobnie nie tylko kamieniste podłoże rzeki i brak miejsc piaszczystych. Koza zło
tawa nie jest w Sanie tak liczna, jak np. w dorzeczu Dniestru, w Strwiążu (R o lik 1967b),
gdzie mimo kamienistego podłoża rzeki i niewielkiej ilości piasku pod kamieniami
koza złotawa jest bardzo liczna (do 3-4 okazów na 1 m2). W dorzeczu Sanu, w stano
wiskach o podobnym charakterze co Strwiąż, kozy złotawej nie spotkałam. S olew ski
(1964) podaje dla górnego biegu Sanu C. taenia L in n a e u s. Tego gatunku nie wy
kryłam w Sanie w ogóle. Misgurnus fossilis (L inn aeus) — piskorz Cottus poecilopus H eckel — głow acz pręgopłetw y Cottus poecilopus H eckel — głow acz pręgopłetw y Rozpatruję oba gatunki głowaczy razem, ponieważ ich występowanie jest ściśle
ze sobą powiązane. Zasięg i charakter występowania obu gatunków na przykładzie
dorzecza Sanu wymaga szerszego omówienia, gdyż danych o tym w literaturze ichtio-
logicznej jest niewiele. Oba gatunki, szczególnie zaś głowacza pręgopłetwego, cechuje
bardzo wyraźna strefowość w występowaniu. Głowacz pręgopłetwy zasiedla górną
część górskich dopływów. W strefie I stanowi 55,3-66,7%. W strefie II także jest liczny
i stanowi ponad 25% wszystkich ryb. W strefie II pojawiają się pojedyncze okazy gło
wacza białopłetwego. W strefie III średnia liczebność obu gatunków jest podobna i wa
ha się w granicach od 1,4 do 15,7% dla głowacza pręgopłetwego i 0,4-12,3% dla gło
wacza białopłetwego. Charakterystyczny przy tym jest fakt, że w stanowiskach o wyższej
liczebności jednego gatunku drugi spotyka się pojedynczo i na odwrót. Dla przykładu
można przytoczyć następujące liczby złowionych okazów. Stanowiska:
19
24
28
29
46
W olosaty
Solinka
W etlinka
Hoczewka
Czarny
Pszczeliny
Buk
Jaworzec
Baligród
Polana
C. poecilopus
8
12
5
35
38
C. gobio
27
8
44
4
1 Stanowiska:
19
24
28
29
46
W olosaty
Solinka
W etlinka
Hoczewka
Czarny
Pszczeliny
Buk
Jaworzec
Baligród
Polana
8
12
5
35
38
27
8
44
4
1 W środkowej części dopływów, lecz w stanowiskach należących do strefy IV, gło
wacz pręgopłetwy nie występuje, a głowacz białopłetwy stanowi l,7-4,7% . Na tym
poziomie utrzymuje się liczebność tego gatunku w dolnych odcinkach dopływów gór
nych oraz w górnym i środkowym biegu Sanu po Dubiecko (stan. 16). Poniżej D u
biecka głowacz białopłetwy w połowach agregatem nie był stwierdzony. Pojedyncze mło
de okazy zbierałam pod Przemyślem po dużym wezbraniu Sanu. W środkowym biegu
Sanu głowacz białopłetwy występuje w odcinkach przełomowych. W dopływach biegu
środkowego występuje pojedynczo (i nie we wszystkich potokach) tylko głowacz bia
łopłetwy. Badania wykazały, że głowacz pręgopłetwy, aczkolwiek ma ograniczony
zasięg, występuje masowo. Natomiast głowacz białopłetwy, o zasięgu znacznie szer
szym, odznacza się niskim zagęszczeniem populacji. Wymienione wyżej stanowiska
19 i 28 o wysokiej liczebności tego gatunku należą do wyjątków. We wszystkich po http://rcin.org.pl 575 Ichtiofauna dorzecza Sanu Acerina cernua (L inn aeus) — jazgarz Acerina cernua (L inn aeus) — jazgarz W dorzeczu górnego Sanu spotkałem go tylko w Osławię w Zagórzu (stan. 35). Liczniejszy w dolnej części środkowego biegu Sanu. Cottus gobio L inn aeus — głow acz białopłetw y Anguilla anguilla (Linnaeus) — węgorz W połowach agregatem nie notowany. Według informacji uzyskanych od węłkarzy
występuje w środkowym i górnym biegu Sanu, przy czym dane odnośnie do wystę
powania węgorza w górnym biegu Sanu pochodzą sprzed 1958 r. ★ Liczba wszystkich gatunków ryb stwierdzonych przeze mnie w górnym i środkowym
biegu Sanu wraz z dopływami wynosi 36 (tabela 2). Z tej liczby Nowicki nie uwzględnił
9 gatunków, wymienił natomiast kilka nie znalezionych przeze mnie (jak Stizcstedion
lucioperca, Leuciscus idus i niektóre inne, właściwe dla biegu dolnego), podając dla Sanu
łącznie 31 gatunków. W górnym biegu Sanu oraz w jego górnych dopływach spośród stwierdzon/ch tu
22 gatunków ryb 11 występuje licznie; są to: Ph. phoxinus, B. petenyi, N. barbatulus,
L. cephalus, S. trutta m. fario, C. poecilopus, G. gobio, A. bipunctatus, A. alournus,
oraz nieco mniej licznie C. gobio i L. leuciscus. Pozostałe 11 gatunków spotyka się w do
rzeczu górnego Sanu nielicznie i sporadycznie. Przyczyny ich niskiej liczebności są
następujące: jedne z nich to gatunki charakterystyczne dla środkowego biegu rzeki
('Ch. nasus, B. barbus, G. kessleri), a nawet gatunki limnofilne {Rh. sericeus cmarus,
T. tinea, R. rutilus, P. fluviatilis, A. cernua), które nie znajdują w górnym biegu odpo
wiednich warunków ekologicznych; drugie — to gatunki odznaczające się niskim zagę
szczeniem populacji, występujące nielicznie także w dorzeczu środkowego biegu Sanu
(C. auratus gibelio, B. cyclolepis waleckii, A. anguilla). W dorzeczu środkowego biegu Sanu stwierdziłam 35 gatunków, w tej liczbie wszyst
kie gatunki wymienione dla dorzecza górnego z wyjątkiem C. poecilopus. Dwa gatunki —
S. erythrophthlmus i M . fossilis — złowiono tylko w starorzeczu w Hurko. Pozostałe
nie spotkane w dorzeczu górnego Sanu to: E. lucius, B. bjoerkna, A. brama, L. deli-
neatus, C. carassius, A. aspius, P. cultratus, V. vimba, C. (S.) aurata, S. glanis, I. nebulosus, L. lota. Porównując uzyskane przeze mnie dane ze stanem ichtiofauny stwierdzonym przez
N o w ickiego (1880a, 1889) należy odnotować brak obecnie ć>. salar, S. trutta i A. sturio. Jeszcze K ołder (1958) zaznaczył na mapie wędrówki S. trutta do dorzecza
Sanu, a S c h r a m m (1957) wspomina o tarliskach S. salar lub S. trutta w dorzeczu
górnego Sanu w latach 30-tych bieżącego stulecia. Prawdopodobnie znacznie rzadszy
jest obecnie A . anguilla, który w drugiej połowie ubiegłego stulecia miał znaczenie
nawet dla rybołówstwa. Kiełbia długowąsego, tj. G. kessleri oraz P. cultratus zaliczył
N o w ic k i do gatunków rzadkich w Galicji. Anguilla anguilla (Linnaeus) — węgorz Nadmienia on, że mimo wyznaczenia wy
sokiej nagrody pieniężnej nie udało mu się otrzymać P. cultratus z karpackich dopływów http://rcin.org.pl H. Rolik 576 zostałych stanowiskach liczebność tego głowacza wahała się w granicach od 1 do 11
okazów. Anguilla anguilla (Linnaeus) — węgorz Anguilla anguilla (Linnaeus) — węgorz http://rcin.org.pl Ichtiofauna dorzecza Sanu 577 T a b e l a 2 . S p i s g a t u n k ó w r y b w y s t ę p u j ą c y c h w g ó r n y m i ś r o d k o w y m b i e g u S a n u o r a z w d o p ł y w a c h z e
s t a w i o n y
n a p o d s t a w i e b a d a ń
w ł a s n y c h w p o r ó w n a n i u z
d a n y m i N o w i c k i e g o ( 1 8 8 0 a , 1 8 8 0 b ,
1 8 8 9 )
Gatunek
San
wg
N owickiego
Górny
bieg
Sanu
Dopływy
górnego
biegu
Sanu
Środkowy
bieg
Sanu
Dopływy
środkowego
biegu
Sanu
1
2
3
4
5
6
Acipenser sturio L. +
Salmo salar L. ~r
—
—
—
_
Salmo trutta L. _ #
—
—
—
_
Salmo trutta nr. fario L. +
+
+
—
+
Esox lucius L. +
-
—
+
( + )
Rutilus rutilus (L.)
+
—
+
+
+
Leuciscus leuciscus (L.)
+
+
+
+
+
Leuciscus cephalus (L.)
+
+
+
+
+
Leuciscus idus (L.)
+
—
—
—
—
Phoxinus phoxinus (L.)
+
+
+
+
+
Scardinius erythrophthalmus
(L.)
___
( + )
Aspius aspius (L.)
+
-
—
+
—
Leucaspius delineatus (IIeck.)
+
-
—
+
—
Tinea tinea (L.)
+
-
+
+
( + )
Chondrostoma tiasus (L.)
+
+
+
+
+
Gobio gobio (L.)
+
+
+
+
+
Gobio kessleri D y b . _j_##
+
+
+
—
Barbus barbus (L.)J
+
+
+
+
+
Barbus petenyi H eck. +
+
+
+
+
Barbus cyclolepis H eck. http://rcin.org.pl -
+
+
+
+
Alburnus alburnus (L.)
+
+
+
+
+
Alburnoides bipunctatus (Bl.)
+
+
+
+
+
Blicca bjoerkna (L.)
-
-
—
+
—
Abramis brama (L.)
+
—
—
+
—
Vimba vimba (L.)
+
—
—
+
—
Pelecus cultratus (L.)
—
—
—
+
—
Rhodeus sericeus amarus (Bl.)
+
—
+
+
(+ )
Carassius carassius (L.)
+
—
—
—
+
Carassius auratus gibelio
(Bl.)
+
+
Cyprinus carpio L. +
-
—
—
—
Nemachilus barbatulus (L.)
+
+
+
+
+
Cobitis (Sabanejewia) aur a ta
(F il.)
+
Misgurnus fossilis (L.)
-
-
—
—
(+ )
Silurus glanis L. +
-
—
+
—
Ictalurus nebulosus (Le Sueur)
. -
-
—
+ ? —
Anguilla anguilla (L.)
+
+ ? —
+ ? —
Lota lota (L.)
+
-
-
+
- T a b e l a 2 . S p i s g a t u n k ó w r y b w y s t ę p u j ą c y c h w g ó r n y m i ś r o d k o w y m b i e g u S a n u o r a z w d o p ł y w a c h z e "
s t a w i o n y
n a p o d s t a w i e b a d a ń
w ł a s n y c h w p o r ó w n a n i u z
d a n y m i N o w i c k i e g o ( 1 8 8 0 a , 1 8 8 0 b ,
1 8 8 9 ) . http://rcin.org.pl H. Rolik 578 tabela 2 c.d. 1
2
3
4
5
6
Stizostedion lucioperca (L.)
+
_
_
_
_
Perea fluviatilis L. +
+
+
+
( + )
Acerina cernua (L.)
+
—
+
+
-
Cottus gobio L. -
+
+
+
+
Cottus poecilopus Heck. —
—
+
—
-
+ — gatunek stwierdzony na podstawie zebranych materiałów
+ ? — gatunek stwierdzony na podstawie informacji wędkarzy i rybaków
(-f)
— gatunek występuje w starorzeczach w okolicy Przemyśla
—
— gatunku nie stwierdzono
*
— N o w i c k i nie wymienia troci a przytacza tylko łososia
* *
— N o w i c k i podaje „kiełbia dlugowąsego” Wisły. W związku z tym znalezienie tego gatunku w Sanie jest bardzo interesujące. Do gatunków wykazanych w dorzeczu Sanu po raz pierwszy należą także S. erythro-
phthalmus, B. bjoerkna i /. nebulosus. ANALIZA EKOLOGICZNA Do charakterystyki ekologicznej ichtiofauny badanego obszaru obrałam zastoso
wany przez P en cza k a (1969) wskaźnik stałości i wartości dominacji (tabela 3) oraz układ
ekologiczny ryb K ryżanow skiego (1949) (tabela 4). Opracowany przez K ryżanow skiego
go układ ekologiczny ryb wielokrotnie był już stosowany do charakterystyki ichtiofauny
słodkowodnej zarówno w Polsce (B a l o n 1964, P en cza k 1969) jak i w innych krajach
(B a l o n 1962, H o lCi' k 1966a i b). H o lCIk (1966a) i P en cza k (1969), analizując kry
tycznie przynależność niektórych gatunków do poszczególnych grup ekologicznych,
nieco ten układ zmodyfikowali, poszerzając znacznie listę gatunków z grupy indyfe-
rentnej. Przynależność niektórych gatunków do odpowiedniej grupy ekologicznej,
jak się wydaje, nadal można uważać za dyskusyjną (np. przynależność L. łeuciscus
do grupy indyferentnej). A . alburnus nie powinien być zaliczany do grupy fitofilnej,
na co zwrócili uwagę H o lCIk i H r uSka (1966). H o lCIk (1966a i b) umieścił ten gatunek
pośród ryb indyferentnych. Także G ąsow ska (1962) podaje, że ukleja w rzekach obie
ra do tarła miejsca kamieniste. Z 36 gatunków odnotowanych w dorzeczu górnego i środkowego biegu Sanu do grupy
litofilnej należy 12 gatunków, do fitofilnej 11 (dwa z nich tylko w starorzeczu), gatun
ków indyferentnych jest 5, psammofilnych 4 i po jednym gatunku z grup: pelagofilnej,
częściowo pelagofilnej, ostrakofilnej i specjalnej (tabela 4). Do gatunków najczęściej spotykanych należy L. cephalus, G. gobio, B. petenyi (od
44 do 47 stanowisk, wskaźnik stałości 77,2-82,4%). Następnie do gatunków częścije
spotykanych należą Ph. phoxinus, N . barbatulus, A. alburnus, A. bipunctatus (36-39 http://rcin.org.pl Obszar
Liczebność
gatunków
Górny
bieg Sanu
Środkowy
bieg Sanu
Dopływy
biegu
górnego
Dopływy
biegu
środkowego
Starorze
cza
Ogółem
Grupa
ekologiczna
n
%
n
%
n
V/o
n
%
n
°//o
n
%
litofilna
9
56,25
10
33,33
10
50,00
9
52,94
12
33,33
indyferentna
3
18,75
5
16,67
4
20,00
3
17,65
1
14,29
5
13,89
fitofilna
—
—
7
23,33
2
10,00
3
17,65
5
71,43
11
30,56
psammofilna
3
18,75
4
13,33
3
15,00
2
11,76
—
—
4
11,11
ostrakcfilna
pelagofilna i częściowo
—
—
1
3,33
1
5,00
—
—
1
14,28
1
2,78
pelagofilna
—
—
2
6,67
—
—
—
—
—
2
5,55
specjalna
1
6,25
1
3,33
1
2,78
ogółem
16
100,00
30
100,00
20
100,00
17
100,00
7
100,00
36
100,00
stanowisk, wskaźnik stałości 63,1-68,4%), CA. nasus, B. barbus, L. leuciscus, C. gobio,
S. trutta m . fario (24-28 stanowisk, wskaźnik stałości 42,1-49,1%). Rzadziej występują
P . cyclolepis waleckii, C. poecillopus, G. kessleri (13-14 stanowisk, wskaźnik stałości
22.8-24,6%), jeszcze rzadziej P. fluviatilis, Rh. sericeus amarus, R. rutilus (4-7 stano
wisk, wskaźnik stałości 7,0-12,3%), T. tinea, C. auratus gibelio, C. carassius, F. vimba,
zł. cernua, L. lota, M. fossilis i S’. erythrophthlamus (1-2 stanowiska, wskaźnik stałości
1.8-3,5%). Najwyższą dominacją charakteryzują się PA. phoxinus i P. petenyi (16,40-18,74%),
następnie P. cephalus i A/", barbatulus (9,76-10,87%). Dominacja G. CA. wasws,
zł. bipunctatus, C. poecillopus, P. barbus, zł. alburnus, S. trulta m. fario waha się w gra
i
h 4 03 6 77%
L
l
i
C
bi
P
l l
i
l
kii
1 64 3 38%
D
i Tabela 4. Liczebność gatunków z określonych grup ekologicznych w poszczególnych partiach dorzecza
Sanu. http://rcin.org.pl Obszar
Liczebność
gatunków
Górny
bieg Sanu
Środkowy
bieg Sanu
Dopływy
biegu
górnego
Dopływy
biegu
środkowego
Starorze
cza
Ogółem
Grupa
ekologiczna
n
%
n
%
n
V/o
n
%
n
°//o
n
%
litofilna
9
56,25
10
33,33
10
50,00
9
52,94
12
33,33
indyferentna
3
18,75
5
16,67
4
20,00
3
17,65
1
14,29
5
13,89
fitofilna
—
—
7
23,33
2
10,00
3
17,65
5
71,43
11
30,56
psammofilna
3
18,75
4
13,33
3
15,00
2
11,76
—
—
4
11,11
ostrakcfilna
pelagofilna i częściowo
—
—
1
3,33
1
5,00
—
—
1
14,28
1
2,78
pelagofilna
—
—
2
6,67
—
—
—
—
—
2
5,55
specjalna
1
6,25
1
3,33
1
2,78
ogółem
16
100,00
30
100,00
20
100,00
17
100,00
7
100,00
36
100,00 Tabela 4. Liczebność gatunków z określonych grup ekologicznych w poszczególnych partiach dorzecza
Sanu. Tabela 4. Liczebność gatunków z określonych grup ekologicznych w poszczególnych partiach dorzecza
Sanu. stanowisk, wskaźnik stałości 63,1-68,4%), CA. nasus, B. barbus, L. leuciscus, C. gobio,
S. trutta m . fario (24-28 stanowisk, wskaźnik stałości 42,1-49,1%). Rzadziej występują
P . cyclolepis waleckii, C. poecillopus, G. kessleri (13-14 stanowisk, wskaźnik stałości
22.8-24,6%), jeszcze rzadziej P. fluviatilis, Rh. sericeus amarus, R. rutilus (4-7 stano
wisk, wskaźnik stałości 7,0-12,3%), T. tinea, C. auratus gibelio, C. carassius, F. vimba,
zł. cernua, L. lota, M. fossilis i S’. erythrophthlamus (1-2 stanowiska, wskaźnik stałości
1.8-3,5%). Najwyższą dominacją charakteryzują się PA. phoxinus i P. petenyi (16,40-18,74%),
następnie P. cephalus i A/", barbatulus (9,76-10,87%). Dominacja G. CA. wasws,
zł. bipunctatus, C. poecillopus, P. barbus, zł. alburnus, S. trulta m. fario waha się w gra
nicach 4,03-6,77%; L. leuciscus, C. gobio, P. cyclolepis waleckii — 1,64-3,38%. Domi
nacja pozostałych gatunków, typowo limnofilnych, spotykanych w badanym terenie
pojedynczo, wynosi setne części procenta. Oba przedstawione parametry, mianowicie wskaźnik stałości i dominacja, bardzo
wyraźnie charakteryzują zarówno frekwencję poszczególnych gatunków, jak i ich
liczebność. Analiza ekologiczna wskazuje na to, że badany obszar dorzecza Sanu jest zamiesz
kały głównie przez ichtiofaunę górską i podgórską. Mimo wysokiej liczebności ga
tunków fitofilnych, wskaźniki stałości i dominacji tych gatunków są niewspółmiernie
niskie w porównaniu z odpowiednimi parametrami w grupie gatunków litofilnych. Instytut Zoologiczny PAN
Warszawa, Wilcza 64 http://rcin.org.pl http://rcin.org.pl Ichtiofauna dorzecza Sanu 579 Tabela 3. Podział gatunków ryb dorzecza górnego i środkowego Sanu według grup ekologicznych
z uwzględnieniem wskaźnika stałości (dla 57 stanowisk) oraz dominacji w procentach. z uwzględnieniem wskaźnika stałości (dla 57 stanowisk) oraz dominacji w procentach. Grupy
Gatunki
Stałość
Dominacja
n
ekologiczne
zakres
M
litofilna
Salmo trutta m. fario
42,1
0-44,70
4,03
241
Leuciscus cephalus
82,5
0-62,50
10,84
649
Phoxinus phoximis
68,4
0-72,98
18,75
1122
Aspius aspius*
—
-
-
-
Chondrostoma nasus
49,1
0-78,12
5,91
354
Barbus barbus
49,1
0-37,14
4,19
251
Barbus petenyi
77,2
0-39,77
16,39
981
Barbus cyclolepis waleckii
35,1
0-12,88
1,64
98
Alburnoides bipunctatus
63,1
0-21,43
5,66
339
Vimba vimba*
1,8
—
—
—
Cołtus gobio
45,6
0 -12,33
2,62
157
Cottus poecilopus
24,6
0-55,30
4,71
282
indyferentna
Leuciscus leuciscus
49,1
0-14,24
(-26,56)
3,38
202
Albnrnus alburnus
65,0
0-80,00
4,13
247
Ictalurus nebulosus*
—
—
—
—
Perea fluviatilis
12,3
0-1,56
0,05
3
Acerina cernua
1,8
0-0,27
0,017
1
fitofilna
Esox lucius*
—
—
—
—
Rutilus rutilus
Scardittius
7,0
0-7,81
0,13
8
eryłhrophthalmus**
1,8
—
—
—
Leucaspius delineaius*
—
—
—
—
Tinea tinca
3,5
0-0,27
0,017
1
Blicca bjoerkna
1,8
0-1,08
0,017
1
Abramis brama*
—
—
—
—
Carassius carassius
1,8
0-3,12
0,033
2
Carassitts auratus gibelio
3,5
0-1,51
0,033
2
Misgurnus fossilis**
1,8
—
—
—
Silurus glanis*
—
—
—
—
psammofilna
Gobio gobio
82,4
0-35,82
6,77
405
Gobio kessleri
22,8
0-32,50
' 0,52
31
Nemachilus barbatulus
68,4
0-28,57
9,76
584
Cobitis aurata*
—
—
—
—
ostrakofilna
Rhodeus sericeus amarus
7,0
0-20,00
0,38
23
pelagohlna
Pelecus cultratus*
—
—
—
i częściowo
pelagofilna
Lota lota
1,8
0-6,25
0,017
1
specjalna
Anguilla anguilla*
—
—
—
—
Ogółem
100,00
5985
* tylko w połowach siecią lub wędką. ** t lk
t
i k
h
ł
h
bli
ń * tylko w połowach siecią lub wędką. ** tylko w stanowiskach wyłączonych z obliczeń. H. Rolik 580 Tabela 4. Liczebność gatunków z określonych grup ekologicznych w poszczególnych partiach dorzecza
Sanu. PIŚM IENNICTW O B a c k i e l T. 1964. Populacje ryb w systemie rzeki Drwęcy. Roczn. Nauk roln., Warszawa, 84 B: 193—
-214, 5 tab., 4 ff. B a l o n E. K. 1962. Prispevok k ekologickej charakteristike ichtyofauny ćeskoslovenskeho useku Dunaja. Biologia, Bratislava, 17: 283-296, 7 tabl., 1 f. 1964. Spis i ekologiczna charakterystyka
słodkowodnych krągłoustych i ryb Polski. Po B a l o n E. K. 1964. Spis i ekologiczna charakterystyka
słodkowodnych krągłoustych i ryb Polski. Pol. Arch. Hydrobiol., Warszawa, 12 (25): 233-251, 2 tab. B o n t e m p s S. 1960. Ocena stanu pogłowia certy w systemie rzeki Wisły. Roczn. Nauk roln., Warszawa,
75 B: 179-211, 16 tab., 7 ff. B o n t e m p s S. 1969. Zagadnienie występowanie lokalnych form certy w systemie Wisły. Zesz. nauk. SGGW , Warszawa, Zootechnika 7, Rybactwo 3: 57-70, 3 tab., 2 ff. C h l e b o w s k i B. i in. 1889. Słownik geograficzny Królestwa Polskiego i innych krajów słowiańskich,
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Warszawa, 8: 435-414, 2 tab., 4 tt. G ą s o w s k a M. i in. 1962. Krągłouste — Cyclostotni, Ryby — Pisces. Klucze d o oznaczania kręgowców
Polski, I. Warszawa-Kraków, 240 pp., 168 ff., 23 mapy. r R. 1858. Die Siisswasserfische der óstreichischen Monarchie. Leipzig, 388 pp., 204 ff. H e c k i e l J., K n e r R. 1858. Die Siisswasserfische der óstreichischen Monarchie. Leipzig, HOLCik J. 1966 a. Vyvoj a formowanie ichtyofauny v Oravskej priehrade. Biológia, Bratislava, 12: 5-75,
357 tab., 8 ff. HOLCik J. 1966 b. Ichtyologicky vyskum Karpatskeho obluka. 4. Ichtyofauna rieky Hornad so zretel’om
na vybudovanie vodneho diela Rużin. Biológia, Bratislava, 12: 76-117, 5 tab., 2 ff., 11 fot. HObClK J., HRUSKA V. 1966. http://rcin.org.pl 581 Ichtiofauna dorzecza Sanu Ichtiofauna dorzecza Sanu PIŚM IENNICTW O Ichtiofauna rzek Wyżyny Łódzkiej i terenów przyległych. Część II. Ekologia. Acta
hydrobiol., Kraków, 11: 313-338, 3 tab., 5 ff. c z a k T. 1969. Ichtiofauna rzek Wyżyny Łódzkiej i terenów przyległych. Część II. Eko
hydrobiol., Kraków, 11: 313-338, 3 tab., 5 ff. P l i s z k a F. 1951. Wędrówki certy (Vimba vimba L.) w Wiśle. Roczn. Nauk roln z k a F. 1951. Wędrówki certy (Vimba vimba L.) w Wiśle. Roczn. Nauk roln., Warszawa, 5
f 1
f. PIŚM IENNICTW O On the spawning substrate of the roach — Rutilus rutiliis ( L i n n a e u s , 1758)
atnd bream — Abramis brama ( L i n n a e u s , 1758) and notes on the ecological characteristic of some
european fishes. VSstn. 2sl. Spol. zool., Praha, 30: 22-29, 3 ff. K l i m a s z e w s k i M. 1937. Z morfologii doliny Sanu między Leskiem a Przemyślem. Pr. geogr., Warszawa,
16 (1936): 107-132, 1 t. 1958. Zarybianie łososiami i trociami w górnej części systemu rzecznego Wisły w łatac
54
R
N
k
l
W
73 B
215 267
16 t b
1 K o ł d e r W. 1958. Zarybianie łososiami i trociami w górnej części systemu rzecznego Wisły w łatach
1879-1954. Roczn. Nauk roln., Warszawa, 73 B: 215-267, 16 tab., 1 mapa. y
g
j
y
g
y
54. Roczn. Nauk roln., Warszawa, 73 B: 215-267, 16 tab., 1 mapa. K R y Z a n o w s k i j S. G. 1949. Bkologo-morfologićeskie zakonomernosti razvitija karpowych, v’juno-
vyh
i somovyh
ryb (Cyprinoidei i Siluroidei). Trudy Ins. Morf. Żiv., Moskva-Leningrad,
1: 5-332, 93 tab., 99 ff. M i k u l s k i Z. 1963. Zarys hydrografii Polski. Warszawa, 288 p p . , 47 ff,. 28 tab., 1 m a p a . M o r a w s k a B. 1964. Płodność certy (Vimba vimba L.) systemu rzeki Wisły. Roczn. Nauk roln., Warsza
wa, 84 B: 315-327, 5 tab., 2 ff. N o w i c k i M. 1880 a. Ryby i wody Galicyi. Kraków, 96 + 10
pp. N o w i c k i M. 1880 b. Dodatki do rozsiedlenia ryb w wodach Galicyi. Kraków, 17 pp N o w i c k i M. 1882. Krainy rybne Wisły. Kraków, 16 pp. N o w i c k i M. 1883. Przegląd
rozsiedlenia ryb w wodach Galicyi według dorzeczy
i krain r
Wiedeń. N o w i c k i M. 1889. O rybach dorzeczy Wisły, Styru, Dniestru i Prutu w Galicyi. Kraków, 4 + 5 4 pp. P a s t e r n a k K. 1964. Geologiczna i gleboznawcza charakterystyka dorzecza rzeki Sanu. Acta hydrobiol.,
Kraków, 6: 289-307, 2 tab. P e n c z a k T. 1969. http://rcin.org.pl H. Rolik 582 P l is z k a F. 1 9 5 3 . Rozród i rozwój certy (Vimba vimba L.). Pol. Arch. Hydrobiol., Warszawa, 1 (14):
1 3 7 -1 6 3 , 3 tab., 7 ff. R o l i k H. 1959. Kiełb Kesslera (Gobio kessleri D y b . ) w Polsce. Fragm. faun. Warszawa, 8: 207-221,
7 tab. R o l i k H. 1960. Cobitis aurata ( F i l i p p i , 1865) — koza złotawa, nowy gatunek w zlewisku Morza Bał
tyckiego. Fragm. faun., Warszawa, 8: 413-420, 2 tab. R o l i k H. 1962. Biometria oraz niektóre zagadnienia biologii i systematyki Leuciscus cephalus (L.) z rzeki
San. Fragm. faun., Warszawa, 9: 355-372, 9 tab., 1 f. R o l i k H. 1965a. Uwagi o stanowisku systematycznym Gobio kessleri D y b . z Sanu. Fragm. faun., War
szawa, 11: 455-465, 4 tab., 1 t. R o l i k H. 1965b. Materiały dotyczące zmienności geograficznej i ekologicznej Gobio gobio (L.) w Polsce. Fragm. faun., Warszawa, 12: 15-29, 7 tab., 1 mapa. R o l i k H. 1967a. O niektórych naturalnych krzyżówkach ryb karpiowatych (Pisces, Cyprinidae) w Polsce. Fragm. faun., Warszawa, 14: 153-167, 6 tab., 2 tt. R o l i k H. 1967b. Materiały do ichtiofauny Strwiąża (dopływ Dniestru) ze szczególnym uwzględnieniem
Gobio gobio (L.) i Cobitis (Sabanejewia) aurata ( F i l . ) . Fragm. faun., Warszawa, 14: 133-151, 6 tab.,
2 tt. R o l i k H. 1970. Barbus cyclolepis waleckii ssp. n . — A New Subspecies o f B. cyclolepis H e c k e l , 1840,
from the Vistula and Dniestr Basins (Pisces, Cyprinidae). Bull. Acad. p o i . Sci., Ser. biol., Warszawa,
18: 401-404. R o l i k H. 1971. Studia nad gatunkami rodzaju Barbus C u v i e r , 1817, z dorzecza Sanu i Wisłoki (Pisces,
Cyprinidae). Ann. zool., Warszawa, 28: 257-330, 30 tab., 6 ff., 1 mapa. Schramm W. 1957. Uwagi do zoogeografii Atlasu Polski. Prz. zool., Wrocław, 1: 81-83. S z y p u ł a J. 1966. Obserwacje nad populacjami narybkowymi górnego Sanu. Prz. zool., Wrocław, 10:
304-307, 2 ff. S o l e w s k i W. 1962. Pstrąg potokowy (Salmo trutta m.fario L.) z dorzecza górnego Sanu. http://rcin.org.pl Acta hydrobiol.,
Kraków, 4: 47-57, 6 tab., 1 f. S o l e w s k i W. 1964. P s t r ą g p o t o k o w y (Salmo trutta m .fario L.) n i e k t ó r y c h r z e k k a r p a c k i c h . A c t a h y d r o
b i o l . , K r a k ó w , 6: 227-253, 12 t a b . , 12 ff. S t a r m a c h K. 1956. Rybacka
i biologiczna charakterystyka rzek. Pol. Arch. Hydrobiol., Warszawa,
3 (16): 307-332, 5 tab., 4 ff. W a j d o w i c z Z. 1966. Zmiany ichtiofauny w rejonie zbiornika w Myczkowcach. Acta hydriobiol., Kraków
8, suppl. 1: 411-424, 4 tab., 3 ff. W a ł e c k i A. 1864. Materyały do fauny ichtyologicznej Polski. II. Systematyczny przegląd ryb krajowych. Warszawa, 6 + 115 pp. W a ł e c k i A. 1864. Materyały do fauny ichtyologicznej Polski. II. Systematyczny przegląd ryb krajowych. Warszawa, 6 + 115 pp. Z a w a d z k i A. 1840. Fauna der galizischen-bukowinischen Wirbelthiere. Stuttgart, pp. 162-182. Z a w a d z k i A. 1840. Fauna der galizischen-bukowinischen Wirbelthiere. Stuttgart, pp. 162-182. PE3I0M E [3 a rjia B H e: H xTH O <|)ayH a Gacceima B e p x n e r o
i i cpe^Hero T e q e H H fl C ana]
MxTiiojiorivieCKHe iiccjieAOBaHHH, npoBefleim bie
b Gacceirae BepxHero
h cp e/u
nero TeqeHHH CaHa no IlmeMbicjiH, G i.t jih nocBHipeHbi H3yqeHHio B H /iO B o r o
c o c i a B a
HXTHopeiiojiorHqecKHX KOMnjiencoB, xapaKTepHbix ajih om ejibiibix yqacTKOB CaHa
h ero npHTOKOB. McnoJib30BaHHbifi b paG oie MaTepnaji c o c t o h j i H3 8500 3K3eMHJiH-
poB, ii3 h h x 7300 3K3eMn.iHpoB GbuiH coGpaHbi npw h o m o i r h ajieKTpiiqecKoro arp e-
r a ia nocTOHimoro Tona. B 3aMeqaHwax k n a p ie (pnc. 1) ynaaaHbi 57 c ia m p m , http://rcin.org.pl http://rcin.org.pl http://rcin.org.pl http://rcin.org.pl 583 Ichtiofauna dorzecza Sanu b KOTopi.ix b nepBoii nojioBHHe hw hh 1966-1968 r r . 6łijih npoH3Be,n;eHbi ajieKTpo-
jiobłi. B iiccjieAOBaHHflx no hhcjiohhocth n n,enojiornnecKHM OTHomeHHHM hc-
nojib30BaH TOJibKO MaTepnaji cofipaniibin npn noMOinn 3JieKTpnnecKoro a rp e ra ia . n p n o63ope
bhaob
ncnojib30Ban Becb
Maxepnaji co6paHHbui Ha npoTHxemiH
1956-1966 rr. TaK>Ke npn noMOipn pa3JiHim oro po^a
ceTOK. p
p
p
p
HxTHopeHOJiornnecKne KOMnjieKCbi yCTanaBJiHBajincb Ha oCHOBaimn yKJiOHa
h reoMop(|)OJiorHqecKoro cipoeHHH /ma penn h /joMHHHpoBaHHH onpe,n;ejieHHbix
BHflOB. Kpowe BH^OB-^OMHHaHTOB, pa3JIHHajIHCb BHflbl COHyTCTByiOIĘHe H BH^H flO-
nojiHHTejibHbie. riocjie^Hno BCTpenajincb He Ha Bcex CTaHRHHx OTHeceHHbix k ,n;aH-
HOMy KOMHJieKCy HJIII BCTpenajIHCb e^HHHHHO. y
y
p
j
B nprnoK ax ropH oro TenerniH CaHa b Eema,n;ax Bbi^ejieHo 5 HXTHOu,eHOJiorn-
necKHX KOMnjieKCOB, b KOTopbix cjie^yn bhh3 no TeneHHio peKH flOMiiHHpoBajra
cjie^yiom ne bh^m: I KOMHJieKC — Cottus poeciłopus, II — Phoxinus phoxinus, III —
Phoxinus phoxinus hjih Barbus petenyi, IV — B. petenyi, V — B. barbus hjih B. petenyi. B npHTOKax cpe^nero TeneHHH, KOTopwe TO/Ke hocht b ochobhom xapaK Tep ro p -
iibix hjih npe^ropH bix noTOKOB, Bbi^eJieinibix Kan VI KOMnJienc,
,n,OMHHHpoBaji
Leuciscus cephalus. B npHTOKax
HH>Kiiero TeneiiHH — V II KOMnjienc, aomhhhpo-
Baji Chondrostoma nasus
(btot KOMnjieKC Tpe6yeT /jonojiHHTejibHbix MaTepnajioB). B BepxHeM TeneHHH C ana (KOMnjienc A), KOTopoe pacnpocTHpaeTcn ao PaficK oro
(noBHT JlecK o), flOMHirapyeT
B. petenyi;
b cpe^HeM
Teneim n, KOTopoe
xapaK -
TepH3yeTCH
HajimmeM
MHoronHCJiemibix
nepejiOMOB,
Me>K,n,y KOTopbiMH
Jie>KaT
Myjib/jooOpasHbie ynacTKH,
mo>kho Bbi^ejiHTb ^Ba KOMnjienca: B — nepejiOMbi,
r^e flOMimaHTOH hbjihgtch Ch. nasus h C — Myjib/i;oo6pa3Hbie yqacTKH — L. cephalus
(TaCji. 1, pnc. 2). H a HCCJieAOBanoM ynacTKe CaHa KOHCTampoBano 36 bh;i,ob pu6 (Ta6ji. 2),
4 H3
h h x: Scardinius erythrophthalmus, Blicca bjoerkna, Pelecus cultratus h Ictalurus
nebulosus He 6h jih oTMeneHbi /jo nacTonmero BpeMeHH b fiacceime Cana. K BH^aM nam e Bcero BCTpenaiomnMCH othochtch L. cephalus, B. petenyi h Gobio
gobio3 a k Han6ojiee MHoroHHCJiemibiM Ph. phoxinus h B. petenyi (Tafiji. 3). X oth
b HCCJie^oBaHHbix BOflax OTMeneHO ^OBojibHO Bbiconyio nucjieHHOCTb (Jjhto^ hjib-
Hbix bhaob (30,56% ) (Tafiji. 4), HXTHO({)ayHa fiacceim a B epxnero Tenennn
CaHa
HBJineTCH THnHHiio ro p iio n , a cpe/jHero TeneiiHH n p e^ ro p n o n ,
nocKOJibKy
$ h-
TO(|)HjibHbie BHflu BCTpenaioTCH TyT cnopa^HnecKH. SUMMARY SUMMARY [Title: Fishes of upper and middle San River and its tributaries] Total number of 7300 specimens for quantitative analysis was collected with
electric current form 57 collecting points (fig. 1) during two first weeks of June in
three consecutive years 1966-1968. To ensure completeness of the list of species http://rcin.org.pl http://rcin.org.pl http://rcin.org.pl H. Rolik 584 an additional material of 1200 specimens fished with nets in the same area in years
1956-1969 was also studied. Altogether 36 species of fishes were found (table 2) out
of which Scardinus erythrophthalmus, Blicca bjoerkna, Pelecus cultratus and Ictalurus
nebulosus are for the first time found in that area. an additional material of 1200 specimens fished with nets in the same area in years
1956-1969 was also studied. Altogether 36 species of fishes were found (table 2) out
of which Scardinus erythrophthalmus, Blicca bjoerkna, Pelecus cultratus and Ictalurus
nebulosus are for the first time found in that area. To express various riverain conditions (unitary slope gradient, character of river
bed) and their relation to fish communities the following river zones are described;
the list of species living in each zone is given on table 1 and on fig 2. 1) Tributaries of upper San River 1) Tributaries of upper San River
Zone I — dominant species — Cottus poecilopus Zone I — dominant species — Cottus poecilopus Zone II — dominant species — Phoxinus phoxinus Zone III — dominant species — either Ph. phoxinus or Barbus petenyi Zone IV — dominant species — B. petenyi Zone V — dominant species — B. barbus or B. petenyi 2) Tributaries of middle San River
Zone VI — dominant species — Leuciscus cephalus 2) Tributaries of middle San River
Zone VI — dominant species — Leuciscus cephalus 3) Tributaries of lower San River
Zone V III — dominant species — Chondrostoma nasus 4) U pper San River — down to village Rajskie
Zone A — domant species B. petenyi 5) M iddle San River — rapids
Zone B — dominant species — Chondrostoma nasus 6) M iddle San River — slowly flowing stretches — down to Przemyśl
Zone C — dominant species — Lcuciscus cephalus T he commonest species were L. cephalus, B. petenyi, and Gobio gobio which were
caught in the biggest number of collecting points, the most numerous, however, were
Ph. phoxinus and B. petenyi (table 3). T he commonest species were L. cephalus, B. SUMMARY petenyi, and Gobio gobio which were
caught in the biggest number of collecting points, the most numerous, however, were
Ph. phoxinus and B. petenyi (table 3). T he fish fauna of San River and of its tributaries has a distinctly rheophil character,
although the number of phytophilous species is rather high and amounts to 30.56%
(table 4) the number of specimens of such species is rather insignificant.
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https://openalex.org/W2602815172
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https://www.frontiersin.org/articles/10.3389/fonc.2017.00040/pdf
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English
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Targeting the Metabolic Reprogramming That Controls Epithelial-to-Mesenchymal Transition in Aggressive Tumors
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Frontiers in oncology
| 2,017
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cc-by
| 18,885
|
Targeting the Metabolic
Reprogramming That Controls
Epithelial-to-Mesenchymal
Transition in Aggressive Tumors
Andrea Morandi1*, Maria Letizia Taddei2, Paola Chiarugi1,3* and Elisa Giannoni1
1 Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy, 2 Department of
Experimental and Clinical Medicine, University of Florence, Florence, Italy, 3 Excellence Centre for Research, Transfer and
High Education DenoTHE, University of Florence, Florence, Italy 1 Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy, 2 Department of
Experimental and Clinical Medicine, University of Florence, Florence, Italy, 3 Excellence Centre for Research, Transfer and
High Education DenoTHE, University of Florence, Florence, Italy The epithelial-to-mesenchymal transition (EMT) process allows the trans-differentiation
of a cell with epithelial features into a cell with mesenchymal characteristics. This process
has been reported to be a key priming event for tumor development and therefore EMT
activation is now considered an established trait of malignancy. The transcriptional and
epigenetic reprogramming that governs EMT has been extensively characterized and
reviewed in the last decade. However, increasing evidence demonstrates a correlation
between metabolic reprogramming and EMT execution. The aim of the current review
is to gather the recent findings that illustrate this correlation to help deciphering whether
metabolic changes are causative or just a bystander effect of EMT activation. The
review is divided accordingly to the catabolic and anabolic pathways that characterize
carbohydrate, aminoacid, and lipid metabolism. Moreover, at the end of each part, we
have discussed a series of potential metabolic targets involved in EMT promotion and
execution for which drugs are either available or that could be further investigated for
therapeutic intervention. Review
published: 14 March 2017
doi: 10.3389/fonc.2017.00040 published: 14 March 2017
doi: 10.3389/fonc.2017.00040 Edited by:
Silvia Giordano,
University of Turin, Italy
Reviewed by:
K. B. Harikumar,
Rajiv Gandhi Centre for
Biotechnology, India
Maria Felice Brizzi,
University of Turin, Italy
*Correspondence:
Andrea Morandi
andrea.morandi@unifi.it;
Paola Chiarugi
paola.chiarugi@unifi.it Edited by:
Silvia Giordano,
University of Turin, Italy Reviewed by:
K. B. Harikumar,
Rajiv Gandhi Centre for
Biotechnology, India
Maria Felice Brizzi,
University of Turin, Italy *Correspondence:
Andrea Morandi
andrea.morandi@unifi.it;
Paola Chiarugi
paola.chiarugi@unifi.it Keywords: epithelial-to-mesenchymal transition, metabolic reprogramming, Warburg metabolism, OXPHOS, TCA
cycle, oncometabolites, amino acid, lipids Abbreviations: ACC, acetyl-CoA carboxylase; ACSLs, acyl-CoA synthetases; AMF, autocrine motility factor; CAFs,
cancer-associated fibroblasts; CAIX, carbonic anhydrase IX; CerS6, ceramide synthase 6; CHC, α-cyano-4-hydroxycinnamate;
ChREBP, carbohydrate-responsive element-binding protein; CSC, cancer stem cell; DHA, docosahexaenoic acid; ECM,
extracellular matrix; EMT, epithelial-to-mesenchymal transition; ETC, electron transport chain; FAs, fatty acids; FASN, fatty
acid synthase; FBP1, fructose-1,6-bisphosphatase; FH, fumarate hydratase; GDH, glutamate dehydrogenase; GLS, glutaminase;
GLUT, glucose transporter; GSK-3β, glycogen synthase kinase 3β; d-2HG, d-2-hydroxyglutarate; HGF, hepatocyte growth
factor; HIF-1α, hypoxia-inducible factor-1α; HK2, hexokinase; IDH, isocitrate dehydrogenase; IL-8, interleukin-8; α-KG,
α-ketoglutarate; LDs, lipid droplets; LDH, lactate dehydrogenase; MCTs, monocarboxylate transporters; ME1, malic enzyme 1;
MET, mesenchymal-to-epithelial transition; MMPs, matrix metalloproteases; mtROS, mitochondrial reactive oxygen species;
NHE1, sodium-proton exchange 1; NSCLC, non-small cell lung cancer; O-GlcNAc, O-linked N-acetyl-glucosamine; OXPHOS,
oxidative phosphorylation; PDAC, pancreatic ductal adenocarcinoma; PDH, pyruvate dehydrogenase; PDK4, pyruvate dehy-
drogenase kinase 4; PGI, glucose phosphate isomerase; PM, plasma membrane; PK, pyruvate kinase; PPARGC1A/PGC-1α,
peroxisome proliferator-activated receptor gamma coactivator 1 alpha; PPP, pentose phosphate pathway; SAH, S-adenosyl
homocysteine; SAM, S-adenosyl methionine; SCD, stearoyl-CoA desaturase-1; SDH, succinate dehydrogenase; SIRT, sirtuin;
SOD2, superoxide dismutase 2; SREBPs, sterol regulatory element-binding proteins; TAGs, triacylglicerols; TCA, tricarboxylic
acid; TDH, threonine dehydrogenase; TET1, ten–eleven translocation 1; TGF-β, transforming growth factor-β; TGIF2, TGF-β-
induced factor homeobox 2; TKTL1, transketolase-like protein 1 isoform; TNBC, triple-negative breast cancer; TNF-α, tumor
necrosis factor-α; TUFM, TU translation elongation factor mitochondrial; VEGF, vascular endothelial growth factor. Specialty section:
This article was submitted to Cancer
Molecular Targets and Therapeutics,
a section of the journal
Frontiers in Oncology Specialty section:
This article was submitted to Cancer
Molecular Targets and Therapeutics,
a section of the journal
Frontiers in Oncology Received: 22 December 2016
Accepted: 28 February 2017
Published: 14 March 2017 Accepted: 28 February 2017
Published: 14 March 2017 Keywords: epithelial-to-mesenchymal transition, metabolic reprogramming, Warburg metabolism, OXPHOS, TCA
cycle, oncometabolites, amino acid, lipids Abbreviations: ACC, acetyl-CoA carboxylase; ACSLs, acyl-CoA synthetases; AMF, autocrine motility factor; CAFs,
cancer-associated fibroblasts; CAIX, carbonic anhydrase IX; CerS6, ceramide synthase 6; CHC, α-cyano-4-hydroxycinnamate;
ChREBP, carbohydrate-responsive element-binding protein; CSC, cancer stem cell; DHA, docosahexaenoic acid; ECM,
extracellular matrix; EMT, epithelial-to-mesenchymal transition; ETC, electron transport chain; FAs, fatty acids; FASN, fatty
acid synthase; FBP1, fructose-1,6-bisphosphatase; FH, fumarate hydratase; GDH, glutamate dehydrogenase; GLS, glutaminase;
GLUT, glucose transporter; GSK-3β, glycogen synthase kinase 3β; d-2HG, d-2-hydroxyglutarate; HGF, hepatocyte growth
factor; HIF-1α, hypoxia-inducible factor-1α; HK2, hexokinase; IDH, isocitrate dehydrogenase; IL-8, interleukin-8; α-KG,
α-ketoglutarate; LDs, lipid droplets; LDH, lactate dehydrogenase; MCTs, monocarboxylate transporters; ME1, malic enzyme 1;
MET, mesenchymal-to-epithelial transition; MMPs, matrix metalloproteases; mtROS, mitochondrial reactive oxygen species;
NHE1, sodium-proton exchange 1; NSCLC, non-small cell lung cancer; O-GlcNAc, O-linked N-acetyl-glucosamine; OXPHOS,
oxidative phosphorylation; PDAC, pancreatic ductal adenocarcinoma; PDH, pyruvate dehydrogenase; PDK4, pyruvate dehy-
drogenase kinase 4; PGI, glucose phosphate isomerase; PM, plasma membrane; PK, pyruvate kinase; PPARGC1A/PGC-1α,
peroxisome proliferator-activated receptor gamma coactivator 1 alpha; PPP, pentose phosphate pathway; SAH, S-adenosyl
homocysteine; SAM, S-adenosyl methionine; SCD, stearoyl-CoA desaturase-1; SDH, succinate dehydrogenase; SIRT, sirtuin;
SOD2, superoxide dismutase 2; SREBPs, sterol regulatory element-binding proteins; TAGs, triacylglicerols; TCA, tricarboxylic
acid; TDH, threonine dehydrogenase; TET1, ten–eleven translocation 1; TGF-β, transforming growth factor-β; TGIF2, TGF-β-
induced factor homeobox 2; TKTL1, transketolase-like protein 1 isoform; TNBC, triple-negative breast cancer; TNF-α, tumor
necrosis factor-α; TUFM, TU translation elongation factor mitochondrial; VEGF, vascular endothelial growth factor. Citation: Morandi A, Taddei ML, Chiarugi P
and Giannoni E (2017) Targeting the
Metabolic Reprogramming That
Controls Epithelial-to-Mesenchymal
Transition in Aggressive Tumors. Front. Oncol. 7:40. doi: 10.3389/fonc.2017.00040 March 2017 | Volume 7 | Article 40 1 Frontiers in Oncology | www.frontiersin.org Targeting EMT-dependent Metabolic Reprogramming Morandi et al. INTRODUCTION current review is on EMT in cancer progression although this
process shares common characteristic with the physiological
EMT program. The molecular mechanisms that an epithelial
cell undergoes during EMT are tightly regulated and influenced
by the cell-to-cell signaling network and by environmental
factors. Independently of the stimuli that induce EMT, loss of
E-cadherin is considered the key event and Snail1, Snail2 (also
known as Slug), Twist, and ZEB1 are established transcription
factors that can regulate E-cadherin expression. These mecha-
nisms have been elegantly reviewed in Ref. (3). Conversely, the
metabolic rewiring that sustains an epithelial cell undergo-
ing EMT has been poorly investigated. However, since the
metabolic reprogramming is now considered a hallmark of The epithelial-to-mesenchymal transition (EMT) process
allows the trans-differentiation of a cell with epithelial features
into a cell with mesenchymal characteristics (Figure 1). This
process has an essential role in physiological conditions (e.g.,
development, wound healing, and stem cell maintenance) and
has been extensively reported to contribute pathologically to
fibrosis and cancer progression. Interestingly, cells can also
undergo the mesenchymal-to-epithelial transition (MET)
process. MET is also essential in physiological conditions
(e.g., during embryogenesis) but a key role in the formation
of secondary metastases was reported (1, 2). The focus of the FIGURE 1 | General features of epithelial-to-mesenchymal transition (EMT). The transition of epithelial cells toward a mesenchymal phenotype, induced by
several environmental or soluble factors, is characterized by the loss of cell–cell contact and cell polarity, which disrupts the epithelial architecture and endows the
mesenchymal cells with migratory and invasive competences. EMT is accompanied by the modulation of well-known markers, among which the loss of epithelial
marker E-cadherin, induced by the upregulation of its transcriptional repressors (i.e., Snail1/2, Twist, ZEB1/2), is one of the priming event. The concomitant
acquisition of mesenchymal markers sustains and stabilizes the newly acquired phenotype. FIGURE 1 | General features of epithelial-to-mesenchymal transition (EMT). The transition of epithelial cells toward a mesenchymal phenotype, induced by
several environmental or soluble factors, is characterized by the loss of cell–cell contact and cell polarity, which disrupts the epithelial architecture and endows the
h
l
ll
ith
i
t
d i
i
t
EMT i
i d b th
d l ti
f
ll k
k
hi h th l
f
ith li l FIGURE 1 | General features of epithelial-to-mesenchymal transition (EMT). Targeting Carbohydrates Metabolic
Reprogramming in EMTf Non-proliferating differentiated cells preferentially metabolize
glucose to pyruvate through glycolysis and then oxidize this
pyruvate via the tricarboxylic acid (TCA) cycle and subsequent
oxidative phosphorylation (OXPHOS). This maximizes the
efficiency of ATP generation from a single molecule of glucose. Otto Warburg first described that most cancer cells show
increased glucose conversion into lactate even in oxygen-rich
condition (aerobic glycolysis) (5). Aerobic glycolysis allow
proliferating cell to satisfy three basic needs of cells in rapid divi-
sion: (i) fast ATP; (ii) carbohydrates redirection to biosynthetic
pathways; and (iii) cellular redox status homeostasis (6). As a
direct consequences of a net increase in glucose consumption,
many cancers exhibit abnormal lactic acid release and a more
acidic extracellular pH (pHe) (7). Moreover, high level of lactate
correlates with metastases of several types of human cancers (8). In this scenario, it is not surprising that hypoxia, low pHe, and
glucose utilization are established features of many solid tumors
and concur to EMT and cancer dissemination. Thus, interfering
with the glycolytic pathway could impair EMT and subsequent
tumor progression and could be a potential anticancer strategy. However, to our knowledge, none of glycolysis inhibitors that
have shown promising results in preclinical models are currently
used in the clinic. Glycolytic metabolism has been also associated to CSC,
a phenotype that shares common molecular pathways with
EMT, and characterizes the cells that within the tumor tissue
are responsible for tumor repopulation, therapy resistance, and
relapse in several cancer types (19). However, contradictory
results described the CSC metabolic phenotype as glycolytic
or OXPHOS addicted not only in various tumor types, but also
within individual cancer types. Some studies have postulated
that glycolysis supports the self-renewal ability of CSCs by
maintaining low ROS levels (19, 20) and in line with this
glucose deprivation reduces the number of CSCs in in vitro
studies (21). In addition, the enhancement of the glycolytic
flux has been shown to be paralleled by a decrease in mito-
chondrial metabolism (i.e., TCA cycle and OXPHOS) with
respect to their differentiated counterparts (22–26). Gammon
et al. demonstrated that the CSC fraction of head and neck
squamous cell carcinoma that undergoes hypoxia-induced
EMT is characterized by glycolysis, a reduction in the oxygen
consumption rate and decreased mitochondrial mass and ROS
levels (27). Notably, Dong et al. demonstrated the concomitant
acquisition of CSC-like properties, EMT, and induction of
glycolysis following fructose-bisphosphatase 1 silencing in
breast cancer (14). INTRODUCTION The transition of epithelial cells toward a mesenchymal phenotype, induced by
several environmental or soluble factors, is characterized by the loss of cell–cell contact and cell polarity, which disrupts the epithelial architecture and endows the
mesenchymal cells with migratory and invasive competences. EMT is accompanied by the modulation of well-known markers, among which the loss of epithelial
marker E-cadherin, induced by the upregulation of its transcriptional repressors (i.e., Snail1/2, Twist, ZEB1/2), is one of the priming event. The concomitant
acquisition of mesenchymal markers sustains and stabilizes the newly acquired phenotype. FIGURE 1 | General features of epithelial-to-mesenchymal transition (EMT). The transition of epithelial cells toward a mesenchymal phenotype, induced by
several environmental or soluble factors, is characterized by the loss of cell–cell contact and cell polarity, which disrupts the epithelial architecture and endows the
mesenchymal cells with migratory and invasive competences. EMT is accompanied by the modulation of well-known markers, among which the loss of epithelial
marker E-cadherin, induced by the upregulation of its transcriptional repressors (i.e., Snail1/2, Twist, ZEB1/2), is one of the priming event. The concomitant
acquisition of mesenchymal markers sustains and stabilizes the newly acquired phenotype. March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 2 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. The importance of glycolysis for EMT is also reinforced by
a metabolite profiling approach conducted in pancreatic ductal
adenocarcinoma (PDAC) cells. This profiling identified three
main subpopulations with different phenotypes: interestingly,
the clone that is enriched for glycolytic-related metabolites is
characterized by a mesenchymal phenotype. Since mesenchymal
properties are positive correlated with cancer aggressiveness
and diseases progression, the study reinforced the functional
relevance of glycolytic metabolism in disease progression (17). In
addition, the exposure of PDAC cells to established EMT induc-
ers [i.e., tumor necrosis factor-α (TNF-α) and TGF-β] increases
glucose uptake and lactate secretion, without affecting OXPHOS
metabolism (18). cancer (4), increasing evidence links metabolic deregulation
to EMT program. This review gathers the recent findings on
EMT and metabolic reprogramming in cancer and discusses
how targeting certain metabolic pathways/hubs may impact
on EMT and therefore on cancer progression. Targeting Carbohydrates Metabolic
Reprogramming in EMTf Several glycolytic enzymes have been found associated with
invadopodia structures, protrusions of the plasma membrane
(PM) that have a major role in extracellular matrix (ECM) deg-
radation, and metastasis (9). In addition, the glycolytic-derived
ATP is the main source for cell survival during metastatic
dissemination (10, 11). Several papers indeed show a strict
correlation between transforming growth factor-β (TGF-β)-
induced EMT, glycolytic switch, and repression of mitochondrial
function (12, 13). Dong et al. have demonstrated that in breast
cancer, loss of fructose-1,6-bisphosphatase together with the loss
of E-cadherin promote cancer stem cell (CSC)-like properties
and cancer cell dissemination by enhancing β-catenin signal-
ing and EMT program. These events are concomitant with the
induction of glycolysis, increase of glucose uptake, and inhibi-
tion of oxygen consumption (14). Additionally, EMT induction
in breast cancer cells is paralleled by the expression of glucose
transporters, lactate dehydrogenase (LDH), monocarboxylate
transporters (MCTs), and glycogen phosphorylase isoforms,
key players in sustaining enhanced aerobic glycolysis (15). Moreover, the acquisition of a malignant and chemo-resistant
phenotype is associated with EMT and aerobic glycolysis in
gastric cancer (16). We now proceed analyzing the principal players involved in
sustaining the Warburg metabolism with a particular focus on the
transporters and glycolytic enzymes whose deregulation has been
linked to the acquisition of EMT traits and subsequent enhanced
metastatic potential in cancer cells. Hypoxia
f In fast-growing solid cancers, the inner zone of the tumor becomes
progressively hypoxic and acid. These environmental inputs
drive the expression and stabilization of the hypoxia-inducible
factor-1α (HIF-1α). HIF-1α is responsible for the transcriptional
activation of genes involved in the restoration of local oxygen
perfusion and in the enhancement of the glycolytic flux (28, 29). In addition to its canonical role, HIF-1α is able to regulate the
expression of matrix metalloproteases (MMPs) (30) and several
hypoxia-inducible EMT-related genes (31) such as Snail1, Slug,
and Twist. In addition, HIF-1α regulates the autocrine motility
factor (AMF), the secreted form of the glycolytic enzyme glucose
phosphate isomerase (PGI). AMF ectopic expression triggers
EMT in breast cancer cells (32). Since hypoxia is a well-known
inducer of EMT in epithelial cancers, such as PDAC (33), March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 3 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. hepatocellular (34), ovarian (35), and lung cancers (36), we
can speculate that this may be partially due to the metabolic
reprogramming orchestrated by HIF-1α. More recently, also in
glioblastoma, it has been demonstrated that HIF-1α silencing is
able to prevent EMT induced by the hypoxic microenvironment
(37). Studies from our laboratory have shown that cancer-asso-
ciated fibroblasts (CAFs) promote EMT of prostate cancer cells
through an ROS-dependent HIF-1α stabilization. This transition
is paralleled by the acquisition of stem-like and metastatic traits
of cancer cell (38, 39). In keeping, hypoxic senescent fibroblasts,
which display a CAF-like phenotype with increased lactate pro-
duction, are also capable of promoting EMT in prostate cancer
cells (40). Due to its important roles, HIF-1α is a potential target
for anticancer therapy. Among the molecules described to be able
to target HIF-1α (41), the compounds BAY87-2243, an inhibi-
tor of its activity and stability, and the antisense oligonucleotide EZN-2968 entered phase I clinical trials (42). Hypoxia induces
EMT not only via the transcriptional regulation of HIF-1α-
dependent EMT genes but also via epigenetic mechanism of
gene regulation (Figure 2). Indeed, HIF-1α is able to increase the
expression of ten–eleven translocation 1 (TET1), which catalyzes
the conversion of 5-methylcytosine to 5-hydroxymethylcytosine,
thus inducing DNA demethylation. Hypoxia-induced expression
of TET1 promotes EMT by complexing with HIF-1α and HIF-1β
and activating the transcriptional activation of HIF-dependent
EMT genes (43, 44). Glucose Transportersh The glucose transporters 1 (GLUT1) and GLUT3 are transcrip-
tionally induced by HIF-1, thus coupling the hypoxia with
increased glucose uptake and glycolysis (45). Interestingly,
overexpression of GLUT1 increases MMP-2 expression/activity FIGURE 2 | The metabolic features of epithelial-to-mesenchymal transition (EMT). Several components of the molecular pathways driving EMT have a clear
impact on cell metabolism and vice versa, resulting in a metabolic rewiring which sustains the EMT transcriptional program. EMT-committed cancer cells could rely
on an aerobic glycolytic metabolism or could shift toward the more efficient oxidative phosphorylation (OXPHOS), dependent on tumor type and/or tumor stage. Here, we have highlighted in red circles/ellipses EMT transcriptional factors that are affected by the metabolites reported in different tumor types. FIGURE 2 | The metabolic features of epithelial-to-mesenchymal transition (EMT). Several components of the molecular pathways driving EMT have a clear
impact on cell metabolism and vice versa, resulting in a metabolic rewiring which sustains the EMT transcriptional program. EMT-committed cancer cells could rely
on an aerobic glycolytic metabolism or could shift toward the more efficient oxidative phosphorylation (OXPHOS), dependent on tumor type and/or tumor stage. Here, we have highlighted in red circles/ellipses EMT transcriptional factors that are affected by the metabolites reported in different tumor types. March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org Targeting EMT-dependent Metabolic Reprogramming Morandi et al. Fiaschi et al. have demonstrated that silencing of MCT1 and
inhibition of MCT1-mediate lactate upload by prostate cancer
cells impairs tumor growth and lung micrometastasis formation
(66). Indeed, the blockade of lactate import and/or export is
an interesting target for alternative therapeutic approaches. The α-cyano-4-hydroxycinnamate (CHC), that targets MCTs,
has been used successfully in preclinical models without major
toxicity in vivo (67–69). The AstraZeneca MCT1 inhibitor
AZD3965 is in phase I/II clinical trials in the United Kingdom. Importantly, MCT4 silencing was shown to reduce tumor
cell migration in vitro and in vivo (62) and the AstraZeneca
MCT4 inhibitor (AZ93) has been reported as an highly effi-
cient and likely selective compound that is currently used in
preclinical studies. and invasiveness of cancer cell (46). Moreover, GLUT3 expres-
sion correlates with both EMT markers (i.e., vimentin, Snail, Slug,
ZEB1, ZEB2, Twist1) and glucose uptake in non-small cell lung
cancer (NSCLC) cells (47). In addition, enhanced GLUT1 and/or
GLUT3 expression is associated with poor prognosis in several
type of human tumors (48). Glucose Transportersh Numerous efforts have been made to
test the efficacy of GLUT inhibitors as anticancer drugs: WZB117
and the natural flavonoid silibinin are under preclinical studies
even if to our knowledge no specific GLUT1 inhibitors have been
identified. Lactate Since lactate is the by-product of Warburg-dependent metabo-
lism and increase of glycolysis is reported in several carcinomas,
high levels of lactate are positively correlate with cancer aggres-
siveness in several tumors (8, 49). In many types of tumor
a significant proportion of pyruvate is reduced into lactate, a
reaction catalyzed by LDH5. Elevated LDH5 expression cor-
relates with unfavorable prognosis in many human tumors and
silencing of LDH5 impairs tumor initiation and progression
(50–53). According to the pivotal role exerted by lactate in
the promotion of the metastatic process, lactate can induce
the upregulation of TGF-β2 expression (54), which in turn
induces a mesenchymal pro-migratory phenotype of glioblas-
toma cells and promotes MMP-2 activation, ECM remodeling,
and metastasis formation (55). It has also been reported that
lactate is able to induce the production of TGF-β1 in vivo
(56). In keeping, it has been shown that lactate administration
promotes in vitro migration of human breast carcinoma cells
as well as experimental lung metastases in mice intraperito-
neally injected with lactate (57). Lactate is also a sensor of
NAD+/NADH ratio, which is crucial for activity of the sirtuins
histone deacetylase, a class of NAD+-dependent enzymes that
possess either mono-ADP-ribosyltransferase or deacetylase
activity (58). High levels of NAD+/NADH ratio correlate with
energy stress and promote sirtuins activity. Some studies show
that SIRT1 induces both EMT and metastasis by suppressing
E-cadherin transcription (59, 60). Furthermore, our group has
recently shown that CAF-induced lactate production increases
activity of SIRT1 with consequent deacetylation/activation of
peroxisome proliferator-activated receptor gamma coactivator 1
alpha (PPARGC1A, also known as PGC-1α), a master regulator
of mitochondrial biogenesis. This activation is crucial for the
achievement of EMT. Accordingly, PGC-1α silencing abolishes
invasion and EMT of prostate cancer cells. This evidence shows
LDH as a possible therapeutic anticancer target. Several dif-
ferent classes of inhibitor have been synthesized, such as the
gossypol derivative FX-11, compound GNE-140, galloflavin,
and others (61), although none of these molecules are under
consideration in the clinic. Acidityh The regulation of lactate transport directly impacts on extracel-
lular acidity and, together with the increase of the glycolytic flux
lead to an unbalance in pH gradient across PM of cancer cells. Indeed, it is well established that tumor tissue are characterized
by low pHe and the role of tumor acidity in cancer invasion
and metastasis is well recognized (70). Indeed, low pHe leads
to apoptosis of normal tissue at the periphery of the tumor
while generating a positive selective pressure on cancer cells,
selecting subclones that are resistant to acidic environment. Furthermore, acidification promotes angiogenesis through the
enhanced expression of the vascular endothelial growth factor
(VEGF) (71) and interleukin-8 (72) induces adherens junctions
dissociation (73) and ECM degradation and remodeling (70, 74). Moreover, low pHe is crucial for the metastasization process
providing secretion/activation of idrolases, such as catepsin (75)
and MMPs (76). Acidic pHe was found to induce EMT in human
melanoma cells (77) and in Lewis lung carcinoma cells (78). Accordingly, exposure of tumor cells to an acidic environment
prior to tail vein injection increases lung colonization in a model
of experimental metastasis (79). Several molecules act as key players of cancer-associated
extracellular acidification, such as the sodium-proton exchange 1
(NHE1), carbonic anhydrase IX (CAIX), the sodium bicarbonate
transporter 1, and anion exchange 2. These molecules are possible
targets for therapy focused on disruption of acid–base balance in
cancer cells. In particular, CAIX, a membrane bound isoform of
CA transcriptionally regulated by HIF-1α, induces extracellular
acidification catalyzing the CO2 hydration and its overexpres-
sion is associated with increased metastasis and poor patients
survival in several cancers (80). Fiaschi et al. have shown that
activation of CAIX in CAFs leads to an extracellular acidification
that enhances MMP-2 and MMP-9 secretion, thereby driving
the stromal-induced EMT in prostate cancer cells. Moreover,
both genetic silencing and pharmacological inhibition of CAIX
are sufficient to impede EMT and CAF-induced invasion of
prostate cancer cells (81). Furthermore, CAIX silencing in
CAFs decreases prostate cancer cell tumor growth and lung
micrometastasis formation, indicating the enzyme as an ideal
target for anticancer therapy. At the moment, indisulam, an
inhibitor of CAIX is tested in clinical trials for the treatment
of NSCLC (82). Glycolytic Enzymesi induce in prostate cancer the oxidation and phosphorylation of
PKM2 prompting the nuclear translocation of the enzyme and
its association with HIF-1α and the differentially expressed in
chondrocyte-1, a transcriptional repressor, which downregulates
miR-205. This transcriptional complex induces EMT execution
(through the upregulation of ZEB2 and Snail1), as well as the
metabolic rewiring of cancer cells toward OXPHOS metabolism
(39). To further corroborate the link between PKM2 and EMT,
Hamabe et al. recently demonstrated that EMT stimulates nuclear
translocation of PKM2 and subsequent interaction with the TGF-
β-induced factor homeobox 2, which promotes histone H3K9
deacetylation and the subsequent downregulation of E-cadherin
expression (104). The PKM2 inhibitor TLN-232 was assessed
for cancer therapy in a phase II clinical trial but despite some
initial promising results the recruitment for a second phase II
trial has halted for legal reasons in 2010 (105). Other efforts have
been made to identify novel small-molecule inhibitors selective
for PKM2 (106). However, emerging data suggest that, at least
some tumors do not require PKM2 (107, 108), thus lowering the
interest about the targeting of PKM2 in cancer. Glycolysis is finely regulated and articulated in 10 different steps
and this could offer an array of potential targets to impair the EMT
process. Moreover, the presence of several potential targets allows
the possibility to interfere with more steps, either simultaneously
or sequentially. Blocking EMT and metabolic reprogramming
could therefore be a potential successful therapeutic approach. Hexokinase (HK2) is a HIF-1 target gene that controls the first
rate-limiting step of glycolysis and is often overexpressed in
cancer (83–86). It has been recently reported that breast CSCs
showed increased HK2 expression and glycolytic rate. Notably,
the glycolytic inhibitor, 2-deoxyglucose counteracts breast
cancer cells undergoing EMT in a dose dependent fashion (87,
88). Furthermore, lonidamine that targets mitochondria-bound
HK is now tested in phase II–III clinical trials (89). The HIF-1
dependent PGI/AMF (see above) acts as a pro-metastatic sign-
aling molecule, due to its ability to promote tumor migration,
invasion, and metastasis (90–92). Indeed, PGI/AMF overexpres-
sion leads to EMT achievement through NF-κB activation and
increased expression of Snail1, ZEB1, and ZEB2 (Figure 2),
downregulation of miR-200s with concomitant loss of E-cadherin
(93). In keeping with these observations, high PGI levels in the
serum positively correlate with metastases in colorectal, esopha-
geal squamous cell, and lung tumors (94–96). Glycolytic Enzymesi However, although
PGI has an established pro-tumorigenic role, to our knowledge,
no selective inhibitors are available for preclinical investigation. g
g
Moreover, the switch between PKM1 and PKM2 couples gly-
colysis to pentose phosphate pathway (PPP), a metabolic pathway
parallel to glycolysis that partially oxidizes glucose to pentoses
and generates NADPH. The PPP has been described to support
tumor cell proliferation (109) and to counteract ROS production,
due to the generation of NADPH. Indeed, several studies show
that PPP is necessary to handle the enhancement of ROS levels
in stress condition, such as anchorage independent growth and
anoikis (110, 111). Despite the PPP has a role in invasion, little is
known about its contribution to EMT. Indeed, scanty data have
shown that 6-phophogluconate dehydrogenase downregulation
reduces in vitro migration of lung carcinoma cells (112) and
high expression of the transketolase-like protein 1 isoform was
positively correlated with invasion and metastasis of several
carcinoma (113–116). Another potential target of the glycolytic cascade is aldolase
that converts the fructose-1,6-bisphosphate to glyceraldehyde-
3-phosphate and dihydroxyacetone phosphate. The aldolase
A isoenzyme is commonly overexpressed in various cancers
(97, 98) and its upregulation has been reported to induce lung
carcinoma cell migration and EMT by decreasing the expression
of E-cadherin and concomitantly increasing those of fibronectin
and vimentin (97). In addition to the aforementioned glycolytic enzymes, eno-
lase-1 has been recently proposed as a therapeutic target for
gene-based therapy. The overexpression of enolase-1 increases
glycolysis, proliferation, migration, and invasion in NSCLC
cells, a process that is in part mediated by the regulation of
EMT genes (99). Indeed, silencing of enolase-1 impaired EMT
execution as shown by Snail1 and N-cadherin downregulation
and the concomitant increase of E-cadherin expression (99). A role for enolase-1 in promoting migration and invasion has
been described also in endometrial carcinoma (100). Small-
molecule inhibitors of enolase are available but none of them
has entered in the clinical practice (101). Finally, the pyruvate
kinase (PK) enzyme has been recently emerged as an important
player in tumor progression. PK catalyzes the dephosphorylation
of phosphoenolpyruvate into pyruvate, resulting in ATP produc-
tion. Differentiated cells primarily express the M1 isoform of the
PK enzyme, whereas tumor cells often express the embryonic
M2 isoform, which can be expressed as a tetramer, in its active
form, or as a dimer with lower affinity for phosphoenolpyruvate. Monocarboxylate Transporters Key elements of the lactate shuttles are the MCTs, being MCT4
mainly involved in export of lactate and MCT1 in the uptake
of this catabolite. Several studies demonstrated that silencing or
inhibition of MCT1 and MCT4 are able to decrease the migra-
tion and invasion of several cancer cells (62–65). In particular, March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 5 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. Glycolytic Enzymesi Dimeric PKM2 (102) acts as a transcriptional coactivator of HIF-
1α in cancer cells, thus promoting glycolysis (103). Recently, our
group demonstrated a peculiar role of PKM2 in inducing EMT
in prostate cancer cells. Indeed, soluble factors secreted by CAF In addition, the hexosamine biosynthetic pathway (117), that
accounts for 2–5% of total glucose metabolism and is intimately
interconnected with the glycolytic pathway, shows a correlation
with EMT. Indeed, glucose that enters the glycolytic pathway
can be diverted to produce O-linked N-acetyl-glucosamine
(O-GlcNAc), a molecule that has a signaling and structural
role in the cells (117). It has been demonstrated that the addi-
tion of an O-GlcNAc motif at serine 112 prevented Snail1
O-phosphorylation-mediated degradation thus promoting its
stabilization (118), thereby providing a direct molecular link
between glucose metabolism and EMT. Frontiers in Oncology | www.frontiersin.org Impact of Amino Acids Metabolism on
EMT and Targeting Approaches Glucose dependency of fast-growing tumor is paralleled by the
higher amino acids requirements of these aggressive cancers. Indeed, targeting amino acid metabolism is now considered
a potential therapeutic approach in many cancer types (119). Particularly, the role of glutamine in cancer progression has
been extensively investigated due to the fact that glutamine is
an essential amino acid for proliferating tumor cells. Glutamine
can enter the oxidative TCA cycle, contribute to reductive March 2017 | Volume 7 | Article 40 6 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. carboxylation, and alter the NADPH production and redox bal-
ance. Glutaminase (GLS) catabolized glutamine conversion into
glutamate, which is then oxidized by glutamate dehydrogenase
(GDH) into α-ketoglutarate (α-KG), an intermediate of the TCA
cycle. Silencing of GLS1 or impairing glutamine metabolism were
able to counteract the induction of EMT mediated by growth
factors (e.g., TGF-β) in breast and colon cancers by increasing
Snail1-targeting miRNA expression and hence impacting on
Snail stability (Figure 2). Importantly, GLS1 silencing impaired
tumor growth and metastases formation (120). Moreover, since
GDH was reported to control tumor aggressiveness and therapy
response (121), Liu and colleagues hypothesized a link between
GDH expression/activity and EMT. Indeed, they reported an
association of GDH overexpression with metastasis formation
and poor prognosis in colorectal cancer patients. These effects
were demonstrated to be dependent, at least in vitro, on the
EMT induction mediated by GDH/STAT3 axis (122). However,
Aguilar and colleagues reported a differential contribution of
amino acid metabolism to EMT. Indeed, prostate cancer cells
that were selected to display a stable EMT phenotype uncoupled
to CSC behaviors were shown to (i) reduce the consumption
rate of glutamine and other ketogenic amino acids (i.e., leucine,
isoleucine, lysine, threonine, tyrosine, tryptophan, and phe-
nylalanine) and (ii) be less sensitive to glutamine metabolism
inhibition (123). The differences observed may be due to the fact
that the model used was peculiar and CSC and EMT programs
were uncoupled. Indeed, it is usually difficult to distinguish
whether the metabolic alterations observed in CSC are depend-
ent of EMT or CSC states, since recent studies have found that
the acquisition of CSC properties may occur in cancer cells
independently of EMT (124). autophagy-deficient mice showed reduced expression levels
of epithelial-related genes and increased of the mesenchymal
related suggesting a direct link between autophagy and EMT. Indeed, induction of autophagy degrades Snail1 thus inhibiting
the TGF-β-mediated EMT (133). Impact of Amino Acids Metabolism on
EMT and Targeting Approaches This inverse correlation was
also reported for glioblastoma cells (134). The diverging results
may be ascribed to the different models used; however the link
between autophagy and EMT seems to be conceivable. Recently,
this connection has been further reinforced by the demonstra-
tion that cadherin-6 (Figure 2), a type 2 cadherin that marks
cells that undergo EMT and correlates with metastatic ability
in papillary thyroid cancers, restrains autophagy, and promotes
reorganization of mitochondrial network (135). Autophagy tar-
geting via hydroxychloroquine has now moved into clinical trials
and a recent review (136) discusses how inducing or inhibiting
autophagy can be achieved in preclinical models. p
gy
p
Finally, amino acid-related metabolic pathways control
the EMT process via epigenetic modifications. Indeed, the
methionine synthesis pathway plays a pivotal role in controlling
promoters’ methylation and subsequent gene expression regula-
tion. DNA methyltransferases and histone methyltransferases
transfer a methyl group from the S-adenosyl methionine (SAM)
to DNA or to positive charged amino acidic residues of histones
with the formation of the by-product S-adenosyl homocysteine
(SAH) (137). Since SAH inhibits DNA methyltransferases and
histone methyltransferases, the ratio between SAM and SAH
is important for regulating DNA and histone methylation
(138). Threonine, glycine, and serine are the amino acids
that sustain SAM generation through the folate metabolism
(139). Threonine dehydrogenase converts threonine into gly-
cine and acetyl-CoA and together with methionine adenosyl
transferase are the key enzymes involved in SAM biosynthesis. Since methionine adenosyl transferase can be localized into
the nucleus, it provides an effective system for chromatin-
localized biosynthesis of metabolites to allow an adequate and
well-organized regulation of the histone and DNA methylation
processes. This mechanism can be exploited for targeting the
epigenetic regulation of EMT-related genes. Indeed, SAM-
competitive inhibitors and SAH hydrolase inhibitors have been
shown to inhibit the methyltransferase EZH2 thus impairing
the overall methylation histone status leading to transcription
reactivation of silenced genes, such as the EMT-related gene
E-cadherin (140, 141). Drugs that target glutamine transport into the cell or the
conversion to α-KG have been designed and tested. GLS
inhibitors have shown positive results in preclinical models;
bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl) ethyl sulfide
impairs cancer cells growth in vitro and in vivo (125, 126),
the GLS inhibitor CB-839 is effective against triple-negative
breast cancer (TNBC) and hematological tumors in preclinical
studies (127, 128) and is currently moved on to clinical trials
(129). Impact of Amino Acids Metabolism on
EMT and Targeting Approaches Therefore, targeting GLS could be an effective strategy to
block the EMT process and therefore impair invasive abilities of
aggressive cancer cells (130). gg
An alternative source of amino acids in cancer cell is due
to degradation and recycling of cellular components (e.g.,
autophagy). A role for protein degradation in controlling EMT
has been postulated recently by investigating how interfering
with lysosomal cathepsin proteases (i.e., lysosomal-dependent
protein turnover) affects TGF-β-induced EMT. Indeed, Kern
and colleagues found increased lysosome activity during EMT
of mammary epithelial cells derived from the MMTV-PyMT
mouse model and showed that cathepsin inhibitor E64d
was able to impair TGF-β-induced EMT and invasion (131). Moreover, Akalay and coworkers reported that tumor cells
undergoing EMT-induced autophagy regulate target recogni-
tion and lysis of cytotoxic T lymphocytes and may be exploited
for immunotherapeutic strategies to block immune escape
(132). However, conflicting results revealed that liver-specific Frontiers in Oncology | www.frontiersin.org Impact of Lipid Metabolism on EMT
and Targeting Approaches Despite glucose and glutamine alterations have been extensively
investigated in the context of cancer metabolism, increasing
evidence are now pointing to a pivotal role of alterations to
lipid-associated pathways (142, 143). Indeed, proliferating can-
cer cells are characterized by increased lipids and cholesterol
dependency, which can be fulfilled by either increasing the
upload of exogenous lipids or by over-activating their endog-
enous synthesis. The excess of lipids in cancer cells are stored in
lipid droplets (LDs) and high LDs and stored-cholesteryl ester
content in tumors are now considered as hallmarks of cancer
aggressiveness (144). March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 7 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. and metastases raises significant attention in considering FASN
targeting and/or other lipogenic enzymes as potential therapeutic
target for cancer treatment. Indeed, this therapeutic intervention,
besides inducing a temporary cancer growth inhibition could, in
the long-term, increase the risk of metastasis. Lipids encompass a vast class of biomolecules of unique
chemical structure. Among their multiple biological functions,
they contribute to cell compartmentalization, cell signaling,
protein trafficking, membrane organization, and energy storage
and production. Despite their established role in regulating a
variety of processes during cancer development (145–147), little
is known about the impact of lipid composition and metabolism
during EMT. Another strategy used by cancer cells to improve FAs avail-
ability for anabolic and catabolic purposes, is to increase the
expression of acyl-CoA synthetases (ACSLs), which converts
the long-chain FAs into acyl-CoA, critical for phospholipids and
TAGs synthesis, lipid modification of proteins as well as for FAs
β-oxidation (154). In colorectal cancer cells, the simultaneous
overexpression of ACSL1, ACSL4, and stearoyl-CoA desatu-
rase-1 (SCD), the rate-limiting enzyme converting saturated FAs
into monounsaturated FAs, induced EMT and increased cellular
migration and invasion (155). Furthermore, the combination
of low doses of pharmacological inhibitors of ACSL and SCD
selectively reduces cell viability of chemotherapy resistant colo-
rectal cells, with no effects on normal colonocytes. The clinical
relevance of these findings is emphasized in colorectal patients
with tumors displaying ACSL1/ACSL4/SCD simultaneous
overexpression, that show a poorer outcome and a higher risk
of relapse compared to other patients within the same clinico-
pathological stage. A lipidomic approach performed on prostate cancer cells that
underwent EMT following TNFα treatment revealed a significant
increase in the synthesis of triacylglicerols (TAGs), sustained by
a concomitant upregulation of fatty acid synthase (FASN) (148). Impact of Lipid Metabolism on EMT
and Targeting Approaches It is conceivable that the newly synthesized TAGs, stored into
LDs, may act not only as a reservoir of energy, but also of fatty
acids (FAs), which could serve for protein modification, cell
membrane remodeling, and the generation of pro-tumorigenic
signals which support EMT-induced cell motility. In keeping
with the increase of FASN during EMT, administration of the
FASN inhibitor osthole to the MCF7 breast cancer cells abol-
ishes cell scattering, EMT execution, migration, and invasion
induced by the hepatocyte growth factor (149). In addition, in
breast cancer cells undergoing EMT an increased expression of
FASN resulted in saturated FAs accumulation, which are then
relocated to the cell membrane and regulate lipid rafts organi-
zation, resulting in the activation of the EMT-inducer VEGF/
VEGFR2 signaling (150). In addition to the above discussed increase in TAGs, a reduc-
tion of C16:0 ceramide levels was also associated with EMT
(148, 156). A recent study documented that TGF-β-induced
EMT in human breast epithelial cells downregulates the expres-
sion of ceramide synthase 6 (CerS6), which produces C16:0
ceramide, further metabolized to C16:0 sphingolipids, such as
sphingomyelin and glycosphingolipids, both enriched in the
PM microdomains (156). CerS6 mRNA and protein levels are
also reduced in TNBC mesenchymal cells as compared with
non-TNBC epithelial cells. Downregulation of CerS6 in TNBC
patients correlates with increased PM fluidity, a required feature
to foster cell motility and metastatic spreading. Hence, CerS6 is
emerging as a novel EMT-regulated gene, whose reduction in
cancer correlates with a mesenchymal phenotype and, in turn,
with tumor aggressiveness and poor prognosis. Accordingly, it
is predictable that in patients with aggressive TNBC character-
ized by CerS6 downregulation, increasing the levels of C16:0
ceramide could impair the metastatic potential of this aggressive
breast cancer subtype. g
g (
)
Although the above-described reports deal with a supporting
role of FASN-driven lipogenesis for EMT and cell migration,
opposing observations are also described in the literature. The
exposure of NSCLC cells to the EMT-inducer TGF-β suppresses
the transcription of enzymes involved in lipogenesis, including
FASN and acetyl-CoA carboxylase (ACC) (151). These enzymes
are controlled by the carbohydrate-responsive element-binding
protein (ChREBP) and sterol regulatory element-binding pro-
teins (SREBPs). Snail1 mediated the suppression of the lipogenic
program by repressing SREBPs and ChREBP levels. In addition
to a decreased lipogenesis, TGF-β-induced EMT also promoted
higher oxygen consumption and elevated intracellular ATP. Frontiers in Oncology | www.frontiersin.org Impact of Lipid Metabolism on EMT
and Targeting Approaches A role of S1P in the modulation of other
MMPs which promote cell invasion, such as MMP-2 and MMP-
9, has also been proposed (163, 164). with a CSC phenotype, and an increase in metastatic potential. Interestingly, stabilization of lipid rafts is emerging as a target for
therapeutic strategies aimed to reduce the metastatic potential
driven by EMT in cancer. The stabilization of lipid raft domains
is a therapeutically attractive approach, since a number of
pharmacological and nutritional factors have been shown to
positively affect raft stability and function (168, 169). Among
them, the essential ω-3 polyunsaturated FA docosahexaenoic
acid (DHA) has been recently discovered to stabilize lipid raft
and to antagonize both EMT and the acquisition of stem-like
features (170). It suggests that the PM properties are subject
to dietary inputs and that long-term perturbations in lipid
metabolism (e.g., hypercholesterolemia, DHA supplementa-
tion) may profoundly affect tumor progression. In addition,
alkylphospholipid drugs (i.e., miltefosine and edelfosine) that
have been described as “dis-rafters” for their ability to affect raft
organization have potent anti-neoplastic activity (169). In parallel with the acquisition of an aggressive phenotype,
the EMT is marked by a profound rewiring of cell signaling
pathways, most of them originating from PM located media-
tors (165, 166). It is therefore conceivable that alterations in
the properties of the PM may impact on the overall signaling
network rearrangements associated with EMT. Recent evidence
showed that EMT is associated with a reorganization of the PM
and in particular with a destabilization of lipid raft domains
(Figure 3) (167). Alterations of the PM biophysical properties
are required to maintain the mesenchymal state, associated To reinforce the link between PM composition and EMT,
alterations in membrane fluidity, e.g., by modulating cholesterol
content (Figure 3), has been shown to induce or inhibit the
conversion toward a mesenchymal phenotype (167). Indeed, FIGURE 3 | Epithelial-to-mesenchymal transition (EMT) and lipid metabolism: interconnection and targeting. Several EMT-induced stimuli modulate the
content of different classes of lipids, with an impact on both cell metabolism and membrane composition. Some EMT inducers [i.e., tumor necrosis factor-α (TNF-α),
hepatocyte growth factor (HGF)] promote lipogenesis by stimulating fatty acid synthase (FASN) and leading to an increase in glicerophospholipids (which are
transferred to the cell membrane and regulate lipid rafts organization) and in triacylglycerol [stored in lipid drops as a reservoir of fatty acids (FAs) for catabolic or
anabolic purpose]. Impact of Lipid Metabolism on EMT
and Targeting Approaches Indeed, decreased FAs synthesis could divert acetyl-CoA into
the catabolic pathways, e.g., TCA cycle and OXPHOS, which
efficiently yield high amount of energy to support the elevated
cell migration required for metastatic spreading. In addition,
an increased availability of acetyl-CoA could foster histone
acetylation/activation of genes responsible for EMT engagement,
i.e., the transcription factors ZEB1, ZEB2, and Slug; the mesen-
chymal markers vimentin, N-cadherin, and fibronectin; and the
CSC markers Sox2 and Nanog (152). Notably, the expression of
the lipogenic enzymes FASN and ACC are downregulated in
circulating cancer cells, when compared to their primary cancer
counterpart (153). Interestingly, this metabolic switch toward
a catabolic signature is reversible upon withdrawal of EMT-
inducing stimuli. The functional role of this metabolic reversion is
that, when metastatic cells finally localize to secondary sites, they
have the potential to re-activate FASN expression, switching their
metabolism back to lipogenic pathways, in order to sustain rapid
cell proliferation and metastatic colonies formation. The obser-
vation that decreased lipogenesis might stimulate cell invasion Further evidence indicates that EMT is associated with
changes in sphingolipids metabolism. Guan and colleagues
reported that EMT is associated with the alteration of ganglioside
(i.e., sialic acid-containing glycosphingolipids) metabolism in
normal mouse mammary gland and bladder cancer (157). Gg4 is
one of the most relevant ganglioside involved in EMT execution,
whose content is reduced upon TGF-β-induced EMT. Gg4 was
reported to physically interact with key epithelial cell molecules,
such as E-cadherin and β-catenin, likely facilitating epithelial
intercellular adhesion via stabilization of the E-cadherin/β-
catenin complex at the cell surface (157). The modulation of
ganglioside pattern upon EMT is related, at least in part, to the
altered expression of genes encoding ganglioside metabolizing
enzymes, which are under the control of different transcription
factors involved in EMT (i.e., Snail1, Twist, ZEB1) (158–160). March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 8 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. Alongside the role of gangliosides, an involvement of sphin-
gosine-1-phosphate (S1P) in EMT has also been reported. In
hepatocellular carcinoma, where high levels of S1P in the serum
correlate with poor prognosis (161), S1P activates the PI3K/AKT
signaling pathway, resulting in MMP-7 upregulation. MMP-7
mediates the shedding of syndecan-1, a transmembrane heparan
sulfate proteoglycan that regulate cell–matrix interaction, with
a consequent increase in TGF-β1 production, a well-established
inducer of EMT (162). Impact of Lipid Metabolism on EMT
and Targeting Approaches On the other hand, other reports support a role for transforming growth factor-β (TGF-β)-derived Snail1 in the suppression of the lipogenic
program by repressing FASN expression, thus diverting acetyl-CoA toward catabolic pathways, such as the tricarboxylic acid (TCA) cycle. TGF-β-induced EMT also
correlates with a reduction in the pool of sphingolipids by (i) the downregulation of the expression of ceramide synthase, which results in the reduction of ceramide
levels; (ii) the lowering of ganglioside content due to the control of EMT-related transcription factors (i.e., SNAIL1, Twist, Zeb1) on the expression of genes encoding
ganglioside metabolizing enzymes. These events result in an increase of plasma membrane fluidity and destabilization of lipid rafts, to which significantly concurs
also the upregulation of cholesterol content. Pharmacological inhibitors of cholesterol synthesis (i.e., statins), the administration of alkyl phospholipid drugs (the
so-called “dis-rafters”), and nutritional factors, such as the ω-3 polyunsaturated FA docosahexaenoic acid, result in the stabilization of lipid raft and in the reduction
of membrane fluidity, thereby counteracting EMT, invasion, and the acquisition of stem-like features. FIGURE 3 | Epithelial-to-mesenchymal transition (EMT) and lipid metabolism: interconnection and targeting. Several FIGURE 3 | Epithelial-to-mesenchymal transition (EMT) and lipid metabolism: interconnection and targeting. Several EMT-induced stimuli modulate the
content of different classes of lipids, with an impact on both cell metabolism and membrane composition. Some EMT inducers [i.e., tumor necrosis factor-α (TNF-α),
hepatocyte growth factor (HGF)] promote lipogenesis by stimulating fatty acid synthase (FASN) and leading to an increase in glicerophospholipids (which are
transferred to the cell membrane and regulate lipid rafts organization) and in triacylglycerol [stored in lipid drops as a reservoir of fatty acids (FAs) for catabolic or
anabolic purpose]. On the other hand, other reports support a role for transforming growth factor-β (TGF-β)-derived Snail1 in the suppression of the lipogenic
program by repressing FASN expression, thus diverting acetyl-CoA toward catabolic pathways, such as the tricarboxylic acid (TCA) cycle. TGF-β-induced EMT also
correlates with a reduction in the pool of sphingolipids by (i) the downregulation of the expression of ceramide synthase, which results in the reduction of ceramide
levels; (ii) the lowering of ganglioside content due to the control of EMT-related transcription factors (i.e., SNAIL1, Twist, Zeb1) on the expression of genes encoding
ganglioside metabolizing enzymes. These events result in an increase of plasma membrane fluidity and destabilization of lipid rafts, to which significantly concurs
also the upregulation of cholesterol content. Impact of Lipid Metabolism on EMT
and Targeting Approaches Pharmacological inhibitors of cholesterol synthesis (i.e., statins), the administration of alkyl phospholipid drugs (the
so-called “dis-rafters”), and nutritional factors, such as the ω-3 polyunsaturated FA docosahexaenoic acid, result in the stabilization of lipid raft and in the reduction
of membrane fluidity, thereby counteracting EMT, invasion, and the acquisition of stem-like features. March 2017 | Volume 7 | Article 40 9 Frontiers in Oncology | www.frontiersin.org 9 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. the lipid compositions of epithelial or mesenchymal cells are
distinct (171) and when a given cell undergo the EMT process
it exhibits a significant increase in cholesterol content, which
strongly contributes, together with the reduction in ceramide
pool, to the enhancement in membrane fluidity. Accordingly,
cancer cells undergoing EMT are more sensitive to cholesterol
lowering drugs, such as statins, that have been reported to
deplete cholesterol content in mesenchymal cells, thereby
reducing PM fluidity and impairing cell motility and metastatic
potential (172). Increased membrane fluidity is an emerging
necessary feature of metastatic cancers that can be controlled by
many currently available drugs, offering a feasible therapeutic
opportunity to prevent cancer metastasis. A fascinating study
has recently identified, using an in silico drug screening, a series
of pharmacological compounds that can repress the metastatic
phenotype of cancer cells by inhibiting a gene expression sig-
nature associated with EMT. The compounds discovered using
this analysis, including previously acknowledged anti-metastatic
drugs, appeared to restrict the metastatic capacity through a
common mechanism, i.e., the ability to modulate the fluidity
of cell membranes (173). In keeping, the treatment of breast
cancer cell lines with some of these anti-metastatic agents (i.e.,
alprostadil, amitriptyline, haloperidol, and maprotiline, as well
as, salinomycin and thioridazine) reduced membrane fluidity,
resulting in EMT impairment, decreased cell motility, and stem
cell-like properties, culminating in the impairment of spontane-
ous metastasis in animal models, thus validating the in silico
analysis. The impact of fluidity on the metastatic behavior and
the strict correlation between membrane fluidity and cholesterol
content was further supported by the finding that in breast can-
cer patients, the overexpression of the cholesterol efflux channel
ABCA1 was associated with increased metastatic success and
was revealed in 41% of metastatic tumors (173). clinical analysis of human invasive breast cancers, the transcrip-
tion coactivator PGC-1α is overexpressed in invasive cancer cells
and stimulates mitochondrial biogenesis and OXPHOS during
their transit to metastatic sites (153). Impact of Lipid Metabolism on EMT
and Targeting Approaches PGC-1α silencing in cancer
cells significantly impaired their invasion ability and decreased
the frequency of metastasis without affecting cell proliferation
and primary tumor growth. Notably, the PGC-1α-induced meta-
bolic conversion toward OXPHOS is synergistically coupled to
a functional EMT program and although PGC-1α-induced
pathways are not essential for cancer cells during the acquisition
of a mesenchymal phenotype, both pathways concur and cor-
relate with the achievement of invasive and metastatic properties
(153). Accordingly, in both mouse melanoma and human breast
cancer models, an overloading of the mitochondrial electron
transport chain (ETC) was reported to promote an invasive
tumor phenotype associated with increased mitochondrial reac-
tive oxygen species (mtROS) generation (174, 175). However, this
study underlined that a dysfunctional mitochondrial activity and
a partial lowering of ETC activity strictly resembled ETC over-
loading, generating a similar pro-oxidant mitochondrial milieu. In both cases, superoxide generation induced mitochondrial Src
activation, which enhanced the expression of Pyk2, a FAK family
member protein tyrosine kinase that was previously reported
to promote EMT and migratory abilities (174, 176). Targeting
mtROS generation by treating with inhibitors of the ETC Complex
I activity (Ebselen) or using specific mitochondria-targeted
antioxidants [MitoTEMPO or MitoQ, a mitochondria-targeted
form of coenzyme Q10 currently tested in clinical trials (177)]
was sufficient to abolish metastasis formation in vivo (174). fi
(
)
Although mitochondrial superoxide acts as an inducer/sus-
tainer of EMT, an uncontrolled production of this pro-oxidant
can shift cell fate toward senescence or cell death (178, 179). To maintain ROS generation to levels that allow EMT execu-
tion, mitochondrial superoxide dismutase 2 (SOD2) have been
found upregulated upon TGF-β-mediated EMT, prompting the
conversion toward a CSC-like phenotype during the early stage
of EMT (180). ZEB2, but not ZEB1, together with NF-κB seems
to control the transcriptional regulation of SOD2. In addition, a
positive feedback loop has been described between SOD2 and
NF-κB during EMT in lung adenocarcinoma cells: NF-κB can
promote SOD2 transcriptional activation and concomitantly
SOD2 induces EMT activating the axis NF-κB/IκB kinase β
(IKKβ) (181). Frontiers in Oncology | www.frontiersin.org Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT Although the contribution of mitochondria to the pathogenesis
of cancer was underestimated for a long time, it is now established
that mitochondria have an essential role during tumorigenesis. Indeed, mitochondrial alterations are essential for the metabolic
rewiring that characterized a given cancer cells and play a role in
a series of additional cellular processes during the development
and progression of cancer.i We have discussed in details in the first paragraph how
aerobic glycolysis is extensively exploited by rapidly proliferating
cancer cells to meet their energetic demands and to accumulate
biosynthetic precursors. However, several reports support the
opposite notion, i.e., that the metabolic requirements of inva-
sive and metastatic cancer cells are characterized by enhanced
mitochondrial respiration and ATP generation. This review
does not discuss whether and what are the circumstances in
which glycolysis or OXPHOS are exploited by cancer cells for
promoting their survival and distant colonization. However, we
would like to emphasize that contradictory results described the
metabolic phenotype of invasive EMT-committed cancer cells
as glycolytic or OXPHOS addicted, not only in various tumor
types, but also within individual cancer types. As confirmed by Since an overloading of the respiratory mitochondrial metab-
olism is often associated with EMT and with the acquisition of
pro-invasive skills by cancer cells, drugs that inhibit OXPHOS
may be proposed as effective strategies to cope with the acquisi-
tion of a motile mesenchymal phenotype endowed with CSC-like
features and metastatic competency. Arsenic trioxide, which
interfere with OXPHOS by inhibiting the complex III of ETC and
metformin, acting on complex I are FDA-approved treatments
already used in the clinical practice (182, 183). Crucially, arse-
nic trioxide has been proposed for the clinical management of
relapsed or refractory acute promyelocytic leukemia (183), while
metformin particularly exerts anticancer activity in patients with
breast, endometrial, or prostate cancer (184, 185). March 2017 | Volume 7 | Article 40 10 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. on the epigenetic control of gene expression (195, 196). High
d-2HG levels paralleled by an altered epigenetic fingerprint
was also described in breast tumors characterized by c-Myc
overexpression and subsequent metabolic reprogramming,
independently of IDH mutations (197). The enantiomer l-2HG
(L2HG) can be generated by the non-canonical activity of malate
dehydrogenase and LDH and not by the IDH1-/IDH2-mutated
enzymes (198, 199). Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT These alterations identify a subset of patients
characterized by poor prognosis (187, 188) and sometimes
specific alterations of these mitochondrial enzymes are directly
linked to EMT induction. Consistent with the frequent metabolic rewiring toward
OXPHOS observed in invasive cancer cells, epithelial cells that
have activated the EMT program may increase the amount of
glucose diverted into the TCA cycle by regulating the pyruvate
dehydrogenase (PDH) complex, the gatekeeper of the cycle. In
particular, in lung cancer cells the execution of the EMT program
is accompanied by a downregulation of pyruvate dehydrogenase
kinase 4 (PDK4) expression levels, which results in an increase
of PDH activity (189). Downregulation of PDK4 is sufficient to
drive EMT and promotes erlotinib resistance in EGFR mutant
lung cancer cells. In keeping, analysis of human lung adenoma
tumor samples reveals PDK4 suppression as a predictor of poor
prognosis, consistent with its role during EMT (189).i The other two oncometabolites that exert their effects out-
side of the conventional metabolic network are succinate and
fumarate, two TCA cycle intermediates which reach elevated
concentrations in some tumors as a consequence of loss-of-
function mutations in the SDH complex or the FH, respectively
(203–205). Like d-2HG, succinate and fumarate have been found
to interfere with dioxygenase activity, underlining the oncogenic
role of both these metabolites in the inhibition of PHDs and
the subsequent stabilization of HIF-1 (206) and supporting the
notion that a general property of oncometabolites is the ability to
regulate epigenetics (191, 207). Besides the deregulation of the expression, specific cancer-
associated mutations of some enzymes of the TCA cycle, associ-
ated with an alteration of their catalytic function, have been also
recognized. Among the mutated enzymes, the most characterized
are those involved in the generation of the so-called oncometabo-
lites, including isocitrate dehydrogenase (IDH) (190), succinate
dehydrogenase (SDH) (191), and fumarate hydratase (FH) (191). Interestingly, fumarate possesses another unique property
linked to its chemical structure. Indeed, fumarate can covalently
bind to cysteine residues of proteins in a process called succina-
tion (208, 209). Several proteins are succinated in FH-deficient
cells, including aconitase (210), Kelch-like ECH-associated
protein 1 (208, 209), and glutathione (211). Notably, in a subset
of FH-deficient human renal cell carcinomas, fumarate at high
concentration directly bounds the antioxidant glutathione
both in vitro and in vivo to produce the metabolite succinated
glutathione (211). Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT Recently, in colorectal cancer cells an
elevation of both d-2HG and its enantiomer l-2HG has been
observed in the absence of IDH mutations and it is promoted
by glutamine anaplerosis. It has been reported that only d-2HG
is able to increase the histone H3 methylation pattern of the
ZEB1 promoter region, resulting in a direct upregulation of
ZEB1 and downregulation of miR-200, two key drivers of the
EMT process (200, 201). Clinical specimens with higher levels
of d-2HG associate with colonization of distant organs, sup-
porting the significant contribution of this metabolite in cancer
metastasis. Treatment strategies designed to reduce the levels of
d-2HG or inhibiting its downstream effects could be effective in
colorectal cancer. Interestingly, 2HG has been reported as one
of the few metabolites whose levels are reduced upon aspirin
administration, currently the most effective drug available for
chemoprevention of colorectal cancer (202), suggesting d-2HG
as a potential target for therapeutic intervention, with low risk
of side effects, since the physiological role of this metabolite is
not known. IDH-mutated tumors represent the perfect scenario
to test the specific targeting of tumor metabolism with minimal
interference with that of normal cells. Inhibitors of the mutant
form of IDH have been tested glioma and AML patients harbor-
ing IDH mutations (202).hf However, a recent study reported data that partially disagree
with the preeminent role of OXPHOS for metastatic potential of
cancer cells. In this study, the authors compared mRNA expres-
sion of metabolic genes in 20 different solid cancers to investigate
the link between metabolic transformation of cancer cells and
patient prognosis (186). Data revealed that the inhibition of
mitochondrial metabolism is a common signature for all the
cancer samples. The downregulation of OXPHOS-related genes
correlates with metastatic potential and poor clinical outcome
across several cancer types and it is associated with the presence
of EMT (186). These results support previous finding that partial
mitochondrial dysfunction increases metastatic potential of
cancer cells (174) and encourage future investigations on the role
of dysregulated metabolism during tumor progression, in order
to suggest more suitable targets for clinical intervention. One of the main metabolic pathways active within mitochon-
dria is the TCA cycle. Numerous enzymes of the TCA cycle show
mutation or deregulated expression pattern in both sporadic and
hereditary cancers. Frontiers in Oncology | www.frontiersin.org Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT Succinated glutathione acts as an alternative
substrate to glutathione reductase to decrease NADPH levels and
enhance mtROS and HIF-1 activation, two mandatory events
fostering EMT execution. Isocitrate dehydrogenase catalyzes the reversible conversion
of isocitrate into α-KG. Three isoforms of the enzymes have
been identified: the NADPH-dependent IDH1 and IDH2 and
the NADH-dependent IDH3. Mutations in the cytoplasmic
IDH1 and in the mitochondrial IDH2 mutations occur in vari-
ous human cancers, including gliomas, glioblastoma, and acute
myelogenous leukemias (AMLs) (192–194). The mutated forms
of the enzymes reduce α-KG into the d-2-hydroxyglutarate
(d-2HG). d-2HG levels are low in normal tissues but can reach
higher concentration levels (up to millimolar) in IDH1 or
IDH2-mutated tumors. High levels of d-2HG interfere with the
function of α-KG-dependent dioxygenases, including prolyl
hydroxylases (PHDs), essential for the regulation of the stability
of HIFs, Jumonji family of histone demethylases, and TET fam-
ily of DNA demethylases, whose inhibition inevitably impacts More recently, Sciacovelli and colleagues demonstrate a
fascinating role of fumarate, which accumulates in FH-deficient
renal cancers, in the epigenetic suppression of miR-200 through March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 11 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. the inhibition of Tet-mediated demethylation of a regulatory
region of the anti-metastatic miRNA cluster mir-200ba429 (212). Fumarate-dependent miR-200 downregulation leads to the
expression of EMT-related transcription factors and enhanced
aggressive features and it is associated with a poor clinical
outcome (212). Although it has been extensively reported that
FH deficiency is associated to fumarate-dependent epigenetic
deregulation, supporting the notion of FH as a tumor suppres-
sor, the molecular mechanisms by which FH gene expression is
controlled is not well clarified. It has been recently reported that
in nasopharyngeal carcinoma, the chromatin remodeling factor
lymphoid-specific helicase binds the FH promoter and recruits
the epigenetic silencer factor G9a to inhibit the transcription of
FH (213). The FH reduction promoted an unbalance in the TCA
intermediates, with a decrease in malate levels and a concomitant
increase in αKG amount. Deregulation of TCA metabolites in
nasopharyngeal carcinoma cells induces the recruitment of IKKα
to the promoter regions of genes involved in the EMT program,
causing the downregulation of E-cadherin and ZO-1 and the
parallel upregulation of the mesenchymal marker vimentin,
leading to EMT, migration and invasion in vitro, and increase
metastasis in vivo (213).f complex. Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT Additionally, several independent reports pointed to a
role of SDH5 as a tumor-suppressor gene and SDH5 deregulated
expression correlate with higher tumor incidence (226–228). An interesting study reported that SDH5 loss in lung cancers
promotes tumor aggressiveness and metastasis, hence providing
an additional functional support on the link between SDH5 and
EMT. In particular, SDH5 acts as a modulator of the glycogen
synthase kinase 3β (GSK-3β)/β-catenin signaling, since the
physical interaction between SDH5 and GSK-3β induces GSK-
3β dephosphorylation/activation, the subsequent decrease of
β-catenin nuclear accumulation and activation, thereby ultimately
inhibiting Wnt-β-catenin signaling (229). The downregulation of
SDH5 observed in lung cancers accounts for the SDH5-mediated
β-catenin stabilization and transcriptional activity, which in turn
induces Slug and Twist1 gene expression, thereby repressing
E-cadherin and contributing to EMT (229). g
Besides SDH5 repression, other mitochondrial signaling
pathways concur to regulate the GSK-3β/β-catenin pathway,
impacting on EMT. In lung cancer cells, the downregulation of
the TU translation elongation factor mitochondrial (TUFM), a
key factor in the translational expression of mitochondrial DNA,
is associated to the maintenance of the mesenchymal pheno-
type of cancer cells and to the acquisition of more aggressive
features. TUFM downregulation results in a reduced expression
of mtDNA, leading to mitochondrial dysfunction and cellular
stresses such as ATP deficiency and ROS production, which
trigger AMPK activation. It has been reported that activated
AMPK induces the phosphorylation of GSK-3β and increases
the nuclear accumulation of β-catenin, leading to the induction
of EMT and increased migration and metastatic competency of
lung cancer cells (230). These results provide a molecular link
between mitochondrial dysfunction and EMT, which is impli-
cated in lung cancer progression. Deregulation of different components of the SDH complex,
enzyme involved in both the ETC and the TCA cycle, have been
reported in several cancer types (214, 215). SDH is composed of
four essential subunits: the flavoprotein SDHA, the iron-sulfur
protein SDHB, and two units anchored to the mitochondrial
membrane SDHC and SDHD. Mutations in SDHx genes (encod-
ing SDH subunits) lead to succinate accumulation that inhibits
DNA demethylases (TET enzymes), leading to a global hyper-
methylation of DNA (191, 216, 217). Crucially, SDHx mutations
have been associated with EMT in hereditary pheochromocy-
toma and paraganglioma, as a result of epigenetic alterations
(218, 219). In particular, Aspuria et al. Role and Targeting of TCA Cycle
Intermediates and OXPHOS in EMT demonstrated that Sdhb
knockdown in mouse ovarian cancer cells resulted in a global
hypermethylation pattern that promotes EMT and induces a
metabolic reprogramming of the central carbon metabolism,
together with additional mitochondrial dysfunction, ultimately
leading to altered glucose and glutamine utilization (220). More
recently, Loriot and colleagues clarified how the succinate-
mediated epigenetic modulation impacts on EMT. First, using
transcriptome profiling of a large cohort of metastatic pheo-
chromocytomas and paragangliomas, the authors reported that
SDHB-malignant samples displayed a change in the expression
pattern of Twist1, Twist2, Snail1, and N-cadherin, all indications
of EMT program activation (221). Moreover, cytokeratin 19
(KRT19), a component of the intermediate filaments, already
implicated in conferring enhanced migratory and invasive
properties in squamous cell carcinomas, neuroblastomas, renal,
and breast cancers (222–225), has been identified as one of the
most significantly hypermethylated and downregulated gene in
SDHB-deficient mouse chromaffin cells, supporting the EMT-
dependent invasive properties and the metastatic behavior of
these neuroendocrine cancer cells (218). Malic enzyme 1 (ME1) is reported to be a major metabolic
enzyme, both localized in cytosol and mitochondria, catalyzing
the oxidative decarboxylation of malate into pyruvate, thus fueling
the TCA cycle, and concomitantly reducing NADP+ into NADPH
and contributing to macromolecular biosynthesis and protec-
tion from excessive oxidative stress. For its essential metabolic
roles, ME1 is ubiquitously expressed in different human tissues. However, up to now, there are no evidence about its oncological
functions and clinic significance. A significant upregulation of
ME1 has been associated to aggressive hepatocellular carcinoma
and to reduced overall survival and progression-free survival
in this tumor type (231). Silencing of ME1 in hepatocellular
carcinoma cells inhibits migration and invasion by blocking the
EMT program (i.e., restoring E-cadherin while downregulating
N-cadherin and vimentin expression) in a ROS-dependent way,
suggesting ME1 as a poor prognostic predictor for hepatocellular
carcinoma-bearing patients (231). Frontiers in Oncology | www.frontiersin.org REFERENCES 12. Lee SY, Jeon HM, Ju MK, Jeong EK, Kim CH, Yoo MA, et al. Dlx-2 is
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malignancy. Semin Radiat Oncol (2004) 14(3):267–74. FUNDING The work was funded by Fondazione Umberto Veronesi to AM,
Associazione Italiana Ricerca sul Cancro (AIRC) (grant 8797 to
PC), AIRC and Fondazione Cassa di Risparmio di Firenze (grant
19515 to PC and AM), Istituto Toscano Tumori (grant 0203607 to
PC), and Programma operativo regionale Obiettivo “Competitività
regionale e occupazione” della Regione Toscana cofinanziato dal
Fondo europeo di sviluppo regionale 2007-2013 (POR CReO FESR
2007-2013, grant to PC). CONCLUDING REMARKS Epithelial-to-mesenchymal transition is a specific program that
confers the cancer cell a series of stem-like properties and migra-
tory abilities thus increasing cancer aggressiveness. The hostile
microenvironment (i.e., hypoxia, low pH, and low glucose) and Besides the four subunits that constitute the SDH complex,
SDH5 has been identified as a mitochondrial protein necessary
for the flavination of SDHA and for the assembly of the SDH March 2017 | Volume 7 | Article 40 12 Targeting EMT-dependent Metabolic Reprogramming Morandi et al. the metabolic requirements of a fast-growing tumor sustain EMT
execution allowing cancer cells to bypass the nutrients and oxygen
supply limitation caused by the rapid primary cancer growth and
colonize secondary sites to secure the adequate support of energy
and nutrients. However, this is not simply a process to acquire
migration and invasion abilities but a more complex rewiring of
signaling, metabolic, genetic, and epigenetic networks that allow
differentiated epithelial cancer cells to revert into a more undiffer-
entiated state and stem cell functions. Additionally, it is plausible
to speculate that aggressive cancer cells are characterized by
the ability to undergoing EMT, acquire stem cell-like traits, and
rewire their metabolism to gain a plethora of strategies to survive
in different environmental conditions and/or in presence of
anticancer drugs. This plasticity reinforces the conclusion that “it
is not the fittest of the species (in this case of the tumor cell bulk) that
survives but the most adaptable.” The challenges we now face are
(i) the identification of distinct metabolic hallmarks exclusive to
cancer-associated EMT and (ii) the identification of therapeutic
window that allows EMT targeting in cancer patients. These are
key questions that need to be addressed to reduce the off target
effects caused by general metabolic therapies and importantly
to avoid the administration of anti-EMT compounds to cause a
MET in cells that have already left the primary site and may give
rise to metastases. However, none of the clinical trials using anti metabolic drugs, extensively reviewed in Ref. (6), have investi-
gated whether the effects exerted by these compounds have an
impact on the EMT process and therefore further confirmation
in preclinical studies and the design of large prospective clinical
trials should be planned. AUTHOR CONTRIBUTIONS AM, MT, PC, and EG were responsible for the conception,
design and drafting of the article, and final approval; agreed to be
accountable for all aspects of the work in ensuring that questions
related to the accuracy or integrity of any part of the work are
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2:21. doi:10.1186/2049-3002-2-21 Conflict of Interest Statement: The authors declare that the research was con-
ducted in the absence of any commercial or financial relationships that could
be construed as a potential conflict of interest. The reviewer MB and handling editor declared their shared affiliation, and the
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in neuroblastoma. BMC Cancer (2011) 11:66. doi:10.1186/1471-2407-11-66 Copyright © 2017 Morandi, Taddei, Chiarugi and Giannoni. This is an open-access
article distributed under the terms of the Creative Commons Attribution License
(CC BY). The use, distribution or reproduction in other forums is permitted,
provided the original author(s) or licensor are credited and that the original
publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with
these terms. 223. Morris MR, Gentle D, Abdulrahman M, Clarke N, Brown M, Kishida T,
et al. Functional epigenomics approach to identify methylated candidate
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98(2):496–501. doi:10.1038/sj.bjc.6604180 March 2017 | Volume 7 | Article 40 Frontiers in Oncology | www.frontiersin.org 19
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Inferências da Organização Mundial da Saúde na promoção da qualidade de vida: uma explanação contemporânea dos instrumentos WHOQOL
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PALAVRAS-CHAVE: Qualidade de vida. Organização Mundial da Saúde. WHOQOL. Programa de Pós-Graduação em Engenharia de Produção - PPGEP
Laboratório de Qualidade de Vida - LaQVida
Universidade Tecnológica Federal do Paraná – UTFPR
Ponta Grossa – PR – Brasil
v. 05, n. 04, out./dez. 2013, p. 19-25
DOI: 10.3895/S2175-08582013000400003 Programa de Pós-Graduação em Engenharia de Produção - PPGEP
Laboratório de Qualidade de Vida - LaQVida
Universidade Tecnológica Federal do Paraná – UTFPR
Ponta Grossa – PR – Brasil
v. 05, n. 04, out./dez. 2013, p. 19-25
DOI: 10.3895/S2175-08582013000400003 Inferências da Organização Mundial da Saúde na promoção da qualidade de
vida: uma explanação contemporânea dos instrumentos WHOQOL
Inferences of the World Health Organization in quality of life promotion: a
contemporary explanation of the WHOQOL instruments Bruno Pedroso
Universidade Estadual de Ponta Grossa – UEPG – Ponta Grossa – Brasil
brunops3@brturbo.com.br Claudia Tania Picinin
Universidade Tecnológica Federal do Paraná – UTFPR – Ponta Grossa – Brasil
claudiapicinin_adm@yahoo.com.br José Roberto Herrera Cantorani
Universidade Estadual de Campinas – UNICAMP – Campinas – Brasil
cantorani@yahoo.com.br Gustavo Luis Gutierrez
Universidade Estadual de Campinas – UNICAMP – Campinas – Brasil
glgutierrez@terra.com.br Gustavo Luis Gutierrez
Universidade Estadual de Campinas – UNICAMP – Campinas – Brasil
glgutierrez@terra.com.br REVISTA BRASILEIRA DE
QUALIDADE DE VIDA Inferências da Organização Mundial da Saúde na promoção da qualidade de
vida: uma explanação contemporânea dos instrumentos WHOQOL
Inferences of the World Health Organization in quality of life promotion: a
contemporary explanation of the WHOQOL instruments ABSTRACT ABSTRACT OBJECTIVE: The present study objectifies to identify the different WHOQOL instruments
presents in literature and also its structures and purposes. OBJECTIVE: The present study objectifies to identify the different WHOQOL instruments
presents in literature and also its structures and purposes. p
p
It is an exploratory and bibliographical research. METHODS: It is an exploratory and bibliographical research. RESULTS: We verified that from the initial project, that resulted in WHOQOL-100, a generic
instrument, the World Health Organization enlarged its objective and started to project new
instruments, addressed to the quality of life assessment of specific groups. It also structured an
abbreviated version of the original instrument. It was also done to some specific instruments. RESULTS: We verified that from the initial project, that resulted in WHOQOL-100, a generic
instrument, the World Health Organization enlarged its objective and started to project new
instruments, addressed to the quality of life assessment of specific groups. It also structured an
abbreviated version of the original instrument. It was also done to some specific instruments. CONCLUSIONS: WHOQOL-100 showed a very effective instrument in quality of life assessment
in a general form. Therefore, it was composed by facets that represent different domains of the life. However, due its generic structure, it was not an instrument with great effectiveness for specific
groups. Aware of this fact, World Health Organization developed instruments to reach efficiency in
quality of life assessment of children, older people, people living with HIV, people with disability
and people
ith chronic pain CONCLUSIONS: WHOQOL-100 showed a very effective instrument in quality of life assessment
in a general form. Therefore, it was composed by facets that represent different domains of the life. However, due its generic structure, it was not an instrument with great effectiveness for specific
groups. Aware of this fact, World Health Organization developed instruments to reach efficiency in
quality of life assessment of children, older people, people living with HIV, people with disability
and people with chronic pain. p
p
p
KEYWORDS: Quality of Life. World Health Organization. WHOQOL. 1. Introdução Não obstante à existência de diversos instrumentos de avaliação da qualidade de vida (QV),
é sabido que a maior parte destes são específicos, avaliando tal variável em função de um aspecto
particular. Em vista da carência de um instrumento que avaliasse a QV de forma global, a
Organização Mundial da Saúde (OMS) empenhou-se na construção do Instrumento de Avaliação da
Qualidade de Vida da Organização Mundial da Saúde (WHOQOL). O ponto de partida para a construção do WHOQOL foi conceituar o termo QV, sendo esta
entendida como “a percepção do indivíduo de sua posição na vida no contexto da cultura e sistema
de valores nos quais ele vive e em relação aos seus objetivos, expectativas, padrões e preocupações”
(FLECK et al., 1999, p. 20). Frente a esse conceito, a OMS inicia o projeto WHOQOL em 1993. O sucesso do
instrumento progenitor, o WHOQOL-100, instigou a OMS a construir diversos outros instrumentos
sob tal metodologia. O presente estudo objetiva identificar os diferentes instrumentos WHOQOL
existentes na literatura e também suas estruturas e propósitos. RESUMO OBJETIVO: O presente estudo tem como objetivo identificar os diferentes instrumentos
WHOQOL existentes na literatura e também suas estruturas e propósitos. É OBJETIVO: O presente estudo tem como objetivo identificar os diferentes instrumentos
WHOQOL existentes na literatura e também suas estruturas e propósitos. MÉTODOS: Trata-se de um estudo exploratório de natureza bibliográfica. RESULTADOS: Constatou-se que a partir do projeto inicial que resultou no WHOQOL-100 um OBJETIVO: O presente estudo tem como objetivo identificar os diferentes instrumentos
WHOQOL existentes na literatura e também suas estruturas e propósitos. MÉTODOS: Trata-se de um estudo exploratório de natureza bibliográfica. RESULTADOS: Constatou-se que a partir do projeto inicial, que resultou no WHOQOL-100, um
instrumento genérico, a Organização Mundial da Saúde ampliou seus objetivos e deu início a
projetos para novos instrumentos, voltados para a avaliação da qualidade de vida de grupos
específicos. Também estruturou uma versão abreviada do instrumento original. E o mesmo foi feito
para alguns instrumentos de caráter específico. MÉTODOS: Trata-se de um estudo exploratório de natureza bibliográfica. RESULTADOS: Constatou-se que a partir do projeto inicial, que resultou no WHOQOL-100, um
instrumento genérico, a Organização Mundial da Saúde ampliou seus objetivos e deu início a
projetos para novos instrumentos, voltados para a avaliação da qualidade de vida de grupos
específicos. Também estruturou uma versão abreviada do instrumento original. E o mesmo foi feito
para alguns instrumentos de caráter específico. p
g
p
CONCLUSÕES: O WHOQOL-100 se mostrou um instrumento muito eficaz na avaliação da
qualidade de vida de forma geral. Por isso este foi composto por facetas que representam os
diferentes domínios da vida. No entanto, em razão de sua estrutura genérica, não se constitui em um
instrumento com grande eficácia para grupos específicos. Ciente deste fato, a Organização Mundial
da Saúde desenvolveu instrumentos para alcançar eficiência na leitura da qualidade de vida de
crianças, idosos, pessoas soropositivas, pessoas com deficiência e pessoas com dor crônica. 2. Projeto WHOQOL O desenvolvimento do WHOQOL teve início em 1994 e foi realizado em 15 centros
simultaneamente, sediados em 14 países. Após o desenvolvimento do projeto WHOQOL, novos
centros foram incorporados e o WHOQOL-100 foi disponibilizado em mais de 20 idiomas. A
metodologia de desenvolvimento do WHOQOL foi seccionada em quatro grandes etapas:
clarificação do conceito de QV, estudo piloto qualitativo, desenvolvimento de um piloto e uma
aplicação de campo. Após a conclusão da elaboração do instrumento, foi estabelecida uma
metodologia que consistia na sua tradução, preparação do teste piloto, desenvolvimento das escalas
de respostas e administração do piloto (GRUPO WHOQOL, 1998). A partir das sugestões formuladas por cada um dos centros de desenvolvimento, totalizou-se
aproximadamente 1800 questões. Destas foram selecionadas 235 questões, seccionadas em 29
facetas, para o instrumento piloto do WHOQOL-100. Após a aplicação do teste piloto, foram
escolhidas as melhores questões para cada faceta, de forma a estabelecer a consistência interna e
validade discriminante do instrumento. Foram selecionadas 100 questões seccionadas em 24
facetas. As facetas foram agrupadas em seis grandes domínios (FLECK et al., 1999). O WHOQOL-100 é seccionado em 24 grupos de quatro questões cada, recebendo a
denominação de “facetas”. Por sua vez, o conjunto de facetas constitui um “domínio”. Ao contrário O WHOQOL-100 é seccionado em 24 grupos de quatro questões cada, recebendo a
denominação de “facetas”. Por sua vez, o conjunto de facetas constitui um “domínio”. Ao contrário 20 da composição das facetas, os seis domínios do WHOQOL-100 não são constituídos pelo mesmo
número de facetas, podendo variar de uma até oito facetas. Há ainda uma faceta que não está
inserida em nenhum domínio. Esta faceta, nomeada “Qualidade de vida global e percepção geral da
saúde”, aborda uma auto avaliação da QV, em que o indivíduo expressa o seu ponto de vista de
satisfação com a sua vida, saúde e QV. ç
Q
As questões que compõem o WHOQOL-100 não estão dispostas no questionário seguindo
uma sequência lógica por domínio ou por faceta, mas sim, por tipo de escala de respostas. 2. Projeto WHOQOL Os
domínios do WHOQOL-100 são: Físico, composto pelas facetas Dor e desconforto, Energia e
fadiga e Sono e repouso; Psicológico, composto pelas facetas Sentimentos positivos, Pensar,
aprender, memória e concentração, Autoestima, Imagem corporal e aparência e Sentimentos
negativos; Nível de independência, composto pelas facetas Mobilidade, Atividades da vida
cotidiana, Dependência de medicação ou de tratamentos e Capacidade de trabalho; Relações sociais,
composto pelas facetas Relações pessoais, Suporte social e Atividade sexual; Ambiente, composto
pelas facetas Segurança física e proteção, Ambiente no lar, Recursos financeiros, Cuidados de saúde
e sociais, Oportunidades de adquirir novas informações e habilidades, Participação em, e
oportunidades
de
recreação/lazer,
Ambiente
físico
e
Transporte
e;
Aspectos
espirituais/Religião/Crenças pessoais, composto pela faceta Espiritualidade/religião/crenças
pessoais. Todas as questões do WHOQOL-100 são fechadas. É utilizada uma escala de respostas do
tipo Likert de cinco pontos, variando entre 1 e 5. Esses extremos representam 0% e 100%,
respectivamente. Existem quatro tipos diferentes de escala de respostas: intensidade, avaliação,
capacidade e frequência. Tendo por base o WHOQOL-100, posteriormente foram construídos outros instrumentos de
avaliação da QV. Com o objetivo de disponibilizar um instrumento que demande menor tempo para
preenchimento e com características psicométricas satisfatórias, o Grupo WHOQOL desenvolveu a
versão abreviada do WHOQOL-100, o WHOQOL-bref. O WHOQOL-bref é composto por 26 questões, sendo duas questões sobre a auto avaliação
da QV e 24 questões representando cada uma das facetas do WHOQOL-100. Para a composição das
questões do WHOQOL-bref foi selecionada a questão de cada faceta que apresentava a maior
correlação com o escore médio de todas as facetas (THE WHOQOL GROUP, 1996). Após a seleção das questões, foi realizada uma análise para verificar se estas, de fato,
representavam as facetas correspondentes. Em seis facetas a questão selecionada foi substituída por
outra questão da faceta correspondente, pois, sob o viés de especialista, havia outra questão que
melhor definisse essas seis facetas (THE WHOQOL GROUP, 1996). As facetas pertencentes ao
domínio Nível de independência foram incorporadas ao domínio Físico, assim como a faceta
pertencente ao domínio Aspectos espirituais/Religião/Crenças pessoais foi incorporada ao domínio
Psicológico. Assim, o WHOQOL-bref é composto por quatro domínios: Físico, Psicológico,
Relações sociais e Meio-ambiente. 2. Projeto WHOQOL No intento de construir um instrumento de avaliação da QV para o Projeto EUROHIS, cujo
objetivo era a elaboração de questionários e entrevistas para avaliação de indicadores da área da
saúde no continente europeu, fora convencionado que se tentaria adaptar um instrumento já
existente para uma versão com um pequeno número de questões, para utilização em pesquisas em
nível nacional, tais quais os censos demográficos (POWER, 2003). O EUROHIS-QOL, posteriormente batizado de WHOQOL-8, é composto por oito questões
derivadas empiricamente do WHOQOL-bref, sendo considerado uma alternativa viável para
utilização em grandes populações ou em situações em que se demande de pouco tempo para o
preenchimento do questionário (AUSTRALIAN WHOQOL FIELD CENTRE, 2006). 3. Prossecução e instrumentos específicos Com o intuito de criar um instrumento de avaliação da QV direcionado para portadores do
HIV, pesquisadores da UNAIDS e da OMS realizaram estudos em portadores do HIV em nove 21 21 países. O resultado desse estudo foi o instrumento WHOQOL-HIV, um suplemento ao WHOQOL-
100 específico para pessoas portadoras do HIV ou AIDS. países. O resultado desse estudo foi o instrumento WHOQOL-HIV, um suplemento ao WHOQOL-
100 específico para pessoas portadoras do HIV ou AIDS. O WHOQOL-HIV avalia a QV a partir de seis domínios e 29 facetas. Os domínios e as
facetas são as mesmas do WHOQOL-100, com o acréscimo de cinco facetas específicas para
portadores do HIV. As facetas específicas para os portadores do HIV, assim como as facetas do
WHOQOL-100, são compostas por quatro questões (O'CONNELL et al., 2004). As facetas adicionais do WHOQOL-HIV se inserem nos domínios já existentes do
WHOQOL-100, sendo o Domínio Físico acrescido da faceta Sintomas de pessoas vivendo com
HIV/AIDS (PVHAs), o Domínio Psicológico acrescido da faceta Inclusão social e o Domínio
Aspectos espirituais/Religião/Crenças pessoais acrescido das facetas Perdão e culpa, Preocupações
sobre o futuro e Morte e morrer. Sob a mesma justificativa da elaboração do WHOQOL-bref, o Grupo WHOQOL
desenvolveu uma versão abreviada do WHOQOL-HIV. O WHOQOL-HIV-bref é baseado no
WHOQOL-bref, da forma que cada faceta é representada por uma única questão, somado a duas
questões gerais da QV. As 26 questões do WHOQOL-bref se repetem no WHOQOL-HIV-bref,
sendo estas acrescidas de cinco questões que representam as facetas adicionais do WHOQOL-HIV
(THE WHOQOL-HIV GROUP, 2002). Ao contrário do que ocorre no WHOQOL-bref, as facetas pertencentes aos domínios nível
de independência e Aspectos espirituais/Religião/Crenças pessoais não são incorporadas aos
domínios Físico e Psicológico, assumindo, assim, a mesma configuração dos domínios presentes no
WHOQOL-100 e WHOQOL-HIV. Tendo em vista que algumas questões dos instrumentos WHOQOL-100 e WHOQOL-bref
não se aplicam às crianças e/ou influenciam a QV destas por meio de outro foco, surge a proposta
da construção de uma versão dos instrumentos WHOQOL com direcionamento para crianças. Inicialmente batizado como C-QOL, tendo em vista a sua derivação do WHOQOL-100,
posteriormente recebeu a denominação de WHOQOL-Children (LUZNY, 2011). p
ç
O WHOQOL-Children contempla os seis domínios presentes no WHOQOL-100, acrescido
do domínio adicional Direitos políticos das crianças. 3. Prossecução e instrumentos específicos As facetas que constituem os domínios já
existentes no WHOQOL-100 são as mesmas, com exceção da faceta Dependência de drogas, que
foi acrescida ao domínio Nível de independência e da faceta Atividade sexual, que foi excluída do
domínio Relações sociais. Com relação ao domínio adicional do WHOQOL-Children, as facetas
que compõem esse domínio são Direito de falar e ser ouvido e Direito à identidade e cidadania. Cada faceta é representada por duas questões, totalizando 54 questões. Ao contrário dos
demais instrumentos WHOQOL, as questões do WHOQOL-Children estão agrupadas por faceta e
domínio (JIROJANAKUL; SKEVINGTON; HUDSON, 2003). Almejando adaptar o WHOQOL para a aplicação em adultos idosos, o Grupo WHOQOL
desenvolveu um módulo adicional denominado WHOQOL-OLD. Ao contrário do WHOQOL-HIV
e WHOQOL-HIV-bref, o WHOQOL-OLD não suplementa o WHOQOL-100 ou o WHOQOL-bref. O WHOQOL-OLD é um módulo complementar, mas que deve ser aplicado em conjunto com o
WHOQOL-100 ou o WHOQOL-bref (POWER et al., 2005). O WHOQOL-OLD é composto por 24 questões seccionadas em seis facetas, formando um
domínio adicional questões relacionadas ao envelhecimento. As facetas adicionais do WHOQOL-
OLD são: Funcionamento dos sentidos, Autonomia, Atividades passadas, presentes e futuras,
Participação social, Morte e morrer e Intimidade (CHACHAMOVICH et al., 2008). Em estudos posteriores ao desenvolvimento do WHOQOL-100 e WHOQOL-bref, foi
constatado que o domínio Aspectos espirituais/Religião/Crenças pessoais nestes instrumentos era
conceitualmente e empiricamente insuficiente. Frente a essa fragilidade, o Grupo WHOQOL
pautou-se no desenvolvimento de um estudo para avaliar a forma com a qual o domínio em exame
está relacionado com a QV (THE WHOQOL-SRPB GROUP, 2006). O WHOQOL-SRPB não é um instrumento para avaliar a espiritualidade, mas sim, um
instrumento que contempla, de forma ampliada, o constructo Aspectos espirituais/Religião/Crenças 22 22 pessoais, representado no WHOQOL-100 e WHOQOL-bref de forma reduzida (THE WHOQOL-
SRPB GROUP, 2006). Nessa perspectiva, o WHOQOL-SRPB suplementa o WHOQOL-100 com oito facetas
adicionais ao domínio Aspectos espirituais/Religião/Crenças pessoais. Os demais domínios do
WHOQOL-100 permanecem inalterados. As facetas adicionais do WHOQOL-SRPB são: Conexão
com o ser ou força espiritual, Sentido na vida, Admiração, Totalidade e integração, Força espiritual,
Paz interior, Esperança e otimismo e Fé. p
ç
Na tentativa de mensurar a QV de adultos portadores de deficiência física e/ou intelectual,
recentemente fora desenvolvido um módulo adicional ao WHOQOL-bref direcionado para tal
público. 3. Prossecução e instrumentos específicos O WHOQOL-DIS é composto por 13 questões: uma geral – contemplando o impacto da
deficiência – e 12 específicas, contemplando os seguintes aspectos: Discriminação, Proteção,
Perspectivas futuras, Controle da vida, Tomada de decisões, Autonomia, Comunicação, Aceitação
social, Respeito, Interação na sociedade, Inclusão social e Capacidade pessoal (POWER et al.,
2010). Fora desenvolvida, também, uma versão do WHOQOL-DIS direcionado para os
responsáveis pelos cuidados da pessoa portadora da deficiência intelectual. A versão do WHOQOL-
DIS direcionada para deficientes intelectuais possui uma escala de resposta diferenciada dos demais
instrumentos WHOQOL, sendo esta do tipo Likert de três pontos. Nesse caso, o WHOQOL-bref
também deve ser aplicado com a adaptação para tal escala. Sob a mesma justificativa que instigou o desenvolvimento do WHOQOL-SRPB – a
insuficiência das questões para avaliar um determinado aspecto da QV –, fora desenvolvido o
instrumento WHOQOL-Pain. Tal qual o WHOQOL-OLD, o WHOQOL-Pain é constituído pelo WHOQOL-100 acrescido
do módulo adicional direcionado às pessoas que sofrem de dor crônica. Este foi desenvolvido pelo
Centro de Estudo de Qualidade de Vida da Universidade de Bath, na Inglaterra. Não foi
desenvolvido paralelamente em outros centros nem traduzido para outros idiomas. O domínio Dor e
desconforto é composto por 16 questões seccionadas em quatro facetas: Alívio da dor, Raiva e
frustração, Vulnerabilidade/medo/preocupação e Incertezas (MASON; SKEVINGTON; OSBORN,
2010). Referências AUSTRALIAN WHOQOL FIELD CENTRE. Use the World Health Organization Quality of Life
(WHOQOL) Instruments. Centre for Participant Report of University of Melbourne,
Melbourne,
2006. Department
of
Psychiatry. Disponível
em:
<http://www.psychiatry.unimelb.edu.au/centres-units/cpro/whoqol/use-whoqol.html>. Acesso em
23 abr. 2012. CHACHAMOVICH, E. et al. Desenvolvimento do instrumento WHOQOL-OLD. In: FLECK, M. P. A. et al. A avaliação de qualidade de vida: guia para profissionais da saúde. Porto Alegre:
Artmed, 2008. p. 102-111. FANG, J. et al. Development of short versions for the WHOQOL-OLD module. The
Gerontologist, Oxford, v. 52, n. 1, p. 66-78, Jan./Feb. 2012. FLECK, M. P. A. et al. Desenvolvimento da versão em português do instrumento de avaliação de
qualidade de vida da OMS (WHOQOL-100). Revista Brasileira de Psiquiatria, São Paulo, v. 21,
n.1, p.19-28, jan./mar. 1999. GRUPO WHOQOL. Versão em português dos instrumentos de avaliação da qualidade de vida
(WHOQOL). Faculdade de Medicina da UFRGS, 1998. Departamento de Psiquiatria. Disponível
em: <http://www.ufrgs.br/psiq/whoqol1.html>. Acesso em: 13 abr. 2008. INTERNATIONAL SOCIETY FOR QUALITY OF LIFE RESEARCH. 2010 conference abstracts. Quality of Life Research, Amsterdam, v. 19, n. Supplement 1, p. 1-144, Oct. 2010. JIROJANAKUL, P.; SKEVINGTON, S. M.; HUDSON, J. Predicting young children’s quality of
life. Social Science and Medicine, New York, v. 57, n. 7, p. 1277-1288, Oct. 2003. LUZNY, J. Quality of life in mentally ill people. In: UEHARA, T. (Org.). Psychiatric disorders:
worldwide advances. Rijeka: Intech, 2011, p. 151-162. MASON, V. L.; SKEVINGTON, S. M.; OSBORN, M. Assessing the properties of the WHOQOL-
pain: quality of life of chronic low back pain patients during treatment. The Clinical Journal of
Pain, Philadelphia, v. 26, n. 7, p. 583-592, Sep. 2010. O'CONNELL, K. et al. WHOQOL-HIV for quality of life assessment among people living with
HIV and AIDS: results from a field test. AIDS Care, Abingdon, v. 16, n. 7, p. 882-889, Oct. 2004. POWER, M. J. Development of a common instrument for quality of life. In: NOSIKOV, A.;
GUDEX, A. (Org.). EUROHIS: developing common instruments for health surveys. 1. ed.,
Amsterdam: IOS Press, 2003. p. 145-163. POWER, M. J et al. Development of the WHOQOL-Old module. Quality of Life Research,
Amsterdam, v. 14, n. 10, p. 2197-2214, 2005. POWER, M. J. et al. Development of the WHOQOL disabilities module, Quality of Life Research,
Amsterdam, v. 19, n. 4, p. 571-584, may 2010. SKEVINGTON, S. M.; GUNSON, K. S.; O'CONNELL, K. A. 4. Considerações finais Transcorrido mais de uma década desde o surgimento do WHOQOL-100 e WHOQOL-bref,
estes permanecem sendo utilizados em grande escala. Atualmente os instrumentos WHOQOL são
disponibilizados em mais de 50 idiomas. O rigor metodológico utilizado no processo de construção e validação dos instrumentos
WHOQOL possibilita a obtenção de características psicométricas satisfatórias, perfazendo com que
os resultados das aplicações de tais instrumentos sejam válidos, consistentes e fidedignos à
população estudada. Na conferência anual de 2010 da Sociedade Internacional para Pesquisas em Qualidade de
Vida (ISOQOL), fora explicitado que a versão abreviada do WHOQOL-SRPB está em
desenvolvimento, e que, futuramente, será construída a versão abreviada do WHOQOL-Pain
(INTERNATIONAL SOCIETY FOR QUALITY OF LIFE RESEARCH, 2010). Segue em
andamento, também, o projeto para a construção da versão abreviada do WHOQOL-OLD (FANG
et al., 2012), e, recentemente, foi anunciada a versão preliminar da variante abreviada do
WHOQOL-SRPB, o WHOQOL-SRPB-bref (SKEVINGTON; GUNSON; O'CONNELL, 2012). A prospecção é de que novos instrumentos WHOQOL sejam criados. Até o momento foram
desenvolvidos 12 instrumentos. Independentemente do surgimento de outros instrumentos, é
pertinente ressaltar que a OMS proferiu, e, continua proferindo, por meio dos instrumentos
WHOQOL, uma contribuição ímpar na promoção da QV. Na conferência anual de 2010 da Sociedade Internacional para Pesquisas em Qualidade de
Vida (ISOQOL), fora explicitado que a versão abreviada do WHOQOL-SRPB está em
desenvolvimento, e que, futuramente, será construída a versão abreviada do WHOQOL-Pain
(INTERNATIONAL SOCIETY FOR QUALITY OF LIFE RESEARCH, 2010). Segue em
andamento, também, o projeto para a construção da versão abreviada do WHOQOL-OLD (FANG
et al., 2012), e, recentemente, foi anunciada a versão preliminar da variante abreviada do
WHOQOL-SRPB, o WHOQOL-SRPB-bref (SKEVINGTON; GUNSON; O'CONNELL, 2012). A prospecção é de que novos instrumentos WHOQOL sejam criados. Até o momento foram
desenvolvidos 12 instrumentos. Independentemente do surgimento de outros instrumentos, é
pertinente ressaltar que a OMS proferiu, e, continua proferindo, por meio dos instrumentos
WHOQOL, uma contribuição ímpar na promoção da QV. A prospecção é de que novos instrumentos WHOQOL sejam criados. Até o momento foram
desenvolvidos 12 instrumentos. Independentemente do surgimento de outros instrumentos, é
pertinente ressaltar que a OMS proferiu, e, continua proferindo, por meio dos instrumentos
WHOQOL, uma contribuição ímpar na promoção da QV. 23 Recebido em: 25 set. 2013.
Aprovado em: 15 nov. 2013. THE WHOQOL GROUP. WHOQOL-bref: introduction, administration, scoring and generic
version of assessment. Geneva: World Health Organization, 1996. Referências Introducing the WHOQOL-SRPB
BREF: developing a short-form instrument for assessing spiritual, religious and personal beliefs
within quality of life. Quality of Life Research, Amsterdam, v. 22, n. 5, p. 1073-1083, jun. 2013. 24 THE WHOQOL GROUP. WHOQOL-bref: introduction, administration, scoring and generic
version of assessment. Geneva: World Health Organization, 1996. THE WHOQOL-HIV GROUP. WHOQOL-HIV instrument user’s manual. Geneva: World
Health Organization, 2002. THE WHOQOL-SRPB GROUP. A cross-cultural study of spirituality, religion, and personal beliefs
as components of quality of life. Social Science and Medicine, New York, v. 62, n. 6, p. 1486-
1497, mar. 2006. THE WHOQOL-SRPB GROUP. A cross-cultural study of spirituality, religion, and personal beliefs
as components of quality of life. Social Science and Medicine, New York, v. 62, n. 6, p. 1486-
1497, mar. 2006. 25 25
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Habitat-specific differences alter traditional biogeographic patterns of life history in a climate-change induced range expansion
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PloS one
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cc-by
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RESEARCH ARTICLE Habitat-specific differences alter traditional
biogeographic patterns of life history in a
climate-change induced range expansion Megan E. Riley1*, Blaine D. Griffen1,2
1 Department of Biological Sciences, University of South Carolina, Columbia, SC, United States of America,
2 School of the Earth, Ocean, and Environment, Marine Science Program, University of South Carolina,
Columbia, SC, United States of America Megan E. Riley1*, Blaine D. Griffen1,2 * riley.megan.e@gmail.com * riley.megan.e@gmail.com a1111111111
a1111111111
a1111111111
a1111111111
a1111111111 Abstract Range shifts and expansions resulting from global climate change have the potential to cre-
ate novel communities with unique plant-animal interactions. Organisms expanding their
range into novel biotic and abiotic environments may encounter selection pressures that
alter traditional biogeographic patterns of life history traits. Here, we used field surveys to
examine latitudinal patterns of life history traits in a broadly distributed ectotherm (mangrove
tree crab Aratus pisonii) that has recently experienced a climate change-induced range
expansion into a novel habitat type. Additionally, we conducted laboratory and field experi-
ments to investigate characteristics associated with these life history traits (e.g. fecundity,
offspring quality, and potential selection pressures). We compared these characteristics in
native mangrove habitats in which the species has historically dwelled and novel salt marsh
habitats into which the species has recently expanded its range. Consistent with traditional
biogeographic concepts (i.e. Bergmann’s clines), size at maturity and mean body size of
reproductive females increased with latitude within the native habitat. However, they de-
creased significantly in novel habitats at the highest latitudes of the species’ range, which
was consistent with habitat-specific differences in both biotic (predation) and abiotic (tem-
perature) selection pressures. Although initial maternal investment (egg volume and weight)
did not differ between habitats, fecundity was lower in novel habitats as a result of differ-
ences in size at reproduction. Offspring quality, as measured by larval starvation resistance,
was likewise diminished in novel habitats relative to native habitats. These differences in off-
spring quality may have enduring consequences for species success and persistence in
novel habitats. Life history characteristics such as those investigated here are fundamental
organismal traits; consequently, understanding the potential impacts of climate change
responses on latitudinal patterns of these traits is key to understanding climate change
impacts on natural systems. Introduction In recent decades, changes in global climate trends have led to radical alterations in natural
ecosystems. A number of systems have experienced losses in biodiversity as well as changes in
the composition and dynamics of communities [1–2], both of which are predicted to continue
in the future [3–4]. One of the most common responses to a warming climate is a shift in the
range or distribution of species. Because the geographic ranges of most plants and animals are
limited to some extent by climatic factors (i.e. temperature, light, precipitation) [3], changes in
global climate trends have caused many species to shift or expand their distributions to include
higher altitudes and more poleward latitudes [5–6]. For example, species whose ranges are
constrained by the cold are expanding their distributions poleward in response to warming
temperatures, with terrestrial species expanding at an average rate of 16.9 km each decade [5]
and marine species expanding at an average rate of 72 km each decade [6]. Such range expan-
sions have been widely documented in an extensive array of taxonomic groups, including
insects [7–8], plants [9], mammals [10], and marine invertebrates [11]. Because different taxonomic groups display variation in their sensitivity and response to cli-
mate changes, species often shift their ranges at different speeds [12]. These differences in the
speed of range expansion have the potential to lead to spatial mismatches between previously
interacting species [12–14]. When rapidly expanding species expand their range faster than
their neighbors, this process creates novel ecosystems with unique community assemblages
and plant-animal interactions [3, 12]. The resulting novel communities often represent a suite
of new selection pressures that may prompt rapid changes in plastic life history traits in range-
expanding organisms [15–17]. Because many organisms already exist across historically broad
ranges in which they display latitudinal clines in life history traits, selection pressures associ-
ated with their range expansion may alter existing biogeographic life history patterns [17]. For example, body size is among the most fundamental life history characteristics of an
organism, influencing key traits such as individual fitness and population dynamics [18]. Biol-
ogists have long documented variations in both body size and size at maturity across latitude,
including increases in body size and size at maturity with latitude (see [19] for a list of reviews)
Bergmann [20] originally addressed this topic by noting that endotherms in cooler tempera-
tures (i.e. Range expansion alters latitudinal life history trends Competing interests: The authors have declared
that no competing interests exist. OPEN ACCESS Citation: Riley ME, Griffen BD (2017) Habitat-
specific differences alter traditional biogeographic
patterns of life history in a climate-change induced
range expansion. PLoS ONE 12(5): e0176263. https://doi.org/10.1371/journal.pone.0176263 Citation: Riley ME, Griffen BD (2017) Habitat-
specific differences alter traditional biogeographic
patterns of life history in a climate-change induced
range expansion. PLoS ONE 12(5): e0176263. https://doi.org/10.1371/journal.pone.0176263
Editor: Stefan Lo¨tters, Universitat Trier, GERMANY
Received: May 11, 2016
Accepted: April 7, 2017
Published: May 4, 2017
Copyright: © 2017 Riley, Griffen. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which
permits unrestricted use, distribution, and
reproduction in any medium, provided the original
author and source are credited. Editor: Stefan Lo¨tters, Universitat Trier, GERMANY
Received: May 11, 2016
Accepted: April 7, 2017
Published: May 4, 2017 Editor: Stefan Lo¨tters, Universitat Trier, GERMANY
Received: May 11, 2016
Accepted: April 7, 2017
Published: May 4, 2017
Copyright: © 2017 Riley, Griffen. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which
permits unrestricted use, distribution, and
reproduction in any medium, provided the original
author and source are credited. Copyright: © 2017 Riley, Griffen. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which
permits unrestricted use, distribution, and
reproduction in any medium, provided the original
author and source are credited. Data Availability Statement: All data files are
available online from the National Science
Foundation database (http://www.bco-dmo.org/
project/562104). Funding: This research was supported by a
National Science Foundation grant to BG (OCE-
1129166), a Smithsonian Institution Predoctoral
Fellowship to MR, a Slocum-Lunz Foundation grant
to MR, and a University of South Carolina SPARC
Graduate Research Fellowship to MR. The funders
had no role in study design, data collection and
analysis, decision to publish, or preparation of the
manuscript. 1 / 19 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Range expansion alters latitudinal life history trends has outpaced that of its native habitat and the species has established itself in novel vegetation
in temperate salt marshes [25]. We identified latitudinal patterns of life history traits (female size at maturity and average
size at reproduction) of A. pisonii across a broad (13.73˚) latitudinal gradient that included its
native mangrove habitat and novel salt marsh vegetation. Additionally, for a representative of
each habitat type (native mangrove and novel mixed marsh) we compared the proportion of
females actively reproducing at multiple time points as well as fecundity and metrics of mater-
nal investment (egg volume and weight) and offspring quality (larval starvation resistance). Finally, because selection pressures such as predation may vary between habitats, and this may
influence the life history traits of interest here (e.g. [27–28]), we used simple metrics designed
to assess the ability of individuals to utilize the vegetation structure in novel habitats as a
potential predator evasion strategy, and compared size-specific predation risk in both native
and novel habitats. 1Laguna de Tacarigua National Park, Venezuela [31] and
2Port Royal Mangroves, Jamaica [32]. Field surveys performed by the authors were supplemented by two sites from the literature Introduction higher latitudes) tend to have larger body sizes than those in warmer temperatures
(i.e. lower latitudes) as a result of thermoregulatory mechanisms. Bergmann’s clines are now
commonly used to describe intraspecific patterns in the body size of both endotherms and
ectotherms across latitude [21–22]. Although there is no definitive consensus on the mecha-
nisms underlying body size clines in ectotherms [19], factors such as temperature, resource
availability, and predation are strong drivers of body size patterns [22–24]. Selection pressures associated with range expansions may influence biogeographic phe-
nomena such as Bergmann clines, as well as trends in other physiological and reproductive
characteristics [17]. Here, we examine the potential for range expansion into a novel habitat
to alter life history traits within the context of existing latitudinal life history clines using the
model organism Aratus pisonii (mangrove tree crab). It is one of numerous species whose
recent climate-induced range expansion has led to a spatial mismatch with a habitat-forming
plant species [25]. Within its native habitat, this neotropical mangrove crab exists in close asso-
ciation with several mangrove species, including red, black, and white mangroves (Rhizophora
mangle, Avicennia germinans, and Laguncularia racemosa, respectively). In particular, the red
mangrove R. mangle provides both food and shelter for this arboreal species [26]. Although
mangroves in Florida have experienced a substantial recent poleward expansion along the At-
lantic coast due to a decrease in the frequency of extreme cold events [9], the climate change-
induced poleward expansion of A. pisonii (estimated at 62 km/decade in the last century [25]) PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 2 / 19 https://doi.org/10.1371/journal.pone.0176263.t001 ezuela [31] and ere supplemented by two sites from the literature y the authors were supplemented by two sites from the literature Determination of latitudinal life history trends In order to determine latitudinal patterns of body size and size at maturity, we conducted pop-
ulation surveys at 12 sites spanning a 13.73˚ latitudinal range in the northern hemisphere
(Table 1, Fig 1). All field research was conducted with permission from the appropriate author-
ity, which included the Florida Department of Environmental Protection (Permit Numbers:
04031310, 04221420), United States National Park Service (Permit Number: BISC-2012-
SCI-0022), Florida Fish and Wildlife Conservation Commission (License Number: SAL-13-
1497-SR), and Belize Fisheries Department (Reference Number: GEN/FIS/15/04/2013 (54)
Vol. IX). No protected species were disturbed or sampled during research activities in the
field. Of the 12 sites surveyed, eight were pure mangrove habitats, three were hybrid mangrove-
salt marsh habitats (hereafter referred to as mixed marsh), and one was a pure salt marsh escription of sites where Aratus pisonii body size distribution surveys were conducted. Table 1. Description of sites where Aratus pisonii body size distribution surveys were conducted. Survey Site
Habitat
Latitude (˚N)
Sample size
Laguna de Tacarigua National Park, Venezuela1
Mangrove
10.87
[31]
Twin Cays, Belize
Mangrove
16.78
132
Port Royal Mangroves, Jamaica2
Mangrove
17.93
[32]
Kemp Channel, FL, USA
Mangrove
24.66
128
Keys Marine Lab, FL, USA
Mangrove
24.83
101
Biscayne Bay National Park, FL, USA
Mangrove
25.46
89
Hobe Sound National Wildlife Refuge, FL, USA
Mangrove
27.09
137
St. Lucie Inlet State Park, FL, USA
Mangrove
27.14
104
Avalon State Park, FL, USA
Mangrove
27.55
127
Sebastian Inlet State Park, FL, USA
Mangrove
27.85
115
Fort Matanzas, FL, USA
Mixed Marsh
29.73
98
Devil’s Elbow, FL, USA
Mixed Marsh
29.75
86
Anastasia State Park, FL, USA
Mixed Marsh
29.88
140
Big Talbot Island State Park, FL, USA
Salt Marsh
30.51
115 Table 1. Description of sites where Aratus pisonii body size distribution surveys were conducted. Field surveys performed by the authors were supplemented by two sites from the literature 3 / 19 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Range expansion alters latitudinal life history trends Fig 1. Map exhibiting the geographic distribution of sampling sites. Circle symbols represent mangrove habitats; triangle symbols represent marsh
habitats. Red symbols represent sites sampled by the authors; blue symbols represent sites for which data was established from the literature [31–32]. Map
was prepared with ESRI ArcGIS v10 software using mangrove distribution data from [33]. https://doi org/10 1371/journal pone 0176263 g001 Fig 1. Map exhibiting the geographic distribution of sampling sites. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Range expansion alters latitudinal life history trends intertidal zone [34]. In the mangroves, crabs were collected by hand from the lower canopy
and prop roots above the water line. In the salt marsh, crabs were collected by hand from
exposed cordgrass. We measured their carapace width (CW) to the nearest tenth of a millime-
ter, identified their sex, and determined the reproductive status (gravid or not gravid) of
females. We considered size at maturity to be the smallest gravid female collected at each site,
consistent with previous methods of determining size at maturity in the species [31–32]. In
order to expand our latitudinal coverage of size at maturity data, we also included published
reports of size at maturity from two additional sites in mangrove habitats [31–32], which both
used the same method for determining size at maturity. We subsequently included size at
maturity values from these literature sources in the same statistical analysis as the values estab-
lished by our surveys. Thus, we maintained methodological consistency by considering size at
maturity to be the smallest gravid female collected at each site, rather than employing alterna-
tive methods for determining size at maturity (e.g. [35]). In order to determine the influence of latitude and habitat type on (1) size at maturity and
(2) average size of ovigerous females, we used separate general linear models for each response
variable. Latitude and habitat type were included as explanatory variables. The interaction
term between these variables was initially considered, but it was not significant for the model
of size at maturity (P = 0.844) or the model of average size of ovigerous females (P = 0.331). Therefore, the interaction was removed to simplify both models. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Determination of latitudinal life history trends Circle symbols represent mangrove habitats; triangle symbols represent marsh
habitats. Red symbols represent sites sampled by the authors; blue symbols represent sites for which data was established from the literature [31–32]. Map
was prepared with ESRI ArcGIS v10 software using mangrove distribution data from [33]. h
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01 6263 001 https://doi.org/10.1371/journal.pone.0176263.g001 habitat. The number of mixed and pure salt marsh sites surveyed was constrained by the extent
of hybrid marsh sites and the limited distribution of A. pisonii in pure salt marsh habitats. The
term “mixed marsh” used here refers to a site that is dominated by salt marsh cordgrass Spar-
tina alterniflora with isolated dwarf mangrove trees occasionally interspersed among the cord-
grass. However, because mangroves themselves have only recently begun expanding their
range into these salt marsh habitats [9, 29], these dwarf trees are younger and smaller than
those found further south in mature mangrove forests. As a result, the mangrove trees in
mixed marshes are highly connected to the surrounding salt marsh, and their small size and
isolated nature likely do not lend themselves to the same microhabitat creation that has been
documented in mature mangrove forests (e.g. [30]). These mixed marshes therefore more
closely resemble salt marshes than mangrove forests. At each site, crabs (mean sample size ± SE = 112.36 ± 6.93 at each site) were randomly col-
lected during daytime high tides, at which time they are the most active foraging above the PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 4 / 19 Comparison of maternal investment, offspring quality, and fecundity at
representative sites In order to determine offspring quality, we collected ovigerous females independent of the
stage of their brood’s embryonic development from the representative mangrove (n = 21) and
mixed marsh (n = 18) sites described in the previous section and transported them live to the
Smithsonian Marine Station in Fort Pierce, FL. Crabs were maintained at ambient temperature
with a natural light: dark cycle in individual plastic aquaria (22.8 cm L x 15.2 cm W x 16.5 cm
H) containing approximately 200–300 mL of filtered seawater (salinity ~31 ppt) and a piece of
plastic mesh (20 cm x 4.5 cm) that provided a substrate on which crabs could exit the water. Crabs were fed plant material (S. alterniflora for crabs collected from the mixed marsh habitat
and R. mangle for crabs collected from the mangrove habitat) ad libitum. Both food and water
were changed every 48 hours. Although there is no published literature documenting the food
sources or dietary preferences of A. pisonii in marsh habitats, M. Riley conducted dissections
of individuals collected from the marsh and observed plant material with striations character-
istic of S. alterniflora in the crabs’ gut contents. In addition, individuals offered only S. alterni-
flora in captivity for up to two weeks produced frass, which further demonstrated that S. alterniflora is a potential food source for the species. Moreover, all crabs included in the off-
spring quality experiment were gravid upon collection from the field, so the diet crabs were
fed in captivity did not influence reproductive investment or offspring quality in the brood
examined. In order to determine offspring quality, we collected ovigerous females independent of the
stage of their brood’s embryonic development from the representative mangrove (n = 21) and
mixed marsh (n = 18) sites described in the previous section and transported them live to the
Smithsonian Marine Station in Fort Pierce, FL. Crabs were maintained at ambient temperature
with a natural light: dark cycle in individual plastic aquaria (22.8 cm L x 15.2 cm W x 16.5 cm
H) containing approximately 200–300 mL of filtered seawater (salinity ~31 ppt) and a piece of
plastic mesh (20 cm x 4.5 cm) that provided a substrate on which crabs could exit the water. Range expansion alters latitudinal life history trends at maturity but not ovigerous) and gravid females in the population in order to elucidate
whether a single cohort or multiple cohorts were reproducing simultaneously in each habitat
type. The proportion of gravid females in each habitat type was determined for each monthly
survey by dividing the total number of gravid females by the total number of females larger
than the site-specific size at maturity from each monthly sampling effort. This data was ana-
lyzed using a generalized linear model with a binomial distribution and month and habitat as
categorical explanatory variables. Comparison of body size distributions and reproductive effort at
representative sites We conducted additional surveys throughout the summer reproductive peak (June, July, and
August 2013) to further investigate body size distributions and reproductive effort. Due to
logistical constraints, these additional surveys were restricted to two representative sites: a
representative mature mangrove forest (Avalon State Park, Fort Pierce, FL, 27.55˚N) and a
representative mixed marsh (Anastasia State Park, St. Augustine, FL, 29.87˚N), which were
chosen after careful consideration of several factors. Both sites (1) exemplify the prominent
characteristics of each habitat type, (2) occur at relatively similar latitudes but are neither the
northernmost nor southernmost site surveyed for each habitat type, and (3) initial surveys
demonstrated that body size patterns at the two sites were consistent with the overall latitudi-
nal patterns described here, which were established based on 12 sampling sites and two addi-
tional sites identified from the literature [31–32]. At each of these representative sites, we collected female crabs (mean sample size ± SE =
57.17 ± 4.14 at each time point) as described previously. This sampling (mangrove: June
(n = 41), July (n = 62), August (n = 53), marsh: June (n = 55), July (n = 58), August (n = 69))
increased our total sample size for the representative mangrove and marsh sites to n = 156 and
n = 182, respectively. For each individual, we measured CW to the nearest tenth of a millimeter
and determined their reproductive status (ovigerous or not ovigerous). Because A. pisonii
carry multiple broods each year, we took advantage of A. pisonii’s reproductive lunar synchro-
nization [32] and conducted all monthly surveys in the week preceding the full moon to cap-
ture periods of maximum reproductive effort for both populations. From these surveys, we
compared body size distribution patterns of reproductively mature and gravid females, as well
as the proportion of females that were gravid at each of the three monthly samples. Body size data was analyzed using a Wilcoxon rank-sum test and Fisher’s F-test in order to
compare mean size and size variability between the two populations, respectively. Addition-
ally, Hartigan’s dip test for unimodality [36] was calculated to determine the body size distri-
bution (unimodal, bimodal, etc.) of both reproductively mature (i.e. above the site-specific size 5 / 19 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Range expansion alters latitudinal life history trends changed and larval mortality was monitored daily to determine starvation resistance (days) as
a proxy for maternal energy provisioning. The impact of habitat on larval starvation resistance
was determined using a mixed effects Cox model, with unique maternal identity included as a
random effect, thus preventing pseudoreplication by accounting for the twelve larvae collected
from each individual female while simultaneously focusing on the importance of maternal
habitat on larval starvation resistance. In order to determine egg weight and fecundity, additional ovigerous females (n = 22 per
site) were collected from the same two representative sites and placed immediately upon ice. Eggs were removed from the pleopods of each female, egg stage (eyed/noneyed) was deter-
mined, and crabs with recently extruded, non-eyed eggs were dissected to ensure that they were
post-vitellogenic (i.e. all eggs were extruded). A subset of eggs was taken from 15 randomly
selected crabs from each habitat type and counted under a dissecting microscope. These subsets
were dried at 65˚C for 48 hours and weighed. We determined the relationship between egg
count and dry egg mass using a linear regression; the slope of this relationship indicates the
average dry mass of a single egg [37]. We then measured the total brood dry weight of the ovi-
gerous individuals collected from both habitats and used the total brood dry weight and the
average dry mass of a single egg to calculate the total fecundity of individuals of varying sizes
collected from both habitats (n = 22 per site). The impact of habitat on individual egg weight
was analyzed with a general linear model with dry mass of the subset of eggs as the response var-
iable and egg count and habitat as explanatory variables. Fecundity was analyzed using a general
linear model with maternal body size (CW) and habitat as explanatory variables. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Comparison of maternal investment, offspring quality, and fecundity at
representative sites A small portion (<1/3) of each female’s egg mass was gently removed from the pleopods
with forceps within 24 hours of collection from the field, placed into filtered sea water
(~31 ppt), and visually examined under a Leica M205C dissecting microscope at 80X to deter-
mine egg developmental stage. Eggs were categorized as non-eyed (mangrove n = 10, marsh
n = 15) or eyed (mangrove n = 11, marsh n = 3) based on visual inspection of each female’s
brood. For females carrying recently extruded, non-eyed eggs, which are reflective of initial
maternal investment in the quantity of egg yolk, a subsample of the removed eggs (n = 20–30)
was photographed using a Jenoptik ProgRes C14+ camera attached to the Leica M205C dis-
secting microscope and ProgRes CapturePro v2.8.0 software. Egg size was then determined by
randomly selecting 10 eggs from the photographed egg mass and measuring the circular area
of each egg with imageJ software. Capitalizing on the spherical nature of A. pisonii eggs, these
measurements of circular area were used to determine egg volume and averaged to provide the
mean egg size for each female. The impact of maternal habitat on mean egg volume was deter-
mined using a general linear model. Other than the non-eyed eggs sampled for egg size determination, the remainder of each
female’s brood (whether non-eyed or eyed eggs), was allowed to remain on the pleopods and
continue normal development. Crabs were monitored twice daily for larval hatching. Twelve
newly hatched larvae from each brood (mangrove n = 21 and mixed marsh n = 18 as described
previously) were randomly selected and pipetted into individual Kimble borosilicate glass cul-
ture tubes (15 mm x 85 mm) with ~80 mL of 0.2 μm filtered seawater. Water was partially 6 / 19 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Range expansion alters latitudinal life history trends (8–15 mm CW) were collected from the representative mixed marsh site and large individuals
(18–25 mm CW) were collected from the representative mangrove site for tethering at both
habitats. We brought A. pisonii from both size classes into the laboratory and affixed monofila-
ment tethers (45 cm of 10 lb test) by tying a loop in the monofilament and attaching this loop
to the back of the carapace using cyanoacrylate glue. Crabs were maintained in individual
aquaria overnight and observed the following morning to ensure that tethers remained secure. Crabs were then transported to the field at low tide. Tethers were attached to mangrove roots
or dead S. alterniflora stalks above the low tide line such that individuals could access the water
to stay moist at low tide and exit the water at high tide. Individuals were left in the field for 24
hours and then checked for survival as well as any indications of predation (e.g. pieces of cara-
pace remaining on tether) during low tide. This tethering protocol was repeated four times in
each habitat for a total of 149 individuals (mangrove n = 78, mixed marsh n = 71). Because the
two habitat types have different physical and structural characteristics, we avoided differences
in tethering artifacts between habitats (see review by [38]) by restricting our analyses to com-
parisons of predation on the two size classes within each habitat using separate generalized lin-
ear models with a binomial distribution. Predator avoidance capability and comparison of size-specific predation
potential In order to investigate the shift in body size distribution to smaller individuals in the mixed
marsh habitats revealed by our population surveys (see Results), we conducted two simple
assessments. First, we assessed the ability of large A. pisonii collected from mangrove habitats
to climb the habitat structure available in novel salt marsh habitats. Arboreality is one of the
most important predator evasion techniques employed by this species in native mangrove sys-
tems [34], where they are preyed upon by larger crabs, mangrove snappers (Lutjanus griseus),
ibises, and raccoons in the lower canopy and on the mud surface [26,32]. The goal of this experiment was to determine whether large crabs are also able to climb the
cordgrass present in salt marsh habitats. Because small crabs are commonly observed climbing
cordgrass in the marsh, they were not included in this experiment. We conducted laboratory trials in which individuals (n = 12, mean ± SE = 22.6 ± 0.4 mm
CW) were placed in 5-gallon buckets containing sediment to a height of 50 mm with a single
dead stalk of S. alterniflora (mean ± SE = 1.0 ± 0.4 mm basal stem diameter) placed upright in
the center of the bucket to simulate the habitat structure available in marsh habitats. Seawater
was then added up to a height of 100 mm to simulate a high tide event, and individuals were
placed in the bucket. Crabs in the salt marsh commonly climb both live and dead S. alterniflora
stalks, which provide similar support to crabs. Dead stalks were used in this microcosm experi-
ment to avoid any potential wilting. Behavior (climbing/not climbing habitat structure) was observed hourly for 6 hours (the
length of a high tide cycle). If a crab had not demonstrated the ability to climb the habitat
structure by the end of the 6 hours, the water was gently disturbed by hand to encourage the
crab to climb the stalk of S. alterniflora and verify its ability to do so. Next, we studied size-specific predation potential in both mangrove and mixed marsh habi-
tats. Because individuals (CW>15 mm) are not present in mixed marshes, small individuals 7 / 19 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Maternal investment, offspring quality, and fecundity at representative
sites Initial maternal investment in eggs, as measured by mean volume of non-eyed eggs, was not
impacted by maternal habitat (GLM, mean slope ± SE = 0.001 ± 0.002, P = 0.759, Fig 5). Similarly, individual egg weight (mean for both habitat types = 0.00729 mg), as measured by
the slope of the regression between egg count and weight, did not differ between habitat
types (GLM, mean slope ± SE = 0.063 ± 0.065, P = 0.343). Larval starvation resistance was
significantly impacted by habitat type, with offspring from mothers in the mangroves dem-
onstrating enhanced starvation resistance compared to those from mothers in the mixed
marsh (mixed effect Cox model, Maternal habitat: Coefficient ± SE = 0.855 ± 0.300, P =
0.004, Fig 6). Fecundity increased significantly with body size but was not explicitly impacted by habitat
type (GLM, Maternal body size: mean slope ± SE = 1754.7 ± 191.3, P<0.001; Habitat: pa-
rameter estimate ± SE = 1394.0 ± 1255.3, P = 0.274, Fig 7). However, due to differences in
maternal body size, fecundity was higher in the mangroves (mean no. eggs/ individual ± SE =
15215.9 ± 1027.6, Fig 7) than in the mixed marsh (mean no. eggs/ individual ± SE = 6649.7 ±
404.9, Fig 7). Latitudinal life history trends Female size at maturity increased significantly with latitude, but this trend was countered
by the species’ range expansion into novel marsh habitats, as size at maturity was smaller in
marshes at the highest latitudes of the species range than in any of the mangrove habitats
surveyed (GLM, Latitude: mean slope ± SE = 0.373 ± 0.097, P = 0.003; Habitat: parameter
estimate ± SE = 7.986 ± 1.202, P<0.001, Fig 2). Similarly, the average size of ovigerous females Fig 2. Latitudinal trends in female size at maturity in A. pisonii in both native and novel habitats. https://doi.org/10.1371/journal.pone.0176263.g002 Fig 2. Latitudinal trends in female size at maturity in A. pisonii in both native and novel habitats. https://doi.org/10.1371/journal.pone.0176263.g002
PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017
8 / 19 Fig 2. Latitudinal trends in female size at maturity in A. pisonii in both native and novel habitats. https://doi.org/10.1371/journal.pone.0176263.g002 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 8 / 19 Range expansion alters latitudinal life history trends increased with latitude (GLM, Latitude: mean slope ± SE = 0.312 ± 0.046, P<0.001), but this
trend was reversed at the highest latitudes of the species’ range, where gravid individuals were
significantly smaller in novel marsh habitats than native mangrove habitats (GLM, Habitat:
parameter estimate ± SE = 8.358 ± 0.313, P<0.001). Body size distributions and reproductive effort at representative sites Female size at maturity occurred at 8.9 mm CW in the representative mixed marsh habitat and
at 14.4 mm CW in the representative mangrove habitat (Figs 2 and 3). In the marsh, the body
size distribution of all females larger than the site-specific size at maturity was unimodal (Har-
tigan’s dip test, D = 0.031, P = 0.227, Fig 3(A)), while body size distribution of these females
was bimodal in the mangroves (Hartigan’s dip test, D = 0.046, P = 0.016, Fig 3(A)). The mean
size of gravid females was smaller in the mixed marsh habitat (mean ± SD = 11.4 ± 1.3 mm
CW, n = 119) than the mangrove habitat (mean ± SD = 19.2 ± 2.6 mm CW, n = 70) (Wilcoxon
rank-sum test, p< 0.001, Fig 3(B)). Additionally, gravid females in the mangrove population
displayed 3.53x more variation in body size than those in the mixed marsh population (Fisher’s
F-test, p<0.001, Fig 3(B)). Although the proportion of females that were gravid increased at both sites over the
monthly summer sampling dates, it was consistently higher in the population from the mixed
marsh habitat than the population from the mangrove habitat (GLM, Month: P<0.001; Site:
parameter estimate ± SD = -1.05 ± 0.25, P<0.001, Fig 4). PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Predator avoidance capability and comparison of size-specific predation
potential All individuals demonstrated the ability to climb S. alterniflora for predator evasion. Field teth-
ering results indicate that predation potential is nearly twice as high on large crabs than small
crabs in novel marsh vegetation (GLM, mean slope ± SE = 0.860 ± 0.501, P = 0.086, Fig 8),
although this was only marginally significant, potentially due to insufficient replication. By
contrast, tethering experiments revealed no differences in size-selective predation in native
mangrove habitats (GLM, mean slope ± SE = 0.041 ± 0.451, P = 0.927, Fig 8). PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 9 / 19 Range expansion alters latitudinal life history trends Fig 3. Body size distribution patterns. Includes all Aratus pisonii (A) females larger than the site-specific
size at maturity and (B) gravid females from June-August 2013. Density curves are included in (A) to highlight
modality differences between habitats. https://doi org/10 1371/journal pone 0176263 g003 Fig 3. Body size distribution patterns. Includes all Aratus pisonii (A) females larger than the site-specific
size at maturity and (B) gravid females from June-August 2013. Density curves are included in (A) to highlight
modality differences between habitats. Fig 3. Body size distribution patterns. Includes all Aratus pisonii (A) females larger than the site-specific
size at maturity and (B) gravid females from June-August 2013. Density curves are included in (A) to highlight
modality differences between habitats. https://doi.org/10.1371/journal.pone.0176263.g003 https://doi.org/10.1371/journal.pone.0176263.g003 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 10 / 19 Range expansion alters latitudinal life history trends Fig 4. Proportion of gravid females. Proportion of ovigerous females in each Aratus pisonii population
throughout the survey period. Bars represent standard error. https://doi.org/10.1371/journal.pone.0176263.g004 Fig 4. Proportion of gravid females. Proportion of ovigerous females in each Aratus pisonii population
throughout the survey period. Bars represent standard error. Fig 4. Proportion of gravid females. Proportion of ovigerous females in each Aratus pisonii population
throughout the survey period. Bars represent standard error. https://doi.org/10.1371/journal.pone.0176263.g004 https://doi.org/10.1371/journal.pone.0176263.g004 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Discussion Our study demonstrates that there is significant variation in key life history traits throughout
the current geographic distribution of the mangrove tree crab Aratus pisonii. In accordance
with traditional biogeographic patterns (i.e. Bergmann’s clines), intraspecific body size, includ-
ing both size at maturity and average size of reproductive females, increased with latitude
within native mangrove habitats. However, this Bergmann’s cline deteriorated in mixed marsh
and pure marsh sites at the northernmost latitudes of the species’ distribution. On average,
gravid females in marshes at the highest latitudes of the species’ range were just 60% as large as
those in the mangroves at lower latitudes. Although it is possible that our method of establish-
ing size at maturity may underestimate intraspecific plasticity within a site, intraspecific body
size, including both size at maturity and average size of reproductive females, clearly differed
between habitats. This life history pattern suggests that A. pisonii at the range edge display
accelerated sexual maturation, as has been seen in a number of other species expanding their
range [39–41]. Although decreased size at maturity at the range edge is consistent with range expansion
predictions [15–17], it is an unusual contrast to the Bergmann’s cline demonstrated by A. piso-
nii throughout its native range. This contrast is likely due to habitat-specific differences associ-
ated with A. pisonii’s range expansion into novel marsh habitat. Organisms at the range edge
often face selective pressures that are unique compared to those experienced by individuals
within the core of the species’ range [15–17, 42]. Just as gradients in biotic and abiotic interac-
tions underlie Bergmann’s clines, habitat-specific differences in these elements may also influ-
ence life history traits such as body size and size at maturity [17]. Factors such as temperature [43–44], resource availability [23], and predation pressure [27,
45] have been shown to impact life history characteristics. Terrestrial multivoltine arthropods, PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 11 / 19 Range expansion alters latitudinal life history trends Fig 5. Mean Aratus pisonii egg volume (mm3) of recently extruded, non-eyed eggs. Bars represent standard error. h
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0176263 005 Fig 5. Mean Aratus pisonii egg volume (mm3) of recently extruded, non-eyed eggs. Bars represent standard Fig 5. Mean Aratus pisonii egg volume (mm3) of recently extruded, non-eyed eggs. Bars represent standard error. Fig 5. Mean Aratus pisonii egg volume (mm3) of recently extruded, non-eyed eggs. Bars represent standard error. https://doi.org/10.1371/journal.pone.0176263.g005 such as the species described here, generally adhere to the temperature-size rule [22]. The tem-
perature-size rule describes the phenotypically plastic relationship between body size at differ-
ent developmental stages (e.g. reproductive maturity) and rearing temperature [46]. As a result
of this phenotypic response to temperature during ontogeny, individuals reared at low temper-
atures delay maturity, reaching maturity at a larger body size and growing to be larger overall
than conspecifics reared at warmer temperatures [43–44]. Although we did not explicitly compare temperature profiles from the two habitats, the
structural complexity of mangrove forests provides numerous shady microhabitats that are
lacking in more structurally simplistic marsh habitats. It is well-established that forest canopies
maintain lower temperature maximums than contiguous forest gaps or grassy habitats ([47]
and references therein), and the temperature profile of red mangrove trees measured in late
summer demonstrates that the underside of red mangrove roots is 6.84˚C cooler than the top
of the roots [30]. Thus, individuals with access to the shaded mangrove understory may experi-
ence lower maximum temperatures than their conspecifics in less structurally complex salt PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 12 / 19 Range expansion alters latitudinal life history trends Fig 6. Comparison of Aratus pisonii larval survival. Kaplan-Meier curves show A. pisonii larval survival
(i.e., starvation resistance in days) as a function of maternal habitat. Solid lines represent mean survival and
dashed lines represent 95% confidence intervals. h
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0176263 006 Fig 6. Comparison of Aratus pisonii larval survival. Kaplan-Meier curves show A. pisonii larval survival
(i.e., starvation resistance in days) as a function of maternal habitat. Solid lines represent mean survival and
dashed lines represent 95% confidence intervals. https://doi.org/10.1371/journal.pone.0176263.g006 https://doi.org/10.1371/journal.pone.0176263.g006 marsh habitats. Consequently, our results are consistent with those predicted by the tempera-
ture-size rule after incorporating habitat-specific differences within its framework. It should be noted that interpretation of the temperature-size rule in this scenario is inher-
ently complicated by habitat-specific differences in resource quality and availability. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 For exam-
ple, diet quality has been shown to reverse the temperature-size rule in insects: in laboratory
experiments, individuals reared on a high quality diet followed the temperature-size rule,
while those reared on a low quality diet showed a reversed response, with smaller final sizes at
higher temperatures [23]. Although there are no studies in the literature detailing the diet of A. pisonii in the marsh, their omnivorous diet in native mangrove habitats is well established. In
native habitats, mangrove leaves comprise a core component of A. pisonii’s diet [48]. In addi-
tion, individuals commonly consume small prey items such as insects and juvenile conspecif-
ics; consumption of animal material is tightly linked to the fitness and physiological condition
of the species [49]. Conversely, mangrove leaves are scarce or entirely absent in novel marsh and mixed marsh
sites, forcing individuals to consume alternative food sources. Although little is known about
the diet of A. pisonii in the marsh, a number of scenarios are possible. First, individuals in the
marsh may consume a diet containing less animal material or poorer quality vegetation than
those in the mangroves. In this case, the crabs in the marsh would be expected to have a slower
growth rate than their conspecifics in the mangroves. Alternatively, crabs in the marsh may
consume more animal material than those in the mangroves, which would be expected to
increase their physiological condition and growth rate [49]. In addition, the quality of the
marsh diet may influence whether individuals there display growth patterns consistent with or PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 13 / 19 Range expansion alters latitudinal life history trends Fig 7. Relationship between maternal carapace width (mm) and fecundity (number of eggs) in Aratus pisonii. Inset shows a
comparison of standardized fecundity (number of eggs per mm of CW) between habitats. https://doi.org/10.1371/journal.pone.0176263.g007 Fig 7. Relationship between maternal carapace width (mm) and fecundity (number of eggs) in Aratus pisonii. Inset shows a
comparison of standardized fecundity (number of eggs per mm of CW) between habitats. https://doi.org/10.1371/journal.pone.0176263.g007 https://doi.org/10.1371/journal.pone.0176263.g007 in opposition to the temperature-size rule (e.g. [23]). Thus, habitat-specific differences in con-
sumption of small prey items such as insects, juvenile conspecifics, or other crustaceans may
influence the strength of the relationship between temperature and body size in these two hab-
itats (e.g. [23]). Further studies are needed to tease apart the relative contribution of these
mechanisms. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Finally, predation pressure has also been shown to drive differences in both size at maturity
and overall body size via phenotypic or genetic processes. For instance, elevated predation
intensity from natural predators or commercial harvesting can drive declines in body size at
reproductive maturity (e.g. [27, 45] but see [50] for an exception). Our tethering trials indi-
cated that predation potential on A. pisonii was almost twice as high on large individuals than
small individuals in the representative mixed marsh habitat. Conversely, there was no evidence
of size-specific predation in the representative mangrove habitat. Because large crabs are not
present in salt marsh habitats, all of the large crabs used in the tethering trials were collected PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 14 / 19 Range expansion alters latitudinal life history trends Fig 8. Size-specific predation pressure on individuals of Aratus pisonii from native mangrove and
novel mixed marsh sites. Bars represent standard error. https://doi org/10 1371/journal pone 0176263 g008 Fig 8. Size-specific predation pressure on individuals of Aratus pisonii from native mangrove and
novel mixed marsh sites. Bars represent standard error. Fig 8. Size-specific predation pressure on individuals of Aratus pisonii from native mangrove and
novel mixed marsh sites. Bars represent standard error. https://doi.org/10.1371/journal.pone.0176263.g008 from mangrove habitats. Although we previously established that large crabs are capable of
climbing cordgrass in the salt marsh, individual inexperience in the salt marsh may have con-
tributed to the increased predation on large individuals. Additionally, habitat may influence
prey palatability, and predator preferences may have been impacted by the inclusion of only
small crabs collected from the salt marsh in the tethering study. Finally, predator community
composition may differ between the two habitat types, which could also impact predation pat-
terns. Nevertheless, even the slight size-specific predation discrepancy noted between large
and small individuals in novel habitats could drive the observed decrease in body size and size
at reproduction (e.g. [27]). Thus, the life history patterns associated with A. pisonii’s range
expansion are consistent with multiple mechanisms presented here, and we are currently
unable to tease apart the relative influence of these explanations. Variation in size at maturity between habitats was also accompanied by differences in the
population structure of reproductive females, which clearly contrasted between native and
novel habitats. Specifically, the body size distribution of reproductive females was bimodal in
the mangroves and unimodal in the mixed marsh habitat. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 Conclusion In this study, we demonstrate that a climate change-induced range expansion into a novel hab-
itat overrules the Bergmann’s cline demonstrated by the species within its native habitat,
potentially due to differences in predation pressure or environmental conditions in the native
and novel habitats. The deterioration of this biogeographic body size pattern has a number of
implications for fecundity and offspring quality in novel habitats at the edge of the species’
range. As climate change advances, spatial mismatches between species distributions will con-
tinue to reshuffle biological communities, leading to novel communities and unique plant-ani-
mal interactions [12–14]. Understanding the potential for life history changes associated with
range expansions, as well as the mechanisms underlying these life history alterations, is critical
for the prediction of key demographic processes [17, 55]. Range expansion alters latitudinal life history trends eyestalks has recently been proposed for cold-water crustaceans [53], but its utility in aging
shorter lived, warm-water species such as A. pisonii requires further examination [53]. In addition to the observed differences in population structure and female size at maturity
in mangrove and marsh habitats, fecundity increased significantly with maternal body size, a
trait commonly observed in crustaceans (e.g. [54]). Differences in the size of reproductive
females in the two habitats therefore resulted in a higher mean nonstandardized fecundity
(mean no. eggs/ individual) in native mangrove habitats than novel marsh habitats. Initial
maternal investment in individual eggs, as measured by the mean egg volume and egg weight
of recently extruded eggs, was not impacted by maternal habitat. However, offspring quality,
as measured by larval starvation resistance, was improved in native mangrove habitats com-
pared to novel salt marsh habitats. Because there was no relationship between egg volume or
weight and maternal habitat, these differences in starvation resistance likely reflect differences
in the quality, but not the quantity, of egg provisioning, which could result from habitat-wide
differences in maternal physiological condition or previous reproductive investment. These
differences in offspring quality may have enduring consequences for the survival and persis-
tence of this species in novel habitats. Acknowledgments This is contribution #TBD of the Smithsonian Marine Station at Fort Pierce, FL and contribu-
tion #1061 of the Smithsonian Institution’s Caribbean Coral Reef Ecosystems (CCRE)
Program. Funding acquisition: MR BG. Investigation: MR. Methodology: MR BG. PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 These distributions suggest that two
cohorts (each represented by a peak in body size distribution) are reproducing simultaneously
in the population from the native mangrove habitat, while in the marsh population, either a
single cohort is reproducing or reduced growth increments prevent the identification of dis-
crete size/age classes. From our data, it is not evident whether differences in the size of oviger-
ous females reflect true differences in body size at a specific age or merely differences in body
size at a particular life stage. Regrettably, a reliable technique to directly determine age in crus-
taceans does not currently exist [51–52]. A method of direct aging using growth rings in the PLOS ONE | https://doi.org/10.1371/journal.pone.0176263
May 4, 2017 15 / 19 Conceptualization: MR BG. Data curation: MR. Formal analysis: MR. Funding acquisition: MR BG. References 1. Walther GR, Berger S, Sykes MT. Ecological responses to recent climate change. Nature. 2002; 416:
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Tomato and Watermelon Production with Mulches and Automatic Drip Irrigation in North Dakota
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Water
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Tomato and Watermelon Production with Mulches and
Automatic Drip Irrigation in North Dakota akash Vaddevolu 1, Justin Lester 1, Xinhua Jia 1,*
, Thomas F. Scherer 1 and Chiwon W. Lee 2 ay Bhanu Prakash Vaddevolu 1, Justin Lester 1, Xinhua Jia 1,*
, Thomas F. Scherer 1 and Chiw 1
Department of Agricultural and Biosystems Engineering, North Dakota State University,
Fargo, ND 58102, USA; uday.vaddevolu@ndsu.edu (U.B.P.V.); justin.lester.1@ndsu.edu (J.L.);
thomas.scherer@ndsu.edu (T.F.S.)
2
Department of Plant Sciences, North Dakota State University, Fargo, ND 58102, USA; chiwon.lee@ndsu.edu
*
Correspondence: xinhua.jia@ndsu.edu 1
Department of Agricultural and Biosystems Engineering, North Dakota State University,
Fargo, ND 58102, USA; uday.vaddevolu@ndsu.edu (U.B.P.V.); justin.lester.1@ndsu.edu (J.L.);
thomas.scherer@ndsu.edu (T.F.S.) Fargo, ND 58102, USA; uday.vaddevolu@ndsu.edu (U.B.P.V.); justin.lester.1@ndsu.edu (J.L.);
thomas.scherer@ndsu.edu (T.F.S.)
2
Department of Plant Sciences, North Dakota State University, Fargo, ND 58102, USA; chiwon.lee@ndsu.edu
*
Correspondence: xinhua.jia@ndsu.edu 2
Department of Plant Sciences, North Dakota State University, Fargo, ND 58102, USA; chiwon.lee@ndsu.edu
*
Correspondence: xinhua.jia@ndsu.edu 2
Department of Plant Sciences, North Dakota State University, Fargo, ND 58102, USA; chiwon.lee@ndsu.edu
*
Correspondence: xinhua.jia@ndsu.edu Abstract: In North Dakota, agriculture contributes a large sector of the state’s economy, but veg-
etable production is limited due to the state’s climate condition. Inadequate soil moisture and low
soil temperature are the two major factors prohibiting quality produce and high-yield vegetable
production. In this study, a soil-water potential, sensor-based drip irrigation system was developed,
designed, and installed to evaluate its application on tomato and watermelon productions in a
two-year field experiment in 2019 and 2020. The experimental treatments were drip irrigation and no
irrigation under three mulches: black plastic, clear plastic, and landscape fabric mulches. Irrigation
was scheduled at 8:00 am for watermelon and 9:00 a.m. for tomato, with the ability for each irrigation
event to be bypassed based on the soil moisture conditions. Due to rainfall differences in the two
years, irrigation was barely needed in 2019, but in 2020, drip irrigation was applied frequently. On
average, for the two-years’ field experiment, the highest yield for tomatoes was obtained from drip
irrigation under black plastic drip irrigation treatment with 40.24 Mg ha−1 in 2020, whereas the
highest yield for watermelon was from drip irrigation under clear plastic mulch with 165.55 Mg ha−1
in 2020. The effect of mulch, irrigation, and combined practices were analyzed based on the average
fruit weight and diameter, electrical conductivity (EC), pH, and sugar content of the samples.
Citation: Vaddevolu, U.B.P.; Lester,
J.; Jia, X.; Scherer, T.F.; Lee, C.W. Tomato and Watermelon Production
with Mulches and Automatic Drip
Irrigation in North Dakota. Water
2021, 13, 1991. https://doi.org/
10.3390/w13141991 Keywords: mulch; automatic; drip irrigation; soil moisture sensor; tomato; watermelon Received: 14 June 2021
Accepted: 17 July 2021
Published: 20 July 2021 Tomato and Watermelon Production with Mulches and
Automatic Drip Irrigation in North Dakota The
results showed that for watermelon, the average weight and diameter were significantly heavier and
higher with irrigation treatments, but the EC and the pH values were significantly higher with mulch
treatments. For tomatoes, the average weight, diameter, pH, and sugar content were all significantly
higher with mulch treatment, but the EC was higher with irrigation treatment. water water 1. Introduction Mulches are available in different colors, thickness, and materials, which can be used
for different purposes, such as earlier maturity, higher yield, reduce evaporation, control
weeds, and root pruning [5,13–16]. The temperature differences on the soil surface and
subsurface under different mulches depend on the thermal properties of the mulches, the
energy partitions of the incoming solar radiation, and the distance between the mulch and
the soil [17,18]. Clear plastic mulch exposed the surface to the incoming shortwave solar
radiation but compressed the outgoing longwave infrared radiation, which causes the soil
surface to be warmer due to the retained infrared radiation. Black plastic mulch completely
covers the soil surface and causes fewer soil temperature changes compared to the clear
mulch. However, the air temperature above the mulch can be increased and stays warmer
due to the released thermal energy or longwave infrared radiation from the black mulch. Weed growth is also reduced due to the lack of sunlight under the black mulch [15,18]. g
g
For high-value crops, drip irrigation is commonly used along with plastic mulch to get
high-quality production. Tomato and watermelon grown under drip irrigation and plastic
mulches can increase the water use efficiency and produce increased yields with less water
compared to those under no mulch treatments [19–21]. Ref. [22] evaluated the impact of
different water tension thresholds in well-drained, clayey soil on tomato growth stages and
found that the highest yield was from soil potential thresholds of 35, 12, and 15 kPa during
vegetative, fruit development, and maturation stages, respectively. Ref. [23] conducted a
field experiment to withhold irrigation at different stages to monitor the tomato water use
efficiency and yield and concluded that withholding drip irrigation between first flower
and first fruit stages can increase tomato marketable yield by 8–15% and save 20% of
irrigation water compared to that with regular irrigation treatment. Ref. [21] researched to
understand the combined effect of irrigation (drip or flood irrigation) and mulch (plastic
or sugarcane trash mulch) on tomato yield and water savings in heavy soils. They found
that the sugarcane trash mulch at 0.4 of pan evaporation level gave the highest fruit yield
with 44% of irrigation water savings. Similarly, plastic mulch can be used to control weed
and increase yield for watermelon. Ref. 1. Introduction Agriculture plays a large role in North Dakota’s (ND) economy and uses around
89 percent of the state’s land [1]. North Dakota has a warm temperature in summer, cold
temperatures in winter, and strong winds throughout the year. Wheat, soybean, and corn
are the major crops grown in ND, but horticulture crops, such as vegetables, can be also
grown in this region though the yields and quality are limited due to low soil temperature
in the root zone and lack of irrigation. Tomato and watermelon production are negligible
when compared to the soybeans, wheat, corn, sunflower, and potato production. With only
63 and 30 farms for tomato and watermelon fresh market sale in 2017 and an average farm
size less than 0.405 ha, the total areas were about 25 and 12 ha for tomato and watermelon
productions, respectively, in North Dakota [2]. However, many people grow vegetables in
their gardens for self-consumption, so currently, the extensions are on master gardens. The
possibilities to improve the vegetable productions are huge. Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article
distributed
under
the
terms
and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/). A warmer soil temperature can advance crop growth and improve crop phenological
development and therefore increase crop yield [3,4]. By adopting some management
practices, such as mulching, it is possible to maintain a warm soil temperature in the root Water 2021, 13, 1991. https://doi.org/10.3390/w13141991 https://www.mdpi.com/journal/water Water 2021, 13, 1991 2 of 22 zone. Mulching is a management practice that is popular all over the world [5–9] and can
be used in agriculture to create favorable conditions, including maintaining warm soil
temperature and suppressing weeds. In arid regions, mulch has been used to conserve
water through reduced evaporation while also increasing yields [8,10]. In cold and humid
regions, such as in North Dakota, the mulch is applied to increase soil temperature and
control weeds [11,12]. In a field experiment, [11] sweet corn was grown under different
mulches and it was found that the soil temperature under clear plastic mulch was 2.31 ◦C
warmer than that under the no mulch treatment for an entire season in Fargo, ND. 2.1. Experimental Site The two-year field experiment was conducted at a research farm located on the west
side of the North Dakota State University (NDSU) campus in Fargo, North Dakota. The field
plot is located at 46◦53′70′′ N and 96◦48′66′′ W, with an elevation of 274 m above sea level. The study area has a warm, humid continental climate [32] with an average temperature
of 5.98 ◦C, a maximal temperature of 28.04 ◦C in July, and a minimum temperature of
−17.02 ◦C in January. The average annual rainfall and potential evapotranspiration are
533–635 mm and 1103–1478 mm, respectively [33]. 1. Introduction [24] evaluated the impact of deficit irrigation on
watermelon yield at different stages and found that irrigation with 100% water requirement
produced the highest yield, whereas irrigation with 50% of irrigation requirement at
vegetative growth and fruit development stages severely reduced the watermelon numbers
and yield, while watermelon at the fruit-ripening stage was less sensitive to water deficit. Since the evaporation from the soil surface is reduced due to mulch cover, irriga-
tion scheduling using traditional crop coefficient (Kc) and reference evapotranspiration
approach becomes challenging due to the reduced or different Kc values [25,26] under
different mulches. Irrigation based on crop water consumption can provide favorable
conditions for higher crop production. In recent years, automatic irrigation based on soil
moisture sensors and controllers has become popular in urban landscape water manage-
ment and turfgrass irrigation for both sprinkler and drip irrigation methods [27]. Using
the smart irrigation technology for high-value crops in agriculture is relatively new, but
it has been shown with positive results of higher yields and better quality [27,28]. There
are several types of irrigation controllers available on the market that can work based
on evapotranspiration (ET) [29], crop coefficient [30], or soil moisture sensor (SMS) [31]. The SMS controller can start irrigation automatically with a soil moisture or a potential
threshold predetermined for a specific crop and soil type. This SMS-based irrigation can Water 2021, 13, 1991 3 of 22 only be applied depending on the soil moisture or potential status, thus can work under
different mulches. In this study, an automatic soil-water potential, sensor-controlled irrigation system
was developed, designed, and installed for a two-year field experiment to test the irrigation
system under different mulches. The objectives of the study were to (1) build and test the
automatic soil-water potential, sensor-controlled drip irrigation system under clear plastic
(CP), black plastic (BP), and landscape fabric (PF) mulches along with no mulch (NM);
(2) compare the irrigation schemes for tomato and watermelon; and (3) evaluate the effect
of irrigation and mulches on tomato and watermelon yield and quality. The hypothesis is
that the combined automatic drip irrigation and mulch only irrigate when soil is drier than
the threshold value and, at the same time, provide the best yield and highest quality of
produce. 2.2. Field Layout The field experiment was conducted in a 1340 m2 (36.6 × 36.6 m) area. The experiment
was designed as a split-split random design in 2019, with the vegetable and irrigation as the
splits and the four mulches randomly arranged (Figure 1a). The main plots were extended
in the west to east direction. In July 2019, a visible soil moisture difference was observed
after rainfall events, with the wetter surface in the west and drier surface in the east. It
was suspected that a potential tile drainage was installed in the east side. Therefore, to
avoid any potential soil moisture difference, the experiment was redesigned as a complete
randomized split-plot in 2020 (Figure 1b) for an area of 740 m2 (48.7 m × 15.2 m), with
the vegetable type as the only split plots. Since the two vegetables were arranged in the
west-east direction, if there are were differences in soil moisture due to the location of the
tile drainage system, there should have been minimal differences in each vegetable due to
the field orientation. Though the dimensions and designs of the experiments changed from 2019 to 2020,
the plot size remained unchanged, with a spacing of 6 m in length and 1.5 m in width [34]. Each plot contained six tomato plants or five watermelon plants placed along the center of
the plot, with a spacing of 1 m for tomatoes and 1.2 m for watermelon, respectively. The
treatments also stayed as the same, with two vegetables, two irrigations, and four mulches. In general, a field layout difference is expected for crop rotation purpose. 4 of 22
of 23 Water 2021, 13, 1991
EW (a)
(b)
Figure 1. Randomized field layout in (a) 2019 field layout and (b) 2020 layout. (a)
(b)
i
1 R
d
i
d fi ld l
i
( ) 2019 fi ld l
d (b) 2020 l
Figure 1. Randomized field layout in (a) 2019 field layout and (b) 2020 layout. (a)
(b)
Figure 1. Randomized field layout in (a) 2019 field layout and (b) 2020 layout. (b) (a) field layout in (a) 2019 field layout and (b) 2020 layout. mensions and designs of the experiments changed from 2019 to 2020,
ed unchanged, with a spacing of 6 m in length and 1.5 m in width [34]. 2.2. Field Layout six tomato plants or five watermelon plants placed along the center
acing of 1 m for tomatoes and 1 2 m for watermelon respectively The
2.3. Soil Analysis
Six soil samples were collected at different locations, with a grid pattern across the
field at 0–15 cm and 15–61 cm depth before the field experiment. The soils were analyzed
for nitrate-nitrogen, potassium, pH, electrical conductivity, and organic matter by the
NDSU soil-testing laboratory following standard protocols. The testing results are shown
in Table 1. mensions and designs of the experiments changed from 2019 to 2020,
ed unchanged, with a spacing of 6 m in length and 1.5 m in width [34]. six tomato plants or five watermelon plants placed along the center
acing of 1 m for tomatoes and 1 2 m for watermelon, respectively The
2.3. Soil Analysis
Six soil samples were collected at different locations, with a grid pattern across the
field at 0–15 cm and 15–61 cm depth before the field experiment. The soils were analyzed
for nitrate-nitrogen, potassium, pH, electrical conductivity, and organic matter by the
NDSU soil-testing laboratory following standard protocols. The testing results are shown
in Table 1. Water 2021, 13, 1991 5 of 22 Table 1. Soil nutrient analysis at the experimental site, while Avg means average value at eight
locations, and Std means standard deviation of values at eight locations. locations, and Std means standard deviation of values at eight locations. 2019
2020
Avg
Std
Avg
Std
NO3-N (kg N/ha)
0–15 cm
18.51
4.51
6.75
2.15
15–61 cm
62.24
8.43
33.95
11.76
P (mg/L)
21.50
4.81
26.63
5.95
K (mg/L)
368.13
35.95
480.63
31.78
pH
6.96
0.09
6.60
0.14
EC (dS/m)
0.41
0.06
0.39
0.03
Organic Matter (%)
7.74
0.99
6.31
0.53 From the soil testing results, the amount of fertilizer can be estimated based on the
recommended 168 kg N/ha for tomato and 134 kg N/ha for watermelon (Cihacek, L., ND’s
soil fertility specialist, personal communication). For the experimental area, with 46% N
content in urea, the urea application on the experimental plots was 31 and 21 kg for tomato
and watermelon in 2019 and 22 and 16 kg for tomato and watermelon in 2020, respectively. The urea was applied using a hand-held fertilizer spreader (Marysville, OH) to maintain
uniform application rate. 2.2. Field Layout pp
Soil core samples were collected at 5, 15, 30, 45, 60, 75, and 90 cm depth from the
study area in spring 2019. Soil bulk density and soil release curve were measured using
HYdraulic PROPerty analyzer (HYPROP) and WP4 dewpoint potentiometer method [35]. The estimated soil volumetric water content at field capacity (−33 kPa), permanent wilting
point (−1500 kPa), and saturation (θs) for the top 45 cm soil layer were averaged and listed
in Table 2, along with other soil physical parameters. These parameters are critical input
data for irrigation scheduling. Table 2. Soil physical parameters at the experimental site. Parameters
Depth
(0–45 cm)
Sand (%)
5
Silt (%)
47
Clay (%)
48
Bulk density (g/cm3)
1.08
Saturation (cm3/cm3)
0.59
Field capacity (cm3/cm3)
0.41
Permanent wilting point (cm3/cm3)
0.26 Table 2. Soil physical parameters at the experimental site. Table 2. Soil physical parameters at the experimental site. Table 2. Soil physical parameters at the experimental site. Parameters
Depth
(0–45 cm)
Sand (%)
5
Silt (%)
47
Clay (%)
48
Bulk density (g/cm3)
1.08
Saturation (cm3/cm3)
0.59
Field capacity (cm3/cm3)
0.41
Permanent wilting point (cm3/cm3)
0.26 2.5.1. Drip Drip tape was categorized based on the flow rate, which is an important factor used
to determine the duration of each irrigation event. For this study, drip tape (Chapin
Watermatics Inc, Watertown, NY, USA) had a 0.05 m emitter space and 5.68 L/min per
30.48 m drip length (0.005 m3/min per 30.48 m) flow-rate capacity. Two drip laterals with
0.4 m space were installed in each plot, which has an estimated flow rate of 106.6 L/min. For each lateral, one end was connected to the mainline using a barbed adaptor, while the
other end was folded and taped to close the flow. The three plots (or six laterals) in each
treatment were connected to one mainline, which then was connected to a control unit. All
mains were laid along the center between the treatments and extended to the controllers,
where the irrigation water was supplied. 2.5.3. Scheduling Irrigation scheduling is one of the most important components in water management
and determines when and how long to irrigate. It can be determined based on the soil
moisture content of the field, crop water requirement, and evapotranspiration rates. The
amount of water to be applied per irrigation can be calculated based on soil physical
properties, area irrigated, drip system characteristics, and management allowable depletion
(MAD). Based on the soil properties in Table 2 for the top 45 cm root zone, the total amount
of water needed to irrigate 60% of the surface area for the three plots in each treatment
(16.73 m2) is 0.113 m3. With an irrigation efficiency of 100% for such a small area and a flow
rate of 5.68 L/min per 30.48 m drip length, the irrigation time was estimated as 17 min for
the 36.6 m drip laterals at about 10% MAD. 2.5.2. Water Supply The water source for the drip irrigation system was a hydrant that is located around
20 m from the field. Water samples were collected in 2020 for chemical analysis and found
that the salinity and sodium concentration are at acceptable levels and were satisfactory
for irrigation according to ND irrigation water standards [36]. Water was diverted to the
field from the hydrant through a hosepipe after it was passed through a filter (Senninger
irrigation Inc., Clermont, FL, USA), pressure regulator (Agricultural products Inc., York,
NE, USA), and two flow meters (Omega Engineering Inc., Norwalk, CT, USA) to monitor
the flow rate to the tomato and watermelon controllers. 2.4. Mulch In this study, tomatoes and watermelons were grown under three different mulches,
including CP, BP, and PF, along with NM as the control. The CP and BP (Dubois Agri. Inc.,
QC, Canada) are 1.22 m wide and 20.32 µm thick, while the PF (Menards, Eau Claire, WI,
USA) was 1.22 m wide and 25 mm thick. A tractor that was mounted with a subcompact
raised bed mulch machine (Berry Hill Irrigation. Inc, Buffalo Junction, VA, USA) was used
to raise the bed height and lay the mulch on the selected plots. Two disks and deflectors
available on the machine were used to raise the beds to 12.7 cm above the ground with a
bed width of 1.0 m. Drip tapes were installed on the beds using drip tape guiders, and
mulch was laid on top of the drip tapes using the disks and press wheels. The side of the
mulch was covered with soil by the mulch machine to secure the mulch in place. Water 2021, 13, 1991 6 of 22 2.5.4. Controller The automatic drip irrigation system was built using a watermark electronic module
battery version (WEM-B, Irrometer Company, Riverside, CA, USA), a node controller
(Hunter, San Marcos, CA, USA), and a solenoid valve to initiate an irrigation event once
the pre-set potential threshold was reached. All components were arranged on two control
boards, with each board consisting of four control units, as shown in Figure 2. A total of
eight control systems were constructed to control each mulch treatment for each crop. In
each treatment, two watermark sensors linked in series were installed in the second plot of
the three plots at 15 and 30 cm depth, while an average for those two values was used to
control irrigation via the controller. The threshold for the irrigation initiation was selected
as 4, which kept the soil in the wet zone based on the switch position (Irrometer Company,
Riverside, CA, USA). The node controller was programmed to set up the time starting at
8:00 and 9:00 am for tomato and watermelon, respectively, with a 17 min irrigation duration
for each plot. 7 of 22
7 of 23 Water 2021, 13, 1991
IEW Figure 2. Irrigation controller board with two control units. Figure 2. Irrigation controller board with two control units. oller board with two control units. Figure 2. Irrigation controller board with two control units. 2.6.1. Rainfall in gauges, an automatic tipping bucket rain gauge, and a manual rain
Alternatives Inc., Fergus Falls, MN, USA) were set up together on the
des of the field plots. Automatic rain gauges were installed to collect
th 10
i
i t
l
Th
l
i
l
t
t t
Two pairs of rain gauges, an automatic tipping bucket rain gauge, and a manual rain
gauge (Productive Alternatives Inc., Fergus Falls, MN, USA) were set up together on the
west and the east sides of the field plots. Automatic rain gauges were installed to collect
the rainfall data with 10 min intervals. The manual rain gauges were also set up next to the
automatic rain gauges to ensure that rainfall measurements were accurate and consistent
during the study period. g
g
gauges to ensure that rainfall measurements w
2.6.2. Soil Water Potential and Soil Temperature Measurements g
g
gauges to ensure that rainfall measurements w
2.6.2. Soil Water Potential and Soil Temperature Measurements g
g
he study period. tential and Soil Temperature Measurements
k sensor measures the soil resistance which is related to soil matric
The watermark sensor measures the soil resistance, which is related to soil matric
potential. Based on the soil release curve, the soil water content can then be estimated
from the measured soil matric potential. As reported by the manufacturer, there are four
equations for the estimates of the soil water potential based on the measured resistance
and soil temperature (Irrometer Company, Riverside, CA, USA): ted
(1) ease curve, the soil water content can then be estimated
I f 1 > R > 0.55, ψ = −20(R ∗(1 + 0.18(T −24)) −0.55)
(1) Water 2021, 13, 1991 8 of 22 I f 1 > R > 0.55, ψ = −20(R ∗(1 + 0.18(T −24)) −0.55)
(2)
I f R > 8, ψ = −2.246 −5.239R(1 + 0.018(T −24)) −0.06756R2(1 + 0.018 (T −24))2
(3) I f 1 > R > 0.55, ψ = −20(R ∗(1 + 0.18(T −24)) −0.55)
(2) (2) I f 1 > R > 0.55, ψ = −20(R ∗(1 + 0.18(T −24)) −0.55)
I f R > 8, ψ = −2.246 −5.239R(1 + 0.018(T −24)) −0.06756R2(1 + 0.018 (T −24))2 (3) I f R < 0.55, ψ = 0
(4) (4) where R is the resistance in kiloohms, ψ is soil water potential in kPa, and T is the tempera-
ture in ◦C. However, for the sensors used on the controller, a default temperature (23.9 ◦C)
was used to estimate the potential instead of the above equations. Thus, the soil water potential was estimated from the resistance measured by the
sensor and the soil temperature measured by a soil temperature sensor, which was con-
structed from a Type E thermocouple (Omega Engineering Inc., Norwalk, CT, USA). The
two watermark sensors were installed in the middle of the plot at 15 and 30 cm depth along
with the two soil temperature sensors in each plot. The four sensors were located on the
opposite side of the plant from the soil water potential sensors used to control irrigation. They were used to monitor the soil matric potential and to ensure that the three replicated
plots in each treatment were all in similar soil moisture status. 2.6.3. Groundwater Three piezometers were installed at different locations in the field to monitor the
groundwater table in the study area (Figure 1). The piezometers were installed at 1.2 m
depth, while the top of the pipe was kept at about 22 cm above the ground. The water
levels were measured using an absolute pressure water-level sensor (Onset computer Corp,
Bourne, MA, USA), recorded in 30-min intervals. The water-level data were compensated
using the barometric pressure readings recorded by the NDAWN station, located about
675 m northwest of the field. g
g
gauges to ensure that rainfall measurements w
2.6.2. Soil Water Potential and Soil Temperature Measurements A total of 96 soil water potential and 96 soil temperature sensors were connected to
three CR1000 dataloggers (Campbell Scientific Inc., Logan, UT, USA) to collect the soil
water potential and soil temperature data for every 15 min such that 32 watermark sensors
and 32 soil temperature sensors were connected to each datalogger with the help of two
AM16/32B multiplexers (Campbell Scientific Inc., Logan, UT, USA), 10 W solar panels,
12 V battery, and other accessories. 2.6.4. Vegetables For the field experiments, Celebrity (Johnny’s Selected Seeds, Winslow, ME, USA) va-
riety was selected for tomato, which is disease resistant, high yield, and semi-determinate. Sangria variety (Agassiz Seed & Supply, West Fargo, ND, USA) was selected for water-
melon, which was chosen due to its suitability in a cold climate. Both seeds were planted
in a greenhouse and then transplanted in the field. The average management timelines in
dates are listed in Table 3 though there were some differences among different treatments. Table 3. Vegetable management timelines in 2019 and 2020. Tomato
Watermelon
2019
2020
2019
2020
Seeded
22 Apr
7 Apr
22 Apr
20 Apr
Transplanted
14 Jun
30 May
14 Jun
30 May
Fruit growth
28 Jun
24 Jun
22 Jul
28 Jun
Start harvest
19 Aug
24 Aug
6 Sep
5 Aug
End harvest
8 Oct
2 Oct
2 Oct
9 Sep
2 7 Vegetable Management Table 3. Vegetable management timelines in 2019 and 2020. 2.8. Field Observations Soil water potential, soil temperature, rainfall, the flow rate from the hydrant, flow
rate from each control unit, and groundwater levels were collected every week. Once signs
of fruit growth were experienced, the number of red and green tomatoes per plant and
number of watermelons were counted, while their sizes were also measured on a weekly
basis during the growing season. At the end of the season, after all fruits were harvested,
the above-ground biomass for each plant and the maximal length for watermelon were
measured in the field. Yield samples were collected from each plot for quality analysis. 2.7. Vegetable Management Topping, staking, fencing, weeding, and watering are some of the management tech-
niques used for tomato plants, whereas curling, weeding, and watering are for watermelon. Irrigation was applied manually until the plants were acclimated to the field conditions Water 2021, 13, 1991 9 of 22 9 of 22 after transplantation. Black plastic and landscape fabric mulches suppressed the weeds
more effectively, while clear plastic mulch did not suppress the weeds until the tempera-
tures increased, and weeds began to die due to high temperature exposure underneath the
mulch. A hand weeder was used to clear the weeds between the rows, while weeds on the
rows were removed manually. Chicken-wire fence was installed around the field to avoid
crop damage due to small animals, such as rabbits. p
g
In the middle of the growing season, when tomato plants grew taller and started
flowering, tomato tops were manually taken off to limit the growth of non-fruit biomass. In 2019, tomato plants were left unstacked, but in 2020, due to high wind damage, small
poles were used to stake and support the tomato plants from strong winds. This step
greatly reduced the tomato plant/twig damages caused by the wind for the remaining
growing season. Watermelons were curled at the vanes around each plant, which helped in
separating the plants to ensure the accuracy of field observations for each individual plant. 2.9. Fruit Quality Analysis Both tomato and watermelon samples were collected and blended to measure pH, elec-
trical conductivity (EC), and sugar content using a handheld pH meter (Hanna Instruments
Inc., Woonsocket, RI, USA), a pH/ISE benchtop multiparameter meter (ThermoFisher
Scientific Inc., Waltham, MA, USA), and a refractometer (Spectrum Technologies Inc., Plain-
field, IL, USA), respectively. The handheld refractometer measures the sugar content of
the sample in percentage (%). Fruit diameter and fruit weight were also measured for
tomatoes, whereas fruit length, diameter, and weight were measured for the watermelon
samples. 2.10. Statistical Analysis The statistical analysis was conducted using the Statistical Analysis System (SAS) to
evaluate the impact of mulch, irrigation, and the combined effect on fruit weight, diameter,
and each quality parameter. Weekly tomato yield and quality parameters were used in the
analysis of the dependent variables (weight, diameter, EC, pH, and sugar content) and the
independent variables (time, mulch, irrigation, and together). Two-way ANOVA analysis
was used to study the impact of mulch, irrigation, and the two together on fruit weight,
length, and quality parameters. 3.1. Weather Conditions Since
mulches can help conserve the soil moisture in the root zone, reduce the evaporation, and
increase the transpiration [37], it further eliminated the need for irrigation in 2019. In 2020,
the PET (977 mm) was much higher (573 mm) than the rainfall (404 mm), which required
much more frequent irrigation in all plots. The vegetables were irrigated a few times under
the DI PF, DI NM, and DI CP treatment in 2019, whereas many irrigation events occurred
in 2020 due to large gaps between the rainfall and the PET. This strongly represented the
need for irrigation in low or variable rainfall seasons and the importance of smart irrigation
to avoid over or under irrigation. 3.2. Soil Moisture and Temperature
Two soil water potential sensors were used monitor the soil water status potential in
each plot, and two sensors were connected to the controller to trigger the drip irrigation. They were same type of sensors and were located at the nearby locations from the plant. The average soil water potential at 15 and 30 cm from the three plots for each treatment
are plotted in Figures 3 and 4 for tomato plants in 2019 and 2020, respectively, whereas
Figures 5 and 6 are for the watermelon plots. The irrigation events are also marked on the
figures. Typically, tomato and watermelon require 400 to 600 mm of water to grow in the
sub-humid region. Due to the frequent and large amount of rainfall in the 2019 growing
season, there was a small difference (281 mm) between the rainfall (558 mm) and the
PET (839 mm), and less irrigation was required to meet the plants’ water demands. Since
mulches can help conserve the soil moisture in the root zone, reduce the evaporation, and
increase the transpiration [37], it further eliminated the need for irrigation in 2019. In 2020,
the PET (977 mm) was much higher (573 mm) than the rainfall (404 mm), which required
much more frequent irrigation in all plots. The vegetables were irrigated a few times under
the DI PF, DI NM, and DI CP treatment in 2019, whereas many irrigation events occurred
in 2020 due to large gaps between the rainfall and the PET. This strongly represented the
need for irrigation in low or variable rainfall seasons and the importance of smart irrigation
to avoid over or under irrigation. 3.1. Weather Conditions During the two-year experiment, the weather conditions were extremely different,
with a very wet year in 2019 and a very dry year in 2020. Thus, the irrigation system was
tested in both wet and dry years. Rainfall, potential evapotranspiration (PET), and air and
soil temperatures are some of the major weather parameters that may have had a certain
impact on this research. The different monthly average weather parameters are shown in
Table 4. 10 of 22 Water 2021, 13, 1991 10 of 22 Table 4. Major weather parameters (Tmax, monthly average maximal air temperature; Tmin, monthly average minimal air
temperature; Tavg, monthly average air temperature; Tsoil, monthly average bare soil temperature at 10 cm depth; Uavg,
monthly average wind speed; Umax, monthly average maximum wind speed; Rs, monthly average incoming shortwave
radiation; PET, monthly average total daily potential evapotranspiration amount; and Rain, monthly average total rainfall)
during the field experiment. 3.1. Weather Conditions Year
Month
Tmax
(◦C)
Tmin
(◦C)
Tavg
(◦C)
Tsoil
(◦C)
Uavg
(m/s)
Umax
(m/s)
Rs
(MJ/m2)
PET
(mm/day)
Rain
(mm/mon)
2019
May
17.57
5.30
11.43
11.28
7.18
21.50
18.79
5.03
69.65
June
25.62
13.71
19.67
19.45
7.03
23.47
20.75
6.41
82.75
July
27.86
17.08
22.47
23.17
5.83
20.08
23.25
6.09
121.26
August
24.77
14.82
19.80
20.82
6.03
19.16
18.91
4.76
89.69
September
21.61
12.10
16.85
16.87
6.99
22.67
12.42
3.24
106.81
October
9.71
2.28
5.99
7.55
8.72
22.60
7.61
1.84
87.78
Avg/Total
21.19
10.88
16.04
16.52
6.96
21.58
16.96
4.56
557.94
2020
May
18.34
6.95
12.64
12.52
8.01
23.32
21.14
5.80
37.92
June
28.29
15.58
21.94
20.20
9.00
27.54
23.63
7.89
66.75
July
28.62
17.72
23.17
23.28
6.38
24.04
24.08
6.51
133.38
August
26.88
15.46
21.17
22.30
5.88
20.90
18.93
4.93
122.33
September
20.81
8.97
14.89
16.26
7.12
22.88
13.61
4.11
22.10
October
9.17
−1.18
4.00
7.79
8.03
23.88
9.44
2.60
21.49
Avg/Total
22.02
10.58
16.30
17.06
7.40
23.76
18.47
5.31
403.97 Year
Month
Tmax
(◦C)
Tmin
(◦C)
Tavg
(◦C)
Tsoil
(◦C)
Uavg
(m/s)
Umax
(m/s)
Rs
(MJ/m2)
PET
(mm/day)
Rain
(mm/mon)
2019
May
17.57
5.30
11.43
11.28
7.18
21.50
18.79
5.03
69.65
June
25.62
13.71
19.67
19.45
7.03
23.47
20.75
6.41
82.75
July
27.86
17.08
22.47
23.17
5.83
20.08
23.25
6.09
121.26
August
24.77
14.82
19.80
20.82
6.03
19.16
18.91
4.76
89.69
September
21.61
12.10
16.85
16.87
6.99
22.67
12.42
3.24
106.81
October
9.71
2.28
5.99
7.55
8.72
22.60
7.61
1.84
87.78
Avg/Total
21.19
10.88
16.04
16.52
6.96
21.58
16.96
4.56
557.94
2020
May
18.34
6.95
12.64
12.52
8.01
23.32
21.14
5.80
37.92
June
28.29
15.58
21.94
20.20
9.00
27.54
23.63
7.89
66.75
July
28.62
17.72
23.17
23.28
6.38
24.04
24.08
6.51
133.38
August
26.88
15.46
21.17
22.30
5.88
20.90
18.93
4.93
122.33
September
20.81
8.97
14.89
16.26
7.12
22.88
13.61
4.11
22.10
October
9.17
−1.18
4.00
7.79
8.03
23.88
9.44
2.60
21.49
Avg/Total
22.02
10.58
16.30
17.06
7.40
23.76
18.47
5.31
403.97
Typically, tomato and watermelon require 400 to 600 mm of water to grow in the
sub-humid region. Due to the frequent and large amount of rainfall in the 2019 growing
season, there was a small difference (281 mm) between the rainfall (558 mm) and the
PET (839 mm), and less irrigation was required to meet the plants’ water demands. 3.2. Soil Moisture and Temperature Two soil water potential sensors were used monitor the soil water status potential in
each plot, and two sensors were connected to the controller to trigger the drip irrigation. They were same type of sensors and were located at the nearby locations from the plant. The average soil water potential at 15 and 30 cm from the three plots for each treatment
are plotted in Figures 3 and 4 for tomato plants in 2019 and 2020, respectively, whereas
Figures 5 and 6 are for the watermelon plots. The irrigation events are also marked on the
figures. 11 of 22
11 of 23 Water 2021, 13, 1991
Water 2021 13 x FO igure 3. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2019 field experim
nder drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c)
lastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF
lack plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). ure 3. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2019 field experime
der drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear plas
lch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f) bla
stic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 3. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2019 field experimen
under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear
plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f
black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 3. 3.2. Soil Moisture and Temperature Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2019 field experiment
under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear plastic
mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f) black
plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). 12 of 22
12 of 23 12 of 22
12 of 23 Water 2021, 13, 1991
Water 2021, 13, x F Figure 4. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2020 field experim
under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) cl
plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF)
black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). ure 4. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2020 field experim
der drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear pla
lch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f) bl
stic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 4. Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2020 field experiment
under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear
plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f)
black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 4. 3.2. Soil Moisture and Temperature Daily average soil matric potential (09:00 am) at 15 and 30 cm deep in tomato plots during 2020 field experiment
under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP), (c) clear plastic
mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI PF), (f) black
plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). 13 of 22
13 of 23 13 of 22
13 of 23 Water 2021, 13, 1991
W t
2021 13
F Figure 5. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 fiel
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulc
(NI PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). gure 5. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
xperiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP)
) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (N
F), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 5. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP),
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch
(NI PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 5. 3.2. Soil Moisture and Temperature Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP),
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI
PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). 14 of 22
14 f 23 Water 2021, 13, 1991 Figure 6. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 fiel
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulc
(NI PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). igure 6. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 fiel
xperiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP
) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (N
F), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 6 Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
Fi
6
D il
il
i
i l (08 00
)
15
d 30
d
i
l
l
d
i
2019 fild Figure 6. Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP),
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch
(NI PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Figure 6. 3.2. Soil Moisture and Temperature In addition, the soil potential values under the
PF are close to those under the NM, but the PF controlled weed growth completely. The
similar water status between PF and NM is probably due to the porous feature of the PF
that did not prevent any water infiltration. In both years’ field experiments, the average soil
water potential was recorded the lowest (driest) in July, which was during the flowering
stage, while the highest average temperature was also recorded in July. Irrigation events in 2020 were plotted along with the average soil water potential
in Figures 4 and 6. For tomatoes, only the DI PF treatment was irrigated three times,
and DI NM treatment two times in 2019, whereas DI BP and DI CP treatments did not
irrigate a single time. This could be due to less evaporation under the plastic mulches
compared to the landscape fabric and bare soil. To evaluate the evaporation from the soil
surface under plastic mulches, Ref. [39] even punched holes through the mulch at different
rates and found that the evaporation rate was reduced 69.26% if there were no holes in
the mulch; if 7.24% of the mulch areas had holes, the evaporation was only reduced by
20.5%. This showed the impact of plastic mulch on reduction of evaporation rate. However,
in 2020, DI NM treatment was irrigated four times, followed by DI PF 7 times, DI CP
13 times, and DI BP 20 times. Total number of irrigation events that occurred in 2020 for BP
mulch treatment was the highest, which could be due to high crop transpiration under the
mulches [36]. Additionally, heavy winds that occurred during the field experiments lead to
mulch flapping, which could increase the evaporation rate whenever the soil surface was
exposed [39]. p
Similarly, the DI PF treatment irrigated three times, and the DI CP treatment irrigated
once in watermelon plots during the 2019 field experiment, whereas in 2020, the irrigation
events occurred many times, with DI CP 12 times, DI NM 19 times, and DI BP and DI
PF 21 times. This clearly showed that the watermelon consumed much more water than
tomato in 2020. In addition, the increase in the number of irrigation events that occurred
for watermelon compared to tomato is possibly due to the higher watermelon transpiration
rate. Ref. 3.2. Soil Moisture and Temperature Daily average soil matric potential (08:00 am) at 15 and 30 cm deep in watermelon plots during 2019 field
experiment under drip irrigation (DI) treatment with (a) landscape fabric mulch (DI PF), (b) black plastic mulch (DI BP),
(c) clear plastic mulch (DI CP), (d) no mulch (DI NM), and no irrigation treatment (NI) with (e) landscape fabric mulch (NI
PF), (f) black plastic mulch (NI BP), (g) clear plastic mulch (NI CP), and (h) no mulch (NI NM). Water 2021, 13, 1991 15 of 22 15 of 22 In most cases, the soil water potential at 15 cm is smaller (drier) than that at 30 cm in
2019 for both tomato and watermelon (Figures 3 and 5), while less irrigation was applied. In contrast, the soil water potential at 30 cm is smaller (drier) than that at 15 cm in 2020
when frequent irrigations were applied (Figures 4 and 6). With less irrigation, the soil
evaporation from the surface can dry up the surface soil, which can result in a drier surface
and low soil moisture at 15 cm depth. With more irrigation, water was applied from the
surface, leaving a wet surface layer; in addition, water was consumed more at the deeper
layer. y
The irrigation events were marked on the figures. Theoretically, irrigation system
is triggered whenever the average soil water potential is less (drier) than the threshold
value. However, the threshold value was not a fixed value but rather ranged from −5 kPa
to −80 kPa if using the soil potential from the average of all six monitoring sensors. The
variable values were probably caused by several reasons. The first would be the variabilities
among the sensors, six monitoring sensors vs. two sensors, linked to the controller to trigger
the irrigation. The second reason was due to the difference in irrigation and crop water
requirement that caused the soil potential difference. The last reason would be a lack of
soil-specific calibration between the set point on the controller and the corresponding soil
potential. As reported by [38], the sensors were calibrated by the manufacture and worked
as well as the on-site calibration and were sufficient for irrigation scheduling. g
g
The soil water potential under plastic mulches (both CP and BP) are larger (wetter)
than those under PF and NM treatments. 3.2. Soil Moisture and Temperature [40] conducted a field experiment to study the effect of plastic mulches and wheat
straw compared to bare soil on water loss through evaporation and transpiration from the
soil surface and found that the water loss due to evaporation was higher under bare soil
than that under plastic mulches. However, the transpiration rate was higher under plastic
mulches than that under bare soil. Overall, the water loss by transpiration loss under mulch
was higher than that by evaporation under bare soil, which results in a higher water loss
with plastic mulch due to a much healthier plant and higher transpiration rate. Compared Water 2021, 13, 1991 16 of 22 16 of 22 to 2019, the yield and quality of the produce from plastic mulches with irrigation treatment
was higher in 2020. to 2019, the yield and quality of the produce from plastic mulches with irrigation treatment
was higher in 2020. g
The soil temperature variation under different mulches was compared, and the results
in 2019 and 2020 with no irrigation (NI) are shown in Table 5. p
p
in 2019 and 2020 with no irrigation (NI) are shown in Table 5. Table 5. Monthly average soil temperature (◦C) under clear plastic mulch (CP), black plastic mulch (BP), landscape fabric
mulch (PF), and no mulch (NM) in no-irrigation plots. 3.2. Soil Moisture and Temperature Crop
Year
Treatment
Depth (cm)
June
July
August
September
October
Average
Tomato
2019
PF
15
20.74
23.09
20.50
16.71
10.24
18.26
BP
15
22.95
25.31
21.81
17.71
11.22
19.80
CP
15
24.05
27.48
24.90
19.13
12.30
21.57
NM
15
21.72
23.23
21.44
17.12
9.87
18.68
PF
30
18.56
21.40
20.33
16.93
11.82
17.81
BP
30
20.10
23.00
20.93
17.51
12.45
18.80
CP
30
22.93
26.00
23.68
18.83
12.78
20.84
NM
30
19.13
21.86
20.70
17.05
10.97
17.94
2020
PF
15
20.45
23.22
21.97
16.42
-
20.52
BP
15
21.55
23.85
22.18
16.65
-
21.06
CP
15
22.71
24.83
23.10
17.77
-
22.10
NM
15
20.69
23.44
21.72
16.07
-
20.48
PF
30
21.63
24.50
22.28
15.81
-
21.05
BP
30
23.01
24.77
22.05
16.20
-
21.51
CP
30
24.17
25.96
23.49
17.43
-
22.76
NM
30
21.50
24.57
22.09
15.35
-
20.88
Watermelon
2019
PF
15
20.49
22.43
20.17
16.68
10.66
18.09
BP
15
23.12
24.20
20.47
17.16
11.09
19.21
CP
15
25.94
25.70
21.13
17.61
11.85
20.45
NM
15
21.29
22.68
19.87
16.44
10.00
18.06
PF
30
18.63
21.23
19.88
16.74
11.68
17.63
BP
30
20.12
22.45
20.33
17.44
12.28
18.53
CP
30
22.28
24.13
20.74
17.55
12.43
19.43
NM
30
19.37
21.56
19.72
16.60
11.02
17.65
2020
PF
15
21.05
22.94
21.64
16.20
-
20.46
BP
15
21.53
22.98
21.62
16.77
-
20.73
CP
15
23.19
23.14
21.81
17.50
-
21.41
NM
15
20.85
22.52
20.81
16.23
-
20.10
PF
30
21.96
22.78
21.13
15.36
-
20.31
BP
30
23.08
23.92
21.91
16.05
-
21.24
CP
30
24.17
23.24
21.82
17.22
-
21.61
NM
30
22.09
22.85
21.08
15.73
-
20.44 Table 5. Monthly average soil temperature (◦C) under clear plastic mulch (CP), black plastic mulch (BP), l
mulch (PF), and no mulch (NM) in no-irrigation plots. verage soil temperature (◦C) under clear plastic mulch (CP), black plastic mulch (BP), landscape fabric
mulch (NM) in no-irrigation plots. During the field experiment, the highest monthly average air temperature was recorded
in July with 22.47 and 23.17 ◦C in 2019 and 2020, respectively, while the minimum air
temperature was found in October (Table 4). During the experiment, soil temperature at
both depths were recorded the highest in July in both 2019 and 2020 for all treatments,
which corresponded with the highest air temperature in July. 3.3. Vegetable Yield Tomato harvesting started in August and ended in October in both years and lasted
for 50 and 39 days in 2019 and 2020, respectively. The short harvesting period in 2020 was
possibly due to the strong winds earlier in the season that damaged some plants and twigs
and delayed the fruit development. For watermelon, the fruit harvest period lasted for 26
and 35 days in 2019 and 2020, respectively. For the yield estimation, considering 9.29 m2 as
the area required for each plot, the total number of plants that can be grown for a hectare
was calculated, and the yield was calculated based on the total number of ripened fruits
per plot, total number of plants per plot, and the average weight of a sample. The total
yields for tomato and watermelon are summarized for each treatment (Table 6). Table 6. Tomato and watermelon yield obtained from drip irrigation (DI) and no irrigation (NI)
under clear plastic mulch (CP), black plastic mulch (BP), landscape fabric mulch (PF), and no mulch
(NM) treatment in 2019 and 2020. Table 6. Tomato and watermelon yield obtained from drip irrigation (DI) and no irrigation (NI)
under clear plastic mulch (CP), black plastic mulch (BP), landscape fabric mulch (PF), and no mulch
(NM) treatment in 2019 and 2020. Treatment
Tomato (Mg ha−1)
Watermelon (Mg ha−1)
Year
2019
2020
2019
2020
DI BP
26.38
40.24
55.11
144.67
DI CP
21.1
22.4
69.43
165.55
DI PF
23.86
34.31
35.01
153.95
DI NM
29.77
34.28
28.15
147.47
NI BP
34.32
27.56
44.75
132.03
NI CP
27.46
25.91
79.30
109.62
NI PF
29.84
23.91
30.89
123.54
NI NM
41.63
18.54
38.72
113.91
Average
29.3
28.4
47.67
136.34 In 2019, with frequent and sufficient rainfall, it was expected that there should be
no yield difference between the DI and the NI treatments. If there were any differences,
it should have been caused by other factors, such as mulch covers, water level, spatial
variation, etc. In 2020, however, since the irrigation was frequently applied, a higher yield
difference was expected between the DI and the NI treatments. For tomato, the yield for
DI treatment was around 36% higher than those with NI, while for watermelon, a 27.6%
higher yield was obtained for DI treatments compared to that for NI treatments. 3.2. Soil Moisture and Temperature The monthly average soil temperature of each NI treatment was used to evaluate
the temperature changes due to different mulches. In tomato plots, the average soil
temperature under the CP treatment recorded the highest soil temperature throughout
the season and among all treatments. The average soil temperature at 15 and 30 cm
under the CP was 2.89 ◦C and 2.9 ◦C higher than that under the NM treatment in 2019. However, for watermelon, the highest temperature increase was in June, where the soil
temperature at 15 cm depth under the CP was 4.65 and 2.34 ◦C higher than that under the
NM field in 2019 and 2020. The soil temperature increase at 30 cm depth was only 2.91
and 2.08 ◦C in 2019 and 2020, respectively. Overall, it was found that the monthly average
soil temperature under the CP was always higher than any other treatments in both years. Thermal properties of the material have a major impact on soil temperature variation on Water 2021, 13, 1991 17 of 22 different mulch types. Black plastic absorbs the longwave radiation and transmits the
absorbed energy to the soil through conduction, whereas clear plastic transmits 85 to 95%
of the absorbed solar radiation and controls the heat loss to the atmosphere by blocking
the longwave infrared radiation which keeps the surface warm [13]. 3.4. Vegetable Quality Fruit-quality parameters, such as EC, pH, and sugar content, were measured from the
tomato and watermelon samples collected during the field experiments, and the average of
each parameter are shown in Tables 7 and 8, respectively. Table 7. Average quality parameters measured from tomato samples collected during 2019 and 2020. pH
EC (mS/cm)
Sugar Content (%)
2019
2020
2019
2020
2019
2020
DI BP
4.1
4.1
4.1
4.0
4.9
5.8
DI CP
4.1
4.0
4.0
4.0
4.9
7.3
DI PF
4.1
4.0
4.0
4.0
4.9
5.6
DI NM
4.1
4.2
4.0
4.1
4.9
5.8
NI BP
4.2
4.0
4.0
4.0
5.4
8.2
NI CP
4.2
3.9
4.0
4.1
4.9
5.7
NI PF
4.2
4.0
4.2
4.0
4.9
6.0
NI NM
4.1
4.2
4.1
4.0
5.1
6.2
Table 8. Average quality parameters measured from watermelon samples collected during 2019
and 2020. pH
EC (dS/cm)
Sugar content (%)
2019
2020
2019
2020
2019
2020
DI BP
5.7
5.2
3.3
3.6
9.4
10.3
DI CP
5.9
5.2
3.4
3.7
10.2
11.0
DI PF
5.4
5.0
3.2
3.8
8.2
10.2
DI NM
5.4
5.2
3.2
3.8
8.7
9.9
NI BP
5.8
5.3
2.9
3.3
10.1
10.2
NI CP
5.8
5.1
3.2
3.5
9.8
10.3
NI PF
5.5
5.2
3.1
3.6
9.1
9.8
NI NM
5.7
5.3
3.2
3.5
9.6
10.2 Table 7. Average quality parameters measured from tomato samples collected during 2019 and 2020. pH
EC (mS/cm)
Sugar Content (%)
2019
2020
2019
2020
2019
2020
DI BP
4.1
4.1
4.1
4.0
4.9
5.8
DI CP
4.1
4.0
4.0
4.0
4.9
7.3
DI PF
4.1
4.0
4.0
4.0
4.9
5.6
DI NM
4.1
4.2
4.0
4.1
4.9
5.8
NI BP
4.2
4.0
4.0
4.0
5.4
8.2
NI CP
4.2
3.9
4.0
4.1
4.9
5.7
NI PF
4.2
4.0
4.2
4.0
4.9
6.0
NI NM
4.1
4.2
4.1
4.0
5.1
6.2
Table 8. Average quality parameters measured from watermelon samples collected during 2019
and 2020 Table 7. Average quality parameters measured from tomato samples collected during 2019 and 2020. Table 8. Average quality parameters measured from watermelon samples collected during 2019
and 2020. Table 8. Average quality parameters measured from watermelon samples collected during 2019
and 2020. 3.3. Vegetable Yield Among
the different treatments, the highest yield for tomato was from the NI NM treatment in 2019
(Table 6), probably due to fewer survival plants in the plot, which resulted in large spaces
between plants. The actual plot with the highest yield was from DI BP, at 40.24 Mg ha−1. The average tomato yield was slightly higher in 2019 than that in 2020, indicating that
sufficient rainfall benefited the crop yield. In addition, the tomatoes were transplanted late
in 2019, when the mulches might not have been really needed. For watermelon, the highest
yield was recorded under the DI CP treatment with 165.55 Mg ha−1 in 2020. The average
yield was also three times higher in 2020 than that in 2019, indicating that irrigation and
warmer weather were the key factors for a higher watermelon yield. However, comparing
to all other treatments, the irrigation demand under the DI CP was the least though it
produced the highest yield. This was similar to what [41] found: the maximum irrigation
water use efficiency for watermelon was found under clear plastic mulch compared to
black plastic and grey-black film mulches. Water 2021, 13, 1991 18 of 22 18 of 22 3.4. Vegetable Quality pH
EC (dS/cm)
Sugar content (%)
2019
2020
2019
2020
2019
2020
DI BP
5.7
5.2
3.3
3.6
9.4
10.3
DI CP
5.9
5.2
3.4
3.7
10.2
11.0
DI PF
5.4
5.0
3.2
3.8
8.2
10.2
DI NM
5.4
5.2
3.2
3.8
8.7
9.9
NI BP
5.8
5.3
2.9
3.3
10.1
10.2
NI CP
5.8
5.1
3.2
3.5
9.8
10.3
NI PF
5.5
5.2
3.1
3.6
9.1
9.8
NI NM
5.7
5.3
3.2
3.5
9.6
10.2 pH is one of the most important parameters for tomatoes as well as sugar content
for watermelon. Soil temperature and soil water content are the most important factors
for yield and the quality of produce. Weather parameters were completely different in
2020 when compared to 2019. Tomato yield in 2020 was heavier and juicier than that in
2019. However, watermelon yield in 2020 was heavier with a higher sugar content (%) than
that in 2019. This could be due to the change in weather parameters, such as precipitation,
temperature, solar radiation, wind speed, etc. p
p
Quality parameters obtained during 2019 and 2020 field experiments were used to
evaluate the effect of mulch and irrigation as well as their interactions. Results obtained
from the statistical analysis are shown in Table 9. For tomatoes, in 2019, most of the quality
parameters were significantly different (p < 0.05) under mulches than the irrigation and
their interactions. The average fruit weight and diameter under the DI treatments are
significantly higher (p < 0.05) than those under irrigation and their interactions. The EC and
pH under the BP treatments are higher (p < 0.05) than those under other treatments. The pH
of the samples under DI BP treatment was better than that under other treatments. Quality
parameters under PF showed no significant difference than that under the NM treatments,
but the BP and CP produced the best quality produce. Overall quality parameters were
better under DI CP treatments, whereas soil temperature was warmer under CP, which
had a positive impact on the quality. Water 2021, 13, 1991 19 of 22 Table 9. Tomato and watermelon quality parameter analysis on mulch, irrigation, and the interaction of mulch and irrigation
in 2019 and 2020 using analysis of variance, with * at p < 0.05, ** at p < 0.01, and *** at p < 0.001. 3.4. Vegetable Quality Thus, polyethylene mulches had a positive
impact on yield, weight, and other quality parameters, and these results were similar to
results of other researches [11,20,42]. In 2020, when less rainfall was received in the field,
irrigation played an important role for the weight, size, and quality parameters. Therefore,
the combined drip and mulch is a proven practice for a successful vegetable growth in
North Dakota. Table 9. Tomato and watermelon quality parameter analysis on mulch, irrigation, and the interaction of mulch and irrigation
in 2019 and 2020 using analysis of variance, with * at p < 0.05, ** at p < 0.01, and *** at p < 0.001. Vegetable
Year
Source
Weight (g)
Diameter (mm)
EC (dS/m)
pH
Sugar (%)
Tomato
2019
Mulch
**
***
**
Irrigation
Interaction
2020
Mulch
**
*
Irrigation
**
***
Interaction
*
Watermelon
2019
Mulch
***
*
**
*
Irrigation
Interaction
2020
Mulch
**
***
*
Irrigation
**
Interaction Table 9. Tomato and watermelon quality parameter analysis on mulch, irrigation, and the interaction of mulch and irrigation
in 2019 and 2020 using analysis of variance, with * at p < 0.05, ** at p < 0.01, and *** at p < 0.001. Table 9. Tomato and watermelon quality parameter analysis on mulch, irrigation, and the interaction of mulch and irrigation
in 2019 and 2020 using analysis of variance, with * at p < 0.05, ** at p < 0.01, and *** at p < 0.001. Similarly, watermelon parameters under DI and NI plots were almost similar in 2019
(Table 9). Whereas, the average watermelon weight and diameter under the CP were
significantly different from the other treatments (p < 0.01). The EC were affected (p < 0.05)
by the irrigation treatment, whereas the sugar content was affected (p < 0.05) by mulch. Watermelon under DI CP is much heavier and produced the highest yield compared to all
other treatments. It is worth noticing that mulch significantly affected the weight, size, and
quality parameters in 2019 for both vegetables. Thus, polyethylene mulches had a positive
impact on yield, weight, and other quality parameters, and these results were similar to
results of other researches [11,20,42]. In 2020, when less rainfall was received in the field,
irrigation played an important role for the weight, size, and quality parameters. Therefore,
the combined drip and mulch is a proven practice for a successful vegetable growth in
North Dakota. 3.4. Vegetable Quality Vegetable
Year
Source
Weight (g)
Diameter (mm)
EC (dS/m)
pH
Sugar (%)
Tomato
2019
Mulch
**
***
**
Irrigation
Interaction
2020
Mulch
**
*
Irrigation
**
***
Interaction
*
Watermelon
2019
Mulch
***
*
**
*
Irrigation
Interaction
2020
Mulch
**
***
*
Irrigation
**
Interaction
Similarly, watermelon parameters under DI and NI plots were almost similar in 2019
(Table 9). Whereas, the average watermelon weight and diameter under the CP were
significantly different from the other treatments (p < 0.01). The EC were affected (p < 0.05)
by the irrigation treatment, whereas the sugar content was affected (p < 0.05) by mulch. Watermelon under DI CP is much heavier and produced the highest yield compared to all
other treatments. It is worth noticing that mulch significantly affected the weight, size, and
quality parameters in 2019 for both vegetables. Thus, polyethylene mulches had a positive
impact on yield, weight, and other quality parameters, and these results were similar to
results of other researches [11,20,42]. In 2020, when less rainfall was received in the field,
irrigation played an important role for the weight, size, and quality parameters. Therefore,
the combined drip and mulch is a proven practice for a successful vegetable growth in
North Dakota. Table 9. Tomato and watermelon quality parameter analysis on mulch, irrigation, and the interaction of mulch and irrigation
in 2019 and 2020 using analysis of variance, with * at p < 0.05, ** at p < 0.01, and *** at p < 0.001. Vegetable
Year
Source
Weight (g)
Diameter (mm)
EC (dS/m)
pH
Sugar (%)
Tomato
2019
Mulch
**
***
**
Irrigation
Interaction
2020
Mulch
**
*
Irrigation
**
***
Interaction
*
Watermelon
2019
Mulch
***
*
**
*
Irrigation
Interaction
2020
Mulch
**
***
*
Irrigation
**
Interaction
Similarly, watermelon parameters under DI and NI plots were almost similar in 2019
(Table 9). Whereas, the average watermelon weight and diameter under the CP were
significantly different from the other treatments (p < 0.01). The EC were affected (p < 0.05)
by the irrigation treatment, whereas the sugar content was affected (p < 0.05) by mulch. Watermelon under DI CP is much heavier and produced the highest yield compared to all
other treatments. It is worth noticing that mulch significantly affected the weight, size, and
quality parameters in 2019 for both vegetables. In 2019, the
the water requir
4. Conclusions the water requirement for the two vegetables. The soil water status, as shown in Figures
3 and 5, also proved that the soil was kept wet and did not go beyond the soil potential
threshold, which restricted the controller to trigger the irrigation. In 2020, with only 366
mm of rainfall during the experimental period and a warmer temperature (Table 4), more
water was needed to meet the crop water demand, while the SMS-based controller
performed the same with frequent irrigation events. For tomato, drip irrigation under the
BP treatment was turned on very often and applied a larger amount of water than any
other treatment, with the order of the irrigation times and amounts as BP > CP > PF > NM. Black mulch and drip irrigation performed better on tomato growth, yield, and quality,
which was reported before in [25,43]. For watermelon, the irrigation times and amounts
followed the order of BP > PF > NM > CP. For watermelon, the accumulative irrigation
amount under the CP treatment was the least compared to all other treatments, but the
CP plots produced the highest yield and best-quality watermelon (Table 6). This was
because the CP thermal properties maintained the warm temperature and increased the
transpiration rate, while watermelon needed the warm temperature and the optimal
amount of water for its production. This finding was different from another study [20] in
which the best yield was obtained from black plastic mulch and drip irrigation. 4. Conclusions
A
fi ld
i
d
d
h
i i i
i
A two-year field experiment was conducted to test the automatic irrigation systems
under clear plastic, black plastic, landscape fabric, and no mulch for tomato and water-
melon production in North Dakota. The irrigation was scheduled based on the preset
soil potential threshold at the wet zone in one of the three plots for each treatment. Two
soil potential and two soil temperature sensors were installed in each plot to monitor
the soil water potential status. The results showed that the irrigation was automatically
triggered each time the soil potential was above the preset threshold. With frequent rainfall
in 2019, the irrigation was barely applied, but the mulch effect was more evident. 3.5. Irrigation Monthly cumulative rainfall (mm) with cumulative irrigation amounts for fabric landscape (PF), clear plastic
(CP), black plastic (BP), and no mulch (NM) during two-year field experiments with (a) 2019 tomato, (b) 2019 watermelon,
(c) 2020 tomato, and (d) 2020 watermelon. Figure 7. Monthly cumulative rainfall (mm) with cumulative irrigation amounts for fabric landscape (PF), clear plastic
(CP), black plastic (BP), and no mulch (NM) during two-year field experiments with (a) 2019 tomato, (b) 2019 watermelon,
(c) 2020 tomato, and (d) 2020 watermelon. 3.5. Irrigation The accumulative rainfall during the field experiments was plotted against the total
amount of irrigation water under each treatment (Figure 7). In 2019, the field received a total of 490 mm of rainfall, which was sufficient to
meet the water requirement for the two vegetables. The soil water status, as shown in
Figures 3 and 5, also proved that the soil was kept wet and did not go beyond the soil
potential threshold, which restricted the controller to trigger the irrigation. In 2020, with
only 366 mm of rainfall during the experimental period and a warmer temperature (Table 4),
more water was needed to meet the crop water demand, while the SMS-based controller
performed the same with frequent irrigation events. For tomato, drip irrigation under the
BP treatment was turned on very often and applied a larger amount of water than any other
treatment, with the order of the irrigation times and amounts as BP > CP > PF > NM. Black
mulch and drip irrigation performed better on tomato growth, yield, and quality, which
was reported before in [25,43]. For watermelon, the irrigation times and amounts followed
the order of BP > PF > NM > CP. For watermelon, the accumulative irrigation amount
under the CP treatment was the least compared to all other treatments, but the CP plots
produced the highest yield and best-quality watermelon (Table 6). This was because the CP
thermal properties maintained the warm temperature and increased the transpiration rate,
while watermelon needed the warm temperature and the optimal amount of water for its
production. This finding was different from another study [20] in which the best yield was
obtained from black plastic mulch and drip irrigation. Water 2021, 13, 1991
W t
2021 13
FOR 20 of 22
20 of 23 20 of 22
20 of 23 Figure 7. Monthly cumulative rainfall (mm) with cumulative irrigation amounts for fabric landscape (PF), clear plastic
(CP), black plastic (BP), and no mulch (NM) during two-year field experiments with (a) 2019 tomato, (b) 2019 watermelon,
(c) 2020 tomato, and (d) 2020 watermelon. Figure 7. Monthly cumulative rainfall (mm) with cumulative irrigation amounts for fabric landscape (PF), clear plastic
(CP), black plastic (BP), and no mulch (NM) during two-year field experiments with (a) 2019 tomato, (b) 2019 watermelon,
(c) 2020 tomato, and (d) 2020 watermelon. Figure 7. In 2019, the
the water requir
4. Conclusions Conflicts of Interest: The authors declare no conflict of interest. Conflicts of Interest: The authors declare no conflict of interest. References 1. NDSU Extension. North Dakota Agriculture. Ag Statistics No. 87. 2018. Available online: https://www.nass.usda.gov/Statistics_
by_State/North_Dakota/Publications/Annual_Statistical_Bulletin/2018/ND-Annual-Bulletin18.pdf (accessed on 8 July 2021). 1. NDSU Extension. North Dakota Agriculture. Ag Statistics No. 87. 2018. Available online: https://www.nass.usda.gov/Statistics_
by_State/North_Dakota/Publications/Annual_Statistical_Bulletin/2018/ND-Annual-Bulletin18.pdf (accessed on 8 July 2021). 2. USDA NASS. Census of Agriculture for Tomato and Watermelon for 2012 and 2017. Available online: https://www.nass.usda. gov/Publications/AgCensus/2017/Full_Report/Volume_1,_Chapter_1_State_Level/North_Dakota/st38_1_0036_0036.pdf (ac-
cessed on 8 July 2021). J
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3. Aguyoh, J.; Taber, H.G.; Lawson, V. Maturity of fresh market sweet corn with direct seeded plants, transplants, clear plastic
mulch, and row cover combinations. HortTechnology 1999, 9, 420–425. [CrossRef] 4. Kwabiah, A.B. Growth, maturity, and yield responses of silage maize (Zea mays L.) to hybrid, planting date and plastic mulch. J. New Seeds. 2005, 7, 37–59. [CrossRef] 5. Dı´az-Pe´rez, J.C.; Batal, K.D. Colored plastic film mulches affect tomato growth and yield via changes in root-zone temperature. J. Amer. Soc. Hort. Sci. 2002, 127, 127–135. [CrossRef] 6. Kara, B.; Atar, B. Effects of mulch practices on fresh ear yield and yield components of sweet corn. Turkish J. Agric. For. 2013, 37,
281–287. 7. Kwabiah, A.B. Growth and yield of sweet corn (Zea mays L.) cultivars in response to planting
short-season environment. Sci. Hortic. 2004, 102, 147–166. [CrossRef] 8. Liu, C.A.; Jin, S.L.; Zhou, L.M.; Jia, Y.; Li, F.M.; Xiong, Y.C.; Li, X.G. Effects of plastic film mulch and tillage on maize productivity
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9. Zhang, S.; Li, P.; Yang, X.; Wang, Z.; Chen, X. Effects of tillage and plastic mulch on soil water, growth and yield of spring-sown
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10. Zhou, L.M.; Li, F.M.; Jin, S.L.; Song, Y. How two ridges and the furrow mulched with plastic film affect soil water, soil temperature
and yield of maize on the semiarid Loess Plateau of China. Field Crop. Res. 2009, 113, 41–47. [CrossRef] 11. Bye, H.D. Sweet Corn Production with Different Mulches, Varieties, and Planting Dates in North Dakota. Master’s Thesis,
Department of Agricultural and Biosystems Engineering, North Dakota State University, Fargo, ND, USA, 2016. 12. Jia, X.; Ransom, J.; Roy, D. Prediction of phenological development and maturity of two corn hybrids in eastern North Dakota. In
Proceedings of the 2014 ASABE Annual International Meeting, Montreal, QC, Canada, 12–16 July 2014. Paper No. 1913820. ament, W.J. Plastic mulches for the production of vegetable crops. In 2019, the
the water requir
4. Conclusions The soil
temperature under clear plastic mulch was always higher than other mulches, with the
highest increase of 4.65 ◦C at 15 cm in June 2019 compared to the no mulch treatment for
watermelon. Tomatoes under black plastic mulch and drip irrigation had the highest yield
compared to all other treatments, with 40.24 Mg ha−1. For watermelon, the highest yield
was from drip irrigation under clear plastic mulch, with 54.67 and 165.55 Mg ha−1 in 2019
and 2020, respectively. The treatment under the clear plastic mulch used the least amount
of irrigation (63 mm) but produced the highest yield for watermelon in 2020. Through
the two-year study, the automatic sensor-controlled irrigation provided a perfect example
on optimal irrigation, with no over irrigation in 2019, a wet year, and no under irrigation
in 2020, a dry year. Future research should focus on testing the irrigation controller for a
specific soil and crop and determining its threshold for higher yield and better quality. Water 2021, 13, 1991 21 of 22 21 of 22 Author Contributions: Conceptualization, U.B.P.V. and X.J.; methodology, U.B.P.V. and X.J.; investi-
gation, U.B.P.V. and X.J.; resources, X.J.; writing—original draft preparation, U.B.P.V.; writing—review
and editing, X.J., J.L., T.F.S. and C.W.L.; supervision, X.J.; funding acquisition, X.J. All authors have
read and agreed to the published version of the manuscript. Author Contributions: Conceptualization, U.B.P.V. and X.J.; methodology, U.B.P.V. and X.J.; investi-
gation, U.B.P.V. and X.J.; resources, X.J.; writing—original draft preparation, U.B.P.V.; writing—review
and editing, X.J., J.L., T.F.S. and C.W.L.; supervision, X.J.; funding acquisition, X.J. All authors have
read and agreed to the published version of the manuscript. Funding: This research was funded by North Dakota Department of Agriculture and ND Water
Resources Research Institute. Institutional Review Board Statement: Not applicable. Informed Consent Statement: Not applicable. Informed Consent Statement: Not applicable. Data Availability Statement: Not applicable. Acknowledgments: This project was supported by the North Dakota Department of Agriculture
(NDDA). Also, we are thankful to North Dakota Water Resources Research Institute (NDWRRI) for
providing the fellowship, which allowed us to complete the experiments. We would like to thank
department staff and students who helped to complete the field experiment successfully. We also
want to thank the reviewers and the associate editor for their valuable comments. The authors would
like to thank Dongqing Lin and Shad Mack for their technical support and help with field work. References HortTechnology 1993, 3, 35–39. [CrossRef] 14. Ibarra-Jiménez, L.; Zermeño-González, A.; Munguía-López, J.; Quezada-Martín, M.A.R.; De La Rosa-Ibarra, M. Photosynthesis,
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field-grown tomato under plastic mulch. Agric. Water Manag. 2007, 87, 285–291. [CrossRef] 24. Abdelkhalik, A.; Pascual-Seva, N.; Nájera, I.; Giner, A.; Baixauli, C.; Pascual, B. Yield response of seedless watermelon to different
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irrigation with black plastic mulch. Agric. Water Manag. 2005, 73, 247–254. [CrossRef] 26. Yang, P.; Hu, H.; Tian, F.; Zhang, Z.; Dai, C. References Crop coefficient for cotton under plastic mulch and drip irrigation based on eddy
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Sci. 2003, 168, 751–758. [CrossRef] 40. Li, S.X.; Wang, Z.H.; Li, S.Q.; Gao, Y.J.; Tian, X.H. Effect of plastic sheet mulch, wheat straw mulch, and maize growth on water
loss by evaporation in dryland areas of China. Agric. Water Manag. 2013, 116, 39–49. [CrossRef] 41. Li, H.; Yang, X.; Chen, H.; Cui, Q.; Yuan, G.; Han, X.; Wei, C.; Zhang, Y.; Ma, J.; Zhang, X. Water requirement characteristics and
the optimal irrigation schedule for the growth, yield, and fruit quality of watermelon under plastic film mulching. Sci. Hortic. 2018, 241, 74–82. [CrossRef] 42. Ray, M.; Biswasi, S. Impact of mulching on crop production: A review. Trends Biosci. 2016, 9, 757–767. 43. Singh, R.; Kumar, S.; Nangare, D.D.; Meena, M.S. Drip irrigation and black polyethylene mulch influence on growth, yield and
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INCONSTITUCIONALIDADE SUPREMA? UMA ANÁLISE ACERCA DA (IN)CONSTITUCIONALIDADE DA DECISÃO MONOCRÁTICA PROFERIDA NA ADPF 701/MG E POSSÍVEIS REFLEXOS EM UM CENÁRIO DE PANDEMIA
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doi.org/10.51891/rease.v7i6.1384 doi.org/10.51891/rease.v7i6.1384 1 Advogado, Mestrando em Direitos Sociais e Processos Reivindicatórios, Especialista em Civil e
Processo Civil e Trabalho e Processo do Trabalho. E-mail: lcsj.adv@gmail.com. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2 http://portal.stf.jus.br/noticias/verNoticiaDetalhe.asp?idConteudo=457782&ori=1 INCONSTITUCIONALIDADE SUPREMA? UMA ANÁLISE ACERCA DA
(IN)CONSTITUCIONALIDADE DA DECISÃO MONOCRÁTICA
PROFERIDA NA ADPF 701/MG E POSSÍVEIS REFLEXOS EM UM CENÁRIO
DE PANDEMIA Luiz Carlos Santos Junior1 RESUMO: Conquanto o Supremo Tribunal Constitucional – STF tenha o papel de
guardião da Constituição, verifica-se cada vez mais a ocorrência de decisões
monocráticas que por vezes são reformadas pelo Plenário, e, que, possivelmente,
estariam eivadas de inconstitucionalidade, já que substituídas por outro
entendimento considerado mais adequado ao ordenamento jurídico constitucional. Outrossim, embora o Plenário do STF não esteja vinculado aos seus próprios
precedentes, eis que possuem o papel de revisá-los, a mesma conclusão não se pode
aferir quanto à imprescindibilidade de observância dos precedentes por parte dos
ministros da Corte ao proferirem decisões monocráticas. Nesse sentido, busca o
presente artigo analisar aspectos da decisão monocrática proferida na ADPF
701/MG, quanto a possível existência de inconstitucionalidade e consequente
corresponsabilização à luz da tese fixada em controle concentrado de
constitucionalidade da MP nº 966/2020 e a necessidade de observância dos
precedentes vinculantes do STF. 433 Palavras-Chave: Decisões monocráticas STF. Inconstitucionalidade. Precedentes
Vinculantes. Pandemia. Responsabilização. ABSTRACT: Although the Brazilian Supreme Court - STF has the role of guardian
of the Constitution, monocratic decisions are occurring more and more, which are
sometimes reformed by the Plenary, and which, possibly, would be riddled with
unconstitutionality, since replaced by another one, understanding considered more
appropriate to the constitutional legal system. Furthermore, although the STF
Plenary is not linked to its own precedents, as they have the role of reviewing them,
the same conclusion cannot be drawn regarding the indispensability of the Court's
ministers to observe the precedents when making monocratic decisions. In this
sense, this article seeks to analyze aspects of the monocratic decision handed down
in ADPF 701/MG, regarding the possible existence of unconstitutionality and Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. ISSN - 2675 – 3375 Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE consequent co-responsibility in light of the thesis established in concentrated control
of constitutionality of MP nº 966/2020 and the need for compliance with binding
precedents of the STF. consequent co-responsibility in light of the thesis established in concentrated control
of constitutionality of MP nº 966/2020 and the need for compliance with binding
precedents of the STF. Keywords: Monocratic decisions STF. Unconstitutionality. Binding precedents. Pandemic. Responsibility. Keywords: Monocratic decisions STF. Unconstitutionality. Binding precedents. Pandemic. Responsibility. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 3 As decisões judiciais devem ser subordinadas à legislação, pois a sociedade deve ser governada por
representantes eleitos e se o juiz criar direito após o caso concreto, a parte perdedora estará sendo
julgada por um fundamento que não existia quando o fato jurídico aconteceu. (DWORKIN, 2002, p.
132) 1 INTRODUÇÃO Em tempos de pandemia e com a crescente judicialização dos conflitos entre
os poderes constituídos e entes federados acerca das suas respectivas competências,
insofismável a digressão pela atuação do Poder Judiciário diante deste cenário,
porquanto como protetor dos direitos fundamentais, no caso deste artigo ver-se-á em
especial o direito à saúde, faz-se necessário perquirir se esse vem exercendo a
jurisdição constitucional à luz da própria Constituição Federal. Tal análise demonstra importância mormente pelo elevado número de
decisões monocráticas proferidas em 20202, as quais representam 82% das decisões
proferidas pelo STF, quantificadas em 81.161 do total de 99.000 decisões, restando
apenas 18.208 decisões colegiadas. 434 Enquanto decisões monocráticas, estas estão sujeitas ao crivo do Plenário do
STF, o qual, enquanto Corte Constitucional, é responsável por apreciar, em última
análise, a constitucionalidade da decisão, não sendo rara as vezes em que uma decisão
monocrática possa vir a ser invalidada, o que se afigura como um oxímoro, tendo em
vista a probabilidade do próprio guardião da Constituição, por meio de seus
membros, exararem decisões inconstitucionais. Do exposto, partir-se-á da premissa da plausibilidade de decisões
monocráticas inconstitucionais no âmbito do STF, analisando a decisão proferida no
seio da ADPF 701/MG, a fim de avaliar possíveis inconstitucionalidades e por
consequência, concebíveis corresponsabilização nos termos propostos pela tese fixada
no controle de constitucionalidade da MP 966/2020. Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE 32)
4 Para Norberto Bobbio (2000, p. 380): “ [...] na democracia moderna, o soberano não é o povo, mas são
todos os cidadãos.”. Dessa maneira, os cidadãos detêm o poder de construir uma sociedade, justa, livre
e igualitária.
5 Grifo do autor. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2.1.
Da Legitimidade do Supremo Tribunal Federal enquanto Guardião da
Constituição Indiscutível a legitimidade do Supremo Tribunal Federal enquanto Guardião
da Constituição, em que pese algumas vezes sejam questionadas se a sua forma de
composição e atuação se encontram em consonância com princípio da democracia,
por não ser os seus representantes escolhidos em processo democrático direto que
prestigie o voto popular, tal qual os membros dos poderes legislativos e executivo34. Tal questionamento tem sentido, e, essa argumentação já fora explorada pelo
jurista Carl Schmitt (2007, págs. 176-177), em sua obra o Guardião da Constituição:
Todavia, um Tribunal exclusivamente com juristas funcionários de
carreira representaria afronta ao princípio democrático, verificando-se a
criação de uma segunda câmara (legislativa), porquanto o exame judicial
constitucional estaria concentrado nas mãos de funcionários inamovíveis e
independentes, de uma configurando-se em uma instância de alta política
dotada de poderes legislativos constitucionais. (...) do ponto de vista
democrático, seria praticamente impossível transferir tais funções a uma
aristocracia da toga5. Tal questionamento tem sentido, e, essa argumentação já fora explorada pelo
jurista Carl Schmitt (2007, págs. 176-177), em sua obra o Guardião da Constituição: 435 Contudo, não é correto afirmar que a configuração com a qual o Supremo
Tribunal Federal é formado afronte de alguma forma o princípio democrático, pois a
Constituição prevê a participação dos poderes executivos e legislativos no processo
de escolha dos membros da Corte, privilegiando dessa forma, a participação popular
por meio dos seus representantes eleitos. Neste sentido, Bobbio (2009, pág. 30) refere-se à construção de uma
definição mínima de democracia como “um conjunto de regras (primárias ou Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE fundamentais) que estabelecem quem está autorizado a tomar as decisões coletivas e
com quais procedimentos”. Assim, para que uma decisão adotada por alguns indivíduos seja aceita como
decisão coletiva é imprescindível que ela seja tomada com base em regras e
procedimentos. Mas isso não basta, porque, além de garantir o direito de participação
a um elevado número de cidadãos e de defender a existência de regras e
procedimentos, ele acrescenta uma terceira condição, “é preciso que aqueles que são
chamados a decidir ou eleger os que deverão decidir sejam colocados diante de
alternativas reais e postos em condição de escolher entre uma e outra” (BOBBIO,
2009, pág. 32). Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2.1.
Da Legitimidade do Supremo Tribunal Federal enquanto Guardião da
Constituição Corrobora o entendimento acerca do caráter democrático do Supremo
Tribunal Federal, o fato de este ser uma instância colegiada, o que reforça a sua
independência e imparcialidade para atuar como Guardião da Constituição,
protegendo-o de pressões externas, e, de certo modo, estabelecendo um sistema de
“freios e contrapesos” em si próprio, eis que a tendência de pluralidade de visões
tende a neutralizar posições extremas e por demais subjetivas, culminando em
debates mais objetivos, bem fundamentados e sensatos. 436 Jurgen Habermas, citado por COHENDET (2006, pág. 713), demonstrou que,
de uma maneira geral, a deliberação traduz-se em um conceito processual de
democracia. Montesquieu, por sua vez, afirma que se a colegialidade é inerente à
nossa tradição jurídica é porque, fundamentalmente, consideramos muito melhor
uma decisão tomada por uma deliberação do que pela decisão de um único. Do exposto, convicto de que o STF se coaduna ao princípio democrático, não
havendo dúvidas quanto a sua legitimidade para a funcionar enquanto Guardião da
Constituição, é mister refletir se de igual modo, a atuação singular dos seus
membros, ao proferir decisões monocráticas sem embasamento anterior da Corte
apto a fundamentar que estes estão decidindo em nome do colegiado, também
possuem o condão de ser consideradas como pronunciamentos do Guardião da
Constituição? Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Ademais, a abusiva e reiterada utilização de decisões monocráticas pode vir a
configurar violação ao princípio da colegialidade e consequentemente ao princípio
democrático, podendo ocasionar a deslegitimação do STF enquanto Guardião da
Constituição? É nessa toada que COHENDET (2006, pág. 736) afirma que a Separação dos
Poderes não seria efetiva se a colegialidade não fosse respeitada. O princípio da
colegialidade seria o corolário necessário de independência e imparcialidade de
justiça, princípio, inequivocamente, de valor constitucional e que reafirma o caráter
democrático de um legítimo Guardião da Constituição. Estas são questões espinhosas que pretendemos abordar e apresentar a nossa
visão no decorrer do artigo. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2.2.
Da necessidade de estabelecimento de limites à atuação monocrática dos
membros do STF 2), considera ativista o juiz
que: 438 a) use o seu poder de forma a rever e contestar decisões dos demais poderes
do estado; b) promova, através de suas decisões, políticas públicas; c) não
considere os princípios da coerência do direito e da segurança jurídica
como limites à sua atividade. 2.2.
Da necessidade de estabelecimento de limites à atuação monocrática dos
membros do STF Com o advento da Constituição de 1988, após a redemocratização do País, os
poderes da República permeavam os desafios da readaptação ao regime democrático,
e o Supremo Tribunal Federal, ainda composto por membros oriundos do regime
militar, possuía atuação mais conversadora, tímida, guiando-se pelo positivismo
jurídico que permanecia vigente, o qual se pautava em separar o direito de outros
ramos do conhecimento, como filosofia, psicologia, política, economia, com o fito de
garantir objetividade às decisões judiciais, tendo o juiz como uma mero tradutor e
aplicador das leis. Contudo, esse contexto mudou. De acordo com Barroso (2015, pág. 437 no início dos anos 2000, essa disfunção foi sendo progressivamente
superada e o STF foi se tornando, verdadeiramente, um intérprete da
Constituição. A partir daí, houve demanda por maior sofisticação teórica
na interpretação constitucional, superadora da visão tradicional de que se
tratava apenas de mais um caso de interpretação jurídica, a ser feita com
base nos elementos gramatical, histórico, sistemático e teleológico. Foi o
início da superação do positivismo normativista e de sua crença de que a
decisão judicial é um ato de escolha política. A partir daí, surge o que chamamos de pós-positivismo, operando-se uma
inovação quanto à hierarquia das normas anteriormente concebida, as normas são Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. ISSN - 2675 – 3375 Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE divididas entre regras (positivadas) e princípios, conferindo-se a estes últimos força
normativa, dotados de superioridade quanto aos primeiros. A Constituição Federal
passa a ser a centralidade de todo o ordenamento jurídico, integrando todas as demais
normas aos seus princípios fundamentais, funcionando como parâmetro
interpretativo. Nesse sentido, amplia-se o papel do judiciário como um todo, sobretudo do
STF, uma vez que todas as normas estão sendo interpretadas à luz dos princípios
fundamentais constitucionais, o que permite maior discricionariedade interpretativa,
já que os princípios são dotados de maior abstração e possibilitam uma valoração
mais subjetiva a critério do julgador, dando ensejo ao que se chama ativismo judicial. Não há consenso na doutrina, havendo conceituações mais favoráveis, outras
menos, levando-se em consideração os efeitos positivos e negativos decorrentes do
ativismo judicial. No entanto, colaciona-se abaixo um entendimento considerado
moderado, de modo que Thamy Pogrebinschi (2000, p. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 Acrescenta ainda a autora: Acrescenta ainda a autora: Não queremos sustentar que os três critérios acima devam ser preenchidos
simultaneamente para que se identifique um caso de ativismo. Um juiz
pode ser considerado ativista pelo exercício em graus diferenciados de
quaisquer das atitudes acima descritas. Porém, a negação de qualquer destas atitudes implica, segundo nossa
definição, que ele não seja considerado um ativista. Isto é, um juiz que se
recuse a exercer seu poder de pelo menos uma das formas acima, não será
considerado, neste artigo, um ativista. Desse modo, perante a sociedade, não se nega a importância de certo grau de
ativismo judicial com vistas à garantia dos direitos fundamentais, porquanto
vivemos em uma situação generalizada de violações de direitos em virtude de
omissões e inconsequências de agentes públicos dos poderes Executivo e Legislativo, contudo, relevante também é buscar limites para a atuação do judiciário, porquanto a
regra é a irresponsabilidade estatal pelas decisões judiciais. Assim, admoesta Marinoni (2013, pág. 64): Assim, admoesta Marinoni (2013, pág. 64): [...] não há poder que não tenha responsabilidade pelas suas decisões. Porém, é pouco plausível que alguém possa justificar a sua
responsabilidade quando decide casos iguais de forma desigual” (...) “tratar
da mesma forma casos similares é algo fundamental para a estabilidade do
Poder. Dessa forma, constata-se a necessidade de criar mecanismos de limitação à
atuação do STF, particularmente no que tange ao exercício da jurisdição
constitucional de forma singular, por meio de decisões monocráticas, pois se
constitui em um elevado poder nas mãos de agentes públicos que dificilmente serão
responsabilizados por eventuais danos ocasionados à sociedade, embora atualmente
funcione muitas vezes como gestor público, como pretende-se demonstrar no
presente artigo, implementando diretamente, ou mesmo restringindo, a adoção das
medidas necessárias ao combate à pandemia. Inconteste o desconforto dos poderes Executivo e Legislativo e a ânsia em
equalizar as forças, arrefecendo-se o ativismo judicial de decisões monocráticas do
STF, primando pela independência e harmonia dos Poderes, tanto que tramita no
Senado Federal a Proposta de Emenda à Constituição – PEC nº 8/20216, que dispõe
sobre a altera da Constituição Federal para dispor sobre os pedidos de vista,
declaração de inconstitucionalidade e concessão de medidas cautelares nos tribunais. 6 Trecho da justificativa da PEC: “É chegada a hora de rediscutir tema de grande importância para o
equilíbrio das instituições no Brasil. Não se pode mais conviver com um modelo em que decisões
judiciais individuais e precárias determinem o futuro de questões de grande relevância nacional.” Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. 7 ADI 6421, 6422, 6424, 6425, 6427, 6428 E 6431 MC/DF. Rel. Min. Roberto Barroso. Julgado em 20 e
21/05/2020. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 Acrescenta ainda a autora: 439 Nesse contexto, revela-se importante a adoção de precedentes judiciais,
sobretudo os vinculantes, devendo estes serem enxergados não como uma limitação
negativa ao poder decisório ou violação à independência de convencimento do juiz,
mas como uma limitação positiva no sentido de garantir ao jurisdicionado uma
jurisprudência íntegra e coerente, além de reforçar o equilíbrio entre os Poderes,
privilegiando os princípios da democracia e separação dos poderes, e, no caso do STF, ,
ISSN - 2675 – 3375 Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE o princípio da colegialidade, respeitado quando os seus próprios membros observam
os precedentes fixados pelo Plenário. 3 DO CARÁTER VINCULANTE DA DECISÃO PROFERIDA NA ADI 6421
MC/DF Preliminarmente, antes de proceder a análise da decisão monocrática
proferida no ADPF 701/MG, mister responder um possível questionamento que
poderá surgir: As teses fixadas pelo STF no controle concentrado de
constitucionalidade de medidas provisórias que perdem a validade continuam sendo
precedentes vinculantes? Essa indagação é justificada pelo fato da Medida Provisória nº 966/2020, que
dispôs sobre a responsabilização de agentes públicos por ação e omissão em atos
relacionados com a pandemia da covid-19, haver perdido a sua validade em
10/09/2020, por não ter sido votada no Congresso Nacional. Todavia, anteriormente a perda de sua validade, a referida MP fora objeto de
controle de constitucionalidade pelo Supremo Tribunal Federal, por meio da ADI
6421 MC/DF7, o qual decidiu pela sua constitucionalidade, realizando uma
intepretação conforme a Constituição, fixando as seguintes teses: 440 1. Configura erro grosseiro o ato administrativo que ensejar violação
ao direito à vida, à saúde, ao meio ambiente equilibrado ou impactos
adversos
à
economia,
por
inobservância:
i)
de
normas
e
critérios
científicos
e
técnicos;
ou
ii) dos princípios constitucionais da precaução e da prevenção. 2. A autoridade a quem compete decidir deve exigir que as opiniões
técnicas
em
que
baseará
sua
decisão
tratem
expressamente:
i) das normas e critérios científicos e técnicos aplicáveis à matéria, tal
como estabelecidos por organizações e entidades internacional e
nacionalmente
reconhecidas;
e ii) da observância dos princípios constitucionais da precaução e da
prevenção, sob pena de se tornarem corresponsáveis por eventuais
violações a direitos. Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Nessa esteira, sobreveio o seguinte questionamento: a tese fixada pelo STF
continua tendo aplicabilidade, em que pese a medida provisória que a originou tenha
perdido a validade, sendo aplicável para fins de aferição de responsabilidade dos
agentes públicos durante à pandemia? Destarte, embora a medida provisória tenha sido extirpada do ordenamento
jurídico pátrio, não se vislumbra nada que possa concluir que o mesmo tenha
ocorrido com a tese fixada pelo STF, porquanto o controle normativo de
constitucionalidade qualifica-se como típico processo de caráter objetivo,
vocacionado, exclusivamente, à defesa, em tese, da harmonia do sistema
constitucional,
tendo
por
função
instrumental
viabilizar
o
julgamento
da validade abstrata do ato estatal em face da Constituição da República8. 8 STF - AG.REG. na ADI 2422 DF, Rel. Min. Celso de Mello, DJE 29/10/2014.
9 ADI nº 3.146/DF, Rel. Min. Joaquim Barbosa, Tribunal Pleno, julgado em 11/05/2006.
10 ADI nº 4.356/CE, Rel. Min. Dias Toffoli, Tribunal Pleno, julgado em 09/02/2011. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 q
j
g
10 ADI nº 4.356/CE, Rel. Min. Dias Toffoli, Tribunal Pleno, julgado em 09/02/2011. 3 DO CARÁTER VINCULANTE DA DECISÃO PROFERIDA NA ADI 6421
MC/DF Ademais, embora o precedente que será trazido à baila não seja próprio da
situação concreta apresentada, acredita-se que esse milita a favor da presunção da
aplicabilidade da tese fixada em sede de controle concentrado de constitucionalidade
conquanto a medida provisória apreciada tenha perdido a sua validade, tendo em
vista que o STF já decidiu anteriormente que não há perda de objeto quando, mesmo
sendo a lei impugnada de vigência temporária e já exaurida, a ADI for proposta antes
de esgotada a sua vigência. 441 A esse respeito, colacionamos trechos das ementas confeccionadas do
julgamento da ADI nº 3.146/MG9 e nº 4.356/CE10: (...) Preliminar de prejudicialidade: dispositivo de norma cuja eficácia foi
limitada até 31.12.2005. Inclusão em pauta do processo antes do
exaurimento da eficácia da norma temporária impugnada. Julgamento
posterior ao exaurimento. Circunstâncias do caso afastam a aplicação da
jurisprudência do Supremo Tribunal Federal sobre a prejudicialidade da
ação, visto que o requerente impugnou a norma em tempo adequado. Conhecimento da ação (...). (ADI 3.146/DF, Rel. Min. Joaquim Barbosa,
DJ 19/12/2006) (...) 1. Singularidades do caso afastam, excepcionalmente, a aplicação da
jurisprudência do Supremo Tribunal Federal sobre a prejudicialidade da
ação, visto que houve impugnação em tempo adequado e a sua inclusão em Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE
pauta antes do exaurimento da eficácia da lei temporária impugnada,
existindo a possibilidade de haver efeitos em curso (art. 7º da Lei
14.506/2009) (...). (ADI 4.356/CE, Rel. Min. Dias Toffoli, DJ 11/05/2011) pauta antes do exaurimento da eficácia da lei temporária impugnada,
existindo a possibilidade de haver efeitos em curso (art. 7º da Lei
14.506/2009) (...). (ADI 4.356/CE, Rel. Min. Dias Toffoli, DJ 11/05/2011) Logo, conclui-se que, se é possível analisar a constitucionalidade de leis
impugnadas ainda que sua vigência já tenha se exaurido, com mais razão deve
prevalecer vigente e aplicável o entendimento fixado pelo STF em sede de controle
concentrado de constitucionalidade, quando da análise de medida provisória que
perdeu a sua validade posteriormente ao julgamento. Quanto ao caráter vinculante da tese fixada, tem-se que a formação de
precedentes busca estabilizar a jurisprudência, privilegiando princípios fundamentais
como o da segurança jurídica e razoável duração do processo, com vistas a ofertar
uma racionalidade normativa para a construção de um modelo de precedentes
embasada na pluralidade de litigiosidade e em conformidade com o processo
democrático, dotando-a de certa previsibilidade. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 3 DO CARÁTER VINCULANTE DA DECISÃO PROFERIDA NA ADI 6421
MC/DF Justifica-se ainda como primordial a formação e respeito aos precedentes
judiciais, pois de acordo com Alexandre Bahia e Dierle Nunes (2014, pág. 433-471), no
cenário atual: 442 parece haver um fomento do desrespeito da opinião de uma corte, mesmo
após a prolação de uma decisão pelo pleno ou corte especial, em prol de um
juízo personalista que despreza o passado institucional (integridade) e que
acaba alimentando a litigiosidade pela evidente possibilidade de êxito
embasada em julgado extraído de nossa jurisprudência lotérica. parece haver um fomento do desrespeito da opinião de uma corte, mesmo
após a prolação de uma decisão pelo pleno ou corte especial, em prol de um
juízo personalista que despreza o passado institucional (integridade) e que
acaba alimentando a litigiosidade pela evidente possibilidade de êxito
embasada em julgado extraído de nossa jurisprudência lotérica. Desse modo, ululante a importância do respeito aos precedentes vinculantes,
pois o que evidentemente se quer combater é a anarquia interpretativa típica de nosso
sistema e que induziu a adoção do presente modelo normativo de precedentes, ora em
comento, para que possamos nos aproximar das virtudes, sem copiar os equívocos, de
um legítimo direito jurisprudencial. (HUMBERTO, NUNES, BAHIA, PEDRON,
pág. 375) Nesse sentido, o novo CPC de 2015 legitimou os chamados precedentes
vinculantes, de aplicação obrigatória pelos Tribunais, os quais abarcam as decisões Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE proferidas em controle concentrado de constitucionalidade, sendo, inclusive, cabível
Reclamação em caso de sua inobservância. Nessa toada, preconiza o CPC: Art. 927. Os juízes e os tribunais observarão: Art. 927. Os juízes e os tribunais observarão:
I – as decisões do Supremo Tribunal Federal em controle concentrado de
constitucionalidade;
Art. 988. Caberá reclamação da parte interessada ou do Ministério Público para:
III – garantir a observância de enunciado de súmula vinculante e de decisão do
Supremo Tribunal Federal em controle concentrado de constitucionalidade; I – as decisões do Supremo Tribunal Federal em controle concentrado de
constitucionalidade; Art. 988. 3 DO CARÁTER VINCULANTE DA DECISÃO PROFERIDA NA ADI 6421
MC/DF Caberá reclamação da parte interessada ou do Ministério Público para: III – garantir a observância de enunciado de súmula vinculante e de decisão do
Supremo Tribunal Federal em controle concentrado de constitucionalidade; Assim, resta claro a força das decisões do Supremo Tribunal Federal em
controle de constitucionalidade, caracterizando-se como precedentes vinculantes de
aplicação obrigatória pelo judiciário, incluindo os próprios Ministros do STF ao
decidirem monocraticamente, pois assim preconiza-se o princípio da colegialidade,
sendo, não uma decisão do Ministro, mas sim, da Corte Constitucional. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 11 ADPF nº 696/DF, Rel. Min. Marco Aurélio, julgado em 19/06/2020. 3 DA ANÁLISE DA DECISÃO MONOCRÁTICA PROFERIDA NA ADPF
701/MG 443 No bojo da APDF nº 701/MG, o Excelentíssimo Ministro Nunes Marques,
do STF, decidiu monocraticamente pela concessão de medida liminar determinando
que os Estados, Distrito Federal e Municípios se abstenham de editar ou de exigir o
cumprimento de decretos ou atos administrativos locais que proíbam completamente
a realização de celebrações religiosas presenciais, por motivos ligados à prevenção da
Covid-19, com a manutenção de protocolos sanitários de prevenção. Nessa vereda, pincelamos aqueles que consideramos os principais argumentos
que consubstanciaram a decisão supramencionada e motivaram o convencimento do
juízo, de forma a viabilizar a análise que será realizada: a) Em preliminar: a) Em preliminar: a) Em preliminar: (...) Não se ignora que, no julgamento do Agravo Regimental na ADPF
703/BA, Rel. Min. Alexandre de Moraes, julgado em 17/02/2021, este
Tribunal considerou que a ora autora não seria parte legítima. Contudo, este caso possui premissas fáticas distintas, a atrair assim o
distinguishing. Com efeito, aqui o provimento buscado pela Associação
guarda relação fundamental com seus objetivos essenciais, consistentes na Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE proteção da liberdade religiosa. Por prudência, ao menos neste momento
processual, esta Suprema Corte deve prestigiar a instrumentalidade do
processo, na medida em que o objeto desta ação diz com a proteção da
liberdade de culto e religião, garantia constitucional. Além disso, é certo que, no Agravo Regimental no ADPF 696, Rel. Min. Marco Aurélio, julgado em 30/11/2020, o Tribunal, ainda que
implicitamente, aceitou a legitimidade da Associação Brasileira de Juristas
Pela Democracia – ABJD. Assim, na existência de aparente divergência
jurisprudencial, deve-se prestigiar a concreção do Acesso à Justiça,
conforme art. 5º, XXXV, Constituição Federal: “(...) a lei não excluirá da
apreciação do Poder Judiciário lesão ou ameaça a direito”; (...) (grifo
nosso) b)
No mérito: [...] Ao tratar o serviço religioso como não-essencial, Estados e municípios
podem, por via indireta, eliminar os cultos religiosos, suprimindo aspecto
absolutamente essencial da religião, que é a realização de reuniões entre os
fiéis para a celebração de seus ritos e crenças. A proibição categórica de cultos não ocorre sequer em estados de defesa
(CF, art. 136, § 1º, I) ou estado de sítio (CF, art. 139). Como poderia ocorrer
por atos administrativos locais? Certo, as questões sanitárias são
importantes e devem ser observadas, mas, para tanto, não se pode fazer
tábula rasa da Constituição. Observa-se, nesse sentido, que diversas atividades também essenciais, tais
como o serviço de transporte coletivo, vêm sendo desenvolvidas ainda que
em contexto pandêmico, demandando para tanto um protocolo sanitário
mínimo que, com as devidas considerações, poderia ser também adotado
no presente caso. 444 p
Entendo por demais gravosa a vedação genérica à atividade religiosa, da
forma como prevista em parte dos diplomas objeto da presente ação,
traduzindo-se
em
medida
atentatória
a
preceito
fundamental
consubstanciado em liberdade religiosa. Reconheço que o momento é de cautela, ante o contexto pandêmico que
vivenciamos. Ainda assim, e justamente por vivermos em momentos tão
difíceis, mais se faz necessário reconhecer a essencialidade da atividade
religiosa, responsável, entre outras funções, por conferir acolhimento e
conforto espiritual. (...) Com todo o respeito ao Excelentíssimo Ministro, mas nenhum dos
argumentos contidos em sua fundamentação é capaz de sustentar minimamente a
constitucionalidade da decisão proferida, conforme ver-se-á mais a frente. Com todo o respeito ao Excelentíssimo Ministro, mas nenhum do constitucionalidade da decisão proferida, conforme ver-se-á mais a frente. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2.3.
Da inconstitucionalidade formal Inicialmente, ressalta-se combalido o distinguinshing realizado pelo Ministro,
pois este se pautou em um “precedente implícito” (ADPF 696/DF)11 anterior ao Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE precedente explícito (ADPF 703/BA)12, o qual versava justamente acerca da
ilegitimidade da Autora da presente ADPF em análise, Associação Nacional de
Juristas Evangélicos (ANAJURE). Dessa forma, a decisão encontra-se totalmente desprovida de fundamentação,
considerando que sequer trouxe as razões da distinção ou superação do precedente
atual, violando o dever constitucional de fundamentação13, padecendo do vício de
nulidade, além de afrontar princípios como o da isonomia e o da segurança jurídica. Outrossim, vale ressaltar que a decisão monocrática que concedeu a medida
liminar vem à luz após o próprio Supremo Tribunal Federal, no julgamento da ADI
6341/DF14, haver decidido pela competência concorrente da União, Estado, Distrito
Federal e Município para adoção de medidas de enfrentamento da emergência da
saúde pública decorrente do coronavírus, autorizando aos demais Entes, mesmo sem
autorização da União, adotar medidas como isolamento, quarentena, exumação,
necropsia, cremação e manejo de cadáver e restrição à locomoção interestadual e
intermunicipal em rodovias, portos ou aeroportos. A decisão conferiu ainda interpretação conforme com a Constituição ao artigo
3º da Lei 13.979/2020, a fim de deixar claro que a União pode legislar sobre o tema,
mas que o exercício desta competência deve sempre resguardar a autonomia dos
demais entes, uma vez que a possibilidade do chefe do Executivo Federal definir por
decreto a essencialidade dos serviços públicos, sem observância da autonomia dos
entes locais, afrontaria o princípio da separação dos poderes. Ora, se o chefe do Poder Executivo não poderia definir por decreto a
essencialidade dos serviços públicos sem observar a autonomia dos entes locais, por
que razão um Ministro do STF o poderia fazer em decisão monocrática, dispondo
sobre a essencialidade da religião e permitindo de cultos e missas presencialmente? É
muito poder para pouca (ou nenhuma) responsabilidade. 445 Esse comportamento inflama ainda mais os ânimos, não se olvidando que o
próprio Plenário do STF havia considerado que uma possível usurpação de
competência dos Estados e Município pela União configuraria afronta ao princípio
da separação dos poderes, inconstitucionalidade material que será tratada mais
adiante. 12 ADPF nº 703/BA, Rel. Min. Alexandre de Moraes, julgado em 11/11/2020.
13 CF, Art. 93, inciso IX - todos os julgamentos dos órgãos do Poder Judiciário serão públicos, e
fundamentadas todas as decisões, sob pena de nulidade, podendo a lei limitar a presença, em
determinados atos, às próprias partes e a seus advogados, ou somente a estes, em casos nos quais a
preservação do direito à intimidade do interessado no sigilo não prejudique o interesse público à
informação.
14 ADI nº 6341/DF, Rel. Min. Marco Aurélio, julgado em 03/07/2020. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 12 ADPF nº 703/BA, Rel. Min. Alexandre de Moraes, julgado em 11/11/2020. 14 ADI nº 6341/DF, Rel. Min. Marco Aurélio, julgado em 03/07/2020. 2.3.
Da inconstitucionalidade formal Destarte, o que ocorreu na realidade foi uma anarquia interpretativa por parte
de um Ministro do STF, tendo em vista que a sua decisão monocrática violou
sistematicamente diversos precedentes da Corte Constitucional conforme exposto
acima. Outrossim, pede-se licença para aprofundar um pouco mais com relação à Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE violação ao precedente firmado na ADI 6.421 MC/DF, retomando o que já fora
abordado no item 3 deste artigo. Em apertada síntese, a tese fixada em sede da ADI 6.421 MC/DF fixou
parâmetros para a responsabilização dos gestores públicos no âmbito de sua atuação
no período da pandemia, preconizando que as medidas de combate ao Covid-19
devem ser precedidas de recomendação técnica e fundamentada. Nessa esteira, no que tange à proteção da saúde e da vida, a jurisprudência do
STF se baseia em dois parâmetros: a) as decisões governamentais devem respeitar
padrões técnicos e evidências científicas sobre a matéria; b) essas questões se
sujeitam ao princípio da prevenção e ao princípio da precaução, ou seja, se existir
alguma dúvida quanto aos efeitos de alguma medida, ela não deve ser aplicada. Isso
significa que a Administração deve se pautar pela autocontenção.15 Nesse contexto, no qual o Brasil já conta com mais de 370.000,00 (trezentos e
setenta mil) mortes decorrentes do covid-1916, imprescindível se faz a reunião de
esforços de todos os Entes e Poderes, cada qual na sua esfera de atuação e com base
em estudos e evidências referendados por organizações e entidades médicas e
sanitárias, reconhecidas nacional e internacionalmente, assim como a observância
dos princípios constitucionais da precaução e da prevenção, conforme citados
alhures. Desta sorte, sem adentrar ao mérito da essencialidade da atividade religiosa
utilizada como fundamento da decisão monocrática do Excelentíssimo Ministro
Nunes Marques, advoga-se pela existência de violação ao precedente vinculante
constituído na ADI 6.421 MC/DF, porquanto ao substituir as ações adotadas pelos
gestores públicos, a decisão do Ministro deveria estar amparada em fortes evidências
técnicas e científicas, o que a priori, não restou configurado no caso concreto, visto
que a decisão não se baseou em nenhuma fonte apta a repelir as justificativas das
ações adotadas pelos gestores públicos. 15https://www.buscadordizerodireito.com.br/jurisprudencia/detalhes/815104ed949f9deaf45165c4b83370
13?palavra-chave=covid+responsabiliza%C3%A7%C3%A3o&criterio-pesquisa=e
16
https://g1.globo.com/bemestar/coronavirus/noticia/2021/04/17/brasil-ultrapassa-371-mil-mortos-
por-covid-pais-registrou-2865-mortes-em-24-horas.ghtml Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 3 p
p
3
7
3
3
p
q
16
https://g1.globo.com/bemestar/coronavirus/noticia/2021/04/17/brasil-ultrapassa-371-mil-morto
por-covid-pais-registrou-2865-mortes-em-24-horas.ghtml 2.3.
Da inconstitucionalidade formal 446 Nesse sentido, não possui lógica que os administradores públicos devam se
submeter aos parâmetros decisórios fixados pelo Plenário do STF para realizar
medidas de combate ao covid-19 sob pena de corresponsabilização, e que os membros
do Judiciário, quando da supressão de tais medidas, não estejam sujeitos aos mesmos
paradigmas, inclusive sob pena de serem responsabilizados. Como diz o dito popular:
“pau que dá em Chico dá em Francisco”! É inconcebível, do ponto de vista do princípio democrático, da separação dos
poderes, da colegialidade, que apenas um Ministro do Supremo tenha o condão de
permitir a realização de cultos e missas presenciais em todo País, quando nem o
Presidente da República o poderia fazer, pois deve respeito ao pacto federativo e a
autonomia dos demais entes, como bem asseverou o Plenário do Supremo, e Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE tampouco o Poder Legislativo tem a prerrogativa de, por meio de apenas um membro
do Congresso Nacional, criar leis que vigorem em todo o território nacional. Do exposto, restam fortes indícios acerca da inconstitucionalidade formal da tampouco o Poder Legislativo tem a prerrogativa de, por meio de apenas um membro
do Congresso Nacional, criar leis que vigorem em todo o território nacional. Do exposto, restam fortes indícios acerca da inconstitucionalidade formal da
decisão monocrática estudada, porquanto viola o dever de observância de diversos
precedentes de caráter vinculante formados pelo Guardião da Constituição. 2.4. Da inconstitucionalidade material decisão monocrática estudada, porquanto viola o dever de observância de diversos
precedentes de caráter vinculante formados pelo Guardião da Constituição. 2.4. Da inconstitucionalidade material p
2.4. Da inconstitucionalidade material A saúde é um direito fundamental, de caráter programático e eficácia
limitada, eis que além de preconizar uma tarefa, uma finalidade a ser atingida,
diferentemente dos direitos fundamentais de defesa que preveem uma abstenção do
Estado, esse requer a intervenção do poder legislativo infraconstitucional para que o
direito alcance a sua eficácia plena, por meio de prestação positiva por parte do
Estado. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 17 Nesse sentido, afirma Ingo Wolfgang Sarlet, em sua obra A Eficácia dos Direitos Fundamentais 10 17 Nesse sentido, afirma Ingo Wolfgang Sarlet, em sua obra A Eficácia dos Direitos Fundamentais 10ª.
ed., rev., atual. e ampl. Porto Alegre: Livraria do Advogado, 2009, p. 297. ,
g
g
g
,
ed., rev., atual. e ampl. Porto Alegre: Livraria do Advogado, 2009, p. 297. 2.3.
Da inconstitucionalidade formal De tal sorte, por ser um direito de prestação positiva, demanda o emprego de
recursos públicos, influenciando tanto na esfera socioeconômica do Ente Público,
quanto na esfera de sua competência, porquanto este é que detém a legitimidade para
a implementação das políticas públicas, dentro de sua possibilidade técnica,
orçamentária e financeira, caracterizando-se esses fatores como atributos do
princípio da reserva do possível, o qual pode vir a representar óbice à realização da
plena efetividade da norma constitucional. 447 Contudo, em que pese o direito a saúde seja considerado norma programática,
de eficácia limitada, convém destacar que a doutrina majoritária, quiçá unanime,
preleciona que todas as normas constitucionais, são dotadas de um mínimo de
eficácia e aplicabilidade, conforme o seu grau de densidade normativa, sendo possível
a sua aplicação direta nos limites da sua normatividade, não constituindo as normas
de eficácia limitada meros conselhos, advertências e/ou proclamações de cunha
ideológico. A esse respeito, trazemos a lição do Professor Ingo Wolfgang Sarlet (2009,
pág. 294), o qual em seu livro a eficácia dos direitos fundamentais, assim apregoa: Todas as normas de direito fundamentais são dotadas de eficácia e, em
certa medida, diretamente aplicáveis já ao nível da Constituição e
independentemente de intermediação legislativa. [...] aqui repisamos para
espancar toda e qualquer incompreensão para com a nossa posição, todas as
normas de direitos fundamentais são diretas (imediatamente) aplicáveis na Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. ISSN - 2675 – 3375 Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE medida de sua eficácia, o que não impede que se possa falar de uma
dimensão “programática” dos direitos fundamentais. medida de sua eficácia, o que não impede que se possa falar de uma
dimensão “programática” dos direitos fundamentais. Nessa esteira, certo da eficácia mínima dos direitos fundamentais, verifica-se
que essa contempla como consequências, dentre outras, a revogação/não recepção de
atos normativos anteriores contrários ao direito fundamental positivado e a
possibilidade de serem utilizados como parâmetros não só para o controle de
constitucionalidade, mas para interpretação, integração e aplicação das normas
jurídicas, vinculação ao legislador quanto à concretização dos programas e
finalidades estabelecidos, além de possuir a chamada dimensão negativa. Para Jorge Miranda (1988, p. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 2.3.
Da inconstitucionalidade formal 219-20), a denominada dimensão negativa dos
direitos a prestações, dispõe que: [...] as normas programáticas, ainda que essencialmente se caracterizem
como preceptivas (e não proibitivas), também possuem um sentido
complementar negativo (ou proibitivo), visto que, além de vedarem a
emissão de atos normativos contrários, proíbem a prática de
comportamentos que tenham por objetivo impedir a produção dos atos
destinados à execução das tarefas, fins ou imposições contidas na norma
programática. [...] as normas programáticas, ainda que essencialmente se caracterizem
como preceptivas (e não proibitivas), também possuem um sentido
complementar negativo (ou proibitivo), visto que, além de vedarem a
emissão de atos normativos contrários, proíbem a prática de
comportamentos que tenham por objetivo impedir a produção dos atos
destinados à execução das tarefas, fins ou imposições contidas na norma
programática. 448 Desta sorte, essa dimensão negativa acaba por atribuir às normas
programáticas, características do direito fundamental de defesa, no que tange a exigir
do Estado uma abstenção de atuar contrariamente ao conteúdo da norma
constitucional que prevê direitos fundamentais. Nessa vereda, a doutrina nacional
consagra, inclusive, que tal exigência de abstenção representa um direito subjetivo do
cidadão. Assim, entendem Meirelles Teixeira, Luís Roberto Barroso e Celso Antônio
Bandeira de Mello, os quais reconhecem ao indivíduo o direito de se opor
judicialmente ao cumprimento de regras ou à sujeição de atos contrários ao sentido
do preceito constitucional que o atingem pessoalmente17. Vale a pena ressaltar que Jorge Miranda (1993, p. 283-4), para além da
dimensão negativa, aponta a existência de uma positiva, quanto à vinculação do
Poder Judiciário aos direitos fundamentais, o qual tem o dever de atribuir-lhes à Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE maior eficácia possível perante o sistema jurídico, através da aplicação, interpretação
e integração. Nesse diapasão, a decisão analisada mostra-se materialmente inconstitucional
e extremamente temerária, porquanto afronta o direito fundamental à saúde em
todas as suas dimensões, na medida em repeliu a atuação de gestores públicos no
combate à pandemia quando vedou a restrição de práticas religiosas presenciais em
templos e igrejas, de forma que impediu a produção de atos, devidamente
fundamentados em evidências técnicas e científicas, destinados a garantir o direito
fundamental à saúde e à vida. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 18http://www.stf.jus.br/portal/cms/verNoticiaDetalhe.asp?idConteudo=441447#:~:text=No%20seu%2
0entendimento%2C%20a%20possibilidade,princ%C3%ADpio%20da%20separa%C3%A7%C3%A3o%20
dos%20poderes. 19 Rcl. nº 41.935/MT. 2.3.
Da inconstitucionalidade formal Outra inconstitucionalidade material detectada e que vale a pena ressaltar,
consiste na afronta a separação dos poderes como assentou o Plenário do STF,
quando a maioria dos Ministros aderiram a interpretação conforme ao art. 3º da Lei
nº 13.979/2020, proposta pelo Ministro Edson Fachin, que no seu entendimento, a
possibilidade do chefe do Executivo Federal definir por decreto a essencialidade dos
serviços públicos, sem observância da autonomia dos entes locais, afrontaria o
princípio da separação dos poderes18. 449 Desta feita, ao substituir os membros do Poder Executivo no combate à
pandemia, em puro ativismo judicial, o Excelentíssimo Ministro Nunes Marques
indubitavelmente fragiliza o processo democrático e viola a separação dos poderes. A
respeito da forma como o ativismo judicial vem ocorrendo no Brasil, alerta Elival
Ramos (2010, p. 120) Com efeito, nos Estados democráticos a subversão dos limites impostos à
criatividade da jurisprudência, com o esmaecimento de sua função
executória, implica deterioração do exercício da função jurisdicional, cuja
autonomia é inafastável sob a vigência de um Estado de Direito, afetando
se, inexoravelmente, as demais funções estatais, máxime a legiferante, o
que, por seu turno, configura gravíssima agressão ao Princípio da
Separação dos Poderes Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE De tal modo, defende o autor que o ativismo acarreta instabilidade à
harmonia necessária entre o Executivo, Legislativo e o Judiciário, desabona o
princípio da separação de poderes e atribui ao juiz um poder criador da norma que é
estranho ao modelo de organização do Estado erigido sob a doutrina da separação de
poderes. No caso dos Ministros do STF, os dota de poderes quase que ilimitados. Corrobora a inconstitucionalidade afirmada, a medida cautelar concedida pelo
Ministro Presidente Dias Toffoli nos autos da Reclamação nº 41.935/MT19, a qual
objetivava suspender os efeitos da decisão judicial que havia relaxado as medidas de
prevenção e contenção do novo coronavírus em Cuiabá (MT), determinando o
aumento da circulação de ônibus e proibindo o governo local de restringir os horários
de funcionamento das atividades consideradas essenciais. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 Colacionamos trecho da decisão do Ministro: Colacionamos trecho da decisão do Ministro: Note-se que embora a decisão de origem consigne que “os decretos
editados nem de longe se enquadram nas Normas Científicas e do Decreto
Estadual”, o que deixaria “claro que as atividades essenciais foram
classificadas de modo aleatório, ao talante do administrador”, não se
observa a devida fundamentação quanto ao ponto, ou seja, não parece ter
havido a efetiva demonstração do porquê os critérios técnicos adotados
pelo estado estariam em posição de maior evidência científica do que os
utilizados pelo Município em seu Decreto. Ausente, assim, fundamentação
apta a justificar a prevalência de uma norma sobre outra, e ausente ainda
indicação de eventual normatização do Município em matéria de
competência estadual, considero ser o caso de concessão da tutela
pretendida. (grifo nosso) 450 De toda a sorte, entendeu o Ministro Presidente que a decisão do juízo de 1º
grau (muito semelhante à do Ministro Nunes Marques) divergia do que foi decido
pelo STF, ressaltando ainda que a decisão não indicou porque os critérios técnicos
adotados pelo estado estariam em posição de maior evidência científica do que os
utilizados pelo município para regular o funcionamento das atividades essenciais,
considerando-a privada de fundamentação. Com base no exposto, evidente também a interferência indevida da decisão
monocrática apreciada no âmbito do Poder Executivo, eis que não se limitou à
análise da legalidade dos atos administrativos que visavam a restrição do Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE funcionamento dos cultos e missas de modo presencial, mas procedeu a um real
exame de mérito, contudo sem considerar o princípio da precaução, o qual restaria
materializado pela indicação dos critérios científicos aptos a fundamentar a decisão,
em patente ofensa ao princípio da separação dos poderes. 21 No caso de decisões judiciais na seara criminal, é possível a responsabilização do Estado por
condenação criminal errônea, contudo o agente público (juiz) só poderá ser regressivamente
responsabilizado em casos de dolo ou fraude. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 20 CF. Art. 37 § 6 º: As pessoas jurídicas de direito público e as de direito privado prestadoras de
serviços públicos responderão pelos danos que seus agentes, nessa qualidade causarem a terceiros,
assegurado o direito de regresso contra o responsável nos casos de dolo ou culpa”. 20 CF. Art. 37 § 6 º: As pessoas jurídicas de direito público e as de direito privado prestadoras de
serviços públicos responderão pelos danos que seus agentes, nessa qualidade causarem a terceiros,
assegurado o direito de regresso contra o responsável nos casos de dolo ou culpa”.
21 No caso de decisões judiciais na seara criminal, é possível a responsabilização do Estado por
condenação criminal errônea, contudo o agente público (juiz) só poderá ser regressivamente
responsabilizado em casos de dolo ou fraude. 3. DA POSSIBILIDADE DE RESPONSABILIZAÇÃO DO ESTADO POR
ATOS DO JUDICIÁRIO EM TEMPOS DE PANDÊMIA A responsabilidade do Estado é prevista na Constituição (art. 37, §6º)20, na
modalidade objetiva, ou seja, independentemente de dolo ou culpa, sendo possível, no
entanto, excludentes de responsabilidade como culpa exclusiva da vítima e caso
fortuito e força maior, sendo adotada a chamada Teoria do Risco Administrativo. Ocorre que, no que tange a responsabilidade do Estado em virtude decisões judiciais
decorrentes da atuação do Poder Judiciário, existem muitas controvérsias, sendo a
regra a irresponsabilidade do Estado por decisões judiciais cíveis21. A esse respeito, o Professor Lucas Rocha Furtado traz razoável entendimento,
o qual preleciona que no caso de trânsito em julgado, não cabe a reparação em
respeito à coisa julgada; mas no caso de decisões liminares incorretas, concedidas ou
não, a indenização é viável. 451 Destarte, não tem como subsistir essa cultura de irresponsabilidade do
Judiciário enquanto Poder Público, que é contrária a ditames como o da
transparência, prestação de contas, dentre outros, pois com o avanço e fortalecimento
da democracia brasileira, veio junto a ideia do accountability. Destaca Anna Maria
Campos (1990) essa relação estreita do accountabilily com a democracia, justificando
que não há como se exigir prestação de contas, nem responsabilização, em regimes
ditatoriais. Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Nas palavras de Luiz Carlos Bresser Pereira (1990, p. 49-96): Desse modo, como visto no decorrer deste artigo, irrefutável que o Judiciário,
amparado pelo ativismo judicial, muitas vezes se comporta como o próprio
governante, decidindo indevidamente acerca de políticas públicas, sem, contudo,
estar submetido ao regime de responsabilização dos outros poderes, o que valida a
necessidade de se discutir com mais afinco o papel do judiciário, os seus limites e a
possibilidade de responsabilização. Nesse sentido, Raup e Pinho (2014) explicam que a questão da prestação de
contas explica apenas parte do conceito de accountability, cujo significado é mais
abrangente,
envolvendo
"responsabilidade
(objetiva
e
subjetiva),
controle,
transparência, obrigação de prestação de contas, justificativas para as ações que foram
ou deixaram de ser empreendidas, premiação e/ou castigo" (RAUP E PINHO, 2014,
p. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 Ato legislativo. Inconstitucionalidade. Responsabilidade civil do Estado.
Cabe responsabilidade civil pelo desempenho inconstitucional da função
do legislador” (STF, RE nº 158.962, Rel. Min. Celso de Mello, RDA
191/175). No mesmo sentido: STF, RE 153.464, Rel. Min. Sepúlveda
Pertence. JSTF 189/14: “O Estado responde civilmente por danos causados
aos particulares pelo desempenho inconstitucional da função do legislar.
(grifo nosso) 3. DA POSSIBILIDADE DE RESPONSABILIZAÇÃO DO ESTADO POR
ATOS DO JUDICIÁRIO EM TEMPOS DE PANDÊMIA 145) 452 Nessa esteira, a nossa construção para o entendimento de que é possível a
responsabilização do Estado por atos do Poder Judiciário, partiu de aspectos
analisados da decisão monocrática proferida na ADPF nº 701 e da premissa do
equilíbrio entre os Poderes, aplicando-se analogicamente o entendimento
jurisprudencial e doutrinário quanto à responsabilidade do Poder Legislativo, em
casos de atos legislativos inconstitucionais, admitida pelo STF: Ato legislativo. Inconstitucionalidade. Responsabilidade civil do Estado. Cabe responsabilidade civil pelo desempenho inconstitucional da função
do legislador” (STF, RE nº 158.962, Rel. Min. Celso de Mello, RDA
191/175). No mesmo sentido: STF, RE 153.464, Rel. Min. Sepúlveda
Pertence. JSTF 189/14: “O Estado responde civilmente por danos causados
aos particulares pelo desempenho inconstitucional da função do legislar. (grifo nosso) Ato legislativo. Inconstitucionalidade. Responsabilidade civil do Estado. Cabe responsabilidade civil pelo desempenho inconstitucional da função
do legislador” (STF, RE nº 158.962, Rel. Min. Celso de Mello, RDA
191/175). No mesmo sentido: STF, RE 153.464, Rel. Min. Sepúlveda
Pertence. JSTF 189/14: “O Estado responde civilmente por danos causados
aos particulares pelo desempenho inconstitucional da função do legislar. (grifo nosso) Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Ademais, a doutrina majoritária admite ainda a responsabilidade legislativa
pela edição das chamadas leis meramente formais ou leis de efeitos concretos. Nesse
caso, o ato apenas formalmente se apresenta como lei; na prática, cuida-se de
verdadeiro ato administrativo que impõe a pessoas determinadas algum encargo
(SCATOLINO; TRINDADE, 2019, p. 1009) Ora vejamos, se da decisão monocrática estudada acabou por substituir as
condutas adotadas pelos gestores públicos quanto à restrição a realização de práticas
religiosas em templos e igrejas, presume-se que o judiciário, em verdade, culminou
realizando um ato administrativo no lugar administrador público (Poder Executivo),
estando, contudo, submetido aos mesmos limites fixados anteriormente pelo STF,
como pautar as suas decisões em recomendações técnicas e fundamentadas, sob pena
de se tornarem corresponsáveis por eventuais violações a direitos. Outrossim, de igual modo, em sendo admissível a responsabilidade legislativa
em casos de atos legislativos inconstitucionais, razoável também admitir-se a
responsabilidade judiciária por decisões monocráticas de Ministros do STF tidas por
inconstitucionais, assim como outras decisões teratológicas oriundas do Poder
Judiciário, flagrantemente violadoras de direitos constitucionais, em função da
dimensão positiva ensinada pelo ilustre mestre Lisboa, a qual vincula a atuação dos
juízes e tribunais, por meio da aplicação, interpretação e integração, a conferir às
normas de direitos fundamentais a maior eficácia possível. 453 Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. ,
ç
,
j
ISSN - 2675 – 3375 Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021. 4. CONCLUSÃO Por todo o exposto, em que pese sejam previstas no ordenamento jurídico e
extremamente necessárias em casos de urgência, as decisões monocráticas em sede
do Supremo Tribunal Federal restaram banalizadas, representando 82% do total das
decisões da Corte Constitucional, enfraquecendo o seu papel enquanto Guardião da
Constituição perante à sociedade, de modo que reforça os argumentos de violação ao
princípio democrático e à separação dos poderes quando desrespeita à colegialidade
da Corte desnecessariamente, refletindo na mitigação de sua independência e Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE imparcialidade, características primordiais para que uma instância funcione como
Guardião da Constituição. Destarte, da análise da decisão monocrática proferida na ADPF 701/MG,
infere-se que está é formalmente inconstitucional, tendo em vista que violou
diversos precedentes vinculantes estabelecidos pelo própria Corte Constitucional, em
contrariedade ao dever dos tribunais de manter a sua jurisprudência estável, íntegra e
coerente. A esse respeito, deixou de considerar a ilegitimidade ativa da ANAJURE nos
termos do ADPF nº 703/BA, quando não, ofendeu a obrigatoriedade de
fundamentação da decisão imposta aos juízes ao não realizar devidamente o
distinguishing do precedente, assim como não observou a decisão do Plenário que
reconheceu a competência concorrente de todos os entes para a adoção de medidas de
enfrentamento à pandemia, quando interferiu diretamente na autonomia de entes
públicos ao impedir a restrição da prática religiosa em cultos e igrejas, em afronta ao
princípio da separação dos poderes. 454 Nesse ponto, constatou-se ainda a inconstitucionalidade material da decisão,
violando o direito fundamental à saúde em sua dimensão negativa e positiva,
atuando o Ministro quando deveria abster-se de praticar atos que causaram
embaraços e dificultaram a busca pela garantia do direito à saúde e à vida, por meio
da adoção de medidas de restrição, com vistas a amenizar os efeitos decorrentes da
Covid-19 e que possui eficácia comprovada pela Organização Mundial da Saúde –
OMS. ,
ç
,
j
ISSN - 2675 – 3375 4. CONCLUSÃO Desta feita, de acordo com os parâmetros fixados pelo STF em sede de
controle de constitucionalidade da MP nº 966/2020, é possível afirmar que o
Ministro incorreu em erro grosseiro, porquanto ao substituir os gestores públicos em
suas ações, não fundamentou a sua decisão com base em normas e critérios
científicos e técnicos, assim como não observou os princípios constitucionais da
prevenção e precaução, tanto que revogou a liminar concedida para se adequar ao
entendimento posteriormente fixado pelo STF em sentido contrário ao que havia ,
ISSN - 2675 – 3375 Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE Revista Ibero- Americana de Humanidades, Ciências e Educação- REASE decidido o Ministro, no entanto, era ululante a inconstitucionalidade, não havia
necessidade do Plenário afirmar o óbvio. Observou-se ainda a inconstitucionalidade material devido à violação do
princípio da separação dos poderes à luz do entendimento fixado pelo Supremo
Tribunal Federal na ADI nº 6341/DF, considerando que decisão monocrática
adentrou no mérito das ações adotadas pelos gestores públicos no combate ao covid-
19, em que pese o STF tenha atribuído competência concorrente a todos os entes,
ressaltando expressamente que o desrespeito a autonomia de quaisquer dos Estados,
DF e Municípios representaria afronta ao princípio da separação dos poderes. Dessarte, no campo da possibilidade de responsabilização do Judiciário,
ressalta-se que a jurisprudência atual e a doutrina caminham para a mitigação da
regra que prevê a irresponsabilidade do Estado por decisões judiciais, tendo em vista
que está em dissonância com os preceitos vitais da democracia. Como dito alhures, se é possível a responsabilização do Estado pelo
desempenho inconstitucional da função de legislar e em decorrência de leis de efeitos
concretos, não se vislumbra óbice para que seja possível a responsabilização do
judiciário por decisões judiciais precárias reputadas flagrantemente inconstitucionais,
que porventura violem direitos fundamentais, em quaisquer de suas dimensões,
negativas e positivas. 455 Assim, no atual contexto pandêmico, levando-se em consideração que o
Supremo Tribunal Federal já consolidou entendimento acerca dos requisitos a serem
observados pelos agentes públicos quando da adoção de medidas de enfrentamento ao
Covid-19, sob pena de se tornarem corresponsáveis por eventuais violações a direitos,
plausível à possibilidade da corresponsabilização de membros do judiciário, enquanto
agentes públicos, quando atuarem fora dos limites da judicatura, em afronta ao
princípio da separação dos poderes. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 REFERÊNCIAS BIBLIOGRÁFICAS DWORKIN. Ronald. Levando os direitos a sério. Tradução de Nelson Boeira. Sã
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disparition?. In Revue Française de droit constitutionnel, nº 68, outubro de 2006. Ed. PUF, Paris, p. 713/736. SCHMITT, Carl. O guardião da Constituição. Trad. Geraldo de Carvalho. Belo
Horizonte: Del Rey, 2007, p. 176-177 MIRANDA, Jorge. Manual de direito constitucional. Volume II. 2. ed. Coimbra:
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dos Tribunais, 2013. p. 64. FURTADO, Lucas Rocha. Curso de Direito Administrativo. Belo Horizonte:
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aposta nos precedentes vs. Uma compreensão constitucionalmente adequada do seu
uso no Brasil. In: MENDES, Aluisio Gonçalvez de Castro; MARINONI, Luiz
Guilherme; WANBIER, Teresa Arruda Alvim (orgs.). Direito Jurisprudencial. São
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informação. Revista Gestão e Planejamento. v. Revista Ibero-Americana de Humanidades, Ciências e Educação. São Paulo, v.7.n.6. jun. 2021.
ISSN - 2675 – 3375 REFERÊNCIAS BIBLIOGRÁFICAS 15, n. 1, jan./abr. 2014, p. 144-161. SARLET, Ingo Wolfgang. A eficácia dos direitos fundamentais. 10ª. ed., rev., atual. e
ampl. Porto Alegre: Livraria do Advogado, 2009, p. 294. SCATOLINO, Gustavo; TRINDADE, João. Manual didático de direito
administrativo. 7ª ed. Ver., ampl. E atual. – Salvador: Ed. JusPodivm, 2019, p. 1009. POGREBINSCHI, Thamy. Ativismo Judicial e Direito: Considerações sobre o
Debate Contemporâneo. Revista Direito, Estado e Sociedade, nº 17, agosto-dezembro de
2000, pág. 2. CAVALCANTE, Márcio André Lopes. Análise da constitucionalidade da MP
966/2020, que dispõe sobre a responsabilização de agentes públicos por atos
relacionados com a pandemia da covid-19. Buscador Dizer o Direito, Manaus. Disponível
em:
<https://www.buscadordizerodireito.com.br/jurisprudencia/detalhes/815104ed949f9
deaf45165c4b8337013>. Acesso em: 19/04/2021 BRASIL. Supremo Tribunal Federal. Ação Direta de Inconstitucionalidade nº
4.356/CE – Distrito Federal. Relator Min. Dias Toffoli. Julgada em 09/02/2011. Disponível em: <https://stf.jusbrasil.com.br/jurisprudencia/19735377/acao-direta-de-
inconstitucionalidade-adi-4356-ce>. Acesso em: 17/04/2021. 457 BRASIL. Supremo Tribunal Federal. Ação Direta de Inconstitucionalidade nº
3.146/DF – Distrito Federal. Relator Min. Joaquim Barbosa. Julgada em 09/02/2011. Disponível em: <https://stf.jusbrasil.com.br/jurisprudencia/14732484/acao-direta-de-
inconstitucionalidade-adi-3146-df>. Acesso em: 17/04/2021. BRASIL. Supremo Tribunal Federal. Ação Direta de Inconstitucionalidade nº
6.341/DF – Distrito Federal. Relator Min. Min. Marco Aurélio. Julgada em
03/07/2020. Disponível
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inconstitucionalidade-adi-6341-df-distrito-federal-0088693-7020201000000>. Acesso
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11/11/2020. Disponível
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<https://stf.jusbrasil.com.br/jurisprudencia/1124668418/arguicao-de-
descumprimento-de-preceito-fundamental-adpf-703-ac-0096437-
1920201000000/inteiro-teor-1124668425>. Acesso em: 17/04/2021. BRASIL. Supremo Tribunal Federal. Arguição de Descumprimento de Preceito
Fundamental nº 696/DF – Distrito Federal. Relator Min. Marco Aurélio. Julgado em
19/06/2020. Disponível
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descumprimento-de-preceito-fundamental-adpf-696-df-0095736-
5820201000000/inteiro-teor-1106712617>. Acesso em: 17/04/2021. <https://stf.jusbrasil.com.br/jurisprudencia/1106712603/arguicao-de-
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5820201000000/inteiro-teor-1106712617>. Acesso em: 17/04/2021. BRASIL. Supremo Tribunal Federal. Ação Direta de Inconstitucionalidade nº
6421/DF – Distrito Federal. Relator Min. Roberto Barroso. Julgado em 20 e
21/05/2020. Informativo
978. Disponível
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<http://www.stf.jus.br/arquivo/informativo/documento/informativo978.htm>. Acesso em: 18/04/2021. BRASIL. Senado Federal. Proposta de Emenda à Constituição nº 8/2021. Distrito
Federal. Disponível
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https://legis.senado.leg.br/sdleg-
getter/documento?dm=8118687&ts=1618604122261&disposition=inline>. Acesso em:
19/04/2021. BRASIL. Supremo Tribunal Federal. Reclamação nº 41.935/MT – Distrito Federal. Relator
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Asymmetry in information acquisition—Exploring the principal–agent dyad of sport organizations and sport management higher education institutions
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German journal of exercise and sport research
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Asymmetry in information
acquisition—Exploring the
principal–agent dyad of sport
organizations and sport
management higher education
institutions Asymmetry in information
acquisition—Exploring the
principal–agent dyad of sport
organizations and sport
management higher education
institutions growth and respond to emerging oppor-
tunities. A specific characterization of
qualifications and competencies needed
for employment in this industry, on the
other hand, is non-existent (Dowling,
2018). Additionally, higher education
tends to slowly anticipate and adapt to
rapidly changing requirements of in-
dustry (Chan, Fong, Luk, & Ho, 2017;
European Commission, 2019). Germany, we acquire and report cru-
cial information for sport management
higher education (SMHE) to use for pro-
gramme design and delivery. In applying
this facet of agency theory to sport man-
agement research, we also contribute to
the discussion and further development
of agency-theory more generally. Fur-
thermore, ourstudyoffersdirectimplica-
tions for sport management faculty and
sport management programme coordi-
nators. Finally, (prospective) sport man-
agement students and graduates, practi-
tioners in the sport management indus-
try and policy makers benefit from our
findingsconcerningthequalificationand
competency requirements of sport man-
agersaswellasthepresupposedrolesport
management HEIs play therein. Introduction Higher education institutions (HEIs)
have reacted to the demand for educa-
tion in the field of sport management
with an increasing number of academic
programmes. To achieve the goals of
the Bologna Declaration, the European
HigherEducationArea requires dialogue
between industry and higher education,
as the latter directly contributes to grad-
uates’ personal development, sustainable
employability
and
active
citizenship
(Vossensteyn et al., 2018). Specialized third parties,
such as
labour market research entities, can help
to mitigate these problems by acquiring
and reporting the necessary information
(Lewis & Sappington, 1997). With this
study we adopt this role and aim to: Our study departs from an agency
theoretical
perspective
and
assumes
a metaphorical
contract
within
the
principal–agent dyad,
wherein sport
organizations
(the
principal)
engage
sport management HEIs (the agent) in
adequately qualifying graduates for em-
ployment. Distinctive for this type of
relationship, problems concerning diver-
gence of interest as well as asymmetrical
distribution of information between the
two are assumed. In addition, we assume
asymmetry in information acquisition,
which results from the agent’s ignorance
and inability to obtain the information
that is necessary to efficiently fulfil the
metaphorical contract(Lewis &Sapping-
ton, 1993). The global sport industry,
on the one hand, requires a highly qual-
ified and flexible workforce to sustain 1. Resolve information acquisition
problems of sport management HEIs
by identifying and exploring which
field-adequate qualifications and
competencies are required of sport
management graduates, and “Sport managers”?—A review
of current literature Olivia Wohlfart
· Sandy Adam · Gregor Hovemann
Faculty of Sport Science, Institute for Sport Psychology and Sport Pedagogy, Department of Sports
Economics and Sports Management, Leipzig University, Leipzig, Germany Olivia Wohlfart
· Sandy Adam · Gregor Hovemann
Faculty of Sport Science, Institute for Sport Psychology and Sport Pedagogy, Department of Sports
Economics and Sports Management, Leipzig University, Leipzig, Germany Ger J Exerc Sport Res 2021 · 51:344–353
https://doi.org/10.1007/s12662-021-00722-w
Received: 8 October 2020
Accepted: 23 April 2021
Published online: 18 June 2021
© The Author(s) 2021 “Sport managers”?—A review
of current literature 2. Clarify the role of sport management
HEIs in diligently fulfilling the
metaphorical contract of preparing
sport management graduates. Induced by increasing commercializa-
tion and professionalization of the sport
industry since the 1970s, academics
around the world concluded the need
for a scientific sport management dis-
cipline and the development of specific
study programmes. As a result, there has
been a steady growth in sport manage-
ment programmes globally (Costa, 2005;
Kaiser & Beech, 2012; Shilbury, Philips,
Karg, & Rowe, 2017). From the outset, Our study contributes to the body of
knowledge, by applying the problem of
asymmetrical information acquisition to
sport management and higher educa-
tion research. Through content analy-
sis of 12 semi-structured interviews with
representatives of sport organizations in 344 German Journal of Exercise and Sport Research 3 · 2021 Assignment to qualify graduates of sport management
programmes adequate to the needs of sport organizations in
the labour market
Incentive
problems
Performance through competency-oriented
qualifications within sports management programmes
Asymmetry in information acquisition
Assigns third party to
acquire information
Insitutional Enviroment
(European, national, federal)
Third Party
(Labour Market
Research Entity)
Agent
(Sport Management
Higher Education
Institutions)
Principal
(Sport
Organizations)
Ignorance
Provides agent crucial
planning information to
be able to fulfil the
contract
Signals expectations
concerning
metaphorical contract
Metaphorical
Contract
Fig. 1 9 Metaphorical con-
tractual relationship be-
tween sport organizations
andsport management
highereducation institu-
tions (ourillustration based
on Jensen &Meckling,
1976;Lewis &Sappington,
1997) Assignment to qualify graduates of sport management
programmes adequate to the needs of sport organizations in
the labour market Performance through competency-oriented
qualifications within sports management programmes Fig. 1 9 Metaphorical con-
tractual relationship be-
tween sport organizations
andsport management
highereducation institu-
tions (ourillustration based
on Jensen &Meckling,
1976;Lewis &Sappington,
1997) Fig. 1 9 Metaphorical con-
tractual relationship be-
tween sport organizations
andsport management
highereducation institu-
tions (ourillustration based
on Jensen &Meckling,
1976;Lewis &Sappington,
1997) Third Party
(Labour Market
Research Entity) (moral
hazard) settings
(Eisenhardt,
1989). and Packheiser and Hovemann (2013),
who analysed the requirements stated in
job advertisements for sport managers in
Australia and Germany, respectively. We
therefore conclude that an information
acquisition problem exists in the prin-
cipal–agent dyad of sport organizations
and sport management HEIs. the legitimacy of these programmes has
been critically discussed (among others:
Chalip, 2006; DeSensi, Kelley, Blanton,
& Beitel, 1990; Dowling, 2018). “Sport managers”?—A review
of current literature Field
dynamics and the heterogeneity of sport
organizations have made it difficult to
clearly define the sport management
profession and related qualifications and
competencies (Dowling, 2018). There-
fore, sport management HEIs find them-
selves in a fierce competition for students
with other vocational training institu-
tions (Kaiser & Schütte, 2012). Common
job requirements revolve around sector-
specific experience, an affinity for sports,
passion, and endurance (Packheiser &
Hovemann, 2013; Wohlfart & Adam,
2019). Therefore, future sport managers
need to possess a professional blend of
generic management competencies and
competencies specific to the sport in-
dustry (Barcelona & Ross, 2004; Emery,
Crabtree, & Kerr, 2012;
Fahrner &
Schüttoff, 2020; Jiang & Alexakis, 2017;
Mathner & Martin, 2012). Lewis and Sappington (1997) suggest
that the principal prefers contracts which
either provide accurate planning infor-
mation to the agent, such as detailed
product specifications,
or encourage
them to gather planning information,
when an optimal level of activity for
the agent is sensitive to the output level
that maximizes the principal’s benefits
and when informational rents from ac-
curate planning information are limited
for the agent. The agent, meanwhile,
decides whether they want to share
the principal’s information or acquire
superior information, and if so, how
much (Kessler, 1998). Devoting more
resources to research would likely pro-
vide the agent with valuable planning
information, but the financial cost of
information acquisition limits the extent
of these activities. Additionally, research
activities might not always result in
desired outcomes, and the agent may re-
main uninformed or ignorant about the
prevailing environment (Kessler, 1998;
Lewis & Sappington, 1993). Abstract This study is based on a metaphorical
contract in sport management, whereby
sport organizations (the principal) engage
sport management higher education
institutions (the agent) in qualifying
graduates according to industry demands. There is asymmetry in the contract due
to the lack of incentives for the principal
to provide specific information and the
agent’s ignorance about this information. A third party can acquire crucial information
that sport management higher education
institutions need to fulfil the contract. Based on a qualitative content analysis
of 12 interviews with sport organization
representatives, the study finds that sport
management higher education institutions
need to focus on either developing generic
competencies or credibly signalling the
value of sport management-specific
competenciesin their curriculum in order to
efficiently fulfil the contract. A principal–agent dyad: sport
organizations and SMHE Althoughagencytheoryoriginatedinthe
field of economics, it has been widely ap-
plied, both theoretically and empirically,
inmanydifferentdisciplines since ‘exam-
ples of agency are universal’ (Ross, 1973,
p. 134). A growing number of authors
are using agency theory as a conceptual
framework, heuristic tool or organizing
concept in the higher education context
(Gornitzka, Stensaker, Smeby, & de Boer,
2004; Kivistö, 2008; Kivistö & Zalyevska,
2015; Liefner, 2003). However, most of
these studies employ common agency
theory models, which assume asymmet-
rical distribution of information rather
than asymmetrical gathering of informa-
tion. Ger J Exerc Sport Res 2021 · 51:344–353
https://doi.org/10.1007/s12662-021-00722-w
© The Author(s) 2021 Ger J Exerc Sport Res 2021 · 51:344–353
https://doi.org/10.1007/s12662-021-00722-w
© The Author(s) 2021 before acting and to subsidize the cost of
information acquisition for the agent. In
most cases, the agent can both acquire
valuable planning information and ful-
fil the duties of the contract, since they
are likely to benefit from economies of
scope. However, Lewis and Sappington
(1997) suggest that separation of the two
activities may eliminate problems of ig-
norance. Ifathirdpartygathersinforma-
tionabouttheenvironment, theprincipal
can ensure the agent is informed while
simultaneously inducing them to work
diligently. This is where our study de-
parts with a focus on the principal–agent
dyad within SMHE. tionallogics, sharingtheneedforahighly
qualified and flexible workforce. While
a sport marketer is interested in profit
maximization, a non-profit sport asso-
ciation aims to maximize utility for its
stakeholders, and a professional sport
club pursues both interests simultane-
ously. The agent in our dyad repre-
sents HEIs with sport management pro-
grammes. While the institutional pres-
sures exerted on higher education (i.e. the Bologna Process)have partlyresulted
inconvergenceofformerlydivergentlog-
ics of research and teaching, diversity
still exists. Whereas the primary role
of public universities remains ensuring
‘excellence, prestige, and influence’ in re-
search (Bowen, 1981, p. 20) and provid-
ing education with academic freedom,
this does notnecessarilycharacterize pri-
vate HEIs with profit-maximizing goals. We assume that SMHEfails to quickly re-
spond to industry demands as a result of
problematic asymmetry in information
acquisition (Crémer et al., 1998; Kessler,
1998; Lewis & Sappington, 1993). O. Wohlfart · S. Adam · G. Hovemann
Asymmetry in information
acquisition—Exploring the
principal–agent dyad of sport
organizations and sport
management higher
education institutions Agency theory Agency theory (Jensen & Meckling,
1976) is a relevant analytical framework
for all kinds of contractual relationships
between at least two parties in which
one party(the principal)engages another
party (the agent) in performing a service
on behalf of the principal. These con-
tractual arrangements are characterized
by so-called agency problems, which
are most often caused by asymmetrical
information distribution between the
two parties (Crémer, Khalil, & Rochet,
1998). Most agency models assume that
the agent is fully informed about their
characteristics, intentions and actions,
whereas the principal is not. This in-
formation structure typically allows the
agent to act opportunistically at the ex-
pense of the principal in precontractual
(adverse selection) and postcontractual Competency research in the sport in-
dustry appears to be a complex endeav-
our, since empirical data have mainly
been generated by individual graduate
studies rather than objective industry re-
quirements. There are only a few excep-
tions, which include Emery et al. (2012) As they are aware of the possibility of
ignorance and undesired outcomes, the
principal is willing to motivate the agent
to acquire valuable planning information 345 German Journal of Exercise and Sport Research 3 · 2021 Abstract Method We employed a purposeful sampling
strategy based on the work of Pat-
ton (2015), selecting individuals with
senior positions such as managing di-
rector, chief executive officer (CEO), and
general secretary. To gain a holistic per-
spective of the sport industry, four main
stakeholder categories were included:
(1) professional sport clubs (C), (2) non-
profit sport clubs and federations (NP),
(3) private sector organizations (P) and
(4) public sector organizations (PU). The selected organizations were repre-
sentative of their sector, and their size
varied from medium to large (Bayle &
Robinson, 2007). In addition to institu-
tionalized internships, the organizations
offerjobopportunitiesforcareerentrants
as well as experts in specific fields. Most
positions focus on general management,
project management or aspects of com-
munication
management (marketing,
sponsoring), and full-time commitment
is expected (. Table 1).h Following a social-constructivist epis-
temology and post-qualitative perspec-
tives on research, we acknowledge that
theory-free knowledge does not ex-
ist (Smith & Hodkinson, 2009). The
lead author analysed and interpreted
the coded segments in regard to the
research aim (qualifications, competen-
cies, role of sport management HEIs,
information asymmetry). To ensure
rigour in the overall research and data
analysis process, we employed ‘critical
friends’ and self-reflexivity, as advo-
cated by Smith and McGannon (2018). The role of a critical friend is ‘not to
“agree” or achieve consensus, but rather
to encourage reflexivity by challenging
each other’s construction of knowledge’
(Cowan & Taylor, 2016, p. 508). This
provides opportunities for rich dialogue
about interpretive possibilities and re-
quires researchers to make their thought
processes explicit. This study involved qualitative data anal-
ysisof12semi-structuredinterviewswith
representatives of sport organizations in
Germany. Thesemi-structuredinterview
guide was developed within the second
phase of the NASME research project
based onreviewed literature as well as the
resultsofthequantitativeanalysis(Wohl-
fart & Adam, 2019; Wohlfart, Adam, &
Hovemann, 2019). Semi-structured in-
terviews have the strength of providing
a detailed understanding of topics and
social settings as well as flexibility in the
interview process based on the intervie-
wees’ background, experience and sta-
tus (Denzin & Lincoln, 2011). Previous
studies withinthe examined principal–a-
gentdyad have applied quantitative ques-
tionnaires (Fahrner & Schüttoff, 2020;
Mathner & Martin, 2012; Schlesinger,
Studer, &Nagel, 2016)ordocumentanal-
ysis (Emery et al., 2012; Packheiser &
Hovemann, 2013). As we aim to better
grasp the relationship between the ac-
tors, we want to understand the how and
the why and therefore apply a qualitative
approach. Asymmetrical information
acquisition Sport organizations’ preferred informa-
tion structure does not provide specific
planning information to sport manage-
mentHEIsorinducethemtoacquiresuch
information. We argue that is due to in-
centive problems for sport organizations,
because they believe that an optimal level
of activity for SMHE is not sensitive to
the output level that maximizes sport or-
ganizations’ benefits. When recruiting
qualified graduates, sport organizations
canselectcandidatesfromawiderangeof
study programmes, not just those offered
by SMHE (Kaiser & Schütte, 2012). In-
deed, sportorganizationshavefewincen-
tives to recruit sport management grad-
uates or inform HEIs about the quali-
fications, competencies, skills and abili-
ties they require. This might be due to
ambiguity about whether SMHE is a dis-
tinct academic discipline or a profession
(Chalip, 2006; Dowling, 2018). Keywords
Agency theory · Agent ignorance · Labour
market · Employability· Competencies Keywords
Agency theory · Agent ignorance · Labour
market · Employability· Competencies Weassumetheexistenceofametaphor-
ical contract within a principal–agent
dyad, wherein sport organizations (the
principal) engage sport management
HEIs (the agent) in adequately qualify-
ing graduates for employment (. Fig. 1). This assumption is grounded in the for-
mal and informal pressures exerted on
both parties by their institutional en-
vironments (European, national, local)
and cultural expectations (DiMaggio
& Powell, 1983). HEIs are responsible
for adequately qualifying graduates to
function as valuable citizens (European
Commission, 2019; Kultusministerkon-
ferenz, 2000).h mation provided by sport organizations
or acquire superior planning informa-
tion. We argue that sport management
HEIs decide to remain uninformed or ig-
norant (Kessler, 1998; Lewis & Sapping-
ton, 1993) due to the information costs
of conducting labour market research,
which are perceived to be excessive. We assert that sport organizations are
willing to subsidize the cost of infor-
mation acquisition for SMHE to avoid
ignorance in sport management HEIs
(Kessler, 1998). However, since sport
organizations assume that unified infor-
mation acquisition and contract fulfil-
ment by these HEIs produce undesired
outcomes, they signal to the institutional
environment that a third party, such as As a consequence of the contract de-
sign, sportmanagementHEIsremainun-
informed about industry demands and
may decide whether to take the infor- The principal in our dyad is a diverse
set of sport organizations with converg-
ing and diverging interests and organiza- 346 German Journal of Exercise and Sport Research 3 · 2021 specialized labour market research en-
tities, should be responsible for infor-
mation acquisition. Asymmetrical information
acquisition The EU-funded re-
search project “New Age of Sport Man-
agementEducationinEurope”(NASME)
was tasked with acquiring information
about qualification and competency re-
quirements as well as sport management
HEIs’ role in developing these require-
ments. From 2017–2019, research teams
from nine European countries partnered
to question and enhance SMHE based
on industry demands. The project was
divided into two consecutive data collec-
tionphases, withanoverall sample of635
responses by experts in the sport man-
agement labour market in the nine part-
ner countries. After acquiring informa-
tion, NASME was to publicly report their
findings so that sport management HEIs
could use this information to diligently
fulfil the metaphorical contract (Lewis &
Sappington, 1997). the world as the informants perceive it
(Brinkman & Kvale, 2014). Data analysis We performed qualitative content analy-
sis of the 12 interview transcripts in ac-
cordancewithMayring(2015). Ofeleven
main questions, six focused on compe-
tencies, qualification requirements and
the role of HEIs in their realization, but
the data analysis included all interview
data. The lead author repeatedly read the
transcripts and coded segments using
MAXQDA Analytics Pro 2018 (VERBI
Software. Consult. Sozialforschung
GmbH, Berlin, Germany). Coding em-
ployed deductive categories identified
from the literature review and theoretical
background (e.g. divergence of interest;
information asymmetry, qualifications;
competencies) as well as inductive cate-
gories related to the main research in-
terest that emerged from the transcribed
interview materials (e.g. expectations,
empathy, assessment). The qualitative
data analysis resulted in 67 codes and
927 coded segments. The interview guide consisted of
11 open-ended questions that related to
four main themes: trends, competencies,
job development and recruitment. In
addition, we used a short questionnaire
to obtain sociodemographic information
about the informants. Three interview-
ers conducted the interviews between
December 2018 and February 2019,
with the lead author assuming the main
responsibility. The interviews lasted be-
tween 34 and 58min, and they were
audio-recorded and transcribed verba-
tim according to a specific transcription
guideline based on that described by
Kuckartz (2010). The lead author then
proofread and anonymized the tran-
scripts. We generated 126 pages of
single-spaced transcribed text and a to-
tal interview time of 550min. Method (Sport Sciences)
20/11
MBA master’s of business administration, CEO chief executive officer, VP vice president
aBased on Bayle & Robinson (2007), organizational size according to the number of paid staff (5–10= small, 11–40= medium, 41–100= big, >100= large) MBA master’s of business administration, CEO chief executive officer, VP vice president
aBased on Bayle & Robinson (2007), organizational size according to the number of paid staff (5–10= small, 11–40= medium, 41–100= big, >100= large) MBA master’s of business administration, CEO chief executive officer, VP vice president
aBased on Bayle & Robinson (2007), organizational size according to the number of paid s sible misunderstandings (Denzin & Lin-
coln, 2011). Qualification and competency
requirements do here, yes. Whether it is someone with
a bachelor’s, a master’s or another degree,
would make no difference to me. (P-3) According to the interviewees, there are
nospecific qualifications required forpo-
sitions at sport organizations: Method When gathering information
using the method of qualitative inter-
views, the researcher tries to understand We extracted quotes from the inter-
view transcripts to illustrate the find-
ings and interpretations. All interviews
were held in German, and a professional
translator translated the quotes into En-
glish. The lead author examined the
translationstoensurethattheinformants’
voices were maintained and avoid pos- The 12 informants were 34–52 years of
age and had academic degrees, mostly in
sport management or sport economics. They had 9–25 years of work experience,
with 3 months to 25 years at their current
organization. 347 German Journal of Exercise and Sport Research 3 · 2021 Table 1
Socio-demographic information of Interviewees (n=12)
Pseudo-
nym
Stakeholder category
Gender
Position
Organizational sizea Education
(Field)
General/specific
work experience
(in years)
C-1
Professional Sport Club
Male
Managing Director
Large (230 staff)
Bachelor
(Economics)
20/5
C-2
Professional Sport Club
Male
Managing Director
Large (160 staff)
MBA
25/9
C-3
Professional Sport Club
Male
Managing Director
Medium (17 staff)
German Diploma
(Sport Education)
12/12
NP-1
Non-Profit Organization
Male
General Secretary
Big (44 staff)
German Diploma
(Sport Sciences)
14/4
NP-2
Non-Profit Organization
Male
Commercial Manager Medium (38 staff)
Magister
(Sport & Economics)
13/7
NP-3
Non-Profit Organization
Female
CEO
Large (170 staff)
German Diploma
(Sport Education)
22/1
P-1
Private Organization
Male
Senior VP
Large (57,016 staff)
German Diploma
(Sport Economics)
24/0.25
P-2
Private Organization
Male
Senior Project Man-
ager
Large (640 staff)
German Diploma
(Sport Economics & Sport
Education)
25/25
P-3
Private Organization
Male
Senior Team Head
Large (12,192 staff)
German Diploma
(Sport Education)
13/6
PU-1
Public Organization
Male
Professor
Medium (24 staff)
Dr. (Sport Management & Sport
Sciences)
9/2
PU-2
Public Organization
Male
Freelancer/Head of
Department
Medium (34 staff)
German Diploma
(Economics)
25/10
PU-3
Public Organization
Male
Professor
Large (341 staff)
Dr. Findings and discussion One interviewee from a non-profit sport
organization (NP-2) mentioned the im-
portance of cooperation with a specific
educational programme for selecting fu-
ture sport managers. The same intervie-
wee negated the importance of certifi-
cates: In this section, we report and discuss the
findings of our qualitative content anal-
ysis. Following the social-constructivist
approach, we analysed and interpreted
the content of the interviews. Following
our research aim of resolving informa-
tion acquisition problems of sport man-
agement HEIs, we examined the per-
spective of key informants from sport
organizations concerning field-adequate
qualifications and competencies as well
as the role of sport management HEIs
in preparing sport management gradu-
ates for employment in these. By pro-
viding planning information to SMHE,
we reduce agent ignorance and promote
fulfilment of the metaphorical contract
described earlier. Well, I believe that sport management cer-
tainly is a good prerequisite. [...] I don’t
believe that what a sport manager brings
along compared to a business administra-
tion graduate is a hiring criterion to the
left or to the right. For us, both are pro-
grammes that offer the prerequisites for
being hired, as well as some other study
programmes. (C-2) I have a look at what he or she has done. And also, the important things, such as
personality. If I read that someone trav-
elled for a year, I think it is sensational. [...] So, I prefer that over someone who
somehow did 10 quick internships in
a row. [...] So, in practical terms: When
it comes to applications, I don’t even have
a look at any certificates. (NP-2) The informants showed nopreference for
graduatesfromaspecifictypeofHEI(e.g. public or private, full-time or part-time). Likewise, theydidnotvaluespecificstudy
programmes and degrees over others: The interviewees did not value a mas-
ter’s degree over a bachelor’s degree in
sport management. This finding sub-
stantiates early research in the field. For To be honest, it would be all the same to
me. [...] What matters to me is that there
is a passion for what he [or she] wants to 348 German Journal of Exercise and Sport Research 3 · 2021 example, DeSensi et al. (1990) pointed
out the conflict between ‘what college/
university faculty suggested as recom-
mended degree levels and what business/
agency personnel suggested as required
degree levels’ (p. 56). Findings and discussion This divergence
of interest has been immanent since the
foundation of the first academic pro-
grammes and presents a typical agency-
problem(Akerlof, 1970). Theprolonging
situationof“over-education”withinsport
managementwould indicate opportunis-
tic behaviour of SMHE in consciously
offering highly specialized graduate pro-
grammes which students do not neces-
sarily need for employability (Meroni &
Vera-Toscano, 2017). supportingtheongoingdiscussiononthe
relevance of sport-management-specific
programmes (Tsitskari, Goudas, Tsalou-
chou, & Michalopoulou, 2017; Zhang,
2015). Leadership, responsibility, pas-
sion and organizational identification
were vital self-competencies for future
sport managers. The interviewees from
non-profit sport organizations empha-
sizedtheimportanceofspecialleadership
skills, since sport managers need to lead
two types of employees: paid staffand
volunteers (NP-1, NP-3). cialized IT skills were less important for
sport management graduates since sport
organizations hire IT specialists. Inconclusion, thekeyinformantsgave
an abundance of in-depth information
concerning field-adequate competence-
requirements for the employment of
sport management graduates. When
and if transmitted to decision makers
within the development of sport man-
agement programmes, this information
can substantially promote the fulfilment
of a competency-based qualification as
required by the metaphorical contract
(Research Aim 1). Most interviewees mentioned soft
skills required of sport managers. This
term was used in various ways, but most
often as a synonym for generic compe-
tencies. We separated the social aspects
of this generic term and highlighted
the complexity of social competencies. Withinthiscategory, teamworkwasiden-
tified as the most important competency
across all sectors: Interestingly, only three interviewees
had a solid understanding of the vari-
ance and diversity of sport management
programmes in Germany (NP-3, PU-1,
PU-3). We interpret this as the re-
sult of asymmetric information between
the contracting partners due to weak
signals from sport management HEIs
concerning the unique value of their
programmes (Jensen & Meckling, 1976;
Spence, 1973). The principal, however,
seems to be highly interested in such
signals: In the attempt to reduce asymmetry
in information acquisition relevant in-
formation for SMHE, we further anal-
ysed the competencies required by in-
terviewees. Based on a modified version
of the competency explorer proposed by
Heyse and Erpenbeck (2004)1 as well as
previous studies (Fahrner & Schüttoff,
2020; Schlesinger et al., 2016), we al-
located coded segments to four compe-
tency classes: self, social and method-
ological competencies (i.e. generic com-
petencies) and professional (sport man-
agement) competencies (i.e. subject-re-
lated competencies). Teamwork [...] is a great skill. 1 We modified the competency classes devel-
opedbyHeyseandErpenbeck(2004)—personal
competencies, activity and action competen-
cies, methods and professional competencies
and socio-communicative competencies—in
order to make the findings comparable to
previousstudies. Findings and discussion On the
other hand, I can tell you that many of our
organizations, you need to picture them as
rather compartmentalized organizations. I will never forget the manager of a re-
gional association who said the only one
to talk to is the mirror because I am the
only one in the office. So, in many places
we also have very, very small organiza-
tions which have one full-time employee,
and the question is, what is teamwork
there? Then, teamwork of course is the co-
operation between full-time and honorary
staff, and then teamwork is one of the key
skills. (NP-3) But if I knew that these skills are taught in
the bachelor’s or master’s degree in sport
management, I might prefer him [/her]
much, much more. Because I say, okay, he
[/she] has exactly these soft skills, team-
work, network, communication, plus can
stand his [/her] ground and can do that
because he [/she] comes from sports. And
has at least basic knowledge in [...] digi-
talization, communication and account-
ing. [...] Then I would much prefer him
[/her] over a pure journalist. Well, but
you don’t know that at that moment. (C-
3) Decision-making capacity was men-
tioned by most interviewees and was
identified as the most critical self-com-
petency. However, the interviewees
ascribed different meanings to decision-
making, such as making firm decisions
(C-2, NP-2, P-1), taking responsibility
for decisions made (C-1, P-3), decision-
making as a team process (C-3, PU-2)
and guiding and preparing decisions as
partofthedecision-makingprocess(NP-
1). Affinity for sport was the only other
crucialself-competencyforallsportorga-
nizations. Interestingly, the interviewees
did not necessarily associate affinity for
sport with a sport management degree, Furthermore, the interviewees from the
professional sport clubs and private-sec-
tor sport organizations highlighted the
importance of (English) communication
skills. Finally, relevant methodological
competencies included applying theoret-
ical knowledge into practice, reflection
and flexibility. The CEO of a national sport federation
agreed: The CEO of a national sport federation
agreed: I’m not interested in whether it’s part-time
or private or at a university [...]. My point
is, if I know this is a sport manager, then
I [should] know that certain competencies
and certain skills have been taught during
their studies and that, so to speak, a min-
imum of quality standards are linked to
them. And, let me give you another exam-
ple: if someone has a doctoral degree, then
it is relatively clear what steps they have Professional competencies concern-
ing sport-management-specific knowl-
edgeweregivenverylittleemphasisinthe
interviews. Specialized knowledge about
sport systems and structures appeared
to be helpful in non-profit and public
sector organizations. Informants from
all sectors regarded generic information
technology (IT) skills as vital. More spe- 349 German Journal of Exercise and Sport Research 3 · 2021 Main Article gone through to get there. And I don’t see
that with the sport manager, and I actu-
ally have to consider and worry that dif-
ferences in quality are not comprehensible
to outside parties. (NP-3) relationship in which a specific sport
management programme allowed em-
ployees to further their education. While
most representatives spoke about part-
nerships with HEIs in general, few of
these partnerships involved sport man-
agement programmes specifically. One
representative highlighted dual-career
opportunities as a form of partnership
with a private HEI offering sport man-
agement (NP-2). Many studies within
and outside of sport settings have em-
phasized the advantages of contractual
cooperation between HEIs and industry
(Hardin, Bemiller, &Pate, 2013; Petersen
& Pierce, 2009). HEIs could contribute
tofulfilmentofthemetaphoricalcontract
by consenting to needs-based qualifi-
cation of sport management students
and being receptive to cooperation with
sport management organizations (i.e. involving sport organizations in cur-
riculum development, hosting career
fairs or creating professorial positions
that combine practice and academia). tory internships as an optimal solution
for both sport management HEIs, as this
increasestheirapplicability, andsportor-
ganizations, as they profit from extra hu-
man resources. One representative ex-
plains: These findings substantiate earlier stud-
ies, which found a lack of clear sig-
nals for sport organizations regarding
sport management programmes (Chalip,
2006; Dowling, Edwards, & Washing-
ton, 2014). Due to this information
asymmetry, the principal is not able to
clearly interpret compliance of SMHE
in fulfilling their assignment within the
contract. This could in parts be due to
the heterogeneity of sport management
programmes in Germany, rendering the
assessment of individual agents nearly
impossible (Eisenhardt, 1989). The CEO of a national sport federation
agreed: Thus,
sport management HEIs need to signal
their unique value (i.e. developing grad-
uates with strong generic competencies)
via reputation assurances, such as the
publication of study documents (e.g. module descriptors), ranking orders and
successful graduate testimonials in order
to reduce information asymmetry and
demonstrate their compliance to the re-
quirements of the metaphorical contract
(Spence, 1973). Interns [...] make an important contri-
bution in many areas. The basic problem
is that the studies are too school-like and
there are no more possibilities to get in-
terns for six months. And we actually only
want to have interns [for six months] be-
cause everything else really doesn’t make
sense. [...] I believe six-month intern-
ships should be integrated and enabled in
all study programmes. (C-3) Study programmes should take this
into account when designing curricula
and make them more flexible to allow
for internships (DeLuca & Braunstein-
Minkove, 2016; Dunkel, Wohlfahrt, &
Wendeborn, 2018). Open and trans-
parent
communication
between
the
principal and agent could help both
sides of the dyad to reduce asymmetry
in information acquisition and, in doing
so, better understand the importance of
practical opportunities. While German
law on minimum wages regulates volun-
tary internship arrangements intended
for students to gain work experience,
we maintain that the principal is just
as responsible for creating appropri-
ate conditions (i.e. compensation) for
internships (mandatory or voluntary). We conclude from the interviews
that the key informants have quite spe-
cific expectations concerning the role of
SMHEin the principal–agent dyad. They
clearlyendorse the idea ofa metaphorical
contract and to that effect the expecta-
tions of qualifying graduates according
to their needs. Notwithstanding, the
key informants also acknowledge the
slow response of HEIs to labour market
demands: Presupposed role of sport
management HEIs Surprisingly, all interviewees expressed
specific expectations for sport manage-
ment HEIs. They are regarded as respon-
sible for developing the aforementioned
generic competencies as well as holisti-
callypreparingstudentsforthe(sport)in-
dustry. As one representative from a pro-
fessional football club explained, ‘I be-
lieve that a university today not only has
the task to prepare [students] academi-
cally for the profession, but to prepare
[them]fortheprofessioninanall-around
way’ (C-2). Voluntaryengagementinsportscould
be advocated within sport management
programmes to improve the practical ap-
plicability of sport management content
(Wicker & Breuer, 2011). As Wallrodt
and Thieme (2020) have shown, sports
volunteering has a positive effect on ap-
plicants’ perceived qualifications and op-
erates as a positive signal of social skills. Integrating such experiences into study
programmes has great potential to raise
awareness of the practical applicability of
content and, thereby, distinguish gradu-
ates. Especially in the areas that are devel-
oping dynamically in sports manage-
ment, universities andespecially the public
ones [...] are very much lagging behind. Because they are simply less able to adapt
quickly to corresponding developments
and to incorporate trends [...] into their
programs. (PU-1) We assert that the metaphorical con-
tract lacks clear information about how
HEIsshouldprofessionallytrainstudents
for employability. HEIs primarily aim
to create and disseminate knowledge. However, sport organizations demand
that sport management HEIs support
students in their personal development
and active citizenship in order to remain
competitive and fulfil supranational and
national economic policy goals (Euro- Furthermore, the interviewees high-
lighted the need for practical applicabil-
ity of learning content, as this leads to
sustainable employability. However, the
interviewees were unable to specifically
describe how practical content can be in-
creasedinhighereducationprogrammes. Most representatives proposed obliga- Finally, possible cooperation between
sport organizations and specific sport
management programmes emerged as
a central theme in the interviews. The
CEO of a large non-profit organiza-
tion (NP-3) spoke about a cooperative 350 German Journal of Exercise and Sport Research 3 · 2021 sport management programmes in the
future. develop higher levels of competence
across all competence blocks compared
with a bachelor’s degree programme
(European Commission, 2008, 2019;
Kultusministerkonferenz, 2017). Finally,
SMHE needs to define the institutional
architecture needed to offer degrees and
determine whether it is more efficient to
engage all types of HEIs in this task or
whether specific HEIs are better suited. pean Commission, 2019; Vossensteyn
et al., 2018). The interests of sport organizations
and sport management HEIs are contra-
dictoryinparts(Akerlof, 1970). Conclusion and future research In our study, we acquired and reported
crucial information for SMHE pro-
gramme design and delivery, focusing
on asymmetry in information acqui-
sition. We omitted pre- and postcon-
tractual information asymmetry in our
dyad that could lead to moral hazard as
well as issues regarding divergence of
interests (Kivistö, 2008; Liefner, 2003). These agency problems present consid-
erable issues that need to be examined
in additional studies. This study contributes to labour market
research on sport management by in-
troducing agency theory as a theoretical
framework. Analysis of 12 semi-struc-
tured interviews with sport organization
representatives revealed no specific re-
quirements regarding the qualifications
of sport managers. We assert that SMHE
needs to credibly signal its unique value
compared to other study programmes
in order to better comply with the
metaphorical contract. Our findings
reinforce the relevance of generic com-
petencies and the possibility of making
the content of sport management study
programmes applicable to the labour
market. Sport management HEIs should
therefore, in turn, concentrate on de-
veloping generic competencies within
study programmes. In clarifying the role of sport man-
agement programmes in fulfilling the
metaphorical contract (Research Aim 2),
our findings offer two major implications
for sport management HEIs. First, sport
management study programmes need to
develop students’ generic competencies
through, for example, group work or
volunteering services. Second, SMHE
needs to establish a clear profile for sport
management programmes. There is cur-
rently no need for sport-management-
specificqualifications, possiblyduetothe
ambiguity surrounding whether sport
management is an academic discipline or
profession (Chalip, 2006; Dowling, 2018;
Dowling et al., 2014). Concise differenti-
ation between the abundance of degrees
seems to be a suitable starting point
in signalling relevance and uniqueness. While the sport management bachelor’s
degree was developed to be an inde-
pendent professional degree (European
Commission, 2019), the master’s degree
was developed to enable graduates to
take on leading positions in the sport
business (e.g. manager, team leader). As the two programmes have different
goals concerning the employability of
graduates, they should develop different
generic and subject-specific competen-
cies. According to educational policy
goals, a master’s degree programme must We used agency theory, as it has par-
ticular strengths for analysing the prin-
cipal–agent dyad of sport organizations
and HEIs. Presupposed role of sport
management HEIs Thistyp-
ical agency problem is exacerbated by the
heterogeneity of the institutions and ex-
isting outcome-uncertainty (Eisenhardt,
1989). Legitimized by the Bologna Re-
form, we recommend SMHE to both ac-
knowledge and fulfil the metaphorical
contract and take a seat at the table, will-
ing to learn more about the requirements
of the principal. This would in the long
run potentially help SMHE in achieving
their goals. Due to their institutional
goals and structures, private HEIs might
be betterprepared forfulfilling the role of
professional trainers, and public univer-
sities’ self-image as educators might be
a considerable hindrance to fulfilment of
their role according to the metaphorical
contract. Our study focused on one specific
dyad in a complex institutional field. Principal–agent relationships, such as
those between students and HEIs as
well as between academic staffand HEIs
were not considered. Future studies can
build on the findings from our study
and examine further principal–agent
relationships in the complex network of
SMHE. Limitations Conclusion and future research Conclusion and future research However, like other theories,
agency theory is subject to criticism, par-
ticularly for its focus on the economic
aspects of contractual relationships and
related assumptions about human moti-
vation and behaviour, which are relevant
to our study. In addition, the theory
does not question whether the princi-
pal’s goals are legitimate (Kivistö, 2008). This is particularly interesting in the con-
text of the controversial debate of the
last two decades concerning the role of
HEIs. Utilitarian arguments (from the
perspective of the sport industry) con-
tend thattheyneed toproduce a qualified
workforce for the labour market, while
cultural arguments (from the perspective
of higher education) advocate for insti-
tutional autonomy and academic free-
dom. Considering the heterogeneity of
sport organizations and the quick pace of
change in this market due to the global
trends of digitalization, commercializa-
tion and internationalization, these de-
mandswillcontinuetochange. However,
it remains uncertain whether society will
accept the demands of sport organiza-
tions as valid and require adaptation of Experimental studies examining the
perceived and measured differences be-
tween graduates of generic management
and sport management programmes
could validate the value of the latter. Standardised and long-term graduate
surveys and tracking studies could offer
vital information regarding the success
of sport management programmes. Col-
loquia with current students, as key
informants, during or after internships
could make tacit information explicit
and, as such, contribute to research in
the field. In addition, we recommend
document analysis of sport management
course content and qualitative inves-
tigation of the actual content that is
taught. HEIs are faced with several chal-
lenges concerning their role within the
metaphorical contract. Following the 351 German Journal of Exercise and Sport Research 3 · 2021 Main Article Journal of Sport Management, 4(1), 31–58. https://doi.org/10.1123/jsm.4.1.31. productioninanymediumorformat,aslongasyou
giveappropriatecredittotheoriginalauthor(s)and
thesource,providealinktotheCreativeCommonsli-
cence,andindicateifchangesweremade. Theimages
orotherthirdpartymaterialinthisarticleareincluded
inthearticle’sCreativeCommonslicence,unlessin-
dicatedotherwiseinacreditlinetothematerial. If
materialisnotincludedinthearticle’sCreativeCom-
monslicenceandyourintendeduseisnotpermitted
bystatutoryregulationorexceedsthepermitteduse,
you willneedtoobtainpermissiondirectlyfromthe
copyrightholder. Toviewacopyofthislicence,visit
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Predicting Safe Parking Spaces: A Machine Learning Approach to Geospatial Urban and Crime Data
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Received: 11 April 2019; Accepted: 13 May 2019; Published: 19 May 2019 Abstract: This research aims to identify spatial and time patterns of theft in Manhattan, NY, to
reveal urban factors that contribute to thefts from motor vehicles and to build a prediction model
for thefts. Methods include time series and hot spot analysis, linear regression, elastic-net, Support
vector machines SVM with radial and linear kernels, decision tree, bagged CART, random forest, and
stochastic gradient boosting. Machine learning methods reveal that linear models perform better
on our data (linear regression, elastic-net), specifying that a higher number of subway entrances,
graffiti, and restaurants on streets contribute to higher theft rates from motor vehicles. Although the
prediction model for thefts meets almost all assumptions (five of six), its accuracy is 77%, suggesting
that there are other undiscovered factors making a contribution to the generation of thefts. As an
output demonstrating final results, the application prototype for searching safer parking in Manhattan,
NY based on the prediction model, has been developed. Keywords: geospatial data; machine learning; Manhattan; prediction model; theft from motor vehicle;
crime prevention through urban planning Sustainability 2019, 11, 2848; doi:10.3390/su11102848 www.mdpi.com/journal/sustainability Predicting Safe Parking Spaces: A Machine Learning
Approach to Geospatial Urban and Crime Data Irina Matijosaitiene 1,2,*, Anthony McDowald 1 and Vishal Juneja 3 1
Data Science Institute, Saint Peter’s University, Jersey City, NJ 07306, USA; amcdowald@saintpeters.edu
2
Centre for Smart Cities and Infrastructure, Kaunas University of Technology, Kaunas 44249, Lithuania
3
Amazon Robotics, Boston, MA 01864, USA; vj2208@columbia.edu
*
Correspondence: imatijosaitiene@saintpeters.edu; Tel.: +1-646-954-2690 1
Data Science Institute, Saint Peter’s University, Jersey City, NJ 07306, USA; amcdowald@saintpeters.edu
2
Centre for Smart Cities and Infrastructure, Kaunas University of Technology, Kaunas 44249, Lithuania
3
Amazon Robotics, Boston, MA 01864, USA; vj2208@columbia.edu
*
Correspondence: imatijosaitiene@saintpeters.edu; Tel.: +1-646-954-2690 sustainability sustainability sustainability sustainability 1. Introduction Safety and security are both considered components of sustainability, as are the quality of air,
land, and water, well-being, economy, education, and health [1]. According to multiple theories and
studies [2–6], minimizing crime improves an area’s sustainability, and safer neighborhoods result in
better social and population health outcomes. Therefore, sustainable development can be achieved
by improving safety and security, and crime can be reduced by the correct implementation of urban
planning and design. The study by Stankevice et al. [7] revealed that when green areas and specialized
areas are incorporated into the residential areas, we observe less crime. Also, public areas combined
with residential and green areas lead to crime reduction. Mixed use areas and commercial areas
create good opportunities for the pickpocketing. Newman [8] proves that “spaces with low urban
development density and single use with a strictly limited access to strangers are less vulnerable to
crime,” whereas Jacobs’ [9] states that “urban spaces with mixed land use and open access to strangers
lead to less crime because they provide more ‘eyes on the street’ and more natural surveillance.” In
the research of Hillier [10] and Monteiro [11], some types of anti-ocial behavior are strongly related
to some urban functional zones. Moreover, Hiller [10] states that low-activity and low-movement
areas are related to crime. In the research performed by Sypion-Dutkowska and Leitner [12], it was
proven that activities such as alcohol outlets, clubs and discos, cultural facilities, municipal housing,
and commercial buildings are strongly related to the higher crime rates, especially in their immediate
surroundings (within 50 m from the activity). On the other hand, they [12] unveil activities that detract
crime, such as grandstands, cemeteries, green areas, allotment gardens, depots, and transport bases. www.mdpi.com/journal/sustainability 2 of 15 Sustainability 2019, 11, 2848 In environmental security, Crime Prevention Through Environmental Design (CPTED) is one
of the most powerful tools to achieve safety goals. CPTED explains the ideas for crime prevention
through urban planning and design [2,3,13–18]. The well-known principles of CPTED that we rely
on in this research include natural surveillance (id est. ‘eyes on the street’), natural access control (id
est. making paths and other spaces visible and easy for communication), and activity support (id
est. urban spaces have to be frequently used by various activities). 1. Introduction These CPTED principles can be
implemented through the proper urban planning and design, regulating land uses and activities on
sites, landscaping, lighting, signage, etc. [3]. The recent examples of successful application of CPTED
principles to safer cities include the COPS project in London, UK [19], demolition and rebuilding of
Bijlmermeer residential area in Netherlands [20], Hammarby Sjöstad in Stockholm, Sweden, Gellerup
and Bispehaven in Aarhus, Denmark, Tingbjerg and Husum North in Copenhagen, Denmark, La
Duchère urban project in Lyon, France, as well as projects in the Arab Emirates [21], Korea [22], and
Botswana [23]. In this paper, we search for patterns in crime time and locations by analyzing geospatial
data, as well as we attempt to predict places of safe and unsafe parking places in Manhattan, NY, and
urban factors as generators of crime through the application of machine learning methods. g
g
pp
g
A review of recent cases of the application of machine learning methods to crime analysis and
predictions demonstrates that despite some interesting examples, machine learning, as well as data
mining and other computational methods, are still not widely used for crime analytics using geospatial
data. Recently, a machine learning algorithm was applied to support the analysis and prediction of
crime patterns in Brazilian cities. During the past few years, Brazilian citizens provided Web-based
systems with a large amount of data. Therefore, the designed machine learning algorithm automatically
acquires data from collaborative sources, generates logical rules and visualizes the found patterns [24]. Boldt and Borg [25] proposed a novel method to approximate offense times for residential burglaries
using a dataset of all Swedish residential burglaries committed from 2010 to 2014 (a total of 103,029). Having access to burglary data over five years (2011—2015), Mburu and Bakillah [26] developed a
series of regression models to identify local indicators of the urban environment (e.g. unemployment,
building density, and type) that increase the risk of burglaries. For the investigation of impact of
vegetation density and transport network on crime, Du and Law [27] employed geographically
weighted regression modeling to unveil the associations across an urban central-peripheral gradient for
the following crime types: Assaults, vehicle theft, sex offences, and drugs. Marco et al. [28] performed
Bayesian analysis with a spatial beta regression model to analyze the association between the spatial
distribution of drug-related police interventions and the neighborhood characteristics. 1. Introduction Their results
indicated the following factors related to the high levels of drug-related police interventions: physical
decay, low socioeconomic status, and high immigrant concentration. 2. Materials and Methods The research subject and area for this research are dependent on the client needs: one of the
biggest insurance companies in the USA was searching for the state-of-the-art data science ideas that
could help them to tailor the insurance price for the car owners based on their parking behavior. Our
idea lays in applying state-of-the-art machine learning methods to predict the rate of theft from motor
vehicle (as one of the components influencing the calculation of price of the car insurance) for each
street segment and to identify urban factors that cause higher rate of theft. Manhattan, NY, which
contains mostly residential, commercial, and manufacturing districts, was selected as an experimental
area for this research. Thefts from motor vehicles in Manhattan are the main subject of this research. The dataset about crimes (including thefts from motor vehicles) comes from the NYC Open Data
“NYPD Complaint Map” (csv file), which contains 25 variables for 478,805 samples [29]. Under NY
Penal Law, the theft from motor vehicle falls under the category of larceny, both grand and petit. However, the yearly percentage of thefts from motor vehicles in Manhattan is not more than 6%
compared to other crimes, with 99.5% of all thefts from motor vehicles occurring in the street. Parking
lots (both public and private) result in 0.496% of all cases, and the rest of urban spaces make up only 3 of 15 Sustainability 2019, 11, 2848 0.004% of all cases. The dataset we use in this research contains the following attributes for each crime:
Latitude and longitude, crime type, date and time when a crime was committed, as well as date and
time when a crime was registered by the police, circumstances of the crime, address, description of
premises, and other variables. The granularity of crime classification is very fine. For instance, theft
from motor vehicle is categorized as grand larceny from vehicle, petty larceny from vehicle, theft of
vehicle accessory, etc. As this fine granularity was not necessary for our research goals, we do not focus
on small classes of crime (for instance, theft of vehicle accessory or similar), and we have aggregated
crime types into bigger groups, such as theft from motor vehicle, burglary, etc., using only crime data
for the years 2015, 2016, and 2017. 2. Materials and Methods The crime data for the recent 2—3 years is considered enough
data for making crime analytics and predictions/forecasting, especially when the crime is analyzed in
terms of urban planning. We also aggregated the new variable “day of week” using R programming
language. Next, we performed data reduction to leave in our dataset only the variables of our interest:
Unique ID, crime type, date and time, day of week, and latitude/longitude. Time series analysis is
applied to analyze the trend of thefts to occur within a day/night time. We used ArcGIS with Python
for the hot spot analysis (Getis-Ord G∗
i) to identify geospatial trends of thefts. For this purpose, the
G∗
i statistic (that is a z-score) is computed for each analyzed feature (independent variable) in the
dataset, using the formulas in Eq. (1) integrated in ArcGIS software. It reveals spatial clusters of high
(statistically significant positive large z-scores) or low (statistically significant negative small z-scores)
values by considering each feature in terms of the neighboring features [30–33]. For instance, only
features with a high value and surrounded by other features with high values will be considered as a
statistically significant hot spot. G∗
i =
Pn
j=1 wi,jxj −X Pn
j=1 wi,j
S
r
n Pn
j=1 w2
i,j−(Pn
j=1 wi,j)2
n−1
X =
Pn
j=1 xj
n
S =
sPn
j=1 x2
j
n
−
X
2A = πr2
(1) (1) where xj is the attribute value for feature j, wi,j is the spatial weight between features i and j, n is the
total number of features. G∗
i statistic is a z-score [30–33]. To proceed with the selection of independent variables (and datasets accordingly) for the geospatial
data analysis, we faced some limitations: (i) Data must be geospatial, or it must at least have coordinates,
longitudes-latitudes, or exact addresses; (ii) data must represent urban features (not social or economic),
such as visibility, location, and operation hours of commercial and public buildings, lighting, graffiti,
vegetation, etc.; (iii) data must be open. These limitations allowed us to select the following datasets
from NYC.gov [34], NYC DOT [35], Open Data NYC [29], and GIS Community [36]: •
Street segment centerlines—As a basis and connective grid for all subsequent datasets (shp). This
dataset contains information about street segment direction, street name, length and width of the
street segment, as well as all generic street and non-street features, along with roadbed. 2. Materials and Methods In the
scope of this research, we use only the data about street segment endpoint coordinates (in order to
map the street segment centerline), as well as street segment length and width. •
Subway entrances as points with their geolocations (shp), having five variables for 1929
samples—Unique ID, URL, name (street or intersection of streets), point geometry (latitude
and longitude), and subway line (A, B, C, D, etc.). This dataset comes from the year 2017. In terms
of this research, we keep for the further analysis only two variables—ID and point geometry. •
Restaurants as points with their geolocations (shp), having 13 variables for 9326 samples—Unique
ID, name, cuisine (Afghan, Chinese, French, etc.), phone, address, point geometry (latitude and
longitude), etc. This dataset comes from the year 2017. We keep for the further analysis only two
variables—ID and point geometry. •
Graffiti as points with their geolocations (shp), having 13 variables for 2226 samples—Unique ID,
address, community, borough, status, date, X and Y coordinates, etc. Only three variables—ID, X, •
Graffiti as points with their geolocations (shp), having 13 variables for 2226 samples—Unique ID,
address, community, borough, status, date, X and Y coordinates, etc. Only three variables—ID, X, 4 of 15 Sustainability 2019, 11, 2848 and Y coordinates—were selected for merging the dataset and proceeding with the research. Also,
we limit the number of samples only to those registered in the years 2015, 2016, and 2017. y
g
y
•
Street pavement rating as lines with the rating being presented as ‘good,’ ‘fair,’ or ‘poor’ (shp),
having 10 variables for 81,210 samples—Unique ID of each segment, line geometry (latitude and
longitude for both start and end points of each street segment), segment length, segment width,
rating word (Good, Fair, Poor, Not Rated (NR)), and rating code as an ordinal variable (0, 1, 2,
3, etc.), rating date and time, etc. This dataset comes from the year 2017. To merge this dataset
with others and to perform the research, we keep only three variables—ID, line geometry, and the
pavement rating code. Street segment was defined as the joining of elements for all six datasets, and further data processing
and research were performed on the microscale, i.e., on the scale of street segment. 2. Materials and Methods According to
the Space Syntax theory, “segment is the shortest path that uses the least number of streets (actually,
the least number of street sections between the crossings) to get to your destination” [37,38]. The
six datasets are to be joined by location method, since they do not have a common key value. Join
by location, or spatial join, uses spatial associations between the layers to append fields from one
layer to another. Therefore, ArcGIS in combination with Python is used for joining six datasets by
following this workflow: (i) All the necessary files (csv and shp) are imported into ArcMap; (ii) for
the thefts dataset that comes as a csv file, the data is geolocated in XY coordinates—id est., the layer
file is created (though, now the geolocated data does not have the unique ID (key) that is required
to proceed with the data processing); (iii) the layer file is converted into shp file and imported into
ArcMap; (iv) each of five shp file datasets, (thefts, restaurants, subway entrances, graffiti, pavement
rating) is joined one-by-one to the street segment centerlines dataset based on spatial location of the
added dataset (layer). Now, each street segment is assigned properties of the joined layer, where each
line (a street segment) is given attributes of the line (pavement rating) and points (thefts, restaurants,
subway entrances, graffiti) that are closest to the street segment centerline. Moreover, for each street
segment, the number of occurrences for each of the following variables was added: Thefts, restaurants,
subway entrances, and graffiti. For instance, for each street segment, the number of thefts that occurred
on that street segment was counted and added as a new generated value. In the same way, new values
were generated while counting the number of restaurants, subway entrances, and graffiti located on
each street segment. Using R programming language, we performed data reduction, leaving only the
variables of our interest, i.e., urban features that might be responsible for the generation of thefts from
motor vehicles. Therefore, the final dataset with 17,060 samples is a geospatial data, where each street
segment carries information about its pavement rating, as well as the number of thefts, restaurants,
subway entrances, and graffiti, as shown in Table 1. Table 1. Final geospatial dataset (extract). •
Multiple linear regression (lm), 2. Materials and Methods Therefore, the goal is find the balance between small
and large C and gamma values, •
Support vector machines (SVM) with the radial kernel with tuned parameters gamma γ, that
is, the Gaussian Kernel hyperparameter (to handle nonlinear classification), which controls the
shape of the peaks of the Radial Basis Function (RBF) kernel, and cost C, that, is the soft margin
hyperparameter, which controls the influence of each individual support vector and how much
we penalize variables. If gamma is large, it leads to high bias and low variance models. High
gamma makes our decision boundary depend on points close to the decision boundary and nearer
points carry more weights than faraway points due to which our decision boundary becomes
wigglier, and small gamma makes faraway points carry more weights than nearer points. Thus,
our decision boundary becomes more like a straight line. The cost of misclassification is low with
small C values (id est. a soft margin), whereas the cost of misclassification high with the large C
values (id est. hard margin). With the large C values, the algorithm tries to explain the input data
stricter, and it usually leads to overfitting. Therefore, the goal is find the balance between small
and large C and gamma values, •
Support vector machines (SVM) with the linear kernel with a tuned parameter C, which defines
how much we want to avoid misclassification, where small C corresponds to small margin between
support vectors, and high C gives bigger margin, •
Decision tree (CART) with a tuned complexity parameter cp that corresponds to the size of the
decision tree. When the cp value is reached for the particular node in the tree, then tree building
stops growing, •
Bagged CART (different from CART), a bootstrapped aggregation is an ensemble method that fits
many trees to bootstrap-resampled versions of the training data to build independent prediction
models, and then combines them using an averaging technique (for instance, a majority vote). 2. Materials and Methods To build a good prediction model we have tried nine machine learning algorithms
on the training set and tested the model performance using the test set data: 5 of 15 Sustainability 2019, 11, 2848 •
Elastic-net (enet) with a tuned hyper-parameter lambda λ, where penalties α for both lasso (here,
L-1 norm is used to shrink the regression coefficients toward zero by penalizing the regression
model, which is the sum of the absolute coefficients) and ridge (here, L-2 norm is used to shrink
the regression coefficients, with minor contribution to the outcome, to be close to zero, which is
the sum of the squared coefficients) regressions are balanced [39], •
Elastic-net (enet) with a tuned hyper-parameter lambda λ, where penalties α for both lasso (here,
L-1 norm is used to shrink the regression coefficients toward zero by penalizing the regression
model, which is the sum of the absolute coefficients) and ridge (here, L-2 norm is used to shrink
the regression coefficients, with minor contribution to the outcome, to be close to zero, which is
the sum of the squared coefficients) regressions are balanced [39], g
•
Support vector machines (SVM) with the radial kernel with tuned parameters gamma γ, that
is, the Gaussian Kernel hyperparameter (to handle nonlinear classification), which controls the
shape of the peaks of the Radial Basis Function (RBF) kernel, and cost C, that, is the soft margin
hyperparameter, which controls the influence of each individual support vector and how much
we penalize variables. If gamma is large, it leads to high bias and low variance models. High
gamma makes our decision boundary depend on points close to the decision boundary and nearer
points carry more weights than faraway points due to which our decision boundary becomes
wigglier, and small gamma makes faraway points carry more weights than nearer points. Thus,
our decision boundary becomes more like a straight line. The cost of misclassification is low with
small C values (id est. a soft margin), whereas the cost of misclassification high with the large C
values (id est. hard margin). With the large C values, the algorithm tries to explain the input data
stricter, and it usually leads to overfitting. 2. Materials and Methods ID
Latitude,
Longitude
Segment
Length
Segment
Width
Thefts
Restaurants
Subway
Entrances
Graffiti
Pavement
Rating
33
−74.25287637435896
40.50762599010005,
−74.25299942180904
40.50688186992653
229
33.58
1
2
0
0
6
35
−74.25308030444408
40.51209961294924,
−74.25299384780556
40.51206137595131
66
30
0
0
0
0
0
All data in the final dataset was divided randomly into the training set (80% of data) and the test
set (20% of data). To build a good prediction model we have tried nine machine learning algorithms
on the training set and tested the model performance using the test set data:
•
Multiple linear regression (lm), Table 1. Final geospatial dataset (extract). ID
Latitude,
Longitude
Segment
Length
Segment
Width
Thefts
Restaurants
Subway
Entrances
Graffiti
Pavement
Rating
33
−74.25287637435896
40.50762599010005,
−74.25299942180904
40.50688186992653
229
33.58
1
2
0
0
6
35
−74.25308030444408
40.51209961294924,
−74.25299384780556
40.51206137595131
66
30
0
0
0
0
0
All data in the final dataset was divided randomly into the training set (80% of data) and the test
set (20% of data). To build a good prediction model we have tried nine machine learning algorithms
on the training set and tested the model performance using the test set data: Table 1. Final geospatial dataset (extract). Table 1. Final geospatial dataset (extract). All data in the final dataset was divided randomly into the training set (80% of data) and the test
set (20% of data). To build a good prediction model we have tried nine machine learning algorithms
on the training set and tested the model performance using the test set data: All data in the final dataset was divided randomly into the training set (80% of data) and the test
set (20% of data). 2. Materials and Methods Because this technique takes many uncorrelated trees to make a final model, it reduces error by
reducing variance [40], •
Random forest (rf) with tuned number of trees in the forest and mtry parameter as a number
of variables randomly taken as candidates from the initial list of all variables to make the split
for building the trees, where usually mtry = n/3 for regression, with n being a number of
independent variables, •
Stochastic gradient boosting (generalized boosted modeling (gbm)), different from bagged methods
and random forest, chooses all features to make a split, and predictors are made sequentially
(not independently, like in random forest and other bagged algorithms), where the subsequent
predictors learn from the errors (residuals) of the previous predictors. It aims to optimize the loss
function (that can be any function, for instance, regression can use a squared error), then uses a
weak learner (such as decision trees) to make a prediction, and finally, it makes an additive model,
where one tree as a weak learner is added at a time without changing existing trees. Here, error
is calculated and gradient decent is used to minimize the error by adding a tree to the model
that minimizes the error [41]. Here, we tune the following parameters: The number of trees, the
maximum depth of variable interactions (interaction.depth), the shrinkage parameter applied
to each tree in the expansion, and the minimum number of observations in trees terminal nodes
(n.minobsinnode). Some algorithms require data preprocessing, such as feature scaling and centering the data points. Therefore, the data preprocessing is performed within the linear regression, elastic-net, and both SVMs. We applied repeated cross-validation of our dataset to select the best prediction model with 10-fold
cross validation and three repeats, which gives us a more robust estimate of the models. RMSE (root 6 of 15 Sustainability 2019, 11, 2848 mean squared error) and R2 (R squared) are used to compare the models. RMSE gives us information
about the prediction model errors (the less the metric the better), and R2 tells us about the amount
of variability of a target (dependent variable) explained by the features (independent variables) (the
closer to 1 the better). Sustainability 2019 11 x FOR PEER REVIEW
6 of 15 To build a good regression model, we checked data for missing values, multicollinearity, outliers,
and normality. 2. Materials and Methods Missing values: After finding out the character of missing values, we excluded them
from analysis. Multicollinearity: We wanted to avoid any two independent variables that are highly
correlated (r > 0.9) because they create a multicollinearity problem in the regression model. Therefore,
we performed a correlation analysis using the Pearson correlation coefficient, which describes the
relation between variables in terms of linearity. However, after checking the variables, we did not
find any independent variables that are strongly correlated. Therefore, we have no multicollinearity
problem in our data. Additionally, checking the training set data for outliers did not result in any
outliers. However, a normality check showed that data of four variables (thefts, restaurants, subway
entrances, and graffiti) is strongly right-skewed (Figure 1A) because many street segments have zero
values of thefts, restaurants, subway entrances, and graffiti. Therefore, we performed a logarithmic
transformation of data within these four variables, previously adding one to all values to avoid log
0 cases. Logarithmic transformation of data significantly improved data distribution, and therefore,
the transformed data in the training set was used to perform a correlation analysis to decide what
variables to include in the regression analysis. Finally, we checked six assumptions for correctness of
the prediction model and tested the prediction model on the test set data. Sustainability 2019, 11, x FOR PEER REVIEW
6 of 15
excluded them from analysis. Multicollinearity: We wanted to avoid any two independent variables
that are highly correlated (r > 0.9) because they create a multicollinearity problem in the regression
model. Therefore, we performed a correlation analysis using the Pearson correlation coefficient,
which describes the relation between variables in terms of linearity. However, after checking the
variables, we did not find any independent variables that are strongly correlated. Therefore, we have
no multicollinearity problem in our data. Additionally, checking the training set data for outliers did
not result in any outliers. However, a normality check showed that data of four variables (thefts,
restaurants, subway entrances, and graffiti) is strongly right-skewed (Figure 1A) because many street
segments have zero values of thefts, restaurants, subway entrances, and graffiti. Therefore, we
performed a logarithmic transformation of data within these four variables, previously adding one
to all values to avoid log 0 cases. 2. Materials and Methods Logarithmic transformation of data significantly improved data
distribution, and therefore, the transformed data in the training set was used to perform a correlation
analysis to decide what variables to include in the regression analysis. Finally, we checked six
assumptions for correctness of the prediction model and tested the prediction model on the test set
data. Figure 1. (A). Histogram of restaurants. The data is strongly right-skewed, with the most of values
equal to zeros. (B). When do thefts happen? The graph shows a slight, although not very significant,
increase in theft rates on Fridays, Saturdays, and Sundays. (C). Where do thefts happen? Almost all
thefts from motor vehicles in Manhattan happen on the street. (D). Where and when thefts happen:
Identifying patterns. The temporal distribution (in terms of crime per hour) and the spatial
distribution of thefts from motor vehicles reveal a common pattern in theft location and time on
Tuesdays, Wednesday, and Thursdays. However, we do not observe any pattern for other weekdays. Figure 1. (A). Histogram of restaurants. The data is strongly right-skewed, with the most of values equal
to zeros. (B). When do thefts happen? The graph shows a slight, although not very significant, increase
in theft rates on Fridays, Saturdays, and Sundays. (C). Where do thefts happen? Almost all thefts from
motor vehicles in Manhattan happen on the street. (D). Where and when thefts happen: Identifying
patterns. The temporal distribution (in terms of crime per hour) and the spatial distribution of thefts
from motor vehicles reveal a common pattern in theft location and time on Tuesdays, Wednesday, and
Thursdays. However, we do not observe any pattern for other weekdays. Figure 1. (A). Histogram of restaurants. The data is strongly right-skewed, with the most of values
equal to zeros. (B). When do thefts happen? The graph shows a slight, although not very significant,
increase in theft rates on Fridays, Saturdays, and Sundays. (C). Where do thefts happen? Almost all
thefts from motor vehicles in Manhattan happen on the street. (D). Where and when thefts happen:
Identifying patterns. The temporal distribution (in terms of crime per hour) and the spatial
distribution of thefts from motor vehicles reveal a common pattern in theft location and time on
Tuesdays, Wednesday, and Thursdays. However, we do not observe any pattern for other weekdays. Figure 1. (A). Histogram of restaurants. 3.1. Temporl and Spatial Distribution of Data
demonstrate any common trends. To check
theft depend on human activities during pa Exploration of day of week and theft variables does not demonstrate any significant trend
(Figure 1B). However, the initial theft data analysis with Tableau revealed that almost all thefts from
motor vehicles happen on streets (Figure 1C). Therefore, to investigate further, we used only thefts
that occurred on streets. Time series analysis of thefts for each day within a week and the geospatial
analysis of theft unveils a common pattern in both time series and theft mapping results for Tuesdays,
Wednesdays, and Thursdays data (Figure 1D) with theft rates increasing from 4 p.m. to 7—8 p.m.,
and the lowest theft rates being at 2—6 a.m. However, we did not observe any pattern for any of the
remaining week days, i.e., neither time series plots nor theft maps for different time ranges demonstrate
any common trends. To check if criminal behaviors or opportunities for committing a theft depend
on human activities during particular hours, we divided a week into time ranges based on human
activities (weekday work hours, weekday commute hours, weekday nights, weekend nights, etc.). For each time range, we generated a map with theft rate for every street segment (Figure 2A). Visual
comparative analysis of maps reveals that thefts from motor vehicles “move” within Manhattan
depending on time range, and the intensity of theft is different depending on human activities. p
g p
g
on human activities (weekday work hours, weekday commute hours, weekday nights, weekend
nights, etc.). For each time range, we generated a map with theft rate for every street segment (Figure
2A). Visual comparative analysis of maps reveals that thefts from motor vehicles “move” within
Manhattan depending on time range, and the intensity of theft is different depending on human
activities. To narrow our search from the most opportune days and time for committing a theft to
territories where a theft is most likely to occur, we performed a hot spot analysis (Figure 2B). Hot
spot analysis demonstrates the most potentially dangerous territories located in East Village and
some spots in Uptown Manhattan, i.e., Washington Heights and Inwood, whereas the safest areas
are Upper East Side and Financial District. The reason for the latter, however, appears to be
restrictions on parking a private vehicle on many streets. 3.1. Temporl and Spatial Distribution of Data
demonstrate any common trends. To check
theft depend on human activities during pa In order to pinpoint the crime locations
more accurately, we performed a correlation and regression analysis to find urban generators of
thefts and, based on these generators, to come closer to the specific street segments with high theft
prediction. Figure 2. (A). Theft rates for street segments within different time ranges. The maps show that thefts
from motor vehicles “move” within Manhattan depending on time, while the intensity of theft is
different depending on human activities. (B). Hot spot analysis of thefts from motor vehicles. Significant theft hot spots (with 90–95–99% confidence level) are found in the East Village,
Washington Heights, and Inwood neighborhoods, whereas significant cold spots (with 90–95–99%
confidence level) are located in the Upper East Side and Financial District of Manhattan, NY. Figure 2. (A). Theft rates for street segments within different time ranges. The maps show that thefts
from motor vehicles “move” within Manhattan depending on time, while the intensity of theft is
different depending on human activities. (B). Hot spot analysis of thefts from motor vehicles. Significant
theft hot spots (with 90–95–99% confidence level) are found in the East Village, Washington Heights,
and Inwood neighborhoods, whereas significant cold spots (with 90–95–99% confidence level) are
located in the Upper East Side and Financial District of Manhattan, NY. Figure 2. (A). Theft rates for street segments within different time ranges. The maps show that thefts
from motor vehicles “move” within Manhattan depending on time, while the intensity of theft is
different depending on human activities. (B). Hot spot analysis of thefts from motor vehicles. Significant theft hot spots (with 90–95–99% confidence level) are found in the East Village,
Washington Heights, and Inwood neighborhoods, whereas significant cold spots (with 90–95–99%
confidence level) are located in the Upper East Side and Financial District of Manhattan, NY. Figure 2. (A). Theft rates for street segments within different time ranges. The maps show that thefts
from motor vehicles “move” within Manhattan depending on time, while the intensity of theft is
different depending on human activities. (B). Hot spot analysis of thefts from motor vehicles. Significant
theft hot spots (with 90–95–99% confidence level) are found in the East Village, Washington Heights,
and Inwood neighborhoods, whereas significant cold spots (with 90–95–99% confidence level) are
located in the Upper East Side and Financial District of Manhattan, NY. 3.2. 2. Materials and Methods The data is strongly right-skewed, with the most of values equal
to zeros. (B). When do thefts happen? The graph shows a slight, although not very significant, increase
in theft rates on Fridays, Saturdays, and Sundays. (C). Where do thefts happen? Almost all thefts from
motor vehicles in Manhattan happen on the street. (D). Where and when thefts happen: Identifying
patterns. The temporal distribution (in terms of crime per hour) and the spatial distribution of thefts
from motor vehicles reveal a common pattern in theft location and time on Tuesdays, Wednesday, and
Thursdays. However, we do not observe any pattern for other weekdays. 7 of 15 7 of 15 Sustainability 2019, 11, 2848 3.2. Selection of the Prediction Model We trained eight different machine learning algorithms for the regression task on the training
dataset with 12,794 samples and 10-fold cross validation with three repeats, tuning the following
parameters: (1) For the linear regression, the intercept is included; (2) for the elastic-net, we use the
tuned best-fit parameters lambda λ = 0 that corresponds to zero amount of shrinkage and the mixing
parameter alpha α = 1 that performs the lasso fit; (3) for the SVM with a radial kernel, we use the
tuned best-fit parameters cost C = 1 and gamma γ = 2.908401; (4) for the SVM with a linear kernel, we
use the tuned best-fit parameter cost C = 1; (5) for the decision tree (CART), we tune the complexity
parameter cp = 0.01185767; (6) for the bagged CART, we do not tune any hyperparameter; (7) for the
random forest, we tune the number of trees to be 500 and mtry = 2; (8) for the stochastic gradient
boosting (gbm), we tune the number of trees equal to 150, the maximum depth of variable interactions
interaction.depth = 3, the shrinkage = 0.1, and the minimum number of observations in trees terminal
nodes n.minobsinnode = 10. The number of thefts from motor vehicles committed in a street segment is
the target variable (dependent variable). The number of subways, restaurants, graffiti, street pavement
rating, and street segment width and length are the features (independent variables) used for the
model selection by applying different machine learning methods. We computed RMSE and R2 for all
built models and compared them in order to select the best prediction model. From the comparison
results presented in the Table 2, it is obvious that linear models perform better on our data (linear
regression, elastic-net), as well as the random forest. For the best regression model, we want to have
the low error (close to zero) and the high variability of a target variable explained by the features (close
to one). Although the lowest value of RMSEmin and the highest value of R2max are observed while
using the random forest regression, we proceed with the mean values of RMSE and R2 obtained from
the 10-fold cross validation with three repeats procedure. Therefore, we consider linear regression
and elastic-net as candidate models having the lowest RMSEmean and the highest R2mean. 3.1. Temporl and Spatial Distribution of Data
demonstrate any common trends. To check
theft depend on human activities during pa The number of subways,
rating, and street segment width and length are the features (i
model selection by applying different machine learning methods
built models and compared them in order to select the best pred
results presented in the Table 2, it is obvious that linear models
regression, elastic-net), as well as the random forest. For the best
the low error (close to zero) and the high variability of a target var
to one). Although the lowest value of RMSEmin and the highes
using the random forest regression, we proceed with the mean va
the 10-fold cross validation with three repeats procedure. Ther
and elastic-net as candidate models having the lowest RMSEmean
elastic-net increases bias to reduce overfitting (and, accordingly,
of our models suffer from overfitting, we choose linear regressi
thefts from motor vehicles. With linear regression, the model p
same RMSE and R2. Table 2. Comparison of prediction models using root mean squa
determination (R2). Model
RMSEmin
RMSEmean
Linear regression
0.4407129
0.6049274
Elastic-net
0.4407129
0.6049274
SVMRadial
0.4548175
0.6387557
SVMLinear
0.4486161
0.6281775
Decision tree (CART)
0.4470269
0.6164779
BaggedCART
0.4393015
0.6114266
Random forest
0.4404932
0.6053636
Stochastic Gradient Boosting
0.4446763
0.6102181
3.3. Prediction Model Using Multiple Linear Regression
The Pearson correlation coefficient demonstrates a moderate p
and subway entrances (s_rho = 0.49, p-value < 2.2 × 10−16, a weak p
and restaurants, as well as thefts and graffiti (for both relations s_rho
weak positive linear relation between thefts and street pavement ra
and a very weak negative positive relation between thefts and s
p-value = 0.01068), as well as thefts and street width (s_rho = − 3.1. Temporl and Spatial Distribution of Data
demonstrate any common trends. To check
theft depend on human activities during pa Selection of the Prediction Model
We trained eight different machine learning algorithms for the regression task on the training
dataset with 12,794 samples and 10-fold cross validation with three repeats, tuning the following
parameters: (1) For the linear regression, the intercept is included; (2) for the elastic-net, we use the
tuned best-fit parameters lambda λ = 0 that corresponds to zero amount of shrinkage and the mixing
parameter alpha α = 1 that performs the lasso fit; (3) for the SVM with a radial kernel we use the
To narrow our search from the most opportune days and time for committing a theft to territories
where a theft is most likely to occur, we performed a hot spot analysis (Figure 2B). Hot spot analysis
demonstrates the most potentially dangerous territories located in East Village and some spots in
Uptown Manhattan, i.e., Washington Heights and Inwood, whereas the safest areas are Upper East
Side and Financial District. The reason for the latter, however, appears to be restrictions on parking a
private vehicle on many streets. In order to pinpoint the crime locations more accurately, we performed 8 of 15 Sustainability 2019, 11, 2848 a correlation and regression analysis to find urban generators of thefts and, based on these generators,
to come closer to the specific street segments with high theft prediction. a correlation and regression analysis to find urban generators of thefts and, based on these generators,
to come closer to the specific street segments with high theft prediction. 3.2. Selection of the Prediction Model
We trained eight different machine learning algorithms for
dataset with 12,794 samples and 10-fold cross validation with
parameters: (1) For the linear regression, the intercept is include
tuned best-fit parameters lambda λ = 0 that corresponds to zero a
parameter alpha α = 1 that performs the lasso fit; (3) for the SV
tuned best-fit parameters cost C = 1 and gamma γ = 2.908401; (4)
use the tuned best-fit parameter cost C = 1; (5) for the decision t
parameter cp = 0.01185767; (6) for the bagged CART, we do not t
random forest, we tune the number of trees to be 500 and mtry
boosting (gbm), we tune the number of trees equal to 150, the max
interaction.depth = 3, the shrinkage = 0.1, and the minimum num
nodes n.minobsinnode = 10. The number of thefts from motor veh
the target variable (dependent variable). 3.2. Selection of the Prediction Model Because the
elastic-net increases bias to reduce overfitting (and, accordingly, to achieve lower variance) and none
of our models suffer from overfitting, we choose linear regression as the final prediction model for
thefts from motor vehicles. With linear regression, the model produces less bias, and preserves the
same RMSE and R2. Table 2. Comparison of prediction models using root mean square error (RMSE) and coefficient of
determination (R2). Table 2. Comparison of prediction models using root mean square error (RMSE) and coefficient of
determination (R2). Model
RMSEmin
RMSEmean
R2mean
R2max
Linear regression
0.4407129
0.6049274
0.6303150
0.8730436
Elastic-net
0.4407129
0.6049274
0.6303150
0.8730436
SVMRadial
0.4548175
0.6387557
0.5693278
0.7638390
SVMLinear
0.4486161
0.6281775
0.6133442
0.8739586
Decision tree (CART)
0.4470269
0.6164779
0.6024365
0.8418731
BaggedCART
0.4393015
0.6114266
0.6154108
0.8664760
Random forest
0.4404932
0.6053636
0.6258950
0.9007527
Stochastic Gradient Boosting
0.4446763
0.6102181
0.6194829
0.8680953
3.3. Prediction Model Using Multiple Linear Regression 3.3. Prediction Model Using Multiple Linear Regression 3.3. Prediction Model Using Multiple Linear Regression The Pearson correlation coefficient demonstrates a moderate positive linear relation between thefts
and subway entrances (s_rho = 0.49, p-value < 2.2 × 10−16, a weak positive linear relation between thefts
and restaurants, as well as thefts and graffiti (for both relations s_rho = 0.19, p-value < 2.2 × 10−16), a very
weak positive linear relation between thefts and street pavement rating (s_rho = 0.02, p-value = 0.02153),
and a very weak negative positive relation between thefts and street segment length (s_rho = −0.02,
p-value = 0.01068), as well as thefts and street width (s_rho = −0.02, p-value = 0.005803). Variables 9 of 15 Sustainability 2019, 11, 2848 having very weak relations with thefts are excluded from the regression (street pavement rating, street
segment length, and street segment width). having very weak relations with thefts are excluded from the regression (street pavement rating, street
segment length, and street segment width). During the building of the linear regression model, we considered the significant variables (subway
entrances, restaurants, and graffiti). Then, if necessary, we dropped the one with the highest p-value
until all p = values were reasonable (p-value < 0.05). The output of the multiple linear regression is
presented in the Table 3. Table 3. Multiple linear regression for prediction of thefts from motor vehicles. Estimate
Std. 3.2. Selection of the Prediction Model Error
t Value
Pr (>|t|)
Intercept
0.051844
0.002463
21.05
<2 × 10−16***
Subway
1.158844
0.018242
63.52
<2 × 10−16***
Graffiti
0.125207
0.007937
15.77
<2 × 10−16***
Restaurant
0.054781
0.004021
13.62
<2 × 10−16*** Table 3. Multiple linear regression for prediction of thefts from motor vehicles. In the Table 3 *** corresponds to the significance level 0.000. In the Table 3 *** corresponds to the significance level 0.000. With p-values < 0.05 for all variables, we conclude there is strong evidence against the null
hypothesis, H0 that the predictors have no effect on the mean level of the response variable (thefts from
motor vehicles). To examine if the prediction model is correct, we checked the following assumptions: motor vehicles). To examine if the prediction model is correct, we checked the following assumptions:
Assumption 1, “Mean of residuals is zero:” In our case, the assumption is met with mean of
residuals equal to −2.278831 × 10−16. Assumption 1, “Mean of residuals is zero:” In our case, the assumption is met with mean of
residuals equal to −2.278831 × 10−16. Assumption 2, “Homoscedasticity of residuals or equal variance” (by checking residuals vs. fitted
values): In our case, the assumption is met because the data is not evenly dispersed and does not form
clusters on the residual vs. fitted graph. Assumption 2, “Homoscedasticity of residuals or equal variance” (by checking residuals vs. fitted
values): In our case, the assumption is met because the data is not evenly dispersed and does not form
clusters on the residual vs. fitted graph. Assumption 3, “No autocorrelation of residuals:” The assumption is met since the residuals are
not autocorrelated. The computed Durbin-Watson statistic = 2.0326, p-value < 0.9717, and therefore,
we accept the null hypothesis H0: Autocorrelation is not greater than 0. Assumption 3, “No autocorrelation of residuals:” The assumption is met since the residuals are
not autocorrelated. The computed Durbin-Watson statistic = 2.0326, p-value < 0.9717, and therefore,
we accept the null hypothesis H0: Autocorrelation is not greater than 0. Assumption 4, “The X variables and residuals are uncorrelatedL” The assumption is met, as all
three variables are uncorrelated with residuals. For subways and residuals cor = 2.440349 × 10−16
with p-value = 1, for graffiti and residuals cor = 1.01497 × 10−15 with p-value = 1, for restaurants and
residuals cor = −2.225019 × 10−16 with p-value = 1. 3.2. Selection of the Prediction Model Assumption 5, “No multicollinearity:” Assumption is met for subways, restaurants, and graffiti
VIF = 1. If VIF of a variable is high, the information in that variable is explained by other X variables
present in the given model and the variable is redundant. Therefore, the lower VIF (<2) the better. Assumption 6, “Normality of residuals:” Assumption is not met, as residuals are not
normally distributed. Since five of six assumptions are met, the prediction model is built correctly. However, the
accuracy of the model is 77% (R-squared = 0.77), which implies that not all variables generating
thefts from motor vehicles are considered in this research. Nonetheless, the test of this model on the
test set demonstrates a good fit regarding error mean (Actual thefts—Predicted theft = 0.04692426)
and correlation between actual and predicted thefts (cor = 0.8999567), even though residuals are not
normally distributed. 3.4. Application of Research Results to Safer Parking Using Shiny package for R, we developed the prototype of the application of our research results
for safer parking that can be used by city dwellers, police, and insurance companies (Figure 3). City
dwellers can find safety information for the selected street segment for the available parking. The
developed Android App would take users’ provided street address or a current location into account
and provide information about the parking safety in that location. It will also guide users to safe
parking streets nearby. Police officers can use the application to allocate forces in risky areas. Insurance
companies can tailor insurance prices for different cases based on users’ parking behavior. For instance, 10 of 15
10 of 15 Sustainability 2019, 11, 2848
S
i
bili
2019 11
FOR if a client lives and parks his motor vehicle on weekday nights and weekend days in Upper East Side,
his motor vehicle is safe. If he works at daytime during weekdays and parks his vehicle close to his
office in Midtown, he is safe too, as no significant hot spots of thefts are identified here. However, if he
drives on weekend nights to East Village and parks his car there, this is where the trouble begins, as
this area is identified as a hot spot of thefts from motor vehicles (with 99–95% confidence). Thus, an
insurance company can offer a client the price regarding his parking behavior and locations during
the week. The application is based on the prediction model, where the predicted theft rate increases with
the increase of number of subway entrances, graffiti, and restaurants. Therefore, all the variables with
their values are mapped. Each street segment is associated with number of subway entrances, graffiti,
and restaurants currently located on that segment. When a user chooses a street segment of his
interest, he sees information about dependent variables there (number of subway entrances, graffiti,
and restaurants), and the predicted value of thefts from motor vehicles is computed (based on data
and prediction model) and displayed. Figure 3. Application of the research results for safer parking. 3.4. Application of Research Results to Safer Parking Also, a user can choose how to display
the location—using a street address or a zip code, and the level of theft rate to display—a high (red
color), medium (yellow), or low (green). 4. Discussion
The comparison of similar studies with our research results demonstrates that the biggest part
of research is performed toward predicting hot sport or areas or neighborhoods, while predicting the
exact location of crime to happen still remains the target for many researchers, data scientists, and
crime analysts. Moreover, the majority of crime prediction models are built using data classification
methods [42–45], differing from our research using regression methods. For instance, Bogomolov et
al [42] applied decision tree classifier based on the Breiman’s random forest and achieved accuracy
The application is based on the prediction model, where the predicted theft rate increases with
the increase of number of subway entrances, graffiti, and restaurants. Therefore, all the variables
with their values are mapped. Each street segment is associated with number of subway entrances,
graffiti, and restaurants currently located on that segment. When a user chooses a street segment of his
interest, he sees information about dependent variables there (number of subway entrances, graffiti,
and restaurants), and the predicted value of thefts from motor vehicles is computed (based on data
and prediction model) and displayed. 3.4. Application of Research Results to Safer Parking When a user chooses a street segment
of his interest, he sees information about dependent variables there (number of subway entrances,
graffiti, and restaurants as Subway, Restaurant, and Graffiti accordingly on the picture above), and
the predicted value of thefts from motor vehicles is computed (based on the data and the prediction
model) and displayed—as Predict theft on the picture above. Also, a user can choose how to display
the location—using a street address or a zip code, and the level of theft rate to display—a high (red
color), medium (yellow), or low (green). Figure 3. Application of the research results for safer parking. When a user chooses a street segment
of his interest, he sees information about dependent variables there (number of subway entrances,
graffiti, and restaurants as Subway, Restaurant, and Graffiti accordingly on the picture above), and
the predicted value of thefts from motor vehicles is computed (based on the data and the prediction
model) and displayed—as Predict theft on the picture above. Also, a user can choose how to display
the location—using a street address or a zip code, and the level of theft rate to display—a high (red
color), medium (yellow), or low (green). Figure 3. Application of the research results for safer parking. When a user chooses a street segment
of his interest, he sees information about dependent variables there (number of subway entrances,
graffiti, and restaurants as Subway, Restaurant, and Graffiti accordingly on the picture above), and
the predicted value of thefts from motor vehicles is computed (based on the data and the prediction
model) and displayed—as Predict theft on the picture above. Also, a user can choose how to display
the location—using a street address or a zip code, and the level of theft rate to display—a high (red
color), medium (yellow), or low (green). Figure 3. Application of the research results for safer parking. When a user chooses a street segment
of his interest, he sees information about dependent variables there (number of subway entrances,
graffiti, and restaurants as Subway, Restaurant, and Graffiti accordingly on the picture above), and
the predicted value of thefts from motor vehicles is computed (based on the data and the prediction
model) and displayed—as Predict theft on the picture above. of 70% while p
and demograp
4. Discussion Street segment
with the third highest theft rate in Manhattan: West 138th Street. Urban reasons of unsafety: No
natural surveillance, no lighting, no pedestrian flows, no closed-circuit television (CCTV) cameras, etc. Nothing meets CPTED (Crime Prevention Through Environmental Design) requirements for safety
and security. Moreover, there is a sign in this tunnel permitting parking, and therefore, contributing to
the availability of motor vehicles as targets for theft. The limitations of this research include an unsatisfactory determination coefficient of the
prediction model (only 77% of variance in thefts from motor vehicles is explained by the independent
variables), as well as automation of loading the updated data into the pipeline for making predictions
up-to-date. The first limitation could be improved by adding more datasets and features in order to
find other important variables that contribute to the theft rate increase or decrease. The second
limitation requires to constantly obtain the newest data about the city and crime, to update the
database, to load the new data into the pipeline, that includes data preprocessing, model building,
testing, validation, and deployment, and to produce the predicted theft rates together with important
factors. Moreover, continuing this research we would like to try other units for data aggregation—
such as a square area within the city (for instance, 3 × 3 m, 10 × 10, or similar), instead of the street
segment that we were using for this current research. As per the most recent research in machine
The limitations of this research include an unsatisfactory determination coefficient of the prediction
model (only 77% of variance in thefts from motor vehicles is explained by the independent variables),
as well as automation of loading the updated data into the pipeline for making predictions up-to-date. The first limitation could be improved by adding more datasets and features in order to find other
important variables that contribute to the theft rate increase or decrease. The second limitation requires
to constantly obtain the newest data about the city and crime, to update the database, to load the
new data into the pipeline, that includes data preprocessing, model building, testing, validation, and
deployment, and to produce the predicted theft rates together with important factors. of 70% while p
and demograp
4. Discussion The prediction model of homicides, a regression approach developed by
Alves et al. [46] while using the random forest regression, achieves 97% accuracy. The most unsafe street segments are revealed by applying the search algorithm in ArcGIS. It is
clear from the photographs (Figure 4A–C) that the urban reasons of these places being unsafe lay in
not meeting the main principles of CPTED (Crime Prevention Through Environmental Design): Lack
of natural surveillance (almost no observation from windows, no entrance doors, i.e., no “eyes on the
street”), the nearest windows are too far to observe the environment or do not exist at all (walls and
fences with no windows, i.e., ‘blind walls’), no inter-visibility of constructions (for instance, the fence is
not inter-visible on Figure 4A), spaces are not frequently used by pedestrians, spaces are dark, and
there is no lighting (Figure 4C), etc. In all the above-mentioned cases, the urban environment should
be redesigned to make parking safer. If redesign of urban spaces is not possible, signs preventing
parking should be placed on unsafe street segments. Sustainability 2019, 11, x FOR PEER REVIEW
11 of 15
be redesigned to make parking safer. If redesign of urban spaces is not possible, signs preventing
parking should be placed on unsafe street segments. Figure 4. (A). Street segments with the highest theft rate in Manhattan: Department of Vascular
Surgery. Urban reasons of unsafety: Lack of natural surveillance (no “eyes on the street”), nearest
windows are far, fence is not inter-visible, and blind wall (wall with no windows or doors). (B) Street
segments with the highest theft rate in Manhattan: Morningside Church & Sakura park. Urban
reasons of unsafety: Pedestrians rarely use this space, no natural surveillance (almost no observation
from windows, no entrance doors) with a church on one side and park on another. (C). Street segment
with the third highest theft rate in Manhattan: West 138th Street. Urban reasons of unsafety: No
natural surveillance, no lighting, no pedestrian flows, no closed-circuit television (CCTV) cameras,
etc. Nothing meets CPTED (Crime Prevention Through Environmental Design) requirements for
safety and security. Moreover, there is a sign in this tunnel permitting parking, and therefore,
contributing to the availability of motor vehicles as targets for theft. A
A
Figure 4. (A). Street segments with the highest theft rate in Manhattan: Department of Vascular
Surgery. of 70% while p
and demograp
4. Discussion and demographic data of London, UK. Antolos et al. [43] achieved accuracy of 74 83% while using
the logistic regression to predict burglary based on the day of the week, time of the day, repeated
victimization, connectors and barriers, id est., and historical crime data from 2010. Other machine
learning approaches demonstrate reasonable accuracy when applying support vector machines
(SVM) to predict crime hot spots [44], as well as Bayesian approach to predict the neighborhood
where the next crime will happen [45]. The prediction model of homicides, a regression approach
developed by Alves et al. [46] while using the random forest regression, achieves 97% accuracy. The most unsafe street segments are revealed by applying the search algorithm in ArcGIS. It is
clear from the photographs (Figure 4A–C) that the urban reasons of these places being unsafe lay in
not meeting the main principles of CPTED (Crime Prevention Through Environmental Design): Lack
of natural surveillance (almost no observation from windows, no entrance doors, i.e., no “eyes on the
street”), the nearest windows are too far to observe the environment or do not exist at all (walls and
fences with no windows, i.e., ‘blind walls’), no inter-visibility of constructions (for instance, the fence
The comparison of similar studies with our research results demonstrates that the biggest part of
research is performed toward predicting hot sport or areas or neighborhoods, while predicting the
exact location of crime to happen still remains the target for many researchers, data scientists, and
crime analysts. Moreover, the majority of crime prediction models are built using data classification
methods [42–45], differing from our research using regression methods. For instance, Bogomolov
et al. [42] applied decision tree classifier based on the Breiman’s random forest and achieved accuracy
of 70% while predicting if a specific urban area will be a crime hotspot or not using mobile phone
and demographic data of London, UK. Antolos et al. [43] achieved accuracy of 74–83% while using
the logistic regression to predict burglary based on the day of the week, time of the day, repeated
victimization, connectors and barriers, id est., and historical crime data from 2010. Other machine
learning approaches demonstrate reasonable accuracy when applying support vector machines (SVM)
to predict crime hot spots [44], as well as Bayesian approach to predict the neighborhood where the Sustainability 2019, 11, 2848 11 of 15 next crime will happen [45]. of 70% while p
and demograp
4. Discussion Urban reasons of unsafety: Lack of natural surveillance (no “eyes on the street”), nearest
windows are far, fence is not inter-visible, and blind wall (wall with no windows or doors). (B) Street
segments with the highest theft rate in Manhattan: Morningside Church & Sakura park. Urban reasons
of unsafety: Pedestrians rarely use this space, no natural surveillance (almost no observation from
windows, no entrance doors) with a church on one side and park on another. (C). Street segment
with the third highest theft rate in Manhattan: West 138th Street. Urban reasons of unsafety: No
natural surveillance, no lighting, no pedestrian flows, no closed-circuit television (CCTV) cameras, etc. Nothing meets CPTED (Crime Prevention Through Environmental Design) requirements for safety
and security. Moreover, there is a sign in this tunnel permitting parking, and therefore, contributing to
the availability of motor vehicles as targets for theft. A A A A Figure 4. (A). Street segments with the highest theft rate in Manhattan: Department of Vascular
Surgery. Urban reasons of unsafety: Lack of natural surveillance (no “eyes on the street”), nearest
windows are far, fence is not inter-visible, and blind wall (wall with no windows or doors). (B) Street
segments with the highest theft rate in Manhattan: Morningside Church & Sakura park. Urban
reasons of unsafety: Pedestrians rarely use this space, no natural surveillance (almost no observation
from windows, no entrance doors) with a church on one side and park on another. (C). Street segment
with the third highest theft rate in Manhattan: West 138th Street. Urban reasons of unsafety: No
natural surveillance, no lighting, no pedestrian flows, no closed-circuit television (CCTV) cameras,
etc. Nothing meets CPTED (Crime Prevention Through Environmental Design) requirements for
safety and security. Moreover, there is a sign in this tunnel permitting parking, and therefore,
contributing to the availability of motor vehicles as targets for theft. Figure 4. (A). Street segments with the highest theft rate in Manhattan: Department of Vascular
Surgery. Urban reasons of unsafety: Lack of natural surveillance (no “eyes on the street”), nearest
windows are far, fence is not inter-visible, and blind wall (wall with no windows or doors). (B) Street
segments with the highest theft rate in Manhattan: Morningside Church & Sakura park. Urban reasons
of unsafety: Pedestrians rarely use this space, no natural surveillance (almost no observation from
windows, no entrance doors) with a church on one side and park on another. (C). 5. Conclusions The geospatial data of thefts from motor vehicles reveal some interesting patterns that suggest
that human activity, as well as features of the urban environment, contribute to the frequency of crime. Time series analysis reveals a common theft pattern for Tuesdays, Wednesdays, and Thursdays with
theft rates increasing from 4 p.m. to between 7 p.m. and 8 p.m., and the lowest theft rates being
between 2 a.m. and 6 a.m. A plausible explanation for these observations is that while thefts might
occur throughout the day, most are discovered and reported when people return to their vehicles after
leaving their places of work. Similarly, when commuters leave the city during evenings and weekends,
the number of cars exposed to theft decreases and crime reporting drops. Clearly, other explanations
might also be possible. The geospatial analysis of theft data demonstrates a spatial pattern of locations where thefts were
committed on Tuesdays, Wednesdays, and Thursdays as well. It suggests that during the middle
of the week, criminals’ behavior regarding thefts from motor vehicles has the same pattern, with
similar opportunities for committing theft dictated by the urban environment, location of vulnerable
motor vehicles, and other factors, such as the presence of witnesses and cameras. Moreover, visual
comparative analysis of theft rates mapped on street segments demonstrates “movement” of thefts
within Manhattan varying over time, such as the shifting intensity of theft, which diminishes during
morning and evening commute hours. It demonstrates the dynamic nature of crime. Hot spot analysis reveals the most crime-ridden areas are located in the East Village and some
areas in Uptown Manhattan: Washington Heights and Inwood. The East Village has become more
gentrified in recent years after previously being a bohemian neighborhood. Nowadays, it is known
for a large variety of nightlife and is home to artists and diverse communities. Washington Heights
used to be a territory for drug dealers with high crime rates, and Inwood is still one of the most unsafe
neighborhoods in Manhattan, having many aging multifamily five- to eight-story buildings and almost
no new construction. Comparing these results with theft rates for street segments within different time
ranges might explain the hot spots of thefts: Once residents leave their apartments for work (around
7 a.m. to 8 a.m.), thefts begin to happen. The safest areas for car theft appear to be the Upper East Side
and Financial District. of 70% while p
and demograp
4. Discussion Moreover,
continuing this research we would like to try other units for data aggregation—such as a square area
within the city (for instance, 3 × 3 m, 10 × 10, or similar), instead of the street segment that we were
using for this current research. As per the most recent research in machine learning/deep learning Sustainability 2019, 11, 2848 12 of 15 and urban science [42,47–51], we assume that using a square area of small granularity will bring our
prediction closer to the exact location where and when a crime can happen. and urban science [42,47–51], we assume that using a square area of small granularity will bring our
prediction closer to the exact location where and when a crime can happen. Despite using several machine learning methods, we were not able to identify other significant
urban factors using geospatial data analysis and machine learning tools because the open datasets on
urban factors are limited or nonexistent. Therefore, in the future, we might implement more complex
machine learning tools, such as object recognition, deep learning, etc., to detect urban factors in street
views, and record, process, and extract knowledge from streetscape photo or video data. However, the
methods we developed thus far should be useful in a general context for insurance companies and
crime prevention in urban areas and have applications in risk management strategies for insurers. 5. Conclusions In the Financial District, this may simply be a result of parking restrictions on
private vehicles on many streets. However, there also appears to be a socioeconomic divide where
thefts occur: Thefts are lower on the Upper East Side, where incomes are higher. This may reflect
a greater concentration of crime prevention resources in these areas, such as the presence of police,
private security, cameras, and doormen on duty. One practical result of these observations is that
crime prevention resources can be concentrated in the appropriate areas at specific times and thereby
increase the efficiency of resource utilization. Applying eight different machine learning methods on the training set data with 10-fold cross
validation with three repeats reveals that linear models perform better on our data (linear regression,
elastic-net), as well as the random forest does. Though, due to random forest’s good performance on
RMSEmin and R2max (but not on mean values of RMSE and R2), and because the elastic-net produces
bias to reduce overfitting (though, overfitting does not occur in our case), we chose a linear regression
to build the final prediction model. Multiple linear regression was useful in unveiling the following 13 of 15 Sustainability 2019, 11, 2848 urban generators of thefts from motor vehicles: A higher number of subway entrances, graffiti, and
restaurants on streets contributes to higher theft rates. Subway entrances, for example, might provide
easy access and escape from an area after a crime is committed, as well as a greater flow of people
who are not resident in the neighborhood. Although the prediction model for thefts meets almost
all assumptions (five of six), its accuracy is not very high (only 77%), suggesting that there are other
undiscovered factors making a contribution to the generation of thefts. Author Contributions: Conceptualization, I.M.; methodology, I.M.; software, I.M., A.M., V.J.; validation, I.M.;
formal analysis, I.M., A.M.; data curation, I.M.; writing—original draft preparation, I.M.; writing—review and
editing, I.M.; visualization, I.M., A.M.; supervision, I.M. Funding: This research received no external funding. Acknowledgments: This work was conducted in collaboration with Liberty Mutual, Boston, MA. A special thank
you for professor Sylvain Jaume for his advice and support. Conflicts of Interest: The authors declare no conflict of interest. Conflicts of Interest: The authors declare no conflict of interest. References 1. King, L.O. Comprehensive Sustainability Indicators: The Houston Sustainability Indicators Program
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International Publishing: London, UK, 2017; pp. 167–184. g
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3856–3866. Available online: https://arxiv.org/pdf/1710.09829.pdf (accessed on 28 March 2019). 51. Lv, Y.; Duan, Y.; Kang, W.; Li, Z.; Wang, F.Y. Traffic flow prediction with big data: A deep learning approach. IEEE Trans. Intell. Transp. Syst. 2015, 16, 865–873. [CrossRef] 51. © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/). References Lv, Y.; Duan, Y.; Kang, W.; Li, Z.; Wang, F.Y. Traffic flow prediction with big data: A deep learning approach. IEEE Trans. Intell. Transp. Syst. 2015, 16, 865–873. [CrossRef] © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/). © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
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Thiamethoxam exposure deregulates short ORF gene expression in the honey bee and compromises immune response to bacteria
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Scientific reports
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Thiamethoxam exposure
deregulates short ORF gene
expression in the honey bee
and compromises immune
response to bacteria Pâmela Decio1,5, Pinar Ustaoglu2,5, Kamila Derecka3, Ian C. W. Hardy3, Thaisa C. Roat1,
Osmar Malaspina1, Nigel Mongan4, Reinhard Stöger3* & Matthias Soller2* Maximizing crop yields relies on the use of agrochemicals to control insect pests. One of the
most widely used classes of insecticides are neonicotinoids that interfere with signalling of the
neurotransmitter acetylcholine, but these can also disrupt crop-pollination services provided by bees. Here, we analysed whether chronic low dose long-term exposure to the neonicotinoid thiamethoxam
alters gene expression and alternative splicing in brains of Africanized honey bees, Apis mellifera,
as adaptation to altered neuronal signalling. We find differentially regulated genes that show
concentration-dependent responses to thiamethoxam, but no changes in alternative splicing. Most
differentially expressed genes have no annotated function but encode short Open Reading Frames,
a characteristic feature of anti-microbial peptides. As this suggested that immune responses may be
compromised by thiamethoxam exposure, we tested the impact of thiamethoxam on bee immunity
by injecting bacteria. We show that intrinsically sub-lethal thiamethoxam exposure makes bees more
vulnerable to normally non-pathogenic bacteria. Our findings imply a synergistic mechanism for the
observed bee population declines that concern agriculturists, conservation ecologists and the public. The western honey bee Apis mellifera is highly beneficial for human societies. Besides honey production, this
semi-domesticated insect plays a critical role in sustaining global food security through the provision of man-
aged pollination services that contribute to increased yields of many crops. Globally, crop productivity is also
enhanced by the application of pesticides, including insecticides. Agrochemicals are, however, among the key
factors implicated in contributing to declining bee health and abundance and there is a need to find a balance
between the necessity of insecticide applications and the unintended effects to non-target species1–6. f
Compared to organophosphate pesticides, neonicotinoids are relatively safe for vertebrates and have been
one of the most important classes of insecticides for the last three decades7. Neonicotinoids can be applied to
seeds and they then spread systemically within growing plants, killing leaf-eating pests. This reduces non-target
effects compared to spraying but still represents a toxic burden for pollinators as neonicotinoids are commonly
found in wildflowers adjacent to treated fields8,9. l
ji
Neonicotinoids act as agonists, competing with the neurotransmitter acetylcholine in binding to nicotinic
acetylcholine receptors (nAChR)10–12. The increased toxicity for insects is thought to be caused by the charac-
teristically high nAChR density within insect nervous systems13. www.nature.com/scientificreports www.nature.com/scientificreports www.nature.com/scientificreports Materials and methods We assessed whether the neonicotinoid thiamethoxam alters gene expression in the brain of Africanized honey
bees, Apis mellifera, after long-term exposure to field-relevant concentrations. Thiamethoxam exposures. Genetically unrelated bees from three unrelated hives from different loca-
tions in the apiary of the Biosciences Institute, UNESP, Rio Claro-SP (Brazil). Brood frames were removed from
three different colonies and kept for 24 h in biochemical oxygen demand (BOD) at 34 °C and relative humidity
of 80 ± 10%. The newly emerged bees (aged 0–24 h) were marked with special non-toxic pen (Posca pen) and
returned to their respective colonies until they were collected after 15 days. All experiments followed the recom-
mendations of the guidelines for xenobiotic assessments on bees45,46.ht g
Thereafter, bees were kept in groups of 20 individuals in small cages (12 cages/treatment) within an incubator
at 32 °C. Bees were fed ad libitum with sugar solution (1:1 water and inverted sugar) from pierced 2 ml Eppendorf
tubes for the control group (dataset A, Supplemental Data 1) and, for toxin exposure treatments, thiamethoxam
(Sigma) was added at 2 ng/ml (low dose, LD, dataset B, Supplemental Data 1) or at 50 ng/ml (high dose, HD,
dataset C, Supplemental Data 1). Sugar-feeders were re-filled before they were empty to guarantee continuous
consumption. The tested concentrations were chosen based on the range of realistic field concentrations found
in pollen and nectar9,47–50. Assuming that caged worker bees consume about 22 µl of 50% sugar per day51 the
bees would have ingested 0.44 ng and 11 ng after ten days for the low and high dose exposure, respectively. After
10 days, bees were cold-anaesthetised and their brains were dissected for RNA extraction. RNA extraction, illumina sequencing, analysis of differential gene expression and splic-
ing. Total RNA was extracted from ten dissected brains per sample and three replicates were done for the
control, the low and the high dose treatment (dataset A, Supplemental Data 1). To extract total RNA, brains
were cracked in liquid nitrogen with a pestle and then homogenized in 50 µl of Tri-reagent (SIGMA). Then the
volume was increased to 500 µl and proceeded following the manufacturer’s instructions. Total RNA was then
stored in 70% ethanol and transported at ambient temperature from Brazil to the UK. www.nature.com/scientificreports/ extensive alternative splicing, they are prime targets for changes induced by xenobiotics17,19. Moreover, sub-lethal
uptake of some neonicotinoids affects the honey bee brain, impairing foraging behaviour, flight, navigation,
communication, learning and memory20–25. Neonicotinoid exposure has also been linked to a decline in bee health, including a reduction of immune
competence26–28. Insects do not have antibodies and rely on the innate immune system to fight microbial infec-
tions; both cellular and humoral responses have been identified29, 30. The cellular response leading to phagocytosis
or encapsulation of pathogens is mediated by three types of haematopoietic cell lineages29,31,32. The humoral
immune response is activated by Toll and Imd33 pathways, leading to the expression of short (10–100 amino
acids) antimicrobial peptides (AMPs) which are highly diverse among insect species34. The Toll-pathway is trig-
gered by Gram-positive bacteria and fungi, ultimately leading to expression of AMPs via the NFkappa-B transcrip-
tion factor Dif, that are then secreted from the fat body into the haemolymph29,35,36. The Immune deficiency (Imd)
pathway leads to expression of a different set of AMPs via the NFkappa-B transcription factor relish in response to
peptidoglycans from primarily Gram-negative bacterial through the activation of pattern recognition receptors
and a complex intracellular signalling cascade29,30,35. In addition to the fatbody, AMP expression activated by the
Imd pathway has also been detected in glial cells in the brain37–40. y
g
We have previously shown that worker-bee larvae in colonies contaminated with the neonicotinoid imida-
cloprid, have altered expression of genes belonging to lipid-carbohydrate-mitochondrial metabolic networks41. We have further demonstrated that sub-lethal exposure to thiamethoxam, another neonicotinoid, can cause
impairment in the midgut and brain of the Africanized Apis mellifera, as well as contribute to a reduction in
honey bee lifespan9,42–44.f y
In this study, we analysed the effects of chronic, sub-lethal, thiamethoxam exposure on genome-wide gene
expression and alternative splicing in the brains of honey bee workers. We found 52 differentially regulated
genes showing a concentration dependent response to thiamethoxam exposure. Most of these genes have no
annotated function but the vast majority are characterized by encoding short Open Reading Frames (sORFs),
half of which are predicted to encode antimicrobial peptides. This result suggested that immune responses may
be compromised by thiamethoxam exposure. Indeed, we found that thiamethoxam-exposed bees that were
also infected with bacteria had greatly decreased viability compared to infected but chemically unexposed bees. www.nature.com/scientificreports/ Overall, our results suggest that thiamethoxam makes bees vulnerable to bacterial infection by compromising
immune responses. Thiamethoxam exposure
deregulates short ORF gene
expression in the honey bee
and compromises immune
response to bacteria Detailed knowledge of cellular and molecular
effects of insecticide exposure is required to mitigate negative effects or refine target specificity. Changes in gene
expression and processing of RNAs, including alternative splicing, are among the mechanisms available to an
organism to adapt to environmental perturbations14,15. Sub-lethal exposure to xenobiotics, such as pesticides,
can alter alternative splicing16–18. Since many ion channel genes and other important neuronal genes undergo 1Institute of Biosciences, São Paulo State University (Unesp), Rio Claro, Brazil. 2School of Biosciences, College of
Life and Environmental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK. 3School of
Biosciences, University of Nottingham, Sutton Bonington, Loughborough LE12 5RD, UK. 4School of Veterinary
Medicine and Science, University of Nottingham, Sutton Bonington, Loughborough LE12 5RD, UK. 5These authors
contributed equally: Pâmela Decio and Pinar Ustaoglu. *email: reinhard.stoger@nottingham.ac.uk; m.soller@
bham.ac.uk | https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 www.nature.com/scientificreports/ Materials and methods p
p
Total RNA was treated with DNase I (Ambion) and stranded libraries for Illumina sequencing were prepared
after poly(A) selection from total RNA (1 μg) with the TruSeq stranded mRNA kit (Illumina) using random
primers for reverse transcription according to the manufacturer’s instructions. Pooled indexed libraries were
sequenced on an Illumina HiSeq2500 to yield 28–41 million paired-end 125 bp reads for three control, three
low dose and three high dose samples. After demultiplexing, sequence reads were aligned to the Apis mellifera
genome (Amel-4.5-scaffolds) using Tophat2.0.652. Differential gene expression was determined by Cufflinks-
Cuffdiff and the FDR-correction for multiple tests to raw p values, with p < 0.05 considered significant53. Illumina
sequencing and differential gene expression analysis was carried out by Fasteris (Switzerland). Sequencing data
are deposited in GEO under GSE132858.i p
Alternative splicing was analysed by rMATS54 and manually inspected by uploading bam files into the Inte-
grated Genome Viewer55 and comparing read frequency. Comparison of gene lists was made with Venny 2.1
(https://bioinfogp.cnb.csic.es/tools/venny/). Protein sequences from differentially expressed genes of bees were https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 | www.nature.com/scientificreports/ obtained from ensemble (http://metazoa.ensembl.org/Apis_mellifera/Info/Index) and blasted against Drosophila
annotated proteins using flybase (http://flybase.org) to assign gene functions. Proteins with no assigned functions
were scanned for motifs using the Interpro server (https://www.ebi.ac.uk/interpro/)56. Short ORFs were analysed
for antimicrobial peptide prediction using the following server: https://www.dveltri.com/ascan/v2/ascan.html57. obtained from ensemble (http://metazoa.ensembl.org/Apis_mellifera/Info/Index) and blasted against Drosophila
annotated proteins using flybase (http://flybase.org) to assign gene functions. Proteins with no assigned functions
were scanned for motifs using the Interpro server (https://www.ebi.ac.uk/interpro/)56. Short ORFs were analysed
for antimicrobial peptide prediction using the following server: https://www.dveltri.com/ascan/v2/ascan.html57. Reverse transcription quantitative polymerase chain reaction (RT‑qPCR). Reverse transcription
was carried out with Superscript II (Invitrogen) as previously described58. PCR was performed as described and
PCR products were analysed on ethidium bromide stained 3% agarose gels19. Neuronal genes used as reference
for gene expression comparison were bee erect wing (ewg, GB44653) and Amyloid Precursor Protein-like (Appl,
GB48454), based on expression analysis in Drosophila59,60. To validate cDNAs obtained from RT, the following
primers were used to amplify bee ewg with AM ewgG F (CCTGATGGTACCGTATCAATTATTCAAGTTG) and
AM ewgH R (CCGTGTCCATCTTCTCCTGTGAGAATGATTTG), bee Appl with AM Appl F (GCGCGATTC
CAGGAAACTGTTGCTGCTC) and AM Appl R2 (CTGCTGTCCAGCAGATGTTTGTAATGAG). Materials and methods Additional
primers were GB45995 (GTTGCATTTTTACGCGTACAGTTACACGACAG) and GB45995 R (GGGAAATCC
CCGGGAAGAGAGCAACTGGAG), GB45995 F (GTTGCATTTTTACGCGTACAGTTACACGACAG) and
GB45995 R (GGGAAATCCCCGGGAAGAGAGCAACTGGAG), and GB47479 F (GGGCTATTTGCTATC
TAAGTGATCCTCC) and GB47479 R (GGGTTTAGGAGTTTTCGTTTTAGCTGCTG) and PCR amplifica-
tions were done by 30 s initial denaturation at 94 °C, and then 40 cycles in total with 30 s at 94 °C, annealing at
48 °C with 30 s extension at 72 °C for 2 cycles, then at 50 °C with 30 s extension at 72 °C for 2 cycles and then at
52 °C with and 60 s extension at 72 °C for 36 cycles with a final extension of 2 min at 72 °C. yi
RNA quality and quantity were assessed using an Agilent 6000Nano Kit. 250 ng of DNAse-treated RNA
(the same samples used for deep seq) was reversed transcribed using a SuperScript III Invitrogen kit at 55 °C,
following manufacturer’s instructions. Primers for quantitative real-time PCR (RT-qPCR) were designed by
Primer Express software (exon/exon junctions included except for GB41813). Amplicon sizes were assessed in
15% acrylamide gel, PCR products were then sequenced and efficiency of every pair of primers was included
in the analysis.i y
Duplicate samples of cDNA were amplified in Real Time PCR with SybrGreen chemistry under following
conditions: denaturation 93 °C for 30 s, annealing 60 °C for 30 s, elongation 72 °C for 30 s. GB41923 was ampli-
fied with primers GB41923 F1 (CGCGTTGATCGTCATGATATTG) and GB41923 R1 (CTATAAGGAAATTTT
GAGCCTTCGA), GB40669 with primers GB40669 F1 (GGCCGGATATCGCTTCAAA) and GB40669 R1 (GTC
TCTTTTATCTTTTCCTCGGAATTC), and GB48969 with primers GB48969 F1 (TTGCAGCCGTAGCAAAAG
GTA), GB48969 R1 (ACCGATTTGAGCACCTTGGT) and bubblegum (bgm, GB51680) with primers bgm F1
(CATGCACAAAGAGTACAAAAATTTCA) and bgm R1 (TGGTCCCAATTCTCCAGTAACA). Analysis of
CT values was performed in Light Cycler480 software and normalization and differential expression was deter-
mined with the 2−∆∆Ct method61 by normalizing to the expression of ewg (GB44653), Appl (GB48454) and actin
(GB44311) genes. Actin and ewg were amplified with primers ewg Fw2 (CCGCGTCTCCTACAGCTCTT) and
ewg Rv2 (TGTAAAACTGCCGTAGGATATTGG) and actF1 (TTCCCATCTATCGTCGGAAGA) and actR1
(TTTGTCCCATGCCAACCAT), and Appl with primers AM appl F and AM appl R2, following41,62. Bacterial infection assays. Thiamethoxam-induced alteration of anti-microbial peptide gene expression
could disrupt bees’ immune response. To evaluate the effect of thiamethoxam on immunity, we adopted an assay
procedure, initially developed for Drosophila, which assesses how efficiently injected non-pathogenic bacteria
are cleared by anti-microbial peptides63. Materials and methods To assay clearance in bees we used Bacillus badius, a normally non-path-
ogenic bacterium commonly found in the environment and Ochrobactrum anthropi, which are Gram-positive
and Gram-negative members of the honey bee microbiome64,65. O. anthropi were isolated from worker bee gut
cultures by plating the gut content on LB agar plates incubated at 30 °C.i y p
g
g
g
p
Bacterial species were identified by colony PCR and ribosomal 16S sequencing: a colony was picked from an
LB agar plate with a yellow tip and placed into 10 µl TE in a PCR tube and heated to 94 °C for 5 min. The PCR
mix was added adjusting the MgCl concentration to 1.5 mM. PCR was done for 20–40 cycles with 54 °C annealing
for 40 s and 1 min extension at 72 °C. A 490 bp fragment of the ribosomal 16S gene was amplified with primers
16S F (ACTGAGACACGGYCCAGACTCCTACGTC) and 16S R (GCGTGGACTACCAGGGTATCTAATCC)
and sequenced with primer 16S Fseq (CTCCTACGGGAGGCAGCAGTRGGGTC). If sequences did not yield
a single species, primers 16S F2 (GTGGACTACCAGGGTATCTAATCCTG) and 16S R2 (CCTACGGTTACC
TTGTTACGACTTCAC) were used for amplification of a 733 bp fragment, which was sequenced by 16S R2seq
(CCATGGTGTGACGGGCGGTGTGTAC). (
)
Forager honey bees for infection assays were collected from colonies of the Winterbourne Garden of the
University of Birmingham (UK). They were kept and injected with bacteria as we described previously19. Bacteria
for injections were freshly plated and grown overnight on LB plates. Then a single colony was used to inoculate a
5 ml LB in a 50 ml Falcon tube and grown overnight to saturation: 2 µl of this culture was then injected. Bacterial
titres of cultures were determined at the time of injections by plating 100 µl of 105, 106 and 107 dilutions in LB and
counting the colonies the next day: this showed that 2–8 × 106 B. badius or O. anthropi were typically injected. In some treatments the injected bacteria were diluted tenfold (Fig. 3). Groups of 8 to 12 bees were exposed to
combinations of bacterial and thiamethoxam doses and survival was assessed at 24 h and 48 h. Data on survival were analysed using backwards stepwise logistic ANOVAs adopting a logit-link function
and assuming quasi-binomial distributed errors. These analyses were carried out using GenStat v.19 (VSN Inter-
national, Hemel Hempstead). Statistical analysis of viability were done with GraphPad prism using ANOVA
followed by Tukey–Kramer post-hoc tests. Results
Ch
i t Chronic thiamethoxam exposure effects gene expression. After low (LD) and high dose (HD)
exposure to thiamethoxam, there were 222 up- and 181 down-regulated genes for LD (Fig. 1) and 233 up-
and 114 down-regulated genes for HD with a 1.5 fold difference in expression compared to the control treat-
ment (Fig. 1; Supplemental Data 1). From these differentially regulated genes, 37 were up-regulated and 15 were
down-regulated in a dose-sensitive manner (Fig. 1; Supplemental Data 1). To validate these results from the Illumina sequencing, we performed RT-qPCR for three of the dose-
responsive genes: GB41923, a putative sodium-chloride co-transporter, and GB48969, GB40669, two genes
with unknown function. We detected an expression difference for all three genes upon thiamethoxam exposure
(Supplemental Fig. 1). We also validated and confirmed differential expression of bubblegum, encoding a very
long-chain acyl-CoA synthetase, which has been found to be down-regulated in honey bee larvae exposed to
the neonicotinoid imidacloprid41 (Supplemental Fig. 1). Alternative splicing has been suggested as a mechanism to adapt gene expression to environmental
changes17,66. We analysed the RNA-seq data for changes in alternative splicing but found no conclusive patterns
(Supplemental Information; Supplemental data 2). Dose‑responsive expression occurs mostly in genes encoding uncharacterized ORFs. Next, we
categorized the genes dose-responsive to thiamethoxam according to their functions, taking into account known
functions of orthologues in Drosophila and functions deduced from annotated protein domains retrieved by
BLAST analysis. Amongst the dose-sensitive genes that were up-regulated (Fig. 1A), 14% (5/37) were assigned
roles in cellular signalling (with potential links to altered neuronal function, such as olfactory and taste percep-
tion) and as structural components (cytoskeleton), and 8% (3/37) were assigned functions in transcriptional
regulation of gene expression (Fig. 2A). However, 59% (22/37) of dose-sensitive up-regulated genes and 73%
(11/15) of dose-sensitive down-regulated genes had neither clear orthologues in Drosophila nor any recogniz-
able protein domains that would indicate a biological function (Fig. 2A), which is in contrast to about 20% of
genes with unknown function in gene expression studies in Drosophila67,68. Most thiamethoxam dose‑responsive genes encode short proteins. Many of the dose-responsive,
differentially expressed genes with unknown function coded for short ORFs. For thiamethoxam-induced differ-
entially up- and down-regulated genes, respectively 73% (27/37) and 87% (13/15) encode for genes with ORFs of
250 amino acids or shorter (Fig. 2). Materials and methods Scientific Reports | (2021) 11:1489 | https://doi.org/10.1038/s41598-020-80620-7 www.nature.com/scientificreports/ Figure 1. Thiamethoxam induces differential expression in a subset of genes. Venn diagrams indicating the
number of differentially expressed genes between control bees and bees exposed to a low dose (left) and high
dose (right) of thiamethoxam that were up- (A) or down- (B) regulated. 37 genes were up-regulated and 15
genes were down-regulated in a dose-sensitive manner. Figure 1. Thiamethoxam induces differential expression in a subset of genes. Venn diagrams indicating the
number of differentially expressed genes between control bees and bees exposed to a low dose (left) and high
dose (right) of thiamethoxam that were up- (A) or down- (B) regulated. 37 genes were up-regulated and 15
genes were down-regulated in a dose-sensitive manner. Results
Ch
i t Using a machine learning algorithm57, we predicted from the 40 genes cod-
ing for peptides of ≤ 250 amino acids, that 17 (43%), have antimicrobial function (11/17 genes were up-regulated
and 6/17 were down-regulated, Fig. 2). Thiamethoxam makes worker bees more vulnerable to bacterial infection. When saturated liq-
uid cultures (2–8 Mio bacteria in 2 µl) of B. badius and O. anthropi were injected into bees that were not exposed
to thiamethoxam, viability was little affected, indicating that the bee immune system usually clears the infection
efficiently (Fig. 3, Table 1). In contrast, co-injection of normally sub-lethal doses of thiamethoxam together with
either B. badius or O. anthropi resulted in bee death (Fig. 3, Table 1). In particular, at a dose of 1 µM thiameth- https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 | www.nature.com/scientificreports/ Figure 2. Classification of thiamethoxam induced differentially expressed genes according to function and size. (A) Numbers of genes are plotted according to functions for thiamethoxam induced differentially expressed
genes with annotated functions for up- (top) and down-regulated (bottom) genes. (B) Pie charts indicating the
fraction of thiamethoxam induced differentially expressed genes according the ORF length for up- (left) and
down- (right) regulated genes. Figure 2. Classification of thiamethoxam induced differentially expressed genes according to function and size. (A) Numbers of genes are plotted according to functions for thiamethoxam induced differentially expressed
genes with annotated functions for up- (top) and down-regulated (bottom) genes. (B) Pie charts indicating the
fraction of thiamethoxam induced differentially expressed genes according the ORF length for up- (left) and
down- (right) regulated genes. oxam (arrow in Fig. 3) bees generally survived for 48 h. In contrast, presence of either B. badius or O. anthropi
together with thiamethoxam killed almost all bees within 48 h. Using data on each bacterial species separately, along with replicates without bacterial exposure, showed
that bee viability was negatively affected by increasing doses of thiamethoxam and of bacteria (Table 1: 2-way
logistic ANOVAs). At 48 h, a synergistic interaction between these main effects was detected when the bacterial
species was B. badius: bee viability declined more rapidly in response to bacterial dose when bees were exposed
to higher doses of thiamethoxam (Table 1, Fig. Results
Ch
i t 3).ff g
g
To test for differences between the effects of the two bacterial species, replicates without bacterial exposure
were excluded and the effects of species, non-zero bacterial dose, thiamethoxam and their interactions were
explored (Table 1: 3-way logistic ANOVAs). At 24 h, bee viability declined with both thiamethoxam dose and
bacterial dose and was lower when the bacterium applied was O. anthropi, without interactions (Table 1). At
48 h, the same dosage effects were found but the species effect was not significant (Table 1). There was, however,
a marginally non-significant interaction between bacterial species and bacterial dose: as probability estimates
from logistic analyses are inexact, we explored the consequences of retaining this interaction in the model. This generated a further significant interaction between thiamethoxam dose and the species of bacteria applied
(F1,21 = 5.85, p = 0.025): O. anthropi affected bees more negatively than did B. badius as pesticide dose increased. We conclude that thiamethoxam suppresses the abilities of bees to cope with natural challenges of the immune
system which would normally not be fatal f
We conclude that thiamethoxam suppresses the abilities of bees to cope with natural challenges of the immune
system, which would normally not be fatal. Discussion Our study addresses the effect of thiamethoxam on gene expression at field relevant doses of thiamethoxam found
in pollen and nectar. In particular we chose for these experiments a low and a high dose, which is about 0.04 ng
and 1.1 ng per bee per day for a 10 day chronic long-term exposure. A number of studies have used thiamethoxam
doses in the same range for analysing changes in gene expression or observing an impact on behaviour following
short term exposure18,20–24,69,70. In fact, it was found that chronic low-dose long-term thiamethoxam exposure
altered bee activity, motor function and movement to light24, but to our knowledge no previous analysis of gene
expression under such conditions has been done.i A key finding of our analysis of honey bee transcriptomes is the highly enriched fraction of dose-responsive,
uncharacterised genes encoding short open reading frames (sORFs). Such sORFs have only recently been rec-
ognized to encode functional peptides71–73, some of which play important roles during development or have a
function in the immune system74,75. Intriguingly, the majority of the dose-responsive sORFs we have identified https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 | www.nature.com/scientificreports/ Figure 3. Thiamethoxam exposure makes bees vulnerable to infection by Bacillus badius and Ochrobactrum
anthropi. Viability of groups of 8–12 bees 24 h (grey bars) and 48 h (black bars) after injection with B. badius
(A) or O. anthropi (B) (2–8 × 106 and diluted 1:10) alone or together with thiamethoxam (at a range of doses,
µM) shown as means and standard errors of three replicates. Arrows indicate injection of a 1 µM thiamethoxam
solution with (A right and B) or without bacteria (A left). Statistical significance is indicated by **(p < 0.01) and
***(p < 0.001). Figure 3. Thiamethoxam exposure makes bees vulnerable to infection by Bacillus badius and Ochrobactrum
anthropi. Viability of groups of 8–12 bees 24 h (grey bars) and 48 h (black bars) after injection with B. badius
(A) or O. anthropi (B) (2–8 × 106 and diluted 1:10) alone or together with thiamethoxam (at a range of doses,
µM) shown as means and standard errors of three replicates. Arrows indicate injection of a 1 µM thiamethoxam
solution with (A right and B) or without bacteria (A left). Statistical significance is indicated by **(p < 0.01) and
***(p < 0.001). are predicted to encode peptides with antimicrobial function (anti-microbial peptides, AMPs). Discussion The sORFs we
identified have not been reported in prior whole-transcriptome evaluations of neonicotinoid exposure in bee
brains despite using similar concentrations of neonicotinoids18,69,70. A possible explanation for the different gene
set we obtained after thiamethoxan exposure is that we analysed changes after long-term, low dose exposure,
while prior studies used shorter exposures.hif p
p
The finding that mostly sORFs, and among them many predicted AMPs are differentially expressed points
to the immune system for being dysregulated by neonicotinoids. The best-characterized AMPs are expressed
in the fat body upon pathogen exposure, but it is now recognized that glial cells in the brain also exert immune
functions and express AMPs37,39,40. Accordingly, the sORFs that we found to be differentially expressed upon
chronic low dose thiamethoxam exposure might form a basal immune defence, similar to the antimicrobial
environment present in saliva in vertebrates which contains many AMPs. y
Glia are support cells of neurons and are protective in a number of ways. Dysregulated AMP’s in the brain
have been linked to neurodegenerative disease37,38,40. Hence, the dysregulation of sORFs in response to thia-
methoxam exposure could indicate disturbed communication between these two cell types beyond immune
functions. Likewise, antimicrobial peptides have also been identified in having a role in learning and memory
in Drosophila, but the molecular mechanism how AMPs enhance learning and memory remains unknown76. p
g
y
We noted that other studies found an overrepresentation of down-regulated genes with known function
in immune and defence processes upon exposure to different types of neonicotinoids18,26,69,70,77. In contrast
we did not find differentially regulated immune genes including the known AMPs in the brain. AMPs are
highly expressed in the fat body. For the analysis of gene expression in bee brains we dissected them from cold-
anaesthetized bees and ensured that all fat body surrounding the brain was removed. It is possible, that freezing Scientific Reports | (2021) 11:1489 | https://doi.org/10.1038/s41598-020-80620-7 www.nature.com/scientificreports/ Source
Survival after 24 h
Survival after 48 h
df
Deviance
F
P
df
Deviance
F
P
2-way logistic ANOVAs
B. badius dose
1
60.69
59.26
< 0.001
1
74.64
50.04
< 0.001
Thiamethoxam dose
1
73.10
71.38
< 0.001
1
49.92
33.47
< 0.001
Bacteria × thiamethoxam
1
0.55
0.54
0.472
1
36.79
24.66
< 0.001
Residual
17
17.41
17
25.36
Total
20
121.44
20
186.72
O. Discussion anthropi dose
1
55.42
18.99
< 0.001
1
76.28
1.76
0.003
Thiamethoxam dose
1
59.99
20.55
< 0.001
1
44.51
11.12
< 0.001
Bacteria × thiamethoxam
1
0.09
0.03
0.865
1
7.06
19.06
0.199
Residual
20
58.38
20
80.04
Total
23
156.51
23
191.16
3-way logistic ANOVAs
Bacterial species
1
15.41
0.021
1
6.21
0.25
0.622
Bacterial dose
1
32.67
0.002
1
14.87
6.28
0.021
Thiamethoxam dose
1
90.19
< 0.001
1
85.56
36.12
< 0.001
Bacterial species × bacteraial dose
1
3.01
0.281
1
8.73
3.68
0.069
Bacterial species × thiamethoxam
1
0.13
0.204
1
6.21
2.62
0.121
Bacterial dose × thiamethoxam
1
4.23
0.824
1
2.56
1.08
0.311
Residual
20
49.08
20
47.39
Total
26
155.30
26
151.08 Table 1. Effects of thiamethoxam dose, bacterial dose and species on bee viability. Table 1. Effects of thiamethoxam dose, bacterial dose and species on bee viability. of brains prior to dissection69 or residual fat body adhering to the brain contributed considerably to a different
profile of immune gene expression in other studies. of brains prior to dissection69 or residual fat body adhering to the brain contributed considerably to a different
profile of immune gene expression in other studies. i
Immunosuppression has also observed been upon exposure to neonicotinoids co-infecting bees with viruses
of Nosema, a unicellular parasite of bees. In particular, this affected the cellular response, but also expression of
AMPs18,26,69,70,77. Since we did not detect differential expression of known immune genes, the bees in our study
were not infected with these known pathogens, but we cannot exclude an impact of other microbiota that are
non-pathogenic in healthy bees. Moreover, our analysis focused on gene expression in the brain, where glial
cells govern immune functions, which is a different humoral response than mediated by fatbody cells secreting
AMPs into the hemolymph. y
p
Bees, including Apis mellifera, are characterised by their limited set of canonical immune genes, compared
to non-social insects, such as the fruit fly D. melanogaster78–80. Currently, only six antimicrobial-peptide genes
comprising four gene families have been described in honey bees80. In contrast, Drosophila has 20 antimicrobial-
peptide genes comprising eight gene families29,80. From these genes, only defensin is conserved between honey
bees and Drosophila, consistent with the idea that antimicrobial peptides evolve fast to adapt to species-specific
environmental conditions34. Received: 5 March 2020; Accepted: 23 December 2020 Table 1. Effects of thiamethoxam dose, bacterial dose and species on bee viability. Discussion Given the low number of known antimicrobial peptides in bees, it is conceivable
that new (currently uncharacterised) genes encoding antimicrobial peptides are evolving. Consistent with this
idea, some of the sORFs differentially regulated by thiamethoxam are bee or hymenopteran specific. f
y
g
y
y
p
pi
Various agrochemicals, have been shown to alter the gut microbiome81,82. Our results are consistent with
previous findings where neonicotinoid exposure adversely affects insect immunity82,83. Specifically, we have
shown that immune challenges from what are normally non-pathogenic bacteria become fatal to bees when
combined with thiamethoxam exposure. How foreign bacteria enter the body of bees appears to be crucial. For
example, Dickel and colleagues found that oral ingestion of Enterococcus faecalis bacteria, increases the survival
neonicotinoid-exposed, caged worker bees84. In our study, the bacteria were injected between segments, which
mimics punctures sustained by attacks of the Varroa mite. We note that pathogens can enter bee haemolymph
through punctures inflicted by Varroa destructor mites65 and thus there may be considerable mortality within
hives infested with Varroa and also exposed to thiamethoxam. p
In summary, the most prominent changes in gene expression upon long-term low-dose thiamethoxam expo-
sure identified mostly genes that encode short ORFs, around half of which are predicted to code for antimicrobial
peptides. Furthermore, thiamethoxam exposure reduced the capacity of bees to withstand microbial infection. Taken together, these findings imply that low doses of neonicotinoids may be intrinsically sub-lethal to bees but
can be ultimately fatal via a weakened immune response to extrinsic pathogens. The roles of the identified genes
in the immune response of bees will need to be identified to establish how bee immunity might be strengthened
to resist bacterial infections. Received: 5 March 2020; Accepted: 23 December 2020 Scientific Reports | (2021) 11:1489 | https://doi.org/10.1038/s41598-020-80620-7 www.nature.com/scientificreports/ References Brandt, A., Gorenflo, A., Siede, R., Meixner, M. & Büchler, R. The neonicotinoids thiacloprid, imidacloprid, and clothianidin affect
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PD PU
d K D
f
d Author contributions
P.D., P.U. and K.D. performed the experiments, T.C.R, I.C.W.H., O.M., N.M. analysed data, R.S. and M.S. super-
vised experiments and analysed data, R.S. and M.S. wrote the manuscript with help from P.D., P.U. and I.C.W.H. P.D., P.U. and K.D. performed the experiments, T.C.R, I.C.W.H., O.M., N.M. analysed data, R.S. and M.S. super-
vised experiments and analysed data, R.S. and M.S. wrote the manuscript with help from P.D., P.U. and I.C.W.H. Competing interests The authors declare no competing interests.Additional information
Supplementary information The online version contains supplementary materials available at https://doi. org/10.1038/s41598-020-80620-7. Correspondence and requests for materials should be addressed to R.S. or M.S. Reprints and permissions information is available at www.nature.com/reprints. Acknowledgements We thank N. Parker and the Winterbourne garden for bees, N. Parker for bee suits, D. Scocchia, V. Soller-
Haussmann and K. Nallasivan for help with bee collections. G. Salmond for bacterial strains, D. Scocchia for
help with bacterial culturing and genotyping, and L. Orsini, E. Davies and I. Haussmann for comments on the
manuscript. For this work we acknowledge funding from the Foundation for Research Support of the State of São
Paulo, FAPESP (2012/13370-8; 2013/07251-9; 2014/23197-7; 2015/22368-5), the Biotechnology and Biological
Sciences Research Council (BBSRC), the Nottingham-Birmingham Fund, and the Sukran Sinan Memory Fund. https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 | www.nature.com/scientificreports/ Reprints and permissions information is available at www.nature.com/reprints. Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and
institutional affiliations. Open Access This article is licensed under a Creative Commons Attribution 4.0 International
License, which permits use, sharing, adaptation, distribution and reproduction in any medium or
format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the
Creative Commons licence, and indicate if changes were made. The images or other third party material in this
article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the
material. If material is not included in the article’s Creative Commons licence and your intended use is not
permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from
the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. © The Author(s) 2021 https://doi.org/10.1038/s41598-020-80620-7 Scientific Reports | (2021) 11:1489 |
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Comparison of obturation quality in natural and replica teeth root-filled using different sealers and techniques
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Clinical oral investigations
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Abstract Objectives This study aims to assess the obturation efficacy of sealers placed with different techniques using microcomputed
tomography (µCT) and assess the influence of µCT testing parameters on the obturation data obtained. l
Materials and methods Incisors and mesial roots of lower molars with standardized root length were scanned using µCT,
and one tooth of each type was 3D printed in acrylic. Two obturation techniques (warm vertical and single cone) and 4
sealer types (AH Plus, BioRoot RCS, Totalfill BC, and Bio-C Sealers) were assessed following storage in Hank’s balanced
salt solution for 3 and 6 months by assessing gap and void volume percentages on both natural and replica incisor and molar
roots. The storage solution was analysed to assess calcium ion leaching. The influence of temperature, tooth positioning,
and moisture content of the teeth while µCT scanning was also investigated. Results The obturation quality in the incisor group was the same using both natural teeth and replicas (p > 0.05). No changes
in void volume were identified when comparing the same sealer using different obturation techniques. The premixed seal-
ers used in single-cone obturation exhibited high void volume in the 3D printed replicas in the long term. The temperature,
positioning, and moisture content of the teeth did not affect the outcome of µCT testing.i f
Conclusions BioRoot RCS, Totalfill BC, and Bio-C Sealers are suitable for obturation of both complex and simple root canal
systems using different obturation techniques with BioRoot RCS exhibiting the highest calcium ion release. 3D printed acrylic
teeth can be used to assess the obturation quality in uncomplicated root canal systems. µCT parameters had no significant
effect on the µCT measurement. f
Clinical relevance The single-cone obturation technique with hydraulic sealer is a simple technique that can be used for
obturation of all root canal systems. f
Clinical relevance The single-cone obturation technique with hydraulic sealer is a simple techniq
obturation of all root canal systems. Keywords Root canal filling quality · Microcomputed tomography · Calcium silicate–based sealer · Obturation technique ·
3D printing Clinical Oral Investigations (2023) 27:2407–2417
https://doi.org/10.1007/s00784-023-04884-9 Clinical Oral Investigations (2023) 27:2407–2417
https://doi.org/10.1007/s00784-023-04884-9 RESEARCH RESEARCH RESEARCH Comparison of obturation quality in natural and replica teeth
root‑filled using different sealers and techniques Chuta Kooanantkul1 · Richard M Shelton1 · Josette Camilleri1 Received: 14 November 2022 / Accepted: 26 January 2023 / Published online: 4 February 2023
© The Author(s) 2023 Received: 14 November 2022 / Accepted: 26 January 2023 / Published online: 4 February 2023
© The Author(s) 2023 1
School of Dentistry, Institute of Clinical Sciences, College
of Medical and Dental Sciences, University of Birmingham,
5, Mill Pool Way, Edgbaston, Birmingham B5 7EG, UK Sealer characterisation sealants are all primarily composed of tricalcium silicate with
a radiopacifier and additives to enhance the material properties
[9]. These sealers exhibit higher release of calcium ions and
demonstrate higher flow rates, higher pH, and lower cytotox-
icity than AH Plus [10–12]. Chemical and micromechanical
bonds have also been identified with the dentine [1, 13]. The
hydrophilicity of the hydraulic sealers has led to some con-
cerns regarding adhesion to the gutta-percha cones. A biocer-
amic coated gutta-percha (GP) cone (BC cone) was introduced
(Brassler, Georgia, USA) to address this issue where the GP
was coated and impregnated with calcium silicate nanopar-
ticles aiming to cause interaction with a hydraulic calcium
silicate sealer and form an actual gap-free seal (monoblock
system). To date, relatively few studies have evaluated this type
of GP in terms of filling quality and ion leaching.i sealants are all primarily composed of tricalcium silicate with
a radiopacifier and additives to enhance the material properties
[9]. These sealers exhibit higher release of calcium ions and
demonstrate higher flow rates, higher pH, and lower cytotox-
icity than AH Plus [10–12]. Chemical and micromechanical
bonds have also been identified with the dentine [1, 13]. The
hydrophilicity of the hydraulic sealers has led to some con-
cerns regarding adhesion to the gutta-percha cones. A biocer-
amic coated gutta-percha (GP) cone (BC cone) was introduced
(Brassler, Georgia, USA) to address this issue where the GP
was coated and impregnated with calcium silicate nanopar-
ticles aiming to cause interaction with a hydraulic calcium
silicate sealer and form an actual gap-free seal (monoblock
system). To date, relatively few studies have evaluated this type
of GP in terms of filling quality and ion leaching.i All sealers were characterised to assess the material micro-
structure and elemental composition using scanning electron
microscopy (SEM; EVO MA10, Carl Zeiss, Cambridge, UK)
and energy-dispersive X-ray spectroscopy (EDS; EVO MA10,
Carl Zeiss, Cambridge, UK). Sealers were injected into cir-
cular rubber moulds 10 mm internal diameter and 2 mm high
and placed on a glass mixing slab kept at 37 °C and 100%
humidity until the surface was resistant to indentation. After setting, each sealer was removed from the rubber
mould and embedded in epoxy resin (EpoFix, Struers, Ballerup,
Denmark). Resin blocks were placed in a fume cupboard at
room temperature for 1 day to allow complete polymerisation. Sealer characterisation After setting, the sample surfaces were ground using 220, 500,
and 1200 grit diamond discs (MD-Piano, Struers ApS, Den-
mark) with water coolant for 1 min each, using an automatic
grinding and polishing machine (Phoenix Beta, Buehler, Lake
Bluff, IL, USA). Polishing was completed with cloth discs (MD
Largo, Dac, Nap, Struers Aps, Ballerup, Denmark) using 9,
3, and 1 µm diamond impregnated polishing liquids (DiaPro,
Struers ApS, Ballerup, Denmark) each for 3 min. All specimens
were mounted on aluminium stubs with double-sided carbon
tape. Each sample was sputter-coated with gold (EMITECH
K550X; Carl Zeiss, UK; needle value adjustment was set to
give 0.1 mbar of argon gas at a nominal 0.3 bar, deposition cur-
rent of 25 mA and 2 min deposition time) before examination
using SEM and EDS for elemental analysis (working distance
8.5 mm, I Probe 1,000 pA and accelerating voltage 20 kV). The
type and weight fraction of the elements present were recorded
from EDS using an area analysis. i
Although no link has been definitively demonstrated
between root canal obturation quality and clinical success
rates, sealing ability has always been used to assess and
compare obturation techniques as a surrogate marker. Dyes,
glucose, protein, or bacterial penetration from coronal to
apical through the obturated root canal and the time taken
has several limitations in assessment of sealing ability. Dyes
and bacterial penetration vary according to the type of dye
and bacteria species [14]. Another method for determining
obturation quality is the use of microcomputed tomography
(µCT) to assess percentage of gaps and voids (%Vgv). µCT
can be used to scan and reconstruct the sample in 3-dime-
sions at µm resolution in a non-invasive manner without
causing sample damage [15–17]. It also shows the volume,
location, and size of gaps (space between the material and
root) and voids (space inside the material). Comparison of
data is still challenging since the natural teeth used for these
measurements have variable anatomy and shapes. Such vari-
ations could be overcome by using standardised 3D printed
teeth with the morphology copied from natural teeth. Preparation of teeth and resin replicas The use of human teeth for research was approved by the
Research and Innovation Department, Birmingham Com-
munity Healthcare Trust (14/EM/1128). Teeth with fully
formed apices and root curvatures less than 30° were
included in the study, whilst teeth with root caries or resto-
rations or root fractures were excluded. Remnants of peri-
odontal ligament (PDL) and bone were gently removed
using a Gracey curette (SG11/12, Hu-Friedy, Chicago, IL,
USA). The crowns were removed using a diamond bur,
and the roots of the upper incisors were standardized to 16
mm in length, while the mesio-buccal roots were sectioned
from lower molars and standardized to 12 mm in length. The objectives of this study were to assess the obtura-
tion quality of sealers used with different techniques and the
validity of using 3D printed teeth for assessment, determined
using µCT. Furthermore, the influence of varying the µCT
testing parameters on the data generated were also examined. Introduction Tulsa, OK, USA) is most commonly used in both techniques due
to its low shrinkage (1.76%), low solubility compared with other
resin-based sealers, and low film thickness complying with the
ISO requirement for root canal sealing materials [1]. However,
AH Plus does not fulfil Grossman’s ideal root canal sealer prop-
erties [2] as it has no bactericidal effect and shrinks when set [3]. Root canal obturation is necessary to prevent reinfection of the
root canal space in the long term. Warm vertically compacted
and laterally condensed gutta-percha techniques have been used
to hermetically fill the root canal using a combination of gutta-
percha and root canal sealant. AH Plus (Dentsply Maillefer, f
More recently, the single cone/sealer-based technique has
become more popular among clinicians due to its simplicity
and not requiring specific armamentarium [4]. This technique
is used with hydraulic calcium silicate cements (HCSCs) as
these sealers have shown antimicrobial characteristics [5–8]
and generation of an appropriate seal [3], thus indicating
suitability for sealer-based techniques. There are a number
of hydraulic cement sealers available clinically which have
similar chemistries but different modes of presentation. These * Josette Camilleri
J.Camilleri@bham.ac.uk
Richard M Shelton
R.M.Shelton@bham.ac.uk
1
School of Dentistry, Institute of Clinical Sciences, College
of Medical and Dental Sciences, University of Birmingham,
5, Mill Pool Way, Edgbaston, Birmingham B5 7EG, UK (0121 3456789)
3 3456789)
3 Clinical Oral Investigations (2023) 27:2407–2417 2408 • AH Plus (Dentsply DeTrey, Konstanz, Germany)
• BioRoot RCS (Septodont, Saint-Maur-des-Fosses,
France)
• Totalfill BC Sealer (FKG, Chaux de Fonds, Switzerland)
• Bio-C Sealer (Angelus, Londrina, Brazil) • Totalfill BC Sealer (FKG, Chaux de Fonds, Switzerland) i
• Bio-C Sealer (Angelus, Londrina, Brazil) • BioRoot RCS (Septodont, Saint-Maur-des-Fosses,
France)i • AH Plus (Dentsply DeTrey, Konstanz, Germany) Materials and methods The following sealers were used with two obturation meth-
ods and tested in human teeth and in replicas. • AH Plus (Dentsply DeTrey, Konstanz, Germany) The root canal therapy was all undertaken by the same
operator, a specialist endodontist. The root canals were
prepared using ProTaper Gold (Dentsply Maillefer, Bal-
laigues, Switzerland) using SX, S1, S2, F1, and F2 series
for the molar roots and to F3 for the incisors and copious • BioRoot RCS (Septodont, Saint-Maur-des-Fosses,
France)i 3 Clinical Oral Investigations (2023) 27:2407–2417 2409 obturated using either a single cone/sealer-based technique
or warm vertical compaction (WVC) with the test sealers. Master cones (ProTaper Gold) F3 and F2 were fitted into the
incisors and molar roots respectively. An additional group
was added where Totalfill BC sealer was used. Besides
using the standard gutta-percha master cone, a bioceramic
coated (BC) gutta-percha (Totalfill) was used in conjunc-
tion with the sealer-based technique. For WVC, the down
packing stage was performed using a presized heat carrier
(EQ-V, Meta Biomed, Chungcheongbuk-do, Republic of
Korea) that precisely fitted in the canal at 3–5 mm from WL,
while the back-filling stage was performed using thermo-
plasicized syringeable gutta-percha (EQ-V, Meta Biomed,
Chungcheongbuk-do, Republic of Korea). The coronal ori-
fices were restored with glass ionomer cement (ChemFil®
Superior, Dentsply DeTrey, Konstanz, Germany). Five roots
per technique and tooth type were used. After completion
of obturation, all teeth were immersed in 1.5 mL of HBSS
at 37 °C. irrigation with 5.25% sodium hypochlorite (NaOCl; Chlo-
raxid, Cerkamed, Stalowa, Wola, Poland) between each
instrumentation. After completing mechanical instru-
mentation, 3 mL 17% ethylenediamine tetraacetic acid
(EDTA; Cerkamed, Stalowa, Wola, Poland) was used to
rinse the canal for 1 min, followed by 3 mL 5.25% NaOCl
for 1 min. To make the 3D printed acrylic replicas, a number of
upper incisors and lower molars were scanned using µCT
(Bruker Skyscan MicroCT model 1172, Billerica, MA,
USA) from apex to crown at 11 µm intervals, using an
accelerating voltage of 70 kV at 140 mA and medium
camera pixels (2 K × 1 K). These parameters were used
for all µCT scans throughout the experiment. The scanned
images were reconstructed using the NRecon software
(Bruker, Billerica, MA, USA) and saved in .tiff file for-
mat. An incisor root with a straight circular continuously
tapered canal from the coronal to the apical part of the
root (Fig. Preliminary study A preliminary study was designed to determine how fluc-
tuations in machine temperature, tooth positioning, and the
moisture conditions of the specimens could have affected the
µCT measurements. The experiments were carried out on
human single-rooted teeth that were prepared as described
above and also on the replicas of the incisor roots. Gap and void volume measurement In order to establish the correct parameters to test the per-
centage of gaps and voids (%Vgv), a preliminary study was
undertaken to establish whether the experimental set-up
influenced the data generated. Materials and methods 1A) was selected as a simple canal morphol-
ogy tooth. A molar with a mesial root having two con-
tinuously tapered canals from the coronal to the apical
part of the root, two straight separate apical foramina,
and a continuous isthmus along the root length (Fig. 1B)
was selected for complex root canal anatomy. 3D root
canals and dentine images of these teeth were created
using CTAn software (Bruker) and were saved in the .stl
file format. The selected digital files were then printed
in resin (RepliDens®, Smartodont, Zurich, Switzerland),
thus providing one round uncomplicated canal and a com-
plex root canal system for obturation (Fig. 1C and D). Root canal obturation The prepared natural roots and the replicas of both the sin-
gle-rooted teeth and the mesial roots of the lower molar were Fig. 1 Digital and 3D printed acrylic replicas; A digital model of the incisor, B digital model of the molar, C printed incisor, and D printed
molar. Note the similarity of the root canal morphology of polymer teeth compared with digital models Fig. 1 Digital and 3D printed acrylic replicas; A digital model of the incisor, B digital model of the molar, C printed incisor, and D printed
molar. Note the similarity of the root canal morphology of polymer teeth compared with digital models 1 3 1 3 Clinical Oral Investigations (2023) 27:2407–2417 2410 To test the effect of the µCT temperature on the readings,
the machine was started and was kept on for 3 h. A thermo-
couple device (data logger model TC-08, Pico® technology,
Saint Neots, UK) was used to measure the temperature inside
the µCT machine by continuously recording it every second
for 5 h. The µCT machine was operated in either an inter-
rupted or continuous scan mode. For the interrupted scan
mode, the X-ray beam was stopped every 53 min, rested for
1 min, and restarted with the method repeated five times to
represent the actual working time for tooth scanning. A con-
tinuous scan mode was also employed where the X-ray beam
was used continuously until the scan was completed. The
initial, maximum, and average working temperatures were
recorded by Picolog version 6.1.14 software (Pico® tech-
nology, Saint Neots, UK). The temperature of the machine
chamber when operated in these modes was recorded. Five
single-rooted of replicas were stored at the average operated
machine temperature and were then scanned. Furthermore,
these replicas were stored at room temperature, scanned, and
then placed at 37 °C in an incubator until reaching a stable
temperature and were rescanned. All teeth were scanned five
times for each temperature and scanning mode used. All
the scanned images were reconstructed using the NRecon
software (ring artefact reduction = 13, beam-hardening cor-
rection = 20% and dynamic image range from 0.001 to 0.05)
and saved in.tiff file format. where canal volume = gap volume +void volume + filling volume. The teeth were assessed immediately after obturation
and again after a further 3 and 6 months. To simulate the
clinical environment, a simulated periodontal ligament was
painted on the natural roots. This consisted of a layer of
0.1–0.4 mm thickness of collagen-agarose gel containing 1
mg/mL of collagen (FibriCol®, Advanced BioMatrix, San
Diego, CA, USA) and 1% w/v agarose (Bioline, London,
UK) made up in HBSS (Sigma-Aldrich, Gillingham, UK)
and the pH adjusted to 7.4. The teeth were then immersed
in 1.5 mL HBSS and stored in an incubator at 37 °C for 3
or 6 months. The replicas were also immersed in HBSS but
without any surface modification. After 3 and 6 months,
the coating was gently removed using wet gauze; the teeth
rescanned, reconstructed, and 3D-segmented using the same
methods as before to enable calculation of %Vgv. The tooth
was recoated with collagen/agarose gel and re-immersed in
HBSS in an incubator at 37 °C. The %Vgv data obtained
immediately after obturation was compared with 3 and 6
months data. To evaluate the effect of tooth positioning within the µCT
chamber on the data generated, replicas were either fixed to
a silicone putty that allowed reorientation after every scan or
were just placed in the µCT machine in random positions. Each
tooth was scanned using the same parameters as for the tem-
perature assessment. All scanned images were reconstructed
using the NRecon software. Tooth position was adjusted with a
3D-segmentation method, where the first scanned tooth image
was imported as a reference, whilst another tooth image was
imported as the target image. The target image was adjusted until
it completely overlapped the reference image before exporting
the adjusted image in a .tiff file format. The root canal volume
of the initial position, altered position, and software-adjusted
position were calculated using ImageJ.f Statistical analysis To assess the effect of dehydration, both replicas and
human teeth were used. These were scanned after storage
in water and therefore fully saturated, storage at room tem-
perature and also in a fully dehydrated state by storing in a
desiccator with silica gel. All statistics were performed using the SPSS software version
23 (IBM, North Castle, NY, USA). A Kolmogorov−Smirnov
test was used to test the normality of the data. Intrarater reli-
ability was calculated by using intraclass correlation coef-
ficient. The Wilcoxon signed-rank test was used to test the
effect of the temperature of the tooth on the precision of µCT. The Friedman test was used to test the effect of the mois-
ture and the position of the tooth on the precision of µCT. The mean canal volumes between each group in the same Root canal obturation All reconstructed images were
imported into the Dataviewer software (Bruker) to 3D-regis-
ter by overlapping two images from different scans together,
and the root canal volume was then calculated.f parameters described for the scans of the teeth used to make
the replicas. %Vgv were assessed with the CTAn software by
determining the thresholding range for each type of sample
which varied depending on the tooth type and the obturation
method. The region of interest (ROI) was identified, follow-
ing the outline of the gap and void area. The total %Vgv was
calculated and compared between each group using these
equations: Leachate analysis After 3 and 6 months, 1 mL HBSS were collected and
diluted 10-fold and leachate analysis was performed to
measure calcium ion leaching using inductively coupled
plasma mass spectroscopy (Optima 8000; Perkin Elmer,
Waltham, MA, USA). %Vgv = gap volume + void volume
canal volume
× 100 where canal volume = gap volume +void volume + filling volume. where canal volume = gap volume +void volume + filling volume. Measurement of %Vgv of test specimens The teeth and replicas for both the incisors and molars were
scanned pre- and post-obturation with µCT using the same 1 3 1 3 Clinical Oral Investigations (2023) 27:2407–2417 2411 Table 1 Semi-quantitative elemental composition of the test sealers
Sealer
Element (mean % weight fraction)
Ca
Si
Zr
W
Al
AH Plus
2.5 ± 0.5 BDL
10.8 ± 0.5 12.7 ± 2.4 BDL
BioRoot
RCS
23.8 ± 2.9 6.5 ± 0.7
17.0 ± 1.5 BDL
BDL
Totalfill
BC
9.5 ± 0.8 2.9 ± 0.2
24.3 ± 2.0 BDL
BDL
Bio-C
Sealer
4.3 ± 2.7 1.2 ± 0.6
17.3 ± 5.7 BDL
0.5 ± 0.2 tooth type were compared using one-way repeated analysis
of variance (ANOVA) test and post hoc analysis. %Vgv at each
period (post-obturation, 3 months, and 6 months) were com-
pared between different sealers using the Kruskal−Wallis test
and Dunn−Bonferroni analysis. %Vgv from the same sealer
group at the different periods were compared between each
sealer using the Friedman test and Dunn-Bonferroni analysis. Discussion %Vgv for each tooth type in both natural teeth and 3D printed
acrylic replicas is shown in Fig. 4. The mean empty root
canal volume was the same for all teeth in a specific tooth
type and also did not vary between the natural teeth and the
3D printed acrylic replicas (p > 0.05). The %Vgv of obturated
human teeth was the same as that of the replicas in the
incisor group (p = 0.159) but not in the molars (p < 0.001)
indicating that acrylic replicas could be used to investigate
obturation quality in teeth with simple canal anatomy but not
in molar teeth with more complex anatomy. The current study investigated the assessment of quality of
obturation using microcomputed tomography to measure
%Vgv of complex and simple root canal systems obturated
using either WVC or single-cone/sealer-based techniques
and four different sealers. The sealers investigated in the
current study included AH Plus, an epoxy-resin-based
sealer which was used as the control as it is widely used
and well established. The test sealers included a water-based
hydraulic cement sealer (BioRoot RCS), and two syringe-
able hydraulic cement sealers that were formulated using a
non-aqueous vehicle. These materials were selected to assess
their interaction with the dentine substrate and whether it
was possible to eliminate the voids and gaps within the obtu-
ration. All the sealers used in the study were characterized to
establish their chemistry and microstructure. Material char-
acterization is key to be able to predict and adequately assess
the clinical behaviour of the sealers under investigation. The quality of obturation in natural teeth was not dependent
on the technique used (p > 0.05) but was affected by the type of
sealer. In both incisors and molar teeth obturated with a single
cone and AH Plus sealer, the %Vgv at 6 months were signifi-
cantly decreased compared with post-obturation (Fig. 4A, super-
script A, and Fig. 4B, superscript B; p < 0.05). All the other
sealers and techniques exhibited no significant changes over
the 6-month period. At 3 months, WVC obturations with AH
Plus and Totalfill BC had less voids than were identified with
the BioRoot RCS single cone/sealer-based technique (Fig. 4A,
superscripts a and b; p < 0.05). g
The advantage of measuring %Vgv using µCT allows
longitudinal testing of the obturation as the technique is
non-invasive unlike other techniques [17] that have been
employed to study leakage. Sealer characterisation The microstructure and the mean elemental composition of
each sealer are shown in Fig. 2 and Table 1, respectively. AH Plus included discrete particles (Fig. 2A) composed of
zirconium and tungsten embedded in a calcium-rich matrix
(calcium tungstate) as indicated in Table 1. All HCSC seal-
ers were composed of numerous calcium-rich particles with
zirconium and silicate particles dispersed within the cement matrix (calcium silicate) (Fig. 2B–D). Particle size varied
with AH Plus and BioRoot RCS having particles 2–10 µm. Totalfill BC particle size was approximately 4–15 µm and
Bio-C Sealer appeared the smallest with an average parti-
cle size of 1 µm. Among all sealers, BioRoot RCS had the
highest calcium level and AH Plus was the only sealer that
contained tungsten (Table 1). A
B
C
D
Fig. 2 Back scattered scanning electron micrographs of the four test sealers showing their microstructure; A AH Plus, B BioRoot RCS, C Total-
fill BC Sealer, and D Bio-C Sealer A D Fig. 2 Back scattered scanning electron micrographs of the four test sealers showing their microstructure; A AH Plus, B BioRoot RCS, C Total-
fill BC Sealer, and D Bio-C Sealer 1 3 2412 Clinical Oral Investigations (2023) 27:2407–2417 higher in the single cone/sealer-based technique compared
with the WVC in natural teeth (p < 0.05). This decreased
after 6 months for all HCSC sealers in both natural teeth and
replicas except for BioRoot RCS in the natural teeth where a
higher calcium ion release was noted. Bio-C sealer released
the lowest amount of calcium ions at each time point. The
Totalfill BC sealer used with the BC cone had a higher cal-
cium ion leaching than when used with a standard cone in
the incisor tooth and not in molars and replicas (p < 0.05). AH Plus released the least amount of calcium ions in every
tooth type (Table 2). This increased after 6 months in the
natural teeth and was reduced in the replicas. Preliminary study In the µCT continuous working mode, it took an hour for
the temperature to stabilize, while in the non-continuous
working mode, there were temperature fluctuations with
each scan (Fig. 3a). There was no significant difference
in root canal volume measured at different temperatures,
moisture, or position within the scanner (p > 0.05) as shown
in Fig. 3b. Discussion In the present study, both natural
teeth and replicas were used as natural teeth are inherently
variable [18–23] making the acquired data difficult to analyse
and interpret. The measurement of %Vgv of replicas generated
similar data as that from natural teeth in the incisor group but
not for the molars. This indicated the suitability of replicas
for assessment of quality of obturation in uncomplicated
root canal systems only. The use of replicas is a convenient
way to replace natural teeth as each have the same root canal
morphology making it easier to compare the data obtained
for the %Vgv assessment. In the 3D printed acrylic incisors, the obturation with
Bio-C Sealer and WVC deteriorated significantly over the
6-month period (Fig. 4C, superscript C; p < 0.05). In the
3D printed acrylic molars, the Totalfill BC WVC obturation
deteriorated at 3 months compared with post-obturation
(Fig. 4D, superscript D; p < 0.05). The higher sealer levels
for the premixed sealers (Bio-C Sealer and Totalfill BC
Sealer) used in single-cone obturation technique led to more
gaps and voids at 6 months (Fig. 4D, superscripts c-j; p <
0.05) and 3 months (Fig. 4D, superscript k and l; p < 0.05)
compared with the same sealers used in WVC. Leachate analysis g
The objectives of obturation materials are to serve as a
barrier to prevent the communication between oral cavity and
periapical area and to entomb any remaining microorganisms
within the root canal [24]. To date, several HCSC sealers have The leachate data is shown in Table 2 where BioRoot RCS
exhibited the highest release of calcium. The leaching was 1 3 Clinical Oral Investigations (2023) 27:2407–2417 2413 2413
23) 27:2407–2417
have been claimed to be stable and
[25, 26] but also have bactericidal
materials are thus suggested for use in
echniques with a single gutta-percha
ot to facilitate removal if necessary. volumes of sealer although having
bactericidal properties, in single matched cone obturation,
may have the disadvantage of less canal space being filled as
less effort is made to compress and manipulate the obturation
material into the complex root canal systems. In the current
study, single-rooted teeth and also mesial roots of lower
molars were used to assess whether the complexity of root
m-
-
he
d a
a-
y
T
l
er-
ure
0.00
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8.00
9.00
10.00
24ºC
37ºC
Inial
Modified
Adjusted
Fully
hydrated
tooth
Room
tooth
Desiccated
tooth
Fully
hydrated
replica
Desiccated
replica
Room
replica
Temperature
Posioning
Moisture
m
m
e
m
ulo
v
la
n
a
c
t
o
o
r
3 Fig. 3 a The µCT working tem-
perature readings from a non-
continuous scan. Red arrows
show the temperature when the
µCT chamber was opened and a
continuous scan. The tempera-
ture was stable approximately
50 min after initiated the µCT
scan. b Changes in root canal
volume as a function of temper-
ature, positioning, and moisture
condition of the tooth 0.00
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8.00
9.00
10.00
24ºC
37ºC
Inial
Modified
Adjusted
Fully
hydrated
tooth
Room
tooth
Desiccated
tooth
Fully
hydrated
replica
Desiccated
replica
Room
replica
Temperature
Posioning
Moisture
m
m
e
m
ulo
v
la
n
a
c
t
o
o
r
3 1 been introduced which have been claimed to be stable and
do not expand when set [25, 26] but also have bactericidal
properties [5–8]. These materials are thus suggested for use in
sealer-based obturation techniques with a single gutta-percha
point that is used as a pilot to facilitate removal if necessary. Leachate analysis 4 Mean percentage of gaps and voids of natural and 3D printed
replica teeth from post-obturation, 3 months and 6 months; A natural
incisor group, B natural molar group, C 3D replica incisor group, and
D 3D replica molar group (n = 5) (mean ± SD). (A, B, C, and D) Same letter means a significant difference (p ≤ 0.05) between each
period within the same sealer type and technique. (a, b, c, d, e, f, g,
h, i, j, k, and l) Same letter means a significant difference (p ≤ 0.05)
between each sealer type and obturation technique at the same period canal system influenced the obturation quality. There was no
significant difference in %Vgv between simple root canals and
complex root canals, which indicated the complexity of root
canal system was not a factor influencing obturation quality. Hydraulic cements release calcium ions which in turn
interact with the clinical environment. For sealers, the
interaction is with the root dentine [27] and the sealer
chemistry has been shown to affect the antimicrobial
properties [7]. EDS showed that all hydraulic cement
sealers were composed primarily of calcium and silicon
which is in agreement with previous studies [28–31]. The use of replicas can potentially impede penetration of
environmental moisture into the root canal thus effecting
the interaction of the sealers. The heat from WVC may also
cause evaporation of moisture within the root canal [32]. The
highest calcium ion content identified using EDS was found
in BioRoot RCS followed by Totalfill BC and Bio-C Sealer,
respectively. This reflected the leachate data that identified
BioRoot RCS, and Bio-C Sealer released the highest and the
lowest amount of calcium ions, respectively. The leaching canal system influenced the obturation quality. There was no
significant difference in %Vgv between simple root canals and
complex root canals, which indicated the complexity of root
canal system was not a factor influencing obturation quality. was higher in sealer-based obturation compared with the
WVC in natural teeth. The replicas showed lower calcium
ion leaching than was measured from natural teeth. This may
be explained by the interference of the hydration reaction
that the sealer would have with the dentine substrate. In this
respect, although no variation was found in %Vgv when using
resin replicas, clearly, the sealer hydration and interaction
with dentine were affected. Leachate analysis (a, b, c, d, e, f, g,
h, i, j, k, and l) Same letter means a significant difference (p ≤ 0.05)
between each sealer type and obturation technique at the same period 0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
Acrylic molar
post-obturaon
3 months
6 months
0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill Totalfill BC
cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
Molar tooth
post-obturaon
3 months
6 months
B
B
c,d
k e,f
D,c,g
d,h
D,k,l e,i
f,j
g,h l
i,j
B
D
Same letter means a significant difference (p ≤ 0.05) between each
period within the same sealer type and technique. (a, b, c, d, e, f, g,
h, i, j, k, and l) Same letter means a significant difference (p ≤ 0.05)
between each sealer type and obturation technique at the same period A
li
l
0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill Totalfill BC
cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
Molar tooth
post-obturaon
3 months
6 months
B
B
B Acrylic incisor
0
2
4
6
8
10
12
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
%
e
m
ulo
v
dio
v
d
n
a
p
a
g
Incisor tooth
post-obturaon
3 months
6 months
A
A
a,b
a
b
A A A 0
2
4
6
8
10
12
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
%
e
m
ulo
v
dio
v
d
n
a
p
a
g
post-obturaon
3 months
6 months
C
C
C 0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
post-obturaon
3 months
6 months
c,d
k e,f
D,c,g
d,h
D,k,l e,i
f,j
g,h l
i,j
D C Fig. Leachate analysis However, the use of high volumes of sealer although having bactericidal properties, in single matched cone obturation,
may have the disadvantage of less canal space being filled as
less effort is made to compress and manipulate the obturation
material into the complex root canal systems. In the current
study, single-rooted teeth and also mesial roots of lower
molars were used to assess whether the complexity of root 1 3 Clinical Oral Investigations (2023) 27:2407–2417 2414 0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
Acrylic molar
post-obturaon
3 months
6 months
0
2
4
6
8
10
12
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
%
e
m
ulo
v
dio
v
d
n
a
p
a
g
Acrylic incisor
post-obturaon
3 months
6 months
0
5
10
15
20
25
30
AH Plus
BioRoot
Totalfill Totalfill BC
cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
gap and void volume %
Molar tooth
post-obturaon
3 months
6 months
0
2
4
6
8
10
12
AH Plus
BioRoot
Totalfill
Totalfill
BC cone
Bio-C
AH Plus
BioRoot
Totalfill
Bio-C
single cone
WVC
%
e
m
ulo
v
dio
v
d
n
a
p
a
g
Incisor tooth
post-obturaon
3 months
6 months
A
A
a,b
a
b
B
B
C
C
c,d
k e,f
D,c,g
d,h
D,k,l e,i
f,j
g,h l
i,j
A
B
C
D
Fig. 4 Mean percentage of gaps and voids of natural and 3D printed
replica teeth from post-obturation, 3 months and 6 months; A natural
incisor group, B natural molar group, C 3D replica incisor group, and
D 3D replica molar group (n = 5) (mean ± SD). (A, B, C, and D)
Same letter means a significant difference (p ≤ 0.05) between each
period within the same sealer type and technique. Leachate analysis In the current study, AH Plus and Totalfill BC sealers 1 3 3 Clinical Oral Investigations (2023) 27:2407–2417 2415 Table 2 Calcium ion concentration in ppm leached from each tooth or replica using different obturation techniques and different sealers
Tooth type
Obturation technique
AH Plus
BioRoot RCS
Totalfill BC
Totalfill BC cone
Bio-C Sealer
3 months
6 months
3 months
6 months
3 months
6 months
3 months
6 months
3 months
6 months
Natural incisor
Single cone
0.0
8.3 ± 3.6
40.3 ± 1.7
99.0 ± 8.1
88.5 ± 6.6
33.2 ± 5.8
231.2 ± 3.3
12.7 ± 13.8
16.7 ± 5.6
5.6 ± 4.1
WVC
0.0
12.2 ± 9.2
19.8 ± 1.9
46.7 ± 6.3
7.1 ± 9.8
9.0 ± 11.5
/
/
7.6 ± 7.5
2.7 ± 2.6
Natural molar
Single cone
1.0 ± 0.2
11.4 ± 1.3
0.8 ± 0.7
159.0 ± 5.3
6.1 ±7.8
82.8 ± 27.7
8.2 ± 2.4
44.7 ± 4.9
13.2 ± 8.5
15.9 ± 15.3
WVC
1.4 ± 0.2
14.9 ± 5.4
1.1 ± 0.9
99.8 ± 26.1
28.7 ± 9.5
24.4 ± 9.9
/
/
0.3 ± 0.03
7.9 ± 4.8
Replica incisor
Single cone
66.5 ± 0.7
7.5 ± 0.2
99.3 ± 6.0
5.8 ± 2.1
87.2 ± 6.2
38.4 ± 2.1
86.8 ± 6.3
14.5 ± 3.6
79.9 ± 19.5
22.9 ± 12.3
WVC
56.0 ± 7.2
17.0 ± 8.7
104.4 ± 6.4
21.2 ± 10.8
77.4 ± 16.2
30.1 ± 2.2
/
/
85.2 ± 1.1
13.1 ± 6.3
Replica molar
Single cone
62.5 ± 4.1
8.6 ± 1.5
250.8 ± 7.8
6.2 ± 0.6
138.8 ± 7.9
6.9 ± 0.5
113.8 ± 22.35
6.1 ± 2.8
78.9 ± 6.5
11.3 ± 10.9
WVC
67.4 ± 7.2
5.7 ± 1.9
225.6 ± 5.4
5.9 ± 1.2
94.5 ± 13.8
4.6 ± 1.2
/
/
75.1 ± 2.5
47.5 ± 3.0 performed well when used with warm vertical obturating
technique. Thee Totalfill BC sealer has been shown to be
heat resistant [37], and this could account for the low gap
and void volume shown in this study.i High %Vgv was identified in most teeth when using Bio-
Root RCS, which may be related to its high viscosity and
manual mixing, where air could be entrapped inside the
material [38]; therefore, the material could contain voids
even before the obturation. Leachate analysis The leaching occurred despite
no significant variation in the %Vgv clearly showing the
distinction between the solubility which is a physical process
off material loss and ion leaching which is a chemical
process. l
Hydraulic cements release calcium ions which in turn
interact with the clinical environment. For sealers, the
interaction is with the root dentine [27] and the sealer
chemistry has been shown to affect the antimicrobial
properties [7]. EDS showed that all hydraulic cement
sealers were composed primarily of calcium and silicon
which is in agreement with previous studies [28–31]. The use of replicas can potentially impede penetration of
environmental moisture into the root canal thus effecting
the interaction of the sealers. The heat from WVC may also
cause evaporation of moisture within the root canal [32]. The
highest calcium ion content identified using EDS was found
in BioRoot RCS followed by Totalfill BC and Bio-C Sealer,
respectively. This reflected the leachate data that identified
BioRoot RCS, and Bio-C Sealer released the highest and the
lowest amount of calcium ions, respectively. The leaching The use of warm vertical compaction aims at having an
obturation that is composed primarily of gutta-percha. The
use of heat can negatively affect the sealer properties. AH
Plus can set faster and decompose when heated excessively
[32–34] although it has been shown that the temperature
inside the root canal does not rise to more than 60 °C [35]
and AH Plus will only deteriorate if heated indiscriminately
[36]. Leachate analysis Another sealer property that could contribute to higher
void volumes is the solubility. Although solubility has
been reported to be high for HCSC sealers [39, 40], this
was sown to depend on the environment the sealers were
placed in. Thus, solubility was shown to be reduced when
in contact with simulated tissue fluids [41, 42]. The AH
Plus was not susceptible to the solubility, and it exhibited
no voids after 6 months immersion. High void percentage was also found in the BC cone
group. BC cones have a taper of 4% which was smaller
than both the ProTaper Gold F2 and F3 (various taper of
8% and 9%, respectively). Hence, the amount of gutta-
percha was insufficient to spread the sealer into voids. This
finding contrasted with the sealer-based concept, where
the gutta-percha is used as a carrier and a path for retreat-
ment only. Conclusions HCSC sealers could be used to obturate both complex and
simple root canals using either WVC or single cone/sealer-
based techniques. BioRoot RCS with sealer-based technique
should be used when calcium ion release is clinically useful. 3D printed acrylic teeth can be used to assess the obturation
quality in uncomplicated root canal systems. Temperature,
position, and moisture of the tooth had no significant effect
on the µCT measurement. Author contribution CK: experimental work, drafting, and checking
manuscript; RMS: conceptualization, finalizing, and final checking of
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approval or consent to participate was necessary. Competing interests The authors declare no competing interests. 1 3 3 Clinical Oral Investigations (2023) 27:2407–2417 2416 16. Grande NM, Plotino G, Gambarini G, Testarelli L, D’Ambrosio
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bution 4.0 International License, which permits use, sharing, adapta-
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need to obtain permission directly from the copyright holder. To view a
copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. phology. Ann Ist Super Sanita. 48:26–34 17. Jung M, Lommel D, Klimek J (2005) The imaging of root canal
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Reproducibility of Glutamate, Glutathione, and GABA Measurements in vivo by Single-Voxel STEAM Magnetic Resonance Spectroscopy at 7-Tesla in Healthy Individuals
|
Frontiers in neuroscience
| 2,020
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cc-by
| 7,839
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ORIGINAL RESEARCH
published: 15 September 2020
doi: 10.3389/fnins.2020.566643 Ofer M. Gonen1,2,3*, Bradford A. Moffat2, Patrick Kwan1,2,3,4, Terence J. O’Brien1,2,3,4,
Patricia M. Desmond2,5 and Elaine Lui2,5 1 Department of Neurology, The Royal Melbourne Hospital, Parkville, VIC, Australia, 2 Department of Medicine and Radiology,
The University of Melbourne, Parkville, VIC, Australia, 3 Department of Neurology, The Alfred Hospital, Melbourne, VIC,
Australia, 4 Department of Neuroscience, Central Clinical School, Monash University, Melbourne, VIC, Australia, 5 Department
of Radiology, The Royal Melbourne Hospital, Parkville, VIC, Australia Edited by:
Xi-Nian Zuo,
Nanning Normal University, China Background and Purpose: Derangements in brain glutamate, glutathione, and
γ-amino butyric acid (GABA) are implicated in a range of neurological disorders. Reliable
methods to measure these compounds non-invasively in vivo are needed. We evaluated
the reproducibility of their measurements in brain regions involved in the default mode
network using quantitative MRS at 7-Tesla in healthy individuals. Reviewed by:
Yuzheng Hu,
Zhejiang University, China
Yan Li,
University of California,
San Francisco, United States Methods: Ten right-handed healthy volunteers underwent 7-Tesla MRI scans on 2
separate days, not more than 2 weeks apart. On each day two scanning sessions took
place, with a re-positioning break in between. High-resolution isotropic anatomical scans
were acquired prior to each scan, followed by single-voxel 1H-MRS using the STEAM
pulse sequence on an 8 mL midline cubic voxel, positioned over the posterior cingulate
and precuneus regions. Concentrations were corrected for partial-volume effects. *Correspondence:
Ofer M. Gonen
Ofer.Gonen@mh.org.au Specialty section:
This article was submitted to
Brain Imaging Methods,
a section of the journal
Frontiers in Neuroscience Specialty section:
This article was submitted to
Brain Imaging Methods,
a section of the journal
Frontiers in Neuroscience Results: Maximal Cramér-Rao lower bounds for glutamate, glutathione, and GABA
were 2.0, 8.0, and 14.0%, respectively. Mean coefficients of variation within sessions
were 5.9 ± 4.8%, 9.3 ± 7.6%, and 11.5 ± 8.8%, and between sessions were
4.6 ± 4.5%, 8.3 ± 5.7%, and 9.2 ± 8.7%, respectively. The mean (±SD)
Dice’s coefficient for voxel overlap was 90 ± 4% within sessions and 86 ± 7%
between sessions. Received: 28 May 2020
Accepted: 21 August 2020
Published: 15 September 2020 Reproducibility of Glutamate,
Glutathione, and GABA
Measurements in vivo by
Single-Voxel STEAM Magnetic
Resonance Spectroscopy at 7-Tesla
in Healthy Individuals Ofer M. Gonen1,2,3*, Bradford A. Moffat2, Patrick Kwan1,2,3,4, Terence J. O’Brien1,2,3,4,
Patricia M. Desmond2,5 and Elaine Lui2,5 Ofer M. Gonen1,2,3*, Bradford A. Moffat2, Patrick Kwan1,2,3,4, Terence J. O’Brien1,2,3,4,
Patricia M. Desmond2,5 and Elaine Lui2,5 Keywords: magnetic resonance spectroscopy, STEAM, default mode network, posterior cingulate cortex,
precuneus, reproducibility Imaging Protocol MRI scans were acquired using a 7T research scanner (Siemens
Healthcare, Erlangen, Germany) with a 32-channel head-coil
(Nova Medical Inc., Wilmington MA, United States). To assist
with localization MP2RAGE was performed prior to each session
at 0.9 mm resolution, TR = 4900 ms, TE = 2.9 ms, TI1 = 700 ms,
FA1 = 5◦, TI2 = 2700 ms, FA2 = 6◦, acquisition time (TA)
5:54 min. After manual shimming, single-voxel 1H MRS was
acquired using the STEAM method with a 20 × 20 × 20 mm
cubic voxel (volume of 8 mL), TR = 8500 ms (except for the
scans of participant 5 on day 1, for which TR = 9300 ms was
used due to reaching a SAR limit), and TE = 6 ms using outer
volume suppression. Thirty-two averages were used for the water-
suppressed sequence (TA = 4:49 min) and four averages for the
unsuppressed sequence (TA = 51 s). In this study we sought to determine the test-retest
reproducibility of quantitative single-voxel MRS of glutamate,
GABA, and GSH acquired from the PCC/precuneus region at
7T using a STEAM sequence with ultrashort TE. We chose to
recruit a broad age cohort, to allow application and comparison
to many neurologic diseases. We also chose to scan participants
twice on two different days to assess variability both within
days and between different days. MRS at 7T field strength offers
higher spectral resolution, but at the cost of increased specific
absorption rate (SAR) (Lei et al., 2013). By using a STEAM
acquisition rather than PRESS we ameliorate some of the inherent
increase in the SAR. We used high bandwidth pulses for the STEAM acquisition
in conjunction with outer volume suppression to minimize the
chemical shift artifact. In addition, we included terms fitting
the remaining lipid and macromolecular contaminations in
accordance with methods published by Hurd (2011). The voxel was placed in the midline, with its anterior inferior
border just above the corpus callosum, its anterior superior
border just dorsal to the marginal branch of the cingulate sulcus,
and its posterior border ventral to the parieto-occipital sulcus, in
a similar manner to the study of Kantarci et al. (2007; Figure 1). INTRODUCTION that
glutamate
and
GABA
in
the
PCC/precuneus
are
related to DMN activity. For example, a study of 24 health
volunteers
at
3T
showed
that
GABA
concentration
in
the PCC/precuneus positively correlated with task-related
DMN deactivation, whereas glutamate negatively correlated
with it (Hu et al., 2013). Therefore, the ability to measure
the concentration of these metabolites reliably within the
PCC/precuneus may assist with studying them as potential
non-invasive
biomarkers
for
medical
conditions
affecting
the
DMN
(e.g.,
epilepsy,
Alzheimer’s
disease),
perhaps
even during a subclinical phase, and for assessment of
treatment response. Glutamate and γ-amino butyric acid (GABA) are the major
excitatory and inhibitory neurotransmitters in the human brain,
respectively (Kocsis and Mattson, 1996; Benarroch, 2010). Glutathione (GSH) is the most important free radical scavenging
compound in the brain and is thought to be implicated in a
wide range of neurological disorders, including epilepsy, multiple
sclerosis, Parkinson’s disease, and motor neuron disease (Rae and
Williams, 2017). All three compounds are metabolically related,
as glutamate is a precursor of both GSH and GABA. Many
CNS drugs exert their effects through the glutamatergic and
GABAergic systems, and therapeutic interventions for increasing
the concentration of GSH in the brain are investigated in
recent years (e.g., dimethyl fumarate) (Kocsis and Mattson, 1996;
Benarroch, 2010; Morris et al., 2014). Citation: Gonen OM, Moffat BA, Kwan P,
O’Brien TJ, Desmond PM and Lui E
(2020) Reproducibility of Glutamate,
Glutathione, and GABA
Measurements in vivo by Single-Voxel
STEAM Magnetic Resonance
Spectroscopy at 7-Tesla in Healthy
Individuals. Conclusion: Glutamate, glutathione, and GABA can be reliably quantified using STEAM
MRS at 7-Tesla from the posterior cingulate and precuneus cortices of healthy human
subjects. STEAM MRS at 7-Tesla may be used to study the metabolic behavior of this
important resting-state hub in various disease states. Keywords: magnetic resonance spectroscopy, STEAM, default mode network, posterior cingulate cortex,
precuneus, reproducibility September 2020 | Volume 14 | Article 566643 1 Frontiers in Neuroscience | www.frontiersin.org MRS Reproducibility at 7T Gonen et al. Participants Interest in quantitative MRS of the human brain is increasing
due to its importance in evaluating drugs that potentially
affect these metabolites. However, individual quantification of
glutamate, GABA, and GSH is limited by overlapping resonances
from other molecules at magnetic fields up to 4-Tesla. High-
field (7-Tesla) MRI offers superior signal to noise ratio and
chemical shift separation which may overcome these limitations
(Tkáˇc et al., 2001). Ten healthy, right-handed, volunteers were recruited. The
participant cohort consisted of six women and four men, with
a mean ± standard deviation (SD) age of 40 ± 14 years. No
participant had any history of neurological or psychiatric disease. The study was approved by the Institutional Review Board (IRB)
of The University of Melbourne and all participants provided
written informed consent. Several reproducibility studies on MRS of these three
metabolites
have
been
performed,
evaluating
metabolite
concentrations and coefficients of variation (CoV) (Wijtenburg
et al., 2013, 2014, 2019a,b; Lally et al., 2016; Terpstra et al., 2016;
Prinsen et al., 2017). Notably, there was variation in the number
of participants, number of scans, field strength, sequences used,
and acquisition parameters. In addition, the size and placement
of the volume of interest were also different across studies (see
Table 1). Most of these studies recruited participants within a
narrow age range (e.g., young adults). Frontiers in Neuroscience | www.frontiersin.org Imaging Protocol The volume of interest was placed in the PCC/precuneus
region because of its importance as a main node of the
resting-state default mode network (DMN) of the brain
and its implication in a wide range of neurological and
psychiatric disorders (Leech and Sharp, 2014). In addition,
the disproportionately high metabolic rate of this region in
comparison with other cortical areas underscores the value of
evaluating the reproducibility of exact MRS quantitation from
this important hub (Buckner et al., 2008). Following several minutes’ break, during which time the
patients were encouraged to move their head and reposition
themselves inside the scanner, 1 mm isotropic MP2RAGE was
acquired (TA = 4:26 min) for voxel placement using the same
landmarks. Identical parameters were used for the second intra-
session MRS. The above was repeated for each participant not
more than 14 days after the first scanning session. We hypothesized that quantitative MRS of these metabolites
at 7T will have high reproducibility. There is some evidence September 2020 | Volume 14 | Article 566643 2 MRS Reproducibility at 7T Gonen et al. TABLE 1 | Examples of previous MRS reproducibility studies. No. of subjects
(repeated scans)
Sessions/Day
No. of days
Acquisition methods/Field strength
Voxel placement
References
4
1
2
STEAM, MEGA-PRESS-IVS/7T
ACC, DLPFC
Wijtenburg et al., 2013
10
1
2
PR-STEAM/3T
ACC, PCC
Wijtenburg et al., 2014
13
2
2
PRESS/7T
ACC
Lally et al., 2016
6
1
4
Semi-LASER/3T + 7T
PCC, cerebellar vermis
Terpstra et al., 2016
5
1
2
Semi-LASER, STEAM/7T
Occipital
Prinsen et al., 2017
10
2
1
PRESS, SPECIAL, PR-STEAM, MEGA-PRESS/7T
ACC
Wijtenburg et al., 2019a
10
1
2
STEAM/7T
ACC, PCC
Wijtenburg et al., 2019b
ACC, Anterior cingulate cortex; DLPFC, Dorsolateral prefrontal cortex; PCC, Posterior cingulate cortex; MEGA-PRESS, Meshcher-Garwood point resolved spectroscopy;
PRESS, Point resolved spectroscopy; PR-STEAM, Phase rotation stimulated echo acquisition mode; Semi-LASER, Semi-localization by adiabatic selective refocusing;
SPECIAL, Spin-echo full-intensity acquired localized spectroscopy. TABLE 1 | Examples of previous MRS reproducibility studies. Image Post-processing and Quantitative
MRS Calculation (version 5.0.10) (Jenkinson et al., 2012). The NMR-visible
water concentration (mM) in the voxel was estimated by
(43300fGM+35880fWM+55556fCSF)/(1-fCSF), where fGM, fWM,
and fCSF are the volume fractions of gray matter, white matter,
and CSF in the voxel, respectively (Lee et al., 2013). Metabolite concentrations were quantified using LCModel
(version 6.3-0B) with a 7T basis set (Provencher, 1993). Eddy-
current correction was used, and the metabolite concentrations
were scaled to unsuppressed water. Cramér–Rao lower bounds
(CRLB), estimated standard deviations, expressed in percent of
the estimated concentrations, were calculated by LCModel. Concentrations of metabolites were reported both as corrected
water-scaled values in mM units based on the above formula
and as dimensionless values based on the raw LCModel output
relative to total creatine (creatine + phosphocreatine) to enable
comparison to previous studies. For partial volume effect correction, binary masks were
created for each MRS voxel acquired using a MATLAB script
created by Mr. Bartosz Kossowski from The Polish Academy of
Sciences1 – see acknowledgments. MATLAB version R2017b was
used (MathWorks, Natick, MA, United States). The dimensions
of the masks were transformed via the ANTs toolbox (version
2.3.1)2 to be consistent with MP2RAGE coordinates (Avants et al.,
2011). The MP2RAGE anatomical images were skull-stripped
and the brain extracted using FMRIB’s Brain Extraction Tool
(Smith, 2002). Reproducibility Assessment and
Statistical Analysis Intra- and inter-session reproducibility of MRS concentrations
was assessed via CoV and Intraclass Correlation Coefficients
(ICC) with two-way random effects for average measures
(absolute agreement) (Shrout and Fleiss, 1979). For evaluation
of reproducibility according to ICCs, ICC <0.40 was considered
poor; between 0.40 and <0.60 fair; between 0.60 and <0.75
good; and between 0.75 and 1.00 excellent (Cicchetti, 1994). Reproducibility of the CSF volume fraction within the MRS voxel
was also assessed with CoV. The Dice similarity coefficient was
used to calculate the spatial overlap between the MRS voxels
in different scans using FMRIB’s Brain Intensity AbNormallity
Classification Algorithm (BIANCA) (Griffanti et al., 2016). Statistical analyses were conducted with Stata 15.0 (StataCorp,
College Station, TX, United States). Each brain was then segmented using FMRIB’s Automated
Segmentation Tool (FAST) into partial volume maps for gray
matter, white matter, and CSF (Zhang et al., 2001). The fractions
of each of these partial volume maps within each MRS voxel
were determined using the fslstats utility of the FSL suite 1https://www.nitrc.org/projects/rda2nifti/
2http://stnava.github.io/ANTs/
FIGURE 1 | T1-weighted MRI with superimposed MRS voxel. left – sagittal;
right – axial). 1https://www.nitrc.org/projects/rda2nifti/
2http://stnava.github.io/ANTs/ RESULTS All participants completed the two scans separated by 4–14 days
(mean ± SD was 8.9 ± 3.1 days). Mean ± SD CRLB for glutamate,
GSH, and GABA were 2.0 ± 0%, 5.0 ± 0.8%, and 8.3 ± 1.6%,
respectively; maximal CRLB were 2, 8, and 14%, respectively. The
mean ± SD FWHM of the 40 spectra was 0.033 ± 0.004 ppm
(9.8 ± 1.2 Hz), which reflects good quality acquisition (Kreis,
2004). The chemical shift artifact was 1.7 mm per ppm and
mean ± SD data shift across all 40 scans was 0.026 ± 0.005 ppm. An example of one of the spectra appears in Figure 2. FIGURE 1 | T1-weighted MRI with superimposed MRS voxel. left – sagittal;
right – axial). Mean ± SD concentrations of glutamate, GSH, and GABA
scaled
to
unsuppressed
water
were
10.18
±
1.00
mM, September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org 3 Gonen et al. MRS Reproducibility at 7T FIGURE 2 | Example of a spectrum. TABLE 2 | Metabolite concentrations. A – Mean concentrations scaled to unsuppressed water (standard deviation)
Metabolite (mM)
Day 1/Scan 1
Day 1/Scan 2
Day 2/Scan 1
Day 2/Scan 2
Overall
Glutamate
10.34 (0.93)
9.69 (0.95)
10.24 (0.66)
10.45 (1.30)
10.18 (1.00)
GSH
1.86 (0.27)
1.83 (0.25)
1.80 (0.21)
1.84 (0.37)
1.83 (0.27)
GABA
1.52 (0.20)
1.48 (0.31)
1.57 (0.31)
1.65 (0.26)
1.56 (0.27)
B – Mean concentrations scaled to creatine (standard deviation)
Metabolite/Cr
Day 1/Scan 1
Day 1/Scan 2
Day 2/Scan 1
Day 2/Scan 2
Overall
Glutamate
1.16 (0.07)
1.13 (0.06)
1.16 (0.07)
1.17 (0.09)
1.16 (0.07)
GSH
0.21 (0.02)
0.22 (0.03)
0.20 (0.03)
0.20 (0.02)
0.21 (0.02)
GABA
0.17 (0.02)
0.17 (0.03)
0.18 (0.03)
0.18 (0.02)
0.18 (0.03)
1.83
±
0.27
mM,
and
1.56
±
0.27
mM,
respectively;
concentrations scaled to total creatine were 1.16 ± 0.07,
0.21 ± 0.02, and 0.18 ± 0.03, respectively (Table 2). The intra
and inter-session coefficients of variation for water-scaled
metabolites ranged between 2.79 and 6.41% for glutamate, 6.66
and 10.82% for GSH, and 10.97 and 11.69% for GABA. ICC
values were 0.32–0.75, 0.20–0.72, and 0.49–0.82, respectively
(Table 3). Our results are compared to similar creatine-scaled
studies in Table 4. RESULTS An analysis of MRS voxel placement
reproducibility between scans is included in Table 5 and
an example of the four scans of a single subject (subject
(CRLB < 20%) in all scans and raw uncorrected water-scaled
data for all the metabolites discussed in this manuscript appear
in Supplementary Tables 1, 2. DISCUSSION
Our study demonstrated good reproducibility of glutamate, GSH,
and GABA concentrations measured from the PCC/precuneus
at 7T using STEAM, as measured by CoV in a population of
subjects with a broader age range in comparison with other FIGURE 2 | Example of a spectrum. Day 2/Scan 2
10.45 (1.30)
1.84 (0.37)
1.65 (0.26)
Day 2/Scan 2
1.17 (0.09)
0.20 (0.02)
0.18 (0.02)
s and raw uncorrected w
discussed in this manuscr
2. od reproducibility of glutam
measured from the PCC/
easured by CoV in a pop
e range in comparison
ranging between poor an
ns, with the lowest value (CRLB < 20%) in all scans and raw uncorrected water-scaled
data for all the metabolites discussed in this manuscript appear
in Supplementary Tables 1, 2. 1.83
±
0.27
mM,
and
1.56
±
0.27
mM,
respectively;
concentrations scaled to total creatine were 1.16 ± 0.07,
0.21 ± 0.02, and 0.18 ± 0.03, respectively (Table 2). The intra
and inter-session coefficients of variation for water-scaled
metabolites ranged between 2.79 and 6.41% for glutamate, 6.66
and 10.82% for GSH, and 10.97 and 11.69% for GABA. ICC
values were 0.32–0.75, 0.20–0.72, and 0.49–0.82, respectively
(Table 3). Our results are compared to similar creatine-scaled
studies in Table 4. An analysis of MRS voxel placement
reproducibility between scans is included in Table 5 and
an example of the four scans of a single subject (subject
10) appears in Figure 3. Concentrations of several other
metabolites that were consistently acquired with good quality DISCUSSION Glutamate
GSH
GABA
Glutamate
GSH
GABA
Glutamate
GSH
GABA
Mean concentration day 1
1.37
0.25
0.20
1.25
0.24
0.34
1.15
0.22
0.17
CoV day 1
6.00%
14.95%
36.89%
N/A
N/A
N/A
3.56%
8.99%
13.11%
ICC (mean) day1
0.88
0.49
−0.17
N/A
N/A
N/A
0.44
0.33
−0.27
Mean concentration day 2
1.33
0.24
0.19
1.16
0.23
0.36
1.17
0.20
0.18
CoV day 2
4.77%
8.54%
29.19%
N/A
N/A
N/A
2.22%
6.67%
8.32%
ICC (mean) day2
0.94
0.88
0.37
N/A
N/A
N/A
0.90
0.59
0.72
Mean concentration 1st scans
1.38
0.25
0.20
N/A
N/A
N/A
1.16
0.21
0.18
CoV both 1st scans
6.48%
11.45%
30.92%
6.20%
14.30%
18.40%
2.82%
5.69%
8.89%
ICC (mean) both 1st scans
0.86
0.65
−0.26
N/A
N/A
N/A
0.85
0.67
0.69
Mean concentration 2nd scans
1.38
0.25
0.18
N/A
N/A
N/A
1.15
0.21
0.18
CoV both 2nd scans
7.95%
8.25%
15.43%
N/A
N/A
N/A
4.00%
10.13%
9.91%
ICC (mean) both 2nd scans
0.68
0.76
0.21
N/A
N/A
N/A
0.51
0.13
−0.35 who were scanned once only on two different days, with mixed
sequences (STEAM for glutamate; Semi-Localized by Adiabatic
Selective Refocusing for GSH and GABA) (Prinsen et al., 2017). Terpstra et al. (2016) did not report exact CoV figures for their
7T PCC study, but judging by their bar charts, our CoV values
seem to be superior. between the sessions of the 2nd day, suggesting a substantial
effect of the random repositioning during that day. However, the
overall ICC values, and in particular the ICCs of the 1st scans
after standard positioning in the scanner on both days ranged,
for the most part, between good and excellent, depending on the
metabolite and scaling. Within- and between-sessions reproducibility measures were
not explored in most other 7T studies. A notable exception is
the study of Lally et al. (2016) who measured creatine-scaled
metabolite ratios in the ACC twice on two different days in
26 subjects ranging in age from 20 to 54 years using PRESS. Other exceptions are the studies of Stephenson et al. (2011)
and Cai et al. (2012) who studied intra-session and inter-session
reproducibility of MRS quantitation of glutamate and GABA,
without GSH quantification. Our study, therefore, is novel as it
is a 7T quantitative STEAM study with a 2∗2 design, assessing
reproducibility by CoV and ICC of all three J-coupled metabolites
(glutamate, GSH, and GABA) in the PCC/precuneus. Wijtenburg et al. DISCUSSION Glutamate
GSH
GABA
Glutamate
GSH
GABA
Glutamate
GSH
GABA
Mean concentration day 1
1.37
0.25
0.20
1.25
0.24
0.34
1.15
0.22
0.17
CoV day 1
6.00%
14.95%
36.89%
N/A
N/A
N/A
3.56%
8.99%
13.11%
ICC (mean) day1
0.88
0.49
−0.17
N/A
N/A
N/A
0.44
0.33
−0.27
Mean concentration day 2
1.33
0.24
0.19
1.16
0.23
0.36
1.17
0.20
0.18
CoV day 2
4.77%
8.54%
29.19%
N/A
N/A
N/A
2.22%
6.67%
8.32%
ICC (mean) day2
0.94
0.88
0.37
N/A
N/A
N/A
0.90
0.59
0.72
Mean concentration 1st scans
1.38
0.25
0.20
N/A
N/A
N/A
1.16
0.21
0.18
CoV both 1st scans
6.48%
11.45%
30.92%
6.20%
14.30%
18.40%
2.82%
5.69%
8.89%
ICC (mean) both 1st scans
0.86
0.65
−0.26
N/A
N/A
N/A
0.85
0.67
0.69
Mean concentration 2nd scans
1.38
0.25
0.18
N/A
N/A
N/A
1.15
0.21
0.18
CoV both 2nd scans
7.95%
8.25%
15.43%
N/A
N/A
N/A
4.00%
10.13%
9.91%
ICC (mean) both 2nd scans
0.68
0.76
0.21
N/A
N/A
N/A
0.51
0.13
−0.35 TABLE 3 | Reproducibility measures. Type of Comparison
Coefficient
Relative to unsuppressed water
Relative to Creatine
Glutamate
GSH
GABA
Glutamate
GSH
GABA
Within Day 1
ICC
0.72
0.66
0.49
0.44
0.33
−0.27
(Scan 1 vs. Scan 2)
CoV (%)
5.91%
7.81%
11.52%
3.56%
8.99%
13.11%
Within Day 2
ICC
0.32
0.20
0.52
0.90
0.59
0.72
(Scan 1 vs. Scan 2)
CoV (%)
5.83%
10.69%
11.42%
2.22%
6.67%
8.32%
Between-session
ICC
0.87
0.72
0.82
0.85
0.67
0.69
(Scan 1)
CoV (%)
2.79%
6.66%
7.35%
2.82%
5.69%
8.89%
Between-session
ICC
0.68
0.69
0.65
0.51
0.13
−0.35
(Scan 2)
CoV (%)
6.40%
9.84%
10.97%
4.00%
10.13%
9.91%
Overall (4 scans)
ICC
0.75
0.68
0.77
0.86
0.59
0.66
CoV (%)
6.41%
10.82%
11.69%
3.00%
8.38%
8.06% TABLE 4 | Comparison to other creatine-scaled MRS studies of the PCC (Lally et al., 2016; Wijtenburg et al., 2019b). TABLE 4 | Comparison to other creatine-scaled MRS studies of the PCC (Lally et al., 2016; Wijtenburg et al., 2019b). Type of Comparison
Lally et al. Wijtenburg et al. Gonen et al. DISCUSSION Our study demonstrated good reproducibility of glutamate, GSH,
and GABA concentrations measured from the PCC/precuneus
at 7T using STEAM, as measured by CoV in a population of
subjects with a broader age range in comparison with other
studies. ICC results varied, ranging between poor and excellent
across different comparisons, with the lowest values recorded Our study demonstrated good reproducibility of glutamate, GSH,
and GABA concentrations measured from the PCC/precuneus
at 7T using STEAM, as measured by CoV in a population of
subjects with a broader age range in comparison with other
studies. ICC results varied, ranging between poor and excellent
across different comparisons, with the lowest values recorded September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org Frontiers in Neuroscience | www.frontiersin.org MRS Reproducibility at 7T Gonen et al. TABLE 3 | Reproducibility measures. Type of Comparison
Coefficient
Relative to unsuppressed water
Relative to Creatine
Glutamate
GSH
GABA
Glutamate
GSH
GABA
Within Day 1
ICC
0.72
0.66
0.49
0.44
0.33
−0.27
(Scan 1 vs. Scan 2)
CoV (%)
5.91%
7.81%
11.52%
3.56%
8.99%
13.11%
Within Day 2
ICC
0.32
0.20
0.52
0.90
0.59
0.72
(Scan 1 vs. Scan 2)
CoV (%)
5.83%
10.69%
11.42%
2.22%
6.67%
8.32%
Between-session
ICC
0.87
0.72
0.82
0.85
0.67
0.69
(Scan 1)
CoV (%)
2.79%
6.66%
7.35%
2.82%
5.69%
8.89%
Between-session
ICC
0.68
0.69
0.65
0.51
0.13
−0.35
(Scan 2)
CoV (%)
6.40%
9.84%
10.97%
4.00%
10.13%
9.91%
Overall (4 scans)
ICC
0.75
0.68
0.77
0.86
0.59
0.66
CoV (%)
6.41%
10.82%
11.69%
3.00%
8.38%
8.06%
TABLE 4 | Comparison to other creatine-scaled MRS studies of the PCC (Lally et al., 2016; Wijtenburg et al., 2019b). Type of Comparison
Lally et al. Wijtenburg et al. Gonen et al. Frontiers in Neuroscience | www.frontiersin.org DISCUSSION Subject
CSF coefficient of variation
Dice coefficient (voxel overlap)
Day 1 (%)
Day 2 (%)
1st scans (%)
2nd scans (%)
Day 1 (%)
Day 2 (%)
1st scans (%)
2nd scans (%)
1
12.3
19.2
8.8
1.8
94.5
93.8
88.8
89.6
2
14.0
12.8
9.5
8.3
90.8
90.6
79.0
76.9
3
20.2
28.0
1.1
7.0
83.9
92.2
86.5
95.6
4
9.9
7.4
0.7
18.0
92.2
93.1
77.5
72.0
5
11.2
36.7
11.0
56.5
85.7
86.2
83.5
75.8
6
14.9
47.1
2.0
61.6
85.7
89.4
87.9
88.3
7
55.6
27.3
18.0
12.9
89.5
87.0
91.5
93.1
8
2.4
2.1
8.2
3.8
83.9
95.2
89.0
96.0
9
34.1
28.8
10.3
4.8
92.2
81.8
86.3
77.3
10
66.9
11.4
27.7
53.3
93.9
92.0
91.0
94.2
Mean
24.1
22.1
9.7
22.8
89.2
90.1
86.1
85.9 LE 5 | MRS voxel composition and overlap of individual subjects. FIGURE 3 | Voxel prescriptions and spectra of a single subject (subject number 10). FIGURE 3 | Voxel prescriptions and spectra of a single subject (subject number 10). FIGURE 3 | Voxel prescriptions and spectra of a single subject (subject number 10). semi-adiabatic LASER sequence is most suitable when TE values
of 25–30 ms are acceptable, whereas the semi-adiabatic SPECIAL
sequence is suitable for shorter TE values but at the expense
of increased susceptibility to subject motion (Öz et al., 2020). Therefore, another advantage of STEAM with ultrashort TE, as
used in our study, is its applicability in cases of subjects prone
to move during the scan (Öz et al., 2020). This provides further
support to the importance of studying MRS reproducibility of
STEAM with ultrashort TE, as the results can be applied in studies
of patients with conditions such as epilepsy and various forms of
dementia, in which subject motion is expected. Our results are,
therefore, encouraging in the sense that despite intrinsically lower
signal to noise ratio, the reproducibility of STEAM at 7T with
ultrashort TE is not inferior. also calculated exact concentrations of metabolites scaled to water
with partial volume correction according to segmentation to gray
matter, white matter, and CSF, as outlined above, which renders
the results more useful for future quantitative MRS biomarker
studies. This also allows comparison to other studies that
evaluated water-scaled metabolite concentrations (Wijtenburg
et al., 2014, 2019a; Terpstra et al., 2016; Prinsen et al., 2017). Frontiers in Neuroscience | www.frontiersin.org DISCUSSION (2013; 2014; 2019a; 2019b) studied the
reproducibility of brain MRS in a series of publications at 3T
and at 7T in groups of volunteers who were scanned twice using
several different sequences. Metabolites acquired via STEAM
from the PCC at 7T were from an older cohort scanned twice
2–3 months apart using a voxel of 1.6 × 2.0 × 2.8 cm and
were reported as ratios relative to total creatine (creatine +
phosphocreatine) without exact quantification in water-scaled
values (Wijtenburg et al., 2019b). No ICC values were reported
for this cohort. In our study we reported ratios of metabolite concentrations
relative to total creatine, to enable comparison with other similar
studies. However, the percentages of gray matter, white matter,
and CSF may vary in the volume of interest as a function of voxel
placement and patient characteristics, and the concentration of
creatine itself can vary with age (Suri et al., 2017). Therefore, we Prinsen et al. (2017) who acquired MRS at 7T from the
midline occipital cortex, reported CoV of 3.2, 7.8, and 9.5%, for
glutamate, GSH, and GABA, respectively. Their cohort consisted
of five participants with a mean age of 32 years (range 24–40) September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org 5 MRS Reproducibility at 7T Gonen et al. TABLE 5 | MRS voxel composition and overlap of individual subjects. Subject
CSF coefficient of variation
Dice coefficient (voxel overlap)
Day 1 (%)
Day 2 (%)
1st scans (%)
2nd scans (%)
Day 1 (%)
Day 2 (%)
1st scans (%)
2nd scans (%)
1
12.3
19.2
8.8
1.8
94.5
93.8
88.8
89.6
2
14.0
12.8
9.5
8.3
90.8
90.6
79.0
76.9
3
20.2
28.0
1.1
7.0
83.9
92.2
86.5
95.6
4
9.9
7.4
0.7
18.0
92.2
93.1
77.5
72.0
5
11.2
36.7
11.0
56.5
85.7
86.2
83.5
75.8
6
14.9
47.1
2.0
61.6
85.7
89.4
87.9
88.3
7
55.6
27.3
18.0
12.9
89.5
87.0
91.5
93.1
8
2.4
2.1
8.2
3.8
83.9
95.2
89.0
96.0
9
34.1
28.8
10.3
4.8
92.2
81.8
86.3
77.3
10
66.9
11.4
27.7
53.3
93.9
92.0
91.0
94.2
Mean
24.1
22.1
9.7
22.8
89.2
90.1
86.1
85.9
FIGURE 3 | Voxel prescriptions and spectra of a single subject (subject number 10). TABLE 5 | MRS voxel composition and overlap of individual subjects. DISCUSSION (2016) is not entirely
clear, and this may affect the ability for direct comparison
between the studies. Regardless, this is not at the expense of
poorer reproducibility. Although some of the intra-session CoV are higher than
between sessions, this is not a consistent finding across the three
metabolites. This is most evident for GABA, with both intra-
session CoV higher than both inter-session CoV, but when scaled
to water, the absolute difference is small (i.e., highest mean intra-
session CoV of 11.5% compared to highest mean inter-session
CoV of 11.0%). GSH also has a mean intra-session CoV higher
than its highest inter-session CoV. Again, the absolute difference
is small (intra-session CoV 10.7 vs. 9.8% inter-session). Compared to Lally’s study, our CoV are lower, for all three
metabolites, and for all intra and inter-sessions, except for inter-
session GSH of 2nd scans, which was 10.1% in our study and 8.3%
in Lally’s. For example, our GABA intra-session CoV are 13.1 and
8.3% compared to Lally’s 36.9 and 29.2%, respectively. Similarly,
our inter-session 1st scan glutamate CoV are 2.8% vs. Lally’s 6.5%,
and our GSH CoV is 5.7% vs. Lally’s 11.5%. Variation in voxel placement and composition is an important
contributing factor to the variation in CoV, particularly within,
but also between, sessions. Three subjects had their lowest Dice
coefficient and four subjects had their highest variation in CSF
composition within the first day’s session, with one subject having
a 55.6% variation in CSF composition in the first day’s session. Four subjects had the lowest Dice coefficients between the 2nd
scans of each session. Not only were these of the lowest values
(Dice between 72 and 77%), but two subjects also had the
highest CSF composition variation (of up to 61.6%) between
the 2nd scans of each session. Although meticulous adherence
to the multiple landmarks was employed and included in our
voxel positioning, our methodology also included intersession
repositioning of the patient’s head in a way that accentuates
interscan rotational differences. Notably, while we positioned the
patients facing up for the first scan of each session, we deliberately
asked them to rotate their heads during the break within sessions
so that they were positioned in random angles during the second
scan of each day. However, even despite this, our intrasession
CoV remain smaller than Lally’s, and our Dice coefficients also
remain similar to limited available literature. Compared to Wijtenburg et al. DISCUSSION (2019b)’s 7T STEAM study,
our metabolite concentrations were also lower for the most
part. Glutamate was 0.10 units lower on the first day and 0.01
units higher on the second day, respectively (8.4 and 0.4%,
respectively). GSH was 0.02 and 0.03 units lower, respectively
(10.4 and 13.0%, respectively). Our GABA concentrations were
substantially lower – 0.17 and 0.18 units, respectively (50.0% for
both). But still, the CoV were bigger in Wijtenburg’ study, and
their mean values for creatine-scaled metabolites in the PCC
region (1.6 × 2.0 × 2.8 cm voxel) were 6.2, 14.3, and 18.4% for
glutamate, GSH, and GABA, respectively. It may be the shorter TE of our STEAM sequence compared
to Wijtenburg et al.’s (2019b) 7T study (6 ms compared to
14 ms) that explains our superior reproducibility, reducing
signal loss due to T2 relaxation. Notably, we used outer volume
suppression to enable shorter echo times (Chen et al., 1997). We also used longer TR (8500 ms compared to 3000 ms)
which diminished signal loss due to T1-weighting. Finally, our
improved reproducibility may also be related to our shorter
interval between scan sessions (8.9 days vs. 2.4 months). A
direct
comparison
of
water-scaled
metabolite
concentrations to Wijtenburg’s 2014 phase rotation STEAM
study is not possible, as values in the latter were reported in
“institutional units,” but we note that their CoV for glutamate,
GSH and GABA between their two scans were 7.2, 8.6, and
10.5%, respectively, which were higher than values between the
1st sessions of each day in our study, but lower than our results
for the 2nd sessions of each day, except for glutamate. Their ICCs
for these metabolites were 0.59, 0.51, and 0.35, respectively, and
in comparison, our ICCs were lower both for the intersession
comparison of the 1st scans of each day, and the comparison of
the 2nd scans of each day. When carefully examining the raw
data of Prinsen et al. (2017) who used a 2 × 2 × 2 cm cubic voxel
positioned in the mid-occipital region but largely consisting of
precuneus tissue, their mean concentrations for glutamate, GSH
and GABA using STEAM were 10.56 ± 0.48, 1.34 ± 0.13, and
1.38 ± 0.26 mM, comparable to our results. Bai et al. DISCUSSION We chose STEAM, which has the advantage of a lower SAR
compared to PRESS because of using 90◦rather than 180◦
excitation pulses, at the expense of a lower signal-to-noise ratio
(SNR) (Öz et al., 2020). A lower SAR is beneficial as it enables
the inclusion of other sequences as part of the MRI session. As
the SNR improves at 7T, STEAM becomes more advantageous
(Lei et al., 2013). Several new MRS sequences were introduced
in recent years such as the spin-echo full-intensity acquired
localized (SPECIAL) and the localization by adiabatic selective
refocusing (LASER) (Lin et al., 2014; Mekle et al., 2009). However,
while these sequences and their semi-adiabatic versions have
superior localization performance, they also have limitations. The Our metabolite ratios, when scaled to total creatine, were
somewhat lower in comparison with Lally et al’s values (Table 4;
Lally et al., 2016). Glutamate was 0.22 and 0.16 units lower on
the first and second days, respectively (16.1 and 12.0% lower,
respectively). Similarly, GSH was 0.03 and 0.04 units lower (12.0 September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org 6 Gonen et al. MRS Reproducibility at 7T and 16.7%, respectively), and GABA was 0.03 and 0.01 units lower
(15.0 and 5.3%, respectively). and 16.7%, respectively), and GABA was 0.03 and 0.01 units lower
(15.0 and 5.3%, respectively). The CoV in our study are not clearly lower when the metabolic
concentrations are scaled to water vs. creatine. The CoV of
glutamate concentration is consistently the lowest for both intra
and inter-session reproducibility comparison. Both GABA and
GSH have generally larger CoV than glutamate. However, GABA
and GSH have much lower concentrations than glutamate. The
mean (±SD) ratio of glutamate to GABA across all days was 6.60
(±0.17) and for glutamate to GSH was 5.58 (±0.33). In addition,
glutamate has the lowest CRLB values. Therefore, the lower CoV
of glutamate was expected. y
Although Lally’s study was also of healthy subjects at 7T, this
is not surprising, with the intrinsic lower SNR of a STEAM
MRS sequence compared to the PRESS. It is also possible to at
least be partially related to the difference in location of voxel
placement, ours in the PCC/precuneus compared to Lally et al’s
pregenual cingulate. Notably, the relative contribution of creatine
and phosphocreatine to the spectral peaks used for metabolite
quantification in the study of Lally et al. September 2020 | Volume 14 | Article 566643 DISCUSSION (2017) measuring the reproducibility of voxel
placement for GABA-edited MRS on 13 healthy volunteers using
9 mL cubic voxels placed over the right sensorimotor and midline
occipital cortices has within-subject Dice coefficients of 86 ± 5%
and 87 ± 5%, respectively, compared to our values of 89 ± 4%
and 90 ± 4% within the 1st and 2nd days, respectively, and
86 ± 5% and 86 ± 9% within the first and second sessions of each
day, respectively. A limitation of this study is the MP2RAGE resolution was
higher in the first scans of each day (0.9 mm isotropic) that the
second scans (1 mm isotropic). This was done for reasons related
to SAR, as MRS was acquired as part of a broader protocol. Nevertheless, the spectra were all acquired with an identical voxel
size, and for the purposes of comparison of voxel placement,
all images of each patient were resliced and co-registered to the
first image of the first scan. Another point to consider is that
metabolite concentrations may fluctuate according to diurnal September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org Frontiers in Neuroscience | www.frontiersin.org 7 MRS Reproducibility at 7T Gonen et al. variation, with our participants scanned at different times of the
day for reasons related to their convenience (Arm et al., 2019). conceptualization,
funding
acquisition,
methodology,
supervision,
and
review
of
manuscript. PD
and
EL:
conceptualization, methodology, supervision, and review of
manuscript. BM: investigation, resources, formal analysis,
methodology, and review of manuscript. All authors contributed
to the article and approved the submitted version. In conclusion, glutamate, GSH and GABA can be reliably
quantified at 7T using STEAM MRS with ultrashort TE from the
posterior cingulate and precuneus cortices of healthy individuals
ranging in age from young adults to elderly. This method strikes
a balance between the signal to noise ratio and susceptibility
to subject motion, which can occur with advanced age and in
certain patient groups. Moreover, the broad age range used in our
study serves as a proof of concept for acquiring MRS biomarkers
in conditions affecting different age groups (e.g., schizophrenia,
epilepsy, Alzheimer’s disease), focusing on the metabolic behavior
of this important resting-state hub in various disease states. DATA AVAILABILITY STATEMENT The datasets for this article are not publicly available because
the raw imaging files are stored on a secure server, as per the
ethics approval. However, the numerical output of the LCModel
computer program for all participants is available upon request. Requests to access the datasets should be directed to Ofer M. Gonen, Ofer.Gonen@mh.org.au. ACKNOWLEDGMENTS We acknowledge the facilities, scientific and technical assistance,
and salary support of BM from the National Imaging Facility, a
National Collaborative Research Infrastructure Strategy (NCRIS)
capability, at the Melbourne Brain Centre Imaging Unit, The
University of Melbourne. OG was supported by the Melbourne
Research Scholarship (MRS), The University of Melbourne and
The Neuroscience Foundation, The Royal Melbourne Hospital. We would also like to thank Mr. Bartosz Kossowski, Laboratory
of Brain Imaging, Nencki Institute of Experimental Biology,
Polish Academy of Sciences, for a tool which creates MRS voxel
masks aligned to imaging data (https://www.nitrc.org/projects/
rda2nifti/). SUPPLEMENTARY MATERIAL The studies involving human participants were reviewed and
approved by the University of Melbourne Institutional Review
Board. The patients/participants provided their written informed
consent to participate in this study. The Supplementary Material for this article can be found
online
at:
https://www.frontiersin.org/articles/10.3389/fnins. 2020.566643/full#supplementary-material The Supplementary Material for this article can be found
online
at:
https://www.frontiersin.org/articles/10.3389/fnins. 2020.566643/full#supplementary-material TABLE S1 | Other consistently acquired metabolites. NAA, N-acetylaspartate;
GLN, Glutamine; INS, Inositol; Total Cr, Creatine + Phosphocreatine. TABLE S1 | Other consistently acquired metabolites. NAA, N-acetylaspartate;
GLN, Glutamine; INS, Inositol; Total Cr, Creatine + Phosphocreatine. AUTHOR CONTRIBUTIONS TABLE S2 | Raw uncorrected water-scaled data. GABA, γ-amino butyric acid;
GLUT, Glutamate; GSH, Glutathione; NAA, N-acetylaspartate; GLN, Glutamine; TABLE S2 | Raw uncorrected water-scaled data. GABA, γ-amino butyric acid;
GLUT, Glutamate; GSH, Glutathione; NAA, N-acetylaspartate; GLN, Glutamine;
INS, Inositol; Total Cr, Creatine + Phosphocreatine. For creatine-scaling divide by
total creatine data columns. OG:
conceptualization,
methodology,
investigation,
formal
analysis,
and
writing
original
draft. TO’B
and
PK: OG:
conceptualization,
methodology,
investigation,
formal
analysis,
and
writing
original
draft. TO’B
and
PK: INS, Inositol; Total Cr, Creatine + Phosphocreatine. For creatine-scaling divide by
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10.1002/nbm.4236 [Epub ahead of print]. Conflict of Interest: The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be construed as a
potential conflict of interest. Frontiers in Neuroscience | www.frontiersin.org September 2020 | Volume 14 | Article 566643 REFERENCES Conflict of Interest: The authors declare that the research was conducted in the
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open-access article distributed under the terms of the Creative Commons Attribution
License (CC BY). The use, distribution or reproduction in other forums is permitted,
provided the original author(s) and the copyright owner(s) are credited and that the
original publication in this journal is cited, in accordance with accepted academic
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its roles and measurement by magnetic resonance spectroscopy. Anal. Biochem. 529, 127–143. doi: 10.1016/j.ab.2016.12.022 September 2020 | Volume 14 | Article 566643 Frontiers in Neuroscience | www.frontiersin.org 9
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Panorama das pesquisas sobre a participação das mulheres na construção da astronomia nos séculos XIX e XX
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1 Universidade Federal do Paraná, Curitiba-PR; Universidade Federal do Sul e Sudeste do Pará, Marabá-PA,
camilasitko@yahoo.com.br. Abstract The women's contribution to the production of knowledge in Astronomy throughout history appears, in general, as
peripheral (and sometimes omitted), even though women have often been protagonists in the construction of this
science. In this work, a state of knowledge research was carried out to assess the presence of articles, theses, and
dissertations that deal with women's contributions to the construction of Astronomy in the 19th and 20th
centuries. The results show few productions, to the detriment of the real participation of women in the area. Thus,
it is necessary that historical reconstructions of this period are carried out better understand the history of
astronomy and to present the women's contributions in the area. Keywords: Women in Astronomy; History of Astronomy; History and Philosophy of Science; Content Analysis. Panorama das pesquisas sobre a participação das mulheres na construção da astronomia nos
séculos XIX e XX Panorama das pesquisas sobre a participação das mulheres na construção da astronomia nos
séculos XIX e XX Camila Maria Sitko1 Resumo A contribuição feminina na produção do conhecimento em Astronomia ao longo da História aparece, em geral,
como periférica (ou em alguns casos é omitida), mesmo que muitas vezes as mulheres tenham sido protagonistas
na construção dessa ciência. Neste trabalho, realizou-se uma pesquisa do tipo estado do conhecimento, a fim de se
avaliar a presença de artigos, teses e dissertações que tratem de contribuições femininas para a construção da
Astronomia nos séculos XIX e XX. Os resultados mostram poucas produções, em detrimento da participação real
das mulheres na área. Assim, faz-se necessário que reconstruções históricas desse período sejam realizadas, para
se conhecer melhor a História da Astronomia e para apresentar as contribuições femininas na área. p
p
ç
Palavras-chave: Mulheres na Astronomia; História da Astronomia; História e Filosofia da Ciência; Análise de
Conteúdo. DOI: https://doi.org/10.24933/horizontes.v40i1.1298 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 Introdução Atualmente, pode-se perceber um aumento expressivo da participação das mulheres no
campo científico. No entanto, conforme apresentam Silva e Ribeiro (2014), ainda há muitas
desigualdades e barreiras na produção e participação científica feminina, devido a, por
exemplo, “modelos masculinos de carreira” (VELHO, 2006). Como mostra Chassot (2004), e sem
ter a intenção de simplificar as razões para uma única resposta (LINO; MAYORGA, 2016), nossa Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [1] DOI: https://doi.org/10.24933/horizontes.v40i1.1298
sociedade não é masculina por acaso, mas advém de uma herança cultural religiosa, entre
outros motivos. DOI: https://doi.org/10.24933/horizontes.v40i1.1298
sociedade não é masculina por acaso, mas advém de uma herança cultural religiosa, entre
outros motivos. sociedade não é masculina por acaso, mas advém de uma herança cultural religiosa, entre
outros motivos. sociedade não é masculina por acaso, mas advém de uma herança cultural religiosa, entre
outros motivos. No século XIX, movimentos por igualdade de direitos para as mulheres começaram a
aparecer, bem como críticas ao modelo masculino de ciência (SILVA, 2008), que fazia com que
estas fossem obrigadas a viver à sombra de homens e por isso tivessem suas investigações
usurpadas, e ainda fossem impedidas de partilhar espaços científicos. No entanto, apesar das críticas dos cientistas e das feministas (cientistas ou não), dos
movimentos de denúncia e reivindicação das mulheres, o mundo científico
continua
reproduzindo
as
práticas
discriminatórias
e
estereotipadas
historicamente e reforçando as desigualdades. Mesmo considerando que,
desde
os
primórdios,
muitas
mulheres
produziram
conhecimento
e
considerando os avanços contemporâneos, na questão da inserção das
mulheres nos centros acadêmicos e nos laboratórios, elas continuam
periféricas ou mesmo invisíveis quanto às revoluções no campo das ciências
(SILVA, 2008, p.138). Ainda tendo grandes desafios a serem superados nesse sentido, a contribuição feminina
ao longo da História da Ciência foi e é essencial para a construção do conhecimento científico. Um exemplo disso é a relevante contribuição das mulheres no desenvolvimento da ciência da
Astronomia nos fins do século XIX e início do século XX, que trouxeram conhecimentos que são
utilizados até os dias de hoje na área, mas que não são, de maneira geral, reconhecidas por tais
feitos. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 Segundo Donna Haraway (1989), não existe apenas uma História da Ciência, mas diversas, que
precisam ser analisadas, pois suas construções são feitas para atender certos interesses sociais,
políticos ou econômicos. Como afirma Silva (2008, p.138), “através destes rápidos destaques,
percebe-se que a construção do conhecimento científico depende dos contextos, pois se trata
de uma prática social com base empírica e carregada de valores e ideologias”. Pode ser que as mulheres astrônomas não tivessem aparecido na versão tradicional da
História, mas é necessário então mostrar outra versão, aquela em que elas são protagonistas de
suas próprias produções. Segundo Silva (2008), há uma distorção na História, no sentido de fazer com que as
mulheres não apareçam como protagonistas da construção de um determinado conhecimento. Os estudos de epistemologia feminina permitem perceber o quanto é importante resgatar esse
protagonismo feminino das mulheres da ciência. Este trabalho procura identificar as pesquisas históricas que envolvem mulheres
astrônomas que trabalhavam no Harvard Observatory, nos séculos XIX e XX, a partir de uma
pesquisa tipo estado do conhecimento, a fim de se compreender como a história é contada, e
como a presença feminina aparece ao longo da história da construção do conhecimento
científico em Astronomia. Esta pesquisa é a etapa inicial de uma pesquisa maior, tem como objetivo incentivar as
meninas à ciência e à astronomia, e futuramente, elaborar materiais didáticos e sugerir
abordagens históricas que coloquem em evidência os méritos femininos na ciência. O objetivo desta etapa é obter um panorama das pesquisas já realizadas nessas
temáticas, para que saibamos por onde começar, ou de onde prosseguir, baseando-nos nas
pesquisas já realizadas por outros autores. Assim, neste trabalho, realizou-se uma pesquisa do
tipo estado do conhecimento sobre artigos, teses e dissertações que abordam a história da
astronomia, apresentando as contribuições femininas nessa área. Introdução Na História e na Filosofia da Ciência (HFC), tratamos da discussão a respeito da
construção do conhecimento, aspectos culturais e sociológicos da ciência, como a relação e
influência de aspectos sociais e econômicos da época influenciaram a construção de um
determinado conhecimento, e também da natureza da ciência (LEDERMAN, 1992), que diz
respeito a uma construção humana, criativa e coletiva, feita por pessoas de uma grande
diversidade, diferentemente da imagem apresentada nos materiais didáticos de genialidade e
de que somente homens, ricos e brancos podem fazê-la. O que era de se imaginar é que as mulheres fossem incluídas então no discurso e nos
estudos de HFC para mostrar melhor essas características, mas em geral, não é o que ocorre. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [2] Um adendo à história das calculadoras de Harvard Aqui fazemos um rápido resumo acerca das contribuições femininas para a Astronom Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [3] DOI: https://doi.org/10.24933/horizontes.v40i1.1298
dos séculos XIX e XX, que não pretende esgotar os feitos e os nomes femininos envolvidos, mas
apenas dar uma breve noção ao leitor acerca da existência dessa história (geralmente) não
contada. dos séculos XIX e XX, que não pretende esgotar os feitos e os nomes femininos envolvidos, mas
apenas dar uma breve noção ao leitor acerca da existência dessa história (geralmente) não
contada. dos séculos XIX e XX, que não pretende esgotar os feitos e os nomes femininos envolvidos, mas
apenas dar uma breve noção ao leitor acerca da existência dessa história (geralmente) não
contada. O astrônomo Edward Charles Pickering, diretor do Harvard College Observatory (OCO),
nos Estados Unidos, na década de 1870, contratou um grupo de cerca de 80 mulheres para
catalogar e analisar seus dados astronômicos. Algumas das razões do trabalho feminino é que
seu custo de trabalho era menor do que empregar homens, e porque os detalhes dos dados
exigiam muita atenção, o que Pickering acreditava que seria mais bem levado em conta por
mulheres (SANTOS, SANTOS; SANTOS, 2018). Esse grupo ficou conhecido como as calculadoras de Pickering, pois faziam cálculos
exaustivos com os dados apresentados nas chapas fotográficas com os dados celestes. Participavam desse grupo Williamina Fleming, Antonia Maury, Cecília Gaposchkin-Payne, Annie
Jump Cannon e Henrietta Swan Leavitt, que são grandes nomes para o desenvolvimento da
Astronomia do final do século XIX e início do século XX (BARROS, 2018). Fleming catalogou muitas das estrelas do catálogo Henry Draper, assim como descobriu
diversas nebulosas. Uma das mais famosas é a Nebulosa Cabeça de Cavalo. Antonia Maury
trabalhou com análises espectroscópicas e contribuiu para as classificações estelares utilizadas
por Ejnar Hertzprung, as quais utilizamos até os dias de hoje. Cecília Payne trabalhou com
estrelas variáveis e em desenvolvimentos para os estudos de evolução estelar, e é um grande
símbolo da luta das mulheres na entrada no mundo científico. Annie Jump Cannon foi
responsável pela catalogação de milhares de estrelas do catálogo Henry Draper, e também se
tornou uma importante personagem para a entrada feminina no campo científico. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 desenvolvimentos na área contribuíram para o que sabemos e como sabemos sobre
Astronomia atualmente. As condições sociais femininas da época, por exemplo, permitiram
alguns desenvolvimentos em detrimento de outros, de forma que se tal ciência fosse construída
apenas por homens seria diferente do que hoje conhecemos (SCHIEBINGER, 2001). Assim, é
extremamente pertinente que conheçamos a história de como o conhecimento em Astronomia
foi construído, e isso necessariamente requer conhecer a história da contribuição e
protagonismo feminino nesse âmbito. Um adendo à história das calculadoras de Harvard Henrietta
Leavitt descobriu a relação período-luminosidade das estrelas variáveis cefeidas (FERNIE, 1969;
OLIVEIRA FILHO; SARAIVA, 2017), que permitiu que posteriormente Hubble tivesse fortes
evidências de que o universo estaria em expansão. Leavitt seria até mesmo indicada ao Nobel,
caso não tivesse falecido antes de ser laureada com o prêmio. O que se quer dizer com essas breves menções a tais mulheres e suas importantes
contribuições para o campo da Astronomia, é que de fato existe uma história não contada,
ocultada, ou deturpada, que as fazem parecer simples assistentes. Mas o fato é que seus Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [4] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 é feita uma leitura preliminar no material, na busca por padrões. No caso específico deste
trabalho, a fase de pré-análise consiste na seleção dos artigos, teses e dissertações a serem
analisados, e numa leitura superficial para determinação de padrões que se tornam as
categorias da etapa seguinte. A segunda etapa é a exploração do material, que consiste na categorização dos
resultados de acordo com padrões encontrados. Tais categorias podem ser determinadas a
priori ou a posteriori. Neste trabalho, as categorias foram estabelecidas a priori, de acordo com
o que foi observado durante a seleção dos materiais. E, por fim, na terceira etapa, ocorre o tratamento dos resultados, em que são feitas
inferências a respeito dos achados. Nesta pesquisa, foram realizadas inferências com relação a
como a área de estudos da História da Astronomia trata a participação feminina na construção
e desenvolvimento da área. Metodologia Esta é uma pesquisa do tipo estado do conhecimento, a qual consiste em mapear e
discutir a produção acadêmica de uma determinada área (FERREIRA, 2002), tentando
reconhecer padrões, aspectos e características das pesquisas em questão. Neste caso,
analisaremos a área da produção de trabalhos na área da História da Astronomia que levem em
conta a participação feminina na construção de tais conhecimentos científicos. Os objetos de estudo são teses dissertações brasileiras e artigos em geral que tratem de
episódios históricos com mulheres na Astronomia, ou que pelo menos as mencionem quando
não forem o alvo principal pesquisado. Foram utilizados como fonte de dados o Catálogo de
Teses e Dissertações da CAPES, o Banco de teses e dissertações de Educação em Astronomia
(BTDEA), e as revistas da área de ensino, encontradas da Plataforma Sucupira, publicados até o
ano de 2019. O referencial metodológico utilizado para tratar os resultados encontrados no estado do
conhecimento é a Análise de Conteúdo Categorial de Bardin (2016). Esta é um conjunto de técnicas de análise das comunicações visando obter, por
procedimentos sistemáticos e objetivos de descrição do conteúdo das
mensagens, indicadores (quantitativos ou não) que permitam a inferência de
conhecimentos
relativos
às condições de produção/recepção (variáveis
inferidas) destas mensagens (BARDIN, 2016, p.38). Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [5] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 Coleta e análise dos dados Inicialmente, na fase de pré-análise foram escolhidas algumas palavras-chave para
serem procuradas nas fontes de dados utilizadas (Catálogo da CAPES, BTDEA, e artigos da área
de Ensino), que são as seguintes: Williamina Fleming, Antonia Maury, Cecília Payne, Annie Jump
Cannon, Henrietta Leavitt, História da Astronomia, mulheres na Astronomia, Harvard
Observatory, Pickering. Nas revistas de língua inglesa ou espanhola, as palavras foram
traduzidas. Após isso, tais palavras-chave foram digitadas tanto no Catálogo Capes, no BTDEA,
quanto em cada uma das revistas da área de Ensino, conforme apresentadas na Plataforma
Sucupira, de acordo com o quadriênio 2013-2016. Já na fase de exploração do material, foram
encontrados apenas 5 textos, dentre milhares de artigos, dissertações e teses, descritos no
Quadro 1. Quadro 1. Artigos, dissertações e teses que trazem as contribuições femininas na Astronomia dos séculos XIX-XX
Fonte
Tipo
Nome do material
Conteúdo
Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [6] DOI: https://doi.org/10.24933/horizontes.v40i1.1298
CAPES
Dissertação
Henrietta Swan Leavitt e a relação
período-luminosidade de estrelas
variáveis
Episódio histórico
com foco em Leavitt
CAPES
Tese
A Contribuição de Annie Jump Cannon
para a Classificação Espectral de
Harvard
Episódio histórico
com foco em
Cannon
CAPES
Dissertação
O grande debate e a descoberta da
expansão do Universo
Episódio histórico
com menção à
contribuição
feminina
BTDEA
Dissertação
Variáveis cefeidas e a contribuição
feminina na ciência: recursos para o
ensino de oscilações, ondas e ópticas
Episódio Histórico
como base para
aplicação de
produto educacional
Revista História da
ciência e ensino:
construindo
interfaces
Artigo
As mulheres do Harvard College
Observatory: Henrietta Swan Leavitt - a
mulher que descobriu como medir a
distância das galáxias
Episódio histórico
com foco em Leavitt
Fonte: A autora. No catálogo Capes foram encontradas duas dissertações de mestrado e uma tese de
doutorado que tratavam de episódios históricos envolvendo as astrônomas em questão. Destes,
dois textos tratavam especificamente de um episódio histórico com protagonismo feminino,
enquanto o outro apenas mencionava Henrietta Leavitt e suas contribuições durante a
explicação de um assunto relacionado, conforme mostra o Quadro 1. Já no BTDEA, foram encontrados quatro textos, sendo que três desses apenas faziam
comentários superficiais envolvendo Henrietta Leavitt, e não chegavam a caracterizar o que se
procurava neste trabalho. O quarto texto encontrado tratava do episódio histórico em si. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Coleta e análise dos dados Com relação às revistas encontradas na plataforma sucupira na área, das 111 revistas
pesquisadas, foram selecionados apenas 38 artigos. Alguns destes tratavam de gênero,
mulheres, meninas e Astronomia, mas não falavam sobre a História da Astronomia, que é o
foco deste estudo. Desses, foram então selecionados nove artigos. Algumas resenhas de livros encontradas nessas revistas mencionam a temática, no
entanto, não são pesquisas no formato de artigo, o que mostra a pouca procura pelo tema para
pesquisa. Os artigos de Astronomia, em geral, mencionam o nome de cientistas como Cannon, Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [7] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 DOI: https://doi.org/10.24933/horizontes.v40i1.1298 Leavitt, Fleming, ao mencionar estudos como espectroscopia e catalogação estelar. No entanto,
quando se trata de um estudo histórico e epistemológico, ao que parece, simplesmente não há
espaço para essas personagens. Dos artigos selecionados, apenas um traz um estudo histórico
apresentando a contribuição feminina na História da Astronomia. A partir daí, inicia-se a fase do
tratamento dos resultados da Análise de Conteúdo. Destaca-se o fato de que três dos cinco materiais encontrados foram elaborados pela
mesma pesquisadora, e quanto aos outros dois, um não trata especificamente do processo de
construção científica, mas de uma aplicação de produto educacional utilizado conhecimentos
da área, enfatizando a presença das mulheres na área, e o outro material apresenta a
reconstrução de um episódio histórico, mas que não trazia a participação feminina como foco. Assim, é possível notar o cenário de desprestígio às produções femininas na história da
Astronomia. Materiais importantes para se avaliar a presença feminina na ciência, como o artigo
Mulheres na ciência: um estudo da presença feminina no contexto internacional (CONCEIÇÃO;
TEIXEIRA, 2018) também apareceram nesta pesquisa, mas por não trazerem o foco
epistemológico e da história das mulheres na construção da Astronomia, ficaram de fora dessa
listagem. É importante observar que em materiais como o de Conceição e Teixeira (2018), e em
muitos outros, parece que as mulheres precisam ser incluídas na ciência, que é necessário
haver uma categoria para abordar suas contribuições, como se elas fizessem uma ciência
diferente da masculina, como se fosse algo especial que merecesse condições especiais. De
nosso ponto de vista, não é dessa forma que a produção feminina deveria ser vista. As mulheres contribuíram, foram protagonistas na construção de um conhecimento que
simplesmente não é trabalhado e apresentado da mesma maneira que o conhecimento
masculino produzido, como episódios de Galileu, Newton, ou até mesmo de Hubble, que
ofereceu contribuições da mesma ordem que as mulheres de sua época. Apresenta-se como
evidência disso o fato de terem sido encontrados apenas cinco textos nessa área, em todo o
corpus da pesquisa. Dessa forma, mais estudos historiográficos são necessários nessa área, não somente ssa forma, mais estudos historiográficos são necessários nessa área, não somente Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [8] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 para se revelar a contribuição feminina, mas também para esclarecer a construção dos
conhecimentos em Astronomia moderna, que não são muito explorados, pelo que se pôde
observar nos artigos analisados. Considerações finais Os estudos relacionados à história da Astronomia que mencionam a contribuição
feminina são ínfimos. Neste trabalho, de todos os bancos de teses e artigos verificados,
encontramos apenas cinco textos que abordam a temática. No entanto, sabemos que a
contribuição feminina para a Astronomia precisaria de muito mais material do que os
encontrados para ser apresentada. Dessa forma, são necessários mais estudos e reconstruções historiográficas na área. Conforme Haraway (1989) menciona, novas histórias da ciência precisam ser contadas, mas do
ponto de vista feminino, livre dos interesses sociais e limitações da época em que esse
conhecimento foi construído. São necessários não somente estudos quanto à contribuição
feminina, mas quanto a como o conhecimento em Astronomia moderna foi e tem sido
construído. De acordo com a análise realizada, a História da Astronomia moderna não tem sido
tão
problematizada
e
nem
apresentada
adequadamente
em
estudos
históricos
e
historiográficos. Como futuras investigações, a partir dos achados, espera-se construir então histórias da
ciência a partir de outros pontos de vistas diferentes do tradicional, onde as mulheres passem a
ser protagonistas de suas histórias. Espera-se trabalhar na reconstrução de episódios históricos
nessa temática, de modo a oferecer visões mais realistas da maneira como a ciência da
Astronomia foi construída ao longo dos séculos XIX e XX, bem como evidenciando o
protagonismo feminino em tais episódios. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 Referências Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [9] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 BARDIN, L. Análise de conteúdo. Trad. Luís Antero Reto e Augusto Pinheiro. São Paulo: Edições
70, 2016. BARROS, M. C. As mulheres do Harvard College Observatory: Henrietta Swan Leavitt - a mulher
que descobriu como medir a distância das galáxias. História da Ciência e Ensino: construindo
interfaces. v. 18 (especial), p.12-21, 2018. CHASSOT, A. A ciência é masculina? é, sim senhora! Contexto e Educação, n.71/72, p.9-28, 2004. CONCEIÇÃO, J. M.; TEIXEIRA, M. R. F. Mulheres na ciência: um estudo da presença feminina no
contexto internacional. Tear: Revista de Educação Ciência e Tecn., Canoas, v.7, n.1, p.1-18, 2018. FERNIE, J. D. The period-luminosity relation: a historical review. Publications of the Astronomical
Society of the Pacific, v.81, n.483, p.707-731, 1969. FERREIRA, N. S. A. As pesquisas denominadas “estado da arte”. Educação & Sociedade, v.23,
p.257-272, 2002. HARAWAY, D. Primate visions. Londres: Routledge, 1989. HARAWAY, D. Primate visions. Londres: Routledge, 1989. LEDERMAN, N. G. Students’ and teachers’ conceptions of the nature of science: a review of the
research. Journal of Research in Science Teaching, v.29, n.4, p.331-360, 1992. LINO, T. R.; MAYORGA, C. As mulheres como sujeitos da ciência: uma análise da participação das
mulheres na ciência moderna. Saúde & Transformação Social, v.7, n.3, p.96-107, 2016. OLIVEIRA FILHO, K. S.; SARAIVA, M. F. O. Astronomia & astrofísica. 4. ed. São Paulo: Livraria da
Física, 2017. SANTOS, E. F.; SANTOS, J. O.; SANTOS, I. F. Astronomia: uma experiência em que mulheres
atuam como protagonistas. Temas em Educação, v.27, n.2, p.134-151, jul./dez. 2018. SCHIEBINGER, L. O feminismo mudou a ciência? Trad. Raul Fiker. Bauru: EDUSC, 2001. SILVA, E. R. A (in)visibilidade das mulheres no campo científico. HISTEDBR On-line, n.30,
p.133-148, 2008. SILVA, F. F.; RIBEIRO, P.R. C. Trajetórias de mulheres na ciência: “ser cientista” e “ser mulher”. Ciência e Educação, Bauru, v.20, n.2, p.449-466, 2014. VELHO, L. Prefácio. In: SANTOS, L. W.; ICHIKAWA, E. Y.; CARGANO, D. F. (org.). Ciência, tecnologia
e gênero: desvelando o feminino na construção do conhecimento. Londrina: IAPAR, 2006,
p.xiii-xviii. Periódico Horizontes – USF – Itatiba, SP – Brasil – e022023 [10] DOI: https://doi.org/10.24933/horizontes.v40i1.1298 Recebido em agosto 2021. Aprovado em abril 2022. [11]
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https://periodicos.ufsc.br/index.php/sequencia/article/download/15182/13808
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Cultura jurídica moderna, humanismo renascentista e reforma protestante
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DOAJ (DOAJ: Directory of Open Access Journals)
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*
Professor titular de “História das Instituições Jurídicas” dos cursos de graduação e pós-graduação
em Direito da UFSC. Doutor em Direito e membro do Instituto dos Advogados Brasileiros (RJ). É
pesquisador do CNPq e consultor da CAPES. Professor visitante de cursos de pós-graduação em
várias universidades do Brasil e do exterior. Autor de diversos livros, dentre os quais: Pluralismo
jurídico – Fundamentos de uma nova cultura no direito. 3. ed. São Paulo: Alfa-Omega, 2001. Antonio Carlos Wolkmer * Sumário: Introdução; 1. Origens da cultura jurídica no Ocidente; 2. Renascimento,
Reforma Protestante e Humanismo jurídico. Conclusão. Referências. Resumo: O artigo busca demonstrar que, nos
primórdios da sociedade moderna, ocorre-
ram transformações que atravessaram as es-
feras econômicas, sociais, políticas e jurídi-
cas. Neste contexto, em um horizonte de con-
tinuidade e de rupturas, forjam-se os pensa-
mentos político e jurídico modernos, influ-
enciados pela força e pela criatividade dos
movimentos culturais do Humanismo Renas-
centista e da Reforma Protestante. Abstract: This paper aims on demonstrating
that, at the beginnings of modern society, there
happened some changes that crossed the
economical, social, political and juridical
spheres. In this context, taking into account a
horizon of continuity and ruptures, political
and juridical modern thoughts are forged, since
they are influenced by force and creativity of
cultural movements from Humanism
Renaissance as well as the Protestant Reform. Palavras-chave: Humanismo renascentista;
Reforma protestante; Cultura jurídica;
modernidade; Direito Moderno. Keywords:
Humanism
Renaissance;
Protestant Reforms; juridical Culture;
modernity; Modern Law. CULTURA JURÍDICA MODERNA,
HUMANISMO RENASCENTISTA E
REFORMA PROTESTANTE Antonio Carlos Wolkmer * Introdução N N
ão há que se desconsiderar na formação das idéias modernas acerca
do Estado e do Direito o legado clássico do pensamento greco-roma-
no. Igualmente, não se pode desconhecer as teses de intérpretes de que as
origens do mundo moderno ocidental prende-se às transformações trazidas
pela Igreja Romana Ocidental (unidade e independência desencadeada Revista Seqüência, nº 50, p. 9-27, jul. 2005 por Gregório VII ante imperadores, reis e senhores feudais) entre fins
dos séculos XI e XII.1 Entretanto, o direcionamento que se assume é pelo
discurso analítico, levando-se em conta que as raízes históricas dos valo-
res político-jurídicos e das instituições modernas irão constituir-se num
período compreendido entre os séculos XIV e XVI. Em tal cenário, ins-
tauram-se a dissolução das instituições até então hegemônicas (Igreja
Romana), o aumento do poder real com o surgimento das monarquias
nacionais (França, Inglaterra), o enfraquecimento do papado, a emer-
gência do reformismo filosófico, o aparecimento cultural do humanismo
renascentista e a secularização da política.2 Antes mesmo de buscar-se as origens da cultura jurídica européia,
trata-se de reproduzir as suas condições e seu desenvolvimento no âmbito
específico das relações histórico-sociais da época e de configurar os pri-
mórdios da sociedade moderna européia engendrada ao longo de um pro-
cesso interativo de transformações estruturais que atravessa as esferas eco-
nômica, social, política, científica e religiosa. Tal dinâmica de mudanças estruturais desencadeou, conjuntamente
com o complexo e plural sistema herdado de legalidade (Direito romano,
canônico, germânico, feudal e mercantil), as bases fundantes da moderna
cultura jurídica européia. Em verdade, nesse horizonte de continuidades
e de rupturas em que se forja os pensamentos político e jurídico modernos,
é que se destacam, com muita força e criatividade, os movimentos do Huma-
nismo Jurídico e da Reforma Protestante. 1
Cf. BERMAN, Harold J. La Formación de la Tradición Jurídica de Occidente. México: Fondo
de Cultura Económica, 1996. p. 55, 289 e 352.
2
Cf. WOLKMER, Antonio C. (Org.). Introdução à História do Pensamento Político. Rio de
Janeiro: Renovar, 2003. p. 51. OLKMER, Antonio C. (Org.). Introdução à História do Pensamento Político. Rio de
: Renovar, 2003. p. 51. 3
Cf. WOLKMER, Antonio C. Pluralismo Jurídico. Fundamentos de uma Nova Cultura no Direi-
to. 3. ed. São Paulo: Alfa-Omega, 2001. p. 28-29.
4
SOMBART, Werner. El Burgués. Contribución a la Historia Espiritual del Hombre Económico
Moderno. Madrid: Alianza, 1998. p. 145.
5
DOBB, Maurice. A Evolução do Capitalismo. São Paulo: Abril Cultural, 1983. p. 51 e ss.
6
Ver: SOMBART, Werner. Op. cit., p. 115-117 e 163; WEBER, Max. A Ética Protestante e o
Espírito do Capitalismo. São Paulo: Abril Cultural, 1974. p. 205 e ss. (Os Pensadores).
7
Ver: VON MARTIN, Alfred. Sociología del Renascimiento. Mexico: Fondo de Cultura
Económica, 1946. p. 24.
8
COING. Helmuth. Derecho Privado Europeo. Tomo I. Madrid: Fundación Cultural del Notariado,
1996. p. 86 e 90. 8
COING. Helmuth. Derecho Privado Europeo. Tomo I. Madrid: Fundación Cultural del Notariado,
1996. p. 86 e 90. p
B, Maurice. A Evolução do Capitalismo. São Paulo: Abril Cultural, 1983. p. 51 e ss.
É g
p
BART, Werner. El Burgués. Contribución a la Historia Espiritual del Hombre Económic
rno. Madrid: Alianza, 1998. p. 145. B, Maurice. A Evolução do Capitalismo. São Paulo: Abril Cultural, 1983. p. 51 e ss.
SOMBART, Werner. Op. cit., p. 115-117 e 163; WEBER, Max. A Ética Protestante e OBB, Maurice. A Evolução do Capitalismo. São Paulo: Abril Cultural, 1983. p. 51 e ss.
r: SOMBART, Werner. Op. cit., p. 115-117 e 163; WEBER, Max. A Ética Protestante
d
l
l
b l
l
l
(
d
) MBART, Werner. Op. cit., p. 115-117 e 163; WEBER, Max. A Ética Protestante e o 3
Cf. WOLKMER, Antonio C. Pluralismo Jurídico. Fundamentos de uma Nova Cultura no Direi-
to. 3. ed. São Paulo: Alfa-Omega, 2001. p. 28-29.
4
SOMBART, Werner. El Burgués. Contribución a la Historia Espiritual del Hombre Económico
Moderno. Madrid: Alianza, 1998. p. 145. SOMBART, Werner. Op. cit., p. 115 117 e 163; WEBER, Max. A Ética Protestante e
ito do Capitalismo. São Paulo: Abril Cultural, 1974. p. 205 e ss. (Os Pensadores). p
p
p
(
7
Ver: VON MARTIN, Alfred. Sociología del Renascimiento. Mexico: Fondo d
Económica, 1946. p. 24.
8 1 Origens da cultura jurídica no Ocidente Preliminarmente importa registrar, no âmbito do sistema produti-
vo, a formação de um capitalismo mercantil como novo modelo de de-
senvolvimento das forças materiais em que o capital é o instrumento es-
sencial das atividades reais. Trata-se da passagem da economia agrário- 10 senhorial para a implementação da produtividade econômica de merca-
do livre, pela sistematização do comércio por meio das trocas monetári-
as e pela força de trabalho assalariado.3 Em fins da Baixa Idade Média,
inaugura-se um processo de crise e de ruptura do Feudalismo que irá
desencadear profundas transformações na vida produtiva, substituindo
a economia agrícola de servidão e de subsistência pela atividade mer-
cantil, financeira e lucrativa. Particularmente, é nas repúblicas mercan-
tis do norte da Itália, desde o século XIII, que se desenvolve o espírito
capitalista.4 Entretanto, o Capitalismo irá constituir-se gradualmente,
consolidando-se e alcançando quase toda a Europa depois dos séculos
XVI e XVII.5 Nas suas origens, a mentalidade capitalista está identificada
às práticas comerciais, ao empreendimento individualista e competitivo,
bem como ao afã de lucro ilimitado, ao cálculo previsível e ao procedi-
mento administrativo racionalizado.6 Quanto à formação da estrutura cultural social, cabe destacar o apa-
recimento de um novo segmento social que se distancia do clero e da no-
breza, adquirindo a propriedade dos meios produtivos e impondo uma
hegemonia política, construída por meio de processos revolucionários (In-
glaterra, Estados Unidos e França). Assim, a burguesia detentora do capi-
tal suplanta a nobreza e adquire o poder graças à força de seu prestígio e
de sua riqueza acumulada.7 Certamente, a burguesia representa uma clas-
se ascendente e livre no bojo da estrutura estamental então dominante, e
que prossegue até o Antigo Regime. Sua influência estará diretamente re-
lacionada à posição profissional que ocupa na cidade onde habita.8 Natu-
ralmente, como escreve Gregorio Peces-Barba, o homem burguês 11 era o habitante do burgo, da cidade, com atividades de mercador, de artesão, de
funcionário, de advogado ou de homem de letras, com direitos reconhecidos na
Carta Jurídica de seu município [...]. De burgueses, como habitantes da cidade,
passaram a ser burgueses cidadãos [...]. A burguesia mais representativa do
nascente espírito capitalista era uma burguesia da cidade, ainda que existia
[...], uma burguesia agrária e campesina, proprietária de terras [...]. 9
PECES-BARBA MARTINEZ, Gregorio (Org.). Historia de los Derechos Fundamentales. Tomo
II: Siclo XVIII. Madrid: Dykinson, 2001. p. 54-55.
10 SOMBART, Werner. El Burgués. Op. cit., p. 115, 163 e 164.
11 TOUCHARD, Jean (Org.). História das Idéias Políticas. V. II. Mira-Sintra: Europa-América,
1991. p. 14-15. HARD, Jean (Org.). História das Idéias Políticas. V. II. Mira-Sintra: Europa-América,
p. 14-15. y
p
ART, Werner. El Burgués. Op. cit., p. 115, 163 e 164. -BARBA MARTINEZ, Gregorio (Org.). Historia de los Derechos Fundamentales. Tomo
o XVIII. Madrid: Dykinson, 2001. p. 54-55. 1 Origens da cultura jurídica no Ocidente Para além da burguesia urbana que irá cada vez mais se dedicar ao
comércio e às finanças, existiu igualmente uma “burguesia profissional
formada principalmente por médicos e advogados que passaram a ter um
prestígio social e uma influência crescente [...]”.9 Adverte Werner Sombart
que, por trás de todo burguês, esconde-se a alma de um perfeito empresá-
rio capitalista que, por ser um tipo de pessoa, não se confunde com uma
classe social: “o dinheiro era seu fim, a criação de empresas seu meio;
especulava e calculava, e finalmente também se apoderaram de sua pes-
soa as virtudes burguesas [...]”.10 No que tange aos aspectos políticos, importa ter em conta o pro-
cesso de secularização e a compreensão do poder a partir das teses
absolutistas. No cenário que abarca o Antigo Regime, muitas das idéias
políticas estão deveras próximas do movimento da Reforma e de “uma
crescente nacionalização dos Estados e da Política”.11 Posteriormente,
a partir do século XVII, resultado das condições materiais emergentes,
das novas relações sociais e do clima de tolerância, aparece a doutrina
do liberalismo-individualista no continente europeu. Assim, o liberalis-
mo torna-se a manifestação mais autêntica de uma ética individualis-
ta, voltada basicamente para a noção de liberdade e que está presente
em todos os aspectos da realidade, desde o filosófico até o social, o eco-
nômico, o político, o religioso etc. Certamente, a filosofia liberal como
ideário dos interesses individuais da sociedade burguesa nascente legi-
tima para Harold Laski as reais 12 necessidades de um mundo [...]. Eram necessárias novas concepções que legi-
timassem as novas potencialidades de riqueza que os homens haviam desco-
berto, pouco a pouco, nas eras precedentes. [...] e, em função disso, desenvol-
veu-se uma nova filosofia para permitir uma justificação racional do novo
mundo que assim nascera.12 Outro movimento de referência política é a dinâmica específica de
estruturação do poder. 12 LASKI, Harold. O Liberalismo Europeu. São Paulo: Mestre Jou, 1973. p. 9, 14-15.
13 WOLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. São Paulo:
Manole, 2005. p. 86.
14 Cf. WEBER, Max. Economía y Sociedad. Mexico: Fondo de Cultura Económica. 1984. p. 1048 e 1056.
15 Ver: WOLKMER, Antonio C. Pluralismo Jurídico. p. 43.
16 WOLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. p. 86. p
WOLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. p. 86. 1 Origens da cultura jurídica no Ocidente Na verdade, “a evolução centralizadora e bu-
rocrática contribui para extinguir a fragmentação corporativa feudal,
favorecendo a criação dos Estados Modernos, unitários e seculariza-
dos”.13 Assinala Weber que o Estado Moderno materializa uma asso-
ciação humana institucionalizada, detendo o “monopólio da coação
física legítima”, fundado na economia capitalista mercantil, na buro-
cracia de agentes profissionais e na construção de uma legalidade for-
mal e racionalizada.14 A organização centralizadora de poder que se
institui sob a forma secularizada de monárquica de Estado absolutista
transforma-se no Estado nacional, liberal e representativo do século
XVIII, gerenciador das leis do livre mercado do liberalismo econômico
e tutor das relações de competição privada.15 Tais processos caracterizadores da passagem para a Modernidade
são corroborados também pelas descobertas, progressos e inovações cien-
tíficas. Naturalmente, o conhecimento científico, engendrado pelos gran-
des avanços experimentais da época (Copérnico, Galileu e Newton), su-
planta as formas dogmáticas do saber anterior.16 Quanto à dimensão teo-
lógica, assinala A. Sánchez Vazquez, a religião deixa de ser a forma ideológica dominante e a Igreja Católica perde a
sua função de guia. Verificou-se os movimentos de reforma, que destroem a
unidade cristã medieval. Na nova sociedade, consolida-se um processo de se-
paração daquilo que a Idade Média unira: a) a razão separa-se da fé (e a filoso- 13 fia, da teologia); b) a natureza, de Deus (e as ciências naturais, dos pressupostos
teológicos); c) o Estado, da Igreja; e d) o homem, de Deus.17 O certo é que a secular organização feudal é sucedida por uma estru-
tura societária marcada por múltiplas transformações em vários setores
do conhecimento e da atividade humana. Tal processo abre os horizontes
para a chamada Modernidade. Na verdade, a Modernidade pode ser com-
preendida não só como um “fluxo histórico do tempo” mas como fenôme-
no cultural que implica “certas características específicas”, vivenciadas
pela sociedade em sua estrutura organizacional.18 Há de se ver a Moder-
nidade como resultante de procedimentos mundializados de racionaliza-
ção, que ocorreram nas áreas socioeconômica, política e cultural.19 Uma vez registradas algumas das condições estruturais que edifica-
ram a sociedade moderna, avança-se, agora, no exame mais específico
das origens da cultura jurídica européia. ZQUEZ, Adolfo Sanchez. Ética. 5. ed. Rio de Janeiro: Civilização Brasileira, 1982. p. 247.
É 18 ADEODATO, João M. Ética e Retórica. Para uma Teoria da Dogmática Jurídica. São Paulo:
Saraiva, 2002. p. 205, 298-299; ROUANET, Sérgio P. Mal-Estar da Modernidade. São Paulo:
Companhia das Letras, 1993. p. 9 e 120. 17 VÁZQUEZ, Adolfo Sanchez. Ética. 5. ed. Rio de Janeiro: Civilização Brasileira, 1982. p. 247.
18 ADEODATO, João M. Ética e Retórica. Para uma Teoria da Dogmática Jurídica. São Paulo:
Saraiva, 2002. p. 205, 298-299; ROUANET, Sérgio P. Mal-Estar da Modernidade. São Paulo:
Companhia das Letras, 1993. p. 9 e 120.
19 Cf. WOLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. p. 86.
20 TIGAR, Michael E.; LEVY, Madeleine R. O Direito e a Ascensão do Capitalismo. Rio de Janeiro:
Zahar, 1978. p. 23-24.
21 WIEACKER, Franz. História do Direito Privado Moderno. Lisboa: Calouste Gulbenkian, 1980. p. 15. 19 Cf. WOLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. p. 86.
20 TIGAR, Michael E.; LEVY, Madeleine R. O Direito e a Ascensão do Capitalismo. Rio de Janeiro:
Zahar, 1978. p. 23-24.
21 p
OLKMER, Antonio C. Fundamentos do Humanismo Jurídico no Ocidente. p. 86. 21 WIEACKER, Franz. História do Direito Privado Moderno. Lisboa: Calouste Gulbenkian, 1980. p. 15. 21 WIEACKER, Franz. História do Direito Privado Moderno. Lisboa: Calouste Gulbenkian, 1 Origens da cultura jurídica no Ocidente Inegavelmente, a combinação de algumas tradições legais advindas da
Alta Idade Média, como o Direito romano, o Direito canônico, as práticas
consuetudinárias germânica e mercantil, bem como a doutrina filosófica do
direito natural, contribuiu para a formação da cultura jurídica moderna.20 Também merece referência a interpretação de Franz Wieacker de que
os primórdios do Direito Moderno europeu presente nas sociedades de
tradição romano-germânica deve-se, sobretudo, aos “restos da organiza-
ção do império romano do ocidente, a igreja romana e a tradição escolar
da antigüidade tardia”.21 O primeiro elemento a ser destacado é o fato de
que, mesmo com sua queda posterior, o Império Romano legou uma for-
ma de burocracia, de organização administrativa e financeira, que será
apropriada e reproduzida pelas nações em gestação, que buscavam sua 14 unidade política. Tais povos irão aprender com os romanos que o Direito
pode também ser “uma criação do poder do Estado e uma tradição cultu-
ral”.22 O segundo fator determinante está representado pela Igreja Ro-
mana, que com a derrocada de Roma, assumiu, com eficácia, “muitas das
tarefas públicas, sociais e morais” do antigo Império. Seus dignatários re-
ligiosos acabaram ocupando a “administração, a autoridade, a cultura, a
jurisdição e as técnicas documentais, processuais e notariais das autorida-
des seculares.”23 Por último, “a organização pedagógica e escolar da an-
tigüidade tardia sobreviveu, apesar da repressão cultural e da crise exter-
na das invasões bárbaras”, influenciando o ensino elementar na base do
estudo do trivium: gramática, lógica (dialética) e retórica.24 Assim, o des-
dobramento desses antecedentes contribuíram para o desenvolvimento
futuro de “uma nova consciência jurídica européia” e o despertar do sen-
timento de unidade comum das novas classes dirigentes.25 Neste novo cenário de rupturas e de gradual secularização, vislum-
bram-se o declínio da canonística católica e a força das fontes romanísticas, não só nas instituições legais de tradição latina, mas também nas práticas judiciais
dos Países Baixos, marcados por seculares experiências consuetudinárias. 22 Idem, p. 16.
23 Idem, p. 17.
24 Idem, p. 19.
25 WIEACKER, Franz. Op. cit., p. 20-21.
26 WOLKMER, Antonio C. Pluralismo Jurídico. p. 47. 22 Idem, p. 16.
23 Idem, p. 17.
24 Idem, p. 19.
25 WIEACKER, Franz. Op. cit., p. 20-21.
26 WOLKMER, Antonio C. Pluralismo Jurídico. p. 47. p
p
26 WOLKMER, Antonio C. Pluralismo Jurídico. p. 47. 1 Origens da cultura jurídica no Ocidente O avan-
ço crescente e o amplo reconhecimento do legado jurídico romano demonstram que
suas preceituações, mais do que as difusas e múltiplas práticas legais medievais,
eram mais adequadas para um período que se instaurava sob o prisma da centra-
lização política, da administração burocrática e da racionalização do poder.26 Clara está que a nascente ciência jurídica moderna não só se revela
como produção de uma específica formação social e econômica mas prin-
cipalmente, consolida-se no processo de junção histórica entre a legalida-
de estatal e a centralização burocrática. O ápice teórico de convergência
entre a unicidade do poder político e a nova ordenação do Direito pode
ser encontrado na filosofia política de pensadores da época, como Thomas 15 Hobbes.27 Certamente, assinala-se que Hobbes não é apenas um dos cons-
trutores do moderno Estado absolutista, mas igualmente um dos primei-
ros intérpretes a identificar o Direito como manifestação do Direito do
soberano. Tratava-se da tendência, que acabará sendo predominante, do
Direito identificado com a legislação posta pela autoridade revestida do
poder máximo e, ainda mais, o Direito como criação do Estado. Assim,
um dos traços marcantes do Direito Moderno emergente entre os séculos
XVI e XVII está na íntima relação do Direito com o poder estatal e na sua
identificação com a lei escrita. Trata-se da instrumentalização do jurídico
como significação dos interesses da burguesia e da dinâmica produtiva capi-
talista. Na verdade, como lembra Capella, o Direito Moderno advindo das sociedades do capitalismo concorrencial, apesar de tomar muitas de suas
categorias do Direito romano [...], difere radicalmente de todo direito anterior
por um projeto de sistematização, de exatidão, de fiabilidade [...], do qual
carece o direito pré-moderno.28 sociedades do capitalismo concorrencial, apesar de tomar muitas de suas
categorias do Direito romano [...], difere radicalmente de todo direito anterior
por um projeto de sistematização, de exatidão, de fiabilidade [...], do qual
carece o direito pré-moderno.28 Ora, o Direito da sociedade moderna é produto de uma construção. “É um direito legislado por parlamentares e interpretado por tribunais,
acadêmicos e juristas práticos.”29 Sem dúvida, trata-se de um tempo his-
tórico em que se intercalam a continuidade de tradições clássicas e medie-
vais com a ruptura e a inovação do mundo moderno. Nesse interregno de
continuidades e de rupturas, emergem novos processos culturais e cientí-
ficos movimentos do Humanismo renascentista e da Reforma Protestante. 27 WOLKMER, Antonio C. Pluralismo Jurídico. p. 48 e 50. p
Direito e do Estado. Porto Alegre: Livraria do Advogado, 2002. p. 130.
29 Ibidem, p. 131. p
28 CAPELLA, Juan Ramón. Fruto Proibido. Uma aproximação histórico-teórica ao , J
p
ç
Direito e do Estado. Porto Alegre: Livraria do Advogado, 2002. p. 130.
29 Ibidem, p. 131. J
p
28 CAPELLA, Juan Ramón. Fruto Proibido. Uma aproximação histórico-teórica ao Estudo do
Direito e do Estado. Porto Alegre: Livraria do Advogado, 2002. p. 130.
29 Ibidem, p. 131. 29 Ibidem, p. 131. p
uan Ramón. Fruto Proibido. Uma aproximação histórico-teórica ao Estudo do 30 Cf. PECES-BARBA MARTINEZ, Gregorio (Org.). História de los Derechos Fundamentales. Tomo
I: Transito a la modernidad. Siglos XVI – XVII. Madrid: Dykinson, 1998. p. 66; FASSÒ, Guido.
Historia de la Filosofía del Derecho. La Edad Moderna. Vol. 2. Madrid: Piramide, 1982. p. 21.
31
Ò p
p
32 TAVARES, José A. Giusti (Org.). Coletânea de Textos Históricos. História do Pensamento
Humano. São Leopoldo: Unisinos, 1973. p. 206. Cf. FASSÒ, Guido. Op. cit., p. 27. 2 Renascimento, Reforma Protestante e Humanismo jurídico Certamente que os arbores iniciais do mundo moderno são gestados
por um processo crescente de secularização e racionalização, proveniente
de fenômenos culturais como o Humanismo do Renascimento e a Refor- 16 ma Protestante. A crise e a derrocada do universo medieval na Europa
central no âmbito da religião, da filosofia, da economia e da política de-
sencadearam os ingredientes para uma nova mentalidade, um novo pen-
samento e novos procedimentos científicos. As emergentes formas cultu-
rais marcadas pelo espírito de ruptura, naturalismo e individualidade es-
tão impregnadas por uma visão clássica do mundo, expressa no que se
convencionou designar Renascimento. A gradual substituição das relações de produção feudal, a formação
da instância estatal nacional-soberana e o ímpeto para uma visão norteado-
ra dominada pela individualidade estão presentes neste momento de
criatividade e de “renascimento” do espírito humano.30 Ora, esse movimento cultural e humanista inicia-se, principalmente,
na Itália do século XV, alcançando, no século seguinte, a culminância de
seu apogeu.31 O Renascimento é a celebração do humano como força autô-
noma e racional, desvinculada de todas as restrições transcendentais que
inviabilizam a criatividade do pensamento e a liberdade da prática objetiva. Naturalmente, o Renascimento se orienta e ordena, como assinala
Giusti Tavares, Giusti Tavares, todas as dimensões da existência humana no sentido antropocêntrico, terreno,
ao mesmo tempo naturalista e humanista, mas, sobretudo, profundamente in-
dividualista e racionalista. O Renascimento rompeu com a concepção medie-
val, teológico-natural, da sociedade como uma ordem natural presidida por
Deus, fundada na fixidez e no imobilismo estamental de status e papéis [...] e
substituiu-a por uma concepção [...] voluntarista da sociedade como constru-
ção histórico-social, cultural e artificial do homem.32 todas as dimensões da existência humana no sentido antropocêntrico, terreno,
ao mesmo tempo naturalista e humanista, mas, sobretudo, profundamente in-
dividualista e racionalista. O Renascimento rompeu com a concepção medie-
val, teológico-natural, da sociedade como uma ordem natural presidida por
Deus, fundada na fixidez e no imobilismo estamental de status e papéis [...] e
substituiu-a por uma concepção [...] voluntarista da sociedade como constru-
ção histórico-social, cultural e artificial do homem.32 Assim, é indiscutível o significado do Renascimento como força de
secularização e inovação, capaz de influenciar a vida social e cultural da 17 época. 33 Idem. Observar também: BURCKHARDT, Jacob. O Renascimento Italiano. Lisboa: Editorial
Presença, 1973. p. 137 e ss.
34 PECES-BARBA MARTINEZ, Gregorio (Org.). Op. cit., Tomo I, Siglos XVI – XVII, p. 112 e 117.
35 FASSÒ, Guido. Op. cit., p. 38.
36 TAVARES, José A. Giusti. Op. cit., p. 209. 2 Renascimento, Reforma Protestante e Humanismo jurídico Compreende-se, desse modo, o papel do Renascimento para liber-
tar não só a sociedade mas também a mundialidade da política e da eco-
nomia do controle da ética e da teologia tradicionais.33 Para além do Renascimento, a Reforma Protestante será um segun-
do evento que influenciará a sociedade, a cultura e o pensamento, rom-
pendo com a civilização medieval e contribuindo para o trânsito ao mun-
do liberal moderno.34 A Reforma, em seu sentido religioso e sem minimizar seus efeitos de
natureza social e política, está, como observa Guido Fassò, “intimamente
unida ao voluntarismo e ao antiintelectualismo; se inspira, realmente, nos
grandes defensores do primado absoluto da fé e da graça, como São Paulo
e Santo Agostinho, em contraposição aos teólogos escolásticos” que havi-
am salvado e revalorizado o racionalismo.35 O movimento da Reforma Protestante, principalmente sob o viés do
luteranismo, expressa o enfrentamento medievalista “às transformações
do mundo moderno; como uma tentativa de recuo e de restauração nos-
tálgica de uma ordem irremediavelmente perdida.” Ainda como relembra
Giusti Tavares, a Reforma em seu aspecto geral (compreendendo o pró-
prio calvinismo) rejeita categoricamente a atmosfera renascentista, nutrin-
do “uma profunda aversão à razão e ao racionalismo [...]. E é precisa-
mente a partir de uma perspectiva irracionalista que Lutero criticara,
num primeiro momento, a teologia e a filosofia escolástica e, logo, todo o
racionalismo renascentista”.36 Eis porque, por seu radicalismo, a Refor-
ma Protestante revela-se antítese ao movimento do humanismo, este como
expressão do naturalismo, do secularismo e da sua aspiração mundia-
lizada. Ora, o caráter intolerante e teocrático da primeira fase da Refor-
ma Protestante é incompatível ao ideário do humanismo renascentista
dos séculos XV e XVI. 37 FASSÓ, Guido. Op. cit., p. 39.
38 Idem.
39 Idem.
40 Cf. PECES-BARBA MARTINEZ, Gregorio (Org.). Op. cit., Tomo I, Siglos XVI – XVII, p. 118-119.
41 FASSÓ, Guido. Op. cit., p. 39. 37 FASSÓ, Guido. Op. cit., p. 39.
38 2 Renascimento, Reforma Protestante e Humanismo jurídico Naturalmente, 18 a negação do livre arbítrio e a afirmação da predestinação se achavam em franco
contraste com o espírito do Humanismo e do Renascimento, para quem o homem
era o primeiro e absoluto valor, o único e livre artífice de seu próprio mundo [...].37 Reconhece Guido Fassò que as formulações teóricas dos primeiros
reformadores, principalmente Lutero e Calvino, detêm traços que se po-
deriam caracterizar como “medievais”, ou seja, “concepção religiosa e ecle-
siástica da vida, intolerância, e, em política, teocracia”.38 Entretanto, ad-
mite-se, posteriormente, uma segunda fase da Reforma Protestante, em
que, herdando ventos flexíveis do espírito renascentista, inclina-se por
posturas mais claramente modernas, “promovendo a tolerância religiosa
e política, e favorecendo o desenvolvimento das idéias liberais”.39 O certo é que uma possível aproximação posterior do novo protestan-
tismo com o humanismo encontra guarida na inclinação comum pelo indi-
vidualismo e pela relevância do sujeito individual no contexto da sociedade. Obviamente que todo esse impulso do protestantismo, voltado para a inici-
ativa individual, deve-se, como destaca Gregorio Peces-Barba, ao processo
de ruptura com a autoridade da Igreja e com a adesão ao livre exame.40 Quanto ao tratamento dispensado pelo protestantismo ao tema da
legalidade e da justiça, importa ter presente a inspiração norteadora das
fontes paulinianas e agostinianas. Os principais representantes da Refor-
ma, como Lutero, Zuinglio e Calvino, compartilham com a noção de um
jusnaturalismo teocêntrico, expressão captada pela consciência moral in-
dividual e munida pela rejeição natural à lei objetiva. Para Martinho Lutero,
a ação humana direcionada à obediência das leis não tem valor para a
salvação, de que só é instrumento a fé. A Justiça cristã não vem da lei,
pois, quem pensa de forma distinta são os juristas, legalistas e papistas. “As leis de Deus [...] são a presença em nossos corações do Espírito Santo,
que infunde o amor, que não deriva da lei”.41 Já a justiça, na interpreta-
ção de Lutero, está identificada com a perfeição, decorrendo “da graça e da 19 fé, a que o homem possui parte do espírito de Deus”.42 De qualquer forma,
justifica-se a existência do Direito positivo e do poder político estatal em
razão de a natureza humana estar dominada pela corrupção. Fica claro,
assim, o desprezo do luteranismo pela legalidade temporal objetiva. A ver-
dadeira lei é aquela que o cristão, sem a mediação dos cléricos, absorve uni-
camente das Sagradas Escrituras. 42 Ibidem, p. 40.
43 TRUYOL Y SERRA, Antonio. História de la Filosofia del Derecho y del Estado. v. 2. Madrid:
Revista de Occidente, 1976. p. 38; RUIZ MIGUEL, Alfonso. Una Filosofia del Derecho en Mode-
los Históricos. De la antigüedad a los inicios del constitucionalismo. Madrid: Trotta, 2002. p. 176.
44 FASSÓ, Guido. Op. cit., p. 45-46. 2 Renascimento, Reforma Protestante e Humanismo jurídico O Direito materializado na sociedade tor-
na-se um instrumento para regrar e para estabelecer limites aos cristãos
desgarrados e de mau comportamento, pois os bons não necessitam da le-
galidade temporal. O voluntarismo e a intolerância de Lutero favoreceu um
ideal social e político de fisionomia autoritária e conservadora.43 Muito próximo das formulações de Lutero, aparecem as doutrinas
do suíço Zuinglio e do francês Calvino que se mantêm na tradição da
espiritualidade cristã, na liberdade de consciência individual e na rejeição
da subordinação do homem à lei positiva temporal. A conduta justa não
está necessariamente no acatamento da lei humana, mas na fé e na revela-
ção. Assim, a norma de conduta para os reformadores clássicos não se
encontra no poder político externo, tampouco num plano de interiorização
subjetiva e racional, mas deduzida de uma consciência moral como “tes-
temunho da vontade de Deus”.44 Certamente que a Reforma, representada por Lutero, Zuinglio e
Calvino, ainda que considerando a perceptível dissonância interpretativa
entre estes, impõe a assertiva político-religiosa de que a aspiração maior
era a autonomia do sujeito moral à subordinação da lei temporal. Trata-se
de reordenar a própria noção de liberdade em seu sentido individual que
não se prendia à autoridade ou ao poder, mas à liberdade religiosa, a li-
berdade que rompe com o pecado. Vê-se, assim, que o que movia os refor-
madores – guardado suas diferenças e complementaridade – não era a
liberdade do homem político, mas a do sujeito cristão, inspirado nas Sa-
gradas Escrituras; era a autonomia do cristão diante da lei objetiva e sua 20 submissão à lei como expressão que revela a vontade de Deus. Entende-se,
desse modo, a ênfase da Reforma em favorecer uma condição que implica a
volta do sujeito às regras da obediência ditada pela autoridade religiosa, por
vezes mais vigorosa e ortodoxa que em grande parte da Idade Média.45 Ademais, as raízes religiosas do reformismo estão presentes num pen-
samento moderno acerca do Direito e do Estado. Se as idéias de Lutero
causaram um impacto maior nos rumos do movimento reformista, a obra
de Calvino foi melhor adequada para o mundo jurídico. 45 Idem, ibidem.
46 ALVAREZ CAPEROCHIPI, Jose A. Reforma Protestante y Estado Moderno. Madrid:
Civitas, 1986. p. 19.
47 Ibidem, p. 20 e 25. Sobre as concepções doutrinárias de Lutero e de Calvino sobre o Direito e o
Direito Natural, consultar também: VILLEY, Michel. La Formation de la Pensée Juridique
Moderne. Paris: Montchretien, 1975. p. 279-307 e 308-337.
48 Cf. RUIZ MIGUEL, Alfonso. Op. cit., p. 175. 2 Renascimento, Reforma Protestante e Humanismo jurídico A influência mais
direta de Calvino, passada por meio da produção de seus discípulos, inse-
re-se “na formação das categorias jurídicas modernas, na origem e no de-
senvolvimento da economia capitalista e na organização política moder-
na [...]”.46 A questão jurídica irá ser tratada de forma distinta pelos dois
expoentes da Reforma. Enquanto o teólogo Lutero desprezava o jurídico e
detestava os juristas tanto quanto a filosofia de Aristóteles e a escolástica
metafísica, Calvino, formado em Direito, aplica no estudo da Bíblia o mé-
todo exegético do mundo jurídico. Trata-se de um jurista que, em Gene-
bra, incorpora e leva adiante os propósitos da Reforma naquilo que Lutero
desconsiderava: a organização da Igreja reformada.47 Ainda que, de um lado, inúmeros teóricos admitam uma certa difi-
culdade em assinalar pontos de interação entre a Reforma e o Humanismo
Renascentista, por outro, não se pode deixar de reconhecer a influência
do protestantismo na gênese do Capitalismo moderno, na formulação da
mentalidade livre individualista, na valoração da consciência moral, na
contribuição da filosofia dos direitos humanos e, fundamentalmente, no
impulso para a moderna concepção de jusnaturalismo. Essa também é a interpretação de Alfonso Ruiz Miguel, para quem
não se pode compreender o jusnaturalismo racionalista sem ter presente o
impacto da Reforma Protestante.48 Além disso, recorda Gregorio Peces- 21 Barba que os grandes teóricos do jusnaturalismo moderno tendem a ser
pertencentes à religião protestante, ainda que não “associados às Igrejas
protestantes majoritárias, luteranismo e calvinismo ortodoxos, senão a se-
tores dissidentes e heterodoxos”.49 Ora, não se pode duvidar dessa influ-
ência direta nas origens do jusnaturalismo moderno, tampouco o fato de
que “a Reforma preparará o positivismo voluntarista e será um elemento
importante nos novos fundamentos da relação entre Direito e Poder”.50
Inegavelmente, toda a problemática própria “do mundo moderno e da
filosofia dos direitos fundamentais não teria sido possível sem este passo
prévio, sem esta secularização do Estado e do Direito, no que foi essencial
a contribuição da Reforma Protestante”.51 Naturalmente, a busca por instaurar uma nova sociedade e um novo
ethos cultural na direção da modernidade não está somente associada à
Reforma Protestante, pois o humanismo renascentista torna-se talvez, com
mais força, um dos forjadores do pensamento político e jurídico moderno. 49 PECES-BARBA MARTINEZ, Gregorio (Org.). Op. cit., tomo I, Siglos XVI – XVII, p. 138.
50 Ibidem, p. 141.
51 Ibidem, p. 145.
52 Ibidem, p. 68-69. p
52 Ibidem, p. 68-69. p
51 Ibidem, p. 145.
52 2 Renascimento, Reforma Protestante e Humanismo jurídico Em tais movimentos, ainda que marcados por especificidades próprias e
passado um primeiro momento conflitivo, emergem algumas preocupa-
ções que se interagem, pois não se pode desconsiderar que “o Humanismo
está presente em amplos setores da Reforma, e o espírito reformista mar-
cará também o Humanismo e seus herdeiros nos séculos XVII e XVIII”.52 É certo, assim, que o Humanismo como ampla manifestação
transformadora nos campos da cultura, da filosofia, das artes e das ciên-
cias proclama os valores que enaltecem o indivíduo, sua vontade, capa-
cidade e liberdade de ação. É a participação direta do homem e o exercí-
cio da crítica direcionada para as mudanças da vida social e política. Obviamente que surgiram tendências dentro do movimento humanista,
diferenças que se distinguiam entre si quer pela tradição filosófica da Antigüidade a que se ligavam (platonismo,
aristotelismo), quer pela temática que abordavam de preferência (estudo da natu- 22 reza, da história, da personalidade humana, da matéria religiosa), quer pela prá-
tica a que se dedicavam (política, pesquisa científica, arte, poesia).53 O ideal transcendente e a perspectiva teocêntrica da Idade Média
são sucedidos pelo racionalismo e pelo antropocentrismo. O humanismo como movimento dos mais significativos e fecundos
em trânsito para os tempos modernos trás em si, como assevera Abelardo
Levaggi, uma forte reação ao princípio da autoridade (teológica e eclesiás-
tica), deslocando a valoração para a autoridade que provém da antigüidade
clássica, e para a busca de conhecimento alcançado por meio da razão e
por preocupações dominadas pela secularização da cultura.54 Trata-se de
consagrar um ideário ideológico de emancipação que serve a um novo
segmento social ascendente (a burguesia), porquanto faz a defesa de
“verdades humanas gerais [...], fundada na capacidade individual e nas
forças próprias de cada indivíduo”, representando “a negação de todos
os privilégios das diferentes ordens, de todas as pretendidas prerrogati-
vas de nascimento e Estado”, e substituindo a “doutrina, mantida pelo
clero, dos poderes sobrenaturais [...]”.55 A orientação humanista incidiu, como não poderia deixar de ser, no
âmbito da teoria e da prática jurídica. Assim, o humanismo no Direito
contribuiu para uma natural e clássica revisão crítica da cultura jurídica
produzida pela Idade Média. 53 SEVCENKO, Nicolau. O Renascimento. São Paulo: Atual; Campinas: Unicamp, 1984. p. 14, 15 e 17.
54 Cf. LEVAGGI, Abelardo. Manual de Historia del Derecho Argentino. Tomo I. Parte General.
Buenos Aires: Depalma, 1991. p. 95-97.
55 VON MARTINS, Alfred. Op. cit., p. 46-47.
56 COING, Helmut. Elementos Fundamentais da Filosofia do Direito. Porto Alegre: Sergio A.
Fabris, 2002. p. 55-57. 53 SEVCENKO, Nicolau. O Renascimento. São Paulo: Atual; Campinas: Unicamp, 1984. p. 14, 15 e 17.
54 Cf. LEVAGGI, Abelardo. Manual de Historia del Derecho Argentino. Tomo I. Parte General.
Buenos Aires: Depalma, 1991. p. 95-97.
55 57 ANDRADE, Fabio S. Da Codificação. Porto Alegre: Livraria do Advogado, 1997. p. 38.
58 COSTA, Mário J. de Almeida. História do Direito Português. Coimbra: Almeida, 1999. p. 322.
59 CAVANNA, Adriano. Storia del Diritto Moderno in Europa. Milano: Giuffrè, 1982. p. 178;
VILLEY, Michel. Op. cit., p. 507-509.
60 COSTA, Mário J. Almeida. Op. cit., p. 324.
61 ANDRADE, Fabio S. Op. cit., p. 41. A, Mário J. de Almeida. História do Direito Português. Coimbra: Almeida, 1999. p. 322.
NNA, Adriano. Storia del Diritto Moderno in Europa. Milano: Giuffrè, 1982. p. 178;
Y, Michel. Op. cit., p. 507-509. DRADE, Fabio S. Da Codificação. Porto Alegre: Livraria do Advogado, 1997. p. 38. 2 Renascimento, Reforma Protestante e Humanismo jurídico O próprio eixo explicativo de sustentação
da doutrina do Direito Natural começa a se deslocar para a sociedade
humana e para a natureza racional do homem, antecipando-se, assim, o
que seria mais tarde a proposição filosófica do contrato social.56 Com ra-
zão entende Fábio S. de Andrade que os humanistas saíram em defesa de
múltiplas reformas, dentre elas, aquelas pertencentes ao campo da juridi-
cidade, consubstanciada num amplo questionamento que abrangia a uti-
lização do latim, a metodologia “de ensino praticada pelos Comentadores” 23 e até o interesse maior “pela sistematização dos princípios jurídicos [...]”.57
Parte dessa censura dos humanistas ao Direito voltava-se contra a tradi-
ção do bartolismo e denunciava a obra dos Comentadores, chamando a
atenção para o estudo erudito das fontes normativas romanas. Observa-
se, nessa linha destacada por Mario J. Almeida Costa, que o humanismo
jurídico constitui-se, deveras, “sob diversas tendências: desde as filológico-
críticas, orientadas para o estudo e reconstrução dos textos clássicos, até
a que reivindica a liberdade e autonomia do jurista na exegese da lei
[...]”, reportando-se “ao livre exame das fontes romanas”.58 Tal postura
correspondeu a uma mudança radical ante a hermenêutica dos
Comentadores. Diversos estudiosos reconhecem que, em seu apogeu no
século XVI, a Escola do Humanismo Jurídico teve, dentre seus principais
expoentes, nomes como o do italiano Andrea Alciato (1492-1550), o ale-
mão Ulrico Zasio (1461-1535) e os franceses Guilherme Budé (1467-1540)
e Jacques Cujas, ou Cujácio (1522-1590). No dizer de Cavanna, Alciato
foi o verdadeiro fundador e a grande “personificação do ideal
humanístico da jurisprudência”, enquanto que Cujácio foi o “maior na
exegese histórica e na filologia dos textos romanos”.59 No que se refere à controvérsia que envolveu a erudição dos humanis-
tas e o pragmatismo dos comentadores, destaca Mário Almeida Costa que,
por serem demasiado especulativos, os humanistas construíram, sobretudo,
“um ‘direito teórico’, de tendência erudita, enquanto os processos dos
Comentadores levaram a um ‘direito prático’”, ou seja, empregava-se a
romanística direcionada para “encontrar soluções para os casos concretos”.60 Indubitavelmente, cumpre registrar na esteira de Fabio S. Conclusão Os primórdios da cultura jurídica moderna no âmbito da civiliza-
ção européia reproduziram as condições e o desenvolvimento de um pro-
cesso particular de transformações que atravessou diferentes campos de
atuação humana, seja religioso e cultural, seja econômico, social, políti-
co e científico. Algumas características da época, como a secularização,
a racionalização, a individualidade e o antropocentrismo, marcam a
passagem para a construção e consolidação de um novo mundo que le-
gitima também uma nova forma de produzir, pensar e praticar o Direi-
to. Assim, a “nova consciência jurídica européia” nasce da convergên-
cia histórica do naturalismo, da individualidade e da centralização polí-
tica burocrática. A par de toda essa dinâmica específica, a nascente cul-
tura jurídica eurocêntrica está profundamente afetada por fenômenos
radicais e criadores que têm suas raízes no Humanismo renascentista e
na Reforma Protestante. Tanto um quanto o outro, desses movimentos,
exerceram uma influência direta nas instituições jurídicas e na moderna
doutrina dos direitos fundamentais. 62 Idem, ibidem. Vide também: COSTA, Mário J. Almeida. Op. cit., p. 324. 2 Renascimento, Reforma Protestante e Humanismo jurídico de Almeida
que o movimento do humanismo jurídico primou por duas direções: “de
um lado, a crítica severa pela ausência de método dos juristas que o ante-
cederam; de outro, a necessidade de preservação do Corpus Juris como
depositário da matéria jurídica européia”.61 24 Resta mencionar, finalmente, que ainda que seu período tenha sido
efêmero, deixaram diretrizes inestimáveis, pois foram os humanistas ju-
rídicos os pioneiros “a se dedicar a construir um edifício sistemático a
partir do ius commune”.62 Referências ADEODATO, João M. Ética e Retórica. Para uma Teoria da Dogmática Jurídica. São
Paulo: Saraiva, 2002. ADEODATO, João M. Ética e Retórica. Para uma Teoria da Dogmática Jurídica. São
Paulo: Saraiva, 2002. ALVAREZ CAPEROCHIPI, José A. Reforma Protestante y Estado Moderno. Madrid: Civitas, 1986. ANDRADE, Fabio S. Da Codificação. Porto Alegre: Livraria do Advogado, 1997. 25 BERMAN, Harold J. La formación de la Tradición Juridica de Occidente. México:
Fondo de Cultura Económica, 1996. BURCKHARDT, Jacob. O Renascimento Italiano. Lisboa: Editorial Presença, 1973. CAPELLA, Juan Ramón. Fruto Proibido. Uma aproximação histórico-teórica ao
Estudo do Direito e do Estado. Porto Alegre: Livraria do Advogado, 2002. CAVANNA, Adriano. Storia del Diritto Moderno in Europa. Milano: Giuffrè, 1982. COING, Helmut. Derecho Privado Europeo. Tomo I. Madrid: Fundación Cultural
del Notariado, 1996. __________. Elementos Fundamentais da Filosofia do Direito. Porto Alegre: Sergio
A. Fabris, 2002. COSTA, Mário J. de Almeida. História do Direito Português. Coimbra: Almeida, 1999. DOBB, Maurice. A Evolução do Capitalismo. São Paulo: Abril Cultural, 1974. (Os Pensadores). FASSÒ, Guido. Historia de la Filosofía del Derecho. La Edad Moderna. Vol. 2. Madrid: Piramide, 1982. LASKI, Harold. O Liberalismo Europeu. São Paulo: Mestre Jou, 1973. LEVAGGI, Abelardo. Manual de Historia del Derecho Argentino. Tomo I. Parte
General. Buenos Aires: Depalma, 1991. PECES-BARBA MARTINEZ, Gregorio (Org.). História de los Derechos
Fundamentales. Tomo I: Transito a la modernidad. Siglos XVI – XVII. Madrid:
Dykinson, 1998. __________. Historia de los Derechos Fundamentales. Tomo II: Siclo XVIII. Madrid:
Dykinson, 2001. ROUANET, Sérgio P. Mal-Estar da Modernidade. São Paulo: Companhia
das Letras, 1993. RUIZ MIGUEL, Alfonso. Una Filosofía del Derecho en Modelos Históricos. De la
antigüedad a los inicios del constitucionalismo. Madrid: Trotta, 2002. SEVCENKO, Nicolau. O Renascimento. São Paulo: Atual; Campinas:
Unicamp, 1984. 26 SOMBART, Werner. El Burgués. Contribución a la Historia Espiritual del Hombre
Económico Moderno. Madrid: Alianza Editorial, 1998. TAVARES, José A. Giusti (Org.). Coletânea de Textos Históricos. História do
Pensamento Humano. São Leopoldo: Unisinos, 1973. TIGAR, Michael E.; LEVY, Madeleine R. O Direito e a Ascensão do Capitalismo. Rio de Janeiro: Zahar, 1978. TOUCHARD, Jean (Org.). História das Idéias Políticas. v. II. Mira-Sintra:
Europa-América, 1991. TRUYOL Y SERRA, Antonio. História de la Filosofía del Derecho y del Estado. V. 2. Madrid: Revista de Occidente, 1976. VÁZQUEZ, Adolfo Sanchez. Ética. 5. ed. Rio de Janeiro: Civilização Brasileira, 1982. VILLEY, Michel. La Formation de la Pensée Juridique Moderne. Paris:
Montchretien, 1975. VON MARTIN, Alfred. Sociología del Renascimiento. Referências México: Fondo de Cultura
Económica, 1946. WEBER, Max. A Ética Protestante e o Espírito do Capitalismo. São Paulo: Abril
Cultural, 1974. (Os Pensadores). WEBER, Max. Economía y Sociedad. México: Fondo de Cultura Económica, 1984. WIEACKER, Franz. História do Direito Privado Moderno. Lisboa: Calouste
Gulbenkian, 1980. WOLKMER, Antonio C. Pluralismo jurídico. Fundamentos de uma Nova Cultura
no Direito. 3. ed. São Paulo: Alfa-Omega, 2001. __________. (Org.). Introdução à História do Pensamento Político. Rio de Janeiro:
Renovar, 2003. __________. (Coord.). Fundamentos do Humanismo Jurídico no Ocidente. São
Paulo: Manole, 2005. 27
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https://projecteuclid.org/journals/electronic-journal-of-statistics/volume-18/issue-1/Efficient-change-point-detection-and-estimation-in-high-dimensional-correlation/10.1214/24-EJS2221.pdf
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Efficient change point detection and estimation in high-dimensional correlation matrices
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Electronic journal of statistics
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Efficient change point detection and
estimation in high-dimensional
correlation matrices Zhaoyuan Li1
and Jie Gao2 2School of Data Science, The Chinese University of Hong Kong, Shenzhen
e-mail: jiegao2@link.cuhk.edu.cn Abstract: This paper considers the problems of detecting a change point
and estimating the location in the correlation matrices of a sequence of high-
dimensional vectors, where the dimension is large enough to be comparable
to the sample size or even much larger. A new break test is proposed based
on signflip parallel analysis to detect the existence of change points. Fur-
thermore, a two-step approach combining a signflip permutation dimension
reduction step and a CUSUM statistic is proposed to estimate the change
point’s location and recover the support of changes. The consistency of the
estimator is constructed. Simulation examples and real data applications
illustrate the superior empirical performance of the proposed methods. Es-
pecially, the proposed methods outperform existing ones for non-Gaussian
data and the change point in the extreme tail of a sequence and become
more accurate as the dimension p increases. Supplementary materials for
this article are available online. MSC2020 subject classifications: Primary 62M10, 62H15; secondary
62P20. Keywords and phrases: Change point detection, change point estima-
tion, signflip parallel analysis, correlation matrix, CUSUM. Received March 2023. arXiv: 2311.02312 Electronic Journal of Statistics
Vol. 18 (2024) 942–979
ISSN: 1935-7524
https://doi.org/10.1214/24-EJS2221 Electronic Journal of Statistics
Vol. 18 (2024) 942–979
ISSN: 1935-7524
https://doi.org/10.1214/24-EJS2221 Electronic Journal of Statistics
Vol. 18 (2024) 942–979
ISSN: 1935-7524
https://doi.org/10.1214/24-EJS2221 Electronic Journal of Statistics
Vol. 18 (2024) 942–979
ISSN: 1935-7524
https://doi.org/10.1214/24-EJS2221 1. Introduction Change point detection is a classical statistical problem aiming to detect if
there is a change in the mean, covariance structure, and distribution along a
sequence of time-ordered observations. It has been an active field of research
in many scientific fields, such as quality control [21], financial market analysis
[28], genetics and medicine [7], psychopathology [22], signal processing [18] and
machine learning [5]. Driven by a wide range of modern scientific applications, change point infer-
ence in high-dimensional data is of significant current interest. A vast part of
the existing literature focuses on detecting changes in the mean [17, 8, 27] and in
the covariance structure [1, 11, 2, 10, 26]. The literature on detecting structural 942 Efficient change point detection and estimation in HD correlation matrices 943 breaks in the correlation matrix is relatively scarce. The correlation coefficient
is a standard measure for linear dependency among variables, and correlation
analysis is reliable only when the data has no change in correlation during the
observation period. Moreover, in a multivariate setting, aside from changes in
univariate features, important events are often marked by abrupt correlation
changes [4]. Change point analysis of correlation has been well studied in the conven-
tional low-dimensional setting, i.e., the dimension is fixed while the sample size
becomes large. [28] constructed a bootstrap variance matrix estimator to detect
a change in the correlation matrix. [4] proposed a Gaussian kernel-based change
point detection method (KCP). Compared to CUSUM-type methods, KCP can
locate multiple change points simultaneously. Change point analysis in time se-
ries has been studied by [19] and [12] and the references therein. However, in
the high-dimensional setting, where the dimension can be much larger than the
sample size, the methods constructed under the low-dimensional setting either
perform poorly or are not even well defined. For example, the method proposed
by [28] is unstable when the dimension is not small relative to the sample size
due to the (near) singularity of the variance estimator. To avoid variance esti-
mation, [9] proposed a break test based on the self-normalization method, which
applies to high-dimensional data. However, there is no estimation method pro-
posed to locate the change point. [10] proposed a two-stage approach based on
bootstrap to estimate a change point in the high-dimensional covariance struc-
ture. 1. Introduction This method can be applied to high-dimensional correlation matrices, as
correlation can be treated as a particular case of the covariance matrix. However,
this method is designed to estimate the location when the change point exists
in the middle of a sequence of data, which is restrictive in real applications. This paper proposes a new test based on signflip parallel analysis to detect
the existence of changes in the correlation structure. Moreover, we construct a
two-step approach combining a signflip permutation dimension reduction step
and a CUSUM statistic to estimate the location of change point and simulta-
neously identify corresponding changing components in the correlation matrix. The theoretical properties analysis is conducted, and the consistency of the pro-
posed estimator is established. In addition, by combing the Synthetic Minority
Oversampling Technique (SMOTE), the estimation accuracy is enhanced when
the change point exists in the extreme tail of a data sequence. We examine the
numerical performance of the proposed detection and estimation methods using
both simulated and real datasets. The numerical results show that the proposed
methods significantly outperform the existing ones for non-Gaussian data and
the change point existing in the extreme tail of a sequence. More importantly, as
the dimension p increases, the proposed methods become more accurate while
existing methods maybe not. The remaining sections are organized as follows. In Section 2, we present the
first main result of the paper. A new break test is proposed based on signflip
parallel analysis to detect change points in the correlation structure of high-
dimensional data. In Section 3, we present the second main result of the paper. After a dimension reduction step based on signflip permutation, a CUSUM Z. Li and J. Gao 944 statistic is used to estimate the location of the change point. At the same time,
the components of the correlation matrix leading to the change point are identi-
fied. The consistency of this estimator is established. In Section 4, an algorithm
combing the proposed estimation procedure and SMOTE is designed to locate
the change point in the extreme tail of a data sequence. We evaluate the finite
performance of two proposed methods compared to several existing methods
by a detailed simulation study in Section 5. Real data analyses are carried out
in Section 6. Some discussions are offered in the last conclusion section. 1. Introduction Due
to the limited space, we relegated the proofs and some numerical results to
supplementary materials. 2. Detection of change point For large p, it can be calculated efficiently by vt =
1
t
t
k=1
vecho(xkx′
k) −
1
T −t
T
k=t+1
vecho(xkx′
k)
◦
1
t
t
k=1
vecho(xkx′
k) −
1
T −t
T
k=t+1
vecho(xkx′
k)
, where vecho(·) indicates the half-vectorization p(p −1)/2 vector by vectorizing
only the lower triangular part without the diagonal of the symmetric matrix,
and “ ◦” is the Hadamard product. If a change point exists at t0, then one can
verify that vt0 is an estimator of vecho(R1−R2)2, which measures the difference
between the two population correlation matrices. In particular, the expectation
of the component vt(i, j) of the vector vt corresponding to the position (i, j) in
the matrices R1 and R2 is (for simplicity, this expectation is calculated under
the assumption that original data {yt} have mean zero and variance one. For
notational convenience, write ρ1 = ρ1(i, j) and ρ2 = ρ2(i, j) here) Evt(i, j) =
⎧
⎪
⎪
⎨
⎪
⎪
⎩
(ρ1 −ρ2)2 + 1
t0
β1 −ρ2
1
+
1
T −t0
β2 −ρ2
2
if t = t0,
(T −t0)2
(T −t)2 (ρ1 −ρ2)2 +
1
t +
t0−t
(T −t)2
β1 −ρ2
1
+
T −t0
(T −t)2
β2 −ρ2
2
if t < t0,
t2
0
t2 (ρ1 −ρ2)2 + t0
t2
β1 −ρ2
1
+
t−t0
t2
+
1
T −t
β2 −ρ2
2
if t > t0. (5) where β1 := E(xkixkj)2 for k ≤t, β2 := E(xkixkj)2 for k > t, and w.l.o.g, we
can assume β1 and β2 are uniformly bounded in this paper (see Assumption 1,
Lemma A.1, and the Proposition 2.7.1 of [25]). Thus, in each of the three cases
of (5), the first term is the main term with order O(1), while the other terms are
all o(1). Therefore, the expectation of vt(i, j) consistently achieves the largest
value at the true change point position t0 because the coefficients (T −t0)2
(T −t)2 and t2
0
t2
before (ρ1 −ρ2)2 are smaller than 1 when t ̸= t0. Moreover, these coefficients are
unrelated to the position (i, j). Therefore, for any fixed t, larger values of vt(i, j)
indicate a significant difference between ρ1(i, j) and ρ2(i, j). 2. Detection of change point Given a sequence of data y1, . . . , yt0, yt0+1, . . . , yT ∈Rp, as a first step, one may
be interested in testing the existence of change points in the correlation struc-
ture. In this section, we aim to test for consistency of the correlation matrices
of observations {yt}1≤t≤T : (1) H0 : corr(y1) = · · · = corr(yT ),
v.s. H1 : not H0. (1) The null hypothesis H0 indicates the consistency of the correlation matrices
during the observation period. The alternative hypothesis allows for one or more
change points in the correlation structure. When there exists a change point,
the parameter t0 defines the true change point, that is, the first t0 samples
y1, . . . , yt0 have a common correlation matrix R1 = (ρ1(i, j)), i, j = 1, . . . , p,
while the last (T −t0) samples yt0+1, . . . , yT have another common correlation
matrix R2 = (ρ2(i, j)), i, j = 1, . . . , p, and R1 ̸= R2. For t = 1, . . . , T, denote by yt = (yt1, . . . , ytp)′ the t-th observation, let
¯y =
1
T
T
i=1 yi = (¯y1, . . . , ¯yp)′ be the sample mean, Sn =
1
T −1
T
i=1(yi −
¯y)(yi−¯y)′ the sample covariance matrix with diagonal elements diag(D). Define
xt = D−1/2(yt −¯y) as the vector of standardized observations. Given a time
point t, define the sample correlation matrices
ˆρt
1(i, j)
p×p := ˆRt
1 = 1
t
t
k=1
xkx′
k,
(2)
ˆρT
t+1(i, j)
p×p := ˆRT
t+1 =
1
T −t
T
k=t+1
xkx′
k. (3) (2) (3)
ˆρT
t+1(i, j)
p×p := ˆRT
t+1 =
1
T −t
k=t+1
xkx′
k. (3) In order to quantify the changes in correlations after and before t, a natural
approach is to compare the sample correlations {ˆρt
1(i, j)} and {ˆρT
t+1(i, j)}, and
we consider the following p(p −1)/2-dimensional vector
ˆρt
1(i, j) −ˆρT
t+1(i, j)
2
1≤i<j≤p := vt
(4) (4) Efficient change point detection and estimation in HD correlation matrices 945 of the squared (componentwise) differences of the elements of the sample corre-
lations. 6
τ1 ←the largest element in W = ( ˜w(1), . . . , ˜w(q)) that is, τ1 = max(W). Let w(i, j) denote the elements of the vector w corresponding to the position
(i, j) in the matrices R1 and R2. We identify all components which are larger
than the threshold τ1, to be precise, define wτ1 = {(i, j) : w(i, j) > τ1, 1 ≤i < j ≤p},
(7) (7) as the index set of all corresponding components. Then, for the testing problem
in (1), the rejection region is as the index set of all corresponding components. Then, for the testing problem
in (1), the rejection region is C = {w : card(wτ1) > 0},
(8) (8) where card(·) is the cardinality of a set. The change point detection approach
based on the rejection region C in Equation (8) will be referred to as the signflip
parallel analysis-based change point detection (SPAD) procedure. where card(·) is the cardinality of a set. The change point detection approach
based on the rejection region C in Equation (8) will be referred to as the signflip
parallel analysis-based change point detection (SPAD) procedure. Remark 2.1. To illustrate why the threshold τ1 selected by signflip permutation
is valid, we examine the expectation of the component ˜vt(i, j) of the vector ˜vt
for signflipped data Rm ◦Y (under the same assumption as (5)). We obtain E˜vt(i, j) =
⎧
⎪
⎪
⎪
⎨
⎪
⎪
⎪
⎩
1
t0 β1 +
1
T −t0 β2
t = t0,
1
t +
t0−t
(T −t)2
β1 +
T −t0
(T −t)2 β2
t < t0,
t0
t2 β1 +
t−t0
t2
+
1
T −t
β2
t > t0. (9) (9) Compared to the expectation of vt(i, j) in (5), the elements in the vectors ˜vt’s
are considerably small for all t both under the null and alternative hypothesis, as
the expectation is not related to the difference between ρ1(i, j) and ρ2(i, j), and
E˜vt(i, j) = o(1). In addition, we illustrate the phenomenon in Figure 1 through
a simulation study. Independent samples are drawn from Gaussian distribution
with p = 100 and T = 100. Under H0, the correlation matrices are R1 = R2 =
Ip, and under H1, R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤p. With
increasing order, we plot the distributions of the elements of w for original data
and the elements of ˜w for signflipped data. 2. Detection of change point In contrast, if there
is no change point in the sequence of correlation matrices, then all elements in
the vectors vt’s (1 ≤t ≤T) are considerably small as Evt(i, j) = o(1) for all t. (
)
(
)
( )
In addition, instead of investigating all values of t separately, we use a
weighted summation w =
1
T −3
T −2
t=2
t(T −t)
T
vt
(6) (6) to identify the largest components among the p(p −1)/2 entries. The weights
t(T −t)
T
are introduced to address the different sizes of the variance of vt for
different values of t. By examining the largest entries in the vector w, we can
detect whether change points exist in the correlation structure. 946 Z. Li and J. Gao Z. Li and J. Gao We propose a threshold based on signflip parallel analysis, and the Algorithm
is as follows. Algorithm 1 : Threshold via Signflip Permutation
Input : Data Matrix Y ∈Rp×T (p variables and T series), number of trials q. Output : Threshold τ1. Algorithm 1 : Threshold via Signflip Permutation
Input : Data Matrix Y ∈Rp×T (p variables and T series), number of trials q. Output : Threshold τ1. f
d 1
for m ←1 to q do 2
Randomly signflip entries of Y : form Rm ◦Y where
/ 2
Randomly signflip entries of Y : form Rm ◦Y where
Rm(ij)
i.i.d. ∼
+1,
with probability 1/2,
−1,
with probability 1/2,
that is Rm ∈Rp×T has i.i.d. Rademacher entries;
3
Calculate vectors {˜v(m)
t
}2≤t≤T −2 defined in (4); ;
3
Calculate vectors {˜v(m)
t
}2≤t≤T −2 defined in (4);
(
)
(
) 4
Calculate the weighted sum ˜w(m) in (6) from {˜v(m)
t
}2≤t≤T −2;
d 4
Calculate the weighted sum ˜w(m) in (6) from {˜v(m)
t
}2≤t≤T −2;
d 5
end 5
end
6
τ1 ←the largest element in W = ( ˜w(1), . . . , ˜w(q)) that is, τ1 = max(W). 6
τ1 ←the largest element in W = ( ˜w(1), . . . , ˜w(q)) that is, τ1 = max(W). In Figure 1(a), it is obvious that Efficient change point detection and estimation in HD correlation matrices 947 Fig 1. The distributions of increasing-order elements of w and ˜w, (a) under H0, (b) un-
der H1. Fig 1. The distributions of increasing-order elements of w and ˜w, (a) under H0, (b) un-
der H1. the two distributions are almost coincident under H0, except that the maximum
value of ˜w may be larger than that of w. In contrast, under H1, the distribution
of the elements of w is always above that of ˜w, as shown in Figure 1(b). More
importantly, the distributions of the elements of ˜w are very similar under both
H0 and H1. Thus, the signflip step breaks the changing structure in the original
data. See [14] for more signflip parallel analysis properties. 3. Estimation of change point location In many applications, if the null hypothesis H0 : corr(y1) = · · · = corr(yT ) is
rejected, it is often of significant interest to further estimate the change point’s
location and investigate at which components the correlation matrices R1 and
R2 differ from each other, that is recovering the support of R1 −R2. Suppose
the change point location in the sequence is t0 = βT, 0 < β < 1, under the alter-
native with one change point, we aim to estimate the change point fraction β. When the null hypothesis H0 in (1) is rejected, the cardinality of wτ1 is
nonzero, and its element {wτ1(i, j)} indicates a significant difference between
ρ1(i, j) and ρ2(i, j). Thus, based on the components corresponding to wτ1, we
can estimate the location of the change point. In order to keep more sufficiently
significant changes, we use a new threshold τ2, which is slightly smaller than τ1. τ2 is generated the same way as τ1 by Algorithm 1 except for Line 6, τ2 is the
α-quantile (95%, 90%) of all elements of W, that is, τ2 = α - quantile of vec(W),
(10) (10) where vec(·) is the vectorization of a matrix. Then, we reduce the p(p −1)/2-
dimensional vector w to a vector of dimension d, wτ2 = {(i, j) : w(i, j) > τ2, 1 ≤i < j ≤p},
(11) (11) and correspondingly, we obtain zi, a d-dimensional subvector of vecho(xix′
i) by
keeping the d components of the index set wτ2. 8
Z. Li and J. Gao Z. Li and J. Gao Z. Li and J. Gao 948 For the estimation of the change point, we consider the CUSUM statistic UT (t) = 1
T 4
t
i,k=1
T
j,l=t+1
(zi −zj)′(zk −zl) = 1
T 4 ||(T −t)t(˜ρt
1 −˜ρT
t+1)||2
2, (12) (12) where || · ||2 represents the Euclidean norm, and ˜ρt
1 and ˜ρT
t+1 represent the
vectors containing the elements of the correlation matrice estimators ˆRt
1 and
ˆRT
t+1 corresponding to positions identified in the dimension reduction index set
wτ2, respectively. 3. Estimation of change point location Then the estimator of the change point fraction β is defined
by where || · ||2 represents the Euclidean norm, and ˜ρt
1 and ˜ρT
t+1 represent the
vectors containing the elements of the correlation matrice estimators ˆRt
1 and
ˆRT
t+1 corresponding to positions identified in the dimension reduction index set
wτ2, respectively. Then the estimator of the change point fraction β is defined
by ˆβ = arg max
1≤t≤T UT (t)/T. (13) (13) This estimator ˆβ will be referred to as the signflip parallel analysis-based CUSUM
estimator (SPACE). The dimension reduction index set wτ2 denotes the support
of R1 −R2, that is, the positions at which the two correlation matrices differ. To illustrate the effectiveness of dimension reduction in (11) and derive the
asymptotic consistency of the proposed estimator ˆβ in (13), we need the follow-
ing assumptions on observations. Assumption 1. Denote xt = (xt1, . . . , xtp)′, t = 1, . . . , T. For any 1 ≤i ≤p,
xti is a sub-Gaussian random variable, that is, there are positive constants C1,
C2 (independent of the indices t and i) such that for every ε > 0, P(|xti > ε|) ≤C1e−C2ε2. Assumption 2. The smallest nonzero entry of the matrix R1 −R2 satisfies |ρ1(i, j) −ρ2(i, j)| > C3
τ2
T · max
T 2
(T −t0)2 , T 2
t2
0
, 1 ≤i ≤j ≤p,
(14) (14) where τ2 is given in (10). Assumption 3. For some positive constant c, we have p2T 2e−c√τ2 = o(1). In the following theorem, we prove that all the entries with no difference are
not included in wτ2. Theorem 3.1. Define N = {(i, j) : 1 ≤i < j ≤p; ρ1(i, j) = ρ2(i, j)} as the set
of indices corresponding to identical elements in the correlation matrices R1
and R2. Then under Assumption 1, P
⎧
⎨
⎩
(i,j)∈N
{w(i, j) > τ2}
⎫
⎬
⎭= P(N ∩wτ2 ̸= ∅)
(15)
≤c1p2T
e−c2T 1/4√τ2 + e−c3 min(T 1/4√τ2,τ2)
, P
⎧
⎨
⎩
(i,j)∈N
{w(i, j) > τ2}
⎫
⎬
⎭= P(N ∩wτ2 ̸= ∅)
(15) (15) ≤c1p2T
e−c2T 1/4√τ2 + e−c3 min(T 1/4√τ2,τ2)
, Efficient change point detection and estimation in HD correlation matrices
949 Efficient change point detection and estimation in HD correlation matrices
949 949 where c1, c2 and c3 are some constants. 3. Estimation of change point location In addition, under Assumption 3 we
have
⎧
⎫ P
⎧
⎨
⎩
(i,j)∈N
{w(i, j) > τ2}
⎫
⎬
⎭→0. (16) (16) In the next theorem, we prove that all the entries with a difference larger
than C3
τ2
T · max
T 2
(T −t0)2 , T 2
t2
0
are kept in wτ2. Theorem 3.2. Define P = {(i, j) : 1 ≤i < j ≤p; |ρ1(i, j) −ρ2(i, j)| > λ} as
the
set
of
components
which
differ
by
more
than
λ,
where
λ = C3
τ2
T · max
T 2
(T −t0)2 , T 2
t2
0
as given in Assumption 2. Then under As-
sumption 1 and Assumption 2, we have Theorem 3.2. Define P = {(i, j) : 1 ≤i < j ≤p; |ρ1(i, j) −ρ2(i, j)| > λ} as
the
set
of
components
which
differ
by
more
than
λ,
where
λ = C3
τ2
T · max
T 2
(T −t0)2 , T 2
t2
0
as given in Assumption 2. Then under As-
sumption 1 and Assumption 2, we have P
⎧
⎨
⎩
(i,j)∈P
{w(i, j) > τ2}
⎫
⎬
⎭= P (P ⊂wτ2) ≥1 −c4p2Te−c5
√τ2T ,
(17) (17) where c4 and c5 are some constants. In addition, under Assumption 3 we have P
⎧
⎨
⎩
(i,j)∈P
{w(i, j) > τ2}
⎫
⎬
⎭→1. (18) (18) Theorem 3.2 shows that under Assumptions 2 and 3, the vector wτ2 recovers
the true support of R1 −R2 exactly with probability tending to 1. ˆ In the next result, we establish the asymptotic consistency of the estimator ˆβ. Here they symbol
p→denotes convergence in probability. Theorem 3.3. Under Assumptions 1 and 2, we have Theorem 3.3. Under Assumptions 1 and 2, we have P
ˆβ
β −1
≥ϵ
≤c6p2T 2e−c7√τ2, where c6 and c7 are some constants. In addition, under Assumption 3 we have where c6 and c7 are some constants. In addition, under Assumption 3 we have ˆβ
p→β. Remark 3.1. The number of trails q in the Algorithm. In a parallel analysis,
the number of trials should be as large as possible to retain a stable and accu-
rate result, such as 50 and 100 trials [15, 16, 24]. However, many repetitions
lead to time-consuming computation, especially for high-dimensional data. We
investigate a suitable number of parallel trails by conducting simulations on the
effect of the number of trials q in both change point detection and estimation. Figure 2 presents the success rate of the proposed detection method, SPAD,
against the number of parallel trails q under the H0 without a change point. The
success rate grows fast when q is still small. Therefore, we recommend a number
of trials of around 30 in the change point detection procedure. Z. Li and J. Gao
Fig 2. Success rate vs. the number of trails q under H0. 950 Z. Li and J. Gao Z. Li and J. Gao Z. Li and J. Gao Fig 2. Success rate vs. the number of trails q under H0. Fig 3. Mean value of the SPACE ˆβ vs. number of trails q. Fig 3. Mean value of the SPACE ˆβ vs. number of trails q. Figure 3 presents the mean values of SPACE’s estimated change point fraction
against the number of parallel trails q for the four cases in Section 5.2 and
different combinations of p and T. Each mean value is calculated from 200
repetitions. The mean values of estimates are all close to the true value and
vary slightly with q. Therefore, the number of trails around 10 to 20 is enough
to estimate the change point location. 5. Finite sample properties In this section, we conduct extensive simulations to examine the proposed SPAD
method’s success rate and the proposed SPACE’s accuracy and compare them
with several existing methods. We consider different dimensions p ranging from
5 to 500 and sample sizes T = 100 and T = 200. All the numerical results below
are calculated from 200 replications. 11
β1 ←the estimator of the change point fraction. 11
β1 ←the estimator of the change point fraction. Similarly, by combining the SMOTE, the proposed SPAD method can also be
enhanced to detect the existence of change points when they are in the extreme
tail of the sequence of data, see Algorithm 3 in the Supplement Materials and
corresponding simulations. 4. Change point in the extreme tail Most existing literature aims to detect and estimate the change point in the
middle of a data sequence. However, the change point hardly arises near the
center and may appear in the tail. Therefore, in this section, we propose com-
bining the SMOTE with SPACE to deal with the challenging situation where
the change point arises in the extreme tail of a data sequence. 951 Efficient change point detection and estimation in HD correlation matrices SMOTE is a viral oversampling method proposed by [6] and designed to deal
with imbalanced data in classification problems by generating synthetic samples
from the minority class. Specifically, for each sample y in the minority class,
the nearest five neighbors with the smallest Euclidean distance are identified. One of them is randomly chosen as y∗
t , based on which a new synthetic SMOTE
sample is produced as ys = yt + u · (y∗
t −yt), where u is randomly chosen from
a standard uniform distribution [3]. To estimate the change point in the extreme tail effectively, the proposed
SPACE can be enhanced by combining the SMOTE, and the Algorithm is as
follows. Algorithm 2 : SMOTE + SPACE
Input : Data Matrix Y ∈Rp×T (p variables and T series), small positive ϵ. Output : the estimator of the change point fraction. 1
β0 ←SPACE result from the original data. 2
Let [γT + 1, T] be the minority class, get (1 −γ)T SMOTE variables,γ ≥0.9. 3
β1 ←SPACE result from the inflated data. 4
eps = abs(β0 −β1). 5
while eps > ϵ do
6
β0 = β1;
7
Repeat the above steps 2–4;
10
end
11
β1 ←the estimator of the change point fraction. 5.1. Detection of change point We first illustrate the performance of the SPAD method under both H0 and H1. Under H0, independent samples are drawn from multivariate Gaussian dis-
tributions with correlation matrice R1 = R2 = Ip. The threshold τ1 is obtained
from q = 30 parallel trials. We compare SPAD with a method proposed by [10],
which is designed to detect a change point in a sequence of covariance matrices. For the reader’s convenience, we recall the method of [10]. Z. Li and J. Gao 952 • The detection method of [10] is based on the following vector. • The detection method of [10] is based on the following vector. D
=
1
T −3
T −2
t=2
t(T −t)
T
1
t(t −1)
i̸=j≤t
vech( ˙yi ˙y′
i) ◦vech( ˙yj ˙y′
j)(19)
+
1
(T −t)(T −t −1)
i̸=j>t
vech( ˙yi ˙y′
i) ◦vech( ˙yj ˙y′
j)
−
2
t(T −t)
i≤t
j>t
vech( ˙yi ˙y′
i) ◦vech( ˙yj ˙y′
j)
, where ˙yi = yi −¯y is the demeaned vector of data, and vech(·) indicates
the half-vectorization p(p+1)/2 vector by vectorizing the lower triangular
part of the symmetric matrix. With a threshold τ0 log(log T), where τ0
is chosen by the bootstrap procedure stated in Section 2.3 of [10], no
component in D is larger than the threshold implies no change point exists
in the sequence of covariance matrices. Table 1
The success rate of detecting no change point under H0. p
20
50
100
200
300
500
SPAD
0.980
0.970
0.985
0.985
0.970
0.990
Dette (2020)
0.850
0.945
0.960
0.965
0.970
0.980 Table 1 Table 1 presents the success rate of detecting no change point. Both meth-
ods have very high accuracies, and SPAD performs slightly better than Dette’s
method. 5.1. Detection of change point Under H1, independent Gaussian samples are considered in five different
settings: • Case 1: β = 0.5, R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤p;
• Case 2: β = 0.5, R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤[p/2],
where [p/2] is the integer part of p/2;
• Case 3: β = 0.75, R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤p;
• Case 4: β1 = 1/3, β2 = 2/3, R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸=
j ≤p, R3 = R1 = Ip;
• Case 5: β1 = 1/3, β2 = 2/3, R1 = Ip, R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤p,
and R3(i, j) = 0.9 for all 1 ≤i ̸= j ≤p. Table 2 presents the success rate of detecting change points under the five cases. Overall, the SPAD method prominently outperforms Dette’s method in all cases. Significantly, as the dimension p becomes larger, the accuracy of SPAD increases
while the accuracy of Dette’s method decreases. When the change point exists
in the middle of the sequence (Cases 1 & 2), a larger magnitude of changes in
the correlation matrices leads to higher detection accuracy, as expected. The
detection accuracies of both methods are also affected by the location of the
change point. Compared to Case 1, the change point location in Case 3 is close
to the tail, so the detection accuracies of both methods are lower. When there is 953 Table 2
The success rate of detection change point under H1. p
20
50
100
200
300
500
Case 1
SPAD
0.770
0.860
0.810
0.865
0.910
0.895
Dette (2020)
0.745
0.555
0.540
0.415
0.360
0.225
Case 2
SPAD
0.525
0.660
0.695
0.700
0.785
0.795
Dette (2020)
0.535
0.465
0.360
0.305
0.255
0.170
Case 3
SPAD
0.495
0.560
0.500
0.575
0.635
0.670
Dette (2020)
0.530
0.365
0.280
0.250
0.215
0.115
Case 4
SPAD
0.070
0.045
0.050
0.055
0.070
0.055
Dette (2020)
0.165
0.075
0.030
0.035
0.020
0.025
Case 5
SPAD
0.965
0.955
0.980
0.980
0.990
0.980
Dette (2020)
0.875
0.770
0.715
0.625
0.560
0.455 Table 2 more than one change point in the sequence (Cases 4 & 5), the performances of
both methods are influenced by the change direction of correlations. 5.1. Detection of change point In Case 4,
the correlation matrix changes from an identity matrix to an equal correlation
matrix and then changes back to the identity matrix. It is hard for both methods
to detect the change points. In Case 5, the correlations increase from 0 to 0.5
and then to 0.9. Both methods can detect the existence of change points. Our
SPAD method performs much better and reaches very high accuracies. 5.2. Estimation of change point We investigate the finite sample properties of the proposed SPACE under dif-
ferent settings and compare it with two alternative methods proposed by [10]
and [4], which are recalled as follows. For the threshold τ2 in SPACE, we set
α = 95% in Equation (10). • The estimator of [10] is defined as follows. First, a critical value τ is con-
structed from the bootstrap approach stated in Section 2.3 of [10], based
on which an index set is obtained from the vector D in (19) Dτ = {(i, j) : D(i, j) > τ, 1 ≤i ≤j ≤p}. Dτ = {(i, j) : D(i, j) > τ, 1 ≤i ≤j ≤p}. Then Dette’s estimator is Then Dette’s estimator is ˆβD = 1
T arg max
1≤t≤T
1
T 4
t
t
(i̸=s)=1
T
T
(j̸=l)=t+1
(˜yi −˜yj)′(˜ys −˜yl),
(20) (20) where ˜yi is a subvector of vech( ˙yi ˙y′
i) by keeping the corresponding com-
ponents of the index set Dτ. • The estimator of [4] is kernel-based. The similarities of all pairs of (yt, yt′)
are measured using Gaussian kernel, g(yt, yt′) = exp
−||yt −yt′||2
2h2
, t, t′ = 1, . . . , T, Z. Li and J. Gao 954 Z. Li and J. Gao Z. Li and J. Gao where h is the bandwidth parameter, equals the median Euclidean distance
between all observations. Then the KCP-raw estimator is ˆβC = 1
T arg min
1≤t≤T
1
T (v1,t + v2,t),
(21) (21) where v1,t = (t −1) −
1
t−1
t
i=2
t
j=2
g(yt, yt′) measures the homogeneousness where v1,t = (t −1) −
1
t−1
t
i=2
t
j=2
g(yt, yt′) measures the homogeneousness of phase before the time point t and v2,t = (T−t)−
1
T −t
T
i=t+1
T
j=t+1
g(yt, yt′) of phase before the time point t and v2,t = (T−t)−
1
T −t
T
i=t+1
T
j=t+1
g(yt, yt′)
measures the homogeneousness of phase after the time point t. j
measures the homogeneousness of phase after the time point t. In the data generating process, {yt}1≤t≤T are generated from two distribu-
tions: (i) Multivariate normal, N(0, Σk), k = 1, 2; and (ii) Student-t, t5; with
covariance matrix Σk = ˜DRk ˜D, where ˜D = diag(σ1, . . . 5.2. Estimation of change point , σp), R1 and R2 are the
correlation matrices before and after the change point, respectively. We con-
sider homoscedastic senario so σj = 1, j = 1, . . . , p. Four different choices for
correlation matrices R1 and R2 are considered. • Case 6: R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤p;
• Case 7: R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤[p/2]; p
(
)
[ / ]
• Case 8: R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤[p/3], R2(i, j) = 0.2
for all [p/3] + 1 ≤i ̸= j ≤[2p/3], R2(i, j) = 0.8 for all [2p/3] + 1 ≤i ̸=
j ≤p; p
• Case 8: R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤[p/3], R2(i, j) = 0.2
for all [p/3] + 1 ≤i ̸= j ≤[2p/3], R2(i, j) = 0.8 for all [2p/3] + 1 ≤i ̸=
j ≤p;
• Case 9: R1(i, j) = 0.5 for all 1 ≤i ̸= j ≤p, and R2(i, i+1) = R2(i, i−1) =
−0.5 for all 1 ≤i ≤p. • Case 8: R1 = Ip, and R2(i, j) = 0.5 for all 1 ≤i ̸= j ≤[p/3], R2(i, j) = 0.2
for all [p/3] + 1 ≤i ̸= j ≤[2p/3], R2(i, j) = 0.8 for all [2p/3] + 1 ≤i ̸=
j ≤p;
• Case 9: R1(i, j) = 0.5 for all 1 ≤i ̸= j ≤p, and R2(i, i+1) = R2(i, i−1) =
−0.5 for all 1 ≤i ≤p. j
• Case 9: R1(i, j) = 0.5 for all 1 ≤i ̸= j ≤p, and R2(i, i+1) = R2(i, i−1) =
−0.5 for all 1 ≤i ≤p. Table 3 shows the mean change point fraction, SD, and MSE of three esti-
mators for all four cases when data are drawn from Gaussian distribution. The
true change point exists in the middle of the sequence. The SPACE and Dette’s
estimators perform better than the KCP-raw estimator in all cases. For Cases
6 and 8, Dette’s estimator is slightly better than SPACE in terms of mean,
while SPACE is better than Dette’s estimator in terms of SD and MSE, which
implies SPACE is more stable. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5184
0.1311
0.0174
0.4430
0.2088
0.0466
0.5292
0.3020
0.0916
50
0.5459
0.1050
0.0131
0.2622
0.2492
0.1184
0.5582
0.2581
0.0696
100
0.5256
0.1100
0.0127
0.1886
0.2398
0.1542
0.5710
0.2512
0.0679
300
0.5312
0.1200
0.0153
0.0909
0.1777
0.1987
0.5812
0.2483
0.0679
500
0.5279
0.1129
0.0135
0.0552
0.1293
0.2145
0.5954
0.2495
0.0710
Case 7
5
0.5028
0.2041
0.0413
0.3172
0.2686
0.1052
0.5460
0.3411
0.1179
50
0.5265
0.1502
0.0232
0.1998
0.2502
0.1524
0.5212
0.3481
0.1210
100
0.5221
0.1413
0.0204
0.1324
0.2121
0.1799
0.5111
0.3467
0.1197
300
0.5174
0.1644
0.0272
0.0658
0.1637
0.2152
0.5814
0.3465
0.1261
500
0.5223
0.1472
0.0220
0.0434
0.1114
0.2209
0.5691
0.3516
0.1278
Case 8
5
0.5363
0.1253
0.0169
0.4472
0.1955
0.0408
0.5460
0.3089
0.0971
50
0.5343
0.1042
0.0120
0.2999
0.2541
0.1042
0.5372
0.3266
0.1075
100
0.5289
0.1119
0.0133
0.1906
0.2412
0.1536
0.5614
0.3221
0.1070
300
0.5287
0.1235
0.0160
0.0866
0.1773
0.2022
0.5951
0.3162
0.1085
500
0.5233
0.1054
0.0116
0.0688
0.1554
0.2100
0.5990
0.2988
0.0986
Case 9
5
0.4936
0.0749
0.0056
0.4852
0.0825
0.0070
0.4992
0.2183
0.0474
50
0.4727
0.0948
0.0097
0.3815
0.2147
0.0599
0.4300
0.2403
0.0623
100
0.4673
0.1030
0.0116
0.2891
0.2471
0.1052
0.3961
0.2400
0.0681
300
0.4649
0.1201
0.0156
0.1902
0.2391
0.1528
0.4391
0.2579
0.0699
500
0.4680
0.1279
0.0173
0.1070
0.1887
0.1899
0.4222
0.2627
0.0747 Efficient change point detection and estimation in HD correlation matrices 955 Table 3
Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point exists in the middle β = 0.5, T = 100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5243
0.0671
0.0051
0.5047
0.0959
0.0092
0.5550
0.2790
0.0805
50
0.5231
0.0290
0.0014
0.5059
0.0720
0.0052
0.6379
0.1941
0.0565
100
0.5231
0.0327
0.0016
0.5036
0.0646
0.0042
0.6208
0.1597
0.0400
300
0.5248
0.0344
0.0018
0.4948
0.1041
0.0108
0.6208
0.1508
0.0372
500
0.5217
0.0312
0.0014
0.4833
0.1202
0.0147
0.6180
0.1554
0.0380
Case 7
5
0.5163
0.1136
0.0131
0.4152
0.2186
0.0548
0.5581
0.3367
0.1162
50
0.5227
0.0294
0.0014
0.4815
0.1263
0.0162
0.6204
0.2847
0.0952
100
0.5236
0.0326
0.0016
0.4786
0.1189
0.0145
0.6647
0.2280
0.0788
300
0.5244
0.0349
0.0018
0.4774
0.1383
0.0195
0.6782
0.1940
0.0692
500
0.5201
0.0288
0.0012
0.4480
0.1574
0.0273
0.6696
0.1934
0.0660
Case 8
5
0.5173
0.0469
0.0025
0.5082
0.0735
0.0054
0.5288
0.3028
0.0921
50
0.5165
0.0241
0.0008
0.5101
0.0588
0.0035
0.6227
0.2211
0.0637
100
0.5130
0.0223
0.0007
0.5139
0.0536
0.0031
0.6534
0.1945
0.0612
300
0.5187
0.0309
0.0013
0.5140
0.0338
0.0013
0.6828
0.1735
0.0634
500
0.5160
0.0249
0.0009
0.5089
0.0700
0.0050
0.6622
0.1714
0.0555
Case 9
5
0.4869
0.0316
0.0012
0.4870
0.0350
0.0014
0.4845
0.2030
0.0412
50
0.4760
0.0351
0.0018
0.4906
0.0195
0.0005
0.3837
0.1732
0.0434
100
0.4759
0.0365
0.0019
0.4909
0.0188
0.0004
0.3737
0.1642
0.0428
300
0.4758
0.0294
0.0014
0.4920
0.0365
0.0014
0.3692
0.1569
0.0416
500
0.4759
0.0337
0.0017
0.4945
0.0146
0.0002
0.3629
0.1686
0.0471 Table 3 Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point exists in the middle β = 0.5, T = 100. Table 4 Student-t. Mean, standard deviation (SD), and mean squared error (MSE) of three
estimators when the change point exists in the middle β = 0.5, T = 100. estimators when the change point exists in the middle β = 0.5, T = 100. 5.2. Estimation of change point Moreover, SPACE performs the best in Case 7
and Dette’s estimator performs the best in Case 9. As expected, the KCP-raw
estimator is only close to the true value when p = 5, but performs poorly when
p is large. Meanwhile, the KCP-raw estimator has the largest SD and MSE. Table 4 shows the mean change point fraction, SD, and MSE of three es-
timators for all four cases when data are drawn from Student-t distribution. SPACE performs best in all cases. With the results in Table 3 (where data are
Gaussian), Table 4 shows the robustness of SPACE concerning population dis-
tributions. In contrast, Dette’s estimator is not robust to non-Gaussian data as
the mean values are far from the true value except for the cases when p = 5. Table 5 and Table 6 present the mean change point fraction, SD, and MSE of
three estimators under four cases for normal and student-t distributed samples,
respectively, when the change point is close to the tail, β = 0.7. For normal
samples, the proposed SPACE performs the best for Cases 6, 7, and 8 and is
very close to the true value with the smallest SD and MSE. Dette’s estimator Efficient change point detection and estimation in HD correlation matrices
955
Table 3
Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point exists in the middle β = 0.5, T = 100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5243
0.0671
0.0051
0.5047
0.0959
0.0092
0.5550
0.2790
0.0805
50
0.5231
0.0290
0.0014
0.5059
0.0720
0.0052
0.6379
0.1941
0.0565
100
0.5231
0.0327
0.0016
0.5036
0.0646
0.0042
0.6208
0.1597
0.0400
300
0.5248
0.0344
0.0018
0.4948
0.1041
0.0108
0.6208
0.1508
0.0372
500
0.5217
0.0312
0.0014
0.4833
0.1202
0.0147
0.6180
0.1554
0.0380
Case 7
5
0.5163
0.1136
0.0131
0.4152
0.2186
0.0548
0.5581
0.3367
0.1162
50
0.5227
0.0294
0.0014
0.4815
0.1263
0.0162
0.6204
0.2847
0.0952
100
0.5236
0.0326
0.0016
0.4786
0.1189
0.0145
0.6647
0.2280
0.0788
300
0.5244
0.0349
0.0018
0.4774
0.1383
0.0195
0.6782
0.1940
0.0692
500
0.5201
0.0288
0.0012
0.4480
0.1574
0.0273
0.6696
0.1934
0.0660
Case 8
5
0.5173
0.0469
0.0025
0.5082
0.0735
0.0054
0.5288
0.3028
0.0921
50
0.5165
0.0241
0.0008
0.5101
0.0588
0.0035
0.6227
0.2211
0.0637
100
0.5130
0.0223
0.0007
0.5139
0.0536
0.0031
0.6534
0.1945
0.0612
300
0.5187
0.0309
0.0013
0.5140
0.0338
0.0013
0.6828
0.1735
0.0634
500
0.5160
0.0249
0.0009
0.5089
0.0700
0.0050
0.6622
0.1714
0.0555
Case 9
5
0.4869
0.0316
0.0012
0.4870
0.0350
0.0014
0.4845
0.2030
0.0412
50
0.4760
0.0351
0.0018
0.4906
0.0195
0.0005
0.3837
0.1732
0.0434
100
0.4759
0.0365
0.0019
0.4909
0.0188
0.0004
0.3737
0.1642
0.0428
300
0.4758
0.0294
0.0014
0.4920
0.0365
0.0014
0.3692
0.1569
0.0416
500
0.4759
0.0337
0.0017
0.4945
0.0146
0.0002
0.3629
0.1686
0.0471
Table 4
Student-t. Mean, standard deviation (SD), and mean squared error (MSE) of three
estimators when the change point exists in the middle β = 0.5, T = 100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5954
0.1710
0.0400
0.4872
0.2634
0.1143
0.5587
0.3283
0.1272
50
0.6262
0.1598
0.0308
0.2358
0.2727
0.2894
0.6408
0.2990
0.0925
100
0.6235
0.1558
0.0300
0.1740
0.2600
0.3440
0.6515
0.3048
0.0948
300
0.6137
0.1778
0.0389
0.0703
0.1703
0.4254
0.6966
0.2714
0.0733
500
0.6363
0.1502
0.0265
0.0370
0.0953
0.4487
0.6984
0.2647
0.0697
Case 7
5
0.5393
0.2279
0.0773
0.3236
0.2819
0.2208
0.5359
0.3454
0.1456
50
0.5846
0.1902
0.0493
0.1734
0.2610
0.3450
0.5352
0.3669
0.1611
100
0.5826
0.1798
0.0460
0.1200
0.2186
0.3839
0.5593
0.3730
0.1583
300
0.5602
0.2120
0.0643
0.0621
0.1671
0.4346
0.6358
0.3625
0.1348
500
0.5825
0.2000
0.0536
0.0418
0.1207
0.4478
0.5956
0.3699
0.1470
Case 8
5
0.6353
0.1584
0.0292
0.5100
0.2680
0.1076
0.5703
0.3236
0.1210
50
0.6561
0.1421
0.0220
0.2893
0.3041
0.2607
0.6012
0.3516
0.1327
100
0.6295
0.1600
0.0304
0.1585
0.2568
0.3588
0.6035
0.3554
0.1350
300
0.6329
0.1755
0.0352
0.0781
0.1788
0.4185
0.6464
0.3374
0.1161
500
0.6314
0.1627
0.0311
0.0489
0.1335
0.4417
0.6405
0.3463
0.1229
Case 9
5
0.6321
0.1222
0.0195
0.6032
0.1604
0.0349
0.6158
0.2619
0.0754
50
0.5763
0.1593
0.0406
0.4105
0.3065
0.1773
0.5565
0.2759
0.0963
100
0.5759
0.1628
0.0418
0.2841
0.3098
0.2684
0.5083
0.2896
0.1202
300
0.5628
0.1687
0.0472
0.1979
0.2897
0.3356
0.5528
0.2977
0.1098
500
0.5708
0.1649
0.0437
0.1137
0.2256
0.3943
0.5074
0.3035
0.1287 956 Z. Li and J. Gao Table 5
Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point fraction is β = 0.7, T=100. Table 5 Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point fraction is β = 0.7, T=100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.6315
0.1072
0.0161
0.5908
0.1700
0.0407
0.5971
0.3093
0.1057
50
0.6980
0.0311
0.0010
0.5701
0.1743
0.0471
0.7632
0.1665
0.0316
100
0.6959
0.0328
0.0011
0.5777
0.1525
0.0381
0.7800
0.1446
0.0272
300
0.7014
0.0265
0.0007
0.5587
0.1753
0.0505
0.7810
0.0944
0.0154
500
0.6987
0.0237
0.0006
0.5359
0.1939
0.0643
0.7881
0.0944
0.0166
Case 7
5
0.5744
0.1518
0.0387
0.4318
0.2545
0.1364
0.5640
0.3384
0.1325
50
0.6763
0.0530
0.0034
0.5098
0.2369
0.0920
0.6664
0.3006
0.0910
100
0.6768
0.0509
0.0031
0.5059
0.2281
0.0894
0.6714
0.2968
0.0884
300
0.6861
0.0446
0.0022
0.5338
0.2116
0.0722
0.7835
0.1801
0.0393
500
0.6855
0.0419
0.0020
0.4842
0.2400
0.1039
0.7765
0.1958
0.0440
Case 8
5
0.6690
0.0738
0.0064
0.6309
0.1339
0.0226
0.5794
0.3196
0.1162
50
0.7034
0.0168
0.0003
0.6393
0.1304
0.0206
0.6993
0.2409
0.0578
100
0.7039
0.0184
0.0004
0.6221
0.1479
0.0278
0.7458
0.2284
0.0540
300
0.7032
0.0115
0.0001
0.6231
0.1250
0.0215
0.8087
0.1191
0.0259
500
0.7060
0.0140
0.0002
0.6082
0.1370
0.0271
0.7920
0.1482
0.0303
Case 9
5
0.6585
0.0622
0.0056
0.6551
0.0661
0.0064
0.6147
0.2390
0.0641
50
0.6410
0.0693
0.0083
0.6633
0.0567
0.0045
0.5434
0.2441
0.0838
100
0.6293
0.0820
0.0117
0.6619
0.0665
0.0058
0.5226
0.2347
0.0863
300
0.6223
0.0922
0.0145
0.6611
0.0711
0.0065
0.5147
0.2303
0.0871
500
0.6239
0.0878
0.0135
0.6455
0.1157
0.0163
0.5038
0.2465
0.0989 Table 6 Student-t. Mean, standard deviation (SD), and mean squared error (MSE) of three
estimators when the change point fraction is β = 0.7, T = 100. estimators when the change point fraction is β = 0.7, T = 100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5184
0.1311
0.0174
0.4430
0.2088
0.0466
0.5292
0.3020
0.0916
50
0.5459
0.1050
0.0131
0.2622
0.2492
0.1184
0.5582
0.2581
0.0696
100
0.5256
0.1100
0.0127
0.1886
0.2398
0.1542
0.5710
0.2512
0.0679
300
0.5312
0.1200
0.0153
0.0909
0.1777
0.1987
0.5812
0.2483
0.0679
500
0.5279
0.1129
0.0135
0.0552
0.1293
0.2145
0.5954
0.2495
0.0710
Case 7
5
0.5028
0.2041
0.0413
0.3172
0.2686
0.1052
0.5460
0.3411
0.1179
50
0.5265
0.1502
0.0232
0.1998
0.2502
0.1524
0.5212
0.3481
0.1210
100
0.5221
0.1413
0.0204
0.1324
0.2121
0.1799
0.5111
0.3467
0.1197
300
0.5174
0.1644
0.0272
0.0658
0.1637
0.2152
0.5814
0.3465
0.1261
500
0.5223
0.1472
0.0220
0.0434
0.1114
0.2209
0.5691
0.3516
0.1278
Case 8
5
0.5363
0.1253
0.0169
0.4472
0.1955
0.0408
0.5460
0.3089
0.0971
50
0.5343
0.1042
0.0120
0.2999
0.2541
0.1042
0.5372
0.3266
0.1075
100
0.5289
0.1119
0.0133
0.1906
0.2412
0.1536
0.5614
0.3221
0.1070
300
0.5287
0.1235
0.0160
0.0866
0.1773
0.2022
0.5951
0.3162
0.1085
500
0.5233
0.1054
0.0116
0.0688
0.1554
0.2100
0.5990
0.2988
0.0986
Case 9
5
0.4936
0.0749
0.0056
0.4852
0.0825
0.0070
0.4992
0.2183
0.0474
50
0.4727
0.0948
0.0097
0.3815
0.2147
0.0599
0.4300
0.2403
0.0623
100
0.4673
0.1030
0.0116
0.2891
0.2471
0.1052
0.3961
0.2400
0.0681
300
0.4649
0.1201
0.0156
0.1902
0.2391
0.1528
0.4391
0.2579
0.0699
500
0.4680
0.1279
0.0173
0.1070
0.1887
0.1899
0.4222
0.2627
0.0747 956
Z. Li and J. Gao
Table 5
Normal. Mean, standard deviation (SD), and mean squared error (MSE) of three estimators
when the change point fraction is β = 0.7, T=100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.6315
0.1072
0.0161
0.5908
0.1700
0.0407
0.5971
0.3093
0.1057
50
0.6980
0.0311
0.0010
0.5701
0.1743
0.0471
0.7632
0.1665
0.0316
100
0.6959
0.0328
0.0011
0.5777
0.1525
0.0381
0.7800
0.1446
0.0272
300
0.7014
0.0265
0.0007
0.5587
0.1753
0.0505
0.7810
0.0944
0.0154
500
0.6987
0.0237
0.0006
0.5359
0.1939
0.0643
0.7881
0.0944
0.0166
Case 7
5
0.5744
0.1518
0.0387
0.4318
0.2545
0.1364
0.5640
0.3384
0.1325
50
0.6763
0.0530
0.0034
0.5098
0.2369
0.0920
0.6664
0.3006
0.0910
100
0.6768
0.0509
0.0031
0.5059
0.2281
0.0894
0.6714
0.2968
0.0884
300
0.6861
0.0446
0.0022
0.5338
0.2116
0.0722
0.7835
0.1801
0.0393
500
0.6855
0.0419
0.0020
0.4842
0.2400
0.1039
0.7765
0.1958
0.0440
Case 8
5
0.6690
0.0738
0.0064
0.6309
0.1339
0.0226
0.5794
0.3196
0.1162
50
0.7034
0.0168
0.0003
0.6393
0.1304
0.0206
0.6993
0.2409
0.0578
100
0.7039
0.0184
0.0004
0.6221
0.1479
0.0278
0.7458
0.2284
0.0540
300
0.7032
0.0115
0.0001
0.6231
0.1250
0.0215
0.8087
0.1191
0.0259
500
0.7060
0.0140
0.0002
0.6082
0.1370
0.0271
0.7920
0.1482
0.0303
Case 9
5
0.6585
0.0622
0.0056
0.6551
0.0661
0.0064
0.6147
0.2390
0.0641
50
0.6410
0.0693
0.0083
0.6633
0.0567
0.0045
0.5434
0.2441
0.0838
100
0.6293
0.0820
0.0117
0.6619
0.0665
0.0058
0.5226
0.2347
0.0863
300
0.6223
0.0922
0.0145
0.6611
0.0711
0.0065
0.5147
0.2303
0.0871
500
0.6239
0.0878
0.0135
0.6455
0.1157
0.0163
0.5038
0.2465
0.0989
Table 6
Student-t. Mean, standard deviation (SD), and mean squared error (MSE) of three
estimators when the change point fraction is β = 0.7, T = 100. 5.2. Estimation of change point p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5954
0.1710
0.0400
0.4872
0.2634
0.1143
0.5587
0.3283
0.1272
50
0.6262
0.1598
0.0308
0.2358
0.2727
0.2894
0.6408
0.2990
0.0925
100
0.6235
0.1558
0.0300
0.1740
0.2600
0.3440
0.6515
0.3048
0.0948
300
0.6137
0.1778
0.0389
0.0703
0.1703
0.4254
0.6966
0.2714
0.0733
500
0.6363
0.1502
0.0265
0.0370
0.0953
0.4487
0.6984
0.2647
0.0697
Case 7
5
0.5393
0.2279
0.0773
0.3236
0.2819
0.2208
0.5359
0.3454
0.1456
50
0.5846
0.1902
0.0493
0.1734
0.2610
0.3450
0.5352
0.3669
0.1611
100
0.5826
0.1798
0.0460
0.1200
0.2186
0.3839
0.5593
0.3730
0.1583
300
0.5602
0.2120
0.0643
0.0621
0.1671
0.4346
0.6358
0.3625
0.1348
500
0.5825
0.2000
0.0536
0.0418
0.1207
0.4478
0.5956
0.3699
0.1470
Case 8
5
0.6353
0.1584
0.0292
0.5100
0.2680
0.1076
0.5703
0.3236
0.1210
50
0.6561
0.1421
0.0220
0.2893
0.3041
0.2607
0.6012
0.3516
0.1327
100
0.6295
0.1600
0.0304
0.1585
0.2568
0.3588
0.6035
0.3554
0.1350
300
0.6329
0.1755
0.0352
0.0781
0.1788
0.4185
0.6464
0.3374
0.1161
500
0.6314
0.1627
0.0311
0.0489
0.1335
0.4417
0.6405
0.3463
0.1229
Case 9
5
0.6321
0.1222
0.0195
0.6032
0.1604
0.0349
0.6158
0.2619
0.0754
50
0.5763
0.1593
0.0406
0.4105
0.3065
0.1773
0.5565
0.2759
0.0963
100
0.5759
0.1628
0.0418
0.2841
0.3098
0.2684
0.5083
0.2896
0.1202
300
0.5628
0.1687
0.0472
0.1979
0.2897
0.3356
0.5528
0.2977
0.1098
500
0.5708
0.1649
0.0437
0.1137
0.2256
0.3943
0.5074
0.3035
0.1287 Efficient change point detection and estimation in HD correlation matrices Efficient change point detection and estimation in HD correlation matrices 957 Table 7 Table 7
Normal. Mean, standard deviation (SD), mean squared error (MSE), and iter
time(ITER) of SPACE+SMOTE algorithm for Case 6 when the change point exi
extreme tail, ϵ = 10−3. 5.2. Estimation of change point p
5
50
100
300
500
T=100
β = 0.8
MEAN 0.7072 0.7904 0.7966 0.8021 0.8019
SD
0.1473 0.0523 0.0447 0.0358 0.0304
MSE
0.0302 0.0028 0.0020 0.0013 0.0009
ITER
0.9450 1.1650 1.4050 1.3050 1.2850
β = 0.9
MEAN 0.7287 0.8306 0.8364 0.8570 0.8617
SD
0.1884 0.1138 0.1077 0.0867 0.0733
MSE
0.0646 0.0177 0.0156 0.0093 0.0068
ITER
1.1350 2.0000 2.4100 2.5000 2.7900
β = 0.95
MEAN 0.6642 0.7656 0.7814 0.8231 0.8393
SD
0.2286 0.1378 0.1260 0.0968 0.0986
MSE
0.1336 0.0529 0.0442 0.0254 0.0216
ITER
0.9650 2.2200 2.7450 3.5500 4.2400
T=200
β = 0.8
MEAN 0.7739 0.8047 0.8041 0.8056 0.8058
SD
0.0758 0.0126 0.0072 0.0089 0.0097
MSE
0.0064 0.0002 0.0001 0.0001 0.0001
ITER
0.7300 0.6100 0.5450 0.4200 0.4555
β = 0.9
MEAN 0.8267 0.9021 0.9012 0.9027 0.9024
SD
0.1320 0.0099 0.0163 0.0053 0.0052
MSE
0.0227 0.0001 0.0003 0.0000 0.0000
ITER
1.3950 1.8350 1.7650 1.6200 1.6250
β = 0.95
MEAN 0.7464 0.8785 0.9113 0.9269 0.9284
SD
0.1889 0.0948 0.0710 0.0343 0.0255
MSE
0.0769 0.0141 0.0065 0.0017 0.0011
ITER
1.0200 2.9600 3.5200 4.0450 4.1250 Normal. Mean, standard deviation (SD), mean squared error (MSE), and iteration
time(ITER) of SPACE+SMOTE algorithm for Case 6 when the change point exists in the
extreme tail, ϵ = 10−3. performs slightly better than SPACE only under Case 9. For student-t samples,
SPACE is the best, while Dette’s estimator performs the worst since the mean
tends to 0 as the dimension increases. 5.3. Change point in the extreme tail To show the performance of the SPACE+SMOTE Algorithm, we explore Case
6 when the change point occurs in the extreme right tail, β = 0.8, 0.9, 0.95. The
mean change point fraction, SD, MSE, and average iterations of the algorithm
are shown in Table 7. As the dimension p increases from 5 to 500, or the sample
size T increases from 100 to 200, the mean values become very close to the
true value, and SD and MSE become smaller as expected. The algorithm is
time-efficient as small number of iterations is enough. To compare the overall performance of SPACE+SMOTE, SPACE, Dette’s
estimator and KCP-raw, Figure 4 shows the variation of the absolute value of
ˆβ −β of these four methods as the true change point fraction β changes from 0.5
to 0.95. A smaller absolute value indicates a more accurate estimated result. We
consider Case 6 and Case 7 with p = 100, T = 100, 200 for normal samples. The
SPACE+SMOTE Algorithm is the most effective one in estimating the location
of the change point in the extreme tail. SPACE is also better than Dette (2020)
and KCP-raw methods. Z. Li and J. Gao 958 Fig 4. The absolute values of β −ˆβ varies with β. Fig 4. The absolute values of β −ˆβ varies with β. 5.4. Heteroscedasticity and dependence 1/2 • Heteroscedasticity serial dependence: ψ = 0.8, et = Σ1/2
t
ηt, σ1i = · · · =
σpi1 are independently from U(1, 21). • Conditional Heteroscedasticity: ψ = 0, et = Σ1/2
t
ηt, σ2
li = (1 −α1 −α2) +
α1a2
l,i−1 + α2σ2
l,i−1, l = 1, . . . , p, (α1, α2) = (0.1, 0.89). / • Conditional Heteroscedasticity: ψ = 0, et = Σ1/2
t
ηt, σ2
li = (1 −α1 −α2) +
α1a2
l,i−1 + α2σ2
l,i−1, l = 1, . . . , p, (α1, α2) = (0.1, 0.89). 1/2 • Unconditional Heteroscedasticity: ψ = 0, et = Σ1/2
t
ηt, σli = σ0(1 + δI(i >
T/2)), σ0 = δ = 1 for all 1 ≤l ≤p. The simulation results are presented in Table 8. Under both homoscedastic-
ity serial dependence and conditional heteroscedasticity scenarios, SPACE and
Dette’s estimator perform better than the KCP estimator. Dette’s estimator
is better than SPACE in terms of mean, while SPACE is better than Dette’s
estimator in terms of SD and MSE. Under the unconditional heteroscedasticity
scenario, the KCP estimator performs the best, and SPACE performs similarly
with Dette’s estimator. 6. Real data analysis Though the proposed SPACE outperforms the existing ones in the simulation
experiments, we now compare SPACE and Dette’s estimator on two real data
sets. The KCP-raw method is omitted here due to its bad performance in the
simulations. 5.4. Heteroscedasticity and dependence Till now, homoscedasticity of random variables is assumed, which is often chal-
lenged in real data application, however. Therefore, we consider the following
heteroscedasticity setting. ˜D = diag(σ1, . . . , σp),
Σ1 = ˜DR1 ˜D,
Σ2 = ˜DR2 ˜D, where σi, i = 1, . . . , p are independently drawn from Uniform distribution U(1, 21)
We compare SPACE with Dette’s estimator and KCP-raw under Cases 6-9 for
normal and student-t distributed data, respectively. The results are similar to
that for the homoscedasticity setting in the Section 5.2. The tables are in the
supplementary materials. where σi, i = 1, . . . , p are independently drawn from Uniform distribution U(1, 21)
We compare SPACE with Dette’s estimator and KCP-raw under Cases 6-9 for
normal and student-t distributed data, respectively. The results are similar to
that for the homoscedasticity setting in the Section 5.2. The tables are in the
supplementary materials. The independence assumption on the sequence of data y1, . . . , yT is also
not common in real applications. We investigate the performance of SPACE
for dependent data. Moreover, heteroscedastic scenarios are also considered. Following [9], we consider a vector autoregression of order 1, yt = ψyt−1 + et, 959 Efficient change point detection and estimation in HD correlation matrices where the error vector et = (et1, . . . , etp)′ has zero mean and covariance matrix
Σt = Dt ˜RtDt with Dt = diag(σt1, . . . , σtp) and correlation matrix at time t, ˜Rt. Here we consider the setting of correlation matrices in Case 6, that is, ˜Rt = R1
for 1 ≤t ≤t0, ˜Rt = R2 for t0 + 1 ≤t ≤T. The following four scenarios are
considered. • Homoscedasticity serial dependence: serial dependence is considered by • Homoscedasticity serial dependence: serial dependence is considered by
ψ = 0.8, et = Σ1/2
t
ηt, σ1i = · · · = σpi = 1 and ηt = (ηt1, . . . , ηtp)′ is a
vector of standard normal random variables. 1/2 • Homoscedasticity serial dependence: serial dependence is considered by
ψ = 0.8, et = Σ1/2
t
ηt, σ1i = · · · = σpi = 1 and ηt = (ηt1, . . . , ηtp)′ is a
vector of standard normal random variables. 6.1. Gene expression profile The first data is a real gene expression profile studied by [23]. The data set
investigated the relationship between peripheral blood gene expression patterns
and coronary artery disease severity. Patients undergoing coronary angiography
were selected according to their coronary artery disease index (CADi), a val-
idated angiographical measure of the extent of coronary atherosclerosis. RNA
was extracted from the blood of 110 patients with at least a stenosis greater
than 50% (CADi ≥23) and from 112 controls without evidence of coronary
stenosis (CADi= 0). Therefore, the total sample size is n = 222. Rearrange the
data so the true change point position is t0 = 110. The raw dataset is available
at the Gene Expression Omnibus repository with the accession number Series
GSE12288. Following [10], we first apply the two sample t-tests with a significance level
0.05 to all genes, and 1410 genes are reserved. As mentioned in [23], many genes 960 Z. Li and J. Gao Table 8
Mean, standard deviation (SD), and mean squared error (MSE) of three estimators und
dependence and heteroscedasticity, β = 0.5. 6.2. FRED-MD data set The second data set is the FRED-MD, a macroeconomic database of 134 monthly
U.S. indicators from eight aspects (output and income, labor market, consump-
tion and orders, orders and inventories, money and credit, interest rate and
exchange rates, prices and stock market) from January 1959, aim to establish
a convenient starting point for empirical analysis that requires “big data” [20]. The data set is updated in a timely manner and can be downloaded for free
from the website.1 [13] used this data set to predict the risk of U.S. bond risk
premia. As the change of correlation coefficient is often accompanied by risk, we
apply the proposed method to estimate the change point location in this data
set. After removing the missing data, 127 macroeconomic indicators are retained
for nearly 20 years, that is, p = 127. The latest month is July 2022. To show
the estimation performance of these two estimators, we change the number of
months T until July 2022 from 50 to 200. Table 10 records the estimated change
point location (the exact month) for different T, with change point fraction in
parentheses. Table 10
The estimated change point month in the FRED-MD data set. T
50
100
130
150
200
start month
June 2018
April 2014
October 2011
February 2010
December 2005
SPACE
Oct. 2021
Apr. 2020
Apr. 2020
Apr. 2020
Sep. 2008
(0.82)
(0.73)
(0.79)
(0.82)
(0.17)
Dette’s
Feb. 2021
Jan. 2021
Oct. 2020
Apr. 2020
Apr. 2020
(0.66)
(0.82)
(0.84)
(0.82)
(0.87) Table 10
The estimated change point month in the FRED-MD data set. Table 10 When T = 50, the period is from June 2018 to July 2022. The SPACE
estimated change point month is October 2021. This coincided with a sharp
drop in U.S. stocks at the end of the third quarter of 2021, with the S&P 500
posting its worst monthly drop since the coronavirus outbreak in March 2020,
and the risk of stagflation loomed in the US economy, according to the reports
in The Wall Street Journal and the BBC. When T = 100, 130, 150, SPACE has been estimating the change point month
to be April 2020, which coincided with the coronavirus outbreak. The World
Health Organization (WHO) announced a global pandemic on March 11, 2020. 6.1. Gene expression profile Efficient change point detection and estimation in HD correlation matrices 961 Table 9
The true change point fraction is β = 110/222 = 0.4955. p
5
20
50
100
200
300
500
1000
1410
SPACE
0.4775
0.3694
0.4820
0.4910
0.4955
0.4955
0.4955
0.4955
0.4955
Dette’s
0.4730
0.4324
0.2342
0.2342
0.4279
0.4279
0.4279
0.2477
0.2477 1http://research.stlouisfed.org/econ/mccracken/sel/ 6.1. Gene expression profile Homoscedasticity
Serial
Dependence
SPACE
p
5
50
100
300
500
Mean
0.5203
0.5503
0.5530
0.5562
0.5594
SD
0.1136
0.0769
0.0701
0.0655
0.0739
MSE
0.0133
0.0084
0.0077
0.0074
0.0090
Dette’s
Mean
0.5093
0.5480
0.5455
0.5494
0.5566
SD
0.1387
0.0883
0.0748
0.0685
0.0818
MSE
0.0192
0.0101
0.0076
0.0071
0.0099
KCP-raw
Mean
0.5180
0.5601
0.5757
0.6306
0.6048
SD
0.2358
0.2055
0.1731
0.1700
0.1633
MSE
0.0557
0.0456
0.0355
0.0458
0.0375
Heteroscedasticity
Serial
Dependence
SPACE
Mean
0.5226
0.5381
0.5436
0.5434
0.5454
SD
0.1049
0.0536
0.0611
0.0453
0.0592
MSE
0.0115
0.0043
0.0056
0.0039
0.0055
Dette’s
Mean
0.5183
0.5357
0.5392
0.5379
0.5370
SD
0.1378
0.0668
0.0665
0.0528
0.0539
MSE
0.0192
0.0057
0.0059
0.0042
0.0043
KCP-raw
Mean
0.5573
0.6021
0.6050
0.6344
0.6252
SD
0.2604
0.2152
0.1958
0.1837
0.1664
MSE
0.0707
0.0565
0.0492
0.0516
0.0432
Conditional
Heteroscedasticity
SPACE
Mean
0.5135
0.5269
0.5242
0.5287
0.5295
SD
0.0894
0.0413
0.0401
0.0435
0.0419
MSE
0.0081
0.0024
0.0022
0.0027
0.0026
Dette’s
Mean
0.5008
0.5227
0.5269
0.5272
0.5341
SD
0.1267
0.0864
0.0886
0.0846
0.0841
MSE
0.0160
0.0079
0.0085
0.0079
0.0082
KCP-raw
Mean
0.5137
0.6094
0.6219
0.6166
0.6016
SD
0.2627
0.1767
0.1683
0.1541
0.1477
MSE
0.0689
0.0430
0.0430
0.0372
0.0320
Unconditional
Heteroscedasticity
SPACE
Mean
0.5372
0.5316
0.5316
0.5292
0.5332
SD
0.0462
0.0414
0.0416
0.0347
0.0473
MSE
0.0035
0.0027
0.0027
0.0020
0.0033
Dette’s
Mean
0.5270
0.5300
0.5326
0.5349
0.5343
SD
0.0372
0.0400
0.0439
0.0544
0.0795
MSE
0.0021
0.0025
0.0030
0.0042
0.0075
KCP-raw
Mean
0.5297
0.5149
0.5139
0.5113
0.5130
SD
0.0622
0.0267
0.0266
0.0318
0.0257
MSE
0.0047
0.0009
0.0009
0.0011
0.0008 Table 8 Mean, standard deviation (SD), and mean squared error (MSE) of three estimators under
dependence and heteroscedasticity, β = 0.5. are not correlated with CADi, and thus it is suitable to do such a preprocessing
step. are not correlated with CADi, and thus it is suitable to do such a preprocessing
step. To show the estimation performance of these two estimators, we change the
number of genes kept in estimation procedures from p = 5 to p = 1410 cor-
responding to the smallest p-values in the two sample t-tests. Table 9 records
the estimated change point fractions for different p. As p becomes large, both
estimators are closer to the true value. SPACE performs better and remains
stable when p is large enough. In particular, SPACE achieves the true value
when p ≥200, while Dette’s estimator never reaches the true value. 6.2. FRED-MD data set The New York Times reported on April 2, 2020 that the United States was
facing a dire situation, with more than 200,000 confirmed cases in the United
States, and the federal medical supplies reserve is almost exhausted. 962 Z. Li and J. Gao When T = 200, the period is from December 2005 to July 2022. The SPACE
estimated change point month is September 2008. This month, the financial
crisis started to spiral out of control and led to the failure or government takeover
of many fairly large financial institutions, triggering an economic recession. By comparison, when T = 50, 100, 130, Dette’s estimates are February 2021,
January 2021 and October 2020, respectively. However, the implications of the
changes are unclear. When T = 150, Dette’s estimator gives the same estimator
as SPACE, the time of the coronavirus outbreak. When T = 200, Detter’s
estimator also gives the timing of the coronavirus outbreak. Appendix A Appendix A
A.1. More simulation results
Algorithm 3 : SPAD+SMOTE
Input : Data Matrix Y(i) ∈Rp×T (p variables and T series), i = 1, . . . , m. small
positive number ϵ. Output : the successful detection rate under H1. 1
for i ←1 to m do
2
a(i) = card(wτ1) > 0, a(i) = 1 when there exists a change point, a(i) = 0,
otherwise. 3
end
4
α0 ←sum(a)
m
, the successful detection rate under H1 for raw data. 5
for i ←1 to m do
6
Y0(i) = Y(i). 7
let [γT + 1, T] columns of Y0(i) be the minority class, get(1 −γ)T SMOTE
variables, γ ≥0.9. 8
Y(i) ←combine Y0(i) and SMOTE variables. 9
a(i) = card(wτ1) > 0 for Y(i). 10
end
11
α1 ←sum(a)
m
, the successful detection rate for inflated data. 12
while α1 −α0 ≥ϵ do
13
α0 = α1;
14
Repeat the above steps 5–11. 15
end
16
α1 ←the successful detection rate under H1. A.1. More simulation results Algorithm 3 : SPAD+SMOTE
Input : Data Matrix Y(i) ∈Rp×T (p variables and T series), i = 1, . . . , m. small
positive number ϵ. Output : the successful detection rate under H1. 1
for i ←1 to m do
2
a(i) = card(wτ1) > 0, a(i) = 1 when there exists a change point, a(i) = 0,
otherwise. 11
α1 ←sum(a)
m
, the successful detection rate for inflated data. 14
Repeat the above steps 5–11. 7. Conclusion In this paper, we propose a break test and a change point location estimator
in the correlation structure of high-dimensional data. The main appeal of our
methods is that they are developed without any assumption on the ratio of
p and T and are thus appropriate for a wide range of large dimensional data
sets. The larger p or T, the better performance of the proposed methods. More-
over, they are also effective when the change point exists in the extreme tail
of a data seqence. Monte Carlo experiments and real data analyses demon-
strated the superiority of the proposed methods over several existing methods. In addition, when a change point exists, our methods automatically provide
support recovery in the index set wτ1 or wτ2. The proposed estimator is robust
to non-Gaussian data, although the theories are developed under the normal
assumption. There are several possible extensions of our methods. A direct extension
is to detect and estimate the change point in the covariance structure of a
sequence of high-dimensional vectors. In addition, as we have presented, signflip
permutation can break the change point structure and construct the behavior
under the null without a change point. Thus, signflip can be used in other change
point problems, including online detection. Efficient change point detection and estimation in HD correlation matrices 963 Appendix A
A.1. More simulation results
Algorithm 3 : SPAD+SMOTE
Input : Data Matrix Y(i) ∈Rp×T (p variables and T series), i = 1, . . . , m. small
positive number ϵ. Output : the successful detection rate under H1. 1
for i ←1 to m do
2
a(i) = card(wτ1) > 0, a(i) = 1 when there exists a change point, a(i) = 0,
otherwise. 3
end
4
α0 ←sum(a)
m
, the successful detection rate under H1 for raw data. 5
for i ←1 to m do
6
Y0(i) = Y(i). 7
let [γT + 1, T] columns of Y0(i) be the minority class, get(1 −γ)T SMOTE
variables, γ ≥0.9. 8
Y(i) ←combine Y0(i) and SMOTE variables. 9
a(i) = card(wτ1) > 0 for Y(i). 10
end
11
α1 ←sum(a)
m
, the successful detection rate for inflated data. 12
while α1 −α0 ≥ϵ do
13
α0 = α1;
14
Repeat the above steps 5–11. 15
end
16
α1 ←the successful detection rate under H1. Appendix A 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5184
0.1311
0.0174
0.4430
0.2088
0.0466
0.5292
0.3020
0.0916
50
0.5460
0.1050
0.0131
0.2622
0.2492
0.1184
0.5582
0.2581
0.0696
100
0.5256
0.1100
0.0127
0.1887
0.2398
0.1542
0.5710
0.2512
0.0679
300
0.5312
0.1200
0.0153
0.0909
0.1777
0.1987
0.5812
0.2483
0.0679
500
0.5279
0.1129
0.0135
0.0552
0.1293
0.2145
0.5954
0.2495
0.0710
Case 7
5
0.5028
0.2041
0.0413
0.3173
0.2686
0.1052
0.5461
0.3411
0.1179
50
0.5265
0.1502
0.0232
0.1998
0.2502
0.1524
0.5212
0.3481
0.1210
100
0.5221
0.1413
0.0204
0.1325
0.2121
0.1799
0.5111
0.3467
0.1197
300
0.5174
0.1644
0.0272
0.0658
0.1637
0.2152
0.5814
0.3465
0.1261
500
0.5223
0.1472
0.0220
0.0433
0.1114
0.2209
0.5691
0.3516
0.1278
Case 8
5
0.5363
0.1253
0.0169
0.4472
0.1955
0.0408
0.5460
0.3089
0.0971
50
0.5343
0.1042
0.0120
0.3000
0.2541
0.1042
0.5372
0.3266
0.1075
100
0.5289
0.1119
0.0133
0.1906
0.2412
0.1536
0.5614
0.3221
0.1070
300
0.5287
0.1235
0.0160
0.0866
0.1773
0.2022
0.5951
0.3162
0.1085
500
0.5233
0.1054
0.0116
0.0687
0.1554
0.2100
0.5990
0.2988
0.0986
Case 9
5
0.4936
0.0749
0.0056
0.4852
0.0825
0.0070
0.4992
0.2183
0.0474
50
0.4728
0.0948
0.0097
0.3815
0.2147
0.0599
0.4300
0.2403
0.0623
100
0.4673
0.1030
0.0116
0.2892
0.2471
0.1052
0.3961
0.2400
0.0681
300
0.4650
0.1201
0.0156
0.1903
0.2391
0.1528
0.4392
0.2579
0.0699
500
0.4680
0.1279
0.0173
0.1071
0.1887
0.1899
0.4222
0.2627
0.0747 Table 12
Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. Table 12 Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5243
0.0671
0.0051
0.4947
0.1209
0.0146
0.5238
0.2903
0.0844
50
0.5231
0.0290
0.0014
0.5000
0.0890
0.0079
0.6377
0.1996
0.0586
100
0.5231
0.0327
0.0016
0.5037
0.0738
0.0054
0.6146
0.1576
0.0378
300
0.5248
0.0344
0.0018
0.5067
0.0682
0.0047
0.6262
0.1547
0.0397
500
0.5217
0.0312
0.0014
0.4947
0.0880
0.0077
0.6168
0.1544
0.0374
Case 7
5
0.5163
0.1136
0.0131
0.4130
0.2365
0.0632
0.5645
0.3325
0.1141
50
0.5227
0.0294
0.0014
0.4883
0.1202
0.0145
0.6006
0.2812
0.0888
100
0.5236
0.0326
0.0016
0.4611
0.1408
0.0212
0.6274
0.2547
0.0808
300
0.5244
0.0349
0.0018
0.4840
0.1243
0.0156
0.6689
0.2198
0.0766
500
0.5201
0.0288
0.0012
0.4657
0.1417
0.0212
0.6624
0.2054
0.0683
Case 8
5
0.5173
0.0469
0.0025
0.5076
0.0797
0.0064
0.5381
0.2965
0.0889
50
0.5165
0.0241
0.0008
0.5086
0.0731
0.0054
0.6304
0.2269
0.0682
100
0.5130
0.0223
0.0007
0.5064
0.0589
0.0035
0.6332
0.1978
0.0567
300
0.5187
0.0309
0.0013
0.5165
0.0379
0.0017
0.6837
0.1771
0.0650
500
0.5160
0.0249
0.0009
0.5099
0.0614
0.0038
0.6636
0.1709
0.0558
Case 9
5
0.4869
0.0316
0.0012
0.4845
0.0410
0.0019
0.4726
0.2160
0.0472
50
0.4761
0.0351
0.0018
0.4841
0.0429
0.0021
0.3944
0.1788
0.0430
100
0.4759
0.0365
0.0019
0.4877
0.0225
0.0007
0.3758
0.1649
0.0425
300
0.4758
0.0294
0.0014
0.4882
0.0252
0.0008
0.3655
0.1574
0.0427
500
0.4760
0.0337
0.0017
0.4890
0.0416
0.0018
0.3605
0.1695
0.0481 Table 13 Heteroscedasticity setting. Student-t distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. 16
α1 ←the successful detection rate under H1. Table 11 shows the result of the success rate of detecting the existence of
the change point over 200 times repetitions under H1 when the change point
occurs in the tail(β = 0.9, 0.95), and the correlation matrix before and after
the change point is the same as Case 6 setting. We let ϵ = 0.05 in order to
improve computing efficiency. Compared with the result of Case 3 in Table 2,
where β = 0.75, when the position of the change point is closer to the tail, the
proportion of successful detection of the change point is greatly increased, which
also verifies the effectiveness of the iterative algorithm proposed in this paper. Table 11 Table 11 Table 11
The success rate of detection change point under H1 and iteration time (ITER) of
SPAD+SMOTE estimator under Case 6 when the change point exists in the extreme tail,
ϵ = 0.05, T=100. p
20
50
100
200
300
500
β = 0.9
Success rate
0.8108
0.8672
0.8961
0.9138
0.9228
0.9327
ITER
3.5650
3.2700
3.1550
3.1150
3.0800
3.0000
β = 0.95
Success rate
0.6656
0.7950
0.8492
0.8879
0.8984
0.9082
ITER
3.9800
4.0800
3.9600
3.7900
3.7650
3.6300 The success rate of detection change point under H1 and iteration time (ITER) of
SPAD+SMOTE estimator under Case 6 when the change point exists in the extreme tail, The following four Tables 12, 13, 14 and 15 contain the simulation results
under heteroscedasticity setting in Section 5.4. 964 Z. Li and J. Gao Table 12
Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5243
0.0671
0.0051
0.4947
0.1209
0.0146
0.5238
0.2903
0.0844
50
0.5231
0.0290
0.0014
0.5000
0.0890
0.0079
0.6377
0.1996
0.0586
100
0.5231
0.0327
0.0016
0.5037
0.0738
0.0054
0.6146
0.1576
0.0378
300
0.5248
0.0344
0.0018
0.5067
0.0682
0.0047
0.6262
0.1547
0.0397
500
0.5217
0.0312
0.0014
0.4947
0.0880
0.0077
0.6168
0.1544
0.0374
Case 7
5
0.5163
0.1136
0.0131
0.4130
0.2365
0.0632
0.5645
0.3325
0.1141
50
0.5227
0.0294
0.0014
0.4883
0.1202
0.0145
0.6006
0.2812
0.0888
100
0.5236
0.0326
0.0016
0.4611
0.1408
0.0212
0.6274
0.2547
0.0808
300
0.5244
0.0349
0.0018
0.4840
0.1243
0.0156
0.6689
0.2198
0.0766
500
0.5201
0.0288
0.0012
0.4657
0.1417
0.0212
0.6624
0.2054
0.0683
Case 8
5
0.5173
0.0469
0.0025
0.5076
0.0797
0.0064
0.5381
0.2965
0.0889
50
0.5165
0.0241
0.0008
0.5086
0.0731
0.0054
0.6304
0.2269
0.0682
100
0.5130
0.0223
0.0007
0.5064
0.0589
0.0035
0.6332
0.1978
0.0567
300
0.5187
0.0309
0.0013
0.5165
0.0379
0.0017
0.6837
0.1771
0.0650
500
0.5160
0.0249
0.0009
0.5099
0.0614
0.0038
0.6636
0.1709
0.0558
Case 9
5
0.4869
0.0316
0.0012
0.4845
0.0410
0.0019
0.4726
0.2160
0.0472
50
0.4761
0.0351
0.0018
0.4841
0.0429
0.0021
0.3944
0.1788
0.0430
100
0.4759
0.0365
0.0019
0.4877
0.0225
0.0007
0.3758
0.1649
0.0425
300
0.4758
0.0294
0.0014
0.4882
0.0252
0.0008
0.3655
0.1574
0.0427
500
0.4760
0.0337
0.0017
0.4890
0.0416
0.0018
0.3605
0.1695
0.0481
Table 13
Heteroscedasticity setting. Student-t distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.5. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5954
0.1710
0.0400
0.4872
0.2634
0.1143
0.5588
0.3283
0.1272
50
0.6262
0.1598
0.0308
0.2359
0.2727
0.2894
0.6408
0.2990
0.0925
100
0.6235
0.1558
0.0300
0.1740
0.2600
0.3440
0.6515
0.3048
0.0948
300
0.6138
0.1778
0.0389
0.0703
0.1703
0.4254
0.6967
0.2714
0.0733
500
0.6363
0.1502
0.0265
0.0369
0.0953
0.4487
0.6984
0.2647
0.0697
Case 7
5
0.5393
0.2279
0.0773
0.3236
0.2819
0.2208
0.5359
0.3454
0.1456
50
0.5846
0.1902
0.0493
0.1735
0.2610
0.3450
0.5353
0.3669
0.1611
100
0.5826
0.1798
0.0460
0.1201
0.2186
0.3839
0.5593
0.3730
0.1583
300
0.5602
0.2120
0.0643
0.0621
0.1671
0.4346
0.6358
0.3625
0.1348
500
0.5826
0.2000
0.0536
0.0417
0.1207
0.4478
0.5956
0.3699
0.1470
Case 8
5
0.6353
0.1584
0.0292
0.5101
0.2680
0.1076
0.5703
0.3236
0.1210
50
0.6562
0.1421
0.0220
0.2894
0.3041
0.2607
0.6012
0.3516
0.1327
100
0.6296
0.1600
0.0304
0.1586
0.2568
0.3588
0.6035
0.3554
0.1350
300
0.6329
0.1755
0.0352
0.0781
0.1788
0.4185
0.6464
0.3374
0.1161
500
0.6314
0.1627
0.0311
0.0489
0.1335
0.4417
0.6405
0.3463
0.1229
Case 9
5
0.6321
0.1222
0.0195
0.6033
0.1604
0.0349
0.6158
0.2619
0.0754
50
0.5763
0.1593
0.0406
0.4105
0.3065
0.1773
0.5565
0.2759
0.0963
100
0.5759
0.1628
0.0418
0.2842
0.3098
0.2684
0.5083
0.2896
0.1202
300
0.5628
0.1687
0.0472
0.1980
0.2897
0.3356
0.5529
0.2977
0.1098
500
0.5708
0.1649
0.0437
0.1138
0.2256
0.3943
0.5074
0.3035
0.1287 Efficient change point detection and estimation in HD correlation matrices 965 ffi
g
p
Table 14
Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the tail β = 0.7. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.6315
0.1072
0.0161
0.5902
0.1725
0.0417
0.6013
0.3078
0.1040
50
0.6981
0.0311
0.0010
0.6116
0.1313
0.0250
0.7564
0.1731
0.0330
100
0.6959
0.0328
0.0011
0.5772
0.1667
0.0427
0.7768
0.1412
0.0257
300
0.7015
0.0265
0.0007
0.5906
0.1634
0.0386
0.7777
0.1052
0.0170
500
0.6988
0.0237
0.0006
0.5821
0.1433
0.0343
0.7847
0.1062
0.0184
Case 7
5
0.5744
0.1518
0.0387
0.4319
0.2590
0.1387
0.5664
0.3363
0.1304
50
0.6764
0.0530
0.0034
0.5308
0.2189
0.0763
0.6371
0.3065
0.0974
100
0.6768
0.0509
0.0031
0.5011
0.2292
0.0918
0.6759
0.2988
0.0894
300
0.6861
0.0446
0.0022
0.5575
0.2030
0.0613
0.7565
0.2159
0.0496
500
0.6855
0.0419
0.0020
0.4975
0.2485
0.1024
0.7678
0.2036
0.0458
Case 8
5
0.6690
0.0738
0.0064
0.6354
0.1379
0.0231
0.5718
0.3155
0.1155
50
0.7034
0.0168
0.0003
0.6394
0.1290
0.0202
0.7012
0.2520
0.0632
100
0.7039
0.0184
0.0004
0.6381
0.1196
0.0181
0.7408
0.2243
0.0517
300
0.7032
0.0115
0.0001
0.6490
0.0915
0.0109
0.8097
0.1221
0.0269
500
0.7060
0.0140
0.0002
0.6380
0.1241
0.0192
0.7920
0.1399
0.0279
Case 9
5
0.6585
0.0622
0.0056
0.6505
0.0796
0.0088
0.6122
0.2382
0.0642
50
0.6410
0.0693
0.0083
0.6375
0.1098
0.0159
0.5339
0.2527
0.0911
100
0.6293
0.0820
0.0117
0.6484
0.0779
0.0087
0.5192
0.2344
0.0873
300
0.6223
0.0922
0.0145
0.6333
0.1132
0.0172
0.5051
0.2329
0.0919
500
0.6239
0.0878
0.0135
0.6187
0.1295
0.0233
0.4964
0.2504
0.1039 Table 14 Table 14
Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the tail β = 0.7. Table 15 Heteroscedasticity setting. Student-t distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.7. squared error (MSE) of three estimators when the change point exists in the middle β
0.7. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5184
0.1311
0.0174
0.4430
0.2088
0.0466
0.5292
0.3020
0.0916
50
0.5460
0.1050
0.0131
0.2622
0.2492
0.1184
0.5582
0.2581
0.0696
100
0.5256
0.1100
0.0127
0.1887
0.2398
0.1542
0.5710
0.2512
0.0679
300
0.5312
0.1200
0.0153
0.0909
0.1777
0.1987
0.5812
0.2483
0.0679
500
0.5279
0.1129
0.0135
0.0552
0.1293
0.2145
0.5954
0.2495
0.0710
Case 7
5
0.5028
0.2041
0.0413
0.3173
0.2686
0.1052
0.5461
0.3411
0.1179
50
0.5265
0.1502
0.0232
0.1998
0.2502
0.1524
0.5212
0.3481
0.1210
100
0.5221
0.1413
0.0204
0.1325
0.2121
0.1799
0.5111
0.3467
0.1197
300
0.5174
0.1644
0.0272
0.0658
0.1637
0.2152
0.5814
0.3465
0.1261
500
0.5223
0.1472
0.0220
0.0433
0.1114
0.2209
0.5691
0.3516
0.1278
Case 8
5
0.5363
0.1253
0.0169
0.4472
0.1955
0.0408
0.5460
0.3089
0.0971
50
0.5343
0.1042
0.0120
0.3000
0.2541
0.1042
0.5372
0.3266
0.1075
100
0.5289
0.1119
0.0133
0.1906
0.2412
0.1536
0.5614
0.3221
0.1070
300
0.5287
0.1235
0.0160
0.0866
0.1773
0.2022
0.5951
0.3162
0.1085
500
0.5233
0.1054
0.0116
0.0687
0.1554
0.2100
0.5990
0.2988
0.0986
Case 9
5
0.4936
0.0749
0.0056
0.4852
0.0825
0.0070
0.4992
0.2183
0.0474
50
0.4728
0.0948
0.0097
0.3815
0.2147
0.0599
0.4300
0.2403
0.0623
100
0.4673
0.1030
0.0116
0.2892
0.2471
0.1052
0.3961
0.2400
0.0681
300
0.4650
0.1201
0.0156
0.1903
0.2391
0.1528
0.4392
0.2579
0.0699
500
0.4680
0.1279
0.0173
0.1071
0.1887
0.1899
0.4222
0.2627
0.0747 Efficient change point detection and estimation in HD correlation matrices
965
Table 14
Heteroscedasticity setting. Normal distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the tail β = 0.7. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.6315
0.1072
0.0161
0.5902
0.1725
0.0417
0.6013
0.3078
0.1040
50
0.6981
0.0311
0.0010
0.6116
0.1313
0.0250
0.7564
0.1731
0.0330
100
0.6959
0.0328
0.0011
0.5772
0.1667
0.0427
0.7768
0.1412
0.0257
300
0.7015
0.0265
0.0007
0.5906
0.1634
0.0386
0.7777
0.1052
0.0170
500
0.6988
0.0237
0.0006
0.5821
0.1433
0.0343
0.7847
0.1062
0.0184
Case 7
5
0.5744
0.1518
0.0387
0.4319
0.2590
0.1387
0.5664
0.3363
0.1304
50
0.6764
0.0530
0.0034
0.5308
0.2189
0.0763
0.6371
0.3065
0.0974
100
0.6768
0.0509
0.0031
0.5011
0.2292
0.0918
0.6759
0.2988
0.0894
300
0.6861
0.0446
0.0022
0.5575
0.2030
0.0613
0.7565
0.2159
0.0496
500
0.6855
0.0419
0.0020
0.4975
0.2485
0.1024
0.7678
0.2036
0.0458
Case 8
5
0.6690
0.0738
0.0064
0.6354
0.1379
0.0231
0.5718
0.3155
0.1155
50
0.7034
0.0168
0.0003
0.6394
0.1290
0.0202
0.7012
0.2520
0.0632
100
0.7039
0.0184
0.0004
0.6381
0.1196
0.0181
0.7408
0.2243
0.0517
300
0.7032
0.0115
0.0001
0.6490
0.0915
0.0109
0.8097
0.1221
0.0269
500
0.7060
0.0140
0.0002
0.6380
0.1241
0.0192
0.7920
0.1399
0.0279
Case 9
5
0.6585
0.0622
0.0056
0.6505
0.0796
0.0088
0.6122
0.2382
0.0642
50
0.6410
0.0693
0.0083
0.6375
0.1098
0.0159
0.5339
0.2527
0.0911
100
0.6293
0.0820
0.0117
0.6484
0.0779
0.0087
0.5192
0.2344
0.0873
300
0.6223
0.0922
0.0145
0.6333
0.1132
0.0172
0.5051
0.2329
0.0919
500
0.6239
0.0878
0.0135
0.6187
0.1295
0.0233
0.4964
0.2504
0.1039
Table 15
Heteroscedasticity setting. Student-t distribution. Mean, standard deviation (SD), and mean
squared error (MSE) of three estimators when the change point exists in the middle β = 0.7. A.2.1. Proof of Theorem 3.1 A.2.1. Proof of Theorem 3.1 In Theorem 3.1, we prove P
(i,j)∈N
{w(i, j) > τ2}
→0 under Assumptions 1
& 3. Without loss of generality, we can assume ρ1(i, j) = ρ2(i, j) = 0 for any
1 ≤i, j ≤p. Observing In Theorem 3.1, we prove P
(i,j)∈N
{w(i, j) > τ2}
→0 under Assumptions 1
& 3. Without loss of generality, we can assume ρ1(i, j) = ρ2(i, j) = 0 for any
1 ≤i j ≤p Observing In Theorem 3.1, we prove P
(i,j)∈N
{w(i, j) > τ2}
→0 under Assumptions 1 In Theorem 3.1, we prove P
(i,j)∈N
{w(i, j) > τ2}
→0 under Assumptions 1
& 3. Without loss of generality, we can assume ρ1(i, j) = ρ2(i, j) = 0 for any
1 ≤i, j ≤p. Observing In Theorem 3.1, we prove P
(i,j)∈N
{w(i, j) > τ2}
→0 under Assumptions 1
& 3. Without loss of generality, we can assume ρ1(i, j) = ρ2(i, j) = 0 for any
1 ≤i, j ≤p. Observing P
⎧
⎨
⎩
(i,j)∈N
{w(i, j) > τ2}
⎫
⎬
⎭≤
(i,j)∈N
P{w(i, j) > τ2} ≤1
2(p2 −p)P{w(i, j) > τ2},
(22) (22) then it is sufficient to verify P{w(i, j) > τ2} = o
1
p2
. then it is sufficient to verify P{w(i, j) > τ2} = o
1
p2
. then it is sufficient to verify P{w(i, j) > τ2} = o
1
p2
.
The components of the vector w corresponding to the entry in the position
(i, j) (1 ≤i, j ≤p) of the correlation matrices ˆR1, ˆR2 are given by The components of the vector w corresponding to the entry in the position
, j) (1 ≤i, j ≤p) of the correlation matrices ˆR1, ˆR2 are given by w(i, j) =
1
T −3
T −2
t=2
t(T −t)
T
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
2 As t varies from 2 to T −2, the weight t(T −t)
T
has relatively larger values when
⌊
√
T⌋+ 1 ≤t ≤T −⌊
√
T⌋−1, compared to its values when 2 ≤t ≤⌊
√
T⌋and
T−⌊
√
T⌋≤t ≤T−2. 16
α1 ←the successful detection rate under H1. p
SPACE
Dette’s
KCP-raw
MEAN
SD
MSE
MEAN
SD
MSE
MEAN
SD
MSE
Case 6
5
0.5954
0.1710
0.0400
0.4872
0.2634
0.1143
0.5588
0.3283
0.1272
50
0.6262
0.1598
0.0308
0.2359
0.2727
0.2894
0.6408
0.2990
0.0925
100
0.6235
0.1558
0.0300
0.1740
0.2600
0.3440
0.6515
0.3048
0.0948
300
0.6138
0.1778
0.0389
0.0703
0.1703
0.4254
0.6967
0.2714
0.0733
500
0.6363
0.1502
0.0265
0.0369
0.0953
0.4487
0.6984
0.2647
0.0697
Case 7
5
0.5393
0.2279
0.0773
0.3236
0.2819
0.2208
0.5359
0.3454
0.1456
50
0.5846
0.1902
0.0493
0.1735
0.2610
0.3450
0.5353
0.3669
0.1611
100
0.5826
0.1798
0.0460
0.1201
0.2186
0.3839
0.5593
0.3730
0.1583
300
0.5602
0.2120
0.0643
0.0621
0.1671
0.4346
0.6358
0.3625
0.1348
500
0.5826
0.2000
0.0536
0.0417
0.1207
0.4478
0.5956
0.3699
0.1470
Case 8
5
0.6353
0.1584
0.0292
0.5101
0.2680
0.1076
0.5703
0.3236
0.1210
50
0.6562
0.1421
0.0220
0.2894
0.3041
0.2607
0.6012
0.3516
0.1327
100
0.6296
0.1600
0.0304
0.1586
0.2568
0.3588
0.6035
0.3554
0.1350
300
0.6329
0.1755
0.0352
0.0781
0.1788
0.4185
0.6464
0.3374
0.1161
500
0.6314
0.1627
0.0311
0.0489
0.1335
0.4417
0.6405
0.3463
0.1229
Case 9
5
0.6321
0.1222
0.0195
0.6033
0.1604
0.0349
0.6158
0.2619
0.0754
50
0.5763
0.1593
0.0406
0.4105
0.3065
0.1773
0.5565
0.2759
0.0963
100
0.5759
0.1628
0.0418
0.2842
0.3098
0.2684
0.5083
0.2896
0.1202
300
0.5628
0.1687
0.0472
0.1980
0.2897
0.3356
0.5529
0.2977
0.1098
500
0.5708
0.1649
0.0437
0.1138
0.2256
0.3943
0.5074
0.3035
0.1287 Z. Li and J. Gao 966 A.2. Proofs of main results We first introduce some auxiliary results that are frequently used in the proofs. Lemma A.1 (Lemma 2.7.7 in [25]). Consider the samples xt = (xt1, . . . , xtp)′,
t = 1, . . . , T satisfying Assumption 1. For any 1 ≤i ≤j ≤p, xti and xtj
are sub-Gaussian random variables, then xtixtj is a sub-exponential random
variable. Lemma A.2 (Proposition 2.7.1 in [25]). Let xti, t = 1, . . . , T be a sub-exponential
random variable, then there exist positive constants c1, c2 > 0 such that for every
a > 0, P(|xti| ≥a) ≤c1e−c2d. Lemma A.3 (Corollary 2.8.3 in [25]). Let x1, . . . , xT be independent, mean zero,
sub-exponential random variables, then there exist positive constants c1, c2 > 0
such that for every a > 0, P
1
T
T
t=1
xt
≥a
≤c1e−c2T ·min(a2,a). A.2.1. Proof of Theorem 3.1 In addition, when T is big enough,
√
T can be pretty small,
and the two tails of t(T −t)
T
show symmetric form. Moreover, both the two tails
only involve (⌊
√
T⌋−1) terms, and thus the coefficient
1
T −3 = ⌊
√
T ⌋−1
T −3
·
1
⌊
√
T ⌋−1 Efficient change point detection and estimation in HD correlation matrices 967 gives an extra factor of order
1
√
T in the calculations. Thus, we can decompose
w(i, j) in terms of t into three parts, w(i, j) = w(1)(i, j) + w(2)(i, j) + w(3)(i, j), where w(1)(i, j) corresponds to t
∈
! 2, ⌊
√
T⌋
"
, w(2)(i, j) corresponds to
t ∈
! ⌊
√
T⌋+ 1, T −⌊
√
T⌋−1
"
, w(3)(i, j) corresponds to t ∈
! T −⌊
√
T⌋, T −2
"
. Therefore, in order to prove 1
2(p2 −p)P{w(i, j) > τ2} →0, it is sufficient to
prove p2P{w(h)(i, j) > cτ2} →0, h = 1, 2, 3. In this inequality and hereafter in the proof, c and ci (i = 1, 2, . . .) indicate some
positive constants that may change from line to line. In this inequality and hereafter in the proof, c and ci (i = 1, 2, . . .) indicate some
positive constants that may change from line to line. The case h = 1, 3:
For the index h = 1 and h = 3 the arguments are quite similar, and for brevity,
we only consider the case h=1. A.2.1. Proof of Theorem 3.1 For the statistic w(1)(i, j), we find that P
w(1)(i, j) ≥cτ2
P
w(1)(i, j) ≥cτ2
= P
⎧
⎨
⎩
⌊
√
T⌋−1
T −3
·
1
⌊
√
T⌋−1
⌊
√
T ⌋
t=2
t(T −t)
T
⎛
⎝1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
⎞
⎠
2
> cτ2
⎫
⎬
⎭
≤
[
√
T ]
t=2
P
⎧
⎨
⎩
1
t
t
i=k
xkixkj −
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎫
⎬
⎭
≤
⌊
√
T ⌋
t=2
⎧
⎨
⎩P
⎛
⎝
1
t
t
k=1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠+ P
⎛
⎝
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠
⎫
⎬
⎭. Since xti and xtj are sub-Gaussian random variables and xtixtj is a sub-exponential
random variable based on Lemma A.1. Then, according to Lemmas A.2 & A.3,
we have P
⎛
⎝
1
t
t
k=1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠≤c1e
−c2t min
(
cτ2T
√
T
t(T −t) , cτ2T
√
T
t(T −t)
≤c1e
−c2 min
T 1/4√τ2,τ2
√
T
≤c1e−c2T 1/4√τ2,
d P
⎛
⎝
1
t
t
k=1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠≤c1e
−c2t min
(
cτ2T
√
T
t(T −t) , cτ2T
√
T
t(T −t) and P
⎛
⎝
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠≤c1e
−c2(T −t) min
(
cτ2T
√
T
t(T −t) , cτ2T
√
T
t(T −t)
≤c1e
−c2 min
(
τ2T (T −
√
T ),τ2T
≤c1e−c2
(
τ2T (T −
√
T ), P
⎛
⎝
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠≤c1e
−c2(T −t) min
(
cτ2T
√
T
t(T −t) , cτ2T
√
T
t(T −t) ⎠
≤c1e
−c2 min
(
τ2T (T −
√
T ),τ2T
≤c1e−c2
(
τ2T (T −
√
T ), Z. Li and J. Gao 968 for t = 2, . . . ,
√
T. A.2.1. Proof of Theorem 3.1 Therefore, P
w(1)(i, j) ≥cτ2
≤
⌊
√
T ⌋
t=2
c1e−c2
(
τ2
√
T + c3e−c4
(
τ2T (T −
√
T )
≤c1
√
Te−c2T 1/4√τ2, as the smallest absolute value between the exponents is T 1/4√τ2. Under As-
sumption 3, we have as the smallest absolute value between the exponents is T 1/4√τ2. Under As-
sumption 3, we have p2 · P
w(1)(i, j) > cτ2
≤c1p2√
T · e−c2T 1/4√τ2 →0. (23) (23) The case h = 2:
For the statistic w(2)(i, j), we have The case h = 2:
For the statistic w(2)(i, j), we have P
w(2)(i, j) > cτ2
P
w(2)(i, j) > cτ2
P
w(2)(i, j) > cτ2
= P
⎧
⎨
⎩
1
T −3
T −⌊
√
T ⌋−1
t=⌊
√
T ⌋+1
t(T −t)
T
⎛
⎝1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
⎞
⎠
2
> cτ2
⎫
⎬
⎭
≤
T −⌊
√
T ⌋−1
t=⌊
√
T ⌋+1
P
⎛
⎝
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
t(T −t)
⎞
⎠
≤
T −⌊
√
T ⌋−1
t=⌊
√
T ⌋+1
⎧
⎨
⎩P
1
t
t
k=1
xkixkj
>
'
cτ2T
t(T −t)
+ P
⎛
⎝
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
t(T −t)
⎞
⎠
⎫
⎬
⎭ Then according to Lemma A.3, we have Then according to Lemma A.3, we have P
1
t
t
k=1
xkixkj
>
'
cτ2T
t(T −t)
≤c1e
−c2t min
(
cτ2T
t(T −t) ,
cτ2T
t(T −t)
≤c1e−c2 min(T 1/4√τ2,τ2), and P
⎛
⎝
1
T −t
T
k=t+1
xkixkj
>
'
cτ2T
√
T
t(T −t)
⎞
⎠≤c1e
−c2(T −t) min
(
cτ2T
t(T −t) ,
cτ2T
t(T −t)
≤c1e
−c2 min
T 1/4√τ2,τ2
, for t = ⌊
√
T⌋+ 1, . . . , T −⌊
√
T⌋−1. Therefore, we have
P
w(2)(i, j) > cτ2
≤c1Te−c2 min(T 1/4√τ2,τ2). Under Assumption 3, we have Under Assumption 3, we have p2 · P
w(2)(i, j) > cτ2
≤c1p2T · e−c2 min(T 1/4√τ2,τ2) →0. We complete the proof of Theorem 3.1. A.2.1. Proof of Theorem 3.1 Efficient change point detection and estimation in HD correlation matrices
969 Efficient change point detection and estimation in HD correlation matrices
969 969 A.2.2. Proof of Theorem 3.2 A.2.2. Proof of Theorem 3.2 A.2.2. Proof of Theorem 3.2 The result of Theorem 3.2 is equivalent to The result of Theorem 3.2 is equivalent to The result of Theorem 3.2 is equivalent to P
⎧
⎨
⎩
(i,j)∈P
(w(i, j) ≤τ2)
⎫
⎬
⎭→0. As P
⎧
⎨
⎩
(i,j)∈P
(w(i, j) ≤τ2)
⎫
⎬
⎭≤
(i,j)∈P
P(w(i, j) ≤τ2) ≤p2 · P(w(i, j) ≤τ2) →0, it is sufficient to prove p2 · P(w(i, j) ≤τ2) →0. A straightforward calculation
gives it is sufficient to prove p2 · P(w(i, j) ≤τ2) →0. A straightforward calculation
gives E
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
=
⎧
⎪
⎨
⎪
⎩
T −t0
T −t (ρ1(i, j) −ρ2(i, j)) ,
t ≤t0
t0
t (ρ1(i, j) −ρ2(i, j)) ,
t > t0
Let
At = 1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj−E
1
t
t
k=1
xkixkj−
1
T −t
T
k=t+1
xkixkj
,
Bt = E
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
, At = 1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj−E
1
t
t
k=1
xkixkj−
1
T −t
T
k=t+1
xkixkj
,
Bt = E
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
, then w(i, j) =
1
T −3
T −2
t=2
t(T −t)
T
(At + Bt)2. When t ≤t0, Bt ≥T −t0
T
(ρ1(i, j) −
ρ2(i, j)), when t > t0, Bt ≥t0
T (ρ1(i, j)−ρ2(i, j)), so Bt ≥min( T −t0
T
, t0
T )(ρ1(i, j)−
ρ2(i, j)) and
1
|Bt| ≥
1
|ρ1(i,j)−ρ2(i,j)|. We have then w(i, j) =
1
T −3
T −2
t=2
t(T −t)
T
(At + Bt)2. When t ≤t0, Bt ≥T −t0
T
(ρ1(i, j) −
ρ2(i, j)), when t > t0, Bt ≥t0
T (ρ1(i, j)−ρ2(i, j)), so Bt ≥min( T −t0
T
, t0
T )(ρ1(i, j)−
ρ2(i, j)) and
1
|Bt| ≥
1
|ρ1(i,j)−ρ2(i,j)|. A.2.1. Proof of Theorem 3.1 We have then w(i, j) =
1
T −3
T −2
t=2
t(T −t)
T
(At + Bt)2. When t ≤t0, Bt ≥T −t0
T
(ρ1(i, j) −
ρ2(i, j)), when t > t0, Bt ≥t0
T (ρ1(i, j)−ρ2(i, j)), so Bt ≥min( T −t0
T
, t0
T )(ρ1(i, j)−
ρ2(i, j)) and
1
|Bt| ≥
1
|ρ1(i,j)−ρ2(i,j)|. We have 1
T −3
T −2
t=2
t(T −t)
T
B2
t ≥
1
T −3
T −2
t=2
t(T −t)
T
min
(T −t0)2
T 2
, t2
0
T 2
· (ρ1(i, j) −ρ2(i, j))2
≥min
(T −t0)2
T 2
, t2
0
T 2
· Tλ2
6
≥min
(T −t0)2
T 2
, t2
0
T 2
· T
6 · cτ2
T · max
T 2
(T −t0)2 , T 2
t2
0
= cτ2
6 ,
(24) (24) according to Assumption 2, and then, with a sufficiently large constant c ≥12,
we obtain
T
2 τ2 ≤
1
2(T −3)
T −2
t=2
t(T −t)
T
B2
t . Z. Li and J. Gao 970 Thus, we can have Thus, we can have P(w(i, j) ≤τ2)
= P
1
T −3
T −2
t=2
t(T −t)
T
(At + Bt)2 ≤τ2
≤P
2
T −3
T −2
t=2
t(T −t)
T
AtBt ≤τ2 −
1
T −3
T −2
t=2
t(T −t)
T
B2
t
≤P
2
T −3
T −2
t=2
t(T −t)
T
AtBt
≥
−
1
2(T −3)
T −2
t=2
t(T −t)
T
B2
t
≤
T −2
t=2
P
|At| ≥
T
2t(T −t)
·
1
|Bt| ·
1
2(T −3)
T −2
t=2
t(T −t)
T
B2
t
. (25) (25) From (24), it is easy to obtain From (24), it is easy to obtain 1
T −3
T −2
t=2
t(T −t)
T
B2
t ≥c · T min
(T −t0)2
T 2
, t2
0
T 2
|ρ1(i, j) −ρ2(i, j)|2, then the right-hand side within the probability function in (25) is T
2t(T −t)
·
1
|Bt| ·
1
2(T −3)
T −2
t=2
t(T −t)
T
B2
t
≥
cT 2
t(T −t) min
(T −t0)2
T 2
, t2
0
T 2
|ρ1(i, j) −ρ2(i, j)|
≥
cT
t(T −t)
τ2T. A.2.1. Proof of Theorem 3.1 Then P(w(i, j) ≤τ2) in (25) becomes Then P(w(i, j) ≤τ2) in (25) becomes
P(w(i, j) ≤τ2) ≤TP
|At| ≥
cT
t(T −t)
τ2T
≤TP
⎧
⎨
⎩
1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj −E
⎛
⎝1
t
t
k=1
xkixkj −
1
T −t
T
k=t+1
xkixkj
⎞
⎠
≥
cT
t(T −t)
τ2T
≤TP
⎧
⎨
⎩
1
t
t
k=1
xkixkj −E
⎛
⎝1
t
T
k=t+1
xkixkj
⎞
⎠
≥
cT
t(T −t)
τ2T
⎫
⎬
⎭
+ TP
⎧
⎨
⎩
1
T −t
T
k=t+1
xkixkj −E
⎛
⎝
1
T −t
T
k=t+1
xkixkj
⎞
⎠
≥
cT
t(T −t)
τ2T
⎫
⎬
⎭
≤TP
1
t
t
k=1
xkixkj
≥cT√τ2T
t(T −t)
+ TP
⎧
⎨
⎩
1
T −t
T
k=t+1
xkixkj
≥cT√τ2T
t(T −t)
⎫
⎬
⎭. Efficient change point detection and estimation in HD correlation matrices Efficient change point detection and estimation in HD correlation matrices
971 971 According to Lemma A.3, for t ∈[2, T −2], we have According to Lemma A.3, for t ∈[2, T −2], we have P
1
t
t
k=1
xkixkj
≥cT√τ2T
t(T −t)
≤c1e
−c2t min
cT
τ2T
t(T −t) ,
cT 3τ2
t2(T −t)2
≤c1e−c2 min(√τ2T ,τ2T ) ≤c1e−c2
√τ2T , P
1
t
t
k=1
xkixkj
≥cT√τ2T
t(T −t)
≤c1e
−c2t min
cT
τ2T
t(T −t) ,
cT 3τ2
t2(T −t)2
≤c1e−c2 min(√τ2T ,τ2T ) ≤c1e−c2
√τ2T , and P
1
T −t
T
k=t+1
xkixkj
≥cT√τ2T
t(T −t)
≤c1e
−c2(T −t) min
cT
τ2T
t(T −t) ,
cT 3τ2
t2(T −t)2
≤c1e−c2 min(√τ2T ,τ2T ) ≤c1e−c2
√τ2T , P
1
T −t
T
k=t+1
xkixkj
≥cT√τ2T
t(T −t)
≤c1e
−c2(T −t) min
cT
τ2T
t(T −t) ,
cT 3τ2
t2(T −t)2 finally, we have P(w(i, j) ≤τ2) ≤c1Te−c2
√τ2T . Then under Assumption 3, we have p2 · P(w(i, j) ≤τ2) ≤c1p2Te−c2
√τ2T →0,
we complete the proof. A.2.3. Proof of Theorem 3.3 To prove the consistency of the estimator ˆt defined in Equation (13), we rewrite P
ˆβ
β −1
≥ϵ
= P
ˆβ ≥(1 + ϵ)β
+ P
ˆβ ≤(1 −ϵ)β
. A.2.1. Proof of Theorem 3.1 We only consider the first term because the second term can be handled simi-
larly. If we want to prove P{ˆβ ≥β(1 + ϵ)} →0, P{ˆβ ≥β(1 + ϵ)} →0, it is sufficient to show that P
⎧
⎨
⎩
ˆt≥(1+ϵ)t0
(UT (t) ≥UT (t0))
⎫
⎬
⎭
≤
T
t≥(1+ϵ)t0
P {UT (t) ≥UT (t0)}
≤
T· P (UT (t) ≥UT (t0)) →0. (26) (26) Z. Li and J. Gao 972 Define the vectors mk = (mk1, . . . , mkd)′ = zk −E(zk), then Define the vectors mk = (mk1, . . . , mkd)′ = zk −E(zk), then k
(
k1,
,
kd)
k
UT (t) =
1
T 4
t
i,k=1
T
j,l=t+1
(zi −zj)′ (zk −zl)
=
1
T 4
t
i,k=1
T
j,l=t+1
[(mi −mj) + (E(zi) −E(zj))]′ [(
=
1
T 4
t
i,k=1
T
j,l=t+1
(mi −mj)′ (mk −ml)
+ 1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (mk −ml)
+ 1
T 4
t
i,k=1
T
j,l=t+1
(E(zk) −E(zl))′ (mi −mj)
+ 1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (E(zk) −E(zl)
= f(t) + g(t) + h(t) + δ(t). UT (t) =
1
T 4
t
i,k=1
T
j,l=t+1
(zi −zj)′ (zk −zl) =
1
T 4
t
i,k=1
T
j,l=t+1
[(mi −mj) + (E(zi) −E(zj))]′ [(mk −ml) + (E(zk) −E(zl))] =
1
T 4
t
i,k=1
T
j,l=t+1
(mi −mj)′ (mk −ml) + 1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (mk −ml) + 1
T 4
t
i,k=1
T
j,l=t+1
(E(zk) −E(zl))′ (mi −mj) + 1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (E(zk) −E(zl)) = f(t) + g(t) + h(t) + δ(t). = f(t) + g(t) + h(t) + δ(t). Let μ1, μ2 be the d-dimensional vectors containing the elements of the correla-
tion matrices R1 and R2, respectively, which correspond to positions (i, j) ∈wτ2. First, we can calculate δ = δ(t0) −δ(t)
= 1
T 4
t0
i,k=1
T
j,l=t0+1
(E(zi) −E(zj))′ (E(zk) −E(zl))
−1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (E(zk) −E(zl))
= 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2. A.2.1. Proof of Theorem 3.1 Then, Then, P{UT (t) ≥UT (t0)} = P{(f(t) −f(t0)) + (g(t) −g(t0)) + (h(t) −h(t0)) ≥δ}
≤P{f(t) −f(t0) ≥cδ} + P{g(t) −g(t0) ≥cδ}
+ P{h(t) −h(t0) ≥cδ}. (27) P{UT (t) ≥UT (t0)} = P{(f(t) −f(t0)) + (g(t) −g(t0)) + (h(t) −h(t0)) ≥δ}
≤P{f(t) −f(t0) ≥cδ} + P{g(t) −g(t0) ≥cδ} (27) + P{h(t) −h(t0) ≥cδ}. (27) For the first term in (27), as For the first term in (27), as f(t) = 1
T 4
t
i,k=1
T
j,l=t+1
(mi −mj)′ (mk −ml)
= (T −t)2
T 4
t
i,k=1
m′
imk −2 · t(T −t)
T 4
t
i=1
T
l=t+1
mi
′ml + t2
T 4
T
j,l=t+1
m′
jml
= f1(t) −2f2(t) + f3(t), = f1(t) −2f2(t) + f3(t), Efficient change point detection and estimation in HD correlation matrices
97 973 we have f(t) −f(t0) = (f1(t) −f1(t0)) −2(f2(t) −f2(t0)) + (f3(t) −f3(t0)). For
f1(t) −f1(t0), we have f(t) −f(t0) = (f1(t) −f1(t0)) −2(f2(t) −f2(t0)) + (f3(t) −f3(t0)). For
f1(t) −f1(t0), P(|f1(t) −f1(t0)| ≥cδ)
= P
⎛
⎝
(T −t)2
T 4
t
i,k=1
m′
imk −(T −t0)2
T 4
t0
i,k=1
m′
imk
≥cδ
⎞
⎠
= P
⎛
⎝
(T −t)2
T 4
−(T −t0)2
T 4
t0
i,k=1
m′
imk
+(T −t)2
T 4
t
i,k=t0+1
m′
imk + 2 · (T −t)2
T 4
t0
i=1
t
k=t0+1
m′
imk
≥cδ
⎞
⎠
= P(−ν1 + ν2 + 2ν3) P(|f1(t) −f1(t0)| ≥cδ)
= P
⎛
⎝
(T −t)2
T 4
t
i,k=1
m′
imk −(T −t0)2
T 4
t0
i,k=1
m′
imk
≥cδ
⎞
⎠
= P
⎛
⎝
(T −t)2
T 4
−(T −t0)2
T 4
t0
i,k=1
m′
imk In order to verify T· P(|f1(t) −f1(t0) ≥cδ|) →0, it is sufficient to prove
T· P(|ν1| ≥cδ) →0, T· P(|ν2| ≥cδ) →0 and T· P(|ν3| ≥cδ) →0. First, P(|ν1| ≥cδ) P(|ν1| ≥cδ)
= P
⎛
⎝
(T −t0)2
T 4
−(T −t)2
T 4
t0
i,k=1
m′
imk
≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
⎞
⎠
≤P
⎛
⎝
1
d
d
s=1
t0
i,k=1
mismks
≥ct2
0
d ||μ1 −μ2||2
⎞
⎠
≤
d
s=1
P
1
t0
t0
k=1
m2
ks
≥ct0
d ||μ1 −μ2||2
≤dt0P
|mks| ≥c
t0
d ||μ1 −μ2||
. A.2.1. Proof of Theorem 3.1 Since mk = zk −E(zk), and zk is the d-dimensional subvector of vecho(xkx′
k),
then mks = xksxks′ −E(xksxks′), we can obtain Since mk = zk −E(zk), and zk is the d-dimensional subvector of vecho(xkx′
k),
then mks = xksxks′ −E(xksxks′), we can obtain Since mk = zk −E(zk), and zk is the d-dimensional subvector of vecho(xkx′
k),
then mks = xksxks′ −E(xksxks′), we can obtain P
|mks| ≥c
t0
d ||μ1 −μ2||
= P
|xksxks′ −E(xksxks′)| ≥c
t0
d ||μ1 −μ2||
≤c1e−c2
(
t0
d ||μ1−μ2|| ≤c1e−c2√τ2, according to Lemma A.2. Hence, P(|ν1| ≥cδ) ≤dt0c1e−c2√τ2 ≤c1pTe−c2√τ2. (28) (28) Z. Li and J. Gao 974 Second, Second, P(|ν2| ≥cδ)
= P
⎛
⎝
(T −t)2
T 4
t
i,k=t0+1
m′
imk
≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
⎞
⎠
≤P
⎛
⎝
1
d
d
s=1
t
i,k=t0+1
mismks
≥ct2
0(t −t0)
d(T −t) ||μ1 −μ2||2
⎞
⎠
≤dP
1
t −t0
t
k=t0+1
m2
ks
≥
ct2
0
d(T −t)||μ1 −μ2||2
≤d(t −t0)P
|mki| ≥ct0||μ1 −μ2||
d(T −t)
,
where
P
|mki| ≥ct0||μ1 −μ2||
d(T −t)
≤P
|xksxks′ −E(xksxks′)| ≥ct0||μ1 −μ2||
d(T −t)
≤c1e
−c2
t0||μ1−μ2||
d(T −t)
≤c1e−c2
√τ2T . H P(|ν2| ≥cδ)
= P
⎛
⎝
(T −t)2
T 4
t
i,k=t0+1
m′
imk
≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
⎞
⎠
≤P
⎛
⎝
1
d
d
s=1
t
i,k=t0+1
mismks
≥ct2
0(t −t0)
d(T −t) ||μ1 −μ2||2
⎞
⎠
≤dP
1
t −t0
t
k=t0+1
m2
ks
≥
ct2
0
d(T −t)||μ1 −μ2||2
≤d(t −t0)P
|mki| ≥ct0||μ1 −μ2||
d(T −t)
, where re
P
|mki| ≥ct0||μ1 −μ2||
d(T −t)
≤P
|xksxks′ −E(xksxks′)| ≥ct0||μ1 −μ2||
d(T −t)
≤c1e
−c2
t0||μ1−μ2||
d(T −t)
≤c1e−c2
√τ2T . Hence, P(|ν2| ≥cδ) ≤c1d(t −t0)e−c2
√τ2T ≤c1pTe−c2
√τ2T . A.2.1. Proof of Theorem 3.1 (29) (29) Third, P(|ν3| ≥cδ) = P(|ν3| ≥cδ) =
≤P
⎧
⎨
⎩
(T −t)2
T 4
t0
i=1
t
k=t0+1
m′
imk
≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
⎫
⎬
⎭
≤dP
⎧
⎨
⎩
t
k=1
mks
2
−
t0
k=1
mks
2
−
⎛
⎝
t
k=t0+1
mks
⎞
⎠
2
≥ct2
0(t −t0)
d(T −t) ||μ1 −μ2||2
⎫
⎬
⎭
≤d
P
1
t
t
k=1
mks
≥ct0
(t −t0)||μ1 −μ2||
t
d(T −t)
+ P
1
t0
t0
k=1
mks
≥c√t −t0||μ1 −μ2||
d(T −t)
+ dP
⎛
⎝
1
t −t0
t
k=t0+1
mks
≥
ct0||μ1 −μ2||
d(T −t)(t −t0)
⎞
⎠,
h P
1
t
t
k=1
mks
≥ct0
(t −t0)||μ1 −μ2||
t
d(T −t)
≤c1e
−c2t min
ct0
(t−t0)||μ1−μ2||
t
d(T −t)
,
ct2
0(t−t0)||μ1−μ2||2
dt2(T −t)
≤c1e−c2
√τ2T , Efficient change point detection and estimation in HD correlation matrices
975 Efficient change point detection and estimation in HD correlation matrices
97 975 P
1
t0
t0
k=1
mki
≥c
(t −t0)||μ1 −μ2||
d(T −t)
≤c1e
−c2t0min
c
(t−t0)||μ1−μ2||
d(T −t)
, c(t−t0)||μ1−μ2||2
d(T −t)
≤c1e−c2
√τ2T ,
P
1
t −t0
t
k=t0+1
mki
≥
ct0||μ1 −μ2||
d(T −t)(t −t0)
≤c1e
−c2(t−t0)min
ct0||μ1−μ2||
d(T −t)(t−t0) ,
ct2
0||μ1−μ2||2
d(T −t)(t−t0)
≤c1e−c2
√τ2T . P
1
t0
t0
k=1
mki
≥c
(t −t0)||μ1 −μ2||
d(T −t)
≤c1e
−c2t0min
c
(t−t0)||μ1−μ2||
d(T −t)
, c(t−t0)||μ1−μ2||2
d(T −t)
≤c1e−c2
√τ2T , and P
1
t −t0
t
k=t0+1
mki
≥
ct0||μ1 −μ2||
d(T −t)(t −t0) ≤c1e
−c2(t−t0)min
ct0||μ1−μ2||
d(T −t)(t−t0) ,
ct2
0||μ1−μ2||2
d(T −t)(t−t0)
≤c1e−c2
√τ2T . Hence, we have Hence, we have P(|ν3| ≥cδ) ≤c1de−c2
√τ2T ≤c1pe−c2
√τ2T . (30)
By combaning (28), (29) and (30), we obtain P(|ν3| ≥cδ) ≤c1de−c2
√τ2T ≤c1pe−c2
√τ2T . (30) (30) By combaning (28), (29) and (30), we obtain P(|f1(t) −f1(t0)| ≥cδ) ≤c1pTe−c2√τ2 + c1p2Te−c2
√τ2T ≤c1pTe−c2√τ2. A.2.1. Proof of Theorem 3.1 For the other two terms (f2(t) −f2(t0)) and (f3(t) −f3(t0)) in (27), following
similar analysis, we can obtain For the other two terms (f2(t) −f2(t0)) and (f3(t) −f3(t0)) in (27), following
similar analysis, we can obtain For the other two terms (f2(t) −f2(t0)) and (f3(t) −f3(t0)) in (27), following
similar analysis, we can obtain P(|f2(t) −f2(t0)| ≥cδ) ≤c1pTe−c2√τ2,
P(|f3(t) −f3(t0)| ≥cδ) ≤c1pTe−c2√τ2. Therefore, we obtain P{f(t) −f(t0) ≥cδ} ≤c1pTe−c2√τ2. (31) (31) For the second term in (27), we have For the second term in (27), we have For the second term in (27), we have g(t) −g(t0) g(t) −g(t0) g(t) −g(t0)
=
1
T 4
t
i,k=1
T
j,l=t+1
(E(zi) −E(zj))′ (mk −ml)
−1
T 4
t0
i,k=1
T
j,l=t0+1
(E(zi) −E(zj))′ (mk −ml)
=
1
T 4
t0
t (μ1 −μ2)′
⎛
⎝t(T −t)2
t
k=1
mk −t2(T −t)
T
l=t+1
ml
⎞
⎠
−1
T 4 (μ1 −μ2)′
⎛
⎝t0(T −t0)2
t0
k=1
mk −t2
0(T −t0)
T
l=t0+1
ml
⎞
⎠
= t0
T 4 (μ1 −μ2)′
[(2T −t −t0)(t0 −t)]
t0
k=1
mk+
[t0(T −t0) −t(T −t)]
T
k=t0+1
mk + T(T −t)
t
k=t0+1
mk
⎫
⎬
⎭ = −η1 + η2 + η3, Z. Li and J. Gao 976 where where η1 = t0
T 4 [(2T −t −t0)(t −t0)] (μ1 −μ2)′
t0
k=1
mk,
η2 = t0
T 4 [t0(T −t0) −t(T −t)] (μ1 −μ2)′
T
k=t0+1
mk,
t η3 = t0
T 3 (T −t)(μ1 −μ2)′
t
k=t0+1
m 0
To prove T· P(|g(t) −g(t0)| ≥cδ) →0, it is sufficient to prove T· P(|η1| ≥cδ) →
0, T· P(|η2| ≥cδ) →0 and T· P(|η2| ≥cδ) →0. First, 0
To prove T· P(|g(t) −g(t0)| ≥cδ) →0, it is sufficient to prove T· P(|η1| ≥cδ) →
0, T· P(|η2| ≥cδ) →0 and T· P(|η2| ≥cδ) →0. First, 0
To prove T· P(|g(t) −g(t0)| ≥cδ) →0, it is sufficient to prove T· P(|η1| ≥cδ) →
0, T· P(|η2| ≥cδ) →0 and T· P(|η2| ≥cδ) →0. Funding Zhaoyuan Li’s research is partially supported by National Natural Science Foun-
dation of China (No. 11901492) and Shenzhen Science and Technology Program
(ZDSYS 20211021111415025). A.2.1. Proof of Theorem 3.1 First,
P(|η1| ≥cδ) P(|η1| ≥cδ) = P
t0
T 4 [(2T −t −t0)(t −t0)] (μ1 −μ2)′
t0
k=1
mk ≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
= P
1
d
d
s=1
t0
k=1
mks
≥ct0||μ1 −μ2||
d
≤dP
1
t0
t0
k=1
mks
≥c||μ1 −μ2||
d
≤c1de
−c2t0 min
c||μ1−μ2||
d
, c||μ1−μ2||2
d2
≤c1p2e−c2 min(√τ2T ,τ2). e
−c2t0 min
c||μ1−μ2||
d
, c||μ1−μ2||2
d2
≤c1p2e−c2 min(√τ2T ,τ2). Second, Second, P(|η2| ≥cδ) P(|η2| ≥cδ)
= P
⎧
⎨
⎩
t0
T 4 [t0(T −t0) −t(T −t)] (μ1 −μ2)′
T
k=t0+1
mk
≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
≤P
⎛
⎝1
d
d
s=1
T
k=t0+1
mks
≥ct0(T −t0)||μ1 −μ2||
d(t0 + t −T)
⎞
⎠
≤dP
⎛
⎝
1
T −t0
T
k=t0+1
mks
≥ct0||μ1 −μ2||
d(t0 + t −T)
⎞
⎠
≤c1de
−c2(T −t0) min
ct0||μ1−μ2||
d(t0+t−T )
, ct2
0||μ1−μ2||2
d2(t0+t−T )2
≤c1p2e−c2 min(√τ2T ,τ2). ≤c1p2e−c2 min(√τ2T ,τ2). Last, ,
P(|η3| ≥cδ) = P
⎧
⎨
⎩
t0
T 3 (T −t)(μ1 −μ2)′
t
k=t0+1
mk ≥c 1
T 4 t2
0(t −t0)(2T −t0 −t)||μ1 −μ2||2
⎫
⎬
⎭
≤P
⎛
⎝
1
d
d
s=1
t
k=t0+1
mks
≥ct0(t −t0)||μ1 −μ2||
dT
⎞
⎠
≤dP
⎛
⎝
1
t −t0
t
k=t0+1
mks
≥ct0||μ1 −μ2||
dT
⎞
⎠
≤c1p2e−c2 min(√τ2T ,τ2). Efficient change point detection and estimation in HD correlation matrices
977 Therefore, we obtain P(|g(t) −g(t0)| ≥cδ) ≤c1p2e−c2 min(√τ2T ,τ2). (32) (32) For the third term in (27), similarly, we can obtain P(|h(t) −h(t0)| ≥cδ) ≤c1p2e−c2 min(√τ2T ,τ2). (33) (33) By combining (31), (32), and (33), we can prove that P(UT (t) ≥UT (t0)) ≤
c1p2e−c2√τ2 and Equation (26) holds under Assumption 3. We complete the
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Fragmentation of natural cover and the state of permanent preserved areas of drainage channels at Mineiros, Goiás state, Brazil
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Ambiência
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cc-by
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1
Ecóloga e Analista Ambiental; Mestranda no Programa de Pós-Graduação em Ciências Ambientais na
Universidade Federal de Goiás, UFG; Endereço: Campus Samambaia, CEP: 74690-900, Goiânia, Goiás,
Brasil; E-mail: tassiaponciano@gmail.com (*) Autora para correspondência.
2
Drª; Geógrafa; Professora Adjunta do Instituto de Estudos Sócio Ambientais e do Programa de Pós
Graduação em Ciências Ambientais da Universidade Federal de Goiás, UFG; Endereço: Campus
Samambaia, CEP 74690-900, Goiânia, Goiás, Brasil; E-mail: karlamsfaria@gmail.com
3
MSc.; Bióloga; Professora Adjunta da Faculdade de Engenharia Ambiental da Universidade de Rio Verde, UNIRV;
Endereço: Fazenda Fontes do Saber, CEP: 75901-970, Rio Verde, Goiás, Brasil; E-mail: mns.mariana@gmail.com
4
Drª; Geógrafa; Professora Titular do Instituto de Estudos Sócio Ambientais do Programa de Pós Graduação
em Ciências Ambientais e de Programa de Pós-Graduação em Geografia da Universidade Federal de Goiás,
UFG; Endereço: Campus Samambaia, CEP 74690-900, Goiânia, Goiás, Brasil; E-mail: selma@ufg.com Fragmentation of natural cover and the state of permanent preserved
areas of drainage channels at Mineiros, Goiás state, Brazil Tássia Andrielle Ponciano1(*)
Karla Maria Faria2
Mariana Nascimento Siqueira3
Selma Simões de Castro4 Tássia Andrielle Ponciano1(*)
Karla Maria Faria2
Mariana Nascimento Siqueira3
Selma Simões de Castro4 Fragmentação da cobertura vegetal e estado das Áreas
de Preservação Permanente de canais de drenagem no
Município de Mineiros, Estado de Goiás Fragmentation of natural cover and the state of permanent preserved
areas of drainage channels at Mineiros, Goiás state, Brazil Ambiência Guarapuava (PR) v.11 n.3 p. 545 - 561 Set./Dez. 2015 ISSN 1808 - 0251
Recebido para publicação em 16/03/2015 e aceito em 08/06/2015
DOI:10.5935/ambiencia.2015.03.03 1
Ecóloga e Analista Ambiental; Mestranda no Programa de Pós-Graduação em Ciências Ambientais na
Universidade Federal de Goiás, UFG; Endereço: Campus Samambaia, CEP: 74690-900, Goiânia, Goiás,
Brasil; E-mail: tassiaponciano@gmail.com (*) Autora para correspondência. Resumo A expressiva ocupação do Cerrado nas últimas quatro décadas trouxe consigo
muitos impactos para o bioma, sendo a fragmentação da paisagem indicada
como uma das principais responsáveis pela sua perda de biodiversidade. Diversas
pesquisas na região da alta bacia do rio Araguaia, adotando imagens de satélite
de média resolução, já foram realizadas e constataram impactos distintos
como erosão, assoreamento e fragmentação, inclusive das áreas de preservação
permanente. Tais impactos foram associados à implementação de políticas
públicas desenvolvimentistas, sobretudo o POLOCENTRO, que, dentre outras
ações, incorporou o Cerrado ao sistema produtivo nacional, a partir da década de
1970. Nesse contexto, o objetivo deste trabalho foi o de identificar e caracterizar
a tipologia da vegetação remanescente de Cerrado, no setor sul da alta bacia do
rio Araguaia, no recorte municipal de Mineiros (GO), enfatizando o estado de
fragmentação e de conservação das Áreas de Preservação Permanente (APP’s)
associadas aos canais de drenagem, com imagens de alta resolução. Os resultados
apontaram que a área já apresenta, como atividade predominante, a monocultura
de cana de açúcar. Quanto aos remanescentes que se encontravam em desacordo Ambiência Guarapuava (PR) v.11 n.3 p. 545 - 561 Set./Dez. 2015 ISSN 1808 - 0251
Recebido para publicação em 16/03/2015 e aceito em 08/06/2015
DOI:10.5935/ambiencia.2015.03.03 com o antigo código florestal, estes já se encontram em conformidade com a atual
legislação (Lei 12.651/12). Os mapeamentos e análises, em nível de detalhe, não
confirmaram o padrão de elevada fragmentação diagnosticado anteriormente em
média resolução. No entanto, as APP’s estão ameaçadas pelo uso intensivo de seu
entorno, necessitando, portanto, de mecanismos de conservação. Palavras-chave: áreas de preservação permanente; alta bacia do rio Araguaia;
fragmentação da paisagem; RapidEye. Abstract The significant occupation of the Cerrado in the last four decades has brought many
impacts to the biome, and the landscape fragmentation is seen as one of the main
contributors to the loss of biodiversity. Several studies in the Upper River Araguaia
Basin, adopting medium resolution satellite images have been performed and found
different impacts such as erosion, siltation and fragmentation, including the areas
of permanent preservation. Such impacts were associated with the implementation
of developmental public policies, especially Polocentro, which, among other actions,
incorporated the Cerrado to the national productive system from the decade of
1970 onwards. In this context, the aim of this study was to identify and characterize
the typology of remnant Cerrado vegetation in the southern portion of the Upper
Araguaia River Basin in the municipal Mineiros area (GO), emphasizing the
state of fragmentation and conservation of Permanent Preserved Areas (PPAs),
associated with drainage channels, with high resolution images. The results showed
that the area already holds the sugarcane monoculture as its main activity. However,
regarding the remnant that was at odds with the old Forest Code, it is currently in
accordance with current legislation (Law 12.651/12). The mapping and analysis at
detail did not corroborate the pattern of high fragmentation diagnosed previously
in medium resolution. Nevertheless, the PPAs are threatened by intensive use of
their surroundings, thus requiring conservation mechanisms. Key words: Permanent Preserved Areas; Upper Araguaia River Basin;
landscape fragmentation; RapidEye. Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 20 Introdução (áreas prioritárias para conservação da
biodiversidade) mundiais em função
do alto grau de endemismo de plantas
(pelo menos 4.400 espécies endêmicas,
representando 1,5% de todas as espécies
da flora), de endemismo de vertebrados
(117 espécies endêmicas - representando O Cerrado, mesmo tendo sido
reduzido a cerca de 50% de sua área por
processos intensos de desmatamento, ainda
é o segundo maior bioma do Brasil em
área. É considerado como um dos hotspots 546 Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015 em áreas antrópicas (SANO et al., 2008),
entretanto para Machado et al. (2004), a
taxa de conversão foi de aproximadamente
55%. Essa expressiva ocupação do Cerrado
trouxe consigo vários impactos para o
bioma, sendo a fragmentação e perda da
vegetação uma das principais responsáveis
pelo depauperamento de biodiversidade
(TABARELLI; GASCON, 2005) em
condições locais e regionais, que a médio
e longo prazo promovem até a extinção
de espécies, além do comprometimento
das estruturas genéticas e facilitarem o
desenvolvimento de espécies invasoras e
oportunistas (SAUNDERS, et al., 1991;
PIVELLO, 2005). Tal processo foi mais
acelerado em áreas que integravam os
polos de atuação das referidas políticas
públicas, principalmente nas doze áreas do
POLOCENTRO. 0,4% das espécies dos vertebrados global)
e altas taxas de desmatamento (MYERS,
2000; COLLI et al., 2003). Sem despertar maiores interesses
durante séculos, a ocupação intensiva do
Cerrado iniciou-se, com maior empenho, ao
fim da década de 1960, por meio da expansão
da fronteira agrícola promovida por políticas
públicas federais de desenvolvimento
baseadas em um modelo de exploração
agropecuário denominado de moderno. O Programa de Desenvolvimento das
Áreas de Cerrado (POLOCENTRO)
e o Programa de Cooperação Nipo -
Brasileira de Desenvolvimento dos
Cerrados (PRODECER) se destacaram
por viabilizarem a ocupação rápida das
terras, favorecerem a concentração fundiária
e estimularem a modernização do campo,
conforme vários autores afirmaram
(DUARTE; THEODORO, 2002;
MACHADO, et al., 2004). Nesse contexto, destaca-se o munícipio
de Mineiros, situado na porção sudeste do
estado de Goiás e do Bioma Cerrado, onde
se concentram vários estudos relevantes
por abrigar, em seu território, as principais
nascentes do rio Araguaia, e que é indicada
como uma representante das transformações
históricas socioeconômicas e ambientais
sofridas pelo Cerrado (FRANCO, 2003;
CASTRO, 2005). Os pólos de atuação desses programas
foram instalados em regiões que já dispunham
de certa infraestrutura para o escoamento
da produção. Introdução Assim, tais programas foram
apontados como responsáveis pela rápida
transformação da base técnica do sistema de
produção agropecuária e, consequentemente,
pelas mudanças notáveis da paisagem
(DUARTE; THEODORO, 2002). Adicionalmente, esses autores ressaltam
que tais transformações foram provenientes
da substituição das áreas de cobertura
vegetal nativa por pastagens destinadas à
bovinocultura de corte e, também, de áreas
extensas destinadas ao cultivo de grãos, em
especial da soja, esta nos melhores solos. Barbalho (2002) e Castro et al. (2004),
entre outros, já constataram a área da alta
bacia do rio Araguaia, no seu setor sul, cujo
uso da terra dominante era pastagem, soja
e algodão, índices e focos expressivos de
erosões lineares, assoreamento e fenômeno de
arenização. Faria et al. (2012), Carneiro (2012)
e Siqueira (2012), em sub-bacias vizinhas
também identificaram elevados índices de
degradação e um estado de conservação da
paisagem comprometido, fornecendo suporte
para análise da complexidade das relações Segundo o Projeto de Conservação
e Utilização Sustentável da Diversidade
Biológica Brasileira (MMA, 2004), cerca de
39,5% do Cerrado já teria sido convertido 547 PONCIANO, T. A. et al. geoecológicas e da estrutura da paisagem
em séries históricas. Faria e Castro (2007)
e Cabacinha e Castro (2009), igualmente já
haviam constatado que durante o processo
de ocupação da bacia do Araguaia e também
do município de Mineiros, áreas inaptas às
práticas agrícolas foram ocupadas (como
as escarpadas) e que os remanescentes
restringiam-se a áreas de Reserva Legal,
Unidades de Conservação e Áreas de
Preservação Permanente (APP’s), mas que
em algumas regiões, nem mesmo as APP’s,
sobretudo as de canais de drenagem, foram
respeitadas. Tais estudos, no entanto, foram
realizados em imagens de média resolução,
como as Landsat TM5. espaciais e temporais da Ecologia de
Paisagem, gerando um produto mais conciso
para o planejamento ambiental. O setor sul da alta bacia do rio
Araguaia, onde se insere Mineiros, no
estado de Goiás, é uma região de extrema
importância ambiental pelo número de
nascentes existentes na área, que auxiliam a
manutenção e a própria recarga hídrica para
abastecer o rio Araguaia (RODRIGUES,
2010). Estudos recentes desenvolvidos no
Laboratório de Geomorfologia, Pedologia
e Geografia Física-Universidade Federal
de Goiás (LABOGEF) indicam alteração
recente na matriz agrícola da região pós-
modernização agrícola citada, com a
introdução de cana de açúcar e eucalipto. Introdução As APP’s de canais de drenagem são
áreas importantes para proteção e equilíbrio
hídrico das bacias hidrográficas e sistema am
biental; é uma região sensível, que apresenta
grande incidência de nascentes, sendo sua
proteção e usos permitidos regulamentados
por lei. O conceito de APP está diretamente
relacionado à proteção das áreas de maior
risco de degradação, onde o manejo incorreto
pode ocasionar erosão, deslizamentos/mo
vimentos de massa, e, consequentemente, o
assoreamento dos rios e diminuição da oferta
e da qualidade das águas. De encontro às mudanças de uso do
solo na região, através da análise espacial
da paisagem, o presente estudo tem como
objetivo identificar e caracterizar a tipologia
da vegetação remanescente de Cerrado no
setor sul da alta bacia do rio Araguaia, no
município de Mineiros (GO). Assim, tal
análise teve ênfase no grau de fragmentação
e de conservação das Áreas de Preservação
Permanente (APP’s) associadas aos canais de
drenagem, em escala de detalhe e com auxílio
de imagens de alta resolução. A fragmentação e a descaracterização
das APP´s por interferências antrópicas
são indicadas por Cerqueira et al. (2005)
e Lang e Blaschke, (2009), como fato
preocupante, demandando estudos em escalas
adequadas, por se tratar de um importante
elemento de conexão na paisagem, ao
desenvolverem função de corredores, em
matrizes antropizadas. Esses estudos foram
favorecidos com o advento e evolução das
geotecnologias cujas ferramentas associadas
aos Sistemas de Informações Geográficas
(SIG’s) são manipuladas nas investigações Área de Pesquisa A alta bacia do rio Araguaia localiza-
se entre as coordenadas geográficas 17º 30’
00” - 18º 00’ 00” S e 53º 00’ 00” – 53º 30’ 00”
W, abrangendo parte dos Estados de Goiás
(municípios de Mineiros e Santa Rita), Mato
Grosso (Alto Araguaia e Taquari) e Mato
Grosso do Sul (município de Costa Rica), com
uma área de aproximadamente 1.500 km². A 548 Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015 presente avaliação restringe-se ao estudo
apenas ao lado goiano, correspondente
ao município de Mineiros, cuja área de
drenagem é de 283,39km2. (Figura 1). Quaternários, hoje correlacionados aos
Latossolos Vermelhos argilosos, depositados
sobre arenitos do Grupo Bauru (Cretáceo
superior) e da Formação Botucatu (Jura-
Triássico), intercalada por basaltos da
Formação Serra Geral (Eo-Cretáceo); Aspectos Geoambientais 2) Zona Escarpada desdobrada em
níveis escalonados, com declives médios
de 25% a 45%, que corresponde aos limites
das Superfície Regional de Aplainamento
(SRA) supracitadas, onde se desenvolvem
Cambissolos, Neossolos Litólicos e
afloramentos de rocha; Resumidamente, com base no exposto
por Latrubesse e Carvalho (2006), a área
caracteriza-se por apresentar: 1) Superfícies tabulares e aplainadas
de baixos declives (em geral menos de 3%),
denominadas de Serra de Caiapó (que
abriga a nascente do rio Araguaia) e Morro
Vermelho, principalmente, sustentadas
por sedimentos inconsolidados Terço- l
3) Relevos convexizados que
se estendem por toda zona rebaixada,
constituindo interflúvios alongados na Figura 1 – Mapa de localização da área de pesquisa Figura 1 – Mapa de localização da área de pesquisa Figura 1 – Mapa de localização da área de pesquisa Figura 1 – Mapa de localização da área de pesquisa Figura 1 – Mapa de localização da área de pesquisa
Fonte: Base de dados do IBGE (s. d.). Nota: Banco de Dados Pamira/LABOEGF (2013). Fonte: Base de dados do IBGE (s. d.). Nota: Banco de Dados Pamira/LABOEGF (2013). 549 PONCIANO, T. A. et al. orbitais do sistema RapidEye (período de
seca de 2012), segmentadas e classificadas
(Classificação não supervisionada) no
softwares SPRING (CAMARA et al.,
1996) e, posteriormente, no ArcMap para a
classificação das imagens com base na chave
de classificação construída especialmente
para a imagem de alta resolução. forma de colinas amplas com vertentes
longas, dispersoras dos fluxos hídricos
com declives intermediários, variando
entre 3 a 20%, correspondentes às SRA
III e IV, correlacionadas aos Neossolos
Quartzarênicos órticos distróficos a
álicos (EMBRAPA, 2006), derivados da
Formação Botucatu; 4) Relevos concavizada nos segmentos
inferiores mais declivosos das vertentes
nos topos e terços médios dos amplos
interflúvios, onde dominam os Neossolos
Quartzarênicos hidromórficos e nos fundos
de vale embaciados e várzeas restritas, os
Gleissolos distróficos (CASTRO, 2005),
respectivamente. Em função da realização de trabalhos
de campo, no nível de detalhe das imagens
inspeção visual e definição de chave de
classificação que inclui as características
tonais, texturais, geométricas de classes de
usos (Quadro 1), foram identificadas as classes
de pastagem, agricultura, silvicultura, cana
de açúcar, solo exposto (áreas sem cobertura
vegetal) e, dentre os remanescentes, as
fitofisionomias do Bioma Cerrado, utilizando-
se a proposta de Ribeiro e Walter (1998; 2008),
sendo elas: Mata Ciliar/Galeria (vegetação de
porte arbóreo, identificada nas proximidades Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015 Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015
550 Procedimentos metodológicos Os procedimentos metodológicos
envolveram a elaboração de mapas de uso
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Quadro 1 - Chave de interpretação
Classes de uso
Características
Textura
Tonalidade
Porte
Aspectos Associados
Mata Ciliar/
Galeria
Homogêneo
Verde escuro
Alto
Fundos de Vale;
cursos d’água
Cerrado
Sentido
Restrito
Homogêneo
Verde médio
Médio/
Alto
Formas irregulares
Campo
Limpo Úmido
Lisa,
homogênea
Arroxeado/ preto
Baixo
Fundos de Vale;
cursos d’água
Agricultura
Fina e
aveludada
Rosa/ verde claro
Rasteiro
Formas geométricas;
carreadores/pivôs. Pastagens
Granular
Clara com
algumas variações
Baixo
Formas geométricas,
trilhas/árvores
isoladas. Cana de
açúcar
Lisa
Verde claro/
marrom
Baixo
Formas geométricas/
talhões
Silvicultura
Grossa
Arroxeado/preto
Alto
Formas geométricas/
alinhamentos
Corpos d’
água
Lisa
Cinza escuro a
negro
-
Áreas rebaixadas e
fundos de vale
Solo exposto
Lisa
Rosa
Baixo
Formas geométricas
Fonte: Ponciano et al. (2015). Nota: Org. Faria (2013). 550 paisagem, sendo o Fragstat (MCGARIGAL;
MARKS, 1995), o mais utilizado para análise
de fragmentação florestal. Desse modo, a
avaliação da fragmentação foi realizada com
métricas que analisam o grau de ruptura de uma
unidade na paisagem (METZGER, 2003),
selecionando-se as métricas disponibilizadas
pelo software FRAGSTATS como: CA (Área
Total da Classe), CPLAND (Porcentagem
de Área Central na Paisagem), NP (Número
de Fragmentos), NP/Área (Número de
Fragmentos/Área), TE (Total de Bordas) e TCA
(Total de Áreas Centrais) (MACGARIAL e
MARKS, 1995), que foram calculadas para o
Município e para as áreas de APP. dos canais de drenagem), Cerrado Sentido
Restrito (vegetação de porte arbóreo-arbustiva
representativa da Formação Savânica), Campo
Limpo Úmido (vegetação campestre, situada
em áreas de fundo de vale com ocorrência de
afloramentos de lençol freático) e Cerrado em
Regeneração/Campo Sujo (vegetação de porte
campestre-graminoso com espécies isoladas). As atividades de campo realizadas na
área foram fundamentais para a validação
dessas classes de usos e remanescentes. O
recente processo de conversão na matriz de
uso econômico da região conduziu durante
o processo de classificação da imagem na
especificação da classe de cana de açúcar a
fim de avaliar a distribuição e situação dos
remanescentes nas áreas vizinhas a essa classe. PONCIANO, T. A. et al. Resultados e Discussão Para a delimitação das Áreas de
Preservação Permanente (APP’s) utilizou-
se a definição legal e jurídica para estas
áreas, contida na Lei No 12.651/2012, Art. 3o inciso II: O mapa de uso do solo da área de
pesquisa com imagem alta resolução, pós-
validado em campo, possibilitou a identificação
das fisionomias do Bioma Cerrado (e.g. Mata
Ciliar e de Galeria; Cerrado sentido restrito;
Campo Limpo úmido), porém permitiu
constatar o predomínio do cultivo da cana de
açúcar na região cobrindo mais de 53% da área
total (Figura 2). “área protegida, coberta ou não por
vegetação nativa, com função ambiental de
preservar os recursos hídricos, a paisagem,
a estabilidade geológica e a biodiversidade,
facilitar o fluxo gênico de fauna e flora,
proteger o solo e assegurar o bem-estar
das populações humanas.” A tabela 1 apresenta as categorias de
uso do solo, indicando que os usos antrópicos
representam, aproximadamente, 58,7% da
área total (dos quais 53,3% cana-de-açúcar),
enquanto os remanescentes da vegetação
natural (fisionomias de Mata Ciliar/Galeria,
Campo Limpo Úmido Cerrado Sentido
Restrito e Campo Sujo - Cerrado em
Regeneração) correspondem juntos, a 41,2
% da área (Tabela 1). A delimitação espacial das faixas
marginais de proteção varia conforme a
largura do canal de drenagem (de 30 a
500 metros) e ainda conforme o tamanho
da propriedade - Módulo fiscal (de 5 a
15 metros), quando atividades antrópicas
presentes na APP tenham se instalado
anteriormente a 2008 (exceção permitida
pela atual legislação ambiental). A cana de açúcar cobre 31.475,56 ha
(53,3% da área), e se estende pelos Latossolos
Vermelhos argilosos e inclusive os Neossolos
Quartzarênicos; as áreas de agricultura
correspondem a 2.160,36 ha (3,7% da Em relação à análise da paisagem, há
uma variedade de softwares desenvolvidos, a fim
de quantificar as características da estrutura da 551 PONCIANO, T. A. et al. Figura 2 - Mapa de Uso e Cobertura da Terra Fonte: Faria (2012). Nota: Interpretação de imagens Rapideye, 2012 e validação em campo. Nota: Interpretação de imagens Rapideye, 2012 e validação em campo. 1 - Métricas da paisagem: Área total da classe (CA) e Porcentagem na paisagem
(CPLAND), da área total Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. Resultados e Discussão 2015
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Tabela 1 Métricas da paisagem: Área total da classe (CA) e Porcentagem na paisagem
(CPLAND), da área total
Classe de Uso
CA
CPLAND
Vegetação
Remanescente
Mata Ciliar
3.927,96
6,7
Campo Limpo úmido
3.742,73
6,3
Cerrado sentido restrito
16.343,20
27,7
Cerrado em Regeneração
304,98
0,5
Subtotal 1
24.318,87
41,2
Áreas Antrópicas
Solo Exposto
357,59
0,6
Agricultura
2.160,36
3,7
Cana de Açúcar
31.475,56
53,3
Pastagem
449,15
0,8
Silvicultura
218,01
0,4
Subtotal 2
34.660,67
58,8
Outro Uso
Água
34,20
0,1
Total (Subtotal 1 + Subtotal 2 + Água)
59.013,74
100
Fonte: Mapa de uso da terra. Organização: Faria (2013). Nota: Unidade métrica: CA: Hectare - CPLAND: %. 552 área), sendo que as maiores extensões estão
localizadas principalmente nos topos das
chapadas na porção sul, onde ainda se
desenvolvem cultivos de algodão e soja. As áreas de pastagem representam apenas
449,15 ha (0,8% da área), sendo que as áreas
mais contínuas são identificadas na porção
norte, próximas ao município de Santa Rita
do Araguaia (no limite da área de pesquisa),
onde também predominam os Neossolos
Quartzarênicos. Outros usos foram ainda
identificados, como a Silvicultura (Eucalipto)
na porção sul (próximo à região das nascentes)
sob o domínio dos Neossolos Quartzarênicos,
ocupando 218,01 ha (0,4% da área); e
também áreas de solo exposto (geralmente
áreas degradadas pelas atividades agrícolas
que se encontram abandonadas ou ainda as
áreas erodidas) correspondentes a 357,59 ha
(0,6 da área). A classe água corresponde a
34,20 ha (0,1) e representam barramentos/
reservatórios artificiais e ainda cursos d`água
em seu canal natural. Ao avaliar a cobertura vegetal original,
destaca-se a participação da classe da
fitofisionomia de Cerrado Sentido Restrito,
ainda com 27,7% da área, que é identificada
em grandes manchas localizadas em maior
parte na região norte da área e nas APP’s
do Ribeirão Queixada e encostas das
escarpas e na porção noroeste, próximo
ao rio Araguaia. Associada aos canais
de drenagem e às áreas de preservação
permanente (APP’s) identificaram-se as
fisionomias de Mata Ciliar e Campo Limpo
Úmido que as cobrem, respectivamente,
3.927,96 ha (6,7 % da área) e 3.742,73 ha
(6,3% da área) apresentando forma linear e
estreita. A classe Cerrado em Regeneração
corresponde a 304,98 ha (0,5 % da área) e
foi identificada em pequenas manchas na
porção norte da área. Resultados e Discussão Em campo, constatou-
se que tal classe corresponde a locais onde
houve o desmatamento do Cerrado, mas não
houve incorporação dos terrenos ao processo
produtivo (seja por pasto ou agricultura),
reestabelecendo a revegetação natural. Para Spera et al. (1999), quando os
Neossolos Quartzarênicos são ocupados
por lavouras perenes (ex. Cana-de-açúcar),
requerem manejo adequado, cuidados
intensivos em termos de adubação e
correção, principalmente em relação aos
nutrientes facilmente lixiviáveis como
nitrogênio, potássio e enxofre, além de
medidas visando ao controle de erosão
hídrica e eólica e medidas de economia
de água. Tais solos são classificados por
Lepsch (1991) como terras que tem riscos
ou limitações muito severas para culturas
anuais e que também não são adequados
para cultivos intensivos e contínuos, devido
à baixa capacidade de retenção de água
aliada a problemas de fertilidade. Portanto,
não se trata da aptidão agrícola elevada a
variável explicativa desses cultivos. A análise comparada das métricas
Total de bordas (TE) e área central total
(TCA) permite constatar que a métrica
TCA correlaciona-se às métricas de TE e
de área das classes (CA), sendo observado
que o aumento da área das classes (CA)
leva, consequentemente, ao aumento de
total de bordas (TE) e de área central total
(TCA). A avaliação dessas métricas indica
que os maiores índices correspondem à
classe de Cerrado Sentido Restrito, pois
essa fisionomia apresenta também a maior
área em relação aos demais remanescentes
da vegetação nativa (Tabela 2). Aquino e Miranda (2008) relatam
que as distintas fitofisionomias do Cerrado
respondem de maneiras diferentes ao
efeito de borda, e Harper et al. (2005) 553 PONCIANO, T. A. et al. Tabela 2 - Métricas da paisagem no
Município de Mineiros (GO):
Total de Bordas (TE) e Total
de Área Central (TCA)
Vegetação
Remanescente
TE (m)
TCA (ha)
Mata Ciliar
1.294.580,0
3.476,6
Campo Limpo
úmido
1.474.140,0
2.953,4
Cerrado sentido
restrito
2.930.620,0
13.928,6
Campo Sujo
191.740,0
159,8
Total
5.891.080,0
20.518,4
Fonte: Mapa de uso da terra (2013). Nota: Organização: Siqueira (2013). et al., 2005; LAURANCE et al., 2007;
CAYUELA et al., 2009). É notável a concentração de fragmentos
de remanescentes com área menor que
10 hectares para todas as fisionomias. O
destaque também é para a classe de Cerrado
Sentido Restrito, com maior presença
tanto de fragmentos com área menor que
10 hectares, mas também por apresentar
maior concentração de fragmentos com área
superior a 50 hectares (Tabela 3). Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015
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554 Resultados e Discussão No entanto, o número de fragmentos não
deve ser analisado isoladamente, pois quando
se avalia a área que os fragmentos inferiores a
10 hectares representam, verifica-se que são
consideravelmente menores do que as áreas dos
fragmentos maiores que 10 hectares (Tabela 4). complementam, expondo que a vegetação
com dossel mais aberto e diversificado
apresentam menos influências de borda, e
que esses efeitos são mais difíceis de detectar
e com menor importância ecológica. g
q
Cullen et al. (2005), considerando
mamíferos de grande porte, constatou que,
para que um fragmento seja considerado
sustentável, é necessário que o mesmo
apresente uma área superior a 100 hectares,
a fim de manter o efeito positivo da
relação existente entre efeito de tamanho
do fragmento e efeito de borda e interior. A área estudada em Mineiros apresenta
fragmentos que comportam desde grupos
de organismos menos exigentes até os
animais de topo de cadeia. A avaliação dos índices selecionados
(CA, PLAND, TE e TCA) indica que
essa área apresenta uma proteção parcial de
sua vegetação nuclear, o que, para fins de
conservação, é de suma importância, pois
quanto maior a área central menor o efeito
de borda nas manchas de remanescentes,
especialmente para as formações florestais,
que sofrem mais esses efeitos (HARPER Tabela 3 - Métricas da paisagem: Número de fragmentos por classe de uso e classe de tamanho Tabela 3 - Métricas da paisagem: Número de fragmentos por classe de uso e classe de tamanh p
g
g
p
Número de fragmentos por classe de uso
Tamanho-Área
(ha)
Cerrado
Sentido
Restrito
Campo em Regeneração Mata Ciliar/
Galeria
Campo
Limpo
Úmido
< 1ha
4.798
290
919
1.751
1 - 10 ha
246
28
88
117
10.01 - 50 ha
40
3
39
30
50.01 - 100 ha
12
1
7
7
> 100 ha
18
0
9
8
Total
5114
322
1062
1913
Fonte: Mapa de uso da terra. Organização: Siqueira (2013). Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. Resultados e Discussão 2015
554 554 Tabela 4 - Métricas da paisagem: Área dos fragmentos (ha) por classe de uso e classe de tamanho p
g
g
p
Área dos fragmentos (ha) por classe de uso
Tamanho-Área
(ha)
Cerrado Sentido
Restrito
Campo em
Regeneração
Mata Ciliar/
Galeria
Campo Limpo
Úmido
< 1ha
505
31,73
124,86
209,11
1 - 10 ha
737,42
98,97
226,51
357,57
10.01 - 50 ha
752,05
76,89
837,69
754,94
50.01 - 100 ha
749,56
97,39
497,53
470,27
> 100 ha
13599,57
0
2241,37
1950,84
Total
16.343,20
304,98
3927,96
3742,73
Fonte: Mapa de uso da terra. Organização: Siqueira (2013). Área dos fragmentos (ha) por classe de uso Fonte: Mapa de uso da terra. Organização: Siqueira (2013). A análise da métrica de isolamento
pela Distância Euclidiana média do vizinho
mais próximo (ENN_MN), que indica
ha capacidade que a paisagem apresenta
em facilitar fluxos biológicos, aponta que
a dentre as fitofisionomias remanescentes,
na área de estudo, a distância média entre
as manchas é de 50 metros, considerada
baixa. Entretanto, as distâncias muito baixas,
identificadas entre fragmentos da mesma
classe, especialmente para os remanescentes
de vegetação nativa, somado à análise do
mapa de uso e remanescentes da região,
indicam uma conectividade muito elevada
das classes de vegetação como um todo e que,
apesar de muitos fragmentos de vegetação
ocorrerem na área, eles estão conectados
acompanhando as redes de drenagem nas
áreas de preservação permanente (APP’s). Essa conectividade corrobora a qualidade
ambiental da área estudada e exclui uma das
principais causas da perda de biodiversidade
decorrentes da fragmentação que, de acordo
com diversos autores (e.g. FARIHG, 2003;
PIRES et al., 2006; SCARIOT et al.,
2005; AQUINO; MIRANDA, 2008)
trata-se do isolamento dos remanescentes
de vegetação. Adicionalmente, entende-
se que a conectividade é o oposto da
fragmentação, determinando o grau que uma
paisagem permite ou restringe o movimento de organismos ao longo dos fragmentos
(FORERO-MEDINA; VIEIRA, 2007). Resultados e Discussão Já em relação à avaliação das APP’s,
estas não são compostas apenas por
fisionomias pertencentes às Formações
Florestais, como as Matas Ciliares/Galeria,
mas também por fisionomias das Formações
Savânicas e Campestres (Tabela 5), o que se
considerou como uma fragmentação natural Tabela 5 - Métricas da Paisagem para as
APP’s: Área total da classe (CA)
e porcentagem da área central na
paisagem (CPLAND) paisagem (CPLAND)
Classes de uso
Geral
CA
CPLAND
Mata Ciliar/Galeria
1071,5
51,1
Campo Limpo
Úmido
522,9
24,9
Cerrado Sentido
Restrito
331,5
15,8
Campo Sujo/
Regeneração
13,9
0,7
Pastagem
3,6
0,2
Solo Exposto
16,3
0,8
Agricultura
5,3
0,3
Água
11,9
0,6
Total
2.097,00
100
Fonte: Mapa de uso da terra. Organização: Ponciano (2013). Nota: Unidade métrica: CA: Hectare/ CPLAND: %. 555 PONCIANO, T. A. et al. de fiscalização constante, diferentemente de
outros setores, como soja e pastagem. de cada fitofisionomia pela presença de outra,
o que é característica do Bioma Cerrado. Esse mosaico heterogêneo de
fisionomias é favorável à biodiversidade
(DEVELEY; PONGILUPPI, 2010). Contudo, em menor frequência, é
possível verificar atividades agropecuárias
fragmentando a vegetação que acompanha as
redes de drenagem. A avaliação da métrica
NP, para as áreas de APP, indicou 36.709
fragmentos, sendo que a maior densidade
de fragmentos por hectare ocorreu para a
classe de Mata Ciliar/Galeria, com 26.268
fragmentos, sendo seguido pelas classes de
Campo Limpo Úmido, Cerrado Sentido
Restrito e Cerrado em Regeneração. Deve-se ressaltar que a área, apesar
de representativa quanto à fragmentação
do Cerrado, quanto às APP’s de drenagem,
principalmente as Matas Ciliares/ Galerias,
não se apresentam com um número
significativo de uso antrópico. A continuidade
dessa fisionomia está sendo interrompida por
outras classes de remanescentes, resultantes
de fragmentação natural e da composição
natural do Cerrado, através de mosaicos de
fitofisionomias. Rodrigues (2010), ao analisar
a área, indicou que a pastagem dominava
as culturas anuais, sobretudo soja. Onde na
presente pesquisa se constatou predomínio de
cana-de-açúcar e essa substituição necessita
de estudos mais aprofundados em termos
de impactos diretos e indiretos da expansão
canavieira, assim como seus efeitos na
vegetação remanescente. Destaca-se, no entanto, que a legislação
atual permite a manutenção dos usos antrópicos
identificados nas áreas de APP, se já estavam
presentes em 28 de julho de 2008. Resultados e Discussão Por outro
lado, embora estejam em conformidade com
a legislação atual, os fragmentos encontram-
se envolvidos por entorno antrópico, o que
significa riscos à integridade dos mesmos, pois
Silva Júnior (2001) já alertava que mesmo
o antigo Código Florestal (mais restritivo)
era ineficiente na proteção da vegetação e
diversidade arbórea de matas de galeria. Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 20 Considerações Finais A partir da caracterização das
tipologias remanescentes de Cerrado e da
análise de fragmentação dos remanescentes,
os resultados obtidos nesta pesquisa levam à
compreensão de que os níveis de fragmentação
da paisagem são elevados e com predomínio
de número de fragmentos pequenos envoltos
por uma matriz antrópica. Essa situação
remete esses fragmentos a maiores efeitos
bióticos e abióticos advindos do processo
de fragmentação da paisagem. Por outro
lado, há uma área superior de fragmentos
grandes, apesar de estes serem em menor
número, amenizando o equilíbrio das relações
ecológicas de organismos do topo da cadeia. Contrariando os resultados obtidos
anteriormente por Rodrigues (2010), para
a região, e por Mascarenhas et al. (2009)
utilizando imagens de média resolução,
nesta avaliação, apesar da alta densidade de
fragmentos (NP), as APP’s se encontram
conservadas e em conformidade com a
atual legislação. Essa diferença pode ser
decorrente dos detalhes fornecidos por uma
imagem de alta resolução, como a RapidEye e,
também, pela mudança de matriz circundante
para cana de açúcar, evidenciando maior
preocupação com as APP’s por parte do
setor sucroalcooleiro, que, por sua vez, é alvo Na matriz, destaca-se o predomínio de
atividades agrícolas, como a cana-de-açúcar,
inclusive em áreas de domínio dos Neossolos 556 Ambiência - Revista do Setor de Ciências Agrárias e Ambientais V.11 N.3 Set./Dez. 2015 Quartzarênicos, considerados solos frágeis e de
baixa aptidão agrícola, geralmente indicados
para pastagens e culturas perenes, sobretudo
arbóreas e com um manejo adequado. remanescentes mostram uma expressiva
fragmentação natural e suas possíveis
consequências ecológicas merecem destaque,
pois há uma proximidade da área estudada
com o Parque Nacional das Emas, a cerca
de 2 km apenas, integrando o Corredor
Ecológico Emas- Pantanal (de fluxo gênico,
sobretudo de animais do topo da cadeia
alimentar). Some-se a isso, que as zonas
escarpadas e os solos frágeis presentes
na paisagem, devem ser alvo de medidas
protetoras devido às consequências deletérias
que sua ocupação pode ocasionar. Nesse
sentido, seria importante prosseguir com
a avaliação aprofundada sobre os impactos
diretos e indiretos da sucessão de usos, tendo
vista que em média resolução, já foi feita e
sugere que estavam em desacordo com o
código florestal de então e não seriam alvo de
conservação na lei atual, o que é inquietante. Considerações Finais Apesar de a área ser representativa
quanto à fragmentação, as APP’s de
drenagem, principalmente nas Matas
Ciliares/ Galerias, não se apresentam com
número significativo de uso antrópico,
pois a continuidade dessa fisionomia está
sendo interrompida por outras classes de
remanescentes, resultantes de fragmentação
natural e da composição natural do Cerrado
através de mosaicos de fitofisionomias. ii
Os remanescentes do Cerrado
necessitam de mecanismos de planejamento,
envolvendo manutenção, fiscalização e
recuperação/recomposição de fragmentos
degradados para fins de conservação. Apesar
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Lmx1b Influences Correct Post-mitotic Coding of Mesodiencephalic Dopaminergic Neurons
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UvA-DARE (Digital Academic Repository)
Lmx1b Influences Correct Post-mitotic Coding of Mesodiencephalic
Dopaminergic Neurons
Wever, I.; Largo-Barrientos, P.; Hoekstra, E.J.; Smidt, M.P.
DOI
10.3389/fnmol.2019.00062
Publication date
2019
Document Version
Final published version
Published in
Frontiers in Molecular Neuroscience
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CC BY
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Citation for published version (APA):
Wever, I., Largo-Barrientos, P., Hoekstra, E. J., & Smidt, M. P. (2019). Lmx1b Influences
Correct Post-mitotic Coding of Mesodiencephalic Dopaminergic Neurons. Frontiers in
Molecular Neuroscience, 12, Article 62. https://doi.org/10.3389/fnmol.2019.00062 UvA-DARE (Digital Academic Repository) Lmx1b Influences Correct Post-mitotic Coding of Mesodiencephalic
Dopaminergic Neurons Citation for published version (APA):
Wever, I., Largo-Barrientos, P., Hoekstra, E. J., & Smidt, M. P. (2019). Lmx1b Influences
Correct Post-mitotic Coding of Mesodiencephalic Dopaminergic Neurons. Frontiers in
Molecular Neuroscience, 12, Article 62. https://doi.org/10.3389/fnmol.2019.00062 Citation for published version (APA):
Wever, I., Largo-Barrientos, P., Hoekstra, E. J., & Smidt, M. P. (2019). Lmx1b Influences
Correct Post-mitotic Coding of Mesodiencephalic Dopaminergic Neurons. Frontiers in
Molecular Neuroscience, 12, Article 62. https://doi.org/10.3389/fnmol.2019.00062 General rights
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published: 14 March 2019
doi: 10.3389/fnmol.2019.00062 Lmx1b Influences Correct
Post-mitotic Coding of
Mesodiencephalic Dopaminergic
Neurons
Iris Wever, Pablo Largo-Barrientos , Elisa J. Hoekstra and Marten P. Smidt*
Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, Netherlands The Lim Homeobox transcription factor 1 beta (LMX1b) has been identified as one of the
transcription factors important for the development of mesodiencephalic dopaminergic
(mdDA) neurons. During early development, Lmx1b is essential for induction and
maintenance of the Isthmic Organizer (IsO), and genetic ablation results in the disruption
of inductive activity from the IsO and loss of properly differentiated mdDA neurons. To study the downstream targets of Lmx1b without affecting the IsO, we generated a
conditional model in which Lmx1b was selectively deleted in Pitx3-expressing cells from
embryonic day (E)13 onward. Supporting previous data, no significant changes could be
observed in general dopamine (DA) marks, like Th, Pitx3 and Vmat2 at E14.5. However,
in depth analysis by means of RNA-sequencing revealed that Lmx1b is important for
the mRNA expression level of survival factors En1 and En2 and for the repression of
mdDA subset mark Ahd2 during (late) development. Interestingly, the regulation of Ahd2
by Lmx1b was found to be Pitx3 independent, since Pitx3 mRNA levels were not altered
in Lmx1b conditional knock-outs (cKOs) and Ahd2 expression was also up-regulated
in Lmx1b/Pitx3 double mutants compared to Pitx3 mutants. Further analysis of Lmx1b
cKOs showed that post-mitotic deletion of Lmx1b additional leads to a loss of TH+
cells at 3 months age both in the ventral tegmental area (VTA) and substantia nigra pars
compacta (SNc). Remarkably, different cell types were affected in the SNc and the VTA. While TH+AHD2+ cells were lost the SNc, TH+AHD2- neurons were affected in the VTA,
reflected by a loss of Cck expression, indicating that Lmx1b is important for the survival
of a sub-group of mdDA neurons. INTRODUCTION Accepted: 25 February 2019
Published: 14 March 2019 Dopamine (DA) is one of the catecholaminergic neurotransmitters found in the central
nervous system. Although cell bodies of DA neurons can be found in several positions
within the mammalian brain, the largest population of DA neurons is located in the ventral
mesencephalon (Björklund and Dunnett, 2007; Smidt and Burbach, 2007; Roeper, 2013). The
mesodiencephalic dopaminergic (mdDA) neuronal population consists of specific subsets with
distinct functions (Smits et al., 2006), including the substantia nigra pars compacta (SNc) and
the ventral tegmental area (VTA). The SNc innervates the dorsolateral striatum and caudate
putamen forming the nigrostriatal pathway. The nigrostriatal pathway is integrated in a complex Keywords: dopamine, development, transcription, LMX1B, substantia nigra Reviewed by: Reviewed by:
Sandra Blaess,
University of Bonn, Germany
Jerome Mertens,
Salk Institute for Biological Studies,
United States
Larissa Traxler,
University of Innsbruck, Innsbruck,
Austria, in
collaboration with reviewer JM *Correspondence:
Marten P. Smidt
m.p.smidt@uva.nl *Correspondence:
Marten P. Smidt
m.p.smidt@uva.nl Keywords: dopamine, development, transcription, LMX1B, substantia nigra Received: 04 December 2018
Accepted: 25 February 2019
Published: 14 March 2019 Edited by: Edited by:
Ildikó Rácz,
Universitätsklinikum Bonn, Germany Citation: In the absence
of Lmx1b the initiation, maintenance and inductive activity
of the IsO were found to be severely impaired, affecting the
development of the midbrain in general (Guo et al., 2007). Further analysis of Lmx1b-/- embryonic midbrains demonstrated
a reduction in TH-expressing cells in the ventral mesencephalon
(Smidt et al., 2000; Deng et al., 2011). Although the loss of
Lmx1b initially seemed to primarily affect the lateral group of
the DA progenitor domain (Deng et al., 2011), the medially
located TH-expressing cells failed to induce Pitx3 and were lost
during further development (Smidt et al., 2000). In contrast to the
Lmx1b null mutant, the conditional deletion of Lmx1b in mdDA
progenitors, but not in the IsO, resulted in normal development
of mdDA neurons (Yan et al., 2011). It was suggested that Lmx1a
compensates for the loss of Lmx1b, since the deletion of both led
to a severe reduction in both DA progenitors and mature mdDA
neurons (Yan et al., 2011). Further studies into the functional
redundancy between Lmx1a and Lmx1b suggest that the function
of Lmx1b in mdDA development and postnatal neuronal survival
can be mostly compensated by Lmx1a (Nakatani et al., 2010; showed that DatCre driven deletion of Lmx1b reduces the protein
levels of DAT and TH in the nerve terminals of mdDA neurons in
the dorsal and ventral Striatum of 18-month-old mutant mice. In
the present study we aimed to further elucidate the role of Lmx1b
in postmitotic development and neuronal survival of mdDA
neurons. We generated a mouse model that deletes Lmx1b in
postmitotic mdDA neurons, by crossing a floxed Lmx1b mutant
(Suleiman et al., 2007) with a Pitx3 driven iCre (Smidt et al.,
2012). The loss of Lmx1b did not affect the development of the
general DA phenotype, however a loss of TH+ cells was observed
in 3-month-old animals, suggesting that Lmx1b plays a role in
the survival and/or maintenance of mdDA neurons. During (late)
development we found reduced mRNA levels of En1 and En2,
which relates to the function of Lmx1b during early development
(Guo et al., 2007). In addition, the rostrolateral subset mark,
Ahd2, was increased and ectopically expressed at embryonic day
(E)14.5, however, the caudomedial mark Cck was unaffected. To
verify whether mdDA subsets were also affected during adult
stages, the amount of TH+AHD2+ cells were analyzed in both the
SNc and the VTA. Citation: Analysis of the Lmx1b
complete knock out mouse revealed that LMX1B is an essential
component of a positive feedback loop required to maintain
genes associated with the formation and functioning of the
Isthmic Organizer (IsO), including Wnt1, En1, En2, Pax2 and
Fgf8 (Adams et al., 2000; Guo et al., 2007). In the absence
of Lmx1b the initiation, maintenance and inductive activity
of the IsO were found to be severely impaired, affecting the
development of the midbrain in general (Guo et al., 2007). Further analysis of Lmx1b-/- embryonic midbrains demonstrated
a reduction in TH-expressing cells in the ventral mesencephalon
(Smidt et al., 2000; Deng et al., 2011). Although the loss of
Lmx1b initially seemed to primarily affect the lateral group of
the DA progenitor domain (Deng et al., 2011), the medially
located TH-expressing cells failed to induce Pitx3 and were lost
during further development (Smidt et al., 2000). In contrast to the
Lmx1b null mutant, the conditional deletion of Lmx1b in mdDA
progenitors, but not in the IsO, resulted in normal development
of mdDA neurons (Yan et al., 2011). It was suggested that Lmx1a
compensates for the loss of Lmx1b, since the deletion of both led
to a severe reduction in both DA progenitors and mature mdDA
neurons (Yan et al., 2011). Further studies into the functional
redundancy between Lmx1a and Lmx1b suggest that the function
of Lmx1b in mdDA development and postnatal neuronal survival
can be mostly compensated by Lmx1a (Nakatani et al., 2010;
Deng et al., 2011; Yan et al., 2011; Doucet-Beaupré et al., 2016). However, a recent study Laguna et al. (2015) identified a critical
role for Lmx1b in the functioning of mdDA neurons. They Animals All lines were maintained on a C57BL/6J background (Charles
River). Lmx1b-floxed animals were generated by R.L. Johnson
and obtained from R.Witzgall (University of Regensburg,
Germany) and have been previously described (Suleiman et al.,
2007). The Pitx3Cre has been previously generated in our lab
(Smidt et al., 2012). Pitx3CreCre; Lmx1b L/+ animals were
crossed with Lmx1b L/+ or Pitx3CreCre; Lmx1b L/+ animals to
generate Pitx3Cre/+; Lmx1b +/+, Pitx3Cre/+; Lmx1b L/L and
Pitx3CreCre; Lmx1b +/+, Pitx3CreCre; Lmx1b L/+ littermates. Embryos were isolated at E14.5, considering the morning of plug
formation as E0.5. Pregnant and adult mice were euthanized by
CO2 asphyxiation and embryos and brain were collected in 1×
PBS and immediately frozen on dry-ice (fresh frozen) or fixed by
immersion in 4% paraformaldehyde (PFA) for 4–8 h at 4◦C. After
PFA incubation, samples were cryoprotected O/N in 30% sucrose
at 4◦C. Embryos and brains were frozen on dry-ice and stored at
−80◦C. Cryosections were slices at 16 µm, mounted at Superfrost
plus slides, air-dried and stored at −80◦C until further use. Citation: Wever I, Largo-Barrientos P,
Hoekstra EJ and Smidt MP
(2019) Lmx1b Influences Correct
Post-mitotic Coding of
Mesodiencephalic
Dopaminergic Neurons. Front. Mol. Neurosci. 12:62. doi: 10.3389/fnmol.2019.00062 March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 1 Dopaminergic Subset Specifciation by Lmx1b Wever et al. network that controls voluntary movement and body posture,
and the degeneration of this pathway is the characteristic of
Parkinson’s disease (Braak et al., 2003). The VTA has projections
to the ventral striatum, the amygdala and the prefrontal
cortex. These pathways are involved in regulating emotion-
related behavior and are linked to addiction, depression and
schizophrenia (Prakash and Wurst, 2006; Smidt and Burbach,
2007). The different subsets of mdDA neurons can already be
distinguished during embryonic development based on their
molecular profile and anatomical position (Smits et al., 2013;
La Manno et al., 2016; Tiklová et al., 2019). It has been shown
that the rostrolateral population will eventually form the largest
part of the SNc, while the mediocaudal population is destined
to become the VTA (Hökfelt et al., 1980; Veenvliet et al., 2013). Recent studies have revealed that each subset is dependent
on a different transcriptional program for their development
and survival (Smits et al., 2006; Jacobs et al., 2011; Veenvliet
et al., 2013; Panman et al., 2014; Veenvliet and Smidt, 2014;
Kouwenhoven et al., 2017). The rostrolateral fate is determined
by a complex interplay between Pitx3 and En1. En1 induces
the general DA phenotype and influences Pitx3 expression,
while Pitx3 is required for the suppression of the caudomedial
phenotype by down-regulating En1 mRNA levels ((Veenvliet
et al., 2013). After the formation of the rostrolateral population
the remaining DA population will obtain a caudomedial fate
under the control of En1 (Bye et al., 2012; Veenvliet et al.,
2013). Another factor that has been shown to be crucial for
mdDA development and neuronal survival is LMX1B (Adams
et al., 2000; Smidt et al., 2000; Deng et al., 2011; Laguna et al.,
2015; Doucet-Beaupré et al., 2016). Analysis of the Lmx1b
complete knock out mouse revealed that LMX1B is an essential
component of a positive feedback loop required to maintain
genes associated with the formation and functioning of the
Isthmic Organizer (IsO), including Wnt1, En1, En2, Pax2 and
Fgf8 (Adams et al., 2000; Guo et al., 2007). Citation: Interestingly the amount of TH+AHD2+ cells
was less in the SNc of the mutant compared to controls, but not
in the VTA. In the VTA, TH+AHD2- cells were affected, which
was reflected in lower mRNA levels of Cck. Taken together, our
data shows that Lmx1b plays a role during the late development
and survival of mdDA neurons. During development Lmx1b is
important for maintaining proper En1 and En2 mRNA levels
and the repression of the subset mark Ahd2, while during adult
stages Lmx1b is important for the survival of a sub-group of
mdDA neurons. network that controls voluntary movement and body posture,
and the degeneration of this pathway is the characteristic of
Parkinson’s disease (Braak et al., 2003). The VTA has projections
to the ventral striatum, the amygdala and the prefrontal
cortex. These pathways are involved in regulating emotion-
related behavior and are linked to addiction, depression and
schizophrenia (Prakash and Wurst, 2006; Smidt and Burbach,
2007). The different subsets of mdDA neurons can already be
distinguished during embryonic development based on their
molecular profile and anatomical position (Smits et al., 2013;
La Manno et al., 2016; Tiklová et al., 2019). It has been shown
that the rostrolateral population will eventually form the largest
part of the SNc, while the mediocaudal population is destined
to become the VTA (Hökfelt et al., 1980; Veenvliet et al., 2013). Recent studies have revealed that each subset is dependent
on a different transcriptional program for their development
and survival (Smits et al., 2006; Jacobs et al., 2011; Veenvliet
et al., 2013; Panman et al., 2014; Veenvliet and Smidt, 2014;
Kouwenhoven et al., 2017). The rostrolateral fate is determined
by a complex interplay between Pitx3 and En1. En1 induces
the general DA phenotype and influences Pitx3 expression,
while Pitx3 is required for the suppression of the caudomedial
phenotype by down-regulating En1 mRNA levels ((Veenvliet
et al., 2013). After the formation of the rostrolateral population
the remaining DA population will obtain a caudomedial fate
under the control of En1 (Bye et al., 2012; Veenvliet et al.,
2013). Another factor that has been shown to be crucial for
mdDA development and neuronal survival is LMX1B (Adams
et al., 2000; Smidt et al., 2000; Deng et al., 2011; Laguna et al.,
2015; Doucet-Beaupré et al., 2016). Frontiers in Molecular Neuroscience | www.frontiersin.org Fluorescence Immunohistochemistry y
Cryosections were blocked with 4% HIFCS in 1× THZT
(50 mM Tris-HCL, pH 7.6; 0.5M NaCl; 0.5% Triton X-100)
and incubated with a primary antibody [Rb-TH (Pelfreeze
1:1,000), Sh-TH (Millipore AB1542, 1:750), Rb-AHD2 (Abcam
AB24343, 1:500)] in 1× THZT overnight at room temperature. The following day the slides were washed and incubated for
2 h at room temperature with secondary Alexafluor antibody
[anti-rabbit, anti-sheep (Invitrogen, 1:1,000)] in 1× TBS. After
immunostaining nuclei were staining with DAPI (1:3,000) and
embedded in Fluorsave (Calbiogen) and analyzed with the use
of a fluorescent microscope (Leica). All washes were done
in TBS and double stainings were performed sequential, with
immunostaining for TH being done first, followed by the
staining for AHD2. The antibody against AHD2 requires antigen
retrieval, which was executed as follows; slides were incubates
10 min in PFA after which they were submersed in 0.1 M citrate
buffer pH 6.0 for 3 min at 1,000 Watts followed by 9 min at
200 Watts. Slides were left to cool down, after which protocol was
followed as described above. RNA-Sequencing RNA was isolated from dissected E14.5 midbrains of Pitx3Cre/+;
Lmx1b +/+ and Lmx1b cKO embryos. RNA was isolated with
Trizol (ThermoFisher) according to manufacturer’s protocol. After isolation RNA clean-up was performed with an RNA
mini kit from Qiagen according to manufacturer’s protocol. E14.5 embryos were obtained from four different litters and
RNA isolated of six wildtype or six mutant embryos from
different nests was pooled to eventually form an n = 3 per
group. Pair-end RNA-sequencing (minimal 2 ∗10∧7 reads per
sample), mapping on the mouse genome and DESeq2 statistical
analysis on read counts was performed by Service XS (Leiden,
Netherlands). The mapping was done with the mouse ensemble
GRCm38.p4 data set. Genotyping Genotyping of the Lmx1b-Lox animals was done by PCR
using primers: forward 5′-AGGCTCCATCCATTCTTCTC, and
reverse: 5′-CCACAATAAGCAAGAGGCAC, resulting in a
wild-type product of 243 bp, or a LoxP-inserted product of March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 2 Dopaminergic Subset Specifciation by Lmx1b Wever et al. 277 bp. Pitx3-Cre genotyping was done by two different PCR’s. To discriminate between wild-type and Pitx3-Cre: forward 5′-GC
ATGATTTCAGGGATGGAC and reverse 5′-ATGCTCCTGT
CTGTGTGCAG, resulting in a product of 750 bp for a mutant
allele, and no product in wild-type animals. To additionally
discriminate between heterozygous and homozygous Pitx3-Cre
animals, primers were designed around Pitx3 exon 2 and exon
3: forward 5′-CAAGGGGCAGGAGCACA and reverse 5′-GTGA
GGTTGGTCCACACCG, resulting in a product of 390 bp for the
wildtype allele and no product for the mutant allele. were made based on anatomical landmarks. Everything rostral
of the supramammillary decussation was considered as SNc and
distinction between the SNc and the VTA was made based on
the tracts medial lemniscus, positioned between the SNc and
VTA. Cell numbers from all sections were pooled and statistical
analysis was performed via a student’s T-test. Graphs represent
the mean of counted cells divided by wildtype ± the SEM. Quantitative PCR (qPCR) (q
)
RNA was isolated from dissected E14.5 midbrains of Pitx3Cre/+;
Lmx1b +/+ and Lmx1b cKO embryos and of E14.5 Pitx3CreCre;
Lmx1b +/+ and Pitx3CreCre; Lmx1b L/L. RNA was isolated
with
Trizol
(ThermoFisher)
according
to
manufacturer’s
protocol. For the Lmx1b cKO three midbrains were pooled per
samples (n = 4 wildtype, n = 4 cKO) and further purified
on a column, according to the manufacturer’s protocol
(Qiagen, RNeasy mini kit). For the Pitx3/Lmx1b mutants,
a single midbrain was used per sample (n = 3 Pitx3CreCre;
Lmx1b+/+, n = 3 Pitx3CreCre; Lmx1b L/L) and no RNA
clean-up was performed. Relative mRNA levels were determined
by using the QuantiTect SYBR green PCR lightCycler kit
(Qiagen) according to the manufacturer’s instructions. For
each reaction 10 ng (dissected midbrain) of total RNA was
used as input. Primers used for Th, Ahd2, En1 and Cck were
previously published (Jacobs et al., 2011), primers for Lmx1b
forward 5′-GAGCAAAGATGAAGAAGCTGGC and reverse
5′-CTCCATGCGGCTTGACAGAA
(product
size
98
bp). Quantitative PCR (qPCR) data was analyzed using the delta-
delta Ct method and statistical analysis was performed via a
student’s T-test Graphs represent the fold change divided by
wildtype ± the SEM. Quantitative Analysis Quantification of TH-expressing neurons, TH+AHD2+ cells and
TH+AHD- cells in 3-month-old and 12-month-old midbrain
was performed in ImageJ as follows. First, TH-positive cells were
counted independently of whether or not they expressed AHD2. Cells were counted in 10–12 matching coronal sections covering
the whole midbrain dopaminergic neuronal pool [3 months old,
n = 3 wildtype, n = 4 conditional Lmx1b knock-out (cKO),
n = 3 for Pitx3 KOs, n = 3 for double Pitx3/Lmx1b KOs;
12 months old, n = 4 wildtype, n = 4 Lmx1b cKO]. Cells
were counted as TH+neurons when TH staining co-localized
with nuclear DAPI staining. For the double stained sections,
a color overlay was made and the double stained cells were
counted as TH+AHD2+ and the green cells were considered
as TH+AHD2- cells. Similar to the single TH quantifications,
cells were counted as neurons when the staining co-localized
with nuclear DAPI staining. The separation of the SNc and VTA In situ Hybridization In situ hybridization with digoxigenin (DIG)-labeled RNA
probes was performed as described (Smits et al., 2003; Smidt
et al., 2004). DIG-Labeled RNA probes for Th, Vmat2, Dat, Aadc,
Pitx3, En1, Ahd2 and Cck have been previously described (Grima
et al., 1985; Smits et al., 2003; Smidt et al., 2004; Jacobs et al.,
2007; Hoekstra et al., 2013). The used Lmx1b probe is a 310 bp
fragment containing exon 4–6 of the Lmx1b coding sequence. Specific Deletion of Lmx1b in Developing
Pitx3 Positive Neurons Studies on Lmx1b null mutants have shown that Lmx1b is
essential for the development of mdDA neurons. In the absence
of Lmx1b TH+ cells fail to induce Pitx3 and are lost during
development (Smidt et al., 2000). In addition to an effect on
the mdDA neuron population, Lmx1b was also found to be
essential for the expression of several genes associated with
IsO functioning, including Fgf8, Wnt1, En1, En2, Pax2 and
Gbx2, and that the loss of Lmx1b severely affects the initiation,
maintenance and the inductive activity of the IsO during
midbrain development (Guo et al., 2007). To study the role
of Lmx1b in mdDA development, without affecting the IsO,
we generated a conditional Lmx1b knock-out. Lmx1b-floxed
animals (Suleiman et al., 2007) were crossed with Pitx3Cre
animals (Smidt et al., 2012), to generate mice that lack Lmx1b
exclusively in Pitx3 positive neurons. Pitx3 expression is initiated
around E11.5 (Smidt et al., 1997) and it has been shown that
the onset of CRE activity of Pitx3-Locus driven Cre expression
is around E13.5 (Smidt et al., 2012), so to guarantee a complete
loss of Lmx1b transcript we decided to study E14.5 embryos. To verify that these animals lack Lmx1b only in the Pitx3
positive domain, we assessed the expression of Lmx1b in
Pitx3Cre/+; Lmx1b L/L compared to Pitx3Cre/+; Lmx1b +/+
littermates by means of in situ hybridization (Figure 1A). A
clear loss of Lmx1b expression could be observed in the mdDA
area (arrowheads) in Lmx1b L/L embryos, while expression
in other regions, like the diencephalon and the hindbrain,
remains unaffected (arrows). To examine whether the loss
of a Pitx3 allele and the complete loss of Lmx1b influenced
Pitx3 expression we analyzed its expression in adjacent slides
(Figure 1B). When comparing wild-type to mutant embryos
no clear difference in Pitx3 expression was observed. Taken
together, our analysis shows that crossing Lmx1b-floxed mice
with Pitx3Cre animals, generates a Lmx1b cKO, in which
the deletion is confined to the mdDA neuronal field, leaving
the rostral and caudal expression domains of Lmx1b intact. FIGURE 1 | Lmx1b expression is specifically lost in the Pitx3 positive domain
of Lmx1b conditional knock-outs (cKOs) embryos. Analysis of Lmx1b and
Pitx3 expression via in situ hybridization in embryonic day (E)14.5 midbrain
sagittal sections. (A) When comparing the Pitx3Cre/+; Lmx1b L/L to the
Pitx3Cre/+; Lmx1b+/+ a clear loss of Lmx1b is observed in the Pitx3 positive
area (arrowheads). Specific Deletion of Lmx1b in Developing
Pitx3 Positive Neurons However Lmx1b expression is maintained in regions that
are negative for Pitx3 (arrows). (B) Pitx3 expression seems unaffected. Scale bar = 300 µM. 2010; Arenas et al., 2015). The expression of these genes is
dependent on the combined functioning of Nurr1 and Pitx3
(Saucedo-Cardenas et al., 1998; Hwang et al., 2003; Nunes
et al., 2003; Smits et al., 2003; van den Munckhof et al., 2003;
Smidt et al., 2004; Jacobs et al., 2009). Loss of function studies
showed that Nurr1 mutants fail to induce Th, Vmat2 and
Dat, leading to a developmental arrest of mdDA progenitors
and cell death (Zetterström et al., 1997; Saucedo-Cardenas
et al., 1998; Smits et al., 2003). While Pitx3 was found to
be crucial for the correct specification of mdDA neurons by
acting as an activator of the NURR1 transcriptional complex MN9D Cell Culture and Transfection MN9D Cell Culture and Transfection
MN9D cells were cultured in Dulbecco’s Modified Eagle Medium
(DMEM) with 4.5 g/L glucose, supplemented with 10% (v/v)
HIFCS, 100 units/mL penicillin, 100 units/ml streptomycin
and 2 mM L-glutamine. Cells were grown on 10 cm-diameter
plates manually coated with poly-D-Lysine, in a humidified
atmosphere with 5% CO2 at 37◦C. Transfection of MN9D cells
was performed using Lipofectamine 2,000 reagent (Invitrogen)
on 10 cm-diameter plates-cultured MN9D cells on 70%–90%
confluency. Lipofectamine 2000 Reagent was diluted 1:50 (v/v)
in high glucose DMEM containing 2.5 µg of DNA vector, March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 3 Wever et al. Dopaminergic Subset Specifciation by Lmx1b in a total volume of 1.2 mL of DMEM. After 20 min of
incubation at room temperature, the DNA-lipid complexes mix
was diluted in medium without antibiotics (DMEM, 4.5 g/L
glucose, 10% HIFCS) to a final volume of 10 mL and MN9D
cells were incubated with this medium for 6 h in a humidified
atmosphere with 5% CO2 at 37◦C. Cells were harvested for
RNA isolation with Trizol (Thermofisher) according to the
manufacturer’s protocol. FIGURE 1 | Lmx1b expression is specifically lost in the Pitx3 positive domain
of Lmx1b conditional knock-outs (cKOs) embryos. Analysis of Lmx1b and
Pitx3 expression via in situ hybridization in embryonic day (E)14.5 midbrain
sagittal sections. (A) When comparing the Pitx3Cre/+; Lmx1b L/L to the
Pitx3Cre/+; Lmx1b+/+ a clear loss of Lmx1b is observed in the Pitx3 positive
area (arrowheads). However Lmx1b expression is maintained in regions that
i
f
Pit 3 (
) (B) Pit 3
i
ff
d Lmx1b Is Not Essential for Early
Post-mitotic Development of mdDA
Neurons As described above, the onset of Pitx3 driven CRE activity is
around E13.5 and by E14.5 most genes that define a mature
mdDA neuron, like Th, Vmat2 and Dat, are expressed (Iversen, March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 4 Wever et al. Dopaminergic Subset Specifciation by Lmx1b Conditional Removal of Lmx1b Results in
the Loss of TH+ Cells in the Adult Midbrain
Previous studies have already shown that Lmx1b expression is
continued throughout life (Smidt et al., 2000; Dai et al., 2008;
Laguna et al., 2015), suggesting that Lmx1b might have a role in
neuronal identity, maintenance and survival. A study performed
by Doucet-Beaupré et al. (2016) demonstrated that the combined
conditional genetic ablation of Lmx1a and Lmx1b under the
control of the Dat promoter causes a degeneration of TH+ cells
in 2-month-old mice in both the SNc and the VTA. To verify
whether Pitx3 driven deletion of Lmx1b also influenced neuronal
identity and/or maintenance, we examined the expression of Th,
Vmat2, Aadc, and Dat in 3-month-old midbrains (Figure 3). When comparing Pitx3Cre; Lmx1b +/+ animals to Pitx3Cre;
Lmx1b L/L animals no obvious alterations in the distribution Conditional Removal of Lmx1b Results in
the Loss of TH+ Cells in the Adult Midbrain (Jacobs et al., 2009). Since we removed Lmx1b in post-mitotic
cells, we hypothesized that any effect on the differentiation
of mdDA neurons would become apparent in the spatial
expression of Th, Vmat2 and Dat at E14.5. In addition to
these three factors, we also examined Aadc, which is also
required for the proper DA neurotransmitter phenotype and
is affected in both Nurr1 and the Pitx3 mutants, similar to
the other marks we examined (Saucedo-Cardenas et al., 1998;
Smidt et al., 2004). However, this gene is induced in an
early state at E10.5 in a Nurr1-independent manner (Smits
et al., 2003). When analyzing the spatial expression of Th via
in situ hybridization no clear differences could be observed
in the lateral or medial sections (Figure 2A). In addition,
the expression of Vmat2, Aadc and Dat also seem unaffected
by the post-mitotic loss of Lmx1b (Figures 2B–D). Together
these results suggest that post-mitotic deletion of Lmx1b does
not influence the expression of general DA marks Th, Vmat2,
Aadc and Dat. (Jacobs et al., 2009). Lmx1b Is Not Essential for Early
Post-mitotic Development of mdDA
Neurons Since we removed Lmx1b in post-mitotic
cells, we hypothesized that any effect on the differentiation
of mdDA neurons would become apparent in the spatial
expression of Th, Vmat2 and Dat at E14.5. In addition to
these three factors, we also examined Aadc, which is also
required for the proper DA neurotransmitter phenotype and
is affected in both Nurr1 and the Pitx3 mutants, similar to
the other marks we examined (Saucedo-Cardenas et al., 1998;
Smidt et al., 2004). However, this gene is induced in an
early state at E10.5 in a Nurr1-independent manner (Smits
et al., 2003). When analyzing the spatial expression of Th via
in situ hybridization no clear differences could be observed
in the lateral or medial sections (Figure 2A). In addition,
the expression of Vmat2, Aadc and Dat also seem unaffected
by the post-mitotic loss of Lmx1b (Figures 2B–D). Together
these results suggest that post-mitotic deletion of Lmx1b does
not influence the expression of general DA marks Th, Vmat2,
Aadc and Dat. FIGURE 3 | Pitx3 driven deletion of Lmx1b results in a change in Aadc and
Dat expression in the adult midbrain. Analysis of Th, Vmat2, Aadc and Dat in
coronal adult section in the Pitx3Cre/+; Lmx1b L/L mutant via in situ
hybridization. (A,B) Expression of Th and Vmat2 seem unaffected in
Pitx3Cre/+; Lmx1b L/L animals, but the Pitx3 driven deletion of Lmx1b results
in alteration in the expression of Aadc (C, arrowheads) and an overall
reduction in Dat expression (D) in both rostral and caudal sections
(arrowheads). Scale bar = 150 µM. FIGURE 2 | Several genes important for the dopamine (DA) neurotransmitter
phenotype are not clearly affected by the Pitx3Cre driven loss of Lmx1b at
E14.5. (A–D) In situ hybridization of Th, Vmat2, Aadc and Dat in E14.5
midbrain sagittal sections. (A–D) In both lateral and medial sections,
expression patterns of Th, Vmat2, Aadc and Dat do not seem to be affected
in Pitx3Cre/+; Lmx1b L/L embryos. Scale bar = 200 µM. FIGURE 2 | Several genes important for the dopamine (DA) neurotransmitter
phenotype are not clearly affected by the Pitx3Cre driven loss of Lmx1b at
E14.5. (A–D) In situ hybridization of Th, Vmat2, Aadc and Dat in E14.5
midbrain sagittal sections. Lmx1b Is Not Essential for Early
Post-mitotic Development of mdDA
Neurons (A–D) In both lateral and medial sections,
expression patterns of Th, Vmat2, Aadc and Dat do not seem to be affected
i
Pit 3C
/
L
1b L/L
b
S
l b
200
M FIGURE 3 | Pitx3 driven deletion of Lmx1b results in a change in Aadc and
Dat expression in the adult midbrain. Analysis of Th, Vmat2, Aadc and Dat in
coronal adult section in the Pitx3Cre/+; Lmx1b L/L mutant via in situ
hybridization. (A,B) Expression of Th and Vmat2 seem unaffected in
Pitx3Cre/+; Lmx1b L/L animals, but the Pitx3 driven deletion of Lmx1b results
in alteration in the expression of Aadc (C, arrowheads) and an overall
reduction in Dat expression (D) in both rostral and caudal sections
(arrowheads). Scale bar = 150 µM. FIGURE 3 | Pitx3 driven deletion of Lmx1b results in a change in Aadc and
Dat expression in the adult midbrain. Analysis of Th, Vmat2, Aadc and Dat in
coronal adult section in the Pitx3Cre/+; Lmx1b L/L mutant via in situ
hybridization. (A,B) Expression of Th and Vmat2 seem unaffected in
Pitx3Cre/+; Lmx1b L/L animals, but the Pitx3 driven deletion of Lmx1b results
in alteration in the expression of Aadc (C, arrowheads) and an overall
reduction in Dat expression (D) in both rostral and caudal sections
(arrowheads). Scale bar = 150 µM. FIGURE 2 | Several genes important for the dopamine (DA) neurotransmitter
phenotype are not clearly affected by the Pitx3Cre driven loss of Lmx1b at
E14.5. (A–D) In situ hybridization of Th, Vmat2, Aadc and Dat in E14.5
midbrain sagittal sections. (A–D) In both lateral and medial sections,
expression patterns of Th, Vmat2, Aadc and Dat do not seem to be affected
in Pitx3Cre/+; Lmx1b L/L embryos. Scale bar = 200 µM. March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 5 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 4 | The number of TH+ cells is decreased in Pitx3Cre/+; Lmx1b L/L
3-month-old midbrains. (A) Protein expression of TH (green) was evaluated
via immunohistochemistry in the adult midbrain of 3-month-old Lmx1b cKO
animals. A loss of TH signal was observed in both the ventral tegmental area
(VTA) and the substantia nigra pars compacta (SNc; white arrowheads). The
white dotted line represents the border between what is considered SNc and
VTA. Lmx1b Is Not Essential for Early
Post-mitotic Development of mdDA
Neurons (B) Quantification of TH+ cells in the adult midbrain of Pitx3Cre/+;
Lmx1b L/L (n = 3, gray bars) and Pitx3Cre/+; Lmx1b +/+ controls (n = 3,
black bars) shows that the total amount of TH+ neurons is significantly lower
(∼15% loss, ∗∗P < 0.05, one-tailed) and that neurons are lost both in the
SNc (∼13% loss, ∗P < 0.05) and the VTA (∼20% loss, ∗∗P < 0.01,
one-tailed). Pitx3Cre/+ Lmx1b+/+ animals were set at 1. Scale bar = 100 µM. cells in our model would also further progress. We quantified
the amount of TH+ cells in the SNc and the VTA in 12-
month-old midbrains (Figure 5A; Supplementary Table S1). A reduction of ∼10% of the total TH+ population is observed
in Pitx3Cre; Lmx1b L/L midbrains (n = 4, P < 0.05, one-
tailed; Figure 5B). In contrast to 3-month-old animals, the
loss of TH+ cells is only observed in the SNc, where the
amount of TH+ cells is reduced with 15% (n = 4, P < 0.05,
one-tailed) in the cKO midbrains, whereas the amount of
TH+ cells in the VTA are not significantly altered (n = 4;
Figure 5B). Together these results demonstrate that ablation
of Lmx1b leads to the loss of TH+ neurons. In the VTA,
an accelerated loss of cells is observed in the Lmx1b cKO,
leading to reduced numbers of TH+ cells at 3 months of age. However, over time the number of cells between the wildtype
and Pitx3Cre/+; Lmx1b L/L animals are equalized, suggesting
that during aging Pitx3Cre/+; Lmx1b +/+ animals also lose
cells in the VTA. Interestingly, the defect observed in the SNc
at 3 months of age, resides with time and is still present at
12 months of age. Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 p
In order to obtain a better insight into which molecular
mechanisms
are
affected
during
development
after
the
conditional removal of Lmx1b, we performed next-generation
RNA-sequencing on dissected E14.5 midbrains of Pitx3Cre/+;
Lmx1b +/+ and Pitx3Cre/ Lmx1b L/L embryos (n = 3; two pooled
embryos per biological replicate; Figure 6A). When using
a P-value cut-off of P < 0.05 285 genes were identified, of
which 105 were up-regulated and 180 are down-regulated. A PANTHER over-representation tests showed that these
genes are mainly associated with head and nervous system
development at E14.5 (Supplementary Figure S1). Among the
20 most regulated genes we found Aldh1a1 (Ahd2), En1 and
En2 (Figure 6B). En1 and En2 have been implicated in mdDA
neuronal survival before Albéri et al. (2004) and Simon et al. (2001). It was shown that within animals heterozygous for En1
lose DA neurons specifically in the SNc (Sonnier et al., 2007). In
addition to a role in neuronal survival En1 has been implicated
to play a role in the specification of the mdDA subpopulations. En1-deficient embryos demonstrate a down-regulation of Th,
Dat, Vmat2 and D2R in rostrolateral sub-population of mdDA
neurons, which are destined to become the SNc (Veenvliet
et al., 2013). Interestingly, this rostrolateral sub-population is
marked by the expression of Ahd2 (Jacobs et al., 2007; Veenvliet
et al., 2013), which shows an upregulation of approximately
two-fold (Figure 6B), suggesting that the loss of Lmx1b might
specifically affect the rostrolateral subset. To identify other
genes associated with mdDA neuronal development and the
rostrolateral subset, we cross-referenced the list of 285 possible
target genes to a list of genes known to be involved in mdDA
development (Figure 6C). In addition, we performed an overlay
of transcripts of Lmx1b target genes with the MAANOVA-FDR
analysis of genes regulated by either En1 (Veenvliet et al., FIGURE 4 | The number of TH+ cells is decreased in Pitx3Cre/+; Lmx1b L/L
3-month-old midbrains. (A) Protein expression of TH (green) was evaluated
via immunohistochemistry in the adult midbrain of 3-month-old Lmx1b cKO
animals. A loss of TH signal was observed in both the ventral tegmental area
(VTA) and the substantia nigra pars compacta (SNc; white arrowheads). The
white dotted line represents the border between what is considered SNc and
VTA. Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 (B) Quantification of TH+ cells in the adult midbrains of Pitx3Cre/+; Lmx1b L/L
(n = 4, gray bars) and Pitx3Cre/+; Lmx1b +/+ controls (n = 4, black bars) shows that the total amount of TH+ neurons is significantly lower (∼10% loss, ∗∗P < 0.05,
one-tailed) and that neurons are lost in the SNc (∼15% loss, ∗P < 0.05), but no difference was observed in the VTA (n = 4, one-tailed). Pitx3Cre/+ Lmx1b +/+
animals were set at 1. Scale bar = 100 µM. FIGURE 5 | In 12-month-old midbrains, a reduction in TH+ neurons is observed in Pitx3Cre driven Lmx1b cKO animals. (A) Protein expression of TH (green) was
evaluated by immunohistochemistry in adult midbrains of 12-month-old mutants and wildtypes. A loss of TH signal was observed in the SNc (white arrowheads). The
white dotted line represents the border between what is considered SNc and VTA. (B) Quantification of TH+ cells in the adult midbrains of Pitx3Cre/+; Lmx1b L/L
(n = 4, gray bars) and Pitx3Cre/+; Lmx1b +/+ controls (n = 4, black bars) shows that the total amount of TH+ neurons is significantly lower (∼10% loss, ∗∗P < 0.05,
one-tailed) and that neurons are lost in the SNc (∼15% loss, ∗P < 0.05), but no difference was observed in the VTA (n = 4, one-tailed). Pitx3Cre/+ Lmx1b +/+
animals were set at 1. Scale bar = 100 µM. 2013) or Pitx3 (Jacobs et al., 2011), two genes that have been
shown to be essential for the formation of rostrolateral mdDA
neurons (Veenvliet et al., 2013). We found nine genes that
were previously associated with mdDA development of which
En1 and Ahd2 were considered for further study (Figure 6D,
red arrows). Furthermore, genes regulated by both Lmx1b
and En1 were found to be regulated in the same direction
(Figure 6E). This in contrast to genes that are regulated by
both Lmx1b and Pitx3, which are mostly regulated reciprocal
(Figure 6F), similar to the Pitx3-En1 interplay (Veenvliet
et al., 2013). En1, En2 and CD9 were found to be regulated
by all three genes, Lmx1b, En1 and Pitx3 (Figure 6G), and while they are up-regulated in the Pitx3 mutant (Figure 6F),
they are down-regulated in both En1 mutants and Pitx3Cre/+;
Lmx1b L/L embryos (Figure 6E). Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 (B) Quantification of TH+ cells in the adult midbrain of Pitx3Cre/+;
Lmx1b L/L (n = 3, gray bars) and Pitx3Cre/+; Lmx1b +/+ controls (n = 3,
black bars) shows that the total amount of TH+ neurons is significantly lower
(∼15% loss, ∗∗P < 0.05, one-tailed) and that neurons are lost both in the
SNc (∼13% loss, ∗P < 0.05) and the VTA (∼20% loss, ∗∗P < 0.01,
one-tailed). Pitx3Cre/+ Lmx1b+/+ animals were set at 1. Scale bar = 100 µM. of Th and Vmat2 can be observed (Figures 3A,B), however
when analyzing the expression of Aadc, alterations can be
observed (Figure 3C, arrowheads) and when comparing Dat
expression between the wildtype and the Lmx1b cKOs an
overall reduction can be seen (Figure 3D, arrow heads). To
further establish whether changes in expression are caused by
the influence of Lmx1b on specific gene expression or on
neuronal survival in general, we aimed to quantify the total
amount of TH+ cells in the mdDA population. We performed
immunohistochemistry for TH and counted the cells in both
the SNc and the VTA (Figure 4A; Supplementary Table S1). The total amount of TH+ cells is reduced with ∼15% (n =
3, ∗∗P < 0.01, one-tailed) in the cKO compared to wildtype
littermates (Figure 4B). The loss of TH+ cells is largest in the
VTA (∼20% loss, ∗∗P < 0.01, one-tailed), while ∼13% of
the cells in the SNc are lost (Figure 4B). Since Laguna et al. (2015) found a progressive loss of cells between 2 months old
cKO mice and 18-month-old DatCre; Lmx1a L/L; Lmx1b L/L
animals, we aimed to investigate whether the reduction in TH+ March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 6 Wever et al. Dopaminergic Subset Specifciation by Lmx1b FIGURE 5 | In 12-month-old midbrains, a reduction in TH+ neurons is observed in Pitx3Cre driven Lmx1b cKO animals. (A) Protein expression of TH (green) was
evaluated by immunohistochemistry in adult midbrains of 12-month-old mutants and wildtypes. A loss of TH signal was observed in the SNc (white arrowheads). The
white dotted line represents the border between what is considered SNc and VTA. Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 Based on the results from
the genome-wide expression analysis and the association of
En1 and Ahd2 with the SNc (Jacobs et al., 2007; Sonnier et al.,
2007; Veenvliet et al., 2013), we decided to examine the spatial
expression of these two genes. In independent sets, the mRNA
levels were confirmed using qPCR and the spatial expression
patterns were studied using in situ hybridization. The levels of
En1 were 30% reduced (n = 4, ∗P < 0.05, one-tailed) in Lmx1b
mutant embryos compared to wildtype (Figure 7A, right panel),
validating our RNA-sequencing results. Interestingly, the spatial March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 7 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 6 | In vivo genome-wide expression analysis reveals that Lmx1b regulates genes involved in neuronal development, including En1 and En2 and acts as a
repressor of Ahd2 expression. (A) Schematic representation of the experimental setup for sample preparation and sample comparison. (B) Among the 20 most
regulated genes are Ahd2 (Aldh1a1, red), which shows an 85% increase, and En1 (red) and En2 (red) which are both down-regulated (En1, ∼25% loss, En2, ∼27%
loss). (C) Venn diagram illustrating that nine genes regulated by Lmx1b are mdDA enriched. (D) Relative expression of nine genes that were mdDA enriched. Among
the 9 genes En1 and Ahd2 were considered most interesting (red arrows). (E) Relative expression of genes regulated by both En1 and Lmx1b demonstrate that
genes that are regulated by the loss of En1 (gray bars) are affected in the same manner in the Pitx3Cre/+ Lmx1b L/L E14.5 embryos (black bars). Wildtype was set
at 1. (F) When looking at the relative expression of genes regulated by Pitx3 and Lmx1b the opposite effect is observed as with the En1 mutant. Genes up-regulated
in the Pitx3 mutant are down-regulated in the Lmx1b depleted embryos and genes up-regulated in the Pitx3Cre driven Lmx1b mutant are down-regulated in
embryos missing Pitx3. (G) Venn diagram showing that Lmx1b, En1 and Pitx3 are all involved in regulating En1, En2 and Cd9. FIGURE 6 | In vivo genome-wide expression analysis reveals that Lmx1b regulates genes involved in neuronal development, including En1 and En2 and acts as a
repressor of Ahd2 expression. (A) Schematic representation of the experimental setup for sample preparation and sample comparison. Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 (B) Among the 20 most
regulated genes are Ahd2 (Aldh1a1, red), which shows an 85% increase, and En1 (red) and En2 (red) which are both down-regulated (En1, ∼25% loss, En2, ∼27%
loss). (C) Venn diagram illustrating that nine genes regulated by Lmx1b are mdDA enriched. (D) Relative expression of nine genes that were mdDA enriched. Among
the 9 genes En1 and Ahd2 were considered most interesting (red arrows). (E) Relative expression of genes regulated by both En1 and Lmx1b demonstrate that
genes that are regulated by the loss of En1 (gray bars) are affected in the same manner in the Pitx3Cre/+ Lmx1b L/L E14.5 embryos (black bars). Wildtype was set
at 1. (F) When looking at the relative expression of genes regulated by Pitx3 and Lmx1b the opposite effect is observed as with the En1 mutant. Genes up-regulated
in the Pitx3 mutant are down-regulated in the Lmx1b depleted embryos and genes up-regulated in the Pitx3Cre driven Lmx1b mutant are down-regulated in
embryos missing Pitx3. (G) Venn diagram showing that Lmx1b, En1 and Pitx3 are all involved in regulating En1, En2 and Cd9. expression of En1 is not altered in the Pitx3Cre/+; Lmx1b L/L
embryos compared to Pitx3Cre/+; Lmx1b +/+ embryos at E14.5
(Figure 7A, left panel), suggesting that Lmx1b is important
for the mRNA expression level of En1 in all mdDA neurons. When examining the expression pattern of Ahd2, an increase
in expression can be found in the entire Ahd2 positive area (Figure 7B) and clear ectopic Adh2 expression is found in
the more medial sections (Figure 7B, left panel, arrowheads). Interestingly, the Ahd2 positive domain seems to exceed that of
Th in the lateral sections of E14.5 Lmx1b cKO midbrains, while
in the medial sections it remains restricted to the Th positive
domain (Figure 7C, arrowhead). Quantification of the mRNA March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 8 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 7 | Ahd2 is up-regulated and ectopically expressed in Pitx3Cre driven Lmx1b mutants, together with a generic reduction in En1 expression in the midbrain
of E14.5 mutant embryos. The expression patterns of En1 and Ahd2 in E14.5 midbrains of Lmx1b cKO and wildtype were analyzed via in situ hybridization, followed
by quantitative PCR (qPCR) to quantify the mRNA levels. Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 (A) The mRNA expression level of En1 is significantly reduced (∼30% loss, ∗p < 0.05, one-tailed) in the
Pitx3Cre/+ Lmx1b L/L E14.5 midbrain (n = 4, gray bars) compared to wildtype littermate controls (n = 4, black bars). Wildtype mRNA levels were set at 1. When
visualizing the expression pattern an overall reduction in En1 expression can be seen in both lateral and medial sections. (B) Ahd2 expression can be found
ectopically in medial section (arrowheads) and an up-regulation can be observed in more lateral sections. Quantification of the mRNA levels via qPCR shows an
increase of ∼130% (n = 4, ∗∗P < 0.01, one-tailed) in the mutant (gray bar) compared to the wildtype (black bar). Wildtype was set at 1. (C) Pseudo-overlay of the
lateral and medial expression pattern of Ahd2 (red) and Th (green). In the lateral sections the Ahd2 expression domain exceeds the Th positive domain (white
arrowhead), while the ectopic expression of Ahd2 in the medial sections is restricted to the spatial expression of Th. Scale bar = 300 µM. FIGURE 7 | Ahd2 is up-regulated and ectopically expressed in Pitx3Cre driven Lmx1b mutants, together with a generic reduction in En1 expression in the midbrain
of E14.5 mutant embryos. The expression patterns of En1 and Ahd2 in E14.5 midbrains of Lmx1b cKO and wildtype were analyzed via in situ hybridization, followed
by quantitative PCR (qPCR) to quantify the mRNA levels. (A) The mRNA expression level of En1 is significantly reduced (∼30% loss, ∗p < 0.05, one-tailed) in the
Pitx3Cre/+ Lmx1b L/L E14.5 midbrain (n = 4, gray bars) compared to wildtype littermate controls (n = 4, black bars). Wildtype mRNA levels were set at 1. When
visualizing the expression pattern an overall reduction in En1 expression can be seen in both lateral and medial sections. (B) Ahd2 expression can be found
ectopically in medial section (arrowheads) and an up-regulation can be observed in more lateral sections. Quantification of the mRNA levels via qPCR shows an
increase of ∼130% (n = 4, ∗∗P < 0.01, one-tailed) in the mutant (gray bar) compared to the wildtype (black bar). Wildtype was set at 1. (C) Pseudo-overlay of the
lateral and medial expression pattern of Ahd2 (red) and Th (green). Lmx1b Regulates Genes Involved in
Neuronal Development and Acts as a
Repressor of Ahd2 In the lateral sections the Ahd2 expression domain exceeds the Th positive domain (white
arrowhead), while the ectopic expression of Ahd2 in the medial sections is restricted to the spatial expression of Th. Scale bar = 300 µM. of the SNc, is marked by the expression of Ahd2 (Jacobs et al.,
2011; Veenvliet et al., 2013), while the caudomedial population,
destined to become the VTA, expresses Cck (Hökfelt et al.,
1980; Veenvliet et al., 2013). It was shown that these subsets are
dependent on different transcriptional programs for their proper
specification and survival (Smits et al., 2006; Jacobs et al., 2011;
Veenvliet et al., 2013; Panman et al., 2014; Kouwenhoven et al.,
2017). The rostrolateral population is dependent on a complex
interplay between Pitx3 and En1, in which En1 is required for the
induction of DA phenotype and influences Pitx3 expression and
Pitx3 is required for antagonizing the caudomedial phenotype
by regulating EN1 functioning (Veenvliet et al., 2013). When the
rostrolateral population is formed, the remaining DA population
will obtain a caudomedial phenotype under the influence of
En1 (Bye et al., 2012; Veenvliet et al., 2013). Our results show
lower mRNA levels of En1 and an expansion of the rostrolateral
mark Ahd2 (Figure 7), suggesting that the specification of
mdDA subsets is affected. In order to determine whether expression level showed a two-fold increase in Adh2 (n = 4,
∗∗p < 0.01, one-tailed) in the mutant compared to wildtype
(Figure 7B, right panel). Interestingly, initial observation in
Lmx1b overexpression studies in MN9D cells pointed already to
this regulatory role of Lmx1b, since we found a 48% reduction
in Ahd2 levels after Lmx1b overexpression (n = 4, ∗∗P < 0.01,
two-tailed; Supplementary Figure S2). In summary, we show
that Lmx1b is important for the regulation of genes associated
with mdDA neuronal development, including En1, and that
Lmx1b acts as a repressor of Ahd2. TH+ Cells Are Lost in the SNc and the VTA
of Pitx3Cre/+ Lmx1b L/L Adult Mice During embryonic development different subsets of mdDA
neurons can already be distinguished based on their molecular
profile and anatomical position (Smits et al., 2013; Panman
et al., 2014; La Manno et al., 2016; Tiklová et al., 2019). As
described above, the rostrolateral subset, that will later form most March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 9 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 8 | Cck expression is not affected in the midbrain of E14.5 Pitx3cre/+ Lmx1b L/L embryos, but shows a clear loss in the adult mutant midbrain. (A) Expression of Cck was examined using in situ hybridization and qPCR. No significant difference in mRNA level was found between Lmx1b conditional mutant
E14.5 midbrain (n = 4, gray bar) and wildtype littermate midbrains (n = 4, black bar). Wildtype mRNA levels were set at 1. In addition, the expression pattern of Cck is
comparable between Pitx3Cre/+; Lmx1b L/L E14.5 midbrain sections and Pitx3Cre/+ Lmx1b +/+ E14.5 midbrain sections. (B) In situ hybridization on the adult
midbrain demonstrates a loss of Cck expression in the lateral parts of the VTA (black arrowheads, n = 4 observation). (C) The expression pattern of Th was visualized
using in situ hybridization on adjacent sections. Scale bars are 200 µM. FIGURE 8 | Cck expression is not affected in the midbrain of E14.5 Pitx3cre/+ Lmx1b L/L embryos, but shows a clear loss in the adult mutant midbrain. (A) Expression of Cck was examined using in situ hybridization and qPCR. No significant difference in mRNA level was found between Lmx1b conditional mutant
E14.5 midbrain (n = 4, gray bar) and wildtype littermate midbrains (n = 4, black bar). Wildtype mRNA levels were set at 1. In addition, the expression pattern of Cck is
comparable between Pitx3Cre/+; Lmx1b L/L E14.5 midbrain sections and Pitx3Cre/+ Lmx1b +/+ E14.5 midbrain sections. (B) In situ hybridization on the adult
midbrain demonstrates a loss of Cck expression in the lateral parts of the VTA (black arrowheads, n = 4 observation). (C) The expression pattern of Th was visualized
using in situ hybridization on adjacent sections. Scale bars are 200 µM. the loss of Lmx1b influences the caudomedial population at
E14.5 we examined the expression of Cck (Figure 8A). TH+ Cells Are Lost in the SNc and the VTA
of Pitx3Cre/+ Lmx1b L/L Adult Mice No
differences in spatial expression could be observed between the
wildtype and Pitx3Cre/+; Lmx1b L/L embryos when analyzing
Cck using in situ hybridization (Figure 8A, left panel). In
addition, the mRNA levels of Cck are not significantly different
between the wildtype and the cKO (n = 4; Figure 8A, right
panel). Next to analyzing the expression at E14.5, we also
decided to examine the spatial expression in 3-month-old
adult midbrains (Figure 8B). Interestingly, a clear loss of
Cck expression can be observed in the more lateral positions
of the VTA (arrowheads; n = 2 observation), which seems
to match the position where we observed the initial loss in
TH+ cells (Figure 4B, arrowheads). However, Th expression
can still be found in corresponding areas in adjacent slides
(Figure 8C), suggesting that Cck is down-regulated in these
areas. To further substantiate the effect of the deletion of Lmx1b
on the adult mdDA subsets, we also examined the expression
of Adh2 in the adult midbrain and quantified the amount of
TH+AHD2+ cells in the SNc and the VTA (Figure 9A). The ectopic Ahd2 expression observed in the embryo could not be
detected in the adult stages (Supplementary Figure S3) and
a ∼18% loss of TH+AHD2+ cells (n = 3, ∗P < 0.05, one-
tailed) is even observed in the SNc (Figure 9B; Supplementary
Table S1). While the TH+AHD2+ are affected in the SNc,
the AHD2 negative population is significantly reduced in the
VTA, where ∼21% of the TH+AHD2- cells are lost (n = 3,
∗∗P < 0.01, one-tailed) in the mutant compared to Pitx3Cre/+;
Lmx1b +/+ midbrains (Figure 9C; Supplementary Table S1). Together, our data shows that the DA-subsets are differential
affected by the loss of Lmx1b in both the embryonic and
adult stage. Frontiers in Molecular Neuroscience | www.frontiersin.org Lmx1b Represses Ahd2 Expression
Independent of Pitx3 The transcriptional activation of Ahd2 in the rostrolateral
population has been demonstrated to be dependent on the
expression of Pitx3 (Jacobs et al., 2007). Pitx3 can interact
with a region close to the transcriptional start site of Ahd2
and loss of expression is observed in Pitx3−/−embryos and March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 10 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 9 | The TH+ cells that are lost in the SNc of 3-month-old mice conditionally depleted of Lmx1b are also AHD2+, while the TH+AHD2- population is
affected in the VTA. Immunohistochemistry of TH (green) and AHD2 (red) in the adult midbrain of 3-month-old animals. (A) Assessment of the TH+AHD2+ (yellow)
and the TH+AHD2- (green) population in both the SNc and VTA of Pitx3Cre/+ Lmx1b L/L animals shows that in the SNc (upper panel) the TH+ AHD2+ neurons are
affected, while TH+ neurons affected in the VTA (lower panel) are AHD2- (white arrowheads). The white dotted line represents the border between what is considered
SNc and VTA. (B) Quantification of the amount of cells in the SNc shows that the number of TH+AHD2+ neurons are reduced (∼18%, n = 3, ∗p < 0.05, one-tailed),
while the number TH+AHD2- cells are similar to the Pitx3Cre/+ Lmx1b +/+ (n = 3, black bar). (C) This in contrast to the VTA, where TH+AHD2- cells are lost (∼21%
loss, n = 3, ∗∗P < 0.01, one-tailed) in the Lmx1b cKO (gray bar) and TH+AHD2+ cells are not significantly reduced. The number of cells of in the midbrain of
Pitx3Cre/+; Lmx1b +/+ animals were set at 1. Scale bar = 100 µM. FIGURE 9 | The TH+ cells that are lost in the SNc of 3-month-old mice conditionally depleted of Lmx1b are also AHD2+, while the TH+AHD2- population is
affected in the VTA. Immunohistochemistry of TH (green) and AHD2 (red) in the adult midbrain of 3-month-old animals. (A) Assessment of the TH+AHD2+ (yellow)
and the TH+AHD2- (green) population in both the SNc and VTA of Pitx3Cre/+ Lmx1b L/L animals shows that in the SNc (upper panel) the TH+ AHD2+ neurons are
affected, while TH+ neurons affected in the VTA (lower panel) are AHD2- (white arrowheads). The white dotted line represents the border between what is considered
SNc and VTA. Lmx1b Represses Ahd2 Expression
Independent of Pitx3 (B) Quantification of the amount of cells in the SNc shows that the number of TH+AHD2+ neurons are reduced (∼18%, n = 3, ∗p < 0.05, one-tailed),
while the number TH+AHD2- cells are similar to the Pitx3Cre/+ Lmx1b +/+ (n = 3, black bar). (C) This in contrast to the VTA, where TH+AHD2- cells are lost (∼21%
loss, n = 3, ∗∗P < 0.01, one-tailed) in the Lmx1b cKO (gray bar) and TH+AHD2+ cells are not significantly reduced. The number of cells of in the midbrain of
Pitx3Cre/+; Lmx1b +/+ animals were set at 1. Scale bar = 100 µM. (Jacobs et al., 2007). In addition to the generation of RA,
Ahd2 has also been found to have a protective role for SNc
neurons by detoxifying aldehydes (Wey et al., 2012). With partial
rescue of Adh2 expression in the Pitx3/Lmx1b double mutant
we hypothesized that this might also rescue part of the TH+
cells that are lost in full Pitx3 mutants in adults. We therefore
analyzed the amount of TH+ and TH+AHD2+ cells in the
adult midbrain of Pitx3 mutants and Pitx3CreCre/Lmx1b L/L
animals by performing immunohistochemistry for TH (green)
and AHD2 (red; Figure 11A). More TH+ cells could be observed
in the lateral tier of the SNc (Figure 11A, arrows) and in
the VTA (Figure 11A, arrowheads). Quantification showed an
∼16% increase in TH+ cells (n = 3, ∗∗p < 0.01, one-tailed) and
an upward trend for the amount of TH+AHD2+ cells (n = 3;
p = 0.12, one-tailed) in Pitx3/Lmx1b double mutant midbrains
compared to Pitx3 mutants (Figure 11B; Supplementary
Table S1). To summarize our data, by conditionally removing
Lmx1b in a Pitx3 mutant background a recovery of Ahd2
expression is observed at E14.5 and an increase in TH+ cells can
be detected in the adult midbrain of Pitx3CreCre; Lmx1b L/L
animals compared to Pitx3 mutants alone. adult midbrains (Jacobs et al., 2007). As mentioned above,
the Pitx3 driven deletion of Lmx1b causes an up-regulation
of Adh2 and to get a better insight into the mechanisms via
which Lmx1b influences Ahd2 expression we generated a double
mutant for Pitx3 and Lmx1b (Pitx3CreCre; Lmx1b L/L). We
analyzed the expression of Ahd2, En1 and Th in Pitx3CreCre;
Lmx1b L/L and Pitx3CreCre; Lmx1b +/+ embryos with in situ
hybridization (Figures 10A–C). Lmx1b Represses Ahd2 Expression
Independent of Pitx3 When analyzing the expression
of Ahd2 we observed a clear increase in signal in several
sections (arrows) in embryos depleted of both Pitx3 and Lmx1b
(Figure 10A). In more lateral sections this increase in Ahd2
was less pronounced (Figure 10A). Importantly, The expression
of En1 and Th seem unaffected (Figures 10B,C), suggesting
that the ablation of Lmx1b only leads to an increase in Ahd2
expression in a Pitx3 mutant background. To quantify the mRNA
levels, we isolated RNA from E14.5 dissected midbrains of
Pitx3CreCre: Lmx1b L/L and Pitx3CreCre: Lmx1b +/+ embryos
(Figure 10D). As expected, the Lmx1b mRNA levels were
severely reduced as a consequence of the conditional ablation
(n = 3, ∗P < 0.05, one-tailed). In addition to the loss of Lmx1b,
Ahd2 was up-regulated to ∼160% (n = 3, ∗p < 0.05, one-
tailed), indicating that repression of Ahd2 by Lmx1b does not
require Pitx3. Frontiers in Molecular Neuroscience | www.frontiersin.org DISCUSSION AHD2 is an aldehyde dehydrogenase and generates RA and in
previous studies it has been shown that a partial recovery of TH+
neurons in the rostrolateral population can be accomplished
by maternal supplementation with RA in Pitx3 ablated animals Lmx1b has been associated with several developmental processes
(Chen et al., 1998; Adams et al., 2000; Smidt et al., 2000;
Guo et al., 2007; Deng et al., 2011; Yan et al., 2011). March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 11 Dopaminergic Subset Specifciation by Lmx1b Wever et al. FIGURE 10 | Lmx1b represses Ahd2 independent of Pitx3. The expression patterns of Adh2, En1 and Th were examined using in situ hybridization and the mRNA
levels for Lmx1b, Ahd2, En1, Cck and Th were verified using qPCR. (A) Pitx3Cre driven loss of Lmx1b in Pitx3CreCre animals caused an increase in Ahd2 expression
in the medial sections of the E14.5 midbrain (black arrowheads). (B,C) The expression pattern of En1 did not show clear differences between Pitx3CreCre; Lmx1b
+/+ and the Pitx3CreCre Lmx1b mutant and neither did Th. (D) The mRNA expression levels of Lmx1b were down-regulated with ∼76% in conditional mutant (gray
bar, n = 3, ∗p < 0.05, one-tailed), while Ahd2 shows an increase of ∼160% in the Pitx3CreCre; Lmx1b L/L (gray bar, n = 3, ∗p < 0.05, one-tailed) compared to
Pitx3CreCre; Lmx1b +/+ littermates (red line, n = 4). The mRNA expression levels of En1, Cck and Th did not show significant changes. Wildtype was set at 1. Scale
bar = 100 µM. FIGURE 10 | Lmx1b represses Ahd2 independent of Pitx3. The expression patterns of Adh2, En1 and Th were examined using in situ hybridization and the mRNA
levels for Lmx1b, Ahd2, En1, Cck and Th were verified using qPCR. (A) Pitx3Cre driven loss of Lmx1b in Pitx3CreCre animals caused an increase in Ahd2 expression
in the medial sections of the E14.5 midbrain (black arrowheads). (B,C) The expression pattern of En1 did not show clear differences between Pitx3CreCre; Lmx1b
+/+ and the Pitx3CreCre Lmx1b mutant and neither did Th. DISCUSSION (D) The mRNA expression levels of Lmx1b were down-regulated with ∼76% in conditional mutant (gray
bar, n = 3, ∗p < 0.05, one-tailed), while Ahd2 shows an increase of ∼160% in the Pitx3CreCre; Lmx1b L/L (gray bar, n = 3, ∗p < 0.05, one-tailed) compared to
Pitx3CreCre; Lmx1b +/+ littermates (red line, n = 4). The mRNA expression levels of En1, Cck and Th did not show significant changes. Wildtype was set at 1. Scale
bar = 100 µM. March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 12 Wever et al. Dopaminergic Subset Specifciation by Lmx1b FIGURE 11 | Additional Lmx1b deletion partially rescues the cell loss observed in Pitx3 mutants alone. TH (green) and AHD2 (red) expression was assessed using
immunohistochemistry in coronal midbrain sections of 3-month-old animals. (A) More TH+ cells were observed in lateral tier of the SNc (white arrows) and in the VTA
(white arrowhead). The white dotted line represents the border between what is considered SNc and VTA. (B) Quantification of the total amount of TH+ cells shows
an increase in TH+ cells of ∼16% (n = 3, ∗∗P < 0.01, one-tailed). The number of TH+AHD2+ cells are not significant increased, but shows an upward trend (n = 3,
P = 0.12, one-tailed). Pitx3CreCre; Lmx1b +/+ was set at 1. Scale bar = 100 µM. During early development Lmx1b is a critical component of
the positive feedback loop that maintains genes associated
Deng et al., 2011). Next to a developmental role, a recent
study also linked Lmx1b to cellular homeostasis of mdDA FIGURE 11 | Additional Lmx1b deletion partially rescues the cell loss observed in Pitx3 mutants alone. TH (green) and AHD2 (red) expression was assessed using
immunohistochemistry in coronal midbrain sections of 3-month-old animals. (A) More TH+ cells were observed in lateral tier of the SNc (white arrows) and in the VTA
(white arrowhead). The white dotted line represents the border between what is considered SNc and VTA. (B) Quantification of the total amount of TH+ cells shows
an increase in TH+ cells of ∼16% (n = 3, ∗∗P < 0.01, one-tailed). The number of TH+AHD2+ cells are not significant increased, but shows an upward trend (n = 3,
P = 0.12, one-tailed). Pitx3CreCre; Lmx1b +/+ was set at 1. Scale bar = 100 µM. ETHICS STATEMENT All
animal
studies
were
performed
in
accordance
with
local animal welfare regulations, as this project has been
approved by the animal experimental committee (Dier ethische
commissie, Universiteit van Amsterdam; DEC-UvA), and
international guidelines. ACKNOWLEDGMENTS We would like to thank Dr. Ralph Witzgall of the University
of Regensburg (Germany) for the kind gift of the Lmx1b-
floxed mouse. DATA AVAILABILITY The datasets generated for this study can be found in
GEO, GSE121120. DISCUSSION These findings corroborated initial observations that Lmx1b
overexpression in a dopaminergic cell line (MN9D) led to the
repression of Ahd2 mRNA expression. Interestingly, the shift
in Ahd2 towards the more medial DA domain did not affect
the expression of Cck during development. The actual increased
level of Ahd2 mRNA could be attributed to a higher level per
cell in addition to the ectopic presence. The transcriptional
activation of Ahd2 has been demonstrated to be dependent on
the expression of Pitx3 and En1 (Jacobs et al., 2007; Veenvliet
et al., 2013). Pitx3 can interact with a region close to the
transcriptional start site of Ahd2 and loss of expression is
observed in Pitx3−/−embryos and adult midbrains (Jacobs
et al., 2007). The ablation of Lmx1b did not influence the
mRNA expression levels of Pitx3 itself (not shown) and also the
spatial expression of Pitx3 was not affected. In addition, spatial
expression of Ahd2 was partially recovered in Pitx3/Lmx1b
double mutants compared to Pitx3 mutants, suggesting that the
regulation of Ahd2 by Lmx1b is a PITX3-independent process. Interestingly, the Ahd2 promoter contains several conserved
FLAT-elements, to which LMX1B can bind directly (Rascle et al.,
2009), suggesting that LMX1B might directly influence Ahd2
expression. AHD2 is involved in the generation of retinoic acid
(RA) from retinol and Jacobs et al. (2007) demonstrated that
administration of RA to the maternal diet of Pitx3−/−animals
led to an increase in TH+ cells during development and an
increased innervation of the dorsal Striatum. Although we only
observed a partial rescue of Ahd2 expression, we did observe
an increase in TH+ cells in the midbrain of adult Pitx3CreCre;
Lmx1b L/L animals. Overall, our data shows that Lmx1b is important for the
mRNA expression level of survival factors En1 and En2 during FUNDING This work was sponsored by the NWO-ALW (Nederlandse
Organisatie
voor
Wetenschappelijk
Onderzoek-Aard
en
Levenswetenschappen) VICI grant (865.09.002) awarded to
Prof. Dr. MS. The funders had no role in study design, data
collection and analysis, decision to publish, or preparation of
the manuscript. AUTHOR CONTRIBUTIONS IW performed the experiments, analyzed the data and wrote
the mansuscript. PL-B and EH performed the experiments. MS
initiated the study, analyzed the data, wrote the manuscript,
funded the study. DISCUSSION During early development Lmx1b is a critical component of
the positive feedback loop that maintains genes associated
with IsO functioning. When studying Lmx1b null mutants, it
was demonstrated that Wnt1, Fgf8, En1 and En2 all require
Lmx1b to maintain their expression at the IsO and in mdDA
progenitors (Adams et al., 2000; Guo et al., 2007; Anderegg
et al., 2013; Sherf et al., 2015). The loss of Lmx1b initially
seems to predominantly affect the lateral positioned mdDA
progenitor pool, however the medial located progenitors are
arrested in their development as they fail to co-express Th
and Pitx3 and are lost during later stages (Smidt et al., 2000; Deng et al., 2011). Next to a developmental role, a recent
study also linked Lmx1b to cellular homeostasis of mdDA
neurons in adult mice (Laguna et al., 2015). By using a Dat
driven Cre, Lmx1b was removed in postmitotic mdDA neurons,
which led to dysregulation of the autophagic-lysosal pathway,
abnormal synaptic responses and reduced protein levels of
striatal TH and DAT expression, all processes that contribute
to a Parkinson’s disease pathology (Laguna et al., 2015). Here,
we showed that Lmx1b has a variety of functions during
postmitotic development, including subset specification and
neuronal survival. In line with its function in early development, During early development Lmx1b is a critical component of
the positive feedback loop that maintains genes associated
with IsO functioning. When studying Lmx1b null mutants, it
was demonstrated that Wnt1, Fgf8, En1 and En2 all require
Lmx1b to maintain their expression at the IsO and in mdDA
progenitors (Adams et al., 2000; Guo et al., 2007; Anderegg
et al., 2013; Sherf et al., 2015). The loss of Lmx1b initially
seems to predominantly affect the lateral positioned mdDA
progenitor pool, however the medial located progenitors are
arrested in their development as they fail to co-express Th
and Pitx3 and are lost during later stages (Smidt et al., 2000; March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 13 Wever et al. Dopaminergic Subset Specifciation by Lmx1b we found that a loss of Lmx1b in postmitotic neurons leads
to reduced mRNA levels of En1 and En2 at E14.5. DISCUSSION Both of
these factors have be associated with the survival of mdDA
neurons (Simon et al., 2001; Albéri et al., 2004) and En1 ±
animals show a progressive loss of TH+ cells in the ventral
midbrain from 3 weeks of age onward (Sonnier et al., 2007). The lower mRNA levels of En1, if equally represented by a lower
protein level, might contribute to the observed reduction in the
amount of TH+ cells in 3-month-old Pitx3Cre/+; Lmx1b L/L
animals, however in heterozygous En1 mice the SNc is mostly
affected, while in our model initially the VTA demonstrates the
largest percentile reduction in TH+ cells. Notably, after 1 year
the defect resides in the SNc suggesting a change in timing
of neuronal survival which culminates in loss of SNc neurons
after 12 months, which is in line with observations done in
En1 mutant animals. In addition to a role in neuronal survival
we also found a role for Lmx1b in subset specification. Pitx3
driven deletion of Lmx1b led to a significant increase in the
rostrolateral mark Ahd2 in vivo and in vitro. In E14.5 embryos,
an extension of the Ahd2 positive domain was observed, as
Ahd2 expression was found ectopically in medial sections of
the ventral mesencephalon, overlapping with the Cck positive
domain, and in lateral sections exceeding the Th positive domain. These findings corroborated initial observations that Lmx1b
overexpression in a dopaminergic cell line (MN9D) led to the
repression of Ahd2 mRNA expression. Interestingly, the shift
in Ahd2 towards the more medial DA domain did not affect
the expression of Cck during development. The actual increased
level of Ahd2 mRNA could be attributed to a higher level per
cell in addition to the ectopic presence. The transcriptional
activation of Ahd2 has been demonstrated to be dependent on
the expression of Pitx3 and En1 (Jacobs et al., 2007; Veenvliet
et al., 2013). Pitx3 can interact with a region close to the
transcriptional start site of Ahd2 and loss of expression is
observed in Pitx3−/−embryos and adult midbrains (Jacobs
et al., 2007). The ablation of Lmx1b did not influence the
mRNA expression levels of Pitx3 itself (not shown) and also the
spatial expression of Pitx3 was not affected. In addition, spatial
expression of Ahd2 was partially recovered in Pitx3/Lmx1b
double mutants compared to Pitx3 mutants, suggesting that the
regulation of Ahd2 by Lmx1b is a PITX3-independent process. Frontiers in Molecular Neuroscience | www.frontiersin.org DISCUSSION Interestingly, the Ahd2 promoter contains several conserved
FLAT-elements, to which LMX1B can bind directly (Rascle et al.,
2009), suggesting that LMX1B might directly influence Ahd2
expression. AHD2 is involved in the generation of retinoic acid
(RA) from retinol and Jacobs et al. (2007) demonstrated that
administration of RA to the maternal diet of Pitx3−/−animals
led to an increase in TH+ cells during development and an
increased innervation of the dorsal Striatum. Although we only
observed a partial rescue of Ahd2 expression, we did observe
an increase in TH+ cells in the midbrain of adult Pitx3CreCre;
Lmx1b L/L animals. Overall, our data shows that Lmx1b is important for the
RNA
i
l
l
f
i
l f
E 1
d E 2 d
i Ahd2 independent of Pitx3. This underscores the presence of
developmental programs that lead to mdDA subsets and that
Lmx1b is part of the complex network of interacting transcription
factors that specify these subsets. we found that a loss of Lmx1b in postmitotic neurons leads
to reduced mRNA levels of En1 and En2 at E14.5. Both of
these factors have be associated with the survival of mdDA
neurons (Simon et al., 2001; Albéri et al., 2004) and En1 ±
animals show a progressive loss of TH+ cells in the ventral
midbrain from 3 weeks of age onward (Sonnier et al., 2007). The lower mRNA levels of En1, if equally represented by a lower
protein level, might contribute to the observed reduction in the
amount of TH+ cells in 3-month-old Pitx3Cre/+; Lmx1b L/L
animals, however in heterozygous En1 mice the SNc is mostly
affected, while in our model initially the VTA demonstrates the
largest percentile reduction in TH+ cells. Notably, after 1 year
the defect resides in the SNc suggesting a change in timing
of neuronal survival which culminates in loss of SNc neurons
after 12 months, which is in line with observations done in
En1 mutant animals. In addition to a role in neuronal survival
we also found a role for Lmx1b in subset specification. Pitx3
driven deletion of Lmx1b led to a significant increase in the
rostrolateral mark Ahd2 in vivo and in vitro. In E14.5 embryos,
an extension of the Ahd2 positive domain was observed, as
Ahd2 expression was found ectopically in medial sections of
the ventral mesencephalon, overlapping with the Cck positive
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Neurosci. SUPPLEMENTARY MATERIAL TABLE S1 | Absolute numbers of cells counted. (A) The actual amount of TH+
cells counted in total and the SNc and VTA in 3-month-old wildtype and Lmx1b SUPPLEMENTARY MATERIAL The Supplementary Material for this article can be found
online
at:
https://www.frontiersin.org/articles/10.3389/fnmol. 2019.00062/full#supplementary-material FIGURE S1 | Lmx1b mostly regulates genes associated with head and nervous
system development. PANTHER over-representation tests show that genetic
ablation of Lmx1b mainly affects genes associated with head and nervous system
development at E14.5. FIGURE S1 | Lmx1b mostly regulates genes associated with head and nervous
system development. PANTHER over-representation tests show that genetic
ablation of Lmx1b mainly affects genes associated with head and nervous system
development at E14.5. FIGURE S2 | Over-expression of Lmx1b reduces Ahd2 expression in MN9D
cells. Ahd2 and Th mRNA levels were examined in MN9D cells over-expressing
Lmx1b. A loss of 48% in Ahd2 mRNA levels was observed in cells
over-expressing Lmx1b (n = 4. ∗∗P < 0.01, two-tailed), while mRNA levels of Th
were not significantly altered (N = 4). Overall, our data shows that Lmx1b is important for the
mRNA expression level of survival factors En1 and En2 during
development and for the survival of a specific group of
mdDA neurons postnatally. Next to a role in survival we
found that Lmx1b functions as a transcriptional repressor of FIGURE S3 | No differences could be detected in the expression pattern
of Ahd2 between Pitx3Cre/+; Lmx1b+/+ 3-month-old animals and Pitx3Cre/+;
Lmx1b L/L animals. Analysis of Ahd2 in coronal adult section in the Frontiers in Molecular Neuroscience | www.frontiersin.org March 2019 | Volume 12 | Article 62 14 Dopaminergic Subset Specifciation by Lmx1b Wever et al. cKO used for Figure 4. (B) The total amount of TH+cells counted in
12-month-old Pitx3Cre/+; Lmx1b +/+ and Pitx3Cre/+; Lmx1b L/L and subdivided
in the SNc and VTA, used for Figure 5. (C) Actual cell numbers used for the
graphs in Figure 9 in 3-month-old Lmx1b cKO and wildtypes. (D) Actual amount
of cells counted in the Pitx3Cre/Cre; Lmx1b +.+ and the Pitx3Cre/Cre; Lmx1b L/L
at 3 months of age and used for Figure 11. Pitx3Cre/+; Lmx1b L/L mutant via in situ hybridization. Expression of Ahd2
seems similar in Pitx3Cre/+; Lmx1b L/L animals compared to wildtype. Scale bar = 200 µM. in the SNc and VTA, used for Figure 5. (C) Actual cell numbers used for the
graphs in Figure 9 in 3-month-old Lmx1b cKO and wildtypes. (D) Actual amount
of cells counted in the Pitx3Cre/Cre; Lmx1b +.+ and the Pitx3Cre/Cre; Lmx1b L/L
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open-access article distributed under the terms of the Creative Commons Attribution
License (CC BY). The use, distribution or reproduction in other forums is permitted,
provided the original author(s) and the copyright owner(s) are credited and that the
original publication in this journal is cited, in accordance with accepted academic
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with these terms. Suleiman, H., Heudobler, D., Raschta, A.-S., Zhao, Y., Zhao, Q., Hertting, I., et al. (2007). The podocyte-specific inactivation of Lmx1b, Ldb1 and E2a yields new
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et al. (2019). Single-cell RNA sequencing reveals midbrain dopamine neuron March 2019 | Volume 12 | Article 62 Frontiers in Molecular Neuroscience | www.frontiersin.org 16
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Analisis Efisiensi Inverter pada Grid-Connected 50 KWp Unpam Viktor
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Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 Analisis Efisiensi Inverter pada Grid-Connected
50 KWp Unpam Viktor Keywords: Efficiency; Grid-Tied Inverter; Pamulang University; Isolarcloud Monitoring Keywords: Efficiency; Grid-Tied Inverter; Pamulang University; Isolarcloud Monitoring Copyright © 2022 Woro Agus Nurtiyanto, Page 348
This Journal is licensed under a Creative Commons Attribution 4.0 International License Analisis Efisiensi Inverter pada Grid-Connected
50 KWp Unpam Viktor Woro Agus Nurtiyanto1*, Perani Rosyani1, Lili Solihin2, Wiji Prayogo2
1Fakultas Teknik, Program Studi Teknik Elektro,Universitas Pamulang, Tangerang Selatan, Indonesia
2Fakultas Teknik, Program Studi Teknik Informatika, Universitas Pamulang, Tangerang Selatan, Indonesia
Email: 1*dosen00855@unpam.ac.id , 2dosen837@unpam.ac.id , 3dosen00860@unpam.ac.id , 4wijiprayogo979@gmail.com
Submitted: 18/08/2022; Accepted: 30/08/2022; Published: 30/08/2022 Abstrak−Efisiensi Grid-tied Inverter dalam mengkonversi energi listrik yang bersumber dari Pembangkit Listrik Tenaga
Surya. Tujuan dari penelitian ini untuk mengetahui seberapa besar efisiensi Grid-tied inverter berdasarkan hasil pengujian di
lapangan, serta bermanfaat dijadikan referensi untuk mengetahui unjuk kerja dari sebuah inverter yang terhubung ke jaringan
utilitas. Metode penelitian deskriptif kuantitatif dengan teknik pengukuran langsung menggunakan alat ukur Seaward PV meter
dan monitoring aplikasi IsolarCloud app dan panel kontrol untuk mengetahui daya output yang diserap oleh beban dengan
lokasi penelitian di ruang inverter lantai 7 kampus Universitas Pamulang Viktor. Hasil penelitian menunjukan bahwa efisiensi
tertinggi tercapai dengan nilai 98,4% di mana dapat dikatakan inverter dapat mengkonversi hampir seluruh energi yang
dihasilkan solar sel, dengan rata-rata arus short circuit (Isc) 16% lebih besar dari arus maximal power (Imp) dan tegangan open
circuit (Voc) lebih besar 21% dari tegangan maximal power. Hasil analisis juga menunjukan bahwa kenaikan arus berpengaruh
terhadap efisiensi inverter walaupun tidak signifikan. Kata Kunci: Efisiensi; Grid-Tied Inverter; Universitas Pamulang; IsolarCloud app Abstract−The efficiency of grid-tied inverters in converting electrical energy sourced from solar power plants. The purpose of
this study is to find out how efficient the Grid-tied inverter is based on the results of field tests, and it is useful as a reference
to determine the performance of an inverter connected to the utility grid. Quantitative research methods with direct
measurement techniques use the Seaward PV meter measuring instrument and the IsolarCloud monitoring application and
control panel to determine the output power absorbed by the load with the research location in the inverter room on the 7th
floor of the Pamulang Viktor University campus. The results show that the highest efficiency is achieved with a value of 98.4%
where it can be said that the inverter can convert almost all of the energy produced by solar cells, with an average short circuit
current (Isc) 16% greater than the maximum current (Imp) and open circuit voltage (Voc) is 21% greater than the maximum
power voltage. The results of the analysis also show that the increase in current has no significant effect on the efficiency of
the inverter. 1. PENDAHULUAN Dalam dokumen Kebijakan Energi Nasional (KEN) Indonesia, yang mana dokumen tersebut mengatur tentang
bidang energi sebagai panduan dengan tujuan untuk mencapai kedaulatan dan ketahanan energi yang bermanfaat
untuk mendukung pembangunan nasional. Salah satu maksud dan tujuan dari KEN yaitu bauran renewable energy
atau EBT nasional dengan persentase minimal sebesar 23% pada tahun 2025 dan meningkat 8% menjadi 31% pada
5 tahun berikutnya dari keseluruhan total konsumsi energi nasional. Namun, kenyataan tersebut masih jauh dari
kata terealisasikan, pasalnya bauran EBT Indonesia pada tahun 2021 masih sangat tergolong rendah, menurut
menteri ESDM Arifin Tasrif bauran EBT pada tahun 2021 tercatat hanya berkisar sebesar 11,2% (Pribadi, 2021;
Putu et al., 2020). Atas dasar itulah universitas pamulang membangun PLTS grid-connected 50 KWp sebagai
bahan pembelajaran dan usaha untuk mempercepat target yang telah ditetapkan. Solar Power plant atau yang biasa disebut (PLTS) merupakan suatu sistem yang mampu mengkonversi
energi cahaya dari matahari menjadi sumber energi listrik, dalam hal ini energi yang diterima oleh photovoltaic
atau solar cell adalah dalam bentuk cahaya atau radiasi untuk menghasilkan listrik arus searah (DC). Inverter
merupakan salah satu subsistem penting di mana perangkat inverter dapat mengubah arus DC (Direct Current)
yang sebelumnya dihasilkan oleh solar cell menjadi arus AC untuk menyuplai beban AC atau jika sistem tersebut
terhubung dengan jaringan PLN yang biasa disebut On-Grid atau Grid-connected (Malik, 2018; Wijaya, 2012). Inverter merupakan komponen paling kompleks dalam sistem PLTS, secara umum bagian utama dari
sebuah inverter adalah DC link bus, kontroler, sistem pengaman dan yang tidak kalah penting adalah
semikonduktor daya sebagai sakelar, di mana komponen elektronika daya yang digunakan dapat berbentuk SCR,
Transistor dan Mosfet yang beroperasi sebagai sakelar dan pengubah (Malik, 2018). Grid-tie inverter adalah salah satu jenis dari inverter yang digunakan untuk menghubungkan PLTS dengan
jaringan listrik, Inverter jenis ini akan secara otomatis mensinkronkan energi yang dihasilkan oleh sel surya dengan
tegangan jala-jala jaringan listrik (Grid) PLN, GTI juga didesain untuk otomatis berhenti menyuplai daya ke
jaringan jika tidak mendeteksi tegangan pada jaringan PLN, hal inilah yang menjadi keunggulan GTI dibanding
dengan jenis inverter yang lainnya (Nahela, 2019). Pada paper kali ini fokus yang dilakukan adalah menganalisa efisiensi grid tie inverter dengan kapasitas 44
KVA dengan sumber PLTS yang terhubung ke dalam jaringan listrik (Grid Connected) PLN. 1. PENDAHULUAN Metode yang
digunakan untuk mendapat parameter efisiensi adalah dengan cara monitoring menggunakan aplikasi Isolarcloud, Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 panel kontrol dan juga pengukuran langsung menggunakan PV meter Seaward dan clamp meter. dengan begitu
analisa dapat dilakukan dengan mendapat nilai input tegangan dan arus dan nilai output tegangan dan arus. Di sisi
lain akan dianalisa juga bagiamana pengaruh naik-turunnya arus terhadap efisiensi 2.1. kerangka penelitian Proses pengujian GTI melibatkan beberapa alat dan bahan yang telah dibahas sebelumnya, pada proses
“ANALISA EFISIENSI INVERTER PADA GRID- CONNECTED 50 KWP UNPAM” terdapat beberapa tahapan
yang harus dilakukan agar penelitian berjalan sesuai dengan rencana. Tahapan penelitian tersebut dimuat dalam
gambar 1. yang berbentuk diagram alir atau flowchart. Gambar 1. Diagram Flowchart (Kerangka pemikiran) Gambar 1. Diagram Flowchart (Kerangka pemikiran) Gambar 1. Diagram flowchart. Berisi penjelasan tentang tahapan penelitian yang menggambarkan urutan
logis untuk mendapatkan hasil penelitian sesuai dengan harapan dan gambaran sistem. Jika ada gambar dan tabel,
itu harus disajikan dengan nama tabel dan gambar yang disertai dengan nomor urut. 2.2. Alat dan Bahan Alat dan bahan sebagai penunjang dalam berjalannya analisa efisiensi inverter. Alat dan bahan sebagai penunjang dalam berjalannya analisa efisiensi inverter. 2.2.1. Alat 7) Perangkat lunak autoCAD 2020, Micosoft Office 2019 yang digunakan sebagai pelengkap dalam menyusun
tugas akhir. 7) Perangkat lunak autoCAD 2020, Micosoft Office 2019 yang digunakan sebagai pelengkap dalam menyusun
tugas akhir. 2.2.1. Alat 4) Seaward Clamp meter, perangkat ini dapat digunakan untuk mengukur arus input DC maupun arus output AC
dengan batas maksimal 300 A. untuk cara penggunaannya harus dikombinasikan dengan Seaward PV meter
200, cara pengukuran cukup dengan Switch on sakelar ke range arus yang paling besar (untuk pencegahan bila
arus yang diukur lebih besar), kemudian kaitkan clamp meter dengan salah satu kabel, untuk DC bisa salah satu
kabel +/- dan untuk AC bisa salah satu kabel Fasa/Netral yang dapat di lihat pada gambar 5. Gambar 4. Seaward PV meter. Gambar 4. Seaward PV meter. 4) Seaward Clamp meter, perangkat ini dapat digunakan untuk mengukur arus input DC maupun arus output AC
dengan batas maksimal 300 A. untuk cara penggunaannya harus dikombinasikan dengan Seaward PV meter
200, cara pengukuran cukup dengan Switch on sakelar ke range arus yang paling besar (untuk pencegahan bila
arus yang diukur lebih besar), kemudian kaitkan clamp meter dengan salah satu kabel, untuk DC bisa salah satu
kabel +/- dan untuk AC bisa salah satu kabel Fasa/Netral yang dapat di lihat pada gambar 5. 4) Seaward Clamp meter, perangkat ini dapat digunakan untuk mengukur arus input DC maupun arus output AC
dengan batas maksimal 300 A. untuk cara penggunaannya harus dikombinasikan dengan Seaward PV meter
200, cara pengukuran cukup dengan Switch on sakelar ke range arus yang paling besar (untuk pencegahan bila
arus yang diukur lebih besar), kemudian kaitkan clamp meter dengan salah satu kabel, untuk DC bisa salah satu
kabel +/- dan untuk AC bisa salah satu kabel Fasa/Netral yang dapat di lihat pada gambar 5. Gambar 5. Seaward Clamp Meter. Gambar 5. Seaward Clamp Meter. 5) Perangkat lunak yang digunakan untuk monitoring daya pada GTI yaitu IsolarCloud APP. Aplikasi ini dapat
menampilkan data-data seperti daya yang dihasilkan oleh inverter, alarm peringatan, total energi yang
dihasilkan perhari, dan informasi nilai arus dan tegangan pada setiap string. Untuk login cukup dengan scan
barcode yang tersedia pada sisi samping inverter dan verifikasi dengan menggunakan Id: user Password:
88888888. Untuk menampilkan nilai tegangan dan arus tiap string dapat menekan pada tab Run information
dapat dilihat pada gambar 6. Gambar 6. Tampilan awal IsolarCloud APP. Gambar 6. Tampilan awal IsolarCloud APP. Gambar 6. Tampilan awal IsolarCloud APP. 6) Perangkat lunak untuk membuat skema rangkaian listrik yaitu proteus 8. 6) Perangkat lunak untuk membuat skema rangkaian listrik yaitu proteus 8. 2.2.1. Alat Alat yang digunakan sebagai penunjang dalam penelitian kali ini adalah: Alat yang digunakan sebagai penunjang dalam penelitian kali ini adalah:
1) Grid-tied inverter, GTI merupakan Instrumen utama dalam penelitian ini yang berfungsi sebagai pemasok
daya AC pada jaringan (grid), inverter yang digunakan yaitu inverter SUNGROW SG40CX. 1) Grid-tied inverter, GTI merupakan Instrumen utama dalam penelitian ini yang berfungsi sebagai pemasok
daya AC pada jaringan (grid), inverter yang digunakan yaitu inverter SUNGROW SG40CX. Gambar 2. Circuit Diagram GTI SUNGROW SG40CX. Gambar 2. Circuit Diagram GTI SUNGROW SG40CX. 2) Panel Kontrol, digunakan untuk memonitoring nilai arus dan tegangan yang dipakai ke baban dalam kata lain
output inverter. Nilai tegangan dan arus ini yang selanjutnya digunakan sebagai bahan penelitian. Gambar 3. Panel Kontrol untuk Monitoring. Gambar 3. Panel Kontrol untuk Monitoring. Copyright © 2022 Woro Agus Nurtiyanto, Page 349 Copyright © 2022 Woro Agus Nurtiyanto, Page 349
This Journal is licensed under a Creative Commons Attribution 4.0 International License py g
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This Journal is licensed under a Creative Commons Attribution 4.0 International License Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 3) PV meter Seaward, digunakan untuk mengukur tegangan dan arus DC pada input inverter. Untuk menyalakan
cukup dengan menekan tombol Riso dan Mode secara bersamaan, kemudian gunakan probe MC4 untuk
mengukur arus dan tegangan, untuk mengetahui nilai arus dan tegangan tekan tombol AUTO, untuk mematikan
perangkat tekan kembali tombol Riso dan Mode secara bersamaan selama beberapa detik sampai display
dalam keadaan off. PV meter Seaward 200 juga dapat digunakan untuk mengukur arus AC pada output dengan
cara dikombinasikan dengan Clamp meter Seaward. Clamp meter seaward ditunjukan pada gambar 4. 3) PV meter Seaward, digunakan untuk mengukur tegangan dan arus DC pada input inverter. Untuk menyalakan
cukup dengan menekan tombol Riso dan Mode secara bersamaan, kemudian gunakan probe MC4 untuk
mengukur arus dan tegangan, untuk mengetahui nilai arus dan tegangan tekan tombol AUTO, untuk mematikan
perangkat tekan kembali tombol Riso dan Mode secara bersamaan selama beberapa detik sampai display
dalam keadaan off. PV meter Seaward 200 juga dapat digunakan untuk mengukur arus AC pada output dengan
cara dikombinasikan dengan Clamp meter Seaward. Clamp meter seaward ditunjukan pada gambar 4. Gambar 4. Seaward PV meter. Gambar 4. Seaward PV meter. 3.1. Pengujian Menggunakan Alat Ukur dan Monitoring Pada bagian ini akan dibahas perbandingan antara tegangan open circuit (Voc) dan arus short circuit dengan
tegangan maximal power (Vmp) dan arus maximal power (Imp), dengan menggunakan alat-alat yang telah
disebutkan sebelumnya pada BAB III. Pada sub bab ini akan dilakukan perbandingan antara pengukuran langsung Copyright © 2022 Woro Agus Nurtiyanto, Page 350
This Journal is licensed under a Creative Commons Attribution 4.0 International License Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 dengan monitoring. Pengukuran langsung dilakukan dengan cara memutus input pada GTI, yang didapat dari
pengukuran langsung adalah arus dan tegangan input tanpa beban (Voc dan Isc) di mana Voc dan Isc lebih besar
dari Vmp dan Imp. Hasil pengukuran dapat di lihat pada table 1. Tabel 1. Pengukuran arus dan tegangan input Tabel 1. Pengukuran arus dan tegangan input Tabel 1. Pengukuran arus dan tegangan input Tabel 1. Pengukuran arus dan tegangan input
Nilai arus dan tegangan input dari Pengukuran Langsung
No. String
Tegangan (Voc)
Arus (Isc)
String 1
876 V
2,74 A
String 2
874 V
2,71 A
String 3
874 V
2,72 A
String 4
873 V
2,66 A
String 5
873 V
2,73 A
String 6
873 V
2,69 A
Nilai arus dan tegangan input dari monitoring
No. String
Tegangan (Vmp)
Arus (Imp)
String 1
738,8 V
2,55 A
String 2
738,8 V
2,42 A
String 3
755,5 V
2,54 A
String 4
755,5 V
2,39 A
String 5
757,6 V
2,31 A
String 6
757,6 V
2,58 A Nilai arus dan tegangan input dari Pengukuran Langsung Pada hasil pengujian tabel 1. 3.1. Pengujian Menggunakan Alat Ukur dan Monitoring menunjukan bahwa Tegangan dan Arus yang terukur menggunakan alat ukur
adalah tegangan open circuit (Voc) dan arus short circuit sedangkan arus dan tegangan yang terukur pada
monitoring IsolarCloud APP adalah tegangan maximal power dan arus maximal power (Vmp dan Imp) pada tabel
4.4 pengambilan data tiap-tiap string dengan monitoring aplikasi terdapat jeda waktu sekitar kurang lebih 10 menit,
pada tegangan Voc dan Vmp string 1 terdapat perbedaan sebesar 137,2 V atau kenaikan pada Voc sebesar 18,6%
kemudian pada string 2 terdapat perbedaan 135,3 V atau kenaikan pada Voc sebesar 18,3%, string 3 terdapat
perbedaan 118,5 V atau kenaikan pada Voc sebesar 15,7%, string 4 terdapat perbedaan sebesar 117,5 V atau
kenaikan pada Voc sebesar 15,5%, string 5 terdapat selisih 115,4 V atau kenaikan pada Voc sebesar 15,2%, string
6 terdapat selisih 115,4 V atau kenaikan pada Voc sebesar 15,2%. tegangan open circuit string 1 s/d string 6 terdapat
perbedaan di mana Voc lebih besar dengan selisih rata-rata sekitar 123,2 V atau dengan kenaikan rata-rata Voc
sebesar 16,4% terhadap Vmp. hal ini membuktikan bahwa Voc > Vmp. Sedangkan untuk arus short circuit string
1 s/d 6 terdapat perbedaan dengan selisih rata-rata sekitar 0,24 A atau Isc > Imp sebesar sekitar 16,4%. Copyright © 2022 Woro Agus Nurtiyanto, Page 351 This Journal is licensed under a Creative Commons Attribution 4.0 International License 3.2. Analisis Efisiensi GTI 4, August 2022, Page 348−355
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DOI 10.47065/josyc.v3i4.2134
Jam 12:00
DC input (V.A)
AC output (V.A)
MPPT 1
713,3 V × 15,9 A = 11 341
L1
234,7 V × 48,35 A = 11 347,7
MPPT 2
711,1 V × 16,7 A = 11 876
L2
237,3 V × 48,04 A = 11 400
MPPT 3
720,8 V × 16,6 A = 11 965
L3
233,3 V × 48,43 A = 11 299
Total
35 182,2
Total
34 064,44
𝜂 = (34 064,44)/(35 182,2)× 100% = 96,8%
Jam 13:00
DC input (V.A)
AC output (V.A)
MPPT 1
723,7 V × 15,3 A = 11 072
L1
231,2 V × 47,61 A = 11 007,4
MPPT 2
722,7 V × 15,8 A = 11 419
L2
233,4 V × 47,30 A = 11 040
MPPT 3
729,1 V × 15,7 A = 11 448
L3
230,0 V × 47,72 A = 10 975,6
Total
33 938,3
Total
33 023
𝜂 =
(33.023)/(33.938,3)× 100% = 97,3%
Jam 14:00
DC input (V.A)
AC output (V.A)
MPPT 1
749,3 V × 11,5 A = 8 617
L1
228,2 V × 38,04 A = 8 680,7
MPPT 2
748,6 V × 12,1 A = 9 058
L2
230,8 V × 37,78 A = 8 719,6
MPPT 3
756,8 V × 11,8 A = 8 930
L3
227,0 V × 38,19 A = 8 669,1
Total
26 605,3
Total
26 070
𝜂 =
(26.070)/(26.605,3)× 100% = 97,9%
Gambar 7. Grafik Efisiensi Terhadap Waktu. Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 Journal of Computer System and Informatics (JoSYC) Gambar 7. Grafik Efisiensi Terhadap Waktu. Gambar 7. Grafik Efisiensi Terhadap Waktu. Hasil data pada gambar 7. diketahui bahwa efisiensi tertinggi terjadi pada pukul 14:00 WIB dengan nilai
efisiensi sebesar 97,9%, sedangkan efisiensi terendah terjadi pada pukul 12:00 yaitu sebesar 96,8%, terjadi
penurunan sebesar 0,06% dari pukul 09:00 ke pukul 10:00 WIB, setelahnya terjadi penurunan sebesar 0,3% dari
pukul 10:00 ke pukul 11:00, kenaikan terjadi dari pukul 11:00 ke pukul 12:00 sebesar 0,2% dari 97% menjadi
97,2%, kemudian naik sebesar 0,1% pada jam 13:00 menjadi 97,3% dan naik sebesar 0,6% menjadi 97,9% pada
pukul 14:00 WIB. Gambar 4.18 menunjukan kurva pengaruh arus terhadap efisiensi. Gambar 8. Grafik Pengaruh Arus Terhadap Efisiensi. Gambar 8. Grafik Pengaruh Arus Terhadap Efisiensi. 3.2. Analisis Efisiensi GTI Efisiensi maksimal dari GTI adalah sebesar 98,7%. Berdasarkan hasil pengujian dengan rentang waktu mulai dari
jam 09:00 s/d jam 14:00 WIB dengan beban yang bervariasi. Nilai efisiensi dihitung dengan hasil yang didapat
adalah sebagai berikut: Tabel 2. Analisis Efisiensi GTI. Copyright © 2022 Woro Agus Nurtiyanto, Page 351
This Journal is licensed under a Creative Commons Attribution 4.0 International License
Tabel 2. Analisis Efisiensi GTI. Jam 09:00
DC input (V.A)
AC output (V.A)
MPPT 1
737,6 V × 12,4 A = 9 146,2
L1
232,4 V × 38,88 A = 9 036
MPPT 2
740,4 V × 12,7 A = 9 403,1
L2
234,9 V × 38,63 A = 9 074,2
MPPT 3
745,0 V × 12,5 A = 9 312,5
L3
231,4 V × 38,97 A = 9 017,6
Total
27.861,82
Total
27.127,55
𝜂 = (27 127,55)/(27 861,82)X 100% = 97,36%
Jam 10:00
DC input (V.A)
AC output (V.A)
MPPT 1
719,2 V × 15,4 A = 11 076
L1
231,2 V × 47,77 A = 11 044,4
MPPT 2
722,8 V × 15,9 A = 11 492
L2
233,2 V × 47,45 A = 11 065,5
MPPT 3
721,3 V × 15,9 A = 11 569
L3
229,5 V × 47,89 A = 10 990,7
Total
34.037
Total
33.100,5
𝜂 = (33.100,5)/(34.037)× 100% = 97,3%
Jam 11:00
DC input (V.A)
AC output (V.A)
MPPT 1
717,2 V × 16,0 A = 11 475
L1
229,6 V × 49,39 A = 11 340
MPPT 2
700,0 V × 16,6 A = 11 620
L2
231,6 V × 49,07 A = 11 364,6
MPPT 3
742,8 V × 16,1 A = 11 960
L3
228,3 V × 49,51 A = 11 303,1
Total
35 054,3
Total
34 008
𝜂 =
(34.008)/(35 054,3)× 100% = 97% Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. py g
©
g
y
,
g
This Journal is licensed under a Creative Commons Attribution 4.0 International Licen Copyright © 2022 Woro Agus Nurtiyanto, Page 352
This Journal is licensed under a Creative Commons Attribution 4.0 International License 3.2. Analisis Efisiensi GTI 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134
Jam 12:00
DC input (V.A)
AC output (V.A)
MPPT 1
713,3 V × 15,9 A = 11 341
L1
234,7 V × 48,35 A = 11 347,7
MPPT 2
711,1 V × 16,7 A = 11 876
L2
237,3 V × 48,04 A = 11 400
MPPT 3
720,8 V × 16,6 A = 11 965
L3
233,3 V × 48,43 A = 11 299
Total
35 182,2
Total
34 064,44
𝜂 = (34 064,44)/(35 182,2)× 100% = 96,8%
Jam 13:00
DC input (V.A)
AC output (V.A)
MPPT 1
723,7 V × 15,3 A = 11 072
L1
231,2 V × 47,61 A = 11 007,4
MPPT 2
722,7 V × 15,8 A = 11 419
L2
233,4 V × 47,30 A = 11 040
MPPT 3
729,1 V × 15,7 A = 11 448
L3
230,0 V × 47,72 A = 10 975,6
Total
33 938,3
Total
33 023
𝜂 =
(33.023)/(33.938,3)× 100% = 97,3%
Jam 14:00
DC input (V.A)
AC output (V.A)
MPPT 1
749,3 V × 11,5 A = 8 617
L1
228,2 V × 38,04 A = 8 680,7
MPPT 2
748,6 V × 12,1 A = 9 058
L2
230,8 V × 37,78 A = 8 719,6
MPPT 3
756,8 V × 11,8 A = 8 930
L3
227,0 V × 38,19 A = 8 669,1
Total
26 605,3
Total
26 070
𝜂 =
(26.070)/(26.605,3)× 100% = 97,9% ournal of Computer System and Informatics (JoSYC)
SSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 3.2. Analisis Efisiensi GTI Pada gambar 8. terlihat bahwa baik arus input maupun output terlihat tidak stabil, arus input maksimum
pada pengujian adalah sebesar 49,2 Ampere terjadi pada pukul 12.00 WIB dan output maksimum sebesar 147,97
Ampere terjadi pada pukul 11.00 WIB, sedangkan arus minimum terjadi pada pukul 14.00 WIB di mana arus input
sebesar 35,4 Ampere dan output sebesar 114,01 Ampere di waktu yang sama. Dilihat pada gambar 8. dapat Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
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DOI 10.47065/josyc.v3i4.2134 disimpulkan bahwa arus memiliki pengaruh terhadap efisiensi, pada saat pukul 09.00 arus output terukur sebesar
116,48 Ampere dengan efisiensi sebesar 97,36%, satu jam kemudian arus output naik menjadi 143,11 Ampere
dengan efisiensi 97,3% dan naik menjadi 147,97 Ampere pada satu jam berikutnya dengan efisiensi 97%, saat
pukul 12.00 WIB terjadi penurunan sebesar 3,15 Ampere menjadi 144,82 Ampere dengan efisiensi sebesar 97,2%
lalu satu jam kemudian turun menjadi 142,63 Ampere pada pukul 13.00 WIB dengan efisiensi 97,3% dan pada
pukul 14.00 arus output turun menjadi 114,01 Ampere dengan efisiensi sebesar 97,6% dari grafik dan penjelasan
yang telah disampaikan pada pengujian dapat disimpulkan bahwa kenaikan arus mempengaruhi efisiensi inverter,
di mana saat arus naik maka efisiensi akan turun sebaliknya ketika arus turun maka efisiensi inverter akan naik. Copyright © 2022 Woro Agus Nurtiyanto, Page 353
This Journal is licensed under a Creative Commons Attribution 4.0 International License 3.3. Pengujian Fitur Anti-Islanding pada GTI Anti-Islanding seperti yang telah diuraikan pada paragraph sebelumnya yang merupakan sebuah fitur yang sangat
berguna bagi inverter jenis on-grid agar dapat memutus suplai daya listrik ke jaringan ketika GTI tidak mendeteksi
tegangan pada jaringan utilitas. Kali ini penulis menguji fitur anti-islanding dengan GTI dalam keadaan sedang
bekerja dalam artian memasok daya ke jaringan utilitas, pengujian dilakukan dengan cara memutus aliran arus yang
terkoneksi ke beban pada panel kontrol. Gambar 9. Panel Kontrol Output Inverter. Gambar 9. Panel Kontrol Output Inverter. Pada gambar 9. terdapat 3 MCCB di mana saat melakukan pengujian fitur anti-islanding tersebut, MCCB
yang terhubung ke jaringan diputus sambungannya, dengan cara menurunkan tuas MCCB tersebut, setelah itu GTI
akan secara otomatis memutus suplai daya listrik walaupun input PV masih memproduksi Daya listrik. Dalam
kondisi ini inverter akan tetap dalam kondisi off selama tidak mendeteksi tegangan atau frekuensi pada jaringan
utilitas. Pada penunjukan ketika GTI sedang dalam kondisi islanding maka led pada bagian depan GTI akan berubah
menjadi warna merah yang ketika normal adalah berwanra biru. Sedangkan pada tampilan IsolarCloud APP setelah
terjadi islanding di mana akan ada fault alarm dengan notifikasi grid power outage atau yang artinya pemadaman
jaringan utilitas, saat jaringan utilitas kembali bertegangan maka GTI akan secara otomatis kembali menyuplai
daya listrik ke jaringan utilitas. Gambar 10. GTI yang sedang dalam kondisi Islanding. Pada gambar 10. terlihat bahwa LED notifikasi yang berada pada sisi depan inverter berwarna merah yang
mengindikasikan bahwa sedang terjadi fault atau kegagalan yang mana GTI tidak mendeteksi tegangan ataupun
frekuensi dari jaringan utilitas, sehingga GTI berhenti menyuplai daya ke jaringan. Gambar 10. GTI yang sedang dalam kondisi Islanding. Gambar 10. GTI yang sedang dalam kondisi Islanding. mbar 10 terlihat bahwa LED notifikasi yang berada pada sisi depaninverter berwarna merah Gambar 10. GTI yang sedang dalam kondisi Islanding. Pada gambar 10. terlihat bahwa LED notifikasi yang berada pada sisi depan inverter berwarna merah yang
mengindikasikan bahwa sedang terjadi fault atau kegagalan yang mana GTI tidak mendeteksi tegangan ataupun
frekuensi dari jaringan utilitas, sehingga GTI berhenti menyuplai daya ke jaringan. Copyright © 2022 Woro Agus Nurtiyanto, Page 353
This Journal is licensed under a Creative Commons Attribution 4.0 International License Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
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DOI 10.47065/josyc.v3i4.2134
Gambar 11. 3.3. Pengujian Fitur Anti-Islanding pada GTI Tampilan dasar IsolarCloud APP ketika terjadi Islanding Journal of Computer System and Informatics (JoSYC)
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DOI 10.47065/josyc.v3i4.2134 al of Computer System and Informatics (JoSYC) Gambar 11. Tampilan dasar IsolarCloud APP ketika terjadi Islanding Sedangkan tampilan pada aplikasi ditampilkan pada gambar 11. di mana ketika terjadi islanding tampilan
awal aplikasi akan menampilkan fault alarm yang mengindikasikan bahwa GTI tidak mendeteksi tegangan di
jaringan utilitas. 4. KESIMPULAN Berdasarkan pengujian berasar efisiensi di lapangan didapat Besar nilai efisiensi maksimal yang terukur dari hasil
pengujian secara langsung yaitu 98,4% yang terjadi pada pengujian pada pukul 14:00 WIB, hasil tersebut berbeda
dengan yang tertulis di manual book Inverter SUNGROW SG40CX di mana spesifikasi tertulis efisiensi maksimal
sebesar 98,7%. Nilai efisiensi rata-rata dari hasil pengujian adalah sebesar 96,9%. Serta terbukti bahwa kenaikan
arus berpengaruh terhadap nilai efisiensi tersebut, di mana ketika arus baik output maupun input naik maka
efisiensi GTI akan turun. Pada pengukuran arus input menggunakan alat ukur dengan kurun waktu pukul 09:00
s/d 14:00 WIB, input arus short circuit (Isc) lebih besar dari arus maximal power (Imp) dengan nilai rata-rata
sebesar Isc 16% > Imp. Pada pengukuran tegangan input meggunakan alat ukur dengan kurun waktu pukul 09:00
s/d 14:00 WIB, tegangan open circuit (Voc) lebih besar dari tegangan maximal power (Vmp) dengan nilai rata-
rata sebesar Voc 21% > Vmp. Salah satu fitur GTI yaitu anti-islanding, yang terdapat pada inverter
SUNGROW SG40CX saat pengujian terbukti efektif ketika dilakukan simulasi islanding, Inverter otomatis
memutus suplai daya dengan selang waktu beberapa detik ketika tidak mendeteksi tegangan pada jaringan utilitas. REFERENCES [1]
Alfayet Umar, R., Adam, K. B., & Aprillia, B. S. (2021). PERANCANGAN INVERTER TENAGA SURYA ON GRID
SATU FASA. [2]
China Power Supply Society, IEEE Power Electronics Society, Institute of Electrical and Electronics Engineers. Nanjing
Section, & Institute of Electrical and Electronics Engineers. (2014). Anti-islanding Modeling of Grid-tied Inverters. https://ieeexplore.ieee.org/document/7037933 p
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Hidayat, F., & Krismadinata, K. (2019). Rancang Bangun VVVF Inverter 3 Fasa untuk Operasi Motor Induksi Tiga Fasa
dengan Antarmuka Komputer. INVOTEK: Jurnal Inovasi Vokasional Dan Teknologi, 19(2), 47–56. https://doi.org/10.24036/invotek.v19i2.609 [4]
Malik, A. (2018). ANALISIS RANGKAIAN INVERTER 12V DC-220V AC DENGAN SUMBER PANEL SURYA
PADA BEBAN MOTOR LISTRIK SATU FASA. [5]
Nahela, S. (2019). 357586-analisa-unjuk-kerja-grid-tied-inverter-t-f0d9b692. ISSN 1858-1463. [6]
Narendiran, S., & School of Electrical Engineering, V. U. V. I. (2013). Grid Tie Inverter and MPPT-A Review [7]
Pribadi, A. (2021, December 14). Upaya Konkrit dan Terencana Capai Target Bauran 23% Di Tahun 2025. Https://Ebtke.Esdm.Go.Id/. [7]
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Putu, G., Nyoman, I., Kumara, S., & Setiawan, N. (2020). Grid Tie Inverter Untuk PLTS Atap Di Indonesia: Review
Standar Dan Inverter Yang Compliance Di Pasar Domestik UNJUK KERJA PEMBANGKIT LISTRIK TENAGA
SURYA ROOFTOP 158 KWP PADA KANTOR GUBERNUR BALI View project [8]
Putu, G., Nyoman, I., Kumara, S., & Setiawan, N. (2020). Grid Tie Inverter Untuk PLTS Atap Di Indonesia: Review
Standar Dan Inverter Yang Compliance Di Pasar Domestik UNJUK KERJA PEMBANGKIT LISTRIK TENAGA
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[10] Sampeallo, A. S., Galla, W. F., & Mbakurawang, F. (2021). ANALISIS KINERJA PLTS 25 KWP DI GEDUNG
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[11] BEBAN. Jurnal Media Elektro |, VII(1). [10] Sampeallo, A. S., Galla, W. F., & Mbakurawang, F. (2021). ANALISIS KINERJA PLTS 25 KWP DI GEDUNG
LABORATORIUM RISET TERPADU LAHAN KERING KEPULAUAN UNDANA TERHADAP VARIASI
[11] BEBAN. Jurnal Media Elektro |, VII(1). [11] BEBAN. Jurnal Media Elektro |, VII(1). |
[12] Sungrow. (2019a). SUNGROW SG40CX Manual Book. https://en.sungrowpower.com/ |
[12] Sungrow. (2019a). SUNGROW SG40CX Manual Book. https://en.sungrowpower.com/ [13] Sungrow. (2019b). SUNGROW SG40CX Spesification. [14] Telkom
University. (2021). Macam-macam
Komponen [14] Telkom
University. (2021). Macam-macam
Komponen
Elektronika. Copyright © 2022 Woro Agus Nurtiyanto, Page 354
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This Journal is licensed under a Creative Commons Attribution 4.0 International License REFERENCES [14] Telkom
University. (2021). Macam-macam
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Elektronika. [15] Https://Fit.Labs.Telkomuniversity.Ac.Id/. [15] Https://Fit.Labs.Telkomuniversity.Ac.Id/. [16] Vernandez, A. B. (2018). DETEKSI ISLANDING PADA PHOTOVOLTAIC TERHUBUNG GRID BERBASIS
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This Journal is licensed under a Creative Commons Attribution 4.0 International License Journal of Computer System and Informatics (JoSYC)
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Volume 3, No. 4, August 2022, Page 348−355
https://ejurnal.seminar-id.com/index.php/josyc
DOI 10.47065/josyc.v3i4.2134 Journal of Computer System and Informatics (JoSYC)
ISSN 2714-8912 (media online), ISSN 2714-7150 (media cetak)
Volume 3, No. 4, August 2022, Page 348−355
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[18] Yasin, M., Samman, F. A., & Sadjad, R. S. (2017). Desain dan Analisis Inverter Tiga Fasa untuk Aplikasi Sistem PLTS
Terhubung Grid PLN sebagai Referensi. In Jurnal Penelitian Enjiniring (Vol. 21, Issue 02). Fakultas Teknik Journal of Computer System and Informatics (JoSYC)
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Terhubung Grid PLN sebagai Referensi. In Jurnal Penelitian Enjiniring (Vol. 21, Issue 02). Fakultas Teknik [17] Wijaya, R. (2012). UNIVERSITAS INDONESIA ANALISIS KARAKTERISTIK GRID-TIE INVERTER SKRIPSI
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https://www.nature.com/articles/s41598-019-56409-8.pdf
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English
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Linking flow-stream variability to grain size distribution of suspended sediment from a satellite-based analysis of the Tiber River plume (Tyrrhenian Sea)
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Scientific reports
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cc-by
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Linking flow-stream variability to
grain size distribution of suspended
sediment from a satellite-based
analysis of the Tiber River plume
(Tyrrhenian Sea) OPEN J. Pitarch1,2, F. Falcini2*, W. Nardin3, V. E. Brando2, A. Di Cicco2 & S. Marullo2,4 Several coastal regions on Earth have been increasingly affected by intense, often catastrophic, flash
floods that deliver significant amounts of sediment along shorelines. One of the critical questions
related to the impact of these impulsive runoffs is “are flash floods more efficient in delivering non-
cohesive sandy sediment along the coasts?” Here we relate flow stages (i.e., from erratic to persistent)
to the grain size distribution of the suspended load, by performing a synergic analysis of in-situ river
discharge and satellite-retrieved grain size distribution, from 2002 to 2014, covering the 2012 Tiber
River (Italy) exceptional flood event. Our analysis shows novel and promising results regarding the
capability of remote sensing in characterizing suspended sediment in terms of grain size distribution
and reveals that erratic stages favour delivering of non-cohesive sandy sediment more than the
persistent stages. This conclusion is supported by numerical simulations and is consistent with previous
studies on suspended sediment rating curves. The hydrologic regime of the Tiber River basin (central Italy; Fig. 1a) plays an important role in sediment trans-
port and, in turn, in the evaluation of medium- and long-term changes of the Mediterranean coastline of central
Italy, called Laurentine Shore by the Romans1,2. The Tiber River is the second largest river in Italy, with a length
of 409 km and a drainage basin of about 17.156 km2; it is sourced from Central Apennines and debouches in the
Tyrrhenian Sea, close to the city of Rome. Mean annual discharge of the Tiber River is about 225 m3/s; maximum
annual discharge exceeds 1500 m3/s and the minimum annual discharge reaches 60 m3/s (ref. 3).h The Tiber River Delta is a progradating, wave dominated system, marked by a complex cusp made up of two
distributary channels (Fig. 1). The last progradation cycle (still ongoing) has particularly intensified over the
last 500 years and isolated the Roman Empire harbors from the sea2. Solid transport of the Tiber River is largely
given by suspended load and only a small fraction by bed load4. In particular, the percentage of sediment load
transported in suspension is more than 90% of the total amount of sediment load3,5. The average annual specific
sediment load, measured from 1934 to 1973 at Ripetta Bridge, located in downtown Rome (43 km upstream the
river mouth; Fig. www.nature.com/scientificreports www.nature.com/scientificreports www.nature.com/scientificreports Linking flow-stream variability to
grain size distribution of suspended
sediment from a satellite-based
analysis of the Tiber River plume
(Tyrrhenian Sea) 1), is about 290 ton/km2.l g
In November 2012, an exceptional flood brought the water discharge of the Tiber River to dangerous levels
and it lasted from 13 to 15 November 2012 (ref. 6) (Fig. 1). The flood wave was originated from the exceptional
river water flow due to the relevant precipitation in the upper part of the Tiber basin, which caused a significant
water level rising between the November 14th afternoon and 15th morning in the lower part of the basin (Fig. 1). Several rain gauges recorded an abundant precipitation, higher than 300 mm/48 hours, in the upper and west-
ern Tiber watershed (Fig. 1). In the following days, a flood event propagated along the Tiber River main stream
flow, reaching a peak in water discharge of 1,933 m3/s on November 15th at 01:00 am (local time) at the water
gauge station of Ripetta Bridge (Fig. 1). Floods like the one occurred in 2012 give us the opportunity to focus on 1NIOZ – Royal Netherlands Institute for Sea Research, Department of Coastal Systems, and Utrecht University,
PO Box 59, 1790 AB, Den Burg, Texel, The Netherlands. 2CNR – Institute of Marine Sciences (ISMAR), Via Fosso del
Cavaliere 100, 00133, Rome, Italy. 3Horn Point Laboratory, University of Maryland Center for Environmental Science,
Cambridge, MD, 21613, USA. 4ENEA – Centro Ricerche Frascati, Frascati, Italy. *email: federico.falcini@cnr.it Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ Figure 1. (a) MODIS Ocean Colour image on 17 November 2012 showing the suspended sediment plume;
the yellow box around the Tiber River mouth represents the area where reflectances (Rrs) were extracted; the
box shows the Tiber river delta evolution2. The image was created using data downloaded from the NASA
(oceancolor.gsfc.nasa.gov) and processed using SeaDAS 7.2 (ref. 55); (b) Qw and the total volume concentration
of suspended particles (VCtot) time series (plot generated by using MATLAB 7.1 http://uk.mathworks.com/
products/matlab); VCtot is obtained from the sum of the two volume concentrations of the grain fractions
considered in this work, i.e., 3.9 µm–62.5 µm and 62.5 µm – 125 µm. Figure 1. (a) MODIS Ocean Colour image on 17 November 2012 showing the suspended sediment plume;
the yellow box around the Tiber River mouth represents the area where reflectances (Rrs) were extracted; the
box shows the Tiber river delta evolution2. The image was created using data downloaded from the NASA
(oceancolor.gsfc.nasa.gov) and processed using SeaDAS 7.2 (ref. Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 In situ hydrological data. In situ hydrological data. Along the Tiber River, the water level is measured at several river flow gauges
combining water level and empirical river discharge. In this work we use Tiber River daily water discharge (Qw)
data at the historic Ripetta flow gauge, matching the period of satellite observations. Ripetta’s station (http://www. idrografico.roma.it), has one of the most complete measurements data set monitoring the river discharge relative
at more than 96% of the Tiber River drainage basin. To estimate the streamflow variability we then calculate
monthly averages from the daily dataset, as well as the coefficient of variation of streamflow,
=
σ
CV
Q
Qw
w, where Qw ll Qw
is the flow discharge monthly mean and σQw its standard deviation18; low (high) CV indicates flow regimes that are
weakly (highly) variable around the mean and are termed as persistent (erratic). The use of monthly averages
allows us to highlight the link between GSD of suspended sediment off the river mouth and the streamflow
variability7. From the comparison between daily water discharge and CV (see Fig. S1 in the Supplementary
Information) we observe that the coefficient of variation marks, properly, those months that were characterized
by strong variability (i.e., erratic flows). Remote estimation of the backscattering coefficient. Light back-scattered by marine particles
depends on size distribution, concentration, refractive index, and the detection wavelength28–30. For a
poly-dispersed assembly of non-absorbing, spherical particles with a Junge-like size distribution (see Eq. 4
below), this dependency is represented by the power law approximation of the spectral particle backscattering
coefficient λ
λ
λ
λ
=
η
−
b
b
( )
(
)
(1)
bp
bp
0
0 (1) where λ0 is a reference wavelength and η the spectral slope (dimensionless). Here we retrieve the bbp from remote-sensing reflectances (Rrs) data, by using the quasi-analytical algorithm
(QAA) (ref. 31), an approach that was validated in a previous work for Italian coastal and open waters32. In particular,
we use observations from the Moderate Resolution Imaging Spectro-radiometer (MODIS) for the period 2002
to 2014, i.e., a data record that includes the November 2012 flood event (see Text S1 in the Supplementary
Information). Rrs data were remapped to a ~ 20 x 20 km2 box centered around the Tiber River mouth (Fig. 1). Linking flow-stream variability to
grain size distribution of suspended
sediment from a satellite-based
analysis of the Tiber River plume
(Tyrrhenian Sea) 55); (b) Qw and the total volume concentration
of suspended particles (VCtot) time series (plot generated by using MATLAB 7.1 http://uk.mathworks.com/
products/matlab); VCtot is obtained from the sum of the two volume concentrations of the grain fractions
considered in this work, i.e., 3.9 µm–62.5 µm and 62.5 µm – 125 µm. amount and characteristics of the suspended load delivered from erratic events in respect to water discharge and
its seasonality. d b
h
b
fl
d h
k
f
h
f
fl
b l Inspired by the 2012 Tiber River flood, here we seek to quantify the impact of stream flow variability, i.e.,
erratic vs. persistent flow stages7, on river plume sediment assemblage, by the analysis of grain size distribution
(GSD) of the suspended sediment, retrieved from satellite. Grain size plays a fundamental role in settling velocity
and dispersal of suspended sediment over shelf areas8,9. Observations showed that formation of sedimentary bot-
tom layers and their related sandy and/or mud deposits strongly depends on sediment (sand, silty and/or clayey
particles) settling properties10–13. In particular, the role of sediment cohesion, which is controlled by sediment
size, influences morphology of deltas and coastal areas14. l
Although the knowledge of sediment characteristics gives a substantial contribution in understanding and
modeling particle dynamics and coastal geomorphology, GSD of suspended particles from river plumes are
still seldom quantified and insufficiently documented. Satellite data can be used as an alternative technique15,
in particular, off river mouth and estuarine environments, where remote sensing algorithms can be applied to
mid-resolution ocean color sensors (e.g., MERIS, MODIS, SeaWiFS, VIIRS, and OLCI) that provide near-daily
data at ~1 km spatial resolution since two decades. Moreover, remote sensing has the additional advantages of
providing a synoptic view of river plumes16–20. Application of in-situ calibrated MODIS satellite data, for instance, Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ allowed capturing the spatio-temporal variability of the distribution of suspended sediment over the coral reefs21
and assessed the impact of these extreme outflows on wetland sedimentation during the historic 2011 Mississippi
River flood18.hfi l
The relationship between the spectral beam attenuation coefficient (cp) and the GSD has been predicted theo-
retically and verified experimentally by many studies22–24. However, those findings had no application to remote
sensing data since the beam attenuation coefficient is not retrievable from space. Linking flow-stream variability to
grain size distribution of suspended
sediment from a satellite-based
analysis of the Tiber River plume
(Tyrrhenian Sea) Fortunately, after commercial
optical backscattering meters became available, research provided evidence that the spectral particle backscatter-
ing coefficient (bbp) and the GSD are also related15,25–28 in a very similar fashion as cp and the GSD were found to
be related. The advantages of this finding are many because bbp can be retrieved from ocean color observations,
thus establishing a link between remote sensing measurements and the spatio-temporal variability of particle
assemblage (i.e., the more or less large grain size fraction) for fluvial geomorphology applications and, in particu-
lar, for understanding and monitoring fate and distribution of riverine suspended load characteristics. All this sets
the base for our investigation on flow-stream variability and GSD of suspended sediment from a satellite-based
analysis off the Tiber River mouth. Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 In situ hydrological data. Its
size was chosen to be 2–3 times greater than the river-induced Rossby radius of deformation in order to ensure
that the whole river plume bulge was considered in our analysis33,34.ht p
g
y
The QAA parameterization was left as originally documented except for η, a parameter that is highly relevant
for the GSD estimations. In particular, the default value is given by35: r
r
2 1
1 2 exp
0 9
(443)
(555)
(2)
rs
rs
η =
−
.
−.
(2) where
=
. +
. r
R
R
/(0 52
1 7
)
rs
rs
rs . where
=
. +
. r
R
R
/(0 52
1 7
)
rs
rs
rs . where
=
. +
. r
R
R
/(0 52
1 7
)
rs
rs
rs . Equation (2) was originally built by using in-situ η vs. rrs(443)/rrs(555) matchups (see below) but the data cloud
was highly scattered around the mean regression curve31 (Fig. 2b), thus having little predictive value; addition-
ally, data were mainly taken from open ocean waters. A biased estimated η does not have a huge impact on the
retrieval of absorption and its various components, which is for what the QAA is mostly used for, but here η is
crucial component of the retrieval. Hence, to provide reliable η estimates, we analyzed matched in-situ spectra of (Rrs) and bbp at various stations,
covering a wide turbidity range, belonging to two field campaigns, conducted in 2013 and 2015 across Italian
seas (Fig. 2a). bbp was measured with an ECO-VSF3 (Wet Labs, Inc.) at the wavelengths 470, 530 and 660 nm. From surface data, the spectral slope (η) was calculated as the slope of the best linear fit of bbp against the wave-
length, both log-transformed. Rrs was measured using the free-profiling technique using OCR-507 radiometers
(Satlantic, Inc.), at the bands 412, 443, 490, 510, 555, 665 and 863 nm using protocols compliant with NASA’s
recommendations32,36,37.l Figure 2b shows the blue-to-green band ratio rrs(443)/rrs(555) from our in-situ reflectances respect to our
measured η. Data shows high scattering but also high overestimation respect to Lee’s curve (Eq. 2). For this Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ Figure 2. (a) Geographical distribution of the in situ Rrs and bbp data. (b) rrs(443)/rrs(555) vs. In situ hydrological data. η and Lee’s curve
(ref Lee), RMSE = 0.93 m−1. (c) rrs(555)/rrs(665) vs. η and the best linear fit for the data: η = 0.2764 rrs(555)/
rrs(665)−1.793, r2 = 0.48, RMSE = 0.51 m−1. Plots are generated by using MATLAB 7.1 http://uk.mathworks. com/products/matlab. Figure 2. (a) Geographical distribution of the in situ Rrs and bbp data. (b) rrs(443)/rrs(555) vs. η and Lee’s curve
(ref Lee), RMSE = 0.93 m−1. (c) rrs(555)/rrs(665) vs. η and the best linear fit for the data: η = 0.2764 rrs(555)/
rrs(665)−1.793, r2 = 0.48, RMSE = 0.51 m−1. Plots are generated by using MATLAB 7.1 http://uk.mathworks. com/products/matlab. reason, we choose the ratio rrs(555)/rrs(665) as predictive variable, which results to be more sensitive to variations
in coastal waters; indeed, the fit is largely improved (Fig. 2c). The best linear fit to this latter data cloud is the
equation that we propose here to estimate the backscattering spectral slope from remote sensing, where we have
replaced the in-situ band 665 nm with the MODIS band 667 nm, assuming negligible difference between them: reason, we choose the ratio rrs(555)/rrs(665) as predictive variable, which results to be more sensitive to variations
in coastal waters; indeed, the fit is largely improved (Fig. 2c). The best linear fit to this latter data cloud is the
equation that we propose here to estimate the backscattering spectral slope from remote sensing, where we have
replaced the in-situ band 665 nm with the MODIS band 667 nm, assuming negligible difference between them: η =
. −
. . r
r
0 2764
(555)
(667)
1 7393
(3)
rs
rs (3) GSD power law approximation. Generally, simple power law approximations of GSD (Eq. 3) can be suc-
cessfully used to describe and assess poly-dispersed particle assemblies in a wide array of oceanic and estuarine
water types and river plumes28,38. =
ξ
−
N D
N D
D
D
( )
(
)
(4)
0
0 (4) where D0 is a reference particle diameter and ξ is a dimensionless parameter called Junge slope39. A higher ξ
implies a higher proportion of smaller particles and vice-versa40. ξ values generally increase from river water to
the open ocean38, clearly indicating that coarser particles are more confined to the near-shore areas.h The full size range can be divided in several sub-ranges to study the proportion between size classes. For
instance, Kostadinov et al. In situ hydrological data. VC estimation can relate to a size fraction like silt (3.9 µm to 62.5 µm), sand (62.5 µm to 125 µm) or to the
global assembly by taking the full size range (3.9 to 125 µm). In this latter case, it is named total volume concen-
tration (VCtot). For these two size classes VCtot is equal to the sum of the particle volume of each class, reported
in parts per million28. A useful parameter, related to a GSD, is the median diameter (D50), which is the diameter at which the lower
and the upper volume fractions of the GSD are equal. It is obtained after the integration of the GSD in Eq. (5),
following the Junge assumption in Eq. (3): ξ
ξ
=
+
≠
=
ξ
ξ
ξ
−
−
−
D
D
D
D
D
2
,
4
,
4
(6)
max
min
min
max
50
4
4
1
4 (6) Remote estimation of the GSD from the backscattering coefficient. Theoretical derivations that
used the Mie scattering theory25 and experimental investigation26,27 showed that the GSD can be associated to bbp by
means of η in Eq. (1). Such research built on previous knowledge that focused on the beam attenuation coefficient
cp instead of on bbp (refs. 23,24,29). Kostadinov et al. (ref. 25) found that if bbp was predicted using Mie calculations,
a relationship between η and ξ also existed, and was similar to that related to cp. Theoretical and empirical studies
agree on the fact that ξ increases as a monotonic function of η and thus, as for ξ, lower η is associated to a higher
proportion of large particles and vice-versa.h p
p
g p
In this work, we use the look-up table (LUT) provided by Kostadinov et al. (ref. 25). The LUT requires the input
of the spectral bbp, which is estimated with the QAA. The LUT returns the Junge slope ξ, and also the GSD value
N0 at a reference size D0, which is relevant for VCtot calculations. In this regard, we notice that VCtot is a concentra-
tion value of particles within a given size range, but is conceptually and methodologically different than the usual
suspended particulate matter concentration which is commonly retrieved by using remote sensing techniques
and expressed in mass density units (e.g.42–44). Remote and hydrological data alignment. In situ hydrological data. (ref. 25) applied this approach to study the particle abundance and volume in the
dimensional ranges corresponding to three main phytoplankton-size classes in the global ocean40,41. Here, we
consider the silt range, between the limits 3.9 µm and 62.5 µm, and the very-fine sand range, between 62.5 µm
and 125 µm. µ
By assuming spherical shape, the volume concentration (VC) between two given edge sizes (Dmin, Dmax) is
equal to: ∫
π
=
. VC
N D
D
dD
( )4
3
2
(5)
D
D
3
min
max (5) Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ Figure 3. (a) Cells size distribution of the computational domain. Red circles show observation points position
at the river mouth and 500 m from the river mouth. (b) Suspended sediment concentration ratio increases and
then decreases during flood events. Colored lines display different duration of floods while continuous line and
dashed shows, respectively, the propagation sediment at the river mouth and at the distance of 500 m. The location
of the observation points is highlighted by red circles in the panel (a). (c) Along-stream velocity profile for the
two flood discharges we consider in our work: 3-day flood (blue solid line) and 10-day flood (red dashed line). The downstream transect begins from the channel mouth and is positioned at the half channel width. Plots are
generated by using MATLAB 7.1 http://uk.mathworks.com/products/matlab. Figure 3. (a) Cells size distribution of the computational domain. Red circles show observation points position
at the river mouth and 500 m from the river mouth. (b) Suspended sediment concentration ratio increases and
then decreases during flood events. Colored lines display different duration of floods while continuous line and
dashed shows, respectively, the propagation sediment at the river mouth and at the distance of 500 m. The location
of the observation points is highlighted by red circles in the panel (a). (c) Along-stream velocity profile for the
two flood discharges we consider in our work: 3-day flood (blue solid line) and 10-day flood (red dashed line). The downstream transect begins from the channel mouth and is positioned at the half channel width. Plots are
generated by using MATLAB 7.1 http://uk.mathworks.com/products/matlab. Results
l Correlation between water discharge (Qw) and the satellite-based total volume concentration of suspended particles
(VCtot) shows a comforting agreement, i.e., r = 0.55 (Fig. 1b); peaks of water discharge are always associated with
high values of VCtot, which ranges from 1200 to 2500 ppm. This qualitatively suggests that the remote sensing
approach, along with the reflectance sampling method we implemented, is a promising tool for complementing
the in situ hydrological dataset.hi y
g
The synergy between remote sensing and hydrologic measurements, specifically, reveals an intriguing relationship
between flow stage characteristics (i.e., flow erraticity) and the mean grain-size population. Here, we use the
monthly particle backscattering spectral slope (η) as an indicator of the GSD. By plotting the monthly η versus
the coefficient of variation (CV) we find that CV and η correlate negatively, thus suggesting that more erratic
river discharges (higher CV) relate to a larger proportion of large particles in suspension. On the contrary, more
persistent flow discharges (i.e., low CV) are related to finer particles in suspension (Fig. 4). Although the dot cloud
of this correlation is rather disperse, this general trend is highlighted by the best linear fit: for a given water dis-
charge, grain-size of the suspended sediment will be larger if the flow is more erratic than if the flow is persistent. Moreover, focusing on seasonality, the correlation shows that erratic events occur mostly in late fall/early winter,
when we observe high proportion of large particles in suspension. g p
p
g p
p
To analyze consistency of our results, we evaluate the Junge slope (ξ) and the median of grain-size diameters
(i.e., the D50) within the plume, for the day that records the maximum concentration of suspended sediment of
the whole time series off the river mouth, i.e., the flood event of 17 November 2012 (Fig. 5). It results that Rrs(667),
which is a good proxy for turbidity, sharply decreases with the distance from the river mouths (Fig. 5a); Fig. 5b,c refer
to particle quality more than quantity, which explains why the plume extent is better delineated than in panel a). Still, particle sedimentation affects GSD within the river plume. Largest, presumably non-cohesive particles (i.e.,
D50 ~ 65 µm) remain in suspension for few hundreds of meters from the two main mouths of the Tiber River,
while the finest (clay and silt) fraction is delivered off shore. Therefore, within the plume, Fig. www.nature.com/scientificreports/ implicit scheme. Here we use the three-dimensional formulation of the hydrodynamic and sediment transport
models, implemented in Delft 3D.tl t
Delft 3D solves the Navier-Stokes equations for an incompressible fluid with the assumptions of shallow water,
Boussinesq, and hydrostatic approximation. The standard k-ε closure model47 is used for the vertical eddy viscos-
ity, and the horizontal eddy viscosity is computed with a large eddy simulation technique. Transport of suspended
sediment is calculated by solving the three-dimensional Advection-Diffusion equation, computed by following
the method of Van Rijn48.l j
For our simulation, the numerical domain is a square (3 km × 3 km), in which there is a river flow in inlet
on the western side (Fig. 3a). The computational grid is composed of squared cells (30 m × 30 m) but, along
the centerline, a refinement occurs, where each cell size is 30 m × 10 m. We simulate a different flood progres-
sion with linear growth and falling limb two times, compared to the rising limb but same peak discharge, i.e.,
Qw = 2000 m3/s. For both study cases, we set up a rising discharge Q and suspended sediment concentration
(SSC) entering in the domain from the river mouth (Fig. 3a). We remark that even though the two flood discharge
evolutions we consider in the numerical experiment are different, they are characterized by similar along-stream
velocity profiles (Fig. 3c). In particular, the two velocity profiles remain identical till the first 500 m downstream,
from the channel mouth (where the GSD analysis is performed). Moreover, we make clearer that our choice is not
“coincidental”: the 3-day flood and a 10-day flood were designed to demonstrate that the ‘erraticity’ (rather than
the velocity) is responsible for the different suspended sediment GSD we observe from remote sensing.i y
pf
p
g
For the simulations we consider the upper limit of very-fine sand size range (i.e., 125 μm), according to the fact
that the suspended fraction recorded during the Tiber River floods ranges from clay to very fine sand (3.9–125 µm)
and that the non-cohesive very fine sand fraction is the most predominant one3,4. At the inlet, we impose a con-
stant input of 0.6 kg/m3 of suspended sediment concentration49. All sediment is characterized by specific density
of 2,650 kg/m3 and dry bed density of 1,600 kg/m3. On the north, south, and eastern boundary we assigned a
water elevation equal to zero (Fig. www.nature.com/scientificreports/ 3a) to reproduce the sea level. A trapezoidal river channel with flow depth h = 3
m is incised into a non-erodible coastline, where we imposed a zero water depth and no flux boundary condition. The bottom roughness is modeled with the Chézy’s formulation, using the constant Chézy coefficient value (i.e.,
65 m1/2/s). The initial condition of the models consists of a constant bathymetry with 5 m of erodible sediment on
the basin bottom and a time step Δt = 9 s. In situ hydrological data. To track the river’s turbid water off the river mouth from
satellite-based images, we select the maximum Rrs(667) value within the box for each day (Fig. 1a). In the prac-
tical implementation, we chose the sampled pixel as the closest to the 99.5th percentile instead of the maximum
to avoid outliers due to inaccuracies in pixel flagging, usually along the coastline or close to clouds. We thus
obtained a time series of daily Rrs spectra (at 443, 488, 555 and 667 nm) over the same period of the collected
water discharge data. From this daily time series, daily time series of η and the GSD were obtained and then
monthly averaged. Numerical experiments. We model river mouth sediment dynamics with Delft 3D45,46, by using different
flood discharge evolution. In particular, we test consistencies between a model of sediment transport based on
remote sensing observations, and a generic numerical model of sediment transport, by exploring the relation
between GSD and stages (erratic vs. persistent) of river outflows (Fig. 3). Delft 3D simulates fluid flow, waves,
sediment transport, and morphological changes at different timescales. The equations of fluid, sediment transport
and deposition are discretized on a 3D curvilinear finite difference grid and solved by an alternating direction Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ Results
l 5b,c shows how the
grain size is decreasing as the distance to the coast increases.hil g
g
The numerical experiment confirms the link between erraticity of the outflow and its ability to suspended
higher concentration of sediment. To assess the behavior of suspended sediment in the river jet plume for dif-
ferent flow stage conditions, we obtain suspended sediment concentration for the entire water column, SSCTOT,
and at 1 m below the water surface, SSC1m, from numerical simulations. The ratio SSC1m / SSCTOT is assumed to
represent the ability of the outflow to keep large (i.e., 125 μm) grain-size in suspension, and thus, to be compa-
rable with the satellite-retrieved Junge slope. Numerical results show that, although reaching the same value of
water discharge, and despite the fact that the two run show similar along-stream velocity profiles, the erratic event
(i.e., the 3-days flood in Fig. 3b) suspends larger concentration of very fine-to-fine sediment, compared to a more
persistent event (i.e., the 10-days flood). Moreover, in both study-cases the sediment concentration in the upper
part of the plume increases due to the jet expansion. In fact, most of the sediment is suspended from the bottom
layers, as bedload, to the top. Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ www.nature.com/scientificreports
tificreports/
Figure 4. Scatter plot of the coefficient of variation of monthly river discharge (CV) vs. the monthly particle
backscattering spectral slope (η). Both variables are dimensionless (see text). The standard deviation of monthly
η is also indicated as vertical bars; dot diameter is proportional to the number of cloud-free satellite images. The best linear fit (y = −0.186 x -1.143; r2 = 0.35) is meant to highlight the general trend that marks the relation
between flow erraticity (CV) and GSD. Months with less than three valid observations were discarded, as
calculation of a standard deviation is not possible for them. Colorbar indicates months for each point (month
numbers are also reported); reddish dots represents late fall/early winter months. Plot generated by using
MATLAB 7.1 http://uk.mathworks.com/products/matlab. Figure 4. Scatter plot of the coefficient of variation of monthly river discharge (CV) vs. the monthly particle
backscattering spectral slope (η). Both variables are dimensionless (see text). The standard deviation of monthly
η is also indicated as vertical bars; dot diameter is proportional to the number of cloud-free satellite images. Results
l The best linear fit (y = −0.186 x -1.143; r2 = 0.35) is meant to highlight the general trend that marks the relation
between flow erraticity (CV) and GSD. Months with less than three valid observations were discarded, as
calculation of a standard deviation is not possible for them. Colorbar indicates months for each point (month
numbers are also reported); reddish dots represents late fall/early winter months. Plot generated by using
MATLAB 7.1 http://uk.mathworks.com/products/matlab. Figure 5. (a) Rrs(667) [sr−1] as a proxy for suspended sediment; (b) GSD Junge slope; (in situ) median particle
diameter (D50) [m] of the GSD, after assuming a Junge slope with a minimum size of 3.9 µm and a maximum
of 125 µm, for the 12 November 2012. The D50 [m] is analytically obtained by assuming a Junge-type GSD
between the range 3.9–125 µm and it is zero out of this range. The three panels confirm that the proportion of
smaller particles increases from river mouth to the open ocean and thus that coarser particles are more confined
to the near-shore areas. Plots are generated by using MATLAB 7.1 http://uk.mathworks.com/products/matlab. Figure 5. (a) Rrs(667) [sr−1] as a proxy for suspended sediment; (b) GSD Junge slope; (in situ) median particle
diameter (D50) [m] of the GSD, after assuming a Junge slope with a minimum size of 3.9 µm and a maximum
of 125 µm, for the 12 November 2012. The D50 [m] is analytically obtained by assuming a Junge-type GSD
between the range 3.9–125 µm and it is zero out of this range. The three panels confirm that the proportion of
smaller particles increases from river mouth to the open ocean and thus that coarser particles are more confined
to the near-shore areas. Plots are generated by using MATLAB 7.1 http://uk.mathworks.com/products/matlab. www.nature.com/scientificreports/ Received: 28 March 2019; Accepted: 21 November 2019;
Published: xx xx xxxx Received: 28 March 2019; Accepted: 21 November 2019;
Published: xx xx xxxx www.nature.com/scientificreports/ spectral slope) time series allows for long-term statistical analyses of both water discharge and suspended
sediment load, thus complementing and integrating GSD in situ measurements; ii) the synoptic property of satel-
lite images allows for a spatial view of the sedimentary characteristics of the plume, thus helping the recognition
of river plume pattern in terms of GSD. Indeed, by analyzing a ten-year-long time series, we assess the ability of
unsteady flow to transport large particle size in suspension in a real field case53. Moreover, by focusing on the 2012
erratic event, we pictured depositional patterns of different grain size off the river mouth, a crucial feature that
may largely impact shoreline restoration strategies54. y
g y
p
g
Previous researches demonstrated the possibility of using satellite to retrieve GSD24–27. Based on those, we
retrieved a GSD long time series within the Tiber River plume off the river mouth. The resulting analysis, which
paired river flow discharge data to satellite-based GSD showed i) an encouraging correlation (r = 0.55) between
water discharge (Qw) and the total volume concentration (VCtot) of suspended particles and ii) a relationship
between flow stage characteristics (i.e., flow erraticity) and the spectral slope of backscattering (η), in turn, related
to the GSD. Finally, by running a numerical model that confirmed what we obtained from remote sensing, there-
fore, assessing the relationship we find between GSD and flow variability. This, in turn, gives also robustness to
our ansatz, that is, the methodology to retrieve GSD from satellite spectral slope of backscattering.lfi gy
p
p
g
Unsteady flow makes the problem of sediment transport very complex and difficult to describe in general laws. Our work presents a novel, satellite-based tool for retrieving GSD of suspended solids debouching from river
mouth, by adapting a semi-analytical technique25,26 to coastal geomorphology and sediment transport applica-
tions. From the new satellite-based tool, we find an emerging relation between GSD and flow stages (erratic vs. persistent floods), also relating this to seasonality. By setting an ad hoc numerical experiment, we then provide
a mechanistic support to our findings, setting novel hypotheses on the role of sharp spatial pressure gradient
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tant than in streams that mainly transport silt and clay51 (Walling et al.40); this will result in high δ-values52. Analysis of matched remote-sensing and in-situ hydrological data, supported by numerical simulations, showed
that streamflow variability (CV) correlates with the grain size of the suspended sediment. Accordingly, Basso et al. (ref. 7) found that large values of coefficient of variation (CV), are associated to large values of the δ -coefficient of
δ the sediment rating curve, i.e., the power law relation between water and sediment discharges,
β
=
δ
Q
Q
s
w (where
𝛿 and β are empirical coefficients)50. In turn, the δ-coefficient represents the erosive power of the river, with large
values being indicative for rivers with a strong increase in erosive power and in sediment transport capacity when
discharge increases38. Moreover, δ-values are also affected by the GSD of the material available for transport, i.e. in
streams characterized by sand sized sediments the power of the stream to transport sediment will be more impor-
tant than in streams that mainly transport silt and clay51 (Walling et al.40); this will result in high δ-values52. l
h
h
h
f
d h d
l
d
d
d
l Our results show that the synergy of remote sensing and hydrologic in situ data expands and complements
these previous findings from two aspects: i) the satellite-based reconstruction of GSD (or particle backscattering Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 References 58, 21–35 15. Pinet, S., Martinez, J. M., Ouillon, S., Lartiges, B. & Villar, R. E. Variability of apparent and inherent optical properties of sediment-
laden waters in large river basins–lessons from in situ measurements and bio-optical modeling. Opt. express 25, A283–A310 (2017). 16. Walker, N. D. Satellite assessment of Mississippi River plume variability: causes and predictability. Remote Sens. Environ. 58, 21–35
(1996).l laden waters in large river basins–lessons from in situ measurements and bio-optical modeling. Opt. express 25, A283–A310 (2017). 16. Walker, N. D. Satellite assessment of Mississippi River plume variability: causes and predictability. Remote Sens. Environ. 58, 21–35
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MODIS AQUA images were provided by NASA Goddard Space Flight Center, Ocean Ecology Laboratory, Ocean
Biology Processing Group; (2014): Moderate-resolution Imaging Spectroradiometer (MODIS) Aqua Ocean
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Agency. Jaime Pitarch received partial funding from the Ministry for Science and Culture of Lower Saxony,
Germany through the “Coastal Ocean Darkening” project (VWZN3175). Partial support was also provided by
European Marine Observation and Data network – Physics, and the Flagship Project RITMARE—The Italian
Research for the Sea—coordinated by the Italian National Research Council and funded by the Italian Ministry
of Education, University and Research within the National Research Program 2011–15. We finally thank D.J. Jerolmack and C. Paola for the insightful discussions at the initial stage of the work. Any use of trade, product, or
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55. https://oceancolor.gsfc.nasa.gov/data/10.5067/AQUA/MODIS/L2/OC/2018/NASA Goddard Space Flight Center, Ocean Ecology
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Laboratory, Ocean Biology Processing Group. Author contributions J.P. performed both remote sensing analysis and hydrographic analysis, contributed to collection and presentation
of the data, contributed to the overall study, participated in data interpretation, and contributed to the writing
of this manuscript. F.F. supervised the research, contributed to the data interpretation and presentation, set the
main, leading hypotheses of the work, led the writing of the main text, and coordinated the overall study. S.M. and 9 Scientific Reports | (2019) 9:19729 | https://doi.org/10.1038/s41598-019-56409-8 www.nature.com/scientificreports/ V.B. co-supervised the research, and participated in data interpretation. A.D. participated in data interpretation,
and contributed to the remote sensing analysis. W.N. performed the numerical experiment and contributed to the
overall study, participated in data interpretation, and contributed to the writing of this manuscript. V.B. co-supervised the research, and participated in data interpretation. A.D. participated in data interpretation,
and contributed to the remote sensing analysis. W.N. performed the numerical experiment and contributed to the
overall study, participated in data interpretation, and contributed to the writing of this manuscript. V.B. co-supervised the research, and participated in data interpretation. A.D. participated in data interpretation,
and contributed to the remote sensing analysis. W.N. performed the numerical experiment and contributed to the
overall study, participated in data interpretation, and contributed to the writing of this manuscript. Competing interests
The authors declare no competing interests. Additional information
Supplementary information is available for this paper at https://doi.org/10.1038/s41598-019-56409-8. Correspondence and requests for materials should be addressed to F.F. Reprints and permissions information is available at www.nature.com/reprints. Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and
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format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Cre-
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article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the
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mitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the
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The authors declare no competing interests. Additional information Supplementary information is available for this paper at https://doi.org/10.1038/s41598-019-56409-8. Correspondence and requests for materials should be addressed to F.F. Reprints and permissions information is available at www.nature.com/reprints. Reprints and permissions information is available at www.nature.com/reprints. Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and
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English
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Effects of paleogeographic changes and CO2 variability on northern mid-latitudinal temperature gradients in the Cretaceous
|
Nature communications
| 2,023
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cc-by
| 7,771
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Article https://doi.org/10.1038/s41467-023-40905-7 1Fredy & Nadine Herrmann Institute of Earth Sciences, Hebrew University of Jerusalem, Edmond J. Safra Campus, Givat Ram, Jerusalem 9190401, Israel.
2School of Geographical Sciences and Cabot Institute, University of Bristol, Bristol BS8 1SS, UK. 3State Key Laboratory of Tibetan Plateau Earth System,
Environment and Resources (TPESER), Institute of Tibetan Plateau Research, Chinese Academy of Sciences, Beijing 100101, China.
e-mail: kaushal.g@mail.huji.ac.il Effects of paleogeographic changes and CO2
variability on northern mid-latitudinal tem-
perature gradients in the Cretaceous Kaushal Gianchandani
1
, Sagi Maor
1, Ori Adam
1,
Alexander Farnsworth
2,3, Hezi Gildor
1, Daniel J. Lunt
2 & Nathan Paldor
1 Kaushal Gianchandani
1
, Sagi Maor
1, Ori Adam
1,
Alexander Farnsworth
2,3, Hezi Gildor
1, Daniel J. Lunt
2 & Nathan Paldor
1 Received: 28 February 2023
Accepted: 15 August 2023
Check for updates The Cretaceous ‘greenhouse’ period (~145 to ~66 million years ago, Ma) in
Earth’s history is relatively well documented by multiple paleoproxy records,
which indicate that the meridional sea surface temperature (SST) gradient
increased (non-monotonically) from the Valanginian (~135 Ma) to the Maas-
trichtian (~68 Ma). Changes in atmospheric CO2 concentration, solar constant,
and paleogeography are the primary drivers of variations in the spatio-
temporal distribution of SST. However, the particular contribution of each of
these drivers (and their underlying mechanisms) to changes in the SST dis-
tribution remains poorly understood. Here we use data from a suite of
paleoclimate simulations to compare the relative effects of atmospheric CO2
variability and paleogeographic changes on mid-latitudinal SST gradient
through the Cretaceous. Further, we use a fundamental model of wind-driven
ocean gyres to quantify how changes in the Northern Hemisphere paleogeo-
graphy weaken the circulation in subtropical ocean gyres, leading to an
increase in extratropical SSTs. Analysis of oxygen isotopes in foraminifera tests (δ18O) and the index
of tetraethers that quantifies the archaeal lipid distributions in marine
sediments (TEX86) indicate that the sea surface temperature (SST)
during the Cretaceous period (~145 to ~66 million years ago, Ma) ran-
ged between 28–35 °C in low latitudes to 11–22 °C in high latitudes
(first and third quartiles of LOESS smoothened data)1. A similar trend is
observed during the Paleocene (~66 to ~56 Ma) and Eocene (~56 to
~34 Ma) epochs of the Paleogene period (~66 to ~23 Ma) when the
tropical and high latitude temperatures were between 31–33 °C and
11–14 °C, respectively2. The persistence of such high globally averaged
SST
and
low
pole-to-tropics
temperature gradient
during
the
Cretaceous–Paleocene–Eocene (CPE) is attributed predominantly to
the strong greenhouse effect ensuing from the higher partial pressure
of CO2 (pCO2) in the atmosphere3,4. Wind-driven gyral circulation in the CPE The mid-latitudinal gyral basin in the northern paleo-Pacific (hereafter
referred to as the North Pacific) retains an approximately rectangular
geometry — similar to Stommel’s idealized ocean — during the CPE,
despite considerable plate tectonic activity25,26. Variations in the North
Pacific’s circulation are expected to strongly affect the climate during
the CPE, since for these ~110 million years much of the planet’s sea-
water was in the large paleo-Pacific27,28. Henry Stommel was the first to formulate a simple mathematical
model comprising five parameters that capture the fundamental
characteristics of the circulation in such gyres17. While the linear
dependence of ψ on the amplitude of zonally averaged wind stress (τ)
is evident from the analytical solution of Stommel’s model, discerning
its dependence on the zonal and meridional extents of the ocean basin
(Lx and Ly, respectively) is rather non-trivial. It should be noted that in
Stommel’s model, the meridional boundaries of the basin are co-
located with the latitudes of zero wind-stress curl. However, in the real
ocean, the two do not necessarily coincide in general, and the extent of
the surface gyres is constrained only by the latitudes corresponding to
the vanishing wind-stress curl. Previous studies established that both ψ
and the poleward heat transport (H) associated with a surface gyre
increase with the horizontal aspect ratio
Ly
Lx
of the ocean basin con-
taining the gyre18–20 (hereafter referred to as the ‘gyral basin’). Thus, the
aspect ratio of any gyral basin is governed by the curl of the overlying
wind stress and the relative position of the zonal boundaries set by the
continents. Based on these first-order constraints, it was recently
postulated that the large meridional SST gradients observed during
some geologic periods may have resulted from the smaller ψ
(and consequently a smaller H) associated with the surface gyres
contained in basins with small aspect ratios20. g p
The Lx of the mid-latitudinal gyral basin in the North Pacific, i.e.,
the typical longitudinal distance between continental margins (see
Methods), remained nearly constant during the CPE (~13,000 km,
Supplementary Fig. 1). On the other hand, Ly of this basin, which is
defined as the distance between the two latitudes where the wind-
stress curl over the surface vanishes, decreased (non-monotonically)
from ~3900 km to ~2800 km and then increased to ~3600 km (Fig. 1a). Variability in meridional SST gradients and its dependence on
atmospheric CO2 and paleogeography To examine the variability in meridional SST gradients during the CPE,
we first consider the SST gradients from the Early to the Late Cretac-
eous and subsequently focus on the trend from the Paleocene to the
Eocene. Specifically, we compare the Maastrichtian to the Valanginian
(rather than the Berriasian, given the paucity of the proxy-derived SST
data in this age1) in the Cretaceous and the Priabonian to the Danian
age (~63 Ma) in the PE. Here we use data generated from two ensembles of paleoclimate
simulations carried out using the HadCM3L (specifically, HadCM3LB-
M2.1aD) model6,7 to better constrain the relative contribution of
changes in pCO2 and paleogeographic effects on the SST distribution
during the CPE. In particular, we develop a simple model that isolates
the role of paleogeographically driven changes in extratropical gyral
circulation and the corresponding effect on the evolution of pole-to-
tropics SST gradient. The model is then applied to the HadCM3L
data to show that paleogeography-driven reduction in ψ (and conse-
quently H) associated with the surface ocean gyres may have been a
key factor contributing to the variability in meridional SST gradients
observed during the CPE1,2. g
In the HadCM3L simulations carried out for an atmospheric CO2
concentration of 560 ppmv, the meridional gradient of the zonally
averaged SST (over all longitudes, ΔSSTy) calculated between 20°N
and 50°N increases from 12.5 °C in the Valanginian to 15.7 °C in the
Maastrichtian, whereas, ΔSSTy decreases from 16.5 °C in the Danian to
15.5 °C in the Priabonian. The same trend persists for an atmospheric
CO2 concentration of 1120 ppmv, ΔSSTy increases from 11.1 °C to
13.9 °C during the Cretaceous and decreases from 14.3 °C to 13.7 °C
during the PE. Figure 2a illustrates the temporal trend in ΔSSTy for the
two atmospheric CO2 concentrations. In both simulation ensembles,
the temporal variation in ΔSSTy from the Valanginian to the Maas-
trichtian is ~3 °C even though the global mean SST varies only slightly
(standard deviation of less than 1 °C about the ensemble averages, see
Supplementary Fig. 3). Alternatively, the temporal variation in ΔSSTy
from the Danian to the Priabonian (~0.8 °C) is comparable to the
standard deviation about the global mean SST for the ensembles. Wind-driven gyral circulation in the CPE These variations in Ly determine the variations in the aspect ratio and
hence the value of ψ in this gyral basin. This is illustrated by comparing
ψ in the longer Valanginian gyral basin (~135 Ma; Fig. 1b), which is
double that of the ~30% narrower Maastrichtian gyral basin (~68 Ma;
Fig. 1c). The ocean during the Valanginian is also characterized by a
much weaker SST gradient, which is consistent with a stronger H
compared to the Maastrichtian. Article Although previous studies have investigated the effect of differ-
ent paleogeographies corresponding to multiple ages in the CPE on
variations in the pole-to-tropics temperature gradient and the ocean
circulation10–12,16, the effect of paleogeographic changes on the surface
ocean circulation specifically has not been examined before. Surface
ocean gyres contained between two meridional continental bound-
aries are prominent features of the present-day ocean. The meridional
volume (mass) transport (ψ) associated with these gyres redistributes
heat in the ocean and is critical to regulating the climate on Earth’s
surface. deep-sea studies6. The solar constant in both ensembles increases
monotonically from the Berriasian age (~143 Ma) to the Priabonian age
(~36 Ma) by 0.9% (see Methods). In practice, the relatively small mag-
nitude of the forcing associated with the increase in solar constant is
dwarfed by changes in pCO2 and paleogeography between simulations
and their relative impact on SST distributions. Variability in meridional SST gradients and its dependence on
atmospheric CO2 and paleogeography Furthermore, comparing the two simulation ensembles shows that
during the CPE, a doubling of pCO2 decreases ΔSSTy between 0.7 °C
(observed in the Albian age, ~106 Ma) and 2.6 °C (observed in the
Selandian age, ~60 Ma), resulting in an average decrease of 1.8 °C
in ΔSSTy. Effects of paleogeographic changes and CO2
variability on northern mid-latitudinal tem-
perature gradients in the Cretaceous However, in addition to changes in
pCO2, the spatiotemporal variability in SST can also stem from
tecton
topogr
Specifi
been d
spread
pics te
the pe
mation
and va
the pa
and La
events
orogra
contrib
Paleoc
1Fredy & Nadine Herrmann Institute of Earth Sciences, Hebrew University of Jerusalem
2School of Geographical Sciences and Cabot Institute, University of Bristol, Bristol BS8
Environment and Resources (TPESER), Institute of Tibetan Plateau Research, Chinese A
e-mail: kaushal.g@mail.huji.ac.il
Nature Communications| (2023) 14:5193 Analysis of oxygen isotopes in foraminifera tests (δ18O) and the index
of tetraethers that quantifies the archaeal lipid distributions in marine
sediments (TEX86) indicate that the sea surface temperature (SST)
during the Cretaceous period (~145 to ~66 million years ago, Ma) ran-
ged between 28–35 °C in low latitudes to 11–22 °C in high latitudes
(first and third quartiles of LOESS smoothened data)1. A similar trend is
observed during the Paleocene (~66 to ~56 Ma) and Eocene (~56 to
~34 Ma) epochs of the Paleogene period (~66 to ~23 Ma) when the
tropical and high latitude temperatures were between 31–33 °C and
11–14 °C, respectively2. The persistence of such high globally averaged
SST
and
low
pole-to-tropics
temperature gradient
during
the
Cretaceous–Paleocene–Eocene (CPE) is attributed predominantly to
the strong greenhouse effect ensuing from the higher partial pressure
of CO2 (pCO2) in the atmosphere3,4. However, in addition to changes in
pCO2, the spatiotemporal variability in SST can also stem from tectonically driven changes in paleogeography, including surface
topography, ocean bathymetry, and the arrangement of continents5–7. Specifically, paleogeographic alterations during the Cretaceous have
been demonstrated to dampen seasonal variations and trigger wide-
spread flooding, ultimately leading to moderation of the pole-to-tro-
pics temperature gradient8,9. The rearrangement of continents during
the period has also been associated with shifts in the deep-water for-
mation sites7,10, alternations in the oceanic overturning circulation11,
and variations in mid-Cretaceous deep-sea ventilation12. Furthermore,
the paleogeographic features associated with certain ages in the Early
and Late Cretaceous have been found to be more conducive to glacial
events than the mid-Cretaceous13. Moreover, changes in Antarctic
orography14 and the opening of the Drake passage15 could have both
contributed to the variability in meridional SST gradients during the
Paleocene–Eocene (PE). Nature Communications| (2023) 14:5193 Nature Communications| (2023) 14:5193 1 https://doi.org/10.1038/s41467-023-40905-7 Article Role of gyral circulation in determining the meridional SST
gradients The decrease in ΔSSTy resulting from an increase in pCO2 of the
model atmosphere can be attributed to polar amplification22, 29–32,
whereas the increase in ΔSSTy from the Early to Late Cretaceous in
the two ensembles of simulations is related to paleogeography-
driven changes. Paleogeography can influence many factors that
could in turn affect polar amplification (e.g., cloud distribution and
other local feedbacks); here, we focus on its influence on oceanic
heat transport (H). Figure 2b shows a strong anti-correlation
between ΔSSTy and the maximal volumetric (mass) transport (ψ)
in mid-latitudes, which indicates that Cretaceous basins with smal-
ler Ly contain surface gyres with smaller ψ. Thus, the weakening of
mid-latitudinal H (Supplementary Fig. 4), caused by the weakened
mid-latitude ocean gyres contributes substantially to the increase of
ΔSSTy in the middle—Late Cretaceous. To quantify this effect, we
assume that: Comparing the typical variation in ΔSSTy from the change in pCO2
with paleogeography-driven variation in ΔSSTy indicates that paleo-
geographic changes had a greater effect on the mid-latitudinal tem-
perature gradient during the Cretaceous than the long-term changes in
pCO2. This is also true for the PE. However, since the overall ΔSSTy
variation during the PE is comparable to the standard deviation in the
ensemble-averaged global mean SST, the analysis that follows only
focuses on the Cretaceous. The differences between the long-term SST gradients in the
simulations and estimates based on paleotemperature reconstructions
may be attributed to the limited spatial distribution of available
proxies. Paleotemperature reconstructions based on δ18O and TEX86
proxies suggest that meridional SST gradients first decreased from
~10–17 °C in the Valanginian to ~3–5 °C in the Aptian (~119 Ma) and
subsequently increased through the middle—Late Cretaceous to reach
a maximum of ~19–21 °C in the Maastrichtian1. However, the trend in
this proxy-derived meridional SST gradient is only expected to capture
the long-term trends in Equator-to-pole temperature difference. It
does not capture the mid-latitudinal temperature gradient in the
Northern Hemisphere (the focus of this study) since no data exists
between 30°N and 80°N and the data for the high latitude paleo-
temperature originates from sites located between 48°S and 54°S
paleolatitudes. Nevertheless, the estimated ΔSSTy variation resulting 1. To first order, ΔSSTy decreases linearly with H and with the
poleward heat transport associated with the atmospheric mer-
idional overturning circulation (MOC). 2. The heat transport associated with the atmospheric MOC is pro-
portional to H33. 3. Numerical simulations All simulations necessary for this work were carried out using the UK
Met Office coupled atmosphere-ocean model, HadCM3L (resolution:
3.75° in longitude × 2.5° in latitude), which includes multiple climate
feedbacks, including vegetation feedbacks6,21. The atmospheric CO2
concentration in the two ensembles is kept constant at 560 ppmv and
1120 ppmv (i.e., ×2 and ×4 preindustrial atmospheric CO2 concentra-
tion) while the geography and the solar constant are varied. The cho-
sen values of atmospheric CO2 provide an idealized framework that
allows us to isolate the effect of CO2 versus paleogeography. In addi-
tion, this range of CO2 values captures the maximum variation
(530 ppmv to 840 ppmv) in the long-term trend of LOESS smoothened
data on estimated atmospheric CO2 concentration during the CPE22. The range does not capture the very lowest values of CO2 seen in some
proxies in the latest Cretaceous23, but these low values are inconsistent
with relatively high reconstructed global mean temperatures at this
time24. The geographies corresponding to different geological ages are
constrained by data aggregated from lithological, tectonic, fossil, and y
The long-term trend of LOESS smoothened data on estimated
atmospheric CO2 concentration suggests that a decrease in atmo-
spheric CO2 from ~700 ppmv in the Valanginian to ~570 ppmv in the
Maastrichtian22 can cause a decrease in global mean SST by ~1.5 °C
(based on a typical climate sensitivity parameter of ~4.5 °C per CO2 Nature Communications| (2023) 14:5193 2 Article https://doi.org/10.1038/s41467-023-40905-7 Fig. 1 | Changes in the meridional extent of the mid-latitudinal gyral basin (Ly)
in the Northern paleo-Pacific during the Cretaceous and Paleogene periods for
an atmospheric CO2 concentration of 560 ppmv (solid blue curve) and
1120 ppmv (dashed red curve). a Change in Ly during the Early-Late Cretaceous,
the Paleocene, and the Eocene epochs. b, c Sea surface temperature (°C, color) and
gyral streamlines (Sv, contour; 1 Sv = 106 m3 s−1) in the ocean during the Valanginian
(~135 Ma) and the Maastrichtian (~68 Ma) ages for an atmospheric CO2 concentra-
tion of 1120 ppmv. Dotted lines mark the latitudes where the wind-stress curl is
zero. For completeness, Supplementary Fig. 2 shows the sea surface temperature
fields and streamlines in the ocean during the Valanginian and the Maastrichtian for
an atmospheric CO2 concentration of 560 ppmv. Fig. Numerical simulations 1 | Changes in the meridional extent of the mid-latitudinal gyral basin (Ly)
in the Northern paleo-Pacific during the Cretaceous and Paleogene periods for
an atmospheric CO2 concentration of 560 ppmv (solid blue curve) and
1120 ppmv (dashed red curve). a Change in Ly during the Early-Late Cretaceous,
the Paleocene, and the Eocene epochs. b, c Sea surface temperature (°C, color) and
gyral streamlines (Sv, contour; 1 Sv = 106 m3 s−1) in the ocean during the Valanginian (~135 Ma) and the Maastrichtian (~68 Ma) ages for an atmospheric CO2 concentra-
tion of 1120 ppmv. Dotted lines mark the latitudes where the wind-stress curl is
zero. For completeness, Supplementary Fig. 2 shows the sea surface temperature
fields and streamlines in the ocean during the Valanginian and the Maastrichtian for
an atmospheric CO2 concentration of 560 ppmv. p
pp
1120 ppmv (dashed red curve). a Change in Ly during the Early-Late Cretaceous,
the Paleocene, and the Eocene epochs. b, c Sea surface temperature (°C, color) and
gyral streamlines (Sv, contour; 1 Sv = 106 m3 s−1) in the ocean during the Valanginian from the combined effect of reduction in atmospheric CO2 con-
centration and paleogeographic changes from the Valanginian to the
Maastrichtian (~3.6 °C) is within 55% of the variation estimated from
proxies (~6.5 °C). doubling7) and an increase in ΔSSTy by ~0.6 °C (since a doubling of
atmospheric CO2 decreases ΔSSTy by 1.8 °C a decrease in CO2 from
~700 ppmv to ~570 ppmv, i.e., a reduction by a factor of 0.8, is
expected to increase ΔSSTy by
logð0:8Þ
logð2Þ
×−1.8 = 0.6 °C). Similarly, a
decrease in atmospheric CO2 from ~590 ppmv in the Danian to
~530 ppmv in the Priabonian22 is expected to cause a 0.7 °C decrease in
global mean SST and an 0.3 °C increase in ΔSSTy. These estimates are
consistent with the inverse relation between ΔSSTy and global mean
SST observed across multiple climate states in Earth’s history with
varied paleogeographies2. It should be noted that changes in atmo-
spheric CO2 may have had a more pronounced effect on ΔSSTy since
this study does not account for the short-term fluctuations in pCO2
trend or the lowest values of CO2 inferred from some proxies23. Nature Communications| (2023) 14:5193 Role of gyral circulation in determining the meridional SST
gradients The ocean circulation, to first order, is similar to the circulation
in a ‘Stommel-like’ gyre. As such, due to rapid advection, there
is negligible loss of heat by warm water along the western
boundary. In contrast, the water mass loses much of its heat
while it traverses the northern and eastern edges of the basin at
a much lower speed. Thus, SST along the southern and western
boundaries is high, whereas SST along the northern and eastern
boundaries is low. Further, the East–West SST gradient is pro-
portional to ΔSSTy. Nature Communications| (2023) 14:5193 3 https://doi.org/10.1038/s41467-023-40905-7 Article Fig. 2 | Meridional temperature gradient and maximal volumetric (mass)
transport in the mid-latitudinal northern paleo-Pacific. a Meridional gradient of
zonal-mean SST (ΔSSTy) between 20°N and 50°N for an atmospheric CO2 con-
centration of 560 ppmv (blue curve) and 1120 ppmv (red curve). The best-fit curves
for the two atmospheric CO2 concentrations (dashed blue and red curves) are
calculated using ΔSSTy and the maximal volume (mass) transport (ψmax). The
shaded region shows the range between which the best-fit curve can vary with 1σ
error in κ and ΔSSTRad
y
. b Decrease in ψmax from the Early to Late Cretaceous and the
subsequent increase from the Paleocene to the Eocene for the two atmospheric
CO2 concentrations atmospheric CO2 of 560 ppmv, and ΔSSTRad
y
= 18:0 ± 0:9 °C and
κ = ð3:7 ± 0:6Þ × 104 Sv−1 °C−1 for an atmospheric CO2 of 1120 ppmv. The best-fit curves (Fig. 2a, dashed curves) illustrate that the reduction
in ψmax explains ~80% and ~75% of the variance in ΔSSTy from the
Berriasian to the Priabonian in the simulation ensemble with an
atmospheric CO2 of 560 ppmv and 1120 ppmv, respectively. This
underscores the role of Ly in the increase in ΔSSTy during the
Cretaceous and in the subsequent decrease in ΔSSTy from the
Paleocene to the Eocene. Though our regression analysis captures the general trend in the
evolution of ΔSSTy during the CPE for both simulation ensembles, we
observe a steep increase in ΔSSTy between ~130 Ma and ~100 Ma for
the simulations with an atmospheric CO2 of 1120 ppmv (Fig. 2a, solid
red curve). This steep increase lies outside the range in which the best-
fit curve is expected to vary based on one standard deviation variation
in κ and ΔSSTRad
y
. Role of gyral circulation in determining the meridional SST
gradients It stems from an abrupt decrease in the SST at 50°N
between the Barremian age (~128 Ma) and the Albian age (Supple-
mentary Fig. 5). However, examining the precise climatic variations
and feedback associated with paleogeographic changes that led to this
abrupt cooling between the Barremian and the Albian are beyond the
scope of this study. The HadCM3L simulations show that geologic ages in the Cre-
taceous with a large Ly (e.g., the Valanginian) are characterized by a
wide Equator-to-pole extent of the ocean basin ( eLy) (Fig. 3a, b), while
geologic ages with a small Ly (e.g., the Maastrichtian) are marked by a
narrower eLy (Fig. 3c, d). Moreover, like Ly, eLy also decreased from the
Early to Late Cretaceous (Supplementary Fig. 6). The correlation
between Ly and eLy can be attributed to the increase in land area in the
polar North Pacific since the mid-Cretaceous (Supplementary Fig. 7),
because the increased land area amplifies the damping effect (friction)
over the surface, which reduces the meridional distance between the
zonal wind extremes (thus reducing the distance between the latitudes
where the wind-stress curl over the surface vanishes). However, mul-
tiple other features of the atmosphere-ocean system not considered
here can affect Ly, e.g., meridional extent and/or intensity of the
Hadley cell, the intensity of the Walker circulation, and exchange of
water masses between ocean basins34. For instance, Ly and eLy do not
coevolve during the PE (Supplementary Fig. 6), which could be related
to one or more of these features. Exploring a causal relation between
changes in these climatic features and Ly on geologic timescales is left
for future study. Fig. 2 | Meridional temperature gradient and maximal volumetric (mass) Fig. 2 | Meridional temperature gradient and maximal volumetric (mass)
transport in the mid-latitudinal northern paleo-Pacific. a Meridional gradient of
zonal-mean SST (ΔSSTy) between 20°N and 50°N for an atmospheric CO2 con-
centration of 560 ppmv (blue curve) and 1120 ppmv (red curve). The best-fit curves
for the two atmospheric CO2 concentrations (dashed blue and red curves) are
calculated using ΔSSTy and the maximal volume (mass) transport (ψmax). The
shaded region shows the range between which the best-fit curve can vary with 1σ
error in κ and ΔSSTRad
y
. b Decrease in ψmax from the Early to Late Cretaceous and the
subsequent increase from the Paleocene to the Eocene for the two atmospheric
CO2 concentrations. Role of gyral circulation in determining the meridional SST
gradients transport in the mid latitudinal northern paleo Pacific. a Meridional gradient of
zonal-mean SST (ΔSSTy) between 20°N and 50°N for an atmospheric CO2 con-
centration of 560 ppmv (blue curve) and 1120 ppmv (red curve). The best-fit curves
for the two atmospheric CO2 concentrations (dashed blue and red curves) are
calculated using ΔSSTy and the maximal volume (mass) transport (ψmax). The
shaded region shows the range between which the best-fit curve can vary with 1σ
error in κ and ΔSSTRad
y
. b Decrease in ψmax from the Early to Late Cretaceous and the
subsequent increase from the Paleocene to the Eocene for the two atmospheric
CO2 concentrations. Discussion In summary, we find that a large portion of the increase in ΔSSTy
observed in the Stommel-like mid-latitudinal North Pacific basin dur-
ing the Cretaceous results from the decrease in poleward ocean heat
transport. This decrease in heat transport is a consequence of a wea-
kened intensity of the gyral circulation, i.e., a reduction in the volu-
metric/mass transport associated with the gyre. This weakening results
from tectonically driven changes in Northern Hemisphere paleogeo-
graphy, which reduces the horizontal aspect ratio of the gyral basin. In summary, we find that a large portion of the increase in ΔSSTy
observed in the Stommel-like mid-latitudinal North Pacific basin dur-
ing the Cretaceous results from the decrease in poleward ocean heat
transport. This decrease in heat transport is a consequence of a wea-
kened intensity of the gyral circulation, i.e., a reduction in the volu-
metric/mass transport associated with the gyre. This weakening results
from tectonically driven changes in Northern Hemisphere paleogeo-
graphy, which reduces the horizontal aspect ratio of the gyral basin. The anti-correlation between oceanic heat transport and ΔSSTy per-
sists in the PE as well, and an increase in oceanic heat transport during
these epochs is accompanied by a decrease in ΔSSTy. However, this
decrease is of the order of the standard deviation of the ensemble-
averaged global mean SST. We note that in contrast to the Northern
Hemisphere, the two ensembles of simulations do not show a dis-
cernible trend in ΔSSTy variations in the Southern Hemisphere, likely
because of the break-up of the Antarctica–Australia landmass during
this period — demonstrating the limitations of the theory pro-
posed here. Nature Communications| (2023) 14:5193 Based on these assumptions, we find (see Methods): Panels b and d show the temperature fields
(°C, color) and streamlines (Sv, contours) of the surface gyre during the two ages; eL
is the typical Equator-to-pole extent of the basin. The yellow-purple loops depic
the meridional extent of the atmospheric meridional overturning circulations. Th
latitude where the winds change from Easterlies to Westerlies marks the polewar
extent of the atmospheric overturning cell40. (°C, color) and streamlines (Sv, contours) of the surface gyre during the two ages; eLy
is the typical Equator-to-pole extent of the basin. The yellow-purple loops depict
the meridional extent of the atmospheric meridional overturning circulations. The
latitude where the winds change from Easterlies to Westerlies marks the poleward
extent of the atmospheric overturning cell40. Fig. 3 | Zonal-mean wind speed and temperature distribution in the northern
paleo-Pacific Ocean for an atmospheric CO2 concentration of 1120 ppmv in the
Valanginian (~142 Ma) and Maastrichtian (~68 Ma) ages. Panels a and c show the
zonally averaged winds in the Valanginian and the Maastrichtian, respectively;
the dotted lines denote the latitudes where the wind-stress curl vanishes and Ly is
the meridional distance between them. Panels b and d show the temperature fields (°C, color) and streamlines (Sv, contours) of the surface gyre during the two ages; eLy
is the typical Equator-to-pole extent of the basin. The yellow-purple loops depict
the meridional extent of the atmospheric meridional overturning circulations. The
latitude where the winds change from Easterlies to Westerlies marks the poleward
extent of the atmospheric overturning cell40. (°C, color) and streamlines (Sv, contours) of the surface gyre during the two ages; eLy
is the typical Equator-to-pole extent of the basin. The yellow-purple loops depict
the meridional extent of the atmospheric meridional overturning circulations. The
latitude where the winds change from Easterlies to Westerlies marks the poleward
extent of the atmospheric overturning cell40. he dotted lines denote the latitudes where the wind-stress curl vanishes and Ly is
he meridional distance between them. Panels b and d show the temperature fields longitudes, λ1–λ4, based on the continental boundaries between the
latitudes ϕ1 and ϕ2 such that the region enclosed between the vertices
P (ϕ1, λ1), Q (ϕ1, λ2), R (ϕ2, λ3) and S (ϕ2, λ4) denotes the ‘gyral basin’. Based on these assumptions, we find (see Methods): The
meridional extent of the gyral basin (Ly) is defined as the latitudinal
distance between the edges PQ and RS and the zonal extent (Lx) is
defined as the cosine(latitude) weighted average of the lengths of PQ
and RS. Supplementary Fig. 1 illustrates the gyral basins during dif-
ferent geologic ages in the Cretaceous and the Paleogene. We note that
both Ly and Lx depend on the grid resolution. Despite the rather
stringent definition of Ly and Lx and their dependence on the grid
resolution, the correlation between Ly and ΔSSTy is very strong, which
further demonstrates the robustness of our analysis. estimated atmospheric CO2 levels23,35 in general and, thus by extension,
in the magnitude of ΔSSTy variation we attribute to pCO2 fluctuations
during the period. In addition to changes in atmospheric CO2 con-
centration and paleogeography, ΔSSTy can also be non-trivially affec-
ted by several other drivers in the climate system, e.g., fluctuations in
deep-sea temperatures22,36, opening/closing/deepening of oceanic
gateways10,11,37 and variations in the extent of marine ice cover8. The temporal variation in meridional temperature gradients
obtained from the HadCM3L model for both the simulation ensembles
agrees with the proxy-inferred increase in temperature gradient from
the Early to the Late Cretaceous1. Since the atmospheric CO2 and
continental arrangement do not coevolve in the current set of simu-
lations, we stop short of comparing the model temperature gradients
with proxy data. This should be addressed in future work using data
from multiple climate models in which atmospheric CO2 and geo-
graphy are varied simultaneously. Additionally, numerical simulations
that use a hierarchy of coupled atmosphere–ocean models with idea-
lized continental configurations can further advance our under-
standing of how paleogeography affects ΔSSTy. Nevertheless, our
analysis is based on foundational models of atmospheric and oceanic
circulation and first-order assumptions to quantify the characteristics
of a given paleoclimate — which are simple and robust. It may, there-
fore, also explain the effects of changes in paleogeography on varia-
tions in ΔSSTy in other geologic periods. Based on these assumptions, we find (see Methods): Based on these assumptions, we find (see Methods): ΔSSTy =
ΔSSTRad
y
1 + κ × ψmax × ΔSSTRad
y
ΔSSTy =
ΔSSTRad
y
1 + κ × ψmax × ΔSSTRad
y
ð1Þ g
p y
p
gy
The anti-correlation between oceanic heat transport and ΔSSTy per-
sists in the PE as well, and an increase in oceanic heat transport during
these epochs is accompanied by a decrease in ΔSSTy. However, this
decrease is of the order of the standard deviation of the ensemble-
averaged global mean SST. We note that in contrast to the Northern
Hemisphere, the two ensembles of simulations do not show a dis-
cernible trend in ΔSSTy variations in the Southern Hemisphere, likely
because of the break-up of the Antarctica–Australia landmass during
this period — demonstrating the limitations of the theory pro-
posed here. where ΔSSTRad
y
is the meridional gradient of SST at radiative equili-
brium ðH = 0Þ, ψmax is the maximal gyral volume (mass) transport in
the mid-latitudinal North Pacific, and κ is an empirical constant. The
above equation is developed based on the paleogeographic con-
siderations relevant to the Cretaceous, but we expect it to be also
applicable to the PE since the majority (>60%) of the volumetric (mass)
transport in the ocean’s surface is concentrated in the North
Pacific until the Priabonian. Performing a regression analysis, based
on this equation, on ΔSSTy and ψmax estimated from the HadCM3L
model data for the 19 ages considered here yields ΔSSTRad
y
= 22:6 ± 1:4
°C
and
κ = ð3:9 ± 0:5Þ × 104 Sv−1 °C−1
(1 Sv = 106 m3 s−1)
for
an We also find that fluctuations in atmospheric CO2 concentration
can substantially affect the ΔSSTy during the Cretaceous and estimate
the magnitude of ΔSSTy variation based on the long-term trend of
atmospheric CO2. However, there is a considerable uncertainty in the Nature Communications| (2023) 14:5193 4 Article https://doi.org/10.1038/s41467-023-40905-7 Fig. 3 | Zonal-mean wind speed and temperature distribution in the northern
paleo-Pacific Ocean for an atmospheric CO2 concentration of 1120 ppmv in the
Valanginian (~142 Ma) and Maastrichtian (~68 Ma) ages. Panels a and c show the
zonally averaged winds in the Valanginian and the Maastrichtian, respectively;
the dotted lines denote the latitudes where the wind-stress curl vanishes and Ly is
the meridional distance between them. Relation between meridional sea surface temperature gradients
and the volume (mass) transport in the ocean Poleward volume (mass) and heat transport associated with the
atmospheric meridional overturning circulation and the gyral circula-
tion in the ocean’s surface lower the meridional sea surface tempera-
ture gradient (ΔSSTy), which is driven by differential solar heating. We
assume that ΔSSTy is given by (see assumption 1, main text): ΔSSTy = ΔSSTRad
y
1 cOHO cAHA
ð2Þ ð2Þ where ΔSSTRad
y
is the meridional gradient of SST at radiative equili-
brium
when
ocean
and
atmosphere
heat
transport
are
zero
ðHO = HA = 0Þ, and cO and cA are the appropriate empirical constants. Assuming that atmosphere and ocean heat transport are proportional
to each other (on average, and not at every latitude)33, we may write
cAHA = ðγ 1ÞcOHO, where γ>1 is an arbitrary constant (see assumption
2, main text). Equation (2) can then be rewritten as: where ΔSSTRad
y
is the meridional gradient of SST at radiative equili-
brium
when
ocean
and
atmosphere
heat
transport
are
zero
ðHO = HA = 0Þ, and cO and cA are the appropriate empirical constants. Assuming that atmosphere and ocean heat transport are proportional
to each other (on average, and not at every latitude)33, we may write
cAHA = ðγ 1ÞcOHO, where γ>1 is an arbitrary constant (see assumption
2, main text). Equation (2) can then be rewritten as: Nature Communications| (2023) 14:5193 Coupled atmosphere–ocean model description The data analyzed in this study was generated from two ensembles of
simulations carried out using the UK Met Office coupled atmosphere-
ocean model, HadCM3L, which includes multiple climate feedbacks,
including vegetation feedbacks6,21. The model has a horizontal reso-
lution of 3.75° in longitude × 2.5° in latitude. The atmosphere and the
ocean are divided into 19 and 20 vertical levels, respectively. The CO2
in the model atmosphere is kept fixed at 560 ppmv and 1120 ppmv for
the two ensembles of simulations, while the paleogeography and solar
constant are varied. The chosen range of atmospheric CO2 captures
the maximum variation (530 ppmv to 840 ppmv) in the long-term
trend of LOESS smoothened data on estimated atmospheric CO2
concentration during the CPE22. The geographies corresponding to
different geological ages are developed by Getech Plc using methods
described in ref. 39, and are constrained by geological data aggregated
from lithological, tectonic, fossil, and deep-sea studies. The geo-
graphies were originally produced at a resolution of 0.5° in both lati-
tude and longitude, which were subsequently used to generate
geographies at model resolution. Additionally, some smoothening was
applied to ensure model stability. A more detailed description is
available in Section 2.1 of ref. 6. Given that the variation in solar con-
stant was quite small (a monotonic increase of ~0.9% from the Ber-
riasian age in the Cretaceous to the Priabonian age in the Paleogene),
we assume that for a given pCO2 in the model atmosphere, any chan-
ges in the spatial distribution of SSTs between simulations arise pri-
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calculated by the HadCM3L model reaches an extremum. These
extrema correspond to the points where the curl of the zonally aver-
aged wind stress vanishes. Subsequently, we determine four typical ΔSSTy = ΔSSTRad
y
1 γcOHO
:
ð3Þ ð3Þ We further assume that the gyral heat transport can be written as
HO = cpρψmaxΔSSTx, where ΔSSTx is the East-West SST gradient38, We further assume that the gyral heat transport can be written as
HO = cpρψmaxΔSSTx, where ΔSSTx is the East-West SST gradient38, Nature Communications| (2023) 14:5193 5 5 Article https://doi.org/10.1038/s41467-023-40905-7 yielding: 5. Goddéris, Y., Donnadieu, Y., le Hir, G., Lefebvre, V. & Nardin, E. The
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:
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long as you give appropriate credit to the original author(s) and the
source, provide a link to the Creative Commons licence, and indicate if
changes were made. The images or other third party material in this
article are included in the article’s Creative Commons licence, unless
indicated otherwise in a credit line to the material. If material is not
included in the article’s Creative Commons licence and your intended
use is not permitted by statutory regulation or exceeds the permitted
use, you will need to obtain permission directly from the copyright
holder. To view a copy of this licence, visit http://creativecommons.org/
licenses/by/4.0/. Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as
long as you give appropriate credit to the original author(s) and the
source, provide a link to the Creative Commons licence, and indicate if
changes were made. The images or other third party material in this
article are included in the article’s Creative Commons licence, unless
indicated otherwise in a credit line to the material. If material is not
included in the article’s Creative Commons licence and your intended
use is not permitted by statutory regulation or exceeds the permitted
use, you will need to obtain permission directly from the copyright
holder. To view a copy of this licence, visit http://creativecommons.org/
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The authors declare no competing interests. The authors declare no competing interests. 33. Held, I. M. The partitioning of the poleward energy transport
between the tropical ocean and atmosphere. J. Atmos. Sci. 58,
943–948 (2001). Additional information
Supplementary information The online version contains
supplementary material available at
https://doi.org/10.1038/s41467-023-40905-7. Acknowledgements K.G. and H.G. were supported by the Joint National Natural Science
Foundation of China–Israel Science Foundation (research grant num-
ber 2547/17) and by the U.S.–Israel Binational Science Foundation
(BSF grant number 2018152). D.J.L. and A.F. acknowledge Leverhulme
grant RPG-2019-365; D.J.L. acknowledges NERC grant NE/X000222/1; © The Author(s) 2023 7 Nature Communications| (2023) 14:5193 7 Nature Communications| (2023) 14:5193 Nature Communications| (2023) 14:5193
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https://openalex.org/W2130393733
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https://malariajournal.biomedcentral.com/counter/pdf/10.1186/1475-2875-12-244
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English
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Genetic diversity of Plasmodium falciparum and distribution of drug resistance haplotypes in Yemen
|
Malaria journal
| 2,013
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cc-by
| 7,493
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* Correspondence: h.babiker@ed.ac.uk
1Department of Biochemistry, Faculty of Medicine and Health Sciences,
Sultan Qaboos University, Alkhod, PO Box 35, Muscat, Oman
6Institute of Immunology and Infection Research, School of Biological
Sciences, University of Edinburgh, Edinburgh EH9 3JT, UK
Full list of author information is available at the end of the article Al-Hamidhi et al. Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 Al-Hamidhi et al. Malaria Journal 2013, 12:244 Open Access Open Access Genetic diversity of Plasmodium falciparum and
distribution of drug resistance haplotypes in
Yemen Salama Al-Hamidhi2, Mohammed AK Mahdy3,4, Zainab Al-Hashami1, Hissa Al-Farsi2, Abdulsalam M Al-mekhlafi4,
Mohamed A Idris2, Albano Beja-Pereira5 and Hamza A Babiker1,6* © 2013 Al-Hamidhi et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the
Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: Despite evident success of malaria control in many sites in the Arabian Peninsula, malaria remains
endemic in a few spots, in Yemen and south-west of Saudi Arabia. In addition to local transmission, imported
malaria sustains an extra source of parasites that can challenge the strengths of local control strategies. This study
examined the genetic diversity of Plasmodium falciparum in Yemen and mutations of drug resistant genes, to
elucidate parasite structure and distribution of drug resistance genotypes in the region. Methods: Five polymorphic loci (MSP-2, Pfg377 and three microsatellites on chromosome 8) not involved in anti-
malarial drug resistance, and four drug resistant genes (pfcrt, pfmdr1, dhfr and dhps) were genotyped in 108 P. falciparum isolates collected in three sites in Yemen: Dhamar, Hodeidah and Taiz. Results: High diversity was seen in non-drug genes, pfg377 (He = 0.66), msp-2 (He = 0.80) and three microsatellites
on chr 8, 7.7 kb (He = 0.88), 4.3 kb (He = 0.77) and 0.8 kb (He = 0.71). There was a high level of mixed-genotype
infections (57%), with an average 1.8 genotypes per patient. No linkage disequilibrium was seen between drug
resistant genes and the non-drug markers (p < 0.05). Genetic differentiation between populations was low (most
pair-wise FST values <0.03), indicating extensive gene flow between the parasites in the three sites. There was a high prevalence of mutations in pfmdr1, pfcrt and dhfr; with four mutant pfmdr1 genotypes (NFCDD
[57%], NFSND[21%], YFCDD[13%] and YFSND[8% ]), two mutant pfcrt genotypes (CVIET[89%] and SVMNT[4%]) and
one mutant dhfr genotype (ICNI[53.7%]). However, no dhps mutations were detected. Conclusion: The high diversity of P. falciparum in Yemen is indicative of a large parasite reservoir, which represents
a challenge to control efforts. The presence of two distinct pfcrt genotype, CVIET and SVMNT, suggests that
chloroquine resistance can possibly be related to a migratory path from Africa and Asia. The absence of the triple
mutant dhfr genotype (IRN) and dhps mutations supports the use of artesunate + sulphadoxine-pyrimethamine as
first-line therapy. However, the prevalent pfmdr1 genotype NFSND [21%] has previously been associated with
tolerance/resistance response to artemisinin combination therapy (ACT). Regular surveys are, therefore, important to
monitor spread of pfmdr1 and dhfr mutations and response to ACT. Keywords: Malaria, Yemen, Arabian Peninsula, Drug resistance, Plasmodium falciparum, Genetic diversity © 2013 Al-Hamidhi et al.; licensee BioMed Central Ltd. Study sites and sampling One hundred and fifteen finger-prick blood samples
were
collected,
from
microscopy-confirmed
malaria
cases, in three localities in Yemen (Dhamar, Hodeidah
and Taiz) (Figure 1) Dhamar (Latitude: 14°32′33″ N,
Longitude: 44°24′18″ E ) is a highland (2407 m above sea)
where transmission occurs throughout the year. Hodeidah
(Latitude: 14°48′08″N, Longitude: 42°57′04″E) is a coastal
area where malaria transmission occurs in winter and Taiz
(Latitude:
13°34′44″N,
Longitude:
44°01′19″E)
is
a
mountainous area (1311 m above sea) and transmission
occurs in summer [2]. Ethical clearance was obtained
from Faculty of Medicine, Sana’a University, Yemen. In-
formed consent was obtained from each participant before
enrolment in the study. In Yemen, over 60% of the population live in areas
with stable malaria transmission [6]. Plasmodium falcip-
arum is responsible for about 90% of malaria cases. Indi-
genous chloroquine resistance (CQR) was reported in
1989 [7,8], and since then it spread to different regions
in the highland and lowland areas in Taiz [7] and
Hodeidah [9]. Cross- sectional surveys in 2008 and 2009
in Taiz, Hodeidah, Dhamar and Rymah showed high
prevalence of the CQR marker pfcrt 76 T [2]. However,
resistance to sulphadoxine-pyrimethamine (SP) is rare in
Yemen [2,10]. Currently SP is used in combination with
artesunate (AS) as first-line treatment of uncomplicated
falciparum malaria. In addition, artemether-lumefantrine
(AL) is used as a second-line therapy for treatment
failure [11]. Background The present study examined genetic diversity of P. falciparum in Yemen, and distribution of genes impli-
cated in resistance to widely used anti-malarial drugs, in-
cluding, chloroquine, SP, mefloquine and artemisinin. Such analysis will provide desirable information on the
genetics structure of P. falciparum in the Arabian
Peninsula, and the prospect of evolution of drug resist-
ance following recent shift of malaria therapy to artemi-
sinin combination therapy (ACT). The Arabian Peninsula lies at the fringes of malaria
endemicity
where
successful
control
measures
have
brought local transmission to halt in many countries in the
region, e.g. Bahrain, Kuwait and United Arab Emirates
(UAE) [1]. However, some sites in Yemen and southern
Saudi Arabia remain malarious, with a high prevalence of
drug-resistant Plasmodium falciparum parasites [2,3]. In
addition, imported malaria cases, via asymptomatic travel-
lers from malaria-endemic areas, sustain a major challenge
for possible rebound of local transmission [4]. Methods The region attracts numerous migrations from Africa
and Asia. Over the past two decades political instability
has resulted in mass movement of people from the
Horne of Africa (Somalia, Eritrea and Ethiopia) into
Yemen and Saudi Arabia [5]. These countries are known
for high prevalence of drug-resistant P. falciparum mal-
aria. In addition, the region attracts a lot of skilled
workers from Asia. However, the impact of these migra-
tions on the gene pool of local malaria parasites in the
Arabian Peninsula remains unclear as information on
malaria parasites genetics is scarce. Abstract This is an Open Access article distributed under the terms of the
Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly cited. Al-Hamidhi et al. Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 Page 2 of 8 Non drug-resistance genes
pfg377 and msp-2 The pfg377 gene primers amplify a polymorphic region,
encoding a seven degenerate amino acid repeats; alleles
of this gene, vary by multiples of 21 base pairs [12]. Polymerase chain reaction (PCR) primers and conditions
for amplification were as described previously [12], with
minor modification where the nested primer 377R3D1
was labelled. Figure 1 Geographical location of the three studied sites in Yemen: Hodeidah (1), Dhamar (2), and Taiz (3). Figure 1 Geographical location of the three studied sites in Yemen: Hodeidah (1), Dhamar (2), and Taiz (3). Page 3 of 8 Page 3 of 8 Page 3 of 8 Al-Hamidhi et al. Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 pfcrt, pfmdr1, dhfr and dhps pfcrt, pfmdr1, dhfr and dhps Alleles of the pfcrt, pfmdr1, dhfr and dhps genes were
amplified using two rounds of PCR [1,16-19]. The ampli-
fied fragments of each gene encompasses mutations as-
sociated with CQR (pfcrt-72,-74,-75, and −76), and
multidrug resistant gene-1 (pfmdr1-86, -184, 1034,1042
and 1246), pyrimethamine resistance (dhfr-51, -59, -108,
and −164), sulphadoxine resistance (dhps-436, -437, -
540, and −613). Principal coordinate analysis (PCoA) was carried out
to examine the genetic similarities (of alleles of non-
drug resistant genes) between P. falciparum isolates in
the three coding sites using Genalex Version 6.5 [28]. The two-dimensional PCoA plot shows the relationships
between the haplotypic variants found in the three
populations. PCR fragments of the isolates, together with control
clones (3D7, Hb3 and Dd2) of known sequence and drug
response, were then sequenced. The parasites were
obtained from The European Malaria Reference Reagent
Repository. 3D7 and Dd2, are chloroquine sensitive and
resistant clones, with pfcrt76K and pfcrt76T codons, re-
spectively [20], and codons pfmdr1 86 N and 86Y associ-
ated
with
susceptible
and
resistance
response
to
choloroquine, respectively [21]. The 3D7 clone is known
to be pyrimethamine and sulphadoxine sensitive carrying
wild-type dhfr and dhps gene, implicated in resistance to
the two drug respectively [22], while Dd2 is a pyrimeth-
amine and sulphadoxine resistance, carrying mutations
in dhfr and dhps associated with in vitro resistance [23]. The PCR products were first purified using ExoSAP-IT
(USB) as described by the manufacturer. Sequencing was
carried out using BigDye Terminator v3.1 cycle (ABI,
UK) and run on a thermocycler. Initial sequences were
compared with reference sequences in the PlasmoDB
database of P. falciparum genomic using BLASTN,
BioEdit and Lasergene software. Statistical analysis msp-2 alleles fall into two sequence types, FC27 and
3D7, which differ in the sequence of the dimorphic
central region, block 3 [13]. PCR primers and conditions
for amplification of each type were described by Aubouy
et al. [14]. The prevalence of an allele and genotype were calculated
as percentage of all the alleles and genotypes detected at
a
given
locus
among
the
examined
isolates. The
expected heterozygosity index (He), which measures the
diversity at a locus examined, was calculated for non-
drug resistant genes using microsatellite Toolkit software
as described elsewhere [24]. The fluorescent PCR products of pfg377 and msp-2
were analysed in an ABI 3130XL Genetic Analyzer, and
alleles were visualized and sized on Genesmapper v. 4
(Applied Biosystems). The multiplicity of infection was defined as the pro-
portion of people who carry more than one allele (geno-
type) for any of the examined genes, and the minimum
number of clones per infection was estimated as the lar-
gest number of alleles at any of the examined loci [25]. Microsatellites Three single copy microsatellites on chr 8 flanking the
dhps (0.8, 4.3 and 7.7 kb from the 3′ end of the gene)
were types. All isolates examined in this study harbour
the wild-type dhps, therefore, the above microsatellites
are not influenced by drug pressure and were used to
examine parasite diversity and population structure. Gamete linkage disequilibrium (LD) values between all
pairs of loci were calculated for all samples GENEPOP
v.4.1 [26]. For these analyses, only the predominant
alleles at each locus, indicating clonal infections, were
used. The microsatellites were examined as described by
Roper et al. [15]. Fluorescent PCR products were
analysed in an ABI 3130XL Genetic Analyzer, and alleles
were visualized and sized on Genemapper v.4 (Applied
Biosystems). To examine whether allele frequencies differ between
parasite populations in Dhamar, Hodeidah and Taiz, FST
indices were calculated using Weir and Cockerham's
method [27] estimator of Wright's F-statistics using the
computer package GENEPOP v.4.1 [26]. A permutation
test (n = 10,000) was applied (permuting alleles over
populations) to test whether FST indices were signifi-
cantly different from zero. Results Out of the 115 samples with microscopy-confirmed mal-
aria parasites, 108 proved to be P. falciparum and pro-
duced the expected PCR amplicons for the examined
genes (pfcrt, pfmdr1, dhfr, dhps, pfg377 and msp-2). The
other seven (6.4%) isolates were most likely to have been
Plasmodium vivax, which is found in Yemen. Diversity and structure of non drug-resistant genes Diversity and structure of non drug-resistant genes
The five non drug-resistant loci (pfg377, msp-2 and three
microsatellites on chr 8) were highly polymorphic,
with a number of alleles per locus ranging from five
(pfg377) to 45 (msp-2). Diversity, expressed as expected
hetrozygosity (He), was high for all loci, ranging from
0.51 to 0.89, with average of 0.66 (for pfg377) and 0.80
(for msp-2) (Table 1), reflecting high diversity among the
examined isolates. No LD was seen between genes involved in drug re-
sistance (pfcrt, pfmdr1 and dhfr) and non-drug loci
(pfg377 and msp-2) (P < 0.001). However, loci involving Al-Hamidhi et al. Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 Page 4 of 8 resistance to one drug (pfcrt and pfmdr1) had a g
average LD than those not involved in drug resi
loci (P < 0.01). Alleles at non drug-resistance loci were distr
widely across parasites in different sites in Y
(Table 1), and any ‘private’ alleles (detected in on
population) existed at very low frequencies (Add
file 1). Comparison of parasites in the three sites s
low Fst values, ranged from 0.141 to 0.003, sugg
frequent gene flow (Table 1). Furthermore, PCoA
lysis showed no pattern of clustering of parasites
of the three geographical sites (Figure 2). Distribution of drug resistance markers
pfcrt Out of the 108 P. falciparum isolates successfully exam-
ined, four (4%), 96 (89%), 96 (89%) and 100 (93%)
harboured mutations at codons 72, 74, 75 and 76 re-
spectively (Table 2). Ninety-six (89%), four (4%) and
eight (7%) isolates carried the pfcrt genotypes CVIET,
SVMNT and
the
wild-type
(CVMNK),
respectively
(Table 2). All isolates carrying the wild-type pfcrt genotype
(CVMNK) were detected in Taiz, except one seen in
Hodeidah. Similarly, isolates with the mutant pfcrt geno-
type SVMNT were only seen in Hodeidah. resistance to one drug (pfcrt and pfmdr1) had a greater
average LD than those not involved in drug resistance
loci (P < 0.01). pfmdr1 Alleles at non drug-resistance loci were distributed
widely across parasites in different sites in Yemen
(Table 1), and any ‘private’ alleles (detected in only one
population) existed at very low frequencies (Additional
file 1). Comparison of parasites in the three sites showed
low Fst values, ranged from 0.141 to 0.003, suggesting
frequent gene flow (Table 1). Furthermore, PCoA ana-
lysis showed no pattern of clustering of parasites in any
of the three geographical sites (Figure 2). There was a, high prevalence of pfmdr1 mutation at
codon N86Y (20%), Y184F (99%), S1034C (70%) and
N1042D (70%), while no mutation was seen at codon
1246 (Table 2). The majority of isolates (57%) carried the
mutant genotype (NFCDD), while 21%, 13%, and 8%
of them harboured the mutant genotype (NFSND),
(YFCDD), and (YFSND) respectively. However, only one
isolate carried the wild-type (1%) (Table 2). There was a, high prevalence of pfmdr1 mutation at
codon N86Y (20%), Y184F (99%), S1034C (70%) and
N1042D (70%), while no mutation was seen at codon
1246 (Table 2). The majority of isolates (57%) carried the
mutant genotype (NFCDD), while 21%, 13%, and 8%
of them harboured the mutant genotype (NFSND),
(YFCDD), and (YFSND) respectively. However, only one
isolate carried the wild-type (1%) (Table 2). Diversity and structure of non drug-resistant genes Multiplicity of Plasmodium falciparum infection
There was a high prevalence of mixed genotype
tion, 57% of the isolates carried more than one gen
Table 1 Allelic diversity (heterozygosity, He) at five
drug resistance loci among Plasmodium falciparum
three sites in Yemen
Dhamar
Hodeidah
Taiz
He
He
He
(n = 29)
(n = 17)
(n = 62)
Chr 8
7.7Kb
0.88
0.08
0.87
4.3Kb
0.77
0.8
0.84
0.8Kb
0.71
0.77
0.66
Chr 2
MSP-2
0.61
0.51
0.89
Chr 12
Pfg377
0.63
0.73
0.66
Mean
0.72
0.72
0.78
Figure 2 Graphical plotting of the PCoA based on genetic
microsatellites in chr 8) among Plasmodium falciparum pa
Blue icon = Taiz, red icon = Dhamar and green icon = Hodeidah Table 1 Allelic diversity (heterozygosity, He) at five non
drug resistance loci among Plasmodium falciparum in
three sites in Yemen
Dhamar
Hodeidah
Taiz
Fst
He
He
He
(n = 29)
(n = 17)
(n = 62)
Chr 8
7.7Kb
0.88
0.08
0.87
0.021
4.3Kb
0.77
0.8
0.84
0.020
0.8Kb
0.71
0.77
0.66
0.024
Chr 2
MSP-2
0.61
0.51
0.89
0.141
Chr 12
Pfg377
0.63
0.73
0.66
0.003
Mean
0.72
0.72
0.78 Table 1 Allelic diversity (heterozygosity, He) at five non
drug resistance loci among Plasmodium falciparum in
three sites in Yemen The number of genotypes per infected person (the max-
imum number of alleles observed at any locus) was esti-
mated as 1.8, assuming that each clone is readily
transmissible to mosquito, this reflects a high rate of
cross-mating [25]. The number of genotypes per infected person (the max-
imum number of alleles observed at any locus) was esti-
mated as 1.8, assuming that each clone is readily
transmissible to mosquito, this reflects a high rate of
cross-mating [25]. Multiplicity of Plasmodium falciparum infection No mutations were seen at codons 59 and 164. However,
58 (54%) isolates showed mutations at codons N51I and
S108N (Table 2). A large proportion of the isolates There was a high prevalence of mixed genotype infec-
tion, 57% of the isolates carried more than one genotype. Figure 2 Graphical plotting of the PCoA based on genetic variation of non drug-resistant genes (Pfg377, MSP-2 and three
microsatellites in chr 8) among Plasmodium falciparum parasites in three sites in Yemen, and variation representation by each axis. Blue icon = Taiz, red icon = Dhamar and green icon = Hodeidah. Figure 2 Graphical plotting of the PCoA based on genetic variation of non drug-resistant genes (Pfg377, MSP-2 and three
microsatellites in chr 8) among Plasmodium falciparum parasites in three sites in Yemen, and variation representation by each axis. Blue icon = Taiz, red icon = Dhamar and green icon = Hodeidah. Figure 2 Graphical plotting of the PCoA based on genetic variation of non drug-resistant genes (Pfg377, MSP-2 and three
microsatellites in chr 8) among Plasmodium falciparum parasites in three sites in Yemen, and variation representation by each axis. Blue icon = Taiz, red icon = Dhamar and green icon = Hodeidah. Al-Hamidhi et al. dhps Unlike the other drug-resistant genes, no mutation was
detected at all codons 436, 437, 540 and 613, which are
implicated in sulphadoxine resistance. Multiplicity of Plasmodium falciparum infection Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 Page 5 of 8 Table 2 Prevalence of mutant alleles of pfcrt, pfmdr1, dhfr
and dhps genes among 108 Plasmodium falciparum in
three sites in Yemen
All
Taiz
Dhamar
Hodeidah
pfcrt
n (%)
n (%)
n (%)
n (%)
Alleles
72S
4 (4)
0
0
4 (4)
74I
96 (89)
55(50.9)
29(26.9)
12(11)
75E
96 (89)
55(50.9)
29(26.9)
12(11)
76T
100 (93)
55(50.9)
29(26.9)
16(14.8)
Genotypes
CVMNK
8 (7)
7(6.5)
0
1(1)
SVMNT
4 (4)
0
0
4 (4)
CVIET
96 (89)
55(50.9)
29(26.9)
12(11)
pfmdr1
Alleles
86Y
22 (20)
18(16.7)
3(3)
1(1)
184 F
107 (99)
62(57)
28(25.9)
17(15.7)
1034C
76 (70)
44(40.7)
20(18.5)
12(11)
1042D
76 (70)
44(40.7)
20(18.5)
12(11)
1246Y
0
0
0
0
Genotypes
NYSND
1 (1)
0
1(1)
0
NFSND
23 (21)
12(11)
6(5.6)
5(4.6)
YFSND
8 (8)
6(5.6)
2(2)
0
NFCDD
62 (57)
32(29.6)
19(17.6)
11(10)
YFCDD
14 (13)
12(11)
1(1)
1(1)
dhfr
Alleles
51I
58 (54)
40(37)
12(11)
6(5.6)
59R
0
0
0
0
108 N
58 (54)
40(37)
12(11)
6(5.6)
164 L
0
0
0
0
Genotypes
NCSI
50 (46)
22(20.4)
17
11(10)
ICNI
58 (54)
40(37)
12(11)
6(5.6)
dhps
Alleles
436 F
0
0
0
0
437G
0
0
0
0
540E
0
0
0
0
613S
0
0
0
0
Genotypes
SAKA
108 (100)
62 (57)
29(26.9)
17(15.7)
n = number of isolates, mutated amino acids are bold and underlined. Table 2 Prevalence of mutant alleles of pfcrt, pfmdr1, dhfr
and dhps genes among 108 Plasmodium falciparum in
three sites in Yemen (54%) carried the mutant genotype (ICNI), while 50
(46%) isolates harboured the wild-type (NCSI) (Table 2). (54%) carried the mutant genotype (ICNI), while 50
(46%) isolates harboured the wild-type (NCSI) (Table 2). (54%) carried the mutant genotype (ICNI), while 50
(46%) isolates harboured the wild-type (NCSI) (Table 2). Discussion The most common orders of muta-
tion are 108 N, 59R, 51I or 108 N, 51I, 59R [57]. These
cumulative mutations in dhfr restore the parasite fitness
and allow it to overcome the effect of the drug [58]. dhfr
mutations appear earlier than dhps mutations in the de-
velopment of resistance to SP combined effect [59], and
P. falciparum has to gain mutations in dhfr and dhps
genes to develop resistance to SP, thus absence of dhps
mutation in Yemen suggest that the SP will be effective
for some years to come. Nonetheless, the occurrence of
high level resistance dhfr and dhps genotypes linked to
SP failure in areas close to Yemen, such as Iran and
Pakistan (100%), the Horn of Africa; Sudan (72.7%) and
Ethiopia (84.5%) [60-64], necessitate the need for regular
surveys to monitor possible emergence of these muta-
tions into Yemen for timely change in drug policy. The distribution of drug resistant genotypes seen in
the present study fits with the history of anti-malarial
usage, and possible migration of parasites from Africa
and Asia into Yemen. The emergence of CQR in Yemen
in 1989 Yemen [2,7,9,38,39], coincided with its appear-
ance in close countries in East Africa [12,16,40] and
Asia, Saudi Arabia [3], Iran and Pakistan [3,41-46]. The
presence of two distinct pfcrt genotype, CVIET (89%)
and SVMNT (4%) suggests that CQR in Yemen has
evolved from at least two different origins. Globally,
there are five genotypes of pfcrt CVIET, SVMNT,
SVIET, CVMNT and CVTNT [47]. The CVIET geno-
type is predominant in Africa [48], while the SVMNT is
common in Asian countries close to the Arabian Penin-
sula, such as Iran [43], India [49] and Pakistan [41,42]. This agrees with the hypothesis of frequent gene flow of
African/Asian parasites into Yemen, and suggests that
the CQR genotype (CVIET) is probably introduced via
Africa, while genotype SVMNT has come from Asia. Analysis of microsatellites around pfcrt will allow further
investigation to the origin of pfcrt resistance genotypes
[29,48]. Similar to pfcrt, high prevalence of pfmdr1 mutations was
seen in Yemen. Mutations in codon 86Y has been linked to
CQR, while those at codons 184, 1034, 1042 and 1246
are related to mefloquine (MQ), amodiaquine (AQ),
halofantrine (HF) and quinine resistance [32,50,51]. In
the present study, 86 (80%), 107 (99%) and 108 (100%)
of the examined P. falciparum isolates carried alleles
86N and 184F and 1246D, respectively. Discussion Malaria Journal 2013, 12:244
http://www.malariajournal.com/content/12/1/244 Page 6 of 8 allele 86 N (80%) in the present study cannot be
explained by withdrawal of CQ in Yemen, as the drug
is still in use [55], and pfcrt76T remains at a prevalence
of 89%. It is therefore, more likely that the pattern of
pfmdr1 polymorphisms, seen in Yemen, is due to
resurgence in use of artesunate-SP combination. In
addition to codon 86 and 184, mutations at the codons
1034 and 1042 were also high in Yemen; nonetheless
no mutation was seen at codon 1246. Further longitu-
dinal surveillance of pfmdr1 mutations, coupled with
in vivo testing, should be considered to examine the impact
of these polymorphisms on efficacy of ACT in Yemen. diversity. However, the lack of genetic differentiation be-
tween parasites in different sites suggests that the pre-
vailing epidemiological and demographical factors in
Yemen are favorable to parasite dispersal. Imported mal-
aria can, therefore, jeopardize the current success of
control in the region despite the presence of a strong
public health infrastructure. For example, countries such
as Oman experience regular epidemics due to imported
malaria [11]. To sustain malaria control and achieve
elimination in the peninsula efforts should be directed
to foci with on-going transmission, such as Yemen and
Saudi Arabia. p y
p
y
With regard to genes controlling P. falciparum re-
sponse to SP, a previous study has detected the mutant
allele dhfr-59R among 4 out of 99 isolates in Lahj gover-
norate, southeast of Yemen, and no information was
given on other codons 51, 108 and 164 [10]. However,
the present study revealed high prevalence of mutations
at codons 51 and 108 of the dhfr, while no mutation was
seen at codon 59. This inconsistency can be attributed
to the probability that some of the samples examined by
Mubjer et al. [10] may have been collected from expatri-
ates. If mutation 59R exists among local parasites in
Yemen, it would have increase in frequency following re-
cent shift to us of artesunate + sulphadoxine-pyrimeth-
amine (SP) as first line therapy [55]. Epidemiological
evidence suggests that dhfr mutations starts at codon
108, yielding low level resistance to pyrimethamine [38],
then the resistance increases with acquisition of extra
mutations at codons 51 or 59 and 164 [56]. Analysis of
evolution pattern of dhfr genotypes shows that the high
resistance genotype (51I, 59R, 108 N) accumulates mu-
tations in sequence. Discussion Selection of
pfmdr1 86N and 184F in recrudescent P. falciparum
parasites following lumefantrine suggested a possible
role of these alleles in the development of tolerance/re-
sistance to lumefantrine [37,52]. Increased prevalence
of pfmdr1 86N, 184F and 1246D haplotype was seen in
Zanzibar
after
several
years
of
extensive
use
of
artesunate-amodiaquine (ASAQ) [53]. A recent study
has confirmed that P. falciparum parasites with the
pfmdr1 N86/184F/D1246 haplotype can withstand 15-
fold higher lumefantrine blood concentrations com-
pared to those with the 86Y/Y184/1246Y haplotype
[54]. The relatively high prevalence of the wild-type Discussion Many sites in the Arabian Peninsula turned into “mal-
aria-free” following intensive control efforts mounted
over a relatively short period of time [1]. In contrast, in
Yemen and southwest Saudi Arabia malaria remains re-
silient to control efforts. Some of the challenges facing
the prospect of elimination, in these sites, is the intro-
duction of drug resistance parasites via asymptomatic
carriers. The present study examined the extent of gen-
etic diversity and distribution of drug-resistant genes
among of P. falciparum in some major foci in Yemen. g
f
j
A high degree of diversity of non-drug-resistant genes
(msp-2 and pfg377 and microsatellites in Chr 8) was seen
among P. falciparum in Yemen. The He index ranged
between 0.51 and 0.89, suggesting a high rate of trans-
mission and a bigger parasite population size than
anticipated in this region. Both allelic diversity and
prevalence of mixed-genotype infection were high, 57%
of the isolates harboured more than one allele at any of
the examined loci, with a mean of 1.8 genotypes per
infected individual. Assuming that each clone is readily
transmissible to mosquito, the rate of cross-mating and
subsequent recombination is expected to be high [25]. This agrees with the observed random association be-
tween loci and absence of geographical differentiation
among
P. falciparum
in
the
three
sites
(Dhamar,
Hodeidah and Taiz) (Fst < 0.05). Such a pattern of para-
site structure is similar to that seen among P. falciparum
populations in Africa [29,30]. These findings imply pres-
ence of a large effective population size (Ne) of P. falcip-
arum in Yemen, as there is a direct relationship between
the level of diversity and Ne [31-33]. This represents a
challenge to control efforts in the whole region, as para-
sites can readily migrate from Yemen into other sites in
the Arabian Peninsula, such as Oman [11], UAE [34]
and Saudi Arabia [35], where transmission has been
interrupted. In addition to local foci, imported malaria,
via numerous migrations from Africa/Asia, can enrich
diversity of parasites in the Arabian Peninsula. Over the
past two decades political instability has resulted in mass
movement of people from the Horn of Africa (Somalia,
Eritrea and Ethiopia) into Yemen and Saudi Arabia
[5]. Travellers from malaria-endemic areas can carry
long-lasting, transmissible, asymptomatic parasitaemia
[36,37]. Multiple
introductions
of
distinct
genetic
sources could in part explain the high level of parasite Al-Hamidhi et al. Additional file 1: Alleles of 3 microsatellites (MS) on chromosomes
8, msp-2 and pfg377 genes among 108 Plasmodium falciparum
isolates in Yemen. References
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http://www.malariajournal.com/content/12/1/244 Author details
1 1Department of Biochemistry, Faculty of Medicine and Health Sciences,
Sultan Qaboos University, Alkhod, PO Box 35, Muscat, Oman. 2Department of
Microbiology and Immunology, Faculty of Medicine and Health Sciences,
Sultan Qaboos University, Muscat, Oman. 3Department of Parasitology,
Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia. 4Department of Parasitology, Faculty of Medicine, Sana’a University, Sana’a,
Yemen. 5Research Centre in Biodiversity and Genetic Resources (CIBIO),
University of Porto, Rua Padre Armando Quintas 7, Vairão, Portugal. 6Institute
of Immunology and Infection Research, School of Biological Sciences,
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LMB, bir Singh Sidhu A, Naudé B, Deitsch KW, Su XZ, Wootton JC, Roepe
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k
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h We acknowledge the support of technical staff of Biochemistry Department,
College of Medicine and Health Sciences, Sultan Qaboos University, Oman,
and staff of the Children Sweden Hospital Taiz, Yemen. This study was
supported by Sultan Qaboos University, Oman, Project IG/MED/BIOC/09/03. 19. Plowe CV, Djimde A, Bouare M, Doumbo O, Wellems TE: Pyrimethamine
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and take full advantage of: 50. Reed MB, Sallba KJ, Caruana SR, Kirk K, Cowman AF: Pgh1 modulates
sensitivity and resistance to multiple antimalarials in Plasmodium
falciparum. Nature 2000, 403:906–909. • Convenient online submission 51. Danquah I, Coulibaly B, Meissner P, Petruschke I, Müller O, Mockenhaupt FP:
Selection of Pfmdr1 and Pfcrt alleles in amodiaquine treatment failure in
north-western Burkina Faso. Acta Trop 2010, 114:63–66. • Thorough peer review 52. Sisowath C, Ferreira PE, Bustamante LY, Dahlström S, Mårtensson A,
Björkman A, Krishna S, Gil JP: The role of Pfmdr1 in Plasmodium falciparum
tolerance to artemether-lumefantrine in Africa. Trop Med Int Health 2007,
12:736–742. 53. Fröberg G, Jörnhagen L, Morris U, Shakely D, Msellem M, Gil J, Björkman A,
Mårtensson A: Decreased prevalence of Plasmodium falciparum resistance
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Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults
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JAMA neurology
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Effect of an 18-Month Meditation Training on Regional
Brain Volume and Perfusion in Older Adults
The Age-Well Randomized Clinical Trial Gael Chételat, PhD; Antoine Lutz, PhD; Olga Klimecki, PhD; Eric Frison, MD, PhD; Julien Asselineau, MSc;
Marco Schlosser, MSc; Eider M. Arenaza-Urquijo, PhD; Florence Mézenge, MSc; Elizabeth Kuhn, PhD;
Inès Moulinet, PhD; Edelweiss Touron, MSc; Sophie Dautricourt, PhD; Claire André, PhD; Cassandre Palix, MSc
Valentin Ourry, PhD; Francesca Felisatti, MSc; Julie Gonneaud, PhD; Brigitte Landeau, MSc;
Géraldine Rauchs, PhD; Anne Chocat, MD; Anne Quillard, MD; Eglantine Ferrand Devouge, MD;
Patrik Vuilleumier, MD; Vincent de La Sayette, MD; Denis Vivien, PhD; Fabienne Collette, PhD;
Géraldine Poisnel, PhD; Natalie L. Marchant, PhD; for the Medit-Ageing Research Group IMPORTANCE No lifestyle-based randomized clinical trial directly targets psychoaffective risk
factors of dementia. Meditation practices recently emerged as a promising mental training
exercise to foster brain health and reduce dementia risk. OBJECTIVE To investigate the effects of meditation training on brain integrity in older adults. DESIGN, SETTING, AND PARTICIPANTS Age-Well was a randomized, controlled superiority trial
with blinded end point assessment. Community-dwelling cognitively unimpaired adults 65
years and older were enrolled between November 24, 2016, and March 5, 2018, in France. Participants were randomly assigned (1:1:1) to (1) an 18-month meditation-based training, (2) a
structurally matched non-native language (English) training, or (3) no intervention arm. Analysis took place between December 2020 and October 2021. INTERVENTIONS Meditation and non-native language training included 2-hour weekly group
sessions, practice of 20 minutes or longer daily at home, and 1-day intensive practices. MAIN OUTCOMES AND MEASURES Primary outcomes included volume and perfusion of
anterior cingulate cortex (ACC) and insula. Main secondary outcomes included a global
composite score capturing metacognitive, prosocial, and self-regulatory capacities and
constituent subscores. RESULTS Among 137 participants (mean [SD] age, 69.4 [3.8] years; 83 [60.6%] female; 54
[39.4%] male) assigned to the meditation (n = 45), non-native language training (n = 46), or
no intervention (n = 46) groups, all but 1 completed the trial. There were no differences in
volume changes of ACC (0.01 [98.75% CI, −0.02 to 0.05]; P = .36) or insula (0.01 [98.75% CI,
−0.02 to 0.03]; P = .58) between meditation and no intervention or non-native language
training groups, respectively. Differences in perfusion changes did not reach statistical
significance for meditation compared with no intervention in ACC (0.02 [98.75% CI, −0.01 to
0.05]; P = .06) or compared with non-native language training in insula (0.02 [98.75% CI,
−0.01 to 0.05]; P = .09). JAMA Neurol. doi:10.1001/jamaneurol.2022.3185
Published online October 10, 2022. Author Affiliations: Author
affiliations are listed at the end of this
article.
Group Information: The
Medit-Ageing Research Group is
listed in Supplement 3.
Corresponding Author: Gael
Chételat, PhD, INSERM Unité U1237,
GIP CYCERON, Bd Henri Becquerel –
BP 5229, 14074 Caen Cedex 5, France
(chetelat@cyceron.fr). Visual Abstract
Supplemental content Research Research Author Affiliations: Author
affiliations are listed at the end of this
article. Downloaded From: https://jamanetwork.com/ on 10/10/2022 Study Design and Participants The design and method of Age-Well have been described
previously.29 The trial protocol and statistical analysis plan are
available in Supplement 1. Briefly, our study was an 18-
month monocentric, randomized, observer-blind controlled
superiority clinical trial with 3 parallel groups: 1 group with a
meditation-based training (intervention group), 1 group with
structurally matched non-native language training (English
learning = active control group), and 1 group with no interven-
tion (passive control group). Participants fulfilling eligibility
criteria were invited to the baseline preintervention visit and
then randomized. Participants were enrolled between Novem-
ber 24, 2016, and March 5, 2018. The 18-month intervention
period started just after randomization, and participants had
amidinterventionvisitat9monthsandapostinterventionvisit
at the end of the intervention. Participants were 65 years or
older,community-dwelling,nativeFrenchspeakers,retiredfor
at least 1 year, had 7 years or more of education, and per-
formed within the normal range for age and educational level
on standardized cognitive tests (see Tables 1 and 2 in Poisnel
et al29 for details). They had no evidence of major neuro-
logical or psychiatric disorders, no history of cerebrovascular
disease, chronic disease or acute unstable illness, and no cur-
rent medication that could interfere with cognitive function-
ing. Full eligibility criteria are listed in eAppendix 1 in Supple-
ment 2. All participants gave their written informed consent
to participate in the study. The Age-Well RCT, sponsored by
Institut National de la Santé et de la Recherche Médicale
(INSERM), was approved by the ethics committee (CPP Nord-
Ouest III, Caen). Thus, meditation appears to be a promising approach to
preserve brain structure and function as well as cognition
and thus to reduce dementia risk by directly targeting psy-
choaffective factors. Faced with methodological limitations
in previous or ongoing studies, the Age-Well RCT of the
Medit-Aging European project was designed to investigate
the impact of an 18-month meditation intervention on the
volume and perfusion of the ACC and insula (coprimary out-
comes) compared with active (non-native language training)
and passive (no intervention) control groups, respectively,
and on a self-report–based global composite score capturing
attention regulation, socioemotional, and self-knowledge
capacities and its constituent subscores (main secondary
outcomes) compared with the active control group to test
for meditation-specific effects. Effect of an 18-Month Meditation Training on Regional
Brain Volume and Perfusion in Older Adults
The Age-Well Randomized Clinical Trial Meditation was superior to non-native language training on
18-month changes in a global composite score capturing attention regulation,
socioemotional, and self-knowledge capacities (Cohen d, 0.52 [95% CI, 0.19-0.85]; P = .002). CONCLUSIONS AND RELEVANCE The study findings confirm the feasibility of meditation and
non-native language training in elderly individuals, with high adherence and very low
attrition. Findings also show positive behavioral effects of meditation that were not reflected
on volume, and not significantly on perfusion, of target brain areas. Author Affiliations: Author
affiliations are listed at the end of this
article. Group Information: The
Medit-Ageing Research Group is
listed in Supplement 3. Corresponding Author: Gael
Chételat, PhD, INSERM Unité U1237,
GIP CYCERON, Bd Henri Becquerel –
BP 5229, 14074 Caen Cedex 5, France
(chetelat@cyceron.fr). TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT02977819 JAMA Neurol. doi:10.1001/jamaneurol.2022.3185
Published online October 10, 2022. E1 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults S
t
l
s S
trategies to prevent dementia are urgently needed. In
later life, the main risk factors for dementia include
smoking, depression, social isolation, physical inactiv-
ity, air pollution, and diabetes.1 There is also evidence for other
potentially modifiable risk factors, including hearing loss, poor
diet, anxiety, neuroticism, repetitive negative thinking, and
sleep disorders.1-3 Recent and ongoing lifestyle-based multi-
domain interventions thus include cognitive stimulation,
physicalactivity,diet,andcardiovascularrecommendations.4-6
However, psychoaffective risk factors, including depression,
stress, and anxiety have not been directly targeted, to our
knowledge. Findings I Findings In this randomized clinical trial that included 137
cognitively unimpaired community-dwelling older individuals, the
18-month meditation-based intervention did not significantly
modify the volume of the anterior cingulate cortex and insula
compared with a passive or active control, respectively. Meaning Future analyses on secondary outcomes will determine
the measures most sensitive to meditation training and the factors
associated with responsiveness to the intervention. Mental training that targets stress and attention regula-
tion has the potential to improve both cognitive and emo-
tional aspects of aging.7,8 Previous studies have shown that
mindfulness meditation improves cognition, specifically
in older adults across multiple domains including atten-
tion, executive functions, and self-awareness or meta-
cognition.9-12 Mindfulness meditation can also reduce
stress, anxiety, and depression,13-15 including in older
adults.16 Moreover, meditation in young adults has been
associated with brain structural and functional changes
mainly in frontal and limbic networks,10,17 with the insula
and anterior cingulate cortex (ACC) being the most sensitive
regions to meditation training according to a recent
meta-analysis.18 These interconnected brain regions
form the salience network and are particularly involved
in self-awareness,19,20 attentional, emotional and empathic
processing, 1 5 and self-regulation of attention and
emotion.20,21 They are consistently reported in task-related
functional magnetic resonance imaging (MRI) studies on
meditation,10,17 showing increased activity during mindful-
ness and/or compassion meditation.7 Interestingly, these
brain areas are also particularly sensitive to aging.22-24 Stud-
ies on meditation in elderly populations are sparse, with
high risk of bias, and there are no randomized clinical trials
(RCTs) with large samples. The 3 RCTs in older adults with
neuroimaging end points used 8-week training and reported
conflicting findings.25-27 Two cross-sectional studies assess-
ing elderly expert meditators showed more age-preserved
gray matter volume and/or glucose metabolism in various
brain regions including the ACC and insula compared with
nonmeditators.22,28 Key Points Question Could meditation, a mental training approach toward
attention and emotion regulation, preserve brain structure and
function in cognitively unimpaired older adults? Study Design and Participants Detailed biological, behavioral, neuroimaging, and sleep
measures were collected in Caen, France, at the preinterven-
tion and postintervention visits (the full list is available from
Poisnel et al29 and eAppendix 2 in Supplement 2), including
the structural T1-weighted MRI and early 18F-florbetapir
(Amyvid;LilyDiagnostics)positronemissiontomography(PET)
scan, used for the primary outcomes, and the self-reported
measuresusedtocomputetheglobalcompositescoreandsub-
scores used as the main secondary outcomes.30 JAMA Neurology
Published online October 10, 2022 jamaneurology.com jamaneurology.com E2 JAMA Neurology
Published online October 10, 2022 Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Original Investigation Research Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Sample Size
h
i l The trial was powered to detect a relevant effect size of 0.75
for the meditation intervention, as suggested by a meta-
analysis of meditation effects on morphometric neuroimag-
ing markers.17 To detect an effect size of 0.75 for each of the 4
comparisons, with 80% power and a 2-sided type I error of
1.25% (Bonferroni correction for test multiplicity), 42 partici-
pantsperarm(126intotal)wererequired.Toaccountforanaly-
ses on secondary outcomes, a total sample size of 150 partici-
pants was fixed. A statistical analysis plan was developed and
validated by the trial steering committee before database lock
and analyses. Outcomes Measures The coprimary outcomes were (1) for the comparison be-
tween meditation and no intervention groups, the 18-month
changes in ACC volume and perfusion (from pre- to postinter-
vention) and (2) for the comparison between meditation and
non-native language training groups, the 18-month changes
in insula volume and perfusion. These brain regions were se-
lected as they are known to be particularly sensitive both to
aging and to meditation practice (see Introduction and Lutz
et al7). The ACC was expected to be modified by both inter-
ventions (compared with no intervention), as this structure is
considered a brain signature of cognitive reserve and brain
maintenanceingeneral,31andtobemoreresistanttoagingwith
bilingual experience in particular.32 In contrast, the insula was
expected to be modified specifically in the meditation com-
pared with the non-native language training group, given its
role in emotional and empathic processing.33 Interventions The meditation and non-native language training interven-
tionswerestructurallyequivalentinoverallcourselength,class
time,andhomeactivities,aswellaslevelofexpertiseandnum-
ber of teachers per class. Thus, they had 2-hour weekly group
sessions,dailyhomepractice(minimum20minutes),and1-day
intensive practice (5 hours) during the intervention. During the
study, participants were strongly encouraged not to practice
the activity proposed in the other group(s). For each intervention, a manual describing the detailed
procedure was written before the study started. Both inter-
ventions have been described in detail previously.29 Briefly,
the meditation intervention consisted of a secular program of
meditation training labeled the Silver Santé Study Medita-
tion Programme especially designed for this study, based
on preexisting interventions, and composed of mindfulness
and loving kindness and compassion meditations. The non-
native language training program consisted of English lan-
guage exercises designed to reinforce each participant’s abili-
ties in comprehension, writing, and speaking. Participants in
the passive control group were requested not to change their
habits, ie, continue living as they used to before entering the
study. Adverse events and serious adverse events were re-
corded throughout the study when reported by the partici-
pants and systematically at each study visit during a consul-
tation with a physician. Randomization and Masking scans, respectively. Note that while early-phase florbetapir has
been shown to be highly correlated with fludeoxyglucose-,
arterial spin‐labeled–, and H2O-PET, it remains a surrogate
marker of perfusion and glucose metabolism. The procedure is
detailed in eAppendix 3 in Supplement 2 and described
elsewhere.34Briefly,T1-weightedimagesweresegmented,nor-
malized,andcorrectedfornonlinearwarpingsothatvalueswere
corrected for brain size. Early 18F-florbetapir-PET images were
normalized and scaled by the white matter to obtain standard-
ized uptake values ratio. Averaged gray matter volume and
perfusionvalueswereextractedfromtheresultingimagesinour
2 regions of interest. Eligible participants from each of the 3 waves were random-
ized after their baseline assessment. They were randomly as-
signed (1:1:1) to the meditation, non-native language training
(active control), or no intervention (passive control) arm
according to a randomization list with permuted blocks of
varying size (6 and 9), which was generated centrally by
a biostatistician at the EUCLID clinical trials platform. All
study personnel, including the investigators and outcome
assessors, were masked to treatment allocation. Only the
meditation and non-native language teachers and the trial-
independent statisticians and data monitoring infrastructure
staff were unmasked or partially unmasked. See further de-
tails in eAppendix 3 in Supplement 2. The main secondary outcomes have been defined after
trial commencement so as to select, among the huge amount
of secondary outcomes (eAppendix 2 in Supplement 2), the
most relevant according to the goal and hypotheses of the
Age-Well RCT. They consisted of the comparison between
meditation and non-native language training on changes from
baseline to 18 months in a self-report–based global composite
score and its constituent subscores capturing (1) attention
regulation (ie, attentional subscore), (2) socioemotional
capacities (ie, constructive subscore), and (3) self-knowledge
capacities to understand one’s own psychological processes
(ie, deconstructive subscore). This global score and its sub-
scores, recently operationalized,30,35 draw on self-report mea-
sures used in Age-Well (eAppendix 4 in Supplement 2).29 Each
composite subscore was computed by averaging the z scores
of the scales that were assigned to the respective composite,
while the global composite score corresponded to the mean of
the 3 composite subscores. Statistical Analyses The primary outcome analysis was performed according to
an intention-to-treat principle, including all randomized par-
ticipants, as planned in the statistical analysis plan. Missing
data on neuroimaging markers were handled using a miss-
ing = failure strategy, where missing values were replaced by
the most detrimental value of change observed in all groups
combined for a given outcome. Outcomes were compared be-
tween groups using a linear regression model adjusted for
baseline prognostic factors (sex, median-centered age, level The ACC and insula volume and perfusion measures were
obtained from T1-weighted MRI and early 18F-florbetapir PET JAMA Neurology
Published online October 10, 2022 jamaneurology.com E3 jamaneurology.com Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Figure 1. Trial Profile
900 Individuals prescreened after recruiting through public advertising
157 Assessed for eligibility
20 Excluded
9 Had clinically significant diagnostic battery findings
4 Had a chronic disease
2 Not available for the trial duration
1 Had history of cerebral disease
1 Had contraindication to MRI
1 Experienced claustrophobia
1 Had hypersensitivity to 18F-florbetapir
1 Withdrew from the trial
137 Randomized
46 Allocated to no intervention arm
45 Received allocated intervention
1 Received non-native language
training intervention
46 Analyzed at 18 mo
1 Died
46 Allocated to non-native language
training arm
45 Received allocated intervention
45 Analyzed at 18 mo
1 Excluded
45 Allocated to meditation arm
45 Received allocated intervention
45 Analyzed at 18 mo
Of 137 randomized participants, 1 was
excluded from all analyses due to
major eligibility criteria not met (not
included in the analyses), 1 died
during follow-up, and 1 revealed not
to have followed his allocated arm
(randomized to no intervention but
attended non-native language
training); those 2 later participants
were retained in the analyses and
treated by the intention-to-treat
principle, as specified in the statistical
analysis plan. MRI indicates magnetic
resonance imaging. Research Original Investigation
Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Of 137 randomized participants, 1 was
excluded from all analyses due to
major eligibility criteria not met (not
included in the analyses), 1 died
during follow-up, and 1 revealed not
to have followed his allocated arm
(randomized to no intervention but
attended non-native language
training); those 2 later participants
were retained in the analyses and
treated by the intention-to-treat
principle, as specified in the statistical
analysis plan. MRI indicates magnetic
resonance imaging. 46 Allocated to no intervention arm
45 Received allocated intervention
1 Received non-native language
training intervention of education, and Mini-Mental State Examination) and base-
line outcome value. Comparisons were made with a 1.25 type
I error rate. Additional analyses were performed, including a
sensitivity analysis to missing data, a “minimum interven-
tion” analysis, and post hoc analyses for additional adjust-
ment, stratification, and subgroup analyses (eAppendix 3 in
Supplement 2). jamaneurology.com Results JAMA Neurology
Published online October 10, 2022 E4 jamaneurology.com Downloaded From: https://jamanetwork.com/ on 10/10/2022 [39.4%] male), among whom 1 participant was excluded from
all analyses because of major eligibility criteria not met (deci-
sion by the trial steering committee blinded to group alloca-
tion for head trauma with loss of consciousness >1 hour;
Figure 1). Among 136 participants included in the analyses, 45
wererandomizedtomeditation,45tonon-nativelanguagetrain-
ing, and 46 to the no intervention groups. One participant died
duringfollow-up,whileanotherparticipantwasrevealedtohave
notfollowedhisallocatedarm;bothwereretainedintheanaly-
sesandincludedusingtheintention-to-treatprinciple,asspeci-
fied in the statistical analysis plan. Forthesecondaryoutcomes,thestatisticalanalysisplanfo-
cused on between-group differences in mean changes in the
global composite score and subscores in meditation vs non-
native language training so as to highlight the specific effects of
meditation compared with its active control. For each compos-
ite score and subscore, we built 1 mixed-effect linear regression
model incorporating data from pre- and postintervention with
aninteractiontermbetweenvisitandgroup,controllingforbase-
linescoresoftheoutcome.Inallmixed-effectsregressionmod-
els, missing data were not replaced and assumed to be missing
at random. Baseline characteristics of the 136 participants included in
the intention-to-treat analysis are detailed in Table 1. There
were no major clinical differences in any demographic or clini-
cal characteristics between groups. The median (IQR) fol-
low-up time between pre- and postintervention visits was 21.1
(20.7-21.7) months. The mean (SD) class attendance for medi-
tation and non-native language groups was 62.0 (8.9) and 57.9
(12.2) of 72 classes, respectively. Of 45 participants, 44 in the
meditation/non-native language groups attended at least 20%
of their intervention classes. AnalyseswereperformedusingSASstatisticalsoftwarever-
sion 9.4 (SAS Institute) for the coprimary outcomes and R ver-
sion 4.0.2 (R Foundation) for the secondary outcomes. Two-
sidedPvalueswerestatisticallysignificantatlessthan.0125for
each coprimary outcome (Bonferroni correction for test multi-
plicity)andlessthan.05forsecondaryoutcomes.Analysistook
placebetweenDecember2020andOctober2021(andApril2022
for the supplementary analyses during the revision process). For the coprimary outcomes, data were missing on base-
line early 18F-florbetapir-PET scans due to extravasation and
technical problems for 2 participants (meditation, n = 1 and no
intervention, n = 1), and data were missing on 18-month MRI
and early 18F-florbetapir-PET scans for 1 participant in the no
intervention group who died during follow-up. Results
Of 157 participants assessed for eligibility, 137 were random-
ized (mean [SD] age, 69.4 [3.8] years; 83 [60.6%] female; 54 Primaryendpointsatbaselineand18monthsandtheirmean
changes are reported in the eTable in Supplement 2, and inter-
vention effects on the coprimary outcome measures in the in-
tention-to-treat analyses are presented in Figure 2. The differ-
ences in the mean volume changes over 18 months between Of 137 randomized participants, 1 was
excluded from all analyses due to
major eligibility criteria not met (not
included in the analyses), 1 died
during follow-up, and 1 revealed not
to have followed his allocated arm
(randomized to no intervention but
attended non-native language
training); those 2 later participants
were retained in the analyses and
treated by the intention-to-treat
principle, as specified in the statistical
analysis plan. MRI indicates magnetic
resonance imaging. Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 ffect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Original Investigation Research Original Investigation Research Table 1. Baseline Characteristics
Characteristic
Mean (SD)
Meditation
(n = 45)
Non-native
language training
(n = 45)
No intervention
(n = 46)
Age, median (IQR), y
68 (67-72)
69 (67-73)
68 (66-70)
Female, No. (%)
31 (69)
24 (53)
28 (61)
Male, No. (%)
14 (31)
21 (47)
18 (39)
Years of education
13 (3)
12 (3)
14 (2)
Years in retirement
7 (5-13)
9 (5-13)
6 (4-10)
Handedness laterality
92 (83-100)
91 (83-100)
92 (83-100)
BP level, mm Hg
Systolic
143.6 (20.1)
145.3 (16.4)
136.7 (20.5)
Diastolic
86.1 (10.5)
86.6 (10.0)
84.8 (11.7)
BMI
26.1 (4.6)
26.5 (4.3)
25.9 (4.0)
Mini-Mental State Examination score
28.9 (1.2)
29.0 (1.0)
29.2 (0.9)
Montgomery–Åsberg Depression Rating Scale score
1.2 (1.2)
1.2 (1.5)
0.7 (1.0)
APOE ε4 carriers (≥1 allele), No. (%)
13 (29)
13 (29)
11 (24)
Positive for brain amyloid,a No. (%)
11 (24)
11 (24)
6 (13)
Amyloid SUVR
1.26 (0.16)
1.27 (0.19)
1.21 (0.10)
Familial history of dementia, No. (%)
14 (31)
11 (24)
14 (30)
Presence of ≥1 Alzheimer disease risk factors,b No. (%)
28 (62)
23 (51)
23 (51)
Abbreviations: APOE, apolipoprotein
E; BMI, body mass index (calculated
as weight in kilograms divided by
height in meters squared); BP, blood
pressure; SUVR, standardized uptake
values ratio. a Methods detailed in eAppendix 3 in
Supplement 2. bAlzheimer disease risk factors
include APOE ε4 (1 allele), brain
amyloid positivity, and familial
history of dementia. Figure 2. Forest Plots for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Meditation vs no intervention for ACC
Meditation vs non-native language
training for insula
Favors
the other
Favors
meditation
ACC and insula volumes
A
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Meditation vs no intervention for ACC
Meditation vs non-native language
training for insula
Favors
the other
Favors
meditation
ACC and insula perfusion
B
β (98.7% CI)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.05)
β (98.7% CI)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
Results of the intention-to-treat
analyses with missing = failure
strategy are shown. Table 1. Downloaded From: https://jamanetwork.com/ on 10/10/2022 Baseline Characteristics
Characteristic
Mean (SD)
Meditation
(n = 45)
Non-native
language training
(n = 45)
No intervention
(n = 46)
Age, median (IQR), y
68 (67-72)
69 (67-73)
68 (66-70)
Female, No. (%)
31 (69)
24 (53)
28 (61)
Male, No. (%)
14 (31)
21 (47)
18 (39)
Years of education
13 (3)
12 (3)
14 (2)
Years in retirement
7 (5-13)
9 (5-13)
6 (4-10)
Handedness laterality
92 (83-100)
91 (83-100)
92 (83-100)
BP level, mm Hg
Systolic
143.6 (20.1)
145.3 (16.4)
136.7 (20.5)
Diastolic
86.1 (10.5)
86.6 (10.0)
84.8 (11.7)
BMI
26.1 (4.6)
26.5 (4.3)
25.9 (4.0)
Mini-Mental State Examination score
28.9 (1.2)
29.0 (1.0)
29.2 (0.9)
Montgomery–Åsberg Depression Rating Scale score
1.2 (1.2)
1.2 (1.5)
0.7 (1.0)
APOE ε4 carriers (≥1 allele), No. (%)
13 (29)
13 (29)
11 (24)
Positive for brain amyloid,a No. (%)
11 (24)
11 (24)
6 (13)
Amyloid SUVR
1.26 (0.16)
1.27 (0.19)
1.21 (0.10)
Familial history of dementia, No. (%)
14 (31)
11 (24)
14 (30)
Presence of ≥1 Alzheimer disease risk factors,b No. (%)
28 (62)
23 (51)
23 (51)
Abbreviations: APOE, apolipoprotein
E; BMI, body mass index (calculated
as weight in kilograms divided by
height in meters squared); BP, blood
pressure; SUVR, standardized uptake
values ratio. a Methods detailed in eAppendix 3 in
Supplement 2. bAlzheimer disease risk factors
include APOE ε4 (1 allele), brain
amyloid positivity, and familial
history of dementia. Figure 2. Forest Plots for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Meditation vs no intervention for ACC
Meditation vs non-native language
training for insula
Favors
the other
Favors
meditation
ACC and insula volumes
A
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Meditation vs no intervention for ACC
Meditation vs non-native language
training for insula
Favors
the other
Favors
meditation
ACC and insula perfusion
B
β (98.7% CI)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.05)
β (98.7% CI)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
Results of the intention-to-treat
analyses with missing = failure
strategy are shown. Figure 2. Forest Plots for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion Favors
meditation Favors
meditation Favors
meditation Favors
the other Results of the intention-to-treat
analyses with missing = failure
strategy are shown. jamaneurology.com Downloaded From: https://jamanetwork.com/ on 10/10/2022 meditation and no intervention in the ACC (0.01 [98.75% CI,
−0.02 to 0.05]) or the non-native language group in the insula
(0.01 [98.75% CI, −0.02 to 0.03]) were not statistically signifi-
cant (P = .36 and P = .58, respectively). As for perfusion, differ-
ences in the mean changes over 18 months, in favor of medita-
tion compared with no intervention in the ACC (0.02 [98.75%
CI,−0.01to0.05])andcomparedwithnon-nativelanguagetrain-
ing in the insula (0.02 [98.75% CI, −0.01 to 0.05]) did not reach
statistical significance (P = .06 and P = .09, respectively). ment wave nor the presence of risk factor(s) for Alzheimer
disease (apolipoprotein E ε4 genotype, brain amyloid positiv-
ity,familialhistoryofdementia,orpresenceofatleast1ofthese
risk factors) affected the results. Regardingsecondaryoutcomes,theglobalcompositescore
and attention regulation subscore were not computed for 1 par-
ticipant because of missing data at baseline on a subscale as-
signed to the attention regulation subscore. Secondary end
points at baseline and 18 months and their standardized mean
changes are reported in Table 2 and in eAppendix 6 and 7 in
Supplement 2. The differences in the mean changes over 18
months between meditation and non-native language training
in the global composite score (0.52 [95% CI, 0.19-0.85];
P = .002), attention regulation (0.38 [95% CI, 0.10-0.67];
P = .009),andsocioemotional(0.31[95%CI,0.06-0.57];P = .01)
subscores were all statistically significant, while it was not sta-
tisticallysignificantfortheself-knowledgesubscore(0.28[95%
CI, −0.01 to 0.58]; P = .06). Results of the sensitivity and post hoc analyses are pre-
sented in Figure 3 and in eAppendix 5 in Supplement 2. Briefly,
results were very similar to those of the main analyses, show-
ingnobetween-groupdifferencesinthevolumechangesofthe
ACC or insula. Changes in the perfusion of the ACC or insula
always favored meditation (compared with no intervention or
non-native language training, respectively), but the between-
group differences never reached statistical significance. Fi-
nally, the subgroup analyses showed that neither the recruit- JAMA Neurology
Published online October 10, 2022 E5 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Sensitivity Analyses for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.01 (–0.02 to 0.05)
0.00 (–0.03 to 0.04)
0.01 (–0.02 to 0.04)
0.01 (–0.02 to 0.04)
0.00 (–0.05 to 0.06)
0.02 (–0.03 to 0.07)
0.01 (–0.05 to 0.06)
0.02 (–0.02 to 0.05)
0.01 (–0.02 to 0.05)
ACC volume
A
Favors
no intervention
Favors
meditation Figure 3. Sensitivity Analyses for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion Adverse Events
One death was reported during study follow-up for a partici-
pant in the no intervention group (myocardial infarction,
not related to the study). A total of 170 adverse events
were recorded, 41 of which were considered serious
(meditation, 13; non-native language training, 15; no inter-
vention, 13). Among these, 7 adverse events (meditation, 3;
non-native language training, 3; no intervention, 1) were
judged to be related to study procedures (procedural com-
plication, 4 [scans had to be redone]; extravasation, 2;
asthenia, 1). No serious adverse event was related to the
intervention. Figure 3. Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults JAMA Neurology
Published online October 10, 2022
(Reprinted)
jamaneurology.com –0.10
0
0.10
0.05
β (98.75% CI)
–0.05
CI)
0 to 0.06)
2 to 0.04)
1 to 0.05)
1 to 0.05)
2 to 0.07)
3 to 0.06)
2 to 0.08)
1 to 0.05)
1 to 0.05)
Favors
no intervention
Favors
meditation –0.10
0
0.10
0.05
β (98.75% CI)
–0.05
)
to 0.03)
to 0.03)
5 to 0.03)
to 0.06)
4 to 0.03)
to 0.03)
to 0.03)
Favors non-native
language training
Favors
meditation Favors non-native
language training –0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.03 (–0.00 to 0.06)
0.02 (–0.01 to 0.05)
0.03 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
0.02 (–0.03 to 0.08)
0.01 (–0.05 to 0.05)
0.04 (–0.01 to 0.09)
0.02 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
Insula perfusion
D
Favors non-native
language training
Favors
meditation
β (98.75% CI)
a No missing outcome values for
participants included in the
comparison. Favors non-native
language training vention, 13). Among these, 7 adverse events (meditation, 3;
non-native language training, 3; no intervention, 1) were
judged to be related to study procedures (procedural com-
plication, 4 [scans had to be redone]; extravasation, 2;
asthenia, 1). No serious adverse event was related to the
intervention. Adverse Events One death was reported during study follow-up for a partici-
pant in the no intervention group (myocardial infarction,
not related to the study). A total of 170 adverse events
were recorded, 41 of which were considered serious
(meditation, 13; non-native language training, 15; no inter- One death was reported during study follow-up for a partici-
pant in the no intervention group (myocardial infarction,
not related to the study). A total of 170 adverse events
were recorded, 41 of which were considered serious
(meditation, 13; non-native language training, 15; no inter- jamaneurology.com E6 Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Sensitivity Analyses for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.03 (–0.00 to 0.06)
0.02 (–0.01 to 0.05)
0.03 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
0.02 (–0.03 to 0.08)
0.01 (–0.05 to 0.05)
0.04 (–0.01 to 0.09)
0.02 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
Insula perfusion
D
Favors non-native
language training
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.03)
–0.01 (–0.05 to 0.03)
0.03 (–0.01 to 0.06)
–0.00 (–0.04 to 0.03)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.03)
Insula volumea
C
Favors non-native
language training
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.03 (–0.00 to 0.06)
0.01 (–0.02 to 0.04)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
0.03 (–0.02 to 0.07)
0.01 (–0.03 to 0.06)
0.03 (–0.02 to 0.08)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
ACC perfusion
B
Favors
no intervention
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.01 (–0.02 to 0.05)
0.00 (–0.03 to 0.04)
0.01 (–0.02 to 0.04)
0.01 (–0.02 to 0.04)
0.00 (–0.05 to 0.06)
0.02 (–0.03 to 0.07)
0.01 (–0.05 to 0.06)
0.02 (–0.02 to 0.05)
0.01 (–0.02 to 0.05)
ACC volume
A
Favors
no intervention
Favors
meditation
a No missing outcome values for
participants included in the
comparison. jamaneurology.com Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Sensitivity Analyses for Anterior Cingulate Cortex (ACC) and Insula Volume and Perfusion
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.03 (–0.00 to 0.06)
0.02 (–0.01 to 0.05)
0.03 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
0.02 (–0.03 to 0.08)
0.01 (–0.05 to 0.05)
0.04 (–0.01 to 0.09)
0.02 (–0.01 to 0.06)
0.02 (–0.01 to 0.05)
Insula perfusion
D
Favors non-native
language training
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.03)
–0.01 (–0.05 to 0.03)
0.03 (–0.01 to 0.06)
–0.00 (–0.04 to 0.03)
0.01 (–0.02 to 0.03)
0.01 (–0.02 to 0.03)
Insula volumea
C
Favors non-native
language training
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.03 (–0.00 to 0.06)
0.01 (–0.02 to 0.04)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
0.03 (–0.02 to 0.07)
0.01 (–0.03 to 0.06)
0.03 (–0.02 to 0.08)
0.02 (–0.01 to 0.05)
0.02 (–0.01 to 0.05)
ACC perfusion
B
Favors
no intervention
Favors
meditation
–0.10
0
0.10
0.05
β (98.75% CI)
–0.05
Source
Missing data = failure
Extreme case 1
Extreme case 2
Minimum intervention
Adjusted by cohort
Cohort 1
Cohort 2
Cohort 3
Adjusted by amyloid
β (98.7% CI)
0.01 (–0.02 to 0.05)
0.00 (–0.03 to 0.04)
0.01 (–0.02 to 0.04)
0.01 (–0.02 to 0.04)
0.00 (–0.05 to 0.06)
0.02 (–0.03 to 0.07)
0.01 (–0.05 to 0.06)
0.02 (–0.02 to 0.05)
0.01 (–0.02 to 0.05)
ACC volume
A
Favors
no intervention
Favors
meditation
a No m
part
com
Research Original Investigation
Effect of an 18 Month Meditation Training on Regional Brain Volum Figure 3. Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Research Original Investigation Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Adverse Events
One death was reported during study follow-up for a partici-
pant in the no intervention group (myocardial infarction,
not related to the study). A total of 170 adverse events
were recorded, 41 of which were considered serious
(meditation, 13; non-native language training, 15; no inter-
vention, 13). Among these, 7 advers
non-native language training, 3; n
judged to be related to study proce
plication, 4 [scans had to be redo
asthenia, 1). No serious adverse ev
intervention. Figure 3. JAMA Neurology
Published online October 10, 2022 Discussion Results indicate that the 18-month meditation-based inter-
ventiondidnotsignificantlymodifythevolumeoftheACCand
insula in older adults compared with a passive or active con-
trol, respectively; the between-group differences did not reach
statistical significance for perfusion either. Regarding the main
secondary outcomes, there were significant effects of medi-
tation compared with non-native language training on the
global composite score reflecting attention regulation, socio-
emotional, and self-knowledge capacities and two-thirds of its
constituent subscores. The fact that no effects were found on anterior cingulate
and insula volumes in our study, despite being identified as
structures most sensitive to meditation,36 might indicate that
18 months of meditation training is not sufficient to alter the
effects of age on their volume. Meditation might alter volume
in younger and more plastic brains but not halt the age- and
disease-related brain volume decreases at older ages. While
larger brain volumes were observed in cross-sectional stud-
ies in older expert meditators vs nonmeditators, especially in
the ACC and insula,22 these differences might reflect medita-
tion effects in younger ages, intense meditation practice ac-
cumulated throughout the adult lifespan, and/or between-
groupdifferencesonothervariables,suchaslifestyle.Notethat
the lack of meditation effect on brain structure is consistent
with the very recent publication from a large and rigorously
controlled study.37 Regarding secondary outcomes, meditation was superior
to non-native language training on changing a global compos-
ite score and 2 of its subscores reflecting attention regulation
and socioemotional capacities. As this composite score and
its subscores are thought to measure core dimensions of
well-being,40 these findings suggest meditation training–
related effects on mental health and human flourishing. The
attention regulation subscore increased after meditation only;
in the context of meditation practices, this capacity allows a
heightened awareness and monitoring of the contents of
experience without becoming absorbed by them. Socioemo-
tional capacities decreased substantially after non-native lan-
guage training but not meditation training, suggesting that
the difference observed may be due to maintenance of skills
by meditation training. Regarding perfusion, both the differences in the anterior
cingulate between the meditation and no intervention groups,
and in the insula between the meditation and non-native lan-
guage training, were not statistically significant but close to
the threshold. Mean differences between groups, in favor of
meditation, were rather low (0.023 and 0.022, respectively)
butcouldrepresentasubstantialgaincomparedwiththemean
18-month change observed in the no intervention group (0.031
for both the ACC and insula). Discussion Assuming a linear loss over 18
months, the 74% and 71% reduction in rate of loss could trans-
late into about 13 months less loss over the course of the in-
tervention, in favor of the meditation group. One could hy-
pothesize that a larger sample size or longer follow-up time
would have yielded a significant effect of the intervention on
perfusion. Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults Original Investigation Research Table 2. Results From Mixed-Effects Models Assessing Change From Baseline to 18 Months
in Meditation and Non-Native Language Training in Composite Scoresa
Outcome
Standardized estimated change (95% CI)
Between-group difference in change
meditation vs non-native language
training
Meditation
Non-native language
training
Mean (95% CI)
P value
Global
0.43 (0.20 to 0.67)
−0.09 (−0.12 to 0.30)
0.52 (0.19 to 0.85)
.002
Attention regulation
capacities
0.48 (0.28 to 0.69)
0.10 (−0.10 to 0.30)
0.38 (0.10 to 0.67)
.009
Socioemotional
capacities
0.04 (−0.15 to 0.23)
−0.27 (−0.46 to −0.08)
0.31 (0.06 to 0.57)
.02
Self-knowledge
capacities
0.27 (0.07 to 0.48)
−0.01 (−0.22 to 0.20)
0.28 (−0.01 to 0.58)
.06
a All analyses were adjusted
baseline scores of the out
Positive (negative) estima
between-group differenc
increases (decreases) in c
scores in the meditation
intervention. Table 2. Results From Mixed-Effects Models Assessing Change From Baseline to 18 Months
in Meditation and Non-Native Language Training in Composite Scoresa The lack of significant effects on the coprimary outcomes
could also be related to the design of the study, eg, the use of
an RCT when the intervention, by its nature and duration,
strongly relies on the motivation, preferences, and adher-
ence of the participants. Alternative trial designs taking into
account patient preference might be particularly relevant in
thecontextofsuchnonpharmacologicalinterventions.38More-
over, as in most preventive trials, our population resulted in
being, through self-selection, enriched with healthy partici-
pants with high education and reserve, and low probability of
cognitive decline, which left limited room for lifestyle changes
and intervention-related improvements.39 Finally, the se-
lected coprimary outcomes were very specific neuroimaging
measures that show variability and significant but protracted
age-related changes from which it might be difficult to show
a deviation in a short period of time in terms of aging effects. Limitations and Strengths Our study has several limitations. The sample size was de-
fined based on an expected effect size of 0.75 on neuro-
imaging measures, which was likely overestimated as it was
defined from a meta-analysis including both preintervention-
postintervention studies and cross-sectional studies in long-
term meditation experts but also due to publication bias, con-
siderable methodological caveats in the existing literature at
that time, and general biases in estimating effect sizes in neu-
roimaging studies.17,41 We were not able to observe this effect
size in the present study and may have been underpowered
to observe a smaller but still clinically significant effect. More- JAMA Neurology
Published online October 10, 2022 jamaneurology.com jamaneurology.com E7 Downloaded From: https://jamanetwork.com/ on 10/10/2022 ARTICLE INFORMATION Author Affiliations: Normandie Univ, UNICAEN,
INSERM, U1237, Physiopathology and Imaging of
Neurological Disorders (PhIND), Institut Blood and
Brain @ Caen-Normandie, Cyceron, France
(Chételat, Arenaza-Urquijo, Mézenge, Kuhn,
Moulinet, Touron, Dautricourt, André, Palix, Ourry,
Felisatti, Gonneaud, Landeau, Rauchs, Chocat,
Quillard, Devouge, Vivien, Poisnel); Lyon
Neuroscience Research Center INSERM U1028,
CNRS UMR5292, Lyon 1 University, Lyon, France
(Lutz); Swiss Center for Affective Sciences,
Department of Neuroscience, University of Geneva
Medical School, Geneva, Switzerland (Klimecki,
Vuilleumier); EUCLID/F-CRIN Clinical Trials
Platform, INSERM, CHU Bordeaux, University of
Bordeaux, CIC1401-EC, Bordeaux, France (Frison,
Asselineau); Division of Psychiatry, University
College London, London, United Kingdom
(Schlosser, Marchant); Department of Psychology,
Faculty of Psychology and Educational Sciences,
University of Geneva, Geneva, Switzerland
(Schlosser); Barcelonabeta Brain Research Center,
Fundación Pasqual Maragall, Barcelona, Spain
(Arenaza-Urquijo); Normandie Univ, UNIROUEN,
Department of General Practice, Rouen, France
(Devouge); Rouen University Hospital, CIC-CRB
1404, F 76000, Rouen, France (Devouge); CHU
Caen-Normandie, Department of Neurology, Caen,
France (de La Sayette); CHU Caen-Normandie,
Department of Clinical Research, Caen, France
(Vivien); GIGA-CRC, In Vivo Imaging, Université
de Liège and Belgian National Fund for Scientific
Research, Liège, Belgium (Collette). y
Critical revision of the manuscript for important
intellectual content: Klimecki, Frison, Asselineau,
Schlosser, Arenaza-Urquijo, Mézenge, Kuhn,
Moulinet, Touron, Dautricourt, André, Palix, Ourry,
Felisatti, Gonneaud, Landeau, Rauchs, Chocat,
Quillard, Ferrand Devouge, Vuilleumier, Vivien,
Collette, Poisnel, Marchant. Statistical analysis: Frison, Asselineau, Schlosser. Obtained funding: Chételat, Lutz, Klimecki,
Arenaza-Urquijo, Vuilleumier, Vivien, Collette,
Poisnel, Marchant. Administrative, technical, or material support: Lutz,
Klimecki, Moulinet, Rauchs, Ferrand Devouge,
Vuilleumier, Vivien, Poisnel. Administrative, technical, or material support: Lutz,
Klimecki, Moulinet, Rauchs, Ferrand Devouge,
Vuilleumier, Vivien, Poisnel. Supervision: Chételat, Lutz, Frison, Vuilleumier,
de la Sayette, Poisnel, Marchant. Funding/Support: This study was funded by
European Union’s Horizon 2020 research and
innovation programme and Fondation d’Entreprise
MMA des Entrepreneurs du Futur. Supervision: Chételat, Lutz, Frison, Vuilleumier,
de la Sayette, Poisnel, Marchant. Role of the Funder/Sponsor: The funders had no
role in the design and conduct of the study;
collection, management, analysis, and
interpretation of the data; preparation, review, or
approval of the manuscript; and decision to submit
the manuscript for publication. Conflict of Interest Disclosures: Drs Chételat,
Lutz, Klimecki, Gonneaud, Poisnel, Collette, and
Marchant have received research support from the
European Union’s Horizon 2020 research and
innovation programme under grant 667696. Drs Chételat, Lutz, Kuhn, Moulinet, and André have
received support from their institution Institut
National de la Santé et de la Recherche Médicale
(Inserm). ARTICLE INFORMATION submitted work. Dr Touron reported grants from
Ministry of Higher Education and Research (thesis
grant, France) outside the submitted work. Dr André has received research support from Fonds
Européen de Développement Régional (payment
made to the institution). Dr Gonneaud reported
grants from Fondation Alzheimer and Fondation
de France, which covered her salary during the
conduct of the study. Dr Vuilleumier reported
grants from Swiss National Science Foundation and
European Commission H2020 during the conduct
of the study. Dr Poisnel reported grants and
nonfinancial support from INSERM during the
conduct of the study; grants from INSERM outside
the submitted work; and has participated in the
data safety monitoring board of the Age-Well trial
and to the executive committee of Medit-Ageing. Dr Marchant received grants from Alzheimer’s
Society and Medical Research Council (payment
made to the institution) outside the submitted
work. No other disclosures were reported. Acquisition, analysis, or interpretation of data:
Chételat, Lutz, Klimecki, Frison, Asselineau,
Schlosser, Arenaza-Urquijo, Mézenge, Kuhn,
Moulinet, Touron, Dautricourt, André, Palix, Ourry,
Felisatti, Gonneaud, Landeau, Rauchs, Chocat,
Ferrand Devouge, Vuilleumier, Collette, Poisnel,
Marchant. Open Access: This is an open access article
distributed under the terms of the CC-BY License. © 2022 Chételat G et al. JAMA Neurology. Ferrand Devouge, Vuilleumier, Collette, Poisnel,
Marchant. Drafting of the manuscript: Chételat, Lutz, Frison,
Asselineau, Schlosser, Mézenge, Landeau,
Vuilleumier, de la Sayette. Critical revision of the manuscript for important
intellectual content: Klimecki, Frison, Asselineau,
Schlosser, Arenaza-Urquijo, Mézenge, Kuhn,
Moulinet, Touron, Dautricourt, André, Palix, Ourry,
Felisatti, Gonneaud, Landeau, Rauchs, Chocat,
Quillard, Ferrand Devouge, Vuilleumier, Vivien,
Collette, Poisnel, Marchant. Statistical analysis: Frison, Asselineau, Schlosser. Obtained funding: Chételat, Lutz, Klimecki,
Arenaza-Urquijo, Vuilleumier, Vivien, Collette,
Poisnel, Marchant. Drafting of the manuscript: Chételat, Lutz, Frison,
Asselineau, Schlosser, Mézenge, Landeau,
Vuilleumier, de la Sayette. Downloaded From: https://jamanetwork.com/ on 10/10/2022 Research Original Investigation Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults over, our sample was not representative of the global aging
population as it included very healthy individuals (see above). Finally,early-phaseflorbetapirisanindirect,surrogatemarker,
of perfusion or fludeoxyglucose-PET, and the optimal process-
ing method (scaling and timeframe) is still unclear. ing whole-brain voxelwise analyses of gray matter volume and
perfusion, but also other brain and behavior modalities, will
allow us to determine the measures most sensitive to medi-
tation practice, and investigate the mechanisms of these
effects. Several strengths must also be noted. The primary end
points were measured blinded to allocated intervention, the
intervention was particularly long, we used both a passive and
a carefully matched active control condition, and the ex-
ceptionally high adherence and low attrition (1 of 137 partici-
pants only) demonstrates the appropriateness of the interven-
tions and feasibility of this approach in healthy and motivated
elderly. The Age-Well clinical trial includes many complemen-
tary biological and behavioral measures of mental health and
well-being. Future analyses on secondary outcomes, includ- Conclusions Future secondary analyses from this trial will allow assess-
ment of the impact of meditation on volume and perfusion
throughout the whole brain and on other measures, and will
examine factors (participants’ characteristics, additional out-
comes, intervention doses) associated with responsiveness to
the intervention. Acquisition, analysis, or interpretation of data:
Chételat, Lutz, Klimecki, Frison, Asselineau,
Schlosser, Arenaza-Urquijo, Mézenge, Kuhn,
Moulinet, Touron, Dautricourt, André, Palix, Ourry,
Felisatti, Gonneaud, Landeau, Rauchs, Chocat,
Ferrand Devouge, Vuilleumier, Collette, Poisnel,
Marchant. Ment Health. 2021;25(7):1181-1190. doi:10.1080/
13607863.2020.1793901 Effect of long-term omega 3
polyunsaturated fatty acid supplementation with or
without multidomain intervention on cognitive
function in elderly adults with memory complaints
(MAPT): a randomised, placebo-controlled trial. Lancet Neurol. 2017;16(5):377-389. doi:10.1016/
S1474-4422(17)30040-6 6. Andrieu S, Guyonnet S, Coley N, et al; MAPT
Study Group. Effect of long-term omega 3
polyunsaturated fatty acid supplementation with or
without multidomain intervention on cognitive
function in elderly adults with memory complaints
(MAPT): a randomised, placebo-controlled trial. Lancet Neurol. 2017;16(5):377-389. doi:10.1016/
S1474-4422(17)30040-6 19. Chong JSX, Ng GJP, Lee SC, Zhou J. Salience
network connectivity in the insula is associated
with individual differences in interoceptive
accuracy. Brain Struct Funct. 2017;222(4):1635-1644. doi:10.1007/s00429-016-1297-7 19. Chong JSX, Ng GJP, Lee SC, Zhou J. Salience
network connectivity in the insula is associated
with individual differences in interoceptive
accuracy. Brain Struct Funct. 2017;222(4):1635-1644. doi:10.1007/s00429-016-1297-7 20. Cauda F, D’Agata F, Sacco K, Duca S, Geminiani
G, Vercelli A. Functional connectivity of the insula in
the resting brain. Neuroimage. 2011;55(1):8-23. doi:
10.1016/j.neuroimage.2010.11.049 7. Lutz A, Chételat G, Collette F, Klimecki OM,
Marchant NL, Gonneaud J. The protective effect of
mindfulness and compassion meditation practices
on ageing: Hypotheses, models and experimental
implementation. Ageing Res Rev. 2021;72:101495. doi:10.1016/j.arr.2021.101495 7. Lutz A, Chételat G, Collette F, Klimecki OM,
Marchant NL, Gonneaud J. The protective effect of
mindfulness and compassion meditation practices
on ageing: Hypotheses, models and experimental
implementation. Ageing Res Rev. 2021;72:101495. doi:10.1016/j.arr.2021.101495 21. Brassen S, Gamer M, Büchel C. Anterior
cingulate activation is related to a positivity bias
and emotional stability in successful aging. Biol
Psychiatry. 2011;70(2):131-137. doi:10.1016/
j.biopsych.2010.10.013 8. Chételat G, Lutz A, Arenaza-Urquijo E, Collette F,
Klimecki O, Marchant N. Why could meditation
practice help promote mental health and well-being
in aging? Alzheimers Res Ther. 2018;10(1):57. doi:10.1186/s13195-018-0388-5 22. Chételat G, Mézenge F, Tomadesso C, et al. Reduced age-associated brain changes in expert
meditators: a multimodal neuroimaging pilot study. Sci Rep. 2017;7(1):10160. doi:10.1038/
s41598-017-07764-x 9. Whitfield T, Barnhofer T, Acabchuk R, et al. The
effect of mindfulness-based programs on cognitive
function in adults: a systematic review and
meta-analysis. Neuropsychol Rev. 2022;32(3):
677-702. doi:10.1007/s11065-021-09519-y 23. Fjell AM, Walhovd KB. Structural brain changes
in aging: courses, causes and cognitive
consequences. Rev Neurosci. 2010;21(3):187-221. doi:10.1515/REVNEURO.2010.21.3.187 24. Kalpouzos G, Chételat G, Baron J-C, et al. Voxel-based mapping of brain gray matter volume
and glucose metabolism profiles in normal aging. Neurobiol Aging. 2009;30(1):112-124. doi:10.1016/
j.neurobiolaging.2007.05.019 10. Tang Y-Y, Hölzel BK, Posner MI. The
neuroscience of mindfulness meditation. ARTICLE INFORMATION Dr Chételat has received research support
from Fondation Alzheimer, Fondation Recherche
Alzheimer, Région Normandie, Association France
Alzheimer et maladies apparentées, and Fondation
Vaincre Alzheimer and personal fees from Caen,
Paris, Lyon and Nice Universities (salary for
lectures), and Fondation Alzheimer (as member of
the operational committee) outside the submitted
work. Drs Chételat and Lutz have received research
support from Fondation d’Entreprise MMA des
Entrepreneurs du Futur and MMA (payments made
to the institution). Dr Klimecki reported consulting
companies and teaching meditation in addition to
her scientific work. Dr Kuhn reported grants from
French Ministry of Higher Education and Research
(3 years of PhD, 2017-2020) during the conduct of
the study and grants from Fondation Philippe
Chatrier (2022 postdoctoral grant) outside the Group Information: The Medit-Ageing Research
Group is listed in Supplement 3. Group Information: The Medit-Ageing Research
Group is listed in Supplement 3. Additional Contributions: We thank D. Roquet,
PhD (Normandie Univ, UNICAEN, INSERM, U1237,
PhIND "Physiopathology and Imaging of
Neurological Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), and Oriane Hébert, MsC (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut
Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), for their help with the
reviewing process; A. Cognet, MSc (Normandie
Univ, UNICAEN, INSERM, U1237, PhIND
"Physiopathology and Imaging of Neurological
Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), C. Gaubert, MSc (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut Author Contributions: Dr Chételat had full access
to all of the data in the study and takes
responsibility for the integrity of the data and the
accuracy of the data analysis. Drs Chételat and Lutz
served as co–first authors. Drs Klimecki and Frison
served as co–second authors. Drs Collette, Poisnel,
and Marchant served as co–last authors. Concept and design: Chételat, Lutz, Klimecki,
Frison, Arenaza-Urquijo, Gonneaud, Quillard,
Vuilleumier, de la Sayette, Vivien, Collette, Poisnel,
Marchant. JAMA Neurology
Published online October 10, 2022 E8 jamaneurology.com Downloaded From: https://jamanetwork.com/ on 10/10/2022 Downloaded From: https://jamanetwork.com/ on 10/10/2022 Data Sharing Statement: See Supplement 4. 26. Cotier FA, Zhang R, Lee TMC. A longitudinal
study of the effect of short-term meditation
training on functional network organization of the
aging brain. Sci Rep. 2017;7(1):598. doi:10.1038/
s41598-017-00678-8 13. Saeed SA, Cunningham K, Bloch RM. Depression and anxiety disorders: benefits of
exercise, yoga, and meditation. Am Fam Physician. 2019;99(10):620-627. Ment Health. 2021;25(7):1181-1190. doi:10.1080/
13607863.2020.1793901 Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), M. Botton, MSc (Normandie
Univ, UNICAEN, INSERM, U1237, PhIND
"Physiopathology and Imaging of Neurological
Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), A. Joret Philippe, MSc (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut
Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), S. Egret, MSc (Normandie
Univ, UNICAEN, INSERM, U1237, PhIND
"Physiopathology and Imaging of Neurological
Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), P. Lacheray, MSc (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut
Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), J. Lebahar, PhD (Normandie
Univ, UNICAEN, INSERM, U1237, PhIND
"Physiopathology and Imaging of Neurological
Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), C. Tomadesso, PhD (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut
Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), F. Allais (EUCLID data
manager; Normandie Univ, UNICAEN, INSERM,
U1237, PhIND "Physiopathology and Imaging of
Neurological Disorders", Institut Blood and Brain
@ Caen-Normandie, Cyceron, 14000 Caen,
France), Marine Faure, MSc (Normandie Univ,
UNICAEN, INSERM, U1237, PhIND "Physiopathology
and Imaging of Neurological Disorders", Institut
Blood and Brain @ Caen-Normandie, Cyceron,
14000 Caen, France), and Jeanne Lepetit MSc
(Mediapilote); M. Ricard, PhD (Mind and Life
Europe, Winterthur, Switzerland & Shechen
Monastery, Kathmandu, Nepal), J. F. Lemoine, MD
(EUROPE1), Jon Kabat-Zinn, PhD (UMass Medical
School, Worcester, MA), Rhonda Smith, MSc
(Minerva), Charlotte Reid (Minerva), and Amanda
Beard (Minerva); the sponsor, Hélène Espérou, MD
(Pôle de recherche clinique Inserm), and the
Cyceron staff members for their help with
recruitment and data acquisition or administrative
support. Compensation was not received. We also
acknowledge all the participants of the study for
their contribution. 17. Fox KCR, Nijeboer S, Dixon ML, et al. Is
meditation associated with altered brain structure? a systematic review and meta-analysis of
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cognitive decline. Alzheimers Dement. 2020;16(7):
1054-1064. doi:10.1002/alz.12116 29. Poisnel G, Arenaza-Urquijo E, Collette F, et al;
Medit-Ageing Research Group. The Age-Well
randomized controlled trial of the Medit-Ageing
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Published online October 10, 2022 jamaneurology.com Downloaded From: https://jamanetwork.com/ on 10/10/2022 jamaneurology.com Effect of an 18-Month Meditation Training on Regional Brain Volume and Perfusion in Older Adults controlled trials. Sci Adv. 2022;8(20):eabk3316. doi:10.1126/sciadv.abk3316 function. Brain Struct Funct. 2010;214(5-6):655-667. doi:10.1007/s00429-010-0262-0 older adults. Alzheimers Dement (N Y). 2018;4:
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neuroplasticity in aging? a systematic review of
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Genetic forms of primary progressive aphasia within the GENetic Frontotemporal dementia Initiative (GENFI) cohort: comparison with sporadic primary progressive aphasia
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BRAIN COMMUNICATIONS Genetic forms of primary progressive aphasia
within the GENetic Frontotemporal dementia
Initiative (GENFI) cohort: comparison with
sporadic primary progressive aphasia Kiran Samra,1 Amy M. MacDougall,2 Arabella Bouzigues,1
Martina Bocchetta,1
David M. Cash,1 Caroline V. Greaves,1 Rhian S. Convery,1 Chris Hardy,1
John C. van Swieten,3
Harro Seelaar,3
Lize C. Jiskoot,3 Fermin Moreno,4,5
Raquel Sanchez-Valle,6 Robert Laforce,7
Caroline Graff,8,9 Mario Masellis,10
Maria Carmela Tartaglia,11 James B. Rowe,12
Barbara Borroni,13 Elizabeth Finger,14
Matthis Synofzik,15,16 Daniela Galimberti,17,18
Rik Vandenberghe,19,20,21
Alexandre de Mendonça,22 Chris R. Butler,23,24
Alexander Gerhard,25,26
Simon Ducharme,27,28 Isabelle Le Ber,29,30,31 Isabel Santana,32,33
Florence Pasquier,34,35,36 Johannes Levin,37,38,39 Markus Otto,40 Sandro Sorbi,41,42
Jason D. Warren,1 Jonathan D. Rohrer,1* and Lucy L. Russell1*
GENetic Frontotemporal dementia Initiative (GENFI) Kiran Samra,1 Amy M. MacDougall,2 Arabella Bouzigues,1
Martina Bocchetta,1
David M. Cash,1 Caroline V. Greaves,1 Rhian S. Convery,1 Chris Hardy,1
John C. van Swieten,3
Harro Seelaar,3
Lize C. Jiskoot,3 Fermin Moreno,4,5
Raquel Sanchez-Valle,6 Robert Laforce,7
Caroline Graff,8,9 Mario Masellis,10
Maria Carmela Tartaglia,11 James B. Rowe,12
Barbara Borroni,13 Elizabeth Finger,14
Matthis Synofzik,15,16 Daniela Galimberti,17,18
Rik Vandenberghe,19,20,21
Alexandre de Mendonça,22 Chris R. Butler,23,24
Alexander Gerhard,25,26
Simon Ducharme,27,28 Isabelle Le Ber,29,30,31 Isabel Santana,32,33
Florence Pasquier,34,35,36 Johannes Levin,37,38,39 Markus Otto,40 Sandro Sorbi,41,42
Jason D. Warren,1 Jonathan D. Rohrer,1* and Lucy L. Russell1*
GENetic Frontotemporal dementia Initiative (GENFI) Primary progressive aphasia is most commonly a sporadic disorder, but in some cases, it can be genetic. This study aimed to under
stand the clinical, cognitive and imaging phenotype of the genetic forms of primary progressive aphasia in comparison to the canonical
nonfluent, semantic and logopenic subtypes seen in sporadic disease. Participants with genetic primary progressive aphasia were re
cruited from the international multicentre GENetic Frontotemporal dementia Initiative study and compared with healthy controls as
well as a cohort of people with sporadic primary progressive aphasia. Symptoms were assessed using the GENetic Frontotemporal
dementia Initiative language, behavioural, neuropsychiatric and motor scales. Participants also underwent a cognitive assessment
and 3 T volumetric T1-weighted MRI. One C9orf72 (2%), 1 MAPT (6%) and 17 GRN (44%) symptomatic mutation carriers
had a diagnosis of primary progressive aphasia. In the GRN cohort, 47% had a diagnosis of nonfluent variant primary progressive
aphasia, and 53% had a primary progressive aphasia syndrome that did not fit diagnostic criteria for any of the three subtypes, called
primary progressive aphasia-not otherwise specified here. Received March 04, 2022. Revised August 26, 2022. Accepted February 15, 2023. Advance access publication February 17, 2023
© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse,
distribution, and reproduction in any medium, provided the original work is properly cited. BRAIN COMMUNICATIONS
https://doi.org/10.1093/braincomms/fcad036
BRAIN COMMUNICATIONS 2023: Page 1 of 18 | 1 https://doi.org/10.1093/braincomms/fcad036 *
These authors contributed equally to this work. BRAIN COMMUNICATIONS 2023: Page 1 of 18 BRAIN COMMUNICATIONS 2023: Page 1 of 18 BRAIN COMMUNICATIONS 1 Dementia Research Centre, Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
2 Department of Medical Statistics, London School of Hygiene and Tropical Medicine, London, UK 1 Dementia Research Centre, Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
2 Department of Medical Statistics, London School of Hygiene and Tropical Medicine, London, UK 1 Dementia Research Centre, Department of Neurodegenerative Disease, UCL Queen Square Institute
2
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3 Department of Neurology, Erasmus Medical Centre, Rotterdam, Netherlands 3 Department of Neurology, Erasmus Medical Centre, Rotterdam, Netherlands 3 Department of Neurology, Erasmus Medical Centre, Rotterdam, Netherlands p
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4 Cognitive Disorders Unit, Department of Neurology, Donostia Universitary Hospital, San Seb g
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5 Neuroscience Area, Biodonostia Health Research Institute, San Sebastian, Gipuzkoa, Spain 6 Alzheimer’s disease and Other Cognitive Disorders Unit, Neurology Service, Hospital Clínic, Institut d’Investigacións
Biomèdiques August Pi I Sunyer, University of Barcelona, Barcelona, Spain 6 Alzheimer’s disease and Other Cognitive Disorders Unit, Neurology Service, Hospital Clín
Biomèdiques August Pi I Sunyer, University of Barcelona, Barcelona, Spain q
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7 Clinique Interdisciplinaire de Mémoire,
Université Laval, Québec City, Canada 7 Clinique Interdisciplinaire de Mémoire, Département des Sciences Neurologiques, CHU de Québec, and Faculté de Médecine,
Université Laval, Québec City, Canada 7 Clinique Interdisciplinaire de Mémoire, D Université Laval, Québec City, Canada 8 Center for Alzheimer Research, Division of Neurogeriatrics, Department of Neurobiology,
Bioclinicum, Karolinska Institutet, Solna, Sweden 8 Center for Alzheimer Research, Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society,
Bioclinicum, Karolinska Institutet, Solna, Sweden Bioclinicum, Karolinska Institutet, Solna, Sweden 9 Unit for Hereditary Dementias, Theme Aging, Karolinska University Hospital, Solna, Sweden 10 Sunnybrook Health Sciences Centre, Sunnybrook Research Institute, University of Toronto, Toronto, Canada 10 Sunnybrook Health Sciences Centre, Sunnybrook Research Institute, University of Toronto, 10 Sunnybrook Health Sciences Centre, Sunnybrook Research Institute, University of g
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12 Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK 12 Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK 13 Neurology Unit, Department of Clinical and Experimental Sciences, University of Brescia, Brescia, 13 Neurology Unit, Department of Clinical and Experimental Sciences, University of Department of Clinical Neurological Sciences, University of Western Ontario, London, ON, Canada 14 Department of Clinical Neurological Sciences, University of Western Ontario, Lon p
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15 Department of Neurodegenerative Diseases, Hertie-Institute for Clinical Brain Research and Center of Neurology, University of
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15 Department of Neurodegenerative Diseases, Hertie-Institute for Clinical Brain Research and Center of Neurology, University of
Tübingen, Tübingen, Germany 15 Department of Neurodegenerative Diseases, Hertie-Institute for Clinical Brain Research and Center
Tübi
Tübi
G 16 Centre for Neurodegenerative Diseases (DZNE), Tübingen, Germany 16 Centre for Neurodegenerative Diseases (DZNE), Tübingen, Germany 16 Centre for Neurodegenerative Diseases (DZNE), Tübingen, Germany g
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17 Fondazione Ca’ Granda, IRCCS Ospedale Policlinico, Milan, Italy g
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17 Fondazione Ca’ Granda, IRCCS Ospedale Policlinico, Milan, Italy 18 University of Milan, Centro Dino Ferrari, Milan, Italy 18 University of Milan, Centro Dino Ferrari, Milan, Italy
19 Laboratory for Cognitive Neurology, Department of Neurosciences, KU Leuven, Leuven, Belgium y
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Laboratory for Cognitive Neurology, Department of Neurosciences, KU Leuven, Leuven, Belgium 19 Laboratory for Cognitive Neurology, Department of Neurosciences, KU Leuven, Leuven, Be boratory for Cognitive Neurology, Department of N 20 Neurology Service, University Hospitals Leuven, Leuven, Belgium Neurology Service, University Hospitals Leuven, Leu 21 Leuven Brain Institute, KU Leuven, Leuven, Belgium y
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23 Nuffield Department of Clinical Neurosciences, Medical Sciences Division, University of Oxford, Oxford, UK y
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23 Nuffield Department of Clinical Neurosciences, Medical Sciences Division, University of Oxford, Oxford, UK y
23 Nuffield Department of Clinical Neurosciences, Medical S y
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23 Nuffield Department of Clinical Neurosciences, Medical Sciences Division, University of 24 Department of Brain Sciences, Imperial College London, London, UK 25 Division of Neuroscience and Experimental Psychology, Wolfson Molecular Imaging Centre, University of Manchester,
Manchester, UK 25 Division of Neuroscience and Experimental Psychology, Wolfson Molecular Imaging Centre, University of Manchester,
Manchester, UK ments of Geriatric Medicine and Nuclear Medicine, University of Duisburg-Essen, Germany 26 Departments of Geriatric Medicine and Nuclear Medicine, University of Duisburg-Essen, Germany 27 Department of Psychiatry, McGill University Health Centre, McGill University, Montreal, Québec, Canada 27 Department of Psychiatry, McGill University Health Centre, McGill University, Montreal, Québec, Canada 29 Sorbonne Université, Paris Brain Institute – Institut du Cerveau – ICM, Inserm U1127, CNRS UMR 7225, AP-HP - Hôpital Pitié-
Salpêtrière, Paris, France 29 Sorbonne Université, Paris Brain Institute – Institut du Cerveau – ICM, Inserm U1127, CNRS UMR 7225, AP-HP - Hôpital Pitié-
Salpêtrière, Paris, France 30 Centre de référence des démences rares ou précoces, IM2A, Département de Neurologie, AP-HP - Hôpital Pitié-Salpêtrière, Paris,
France 30 Centre de référence des démences rares ou précoces, IM2A, Département de Neurologie, AP-HP - Hôpital Pitié-Salpêtrière, Paris,
France p
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32 University Hospital of Coimbra (HUC), Neurology Service, Faculty of Medicine, University of Coimbra, Coimbra, Portugal
33 Center for Neuroscience and Cell Biology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal
i
ill
ill 32 University Hospital of Coimbra (HUC), Neurology Service, Faculty of Medicine, University of Coimbra, Coimbra, Portugal 32 University Hospital of Coimbra (HUC), Neurology Service, Faculty of Medicine, University of Coimbra, Coimbra, Port 32 University Hospital of Coimbra (HUC), Neurology Service, Faculty of Medicine, University of Coimbra, Coimbra, Portugal
33 Center for Neuroscience and Cell Biology Faculty of Medicine University of Coimbra Coimbra Portugal 34 Univ Lille, Inserm 1172, Lille, France 35 Inserm 1172, Lille, France 36 CHU, CNR-MAJ, Labex Distalz, LiCEND Lille, France 36 CHU, CNR-MAJ, Labex Distalz, LiCEND Lille, France 37 Department of Neurology, Ludwig-Maximilians Universität München, Munich, Germany 37 Department of Neurology, Ludwig-Maximilians Universität München, Munich, Germany 37 Department of Neurology, Ludwig-Maximilians Universität Mün p
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38 German Centre for Neurodegenerative Diseases (DZNE), Munich, Germany p
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38 German Centre for Neurodegenerative Diseases (DZNE), Munich, German g
39 Munich Cluster of Systems Neurology (SyNergy), Munich, Germany 39 Munich Cluster of Systems Neurology (SyNergy), Munich, Germany 40 Department of Neurology, University of Ulm, Ulm, Germany 40 Department of Neurology, University of Ulm, Ulm, Germany 41 Department of Neurofarba, University of Florence, Florence, Italy 41 Department of Neurofarba, University of Florence, Florence, Italy 42 IRCCS Fondazione Don Carlo Gnocchi, Florence, Italy 42 IRCCS Fondazione Don Carlo Gnocchi, Florence, Italy BRAIN COMMUNICATIONS The phenotype of the genetic nonfluent variant primary progressive aphasia
group largely overlapped with that of sporadic nonfluent variant primary progressive aphasia, although the presence of an associated
atypical parkinsonian syndrome was characteristic of sporadic and not genetic disease. The primary progressive aphasia -not other
wise specified group however was distinct from the sporadic subtypes with impaired grammar/syntax in the presence of relatively in
tact articulation, alongside other linguistic deficits. The pattern of atrophy seen on MRI in the genetic nonfluent variant primary
progressive aphasia group overlapped with that of the sporadic nonfluent variant primary progressive aphasia cohort, although
with more posterior cortical involvement, whilst the primary progressive aphasia-not otherwise specified group was strikingly asym
metrical with involvement particularly of the insula and dorsolateral prefrontal cortex but also atrophy of the orbitofrontal cortex and
the medial temporal lobes. Whilst there are overlapping symptoms between genetic and sporadic primary progressive aphasia BRAIN COMMUNICATIONS 2023: Page 2 of 18 2 | 2 K. Samra et al. syndromes, there are also distinct features. Future iterations of the primary progressive aphasia consensus criteria should encompass
such information with further research needed to understand the earliest features of these disorders, particularly during the prodromal
period of genetic disease. Correspondence to: Dr Lucy L. Russell Dementia Research Centre, Department of Neurodegenerative Disease UCL Institute of Neurology, Queen Square London, UK, WC1N 3BG London, UK, WC1N 3BG E-mail: l.russell@ucl.ac.uk Keywords: primary progressive aphasia; GRN; c9orf72; MAPT Keywords: primary progressive aphasia; GRN; c9orf72; MAPT BRAIN COMMUNICATIONS 2023: Page 3 of 18 | Comparison of genetic PPA and sporadic PPA 3 Abbreviations: AD = Alzheimer’s disease; AMY amygdala; AC anterior cingulate; AI anterior insula; bvFTD behavioural variant
frontotemporal dementia; BNT Boston Naming Test; CAU caudate; C9orf72 chromosome 9 open reading frame 72; CDR Clinical
Dementia Rating; CBS corticobasal syndrome; CWIT Color-Word Interference Test; DLPFC dorsolateral prefrontal cortex;
FCSRT Free and Cued Selective Reminding Test; FTD frontotemporal dementia; GENFI GENetic Frontotemporal dementia
Initiative; GIF geodesic information flow; GP globus pallidus; HIP hippocampus; LOC lateral occipital; LP lateral parietal; LT
lateral temporal; lvPPA logopenic variant primary progressive aphasia; LIFTD longitudinal investigation of frontotemporal
dementia; MOC medial occipital; MP medial parietal; MT medial temporal; MAPT microtubule-associated protein tau; MMSE
Mini-Mental State Examination; mini-SEA Mini-Social cognition and Emotion Assessment; mCCT Modified Camel and Cactus
Test; MT motor; nfvPPA nonfluent variant primary progressive aphasia; NA nucleus accumbens; OF orbitofrontal; OP opercular;
PC posterior cingulate; PI posterior insula; PPA primary progressive aphasia; PPA -NOS Primary progressive aphasia-not otherwise
specified; GRN progranulin; GRN-NFV progranulin nonfluent variant; GRN-NOS progranulin-not otherwise specified; PASS
Progressive Aphasia Severity Scale; PSP progressive supranuclear palsy; PUT putamen; svPPA semantic variant primary progressive
aphasia; S somatosensory; ST supratemporal; THA thalamus; TP temporal pole; TIV total intracranial volume; TMT Trail Making
Test; VMPFC ventromedial prefrontal cortex; DSF/DSB WMS-R Digit Span Forward/Backward into any of the three main subtypes, a disorder often called
mixed PPA or PPA-not otherwise specified (PPA-NOS).3,4 Graphical Abstract Introduction Frontotemporal dementia (FTD) is a neurodegenerative dis
order characterised by progressive changes in behaviour, lan
guage and motor function.1 Although the most common
presenting symptom is a change in behaviour (behavioural
variant FTD, bvFTD), many people’s initial difficulties are
with language, a condition termed primary progressive apha
sia (PPA). Three subtypes of PPA are described:2 semantic
variant PPA (svPPA) is characterised by anomia and im
paired word comprehension with later non-verbal semantic
impairment; nonfluent variant PPA (nfvPPA) presents with
apraxia of speech and/or impaired grammar; and logopenic
variant PPA (lvPPA) is associated with word retrieval diffi
culties as well as impaired sentence repetition and compre
hension. However, a number of studies have now shown
that a substantial minority of people with PPA do not fit Whilst the majority of people with PPA have a sporadic
disease, cases of familial PPA have been described for
many years.5,6 Overall, between a third and a half of indivi
duals with FTD have a familial disorder7 with mutations in
progranulin (GRN), microtubule-associated protein tau
(MAPT) and chromosome 9 open reading frame 72
(C9orf72) being the commonest genetic causes. However,
the heritability of PPA is less than that of bvFTD, with
only 30% of nfvPPA, 22% of svPPA and 20% of lvPPA being
found to have a family history (of at least one first-degree
relative with dementia) in one previous study.7 GRN muta
tions have previously been described as the most common
genetic cause of PPA,8 with fewer reports of PPA in the other
genetic groups.9 The phenotype of such familial PPA cases
has been poorly studied but has been noted, particularly in BRAIN COMMUNICATIONS 2023: Page 4 of 18 4 K. Samra et al. those with GRN mutations, as often not easily characterized
into one of the three canonical subtypes8,10–15, i.e. to fall into
the PPA-NOS group. An initial comparison group was generated from the
GENFI healthy controls (i.e. non-mutation carriers) includ
ing those who were matched on age, sex and years of educa
tion—this formed a control group of 50 participants (all with
CDR® plus NACC FTLD of 0 or 0.5). A second set of com
parison groups were generated from the UCL Longitudinal
Investigation of Frontotemporal Dementia (LIFTD) study
of sporadic FTD (i.e. all cases in the study tested negative
for FTD-causing mutations), including participants with a
PPA diagnosis and a CDR® plus NACC FTLD global score
of 1 or 2. Participants Participants Participants were recruited from the fifth data freeze of the
GENFI study between 20 January 2012 and 30 May 2019,
including sites in the UK, Canada, France, Belgium,
Germany, Italy, Netherlands, Portugal, Spain and Sweden. All aspects of the study were approved by local ethics com
mittees and written informed consent obtained from all
participants. Behavioural, neuropsychiatric and motor symptoms were as
sessed using the standardised GENFI clinical questionnaire
(which uses the CDR scale of 0–3). This includes seven
behavioural symptoms: disinhibition, apathy, loss of sym
pathy/empathy, ritualistic/compulsive behaviour, hyperoral
ity and appetite changes, poor response to social/emotional
cues and inappropriate trusting behaviour. Fourteen neuro
psychiatric symptoms were assessed: visual, auditory and
tactile hallucinations, delusions, depression, anxiety, irrit
ability/lability, agitation/aggression, euphoria/elation, aber
rant motor behaviour, hypersexuality, hyperreligiosity,
impaired sleep and altered sense of humour. Lastly, eight
motor symptoms were enquired about: dysarthria, dyspha
gia, tremor, slowness, weakness, gait disorder, falls and
functional difficulties using hands. Participants underwent a standardised clinical assessment
including a clinical history and neurological examination,
neuropsychometric assessment, the Mini-Mental State
Examination (MMSE) and the CDR® plus NACC
FTLD.21 The CDR® plus NACC FTLD was used to classify
mutation carriers as asymptomatic (global score of 0), pro
dromal (score 0.5) or symptomatic (score ≥1). Initially, we
reviewed all symptomatic mutation carriers recruited to the
study and excluded those severely affected (CDR® plus
NACC FTLD score of 3, i.e. only included those with a score
of 1 or 2). From this remaining group of 103 participants,
there were 39 GRN, 46 C9orf72 and 18 MAPT mutation
carriers (Supplementary Fig. 1). A diagnosis of PPA was
made in the study by clinician assessment according to the
international consensus PPA criteria.22 An overall PPA diag
nosis was made and then, if possible, a subtype (svPPA,
nfvPPA or lvPPA) was allocated to each participant. If parti
cipants did not fit specific criteria for one of the subtypes,
they were diagnosed as PPA-NOS. Introduction In total 45 people with sporadic PPA were in
cluded: 19 with svPPA, 16 with nfvPPA and 10 with lvPPA
(Supplementary Fig. 1). Demographics are shown in Table 1. In the sporadic forms of PPA, language impairment can be
the only feature for a number of years into the illness.16,17
However, other cognitive, behavioural, neuropsychiatric
and motor features can be associated with each of the different
variants with increasing disease progression. People with
svPPA may develop non-verbal semantic impairment and be
havioural symptoms,18 whilst people with nfvPPA can de
velop dyscalculia and limb apraxia as well as parkinsonism,
including features consistent with either a corticobasal syn
drome (CBS) or progressive supranuclear palsy (PSP).19,20 In
contrast, people with lvPPA often develop episodic memory
and posterior cortical deficits, in line with the disorder being
commonly an atypical form of Alzheimer’s disease (AD).2
As with the linguistic phenotype, little is known about the
non-language features of the genetic forms of PPA. Linguistic symptoms Language was assessed by a clinician using the GENFI linguis
tic symptom scale, which is based on the Progressive Aphasia
Severity Scale (PASS).23 This contains 10 language symptoms
scored as per a Clinical Dementia Rating (CDR) scale, i.e. 0 =
asymptomatic, 0.5 = questionable/very mild, 1 = mild, 2 =
moderate and 3 = severe: impaired articulation, decreased
fluency, impaired grammar/syntax, impaired word retrieval,
impaired speech repetition, impaired sentence comprehen
sion, impaired single word comprehension, dyslexia, dys
graphia and impaired functional communication. With these findings in mind, we therefore aimed to under
stand the linguistic and non-linguistic phenotype of the gen
etic forms of PPA within the Genetic Frontotemporal
dementia Initiative (GENFI) cohort, by investigating the cog
nitive, behavioural, neuropsychiatric and motor features of
the condition, in comparison with healthy controls as well
as sporadic forms of PPA. Non-linguistic (behavioural,
neuropsychiatric, and motor)
symptoms Cognitive assessment Within the GENFI neuropsychology battery, the 30-item
version of the Boston Naming Test24,25 (BNT) and the modi
fied Camel and Cactus Test26 (mCCT) were the linguistic
measures used. Cognitive assessment The rest of the GENFI neuropsychology bat
tery includes tests of attention and executive function includ
ing the Trail Making Test Parts A and B (TMTA and TMTB),
D-KEFS Color-Word Inference Test (CWIT), WAIS-R Digit
Symbol test, WMS-R Digit Span Forwards (DSF) and BRAIN COMMUNICATIONS 2023: Page 5 of 18 | 5 Comparison of genetic PPA and sporadic PPA Table 1 Demographics, clinical and linguistic symptom data for the genetic and sporadic PPA subgroups as well as
healthy controls Table 1 Demographics, clinical and linguistic symptom data for the genetic and sporadic PPA subgroups as well as
healthy controls Table 1 Demographics, clinical and linguistic symptom data for the genetic and sporadic PPA subgroups as well as
healthy controls
Controls
Genetic PPA
Sporadic PPA
GRN-NFV
GRN-NOS
LV
NFV
SV
Number of participants
50
8
9
10
16
19
% Male
34
50
33
70
63
63
Age (years)
65.4 (4.5)
67.4 (7.9)
63.5 (8.2)
68.7 (6.4)
67.3 (6.1)
64.0 (6.8)
Education (years)
13.2 (2.9)
13.6 (2.7)
12.7 (3.9)
14.4 (2.4)
14.0 (2.3)
14.4 (3.1)
MMSE
29.0 (1.4)
22.4 (6.1)
20.9 (9.8)
19.1 (5.5)
22.4 (8.2)
23.0 (7.8)
CDR® plus NACC FTLD Global score
0.1 (0.2)
1.4 (0.2)
1.4 (0.5)
1.4 (0.5)
1.8 (0.5)
1.6 (0.5)
CDR® plus NACC FTLD Sum of boxes
0.2 (0.4)
6.0 (3.7)
6.6 (2.9)
6.8 (2.7)
7.3 (3.1)
8.0 (3.7)
Progressive aphasia Severity Scale Sum of Boxes
0.1 (0.2)
16.1 (5.9)
12.3 (6.0)
11.9 (3.3)
15.8 (5.7)
10.4 (4.0)
Impaired articulation
0.0 (0.0)
1.9 (1.0)
0.4 (0.7)
0.5 (0.4)
2.2 (0.9)
0.1 (0.2)
Decreased fluency
0.0 (0.0)
2.4 (0.5)
1.8 (0.9)
1.3 (0.7)
2.4 (0.6)
0.6 (0.6)
Impaired grammar/syntax
0.0 (0.0)
2.1 (0.6)
1.6 (1.0)
0.6 (0.6)
1.7 (1.0)
0.2 (0.4)
Impaired word retrieval
0.0 (0.1)
1.8 (0.9)
1.8 (0.9)
2.0 (0.5)
1.6 (1.0)
1.8 (0.5)
Impaired speech repetition
0.0 (0.0)
1.6 (1.1)
1.3 (1.1)
1.6 (0.7)
1.4 (1.1)
0.3 (0.6)
Impaired sentence comprehension
0.0 (0.0)
0.8 (0.7)
1.6 (0.8)
1.2 (0.5)
1.0 (1.0)
1.4 (0.9)
Impaired single word comprehension
0.0 (0.1)
0.4 (0.7)
0.8 (1.0)
1.1 (0.6)
0.6 (0.8)
1.9 (0.6)
Dyslexia
0.0 (0.0)
1.0 (0.7)
0.6 (0.7)
1.2 (0.8)
1.0 (0.9)
1.4 (1.0)
Dysgraphia
0.0 (0.1)
2.1 (0.9)
0.8 (0.6)
0.9 (0.5)
1.5 (1.1)
1.0 (0.9)
Impaired functional communication
0.0 (0.0)
2.3 (0.7)
1.5 (0.8)
1.7 (0.5)
2.3 (0.5)
1.6 (0.6)
Age, education, Mini-Mental State Examination (MMSE) and clinical rating scale scores are shown as mean (standard deviation). Cognitive assessment GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. Bold items are significantly different to controls. linical and linguistic symptom data for the genetic and sporadic PPA subgroups as well as Age, education, Mini-Mental State Examination (MMSE) and clinical rating scale scores are shown as mean (standard deviation). GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. Bold items are significantly different to controls. Volumes for subcortical regions were also calculated in
cluding the amygdala, hippocampus, nucleus accumbens,
caudate, globus pallidus, putamen, thalamus, together
with the cerebellum. All measures were expressed as a per
centage of total intracranial volume (TIV) computed with
SPM12 v6470 (29
Statistical Parametric Mapping,
Wellcome Trust Centre for Neuroimaging, London, UK)
running under Matlab R2014b (Math Works, Natick,
MA, USA).30 Backwards (DSB) and category fluency (animals) as well as
tests of visuospatial skills (WASI Block Design), episodic
memory (Free and Cued Selective Reminding Test, FCSRT)
and social cognition (mini-Social Cognition and Emotion
Assessment, mini-SEA, which includes a Faux Pas test of the
ory of mind and a Facial Emotion Recognition Test). The
LIFTD cohort underwent a subset of these tests, with none
of the participants having undergone the FCSRT or
mini-SEA. Neuroimaging analysis All statistical analyses were performed using Stata/MP
16.1. Statistical tests of normality were performed using
the Shapiro–Wilk test. Demographics were compared be
tween groups using either linear regression (age and educa
tion) or a chi-squared test (sex). Linear regressions
adjusting for age and sex were used to analyse the
MMSE, CDR® plus NACC FTLD and PASS scores. Both
linguistic and non-linguistic symptoms were compared in
each disease group versus controls using linear regressions
adjusting for age and sex and 95% bias-corrected boot
strapped confidence intervals with 2000 repetitions (as
there was minimal variation from 0 in severity scores for
the control group). Comparison of these symptoms be
tween groups used an ordinal logistic regression adjusting
for age and sex. The neuropsychological assessments and
cortical and subcortical volumes were compared using lin
ear regression models adjusting for age and sex, as well as
scanner type for the imaging analysis; 95% bias-corrected
bootstrapped confidence intervals with 2000 repetitions
were used if data were not normally distributed. Participants underwent a 3T volumetric T1-weighted MRI
scan as per the harmonized GENFI protocol.27 The majority
of participants had a scan of sufficient quality to be analysed
(42 on a Siemens Prisma, 31 on a Siemens Trio, 6 on a
Siemens Skyra, and 24 on a Philips Achieva): 45 of 50 con
trols, 14 of 17 genetic PPA, 9 of 10 lvPPA, all 16 nfvPPA
and all 19 svPPA participants (see Supplementary Table 4). Those without scans had either not been scanned due to con
traindications or had a poor-quality scan due to movement
or other artefacts. Volumetric MRI scans were first bias field corrected and
whole brain parcellated using the geodesic information
flow (GIF) algorithm,28 which is based on atlas propaga
tion and label fusion. We combined regions of interest to
calculate grey matter volumes of the cortex for 15 regions:
orbitofrontal, dorsolateral (DLPFC) and ventromedial pre
frontal (VMPFC), motor, opercular, anterior and posterior
insula, temporal pole, lateral and medial temporal, supra
temporal, anterior and posterior cingulate, sensory, medial
and lateral parietal, and medial and lateral occipital cortex. 6 | 6 BRAIN COMMUNICATIONS 2023: Page 6 of 18 K. Samra et al. contrast, all of the linguistic symptoms were present within
each of the PPA subtypes in at least some of the patients in
each group (Fig. 1, Table 1). Disease severity CDR® plus NACC FTLD sum of boxes and MMSE scores
were all significantly different to controls in each of the dis
ease groups, but there were no significant differences be
tween the PPA groups. Results comparing each linguistic symptom across the
PPA groups are shown in Supplementary Table 2. Characteristic between-group differences were seen in spor
adic PPA, i.e. more severe impairments of articulation and
grammar/syntax in nfvPPA, single word comprehension in
svPPA and speech repetition in lvPPA. Within the genetic
PPA groups, GRN-NFV had more severely impaired articu
lation (P = 0.001) and dysgraphia (P = 0.002) than the
GRN-NOS group, whilst the GRN-NOS group had more
severely impaired sentence comprehension than GRN-NFV
(P = 0.026). Frequency of PPA in the GENFI
cohort In total 1 C9orf72 (2%), 1 MAPT (6%) and 17 GRN (44%)
mutation carriers from within the total mild to moderate
symptomatic mutation carrier population had a PPA diagno
sis. On assessment of their specific subtype, the C9orf72 mu
tation carrier had a diagnosis of nfvPPA, the MAPT
mutation carrier had a diagnosis of svPPA, and the GRN mu
tation carriers split into eight with a diagnosis of nfvPPA
(termed GRN-NFV from here) and nine who did not fit diag
nostic criteria for any specific PPA variant, i.e. PPA-NOS
(termed GRN-NOS from here). Due to the small numbers
in the C9orf72 and MAPT mutation groups, we focused fur
ther analyses on the GRN-PPA group—details of the
non-GRN cases are shown in Supplementary Table 1. The genetic nfvPPA group (GRN-NFV) had an overlap
ping pattern of symptoms to the sporadic nfvPPA group
(Fig. 1A and B, Table 1): the most frequent and severe symp
toms were decreased fluency [2.4 (0.5), 100%], impaired
grammar/syntax [2.1 (0.6), 100%] and impaired articulation
[1.9 (1.0), 88%] as with sporadic nfvPPA, with the addition
in the GRN-NFV group of dysgraphia [2.1 (0.9), 100%],
whilst the least frequent and severe symptom was impaired
single word comprehension [0.4 (0.7), 38%]. This latter
symptom being the only one not significantly different in se
verity to controls (P = 0.186). Demographics The genetic PPA-NOS group (GRN-NOS) had a different
pattern to any of the sporadic PPA groups or the GRN-NFV
group. The most frequent and severe symptoms were de
creased fluency [1.8 (0.9), 100%], impaired word retrieval
[1.8 (0.9), 100%], impaired grammar/syntax [1.6 (1.0),
100%] and impaired sentence comprehension [1.6 (0.8),
100%], whilst the least frequent and severe symptom was
impaired articulation [0.4 (0.7), 44%], this latter symptom
being the only one not significantly different in severity to
controls (P = 0.063). Of note, impaired speech repetition oc
curred in 89% [1.3 (1.1)], and impaired single word compre
hension occurred in 67% [0.8 (1.0)] in this group. No significant differences were seen between the GRN-PPA
group and controls in age, sex or education. However, the
sporadic lvPPA, nfvPPA and svPPA groups had a significant
ly higher percentage of males than the control group (Chi2 =
4.50, P = 0.034; Chi2 = 4.07, P = 0.044; Chi2 = 4.80, P =
0.028, respectively). The sporadic lvPPA cohort was older
than the svPPA group (P = 0.045), but there were no other
differences in age and no significant differences in education
between the groups. Ethics approval and consent to
participate All GENFI sites had local ethical approval for the study, and
all participants gave written informed consent. As expected, the most frequent and severe symptoms in
the sporadic forms of PPA were decreased fluency [mean
(SD) 2.4 (0.6)], impaired articulation [2.2 (0.9)] and im
paired grammar/syntax [1.7 (1.0)] in the nfvPPA group, im
paired single word comprehension [1.9 (0.6)] and impaired
word retrieval [1.8 (0.5)] in the svPPA group, and impaired
word retrieval [2.0 (0.5)], impaired speech repetition [1.6
(0.7)], impaired sentence comprehension [1.2 (0.5)] and dys
lexia [1.2 (0.8)] in the lvPPA group. Apart from impaired
speech repetition in lvPPA (occurring in 90%), these symp
toms occurred in 100% of patients with these PPA subtypes. The least frequent and severe symptoms in the sporadic
forms of PPA were also as expected: impaired single word
comprehension in nfvPPA [0.6 (0.8), 56%] and both im
paired articulation [0.1 (0.2), 16%; 0.5 (0.4), 70%] and im
paired grammar/syntax [0.2 (0.4), 26%; 0.6 (0.6), 70%] in
the svPPA and lvPPA groups. Nonetheless, in comparing se
verity of symptoms versus controls, all were significant ex
cept impaired articulation for svPPA (P = 0.144). Data availability The datasets used and/or analysed during the current study
are available from the corresponding author on reasonable
request. Linguistic symptoms The majority of controls showed no symptoms of language
impairment, with only 6% having impaired word retrieval,
2% having impaired single word comprehension and 2%
with dysgraphia (all at a 0.5 very mild/questionable severity). All other linguistic symptoms did not occur in controls. In BRAIN COMMUNICATIONS 2023: Page 7 of 18 | 7 Comparison of genetic PPA and sporadic PPA A
B
C
D
E
uistic symptoms showing the percentage of participants in each of the PPA groups who score 0 =
stionable, 1 = mild, 2 = moderate or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C
Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any s
risk above the bar indicates that the symptom severity is significantly greater than controls. Linear regressions
ed to compare groups for each symptom: impaired articulation [Wald chi2(7)
146 7 P < 0 001; R2
0 761]; dec
netic PPA and sporadic PPA BRAIN COMMUNICATIONS 2023: P A B B B C D D C E E E Figure 1 Linguistic symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent, 0.5 =
very mild/questionable, 1 = mild, 2 = moderate or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS; D,
Sporadic SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category
(0.5–3). An asterisk above the bar indicates that the symptom severity is significantly greater than controls. Linguistic symptoms Linear regressions adjusting for age
and sex were used to compare groups for each symptom: impaired articulation [Wald chi2(7) = 146.7, P < 0.001; R2 = 0.761]; decreased fluency
[Wald chi2(7) = 536.0, P < 0.001; R2 = 0.811]; impaired grammar/syntax [Wald chi2(7) = 176.1, P < 0.001; R2 = 0.682]; impaired word retrieval
[Wald chi2(7) = 538.8, P < 0.001; R2 = 0.744]; impaired speech repetition [Wald chi2(7) = 116.3, P < 0.001; R2 = 0.535]; impaired sentence
comprehension [Wald chi2(7) = 155.4, P < 0.001; R2 = 0.521]; impaired single word comprehension [Wald chi2(7) = 250.2, P < 0.001; R2 = 0.638];
dyslexia [Wald chi2(7) = 111.3, P < 0.001; R2 = 0.470]; dysgraphia [Wald chi2(7) = 146.2, P < 0.001; R2 = 0.537]; impaired functional communication
[Wald chi2(7) = 688.6, P < 0.001; R2 = 0.830]. Abbreviations: GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not
otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV, semantic variant PPA. Figure 1 Linguistic symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent, 0.5 =
very mild/questionable, 1 = mild, 2 = moderate or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS; D,
Sporadic SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category
(0.5–3). An asterisk above the bar indicates that the symptom severity is significantly greater than controls. Linear regressions adjusting for age
and sex were used to compare groups for each symptom: impaired articulation [Wald chi2(7) = 146.7, P < 0.001; R2 = 0.761]; decreased fluency
[Wald chi2(7) = 536.0, P < 0.001; R2 = 0.811]; impaired grammar/syntax [Wald chi2(7) = 176.1, P < 0.001; R2 = 0.682]; impaired word retrieval
[Wald chi2(7) = 538.8, P < 0.001; R2 = 0.744]; impaired speech repetition [Wald chi2(7) = 116.3, P < 0.001; R2 = 0.535]; impaired sentence
comprehension [Wald chi2(7) = 155.4, P < 0.001; R2 = 0.521]; impaired single word comprehension [Wald chi2(7) = 250.2, P < 0.001; R2 = 0.638];
dyslexia [Wald chi2(7) = 111.3, P < 0.001; R2 = 0.470]; dysgraphia [Wald chi2(7) = 146.2, P < 0.001; R2 = 0.537]; impaired functional communication
[Wald chi2(7) = 688.6, P < 0.001; R2 = 0.830]. Cognitive assessment In contrast to the sporadic PPA groups, the most common
and severe behavioural symptom in both GRN-NFV and
GRN-NOS groups was apathy [75%, 0.9 (0.6); 67%, 0.8
(0.8), respectively]. Both groups also had significantly more
severe loss of sympathy/empathy [50%, 0.4 (0.4); 56%,
0.4 (0.5), respectively] and hyperorality and appetite changes
[75%, 0.8 (0.8); 44%, 0.7 (0.9), respectively] compared with
controls. Additionally, in the GRN-NOS group, poor re
sponse to emotional cues was more severely affected than
controls [56%, 0.6, (0.7)]. The mean score for the BNT was significantly different to con
trols for all PPA groups apart from the sporadic nfvPPA co
hort where there was a trend for a lower score (P = 0.055)
(Table 2). The sporadic svPPA group scored the lowest at
8.2 (8.1) and was significantly different to GRN-NFV,
GRN-NOS and sporadic nfvPPA groups (P = 0.005, P =
<0.001 and P = <0.001, respectively). The sporadic lvPPA
group also had a significantly lower score of 12.7 (6.8)
compared to GRN-NOS and sporadic nfvPPA groups (P =
0.014, P = <0.001, respectively). Scores in the two genetic
PPA groups were lower than controls but lay between the
sporadic nfvPPA group (23.6 (5.8)) and the other two sporad
ic PPA groups: GRN-NFV 18.0 (8.5), GRN-NOS 21.4 (7.0).i Neuropsychiatric symptoms occurred commonly in pa
tients in each group compared with only 16% of controls:
100% of patients in all groups had symptoms except genetic
nfvPPA (88%). The mean score for the mCCT was significantly lower in
the GRN-NOS group compared with controls [22.0 (8.1),
P = 0.001] with a trend in the svPPA group [25.2 (4.6),
P = 0.073]. The GRN-NOS group scored significantly lower
than the lvPPA [29.5 (0.7), P = 0.007] and sporadic nfvPPA
groups [28.5 (4.5), P = 0.025]. For all groups, depression, anxiety and irritability/lability
were the most frequent and severe symptoms (Fig. 3A–E,
Supplementary Table 3). Depression occurred most fre
quently in the lvPPA group (80%) followed by sporadic
nfvPPA (75%) and svPPA (68%), with the two genetic
groups showing the lowest frequency and severity
(GRN-NFV 63%, GRN-NOS 44%). Depression severity
was more significant than controls in all groups except
GRN-NOS. BRAIN COMMUNICATIONS 2023: Page 8 of 18 K. Samra et al. 8 lvPPA (70%), genetic PPA-NOS (56%), genetic nfvPPA
(50%) and svPPA (32%). Non-linguistic symptoms Behavioural symptoms occurred commonly in patients in
each group compared with only 10% of controls: svPPA
(95% of patients), sporadic nfvPPA (94%), genetic nfvPPA
(88%), genetic PPA-NOS (78%) and lvPPA (70%)
(Fig. 2A–E). All motor symptoms apart from weakness occurred fre
quently and more severely than controls in the sporadic
nfvPPA group with the most frequent and severe being dys
arthria [75%, 1.6 (1.2)], slowness [56%, 0.8 (1.0)] and
gait disorder [56%, 0.8 (0.8)] (Fig. 4A–E, Supplementary
Table 3). Apart from dysarthria in the lvPPA group, no other
symptom in any of the groups was significantly more severe
than controls. In particular, although dysarthria (25%), gait
disorder (38%) and slowness (25%) all occurred in the
GRN-NFV group, this was less frequent than the sporadic
nfvPPA group. Furthermore, 50% of the sporadic nfvPPA
group symptoms met consensus criteria for one of the atyp
ical parkinsonian disorders (four with PSP, three with CBS
and one with a PSP/CBS overlap syndrome), whilst none of
the other patients in any of the other groups met criteria. In svPPA the most frequent and severe symptoms were loss
of sympathy/empathy [84%, 1.1 (0.8)], hyperorality and ap
petite changes [74%, 1.1 (0.9)] and disinhibition [74%, 1.0
(0.8)], but all symptoms were significantly more severe than
controls (Fig. 2A–E, Supplementary Table 3). In sporadic
nfvPPA, the most frequent and severe symptoms were hyper
orality and appetite changes [69%, 0.9 (0.9)], apathy [69%,
0.6 (0.5)] and poor response to emotional cues [56%, 0.9
(0.9)], with all symptoms except disinhibition and inappro
priate trusting behaviour significantly more severe than con
trols. The lvPPA group had the least frequent and severe
symptoms, with only apathy significantly more severe than
controls [50%, 0.7 (0.8)]. Linguistic symptoms Abbreviations: GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not
otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV, semantic variant PPA. BRAIN COMMUNICATIONS 2023: Page 8 of 18 Cognitive assessment Anxiety and irritability/lability were also com
mon (and significantly more severe than controls) in all
groups: 88% and 88%, respectively, in sporadic nfvPPA,
80% and 70% in lvPPA, 78% and 44% in GRN-NOS,
75% and 63% in GRN-NFV and 68% and 53% in svPPA. Apart from agitation/aggression in sporadic nfvPPA and
GRN-NOS, and aberrant motor behaviour in sporadic
nfvPPA, all other symptoms occurred at a low frequency
and were not significantly different to controls. Of note,
however, delusions occurred in 22% of GRN-NOS, and
13% of GRN-NFV but did not occur in the sporadic PPA
groups. All groups performed significantly worse than controls on
the category fluency task, with the lowest scores being found
in the lvPPA [4.4 (3.3)] and svPPA [6.8 (5.1)] groups. In the other cognitive tests, executive dysfunction was pre
sent in all groups, although to a lesser extent in the svPPA
group, with significantly worse performance than controls
in the D-KEFS CWIT and Digit Symbol test for all groups
and for the TMTB and DSB in all groups except svPPA. Similarly, all groups except the svPPA cohort showed signifi
cant impairment compared to controls on DSF and the Block
Design task. In the tests performed only in the genetic PPA cohort, there
was evidence of impaired episodic memory in the GRN-NOS
group, with significantly lower scores compared to controls
for both FCSRT free recall and free and cued recall (P =
0.034 and P = 0.009, respectively), and significantly lower
scores than GRN-NFV individuals for all FCSRT tests ex
cept free delayed recall (P = 0.073). Both the GRN-NFV Motor symptoms occurred in each of the PPA groups (and
in 6% of controls), most commonly in the sporadic nfvPPA
group (100%) and less frequently in the other groups: BRAIN COMMUNICATIONS 2023: Page 9 of 18 | 9 Comparison of genetic PPA and sporadic PPA A
B
C
D
E
viour symptoms showing the percentage of participants in each of the PPA groups who score 0
tionable 1
mild 2
moderate or 3
severe for each symptom A GRN NFV; B Sporadic NFV
etic PPA and sporadic PPA BRAIN COMMUNICATIONS 2023 B A B A B
D
E
oms showing the percentage of participants in each of the PPA groups who score 0 = absent, 0.5 =
mild, 2 = moderate, or 3 = severe for each symptom. Cognitive assessment A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS; D,
lues along the x-axis represent the frequency (%) with which the symptom is present in any severity category (0.5–
dicates that the symptom severity is significantly greater than controls. Linear regressions adjusting for age and sex
or each symptom: disinhibition [Wald chi2(7) = 44.9, P < 0.001; R2 = 0.494]; apathy [Wald chi2(7) = 70.6, P < 0.001;
mpathy [Wald chi2(7) = 73.7, P < 0.001; R2 = 0.436]; ritualistic/compulsive [Wald chi2(7) = 35.4, P < 0.001; R2 =
hanges [Wald chi2(7) = 66.0, P < 0.001; R2 = 0.377]; poor social/emotional responses [Wald chi2(7) = 50.0, P <
te trusting [Wald chi2(7) = 28.5, P < 0.001; R2 = 0.400]. Abbreviations: GRN-NFV, nonfluent variant PPA due to
OS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant D C C D E E E Figure 2 Behaviour symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent, 0.5 =
very mild/questionable, 1 = mild, 2 = moderate, or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS; D,
Sporadic SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category (0.5–
3). An asterisk above the bar indicates that the symptom severity is significantly greater than controls. Linear regressions adjusting for age and sex
were used to compare groups for each symptom: disinhibition [Wald chi2(7) = 44.9, P < 0.001; R2 = 0.494]; apathy [Wald chi2(7) = 70.6, P < 0.001;
R2 = 0.348]; loss of sympathy/empathy [Wald chi2(7) = 73.7, P < 0.001; R2 = 0.436]; ritualistic/compulsive [Wald chi2(7) = 35.4, P < 0.001; R2 =
0.358]; hyperorality/appetite changes [Wald chi2(7) = 66.0, P < 0.001; R2 = 0.377]; poor social/emotional responses [Wald chi2(7) = 50.0, P <
0.001; R2 = 0.359]; inappropriate trusting [Wald chi2(7) = 28.5, P < 0.001; R2 = 0.400]. Abbreviations: GRN-NFV, nonfluent variant PPA due to
progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. A
B
C
D
E
Neuropsychiatric symptoms showing the percentage of participants in each of the PPA groups who score 0 =
mild/questionable, 1 = mild, 2 = moderate, or 3 = severe for each symptom. Cognitive assessment A, GRN-NFV; B, Sporadic NFV; C, G
SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity
asterisk above the bar indicates that the symptom severity is significantly greater than controls. Linear regressions adjusting fo
ed to compare groups for each symptom: visual hallucinations [Wald chi2(7) = 3.3, P = 0.856; R2 = 0.117]; delusions [Wald
08; R2 = 0.084]; depression [Wald chi2(7) = 74.8, P < 0.001; R2 = 0.417]; anxiety [Wald chi2(7) = 118.4, P < 0.001; R2 = 0.47
ability [Wald chi2(7) = 77.4, P < 0.001; R2 = 0.454]; agitation/aggression [Wald chi2(7) = 21.2, P = 0.004; R2 = 0.232]; euphor
7) = 7.8, P = 0.354; R2 = 0.080]; aberrant motor behaviour [Wald chi2(7) = 16.1, P = 0.025; R2 = 0.248]; hypersexuality [Wa
81; R2 = 0.140]; hyperreligiosity [Wald chi2(7) = 4.3, P = 0.749; R2 = 0.157]; impaired sleep [Wald chi2(7) = 28.5, P < 0.001
red sense of humour [Wald chi2(7) = 60.3, P < 0.001; R2 = 0.434]. Abbreviations: GRN-NFV, nonfluent variant PPA due to pr
RN NOS
h
i
ifi d PPA d
li
i
LV l
i
i
PPA NFV
fl
i
PP
N COMMUNICATIONS 2023: Page 10 of 18 K. BRAIN COMMUNICATIONS 2023: Page 10 of 18 10 | K. Samra et al. 10 A B D C D C E E E Figure 3 Neuropsychiatric symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent,
0.5 = very mild/questionable, 1 = mild, 2 = moderate, or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS;
D, Sporadic SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category
(0.5–3). An asterisk above the bar indicates that the symptom severity is significantly greater than controls. Cognitive assessment Linear regressions adjusting for age and
sex were used to compare groups for each symptom: visual hallucinations [Wald chi2(7) = 3.3, P = 0.856; R2 = 0.117]; delusions [Wald chi2(7) =
4.6, P = 0.708; R2 = 0.084]; depression [Wald chi2(7) = 74.8, P < 0.001; R2 = 0.417]; anxiety [Wald chi2(7) = 118.4, P < 0.001; R2 = 0.479];
irritability/lability [Wald chi2(7) = 77.4, P < 0.001; R2 = 0.454]; agitation/aggression [Wald chi2(7) = 21.2, P = 0.004; R2 = 0.232]; euphoria/elation
[Wald chi2(7) = 7.8, P = 0.354; R2 = 0.080]; aberrant motor behaviour [Wald chi2(7) = 16.1, P = 0.025; R2 = 0.248]; hypersexuality [Wald chi2(7)
= 4.0, P = 781; R2 = 0.140]; hyperreligiosity [Wald chi2(7) = 4.3, P = 0.749; R2 = 0.157]; impaired sleep [Wald chi2(7) = 28.5, P < 0.001; R2 =
0.244]; altered sense of humour [Wald chi2(7) = 60.3, P < 0.001; R2 = 0.434]. Abbreviations: GRN-NFV, nonfluent variant PPA due to progranulin
mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV,
semantic variant PPA. Figure 3 Neuropsychiatric symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent,
0.5 = very mild/questionable, 1 = mild, 2 = moderate, or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS;
D, Sporadic SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category
(0.5–3). An asterisk above the bar indicates that the symptom severity is significantly greater than controls. Cognitive assessment Linear regressions adjusting for age and
sex were used to compare groups for each symptom: visual hallucinations [Wald chi2(7) = 3.3, P = 0.856; R2 = 0.117]; delusions [Wald chi2(7) =
4.6, P = 0.708; R2 = 0.084]; depression [Wald chi2(7) = 74.8, P < 0.001; R2 = 0.417]; anxiety [Wald chi2(7) = 118.4, P < 0.001; R2 = 0.479];
irritability/lability [Wald chi2(7) = 77.4, P < 0.001; R2 = 0.454]; agitation/aggression [Wald chi2(7) = 21.2, P = 0.004; R2 = 0.232]; euphoria/elation
[Wald chi2(7) = 7.8, P = 0.354; R2 = 0.080]; aberrant motor behaviour [Wald chi2(7) = 16.1, P = 0.025; R2 = 0.248]; hypersexuality [Wald chi2(7)
= 4.0, P = 781; R2 = 0.140]; hyperreligiosity [Wald chi2(7) = 4.3, P = 0.749; R2 = 0.157]; impaired sleep [Wald chi2(7) = 28.5, P < 0.001; R2 =
0.244]; altered sense of humour [Wald chi2(7) = 60.3, P < 0.001; R2 = 0.434]. Abbreviations: GRN-NFV, nonfluent variant PPA due to progranulin
mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV,
semantic variant PPA. BRAIN COMMUNICATIONS 2023: Page 11 of 18 | 11 Comparison of genetic PPA and sporadic PPA A
B
C
D
E
Motor symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent
ti
bl
1
ild 2
d
t
3
f
h
t
A GRN NFV B S
di NFV C GRN NOS
of genetic PPA and sporadic PPA BRAIN COMMUNICATIONS 2023: Page 1 B A B C D D C E E Figure 4 Motor symptoms showing the percentage of participants in each of the PPA groups who score 0 = absent, 0.5 = very
mild/questionable, 1 = mild, 2 = moderate, or 3 = severe for each symptom. A, GRN-NFV; B, Sporadic NFV; C, GRN-NOS; D, Sporadic
SV; E, Sporadic LV. Values along the x-axis represent the frequency (%) with which the symptom is present in any severity category (0.5–3). An
asterisk above the bar indicates that the symptom severity is significantly greater than controls. Cognitive assessment Linear regressions adjusting for age and sex were
used to compare groups for each symptom: dysarthria [Wald chi2(7) = 38.9, P < 0.001; R2 = 0.495]; dysphagia [Wald chi2(7) = 9.1, P = 0.244; R2 =
0.263]; tremor [Wald chi2(7) = 8.2, P = 0.312; R2 = 0.146]; slowness [Wald chi2(7) = 23.3, P = 0.002; R2 = 0.297]; weakness [Wald chi2(7) = 4.4,
P = 0.734; R2 = 0.130]; gait disorder [Wald chi2(7) = 25.1, P = 0.001; R2 = 0.350]; falls [Wald chi2(7) = 14.8, P = 0.038; R2 = 0.180]; functional
difficulties using hands [Wald chi2(7) = 8.8, P = 0.270; R2 = 0.246]. Abbreviations: GRN-NFV, nonfluent variant PPA due to progranulin mutation;
GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV, semantic variant
PPA. BRAIN COMMUNICATIONS 2023: Page 12 of 18 K. Samra et al. Cognitive assessment GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. FCSRT, Free and Cued Selective Reminding Test; Mini-SEA; Mini-Social Cognition and Emotion Assessment. aBold items are significantly different to controls. bSignificantly impaired compared to svPPA. cSignificantly impaired compared to nfvPPA. dSignificantly impaired compared to GRN-NOS. eSignificantly impaired compared to GRN-NFV. fSignificantly impaired compared to lvPPA. significantly different to controls included the amygdala
and hippocampus as well as the basal ganglia. and GRN-NOS groups scored significantly lower on both
parts of the mini-SEA: 25.0 (7.1) and 25.5 (8.9) for the
Faux Pas test and 21.3 (5.3) and 22.1 (7.5) for the Facial
Emotion Recognition Test. Cognitive assessment 12 | 12 Table 2 Neuropsychological assessment scores in the genetic and sporadic PPA subgroups as well as healthy controls
Controls
Genetic PPA
Sporadic PPA
GRN-NFV
GRN-NOS
LV
NFV
SV
Boston Naming Test (/30)
27.3 (2.1)
18.0a (8.5)
21.4 (7.0)b
12.7 (6.8)c,d
23.6 (5.8)
8.2 (8.1)c,d,e
Modified Camel and Cactus Test (/32)
29.6 (1.7)
26.6 (6.6)
22.0 (8.1)c,f
29.5 (0.7)
28.5 (4.5)
25.2 (4.6)
Trail Making Test Part A (max 150 s)
36.6 (17.5)
73.8 (41.3)
69.1 (40.5)b
101.0 (72.3)b
60.3 (37.0)
41.9 (20.0)
Trail Making Test Part B (max 300 s)
88.4 (45.8)
230.0 (85.4)b
244.0 (93.8)b,c
225.2 (83.0)b
145.1 (83.2)
121.6 (99.5)
D-KEFS Color-Word Inference Test (max 180 s)
53.6 (13.2)
132.8 (49.9)b
127.4 (55.7)b
145.7 (42.6)b
116.2 (37.6)b
78.7 (34.8)
Digit Symbol test (max in 90 s)
49.2 (12.1)
25.3 (15.9)b
28.1 (15.2)b
20.6 (10.3)b
33.8 (13.4)
41.8 (15.6)
Digit Span Forwards (/12)
7.3 (1.8)
3.1 (1.5)b
5.0 (2.8)b
3.2 (2.3)b
5.1 (2.2)b
8.1 (3.0)
Digit Span Backwards (/12)
6.4 (1.9)
2.7 (1.7)b
3.1 (2.6)b
2.2 (1.2)b
3.5 (1.6)b
6.5 (3.2)
Category fluency (max in 60 s)
22.7 (6.1)
13.9 (5.4)
11.8 (6.5)
4.4 (3.3)c,d,e
10.1 (6.4)
6.8 (5.1)e
Block Design (/71)
41.7 (11.4)
20.3 (17.4)b
22.3 (16.2)b
13.2 (10.7)b,c
26.3 (20.3)
37.8 (18.8)
FCSRT—free recall (/48)
27.3 (7.9)
33.5 (7.8)
19.6 (5.9)e
FCSRT—free + cued recall (/48)
44.3 (5.9)
46.5 (2.1)
40.2 (4.1)e
FCSRT—free delayed recall (/16)
10.8 (3.1)
12.5 (4.9)
8.4 (2.3)
FCSRT—free + cued delayed recall (/16)
14.9 (1.8)
16.0 (0.0)
13.6 (2.1)e
Mini-SEA: Faux Pas test (/40)
34.3 (5.2)
25.0 (7.1)
25.5 (8.9)
Mini-SEA: Facial Emotion Recognition Test (/35)
27.9 (3.4)
21.3 (5.3)
22.1 (7.5)
All data are shown as mean (standard deviation). GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. FCSRT, Free and Cued Selective Reminding Test; Mini-SEA; Mini-Social Cognition and Emotion Assessment. aBold items are significantly different to controls. bSignificantly impaired compared to svPPA. cSignificantly impaired compared to nfvPPA. dSignificantly impaired compared to GRN-NOS. eSignificantly impaired compared to GRN-NFV. fSignificantly impaired compared to lvPPA. Table 2 Neuropsychological assessment scores in the genetic and sporadic PPA subgroups a l assessment scores in the genetic and sporadic PPA subgroups as well as healthy controls All data are shown as mean (standard deviation). Discussion In this study, we have shown that, although there are over
lapping symptoms between genetic and sporadic PPA syn
dromes, there are also distinct features. PPA is a common
clinical syndrome in those with GRN mutations (44% in
the GENFI cohort) but uncommon in both C9orf72 and
MAPT mutations (only one case in the GENFI cohort in
both genetic groups). Within the GRN mutation carriers,
there seem to be two distinct groups, a nfvPPA group which
overlaps to a large extent with sporadic nfvPPA, although at
least in this cohort with fewer atypical parkinsonian motor
features, and a PPA-NOS group, not neatly fitting into any
of the canonical sporadic syndromes, with highly asymmet
rical left hemisphere atrophy on brain imaging. All data are shown as mean (standard deviation).
GRN-NFV, nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant
PPA; SV, semantic variant PPA. FCSRT, Free and Cued Selective Reminding Test; Mini-SEA; Mini-Social Cognition and Emotion Assessment.
aBold items are significantly different to controls.
bSignificantly impaired compared to svPPA.
cSignificantly impaired compared to nfvPPA.
dSignificantly impaired compared to GRN-NOS.
eSignificantly impaired compared to GRN-NFV.
fSignificantly impaired compared to lvPPA. Imaging analysis Patterns of brain atrophy in the sporadic PPA cohorts were
as expected with the most atrophied regions being the left an
terior insula (83% of mean control volume in that region),
left motor (85%) and dorsolateral prefrontal cortices
(87%), and the left basal ganglia (globus pallidus 79%, pu
tamen 87%) in nfvPPA; left > right temporal pole (52%,
73%), left amygdala (58%), left hippocampus (77%) and
left nucleus accumbens (75%) in svPPA; and left lateral tem
poral (74%), supratemporal (77%) and lateral parietal cor
tices (80%) in lvPPA (Fig. 5, Supplementary Table 4). The pattern of atrophy was similar in the GRN-NFV
group compared with the sporadic nfvPPA group with the
most affected regions being the left anterior insula (67%),
left motor (80%) and left dorsolateral prefrontal cortices
(76%) as well as basal ganglia involvement (globus pallidus
86%, putamen 80%). More posterior involvement was also
seen in the GRN-NFV group with left lateral parietal volume
significantly lower than controls (83%). The linguistic symptoms in our comparator sporadic PPA
cohort were consistent with previously described patterns of
impairment.2,22 Prior studies of PPA in GRN mutations have
been relatively small, but here, we showed that two groups
emerge, one with nfvPPA and one with a PPA-NOS syn
drome. The pattern of deficits in the genetic nfvPPA group
was similar to that of sporadic nfvPPA although with quite
severe dysgraphia also in the genetic group. The PPA-NOS
group was distinct—features seen in each of the canonical
PPA were present: impaired grammar/syntax similar to
nfvPPA (but no deficit in articulation), impaired word re
trieval and speech repetition similar to lvPPA and impaired The pattern of atrophy was different in the GRN-NOS
group with a highly asymmetrical pattern affecting the left
hemisphere particularly anterior insula (75%), dorsolateral
prefrontal cortex (82%) and, unlike the other PPA groups,
the orbitofrontal cortex (85%). Other areas that were BRAIN COMMUNICATIONS 2023: Page 13 of 18 | 13 Comparison of genetic PPA and sporadic PPA 13 Figure 5 Percentage regional brain volumes in each PPA group compared with controls. These were calculated by dividing the mean of each regional brain volume in each PPA group by
the mean of the same regional brain volume in the control group and multiplying by 100. The darkest colours represent areas of lowest brain volume as per the key. Imaging analysis Abbreviations: GRN-NFV,
nonfluent variant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV, semantic variant
PPA; AMY, amygdala; AC, anterior cingulate; AI, anterior insula; CAU, caudate; DLPFC, dorsolateral prefrontal cortex; GP, globus pallidus; HIP, hippocampus; LOC, lateral occipital; LP, lateral
parietal; LT, lateral temporal; MOC, medial occipital; MP, medial parietal; MT, medial temporal; MOT, motor; NA, nucleus accumbens; OF, orbitofrontal; OP, opercular; PC, posterior cingulate; PI,
posterior insula; PUT, putamen; S, somatosensory; ST, supratemporal; THA, thalamus; TP, temporal pole; VMPFC, ventromedial prefrontal cortex. ercentage regional brain volumes in each PPA group compared with controls. These were calculated by dividing the mean of each regional brain volume in each PPA group by
f the same regional brain volume in the control group and multiplying by 100. The darkest colours represent areas of lowest brain volume as per the key. Abbreviations: GRN-NFV,
ariant PPA due to progranulin mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PPA; NFV, nonfluent variant PPA; SV, semantic variant
amygdala; AC, anterior cingulate; AI, anterior insula; CAU, caudate; DLPFC, dorsolateral prefrontal cortex; GP, globus pallidus; HIP, hippocampus; LOC, lateral occipital; LP, lateral
lateral temporal; MOC, medial occipital; MP, medial parietal; MT, medial temporal; MOT, motor; NA, nucleus accumbens; OF, orbitofrontal; OP, opercular; PC, posterior cingulate; PI,
sula; PUT, putamen; S, somatosensory; ST, supratemporal; THA, thalamus; TP, temporal pole; VMPFC, ventromedial prefrontal cortex. an of each regional brain volume in each PPA group by
ain volume as per the key. Abbreviations: GRN-NFV,
PPA; NFV, nonfluent variant PPA; SV, semantic variant
HIP, hippocampus; LOC, lateral occipital; LP, lateral
bitofrontal; OP, opercular; PC, posterior cingulate; PI,
cortex. olumes in each PPA group compared with controls. These were calculated by dividing the mean
me in the control group and multiplying by 100. The darkest colours represent areas of lowest brain
mutation; GRN-NOS, not otherwise specified PPA due to progranulin mutation; LV, logopenic variant PP
ate; AI, anterior insula; CAU, caudate; DLPFC, dorsolateral prefrontal cortex; GP, globus pallidus; HI
l occipital; MP, medial parietal; MT, medial temporal; MOT, motor; NA, nucleus accumbens; OF, orbito
osensory; ST, supratemporal; THA, thalamus; TP, temporal pole; VMPFC, ventromedial prefrontal co e calculated by dividing the me
represent areas of lowest bra ols. Imaging analysis These were calculated by dividing the me
rkest colours represent areas of lowest bra
progranulin mutation; LV, logopenic variant
eral prefrontal cortex; GP, globus pallidus;
OT, motor; NA, nucleus accumbens; OF, or BRAIN COMMUNICATIONS 2023: Page 14 of 18 K. Samra et al. 14 single word comprehension and semantic deficits (seen on
the mCCT) similar to svPPA. Sentence comprehension defi
cits (seen in both nfvPPA and lvPPA) were also prominent
in the GRN-NOS group. In other words, the PPA-NOS
group here have mixed features of the different PPA canon
ical syndromes but distinctively have impaired grammar/syn
tax in the presence of relatively intact articulation. of the GRN-NOS group in the mCCT, which is known to
have an executive component.40,41 However, there was
also impairment in Digit Span Forwards in both GRN-PPA
groups, particularly in the GRN-NOS group, potentially in
dicative of more lateral parietal cortical deficits in these
groups.42 Interestingly, the GRN-NOS group had lower scores for
the FCSRT suggesting more episodic memory impairment
in this group compared with GRN-NFV individuals and
consistent with the finding of hippocampal atrophy in this
cohort. Although not tested here, impairment of episodic
memory is more common in lvPPA and not generally seen
in nfvPPA and svPPA.43,44 Our findings contrast with a previous study of
GRN-associated PPA in which they found 28% of the cohort
had a diagnosis of nfvPPA, 25% had a ‘mixed’ PPA and 41%
held a diagnosis of lvPPA.8 One reason for this may be that
people with lvPPA are often assumed to have AD pathologic
ally and so less genetic testing is done in some centres. However, this study was a retrospective analysis, and half
of their lvPPA patients were said to have an ‘lvPPA plus’ syn
drome, i.e. features of another form of PPA including seman
tic impairment and syntax difficulties. In fact, their findings
overlap with this study in that both the ‘mixed’ and ‘lvPPA
plus’ groups have a similar pattern of deficits to those seen
in the PPA-NOS group here. Both GRN-PPA groups scored significantly worse than
controls on the tests of social cognition, the Faux Pas test
(testing theory of mind) and Facial Emotion Recognition
Test. Imaging analysis This is consistent with prior studies showing impaired
social cognition in PPA.45–47 Patterns of atrophy in the sporadic PPA subtypes were as
previously
described.48,49
In
previous
studies
of
GRN-associated PPA, a highly asymmetrical left-sided pre
dominant atrophy pattern has been described,1,50 with the
insula being the first brain region to be affected, up to 15
years prior to onset of symptomatic disease.51 However, in
our study, we were able to separate out two groups
(GRN-NFV and GRN-NOS). The pattern in the
GRN-NFV group overlapped with that of the sporadic
nfvPPA cohort, although with more posterior cortical in
volvement of the lateral parietal cortex. In contrast, the
GRN-NOS group was strikingly asymmetrical with involve
ment of the insula and dorsolateral prefrontal cortex (similar
to the GRN-NFV group) but also atrophy of the orbitofron
tal cortex (in distinction to the other forms of PPA) and the
medial temporal lobes (amygdala and hippocampus). Behavioural symptoms were present in the GRN-PPA
groups with a similar frequency and severity to sporadic
svPPA and nfvPPA but more frequent than lvPPA (which is
usually an atypical AD syndrome). Apathy was the most
common behavioural symptom in both GRN-NFV and
GRN-NOS, consistent with prior reports of this being a fre
quent feature in people with GRN mutations.31 Like the other PPA groups, depression, anxiety and irrit
ability/lability were common symptoms in the GRN-PPA
groups, although depression was less frequent in the
GRN-NOS group. Strikingly, although occurring at a low
frequency, delusions were only seen in the two GRN-PPA
groups and not the sporadic PPA groups. Although often
felt to be characteristic of C9orf72 mutations, delusions
have previously been reported in a significant minority of
people with GRN mutations.32 p
yg
pp
p
In summary, we have described a genetic nfvPPA group
similar to sporadic nfvPPA with decreased fluency, impaired
grammar/syntax and impaired articulation, but with relatively
intact single word comprehension, along with an overlapping
pattern of atrophy. The only distinctions in these syndromes
were in the non-linguistic features, with delusions occurring
only in the GRN group and more prominent motor features
and atypical parkinsonian syndromes in the sporadic
nfvPPA group. We have also described a GRN-NOS group,
similar to what has been described previously as a mixed
form of PPA, with linguistic features that are seen in each of
the sporadic PPA syndromes but distinctively grammar/syn
tax problems alongside relatively intact articulation. Funding The Dementia Research Centre is supported by Alzheimer’s
Research UK, Alzheimer’s Society, Brain Research UK and
The Wolfson Foundation. This work was supported by the
National Institute for Health Research University College
London/Hospitals
Biomedical
Research
Centre,
the
Leonard Wolfson Experimental Neurology Centre Clinical
Research Facility and the UK Dementia Research Institute,
which receives its funding from UK Dementia Research
Institute Ltd, funded by the UK Medical Research Council,
Alzheimer’s Society and Alzheimer’s Research UK. This
work was also supported by the EU Joint Programme—
Neurodegenerative Disease Research GENFI-PROX grant
[2019-02248; to J.D.R., MO, BB, CG, JvS and MS. (latter
via DLR/DFG 01ED2008B)]. J.D.R. is supported by the
Miriam Marks Brain Research UK Senior Fellowship and
has received funding from Medical Research Council
Clinician Scientist Fellowship (MR/M008525/1) and the
National Institute for Health Research Rare Disease
Translational Research Collaboration (BRC149/NS/MH). This work was also supported by the Medical Research
Council UK GENFI grant (MR/M023664/1), the Bluefield
Project and the EU Joint Programme—Neurodegenerative
Disease Research GENFI-PROX grant (2019-02248). Several authors of this publication are members of the
European Reference Network for Rare Neurological
Diseases - Project ID No 739510. R.C./C.G. are supported
by a Frontotemporal Dementia Research Studentships in
Memory of David Blechner funded through The National
Brain Appeal (RCN 290173). M.B. is supported by a
Fellowship award from the Alzheimer’s Society, UK
(AS-JF-19a-004-517). M.B.’s work is also supported by the
UK Dementia Research Institute which receives its funding
from Dementia Research Institute Ltd, funded by the UK
Medical Research Council, Alzheimer’s Society and
Alzheimer’s Research UK. C.H. was supported by Royal p
g
In conclusion, we examined the presence of PPA syn
dromes in the GENFI cohort and found that the language
network was strikingly more vulnerable to GRN mutations
than either C9orf72 or MAPT mutations. Two distinct
PPA syndromes were found in association with GRN muta
tions - those with nfvPPA, similar to sporadic nfvPPA, and
those with a PPA-NOS, who had mixed features overlapping
with the canonical sporadic disorders but, distinctively, im
paired grammar/syntax in the presence of relatively intact ar
ticulation. Better recognition of the phenotypic presentations
of GRN-related PPA are likely to trigger earlier genetic test
ing and in turn lead to improvements in management, includ
ing allowing people to enter clinical trials for those with
GRN mutations and family members to access genetic coun
selling. Imaging analysis As with
GRN-NFV, delusions are seen in a minority of individuals,
but other behavioural and neuropsychiatric features are not
particularly specific (although are more prominent than in
lvPPA which this may be mistaken for clinically, particularly
as there can be episodic memory impairment). The pattern
of atrophy is also overlapping but with unique features includ
ing involvement of the orbitofrontal cortex as well as the med
ial temporal lobe. As with previous reports, this study found that sporadic
nfvPPA was associated with the most frequent and severe
motor symptoms, with 50% having features consistent
with an atypical parkinsonian syndrome that developed be
tween 1 and 5 years after initial language symptoms, either
PSP or CBS (or in one case an overlap of both).33–36
Similar motor features were seen in the GRN-NFV group
but to a lesser extent and without anyone in this study meet
ing criteria for an atypical parkinsonian syndrome. Extrapyramidal features have been previously described in
GRN mutations (in up to 40–60% of patients37,38), and peo
ple with overlapping syndromes of PPA and CBS have also
been seen, but PSP is not a syndrome seen in GRN muta
tions.39 The less frequent occurrence of motor features and
full-blown parkinsonian syndromes (particularly PSP) ap
pears to be a distinct feature of the GRN nfvPPA form.i Non-linguistic cognitive deficits in the GRN-PPA groups
were similar to the other groups, particularly the nfvPPA co
hort and to a lesser extent the lvPPA cohort, with significant
executive dysfunction and impaired category fluency. It may
be that this deficit also contributed to the poor performance We only found one patient with a C9orf72 mutation and
one patient with a MAPT mutation who had a PPA BRAIN COMMUNICATIONS 2023: Page 15 of 18 | 15 Comparison of genetic PPA and sporadic PPA 15 syndrome (Supplementary Table 1). C9orf72-associated
PPA is very uncommon39 with a recent review finding only
a small number of cases reported in the literature.9 The clin
ical syndrome is usually nfvPPA but can also be svPPA or,
like here, a PPA-NOS. Similarly, MAPT-associated PPA is
also very uncommon. Whilst many people with a bvFTD
syndrome have associated semantic deficits,52 it is rare for
such an impairment to be the primary symptom, leading to
a diagnosis of svPPA as seen here. Limitations Firstly, despite the large numbers overall compared with pre
vious studies, when stratified, the numbers in the individual
groups are small. Nonetheless, this work provides important
evidence about the phenotype of GRN-related PPA which
will need to be substantiated in future prospective cohort
studies. Secondly, linguistic symptom data was limited to
the 10 items of the PASS, and whilst this encompasses the
majority of symptoms relevant to the canonical PPA syn
dromes, it does not separate out single word and sentence
repetition (with just a single ‘speech repetition’ item), which
can be helpful in differentiating nfvPPA and lvPPA. Furthermore, we only had data from two linguistic neurop
sychometric tests (BNT and mCCT). This is because the
GENFI neuropsychology battery was designed to include
mainly visual tasks due to the involvement of countries
speaking eight different languages and the difficulty of trans
lating and validating verbal tasks across those languages. In
future studies it would be helpful to consider developing a
multilanguage verbal linguistic battery to study cognitive
deficits further in this cohort. Thirdly, we only had data on
episodic memory and social cognition in the genetic groups
which limited the comparison with the canonical groups. Lastly, we focused on cross-sectional differences, but longi
tudinal analysis would be helpful in future studies to under
stand the progression of disease with additional symptoms
and also the presence of very early deficits, particularly in
the prodromal genetic cohort. Supplementary material Supplementary material is available at Brain Communications
online. Acknowledgements We thank the research participants and their families for
their contribution to the study. Several authors of this publi
cation are members of the European Reference Network for
Rare Neurological Diseases - Project ID No 739510. Imaging analysis understand the earliest stages of these disorders and how
they initially present in the prodromal period within the
GENFI cohort, allowing us to better stage these disorders
and identify people at a time when they can potentially
most benefit within clinical trials. Funding We recommend that future iterations of the consen
sus criteria for PPA encompass such information, with the
suggestion of testing for a GRN mutation, particularly in
those with such a phenotype. Future work will need to BRAIN COMMUNICATIONS 2023: Page 16 of 18 16 | 16 K. Samra et al. National Institute for Deaf People Dunhill Medical Trust
Pauline Ashley Fellowship (PA23_Hardy) and Wellcome
Institutional Strategic Support Fund (204841/Z/16/Z). J.C.V.S. was supported by the Dioraphte Foundation grant
09-02-03-00, Association for Frontotemporal Dementias
Research Grant 2009, Netherlands Organization for
Scientific Research grant HCMI 056-13-018, ZonMw
Memorabel (Deltaplan Dementie, project number 733 051
042), Alzheimer Nederland and the Bluefield project. F.M. received funding from the Tau Consortium and the Center
for Networked Biomedical Research on Neurodegenerative
Disease. R.S-V. is supported by Alzheimer’s Research UK
Clinical Research Training Fellowship (ARUK-CRF2017B-2)
and has received funding from Fundació Marató de TV3,
Spain (grant no. 20143810). C.G. received funding from EU
Joint Programme—Neurodegenerative Disease Research
-Prefrontals VR Dnr 529-2014-7504, VR 2015-02926
and 2018-02754, the Swedish FTD Inititative-Schörling
Foundation, Alzheimer Foundation, Brain Foundation and
Stockholm County Council ALF. M.M. has received funding
from a Canadian Institute of Health Research operating grant
and the Weston Brain Institute and Ontario Brain Institute. J.B.R. has received funding from the Welcome Trust
(103838) and is supported by the Cambridge University
Centre for Frontotemporal Dementia, the Medical Research
Council (SUAG/051 G101400) and the National Institute for
Health Research Cambridge Biomedical Research Centre
(BRC-1215-20014). E.F. has received funding from a
Canadian Institute of Health Research grant #327387. D.G. received
support
from
the
EU
Joint
Programme—
Neurodegenerative Disease Research and the Italian Ministry
of Health (PreFrontALS) grant 733051042. R.V. has received
funding from the Mady Browaeys Fund for Research into
Frontotemporal Dementia. M.O. has received funding from
Germany’s Federal Ministry of Education and Research
(BMBF), @@@FTLDc. J.L. received funding for this work by
the Deutsche Forschungsgemeinschaft German Research
Foundation under Germany’s Excellence Strategy within the
framework of the Munich Cluster for Systems Neurology
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HUBUNGAN LAMA TERAPI HEMODIALISIS DENGAN DUKACITA KRONIS PADA PASIEN CHRONIC KIDNEY DISEASE (CKD) (The Association between Duration of Hemodialysis Therapy with Chronic Sorrow in Patients with Cronic Kidney Disease (CKD))
|
Zenodo (CERN European Organization for Nuclear Research)
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cc-by
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ABSTRAK Manifestasi klinis pada pasien CKD (Chronic Kidney Diseases) dan
dampak terapi hemodialisis dapat menimbulkan masalah psikologis pada pasien
karena proses adaptasi terhadap penyakit akibat perubahan yang dialami pasien
memerlukan waktu. Pasien tersebut akan kehilangan fungsi tubuhnya dan juga
harus mendapatkan tindakan invasive yang dialami berulang ulang. Semakin lama
pasien di diagnosis menderita CKD juga akan semakin lama mengalami tindakan
hemodialisis. Keadaan ini akan memepengaruhi psikologis pasien, salah satunya
pasien dapat mengalami berduka kronis. Tujuan penelitian ini menganalisis
hubungan antara lama terapi hemodialisis dengan berduka kronis. Metode penelitian yang digunakan dalam penelitian ini adalah korelasi dengan
pendekatan cross sectional. Populasi pada penelitian adalah pasien hemodialisis di RSUD
Kabupaten Kediri dengan teknik total sampling sebanyak 72 responden. Variabel dalam
penelitian ini adalah Lama terapi hemodialisis dan duka cita kronis. Tehnik
pengumpulan data menggunakan kuesioner yaitu Kendalls Chronic Sorrow
Instrument. Hasil identifikasi lama terapi hemodialisis, Sebagian besar responden (64%)
termasuk kategori lama. Hasil identifikasi dukacita kronis, sebagian besar responden
(61%) mengalami dukacita kronis. Hasil uji korelasi antar variabel yaitu 0,003 < 0,05
artinya ada hubungan antara lama terapi hemodialisis dengan berduka kronis pada pasien
CKD dengan tingkat koefisien korelasi lemah. Hal tersebut dikarenakan kondisi berduka
kronis dapat dinormalkan apabila metode penanganan internal dan eksternal berjalan
dengan efektif. Keluarga diharapkan memberikan dukungan kepada pasien CKD yang menjalani
terapi hemodialisis supaya pasien tidak terus mengalami kondisi berduka dalam waktu
yang lama dan mampu menerima keadaanya saat ini. Volume 11, Nomor 01, Juni 2020
Hal. 69 - 78 Widyasih Sunaringtyas*, Diana Rachmania* STIKes Karya Husada Kediri, Jl. Soekarno Hatta No. 07, Pare Kediri, Jawa Timur
email: sihwidya123@gmail.com ABSTRACT Clinical manifestations in CKD (Chronic Kidney Diseases) patients and the
impact of hemodialysis therapy can cause psychological problems in patients because the
process of adaptation to the disease due to changes experienced by patients requires
time. Patients with this disease can experience chronic sorrow and sorrow, because loss
of bodily functions and invasive actions must be experienced repeatedly. The longer the
patient which diagnosed with CKD, the longer the hemodialysis action will take. This
situation will affect the patient's psychological, one of which patients can experience
chronic grieving. The purpose of this study was to analyze the relationship between
duration of hemodialysis therapy and chronic grieving. The research method used in this study, correlation with cross sectional
approach. The population in this study was hemodialysis patients in RSUD Kabupaten
Kediri with a total sampling technique of 72 respondents. The variables in this study 69 were the duration of hemodialysis therapy and chronic sorrow. Data collection
techniques using a questionnaire that was Kendalls Chronic Sorrow Instrument. were the duration of hemodialysis therapy and chronic sorrow. Data collection
techniques using a questionnaire that was Kendalls Chronic Sorrow Instrument. Identification result of the hemodialysis therapy duration was some respondent
(64%) included in the old category. Identification result of chronic sorrow was almost
some respondents (61%) experienced chronic sorrow. The result of correlation between
variables is 0.003 <0.05, which means there is a relationship between the duration of
hemodialysis therapy with chronic grieving in CKD patients with the closeness of a weak
relationship. That is because the condition of chronic sorrow can be normalized if
internal and external treatment methods work effectively. Family support was very important for Chronic Kidney Disease patients
undergoing hemodialysis therapy so that patients do not continue to experience a long
period of grieving and are able to accept their present condition. Keywords: Chronic Kidney Disease, Chronic Sorrow, Duration of Hemodialysis PENDAHULUAN adaptasi terhadap penyakit akibat
perubahan
yang
dialami
pasien
memerlukan waktu. Pasien dengan
penyakit
ini
dapat
mengalami
dukacita kronis, karena kehilangan
fungsi tubuh dan tindakan invasive
harus dialami berulang ulang. Gangguan fungsi organ pada
penderita chronic kidney desease
(CKD)
menunjukkan
berbagai
manifestasi klinis. Apabila penderita
CKD didiagnose stadium terminal
End-Stage Renal Disesase
maka
memerlukan terapi hemodialisis yang
dilakukan dalam jangka panjang atau
terapi
permanen. Hemodialisis
merupakan terapi pengganti untuk
mengeluarkan sisa-sisa metabolisme
atau racun tertentu, akan tetapi juga
terdapat
resiko
akibat
tidakan
hemodialysis (Himmelfarb, 2005). Tindakan ini dilakukan bervariasi
antara 2 sampai 4 kali setiap bulan,
tergantung kondisi pasien. Setiap kali
tindakan memerlukan waktu 4 sampai
5 jam (Mailani, 2015). Semakin lama
pasien di diagnosis menderita CKD
juga akan semakin lama mengalami
tindakan hemodialisis. Manifestasi
klinis dan dampak tindakan tersebut
dapat
menimbulkan
masalah
psikologis pada pasien karena proses Berdasarkan
WHO,
secara
global lebih dari 500 juta orang
mengalami penyakit gagal ginjal. Sekitar 1,5 juta orang harus menjalani
hidup bergantung pada cuci darah. Menurut Indonesian Renal Registry
(IRR) tahun 2016, sebanyak 98%
penderita
gagal
ginjal
menjalani
terapi hemodialisis dan 2 % menjalani
terapi peritoneal dialisis. Hasil studi
pendahuluan di ruang hemodialisis
Rumah Sakit Umum Daerah Kediri
terdapat
72
pasien
CKD
dan
menjalani hemodialisis. Sejak
pasien
dinyatakan
menderita CKD dan harus dilakukan
hemodialisis,
maka
secara
berulangkali pasien harus menjalani
terapi
hemodialisis
agar
dapat
mempertahankan kehidupan. Aspek 70 terikat yaitu dukacita kronis. Populasi
dalam penelitian ini adalah seluruh
pasien hemodialisis pada bulan Maret
2020 di RSUD Kabupaten Kediri
sebanyak 72 orang. Sampel diambil
dengan teknik total sampling dengan
jumlah 72 responden sebagai sampel
penelitian. Penelitian dilaksanakan
pada tanggal 10-17 Maret 2020. Instrumen
dalam
penelitian
ini
dengan menggunakan kuesioner yaitu
Kendalls Chronic Sorrow Instrument. Data dikumpulkan dan selanjutnya
dianalisa
dengan
uji
statistik
Spearman’s rho dengan nilai apabila
ρ value ≤ α (0,05). psikologis pasien mengalami fase
berduka
untuk
dapat
menerima
perubahan fungsi tubuhnya. Pasien
dapat mengalami kesedihan kronis,
merasa
bersalah,
menyalahkan,
bahkan merasa tidak ada harapan
(PPNI, 2017). Masalah psikologis ini
dapat terjadi berulang dan progresif,
yang
mengakibatkan
pasien
mengalami berduka kronis. Maka dari
itu, agar perubahan fungsi fisik ini
dapat
diadaptasi
dengan
cepat
diperlukan dukungan eksternal dari
pihak perawat maupun keluarga,
sehingga kondisi psikologis pasien
dapat melalui tahap tahap kehilangan
lebif
efektif. Berdasarkan
latar
belakang di atas maka peniliti perlu
meninjau lebih lanjut hubungan lama
terapi hemodialysis dengan dukacita
kronis pada pasien CKD di RSUD
kabupaten
kediri. HASIL DAN PEMBAHASAN Penelitian
yang
dilakukan
pada bulan Maret 2020 ini didapatkan
hasil distribusi frekuensi data umum
responden yang bisa dilihat pada tabel
1. Distribusi frekuensi menunjukkan
bahwa sebagian responden (50,0%)
berusia di kisaran 46-60 tahun, dan
sebagian besar responden (55,6%)
berjenis kelamin perempuan, sebagian
kecil
responden
(36,1
%)
berpendidikan
terakhir
SMA,
sebagian kecil responden (30,6%)
bekerja wiraswasta, sebagian besar
responden (69,4%) memiliki riwayat
penyakit dahulu diabetes melitus
sebanyak 31,9%, dan pola konsumsi
dikarenakan jamu-jamuan sebanyak
37,5%, lebih dari setengah responden PENDAHULUAN Tujuan
dari
penelitian ini adalah mengidentifikasi
lama terapi hemodialisis pasien CKD
dan dukacita kronis pasien CKD serta
menganalisis hubungan antara lama
terapi hemodialisis dengan dukacita
kronis pasien CKD. METODE DAN ANALISA Desain
penelitian
yang
digunakan adalah desain korelasional
dengan
menggunakan
pendekatan
cross sectional. Variabel penelitian
terdiri dari variabel bebas yaitu lama
terapi
hemodialisis
dan
variabel 71 (97,5%) frekuensi HD seminggu 2
kali. Hasil identifikasi lama terapi
hemodialisis pada pasien CKD di
Ruang
Hemodialisa
RSUD
Kabupaten Kediri dapat dilihat pada
tabel 2 yang mana didapatkan data
bahwa
sebagian
besar
(64%)
responden lama terapi hemodialisis pada kategori lama, yaitu lebih dari
12 bulan. Hasil identifikasi dukacita
kronis pada pasien CKD di RSUD
Kabupaten Kediri terlihat pada tabel 3
yang
mana
dari
tabel
tersebut
didapatkan data bahwa sebagian besar
(61%) responden mengalami dukacita
kronis. Tabel 1. Karakteristik Responden Penelitian Hubungan Lama terapi hemodialisis
dengan Duka cita kronis pada Pasien CKD
Karakteristik
Jumlah
Presentase
Usia
<30 tahun
1
1,4 %
30-45 tahun
26
36,1%
46-60 tahun
36
50.0 %
>60 tahun
9
12,5%
Jenis Kelamin
Laki – laki
32
44,4 %
Perempuan
40
55,6 %
Pendidikan
SD
16
22,2%
SMP
22
30,6 %
SMA
26
36,1 %
PT
8
11,1 %
Pekerjaan
IRT
16
22,2 %
Pedagang
18
25,0 %
Wiraswasta
22
30,6 %
PNS
7
9,7 %
Petani
9
12,5 %
Riwayat Penyakit Dahulu
Diabetes Melitus
23
31,9 %
Kolesterol Tinggi
12
16,7 %
Hipertensi
17
23,6 %
Penyakit Ginjal
20
27,8 %
Pola Konsumsi Tertentu
Makan Minum Instan
16
22,2 %
Minuman Suplemen
26
36,1 %
Jamu – jamuan
27
37,5 %
Obat – obatan
3
4,2 %
Frekuensi HD
Seminggu 1 kali
2
2,8 %
Seminggu 2 kali
70
97,2 %
Durasi HD
4 jam
72
100,0 % Tabel 1. Karakteristik Responden Penelitian Hubungan Lama terapi hemodialisis
dengan Duka cita kronis pada Pasien CKD Setelah dilakukan identifikasi
pada
masing-masing
variabel,
selanjutnya
dilakukan
analsisis
hubungan lama hemodialisis dengan Setelah dilakukan identifikasi
pada
masing-masing
variabel,
selanjutnya
dilakukan
analsisis
hubungan lama hemodialisis dengan 72 dukacita kronis pada pasien CKD
menggunakan
uji
statistik
yaitu
Spearman’s rho. Hasil tabulasi silang lama
terapi
hemodialisis
dengan
dukacita kronis dapat dilihat pada
tabel 4. Tabel 2. Identifikasi Lama Terapi Hemodialisis pada Pasien CKD Tabel 2. Identifikasi Lama Terapi Hemodialisis pada Pasien CKD
Lama terapi hemodialisis
Frekuensi
Presentase
Baru : <12 Bulan
26
36 %
Lama : ≥ 12 Bulan
46
64 %
Total
72
100 %
Tabel 3. Identifikasi Dukacita Kronis pada Pasien CKD
Dukacita Kronis
Frekuensi
Presentase
Tidak Dukacita
28
39 %
Dukacita Kronis
44
61 %
Total
72
100 % Tabel 3. METODE DAN ANALISA Identifikasi Dukacita Kronis pada Pasien CKD Pemrograman
pelaksanaan
terapi
hemodialisis diatur oleh specialist
penyakit dalam konsultan ginjal untuk
menentukan
berbagai
parameter
proses dialysis seperti frekuensi, lama
HD, kecepatan aliran darah, cairan
dialisat dan konsentrasi dialisat yang
dipakai. Pasien
CKD
biasanya
mendapat terapi hemodialisi sebanyak
2-3 kali perminggu selama 4-5 jam
(Tjokroprawiro,
2015). Penelitian
yang dilakukan oleh Wahyuni dkk
(2018) menyebutkan semakin lama
pasien menjalani HD, maka pasien
semakin patuh untuk menjalani HD
karena
biasanya
pasien
telah
mencapai
tahap
menerima
dan
kemungkinan pasien telah banyak
mendapatkan pendidikan kesehatan
dari perawat dan juga dokter tentang
penyakit dan pentingnya menjalani Lama terapi hemodialisis pada
pasien CKD di RSUD Kabupaten
Kediri Hasil penelitian untuk variabel
lama terapi hemodialisis pasien CKD
di didapatkan data bahwa sebagian
besar (64%) responden lama terapi
hemodialisis pada kategori lama,
yaitu lebih dari 12 bulan dan hampir
setengah responden (36%) terapi
hemodialisis dalam kategori baru
yaitu kurang dari 12 bulan. Dialisis
merupakan salah satu cara terapi
pengganti
ginjal
akibat
tidak
berfungsinya organ ginjal. Tujuan
utama dari Hemodialisis adalah untuk
mengeluarkan toksin uremik dan
cairan serta pengaturan keseimbangan
cairan, elektrolit dan asam basa. Penderita CKD membutuhkan terapi
hemodialisis seumur hidupnya guna
meningkatkan kualitas hidup pasien. 73 73 HD secara teratur (Wahyuni dkk,
2018). mampu menjalankan fungsinya untuk
mengeluarkan
racun-racun
dalam
tubuh serta tidak mampu menjaga
keseimbangan cairan di dalam tubuh. Jika penderita CKD tidak melakukan
terapi hemodialisis maka akan timbul
komplikasi seperti kelebihan cairan
dalam tubuh yang bisa menyebabkan
gangguan
system
pernafasan,
ketidakseimbangan asam basa yang
bisa menyebabkan kematian. Dengan
demikian
seseorang
yang
telah
terdiagnosa CKD maka hidupnya pun
akan bergantung pada terapi dialysis. Lama seseorang menjalani terapi
hemodialisis
menunjukkan
bahwa
penderita telah lama terdiagnosa CKD
dan perlu rutin untuk melaksanakan
terapi hemodialisis. Hasil penelitian didapatkan
data bahwa sebagian besar responden
masuk dalam kategori lama menjalani
terapi hemodialysis yaitu lebih dari
12
bulan,
dan
hampir
seluruh
responden
menjalani
hemodialisis
dengan frekuensi 2 kali setiap minggu
selama 4 jam dalam satu kali terapi
hemodialisis. Sesuai dengan yang
dikemukakan
oleh
Tjokroprawiro
(2015) bahwa seorang penderita CKD
perlu
mendapatkan
terapi
hemodialisis seumur hidupnya untuk
meningkatkan kualitas hidupnya. Hal
tersebut dikarenakan seseorang yang
mengalami CKD telah mengalami
kerusakan ginjal sehingga ginjal tidak Tabel 4. Tabulasi silang lama terapi hemodialisis dengan Dukacita Kronis pada
Pasien CKD
Lama Terapi Hemodialisis
Dukacita Kronis
Total
Tidak Dukacita
Dukacita Kronis
f
%
f
%
Baru < 12 bulan
16
22
10
14
26
Lama > 12 bulan
12
17
34
47
46
Total
28
39
44
61
72
= 0,05 p Value= 0.003 r = 0,349 Dukacita Kronis pada pasien CKD
di RSUD RSUD Kabupaten Kediri Dukacita kronis pada hasil
penelitian ini juga tidak dipengaruhi
jenis kelamin, karena setiap individu 75 0,003 < 0,05 artinya ada hubungan
antara
lama
terapi
hemodialisis
dengan berduka kronis pada pasien
CKD. Sedangan koefisien korelasi
sebesar 0,349 menunjukkan kekuatan
hubungan lemah dengan arah positif. Pengalaman kehilangan berlaku untuk
individu yang mengalami maupun
pemberi
asuhan
dapat
terjadi
kesenjangan
karena
tidak
sesuai
harapan sehingga mengalami dukacita
kronis,
persaan
dukacita
meresap/mendalam,
menetap/
permanen,
berkala/periodik
dan
berpotensi progresif (Alligood, 2017). Chronic kidney deseases merupakan
kondisi
kehilangan
fungsi
tubuh
sehingga
memerlukan
terapi
hemodialisis
untuk
menggantikan
fungsi ginjal. akan tetapi terapi ini
juga
terdapat
berbagai
resiko,
hipotensi, kram otot, mual muntah,
perdarahan, hipertensi (Himmelfarb,
2005). Selaras dengan teori tersebut
pengalaman kehilangan fungsi organ
tubuh
dan
terapi
hemodialisis
dilakukan secara rutin dan berulang
ulang merupakan keadaan yang
menimbulkan rasa nyeri. Pengalaman
ini tidak hanya mempengaruhi fisik
tapi juga mempengaruhi psikologis
responden sehingga timbul perasaan
sedih berulang ulang dan meresap
sesuai dengan parameter berduka menyatakan tidak akan menyerah
terhadap kondisi yang dialaminya. Sesuai penelitian Sunaringtyas dan
Rachmania (2019) perasaan berduka
kronis dialami oleh pasien CKD yang
menjalani terapi hemodialisis. Pasien
dengan
penyakit
ini
mengalami
gangguan fungsi
ginjal sehingga
merasa kehilangan fungsi tubunya. Perasaan berduka diakibatkan oleh
manifestasi kegagalan fungsi ginjal
yang dirasakan pasien. Keluhan yang
sering dialami pasien antara lain:
tubuh merasa tidak nyaman, mual,
pusing, sesak nafas malaise yang
dirasakan dalam waktu yang panjang. Seolah olah penyakit yang dirasakan
terus menerus dan tidak ada akhirnya. Namun demikian, Sebagian kecil
responden tidak mengalami dukacita
kronis. Hal ini mungkin disebabkan
oleh mekanisme koping individu yang
sudah
mulai
bisa
menerima
kondisinya meskipun harus menjalani
terapi hemodialisis secara rutin. Dukacita Kronis pada pasien CKD
di RSUD RSUD Kabupaten Kediri perasaan berduka atau kesedihan lain
yang
terkait
dengan
adanya
kesenjangan yang sedang berlangsung
yang ditimbulkan oleh pengalaman
kehilangan
(Eakes,
Burke,
&
Hainsworrth dalam Alligood, 2014). Gejala
berduka
yang
disebutkan
dalam
Standar
Diagnosis
Keperawatan Indonesia dapat berupa Hasil penelitian untuk variable
dukacita kronis yaitu sebagian besar
responden (61%) mengalami dukacita
kronis
dan
hamper
setengah
responden (39%) tidak mengalami
dukacita kronis. Dukacita kronis
merupakan kesedihan mendalam, atau 74 merasa sedih, merasa bersalah, tidak
menerima kehilanagn, merasa tidak
ada harapan, menangis, pola tidur
berubah, tidak mampu berkonsentrasi,
merasa tidak berguna dan juga marah
(Tim POkja SDKI DPP PPNI, 2017). Duka kronis dapat dialami pasien
dengan
kasus
paliatif,
misalnya
kanker,
penyakit
degeneratife,
penyakit paru obstruksi kronis, cystic
fibrotic, stroke, parkinson, gagal
jantung,
penyakit
genetika
dan
penyakit infeksi seperti HIV/AIDS,
CKD
(Kemenkes
No.812
tahun
2007). Pada kasus CKD memerlukan
tindakan
invasive,
salah
satunya
adalah
hemodialisis. Hemodialis
merupakan tindakan yang dilakukan
berulang yang dapat menimbulkan
efek secara fisik maupun psikologia. Efek fisik yang terjadi adalah rasa
nyeri,
gangguan
tekanan
darah,
anemia,
kekakuan
otot,
pembengkakan. Sedangkan
efek
psikologis antara lain: Stres, rasa
takut, depresi, kecemasan, putus asa. Oleh sebab itu perasaan berduka
kronis dialami oleh pasien CKD yang
menjalani
terapi
hemodialisis
(Sunaringtyas dan Rachmania, 2019). Dukacita
Kronis
pada
responden di alami pada semua baik laki-laki maupun perempuan
mempunyai peran masing masing
untuk menjalani kehidupan sehingga
apabila
individu
tidak
mampu
memenuhi kebutuhan sesuai peran
yang diharapkan dapat mengalami
kesedihan dan berlangsung lama
karena seperti yang diketahui bahwa
penderita CKD perlu melakukan
hemodialisis seumur hidupnya guna
meningkatkan
kualitas
hidupnya,
namun disisi lain prosedur invasive
akibat
hemodialisis
sendiri
menimbulkan efek samping berupa
efek
fisik
dan
psikologis
yang
mendalam bagi responden. Bentuk dukacita kronis yang
dialami
oleh
responden
dalam
penelitian ini sesuai dengan yang
beberapa gejala yang disebutkan
dalam
Standar
Diagnosis
Keperawatan
Indonesia
dimana
dukacita kronis yang dialami oleh
responden dalam penelitian ini dari
hasil rekapitulasi Kendalls Chronic
Sorrow
Instrument
yaitu
berupa
sebagian besar responden merasakan
sedih dengan kondisi penyakit yang
dialaminya, perasaan sedih tersebut
datang dan pergi, Sebagian responden
menyatakan bahwa hidupnya sudah
tidak lagi sesuai harapan sejak jatuh
dalam kondisi sakit dan menjalani
terapi hemodialisis, bingung dan
khawatir dengan kondisi hidupnya
setelah
mengalami
kejadian
ini. Namun
sebagian
responden Dukacita
Kronis
pada
responden di alami pada semua
tingkat usia. berdasarkan data tersebut
perasaan dukacita tidak dipengaruhi
factor usia. KESIMPULAN DAN SARAN Creswell, J.W. (2010). Research
design:
Qualitative
and
quantitative
approaches. Third
edition. California:
SAGE Publication. Saran Pasien dengan CKD sangat
membutuhkan
dukungan
dari
keluarga agar bisa menjalani terapi
Hemodialisa
dengan
baik. Oleh
karena itu kepada keluarga pada
pasien dengan CKD penting untuk
selalu
memberikan
dukungannya
kepada responden dengan CKD yang
menjalani terapi hemodialisis. Hal
tersebut
dikarenakan
adanya
dukungan keluarga membantu pasien
untuk
bisa
segera
beradaptasi/menerima kondisi yang
bisa dialaminya saat ini Hubungan lama terapi hemodialisis
dengan
dukacita
kronis
pada
pasien CKD di RSUD Kabupaten
Kediri Hasil tabulasi silang antar
variabel menujukkan data bahwa
hampir setengah responden (47%)
responden
mengalami
terapi
hemodialisis dengan kategori lama
(lebih dari 12 bulan) dan mengalami
dukacita
kronis. Nilai
signifikan 76 keadaan psikologis pasien, gangguan
proses berfikir dan konsentrasi serta
gangguan dalam berhubingan social. dukacita
kronis
dengan
tingkat
koefisien korelasi lemah. Sebagian
besar
responden
dengan
CKD
menjalani terapi hemodialisis pada
kategori lama (> 12 bulan) dan
sebagian besar responden mengalami
dukacita kronis. Hasil penelitian ini menjukkan
hubungan dua variabel pada kategori
lemah karena terdapat 12 responden
(17%) menjalani terapi hemodialisis
dengan kategori lama tetapi tidak
mengalami berduka kronis dan 10
responden (14 %) menjalani terapi
hemodialisis termasuk kategori baru
akan
tetapi
mengalami
berduka
kronis. Hubungan antar variabel ini
sesuai dengan teori Eakes bahwa
kondisi kehilangan dapat dinormalkan
apabila metode penanganan internal
dan
eksternal
efektif. Sesuai
pengamatan peneliti adanya support
system dari keluarga maupun praktisi
kesehatan mendukung penanganan
kehilangan
pengalaman
pada
responden. Pengalaman kehilangan
yang dialami oleh setiap individu
dipersepsikan bervariasi dan berbeda
oleh setiap individu. Hal ini terbukti
dukungan
yang
diberikan
oleh
keluarga berupa kehadiran anggota
keluarga pada saat proses terapi
sampai selesai, dapat memberikan
ketenangan bagi pasien sehingga
tercipta kenyamanan. KEPUSTAKAAN Alligood Martha R.(2017). Pakar
teori Keperawatan dan Karya
mereka,8th Indonesia edition,
by Achir Yani S.Hamid and
Kusman Ibrahim Atimiati
WD
.(2012). Tingkat
kualitas hidup pasien gagal
ginjal
kronik
terapi
hemodialysis, KEMAS 1 (2):
EMAS. 1(2):1047–53. Kesimpulan Terdapat
hubungan
antara
lama
terapi
hemodialisis
dengan 77 Kesehatan
Andalas. http://jurnal.fk.unand.ac.id/in
dex.php/jka/article/download/
905/759. (7 Mei 2020) Kesehatan
Andalas. http://jurnal.fk.unand.ac.id/in
dex.php/jka/article/download/
905/759. (7 Mei 2020) Himmelfarb, Jonathan. (2005). Core
Curriculum
In
Nephrology
Hemodialysis
Complications.National
Kidney Foundation. N Eng J
M. Doi : 10.1053 http : //
www.nejm.org/content/full
article.htm (30 April 2008) Kartika, Dani. (2017). Hubungan
Lama
menjalani
Terapi
Hemodialisis dengan Kualitas
Hidup Pasien Penyakit Ginjal
Kronik
di
Instalasi
Hemodialisis
RSUD
Abdul
Moeloek
http://digilib.unila.ac.id/id/epr
int/25325 Keputusan Menteri Kesehatan RI
NOMOR
:
812/Menkes/SK/VII/2007
Tentang Kebijakan Perawatan
Paliatif NANDA-I,
Nursing
Diagnosis
Definition and Clasification,
2018-2020 NANDA-I,
Nursing
Diagnosis
Definition and Clasification,
2018-2020 Sunaringtyas W.& Rachmania D. (2019). Monograf
Chronic
Sorrow
dan
Hynotherapy,
ISBN:
978-602-462—375-3,
MNC, Malang. Tim Pokja SDKI DPP PPNI. (2017). Standar
Diagnosis
Keperawatan
Indonesia:
Definisi
dan
Indikator
Diagnostik. Jakarta: Dewan
Pengurus Pusat PPNI
Tjokroprawiro,
Askandar. (2015). Buku
ajar
ilmu
penyakit
dalam. Ed.2. Airlangga
University Press
Wahyuni,
Putri;
Miro,
Septino;
Kuniawan,
Eka. (2018). Hubungan Lama Menjalani
Hemodialisis dengan Kualitas
Hidup Pasien Penyakit Ginjal
Kronik
dengan
Diabetes
Melitus di RSUP Dr. M
Djamil
Padang. Jurnal Tim Pokja SDKI DPP PPNI. (2017). Standar
Diagnosis
Keperawatan
Indonesia:
Definisi
dan
Indikator
Diagnostik. Jakarta: Dewan
Pengurus Pusat PPNI
Tjokroprawiro,
Askandar. (2015). Buku
ajar
ilmu
penyakit
dalam. Ed.2. Airlangga
University Press
Wahyuni,
Putri;
Miro,
Septino;
Kuniawan,
Eka. (2018). Hubungan Lama Menjalani
Hemodialisis dengan Kualitas
Hidup Pasien Penyakit Ginjal
Kronik
dengan
Diabetes
Melitus di RSUP Dr. M
Djamil
Padang. Jurnal Tim Pokja SDKI DPP PPNI. (2017). Standar
Diagnosis
Keperawatan
Indonesia:
Definisi
dan
Indikator
Diagnostik. Jakarta: Dewan
Pengurus Pusat PPNI 78
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https://openalex.org/W2132474561
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English
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The role of pulmonary arterial stiffness in COPD
|
Respiratory medicine
| 2,015
|
cc-by
| 14,746
|
a r t i c l e i n f o 1386 (Cor Pulmonale when associated with right heart failure in COPD) is
relatively common in severe COPD and is present in >50% of pa-
tients awaiting lung transplantation [2]. The incidence increases
with increasing severity of COPD, with pulmonary hypertension
present in 5% of moderate (GOLD stage II), 27% of severe (GOLD
stage III) and 53% of very severe (GOLD stage IV) COPD [3]. The true
incidence in mild to moderate COPD however is poorly appreciated
due to the absence of large scale epidemiological studies [4]. In Respiratory Medicine 109 (2015) 1381e1390 Respiratory Medicine 109 (2015) 1381e1390 a r t i c l e i n f o . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1385
2. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1386
Acknowledgements: Funding, disclosures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1386
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . * Corresponding author. Division of Cardiovascular and diabetic medicine,
Ninewells Hospital, Dundee DD1 9SY, United Kingdom.
E-mail address: j.weirmccall@dundee.ac.uk (J.R. Weir-McCall). The role of pulmonary arterial stiffness in COPD
Jonathan R. Weir-McCall a, *, Allan D. Struthers a, Brian J. Lipworth b, J. Graeme Houston a
a Division of Cardiovascular and Diabetes Medicine, Medical Research Institute, University of Dundee, Dundee, United Kingdom
b Scottish Centre for Respiratory Research, Medical Research Institute, University of Dundee, Dundee, United Kingdom The role of pulmonary arterial stiffness in COPD
Jonathan R. Weir-McCall a, *, Allan D. Struthers a, Brian J. Lipworth b, J. Graeme Houston a
a Division of Cardiovascular and Diabetes Medicine, Medical Research Institute, University of Dundee, Dundee, United Kingdom
b Scottish Centre for Respiratory Research, Medical Research Institute, University of Dundee, Dundee, United Kingdom a r t i c l e i n f o . . . . . . . . . . . . . . . . . . 1385
2. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1386
Acknowledgements: Funding, disclosures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1386
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1386 Contents
1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a r t i c l e i n f o . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1381
1.1. Vascular changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1382
1.1.1. The right ventriculo-arterial model . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1382
1.1.2. Measuring pulmonary artery stiffness . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1384
1.1.3. Pulmonary arterial stiffness as a treatment target . . . . . . . . . . . . . . . . . . . . . . . . . . . a r t i c l e i n f o Article history:
Received 10 January 2015
Received in revised form
10 May 2015
Accepted 10 June 2015
Available online 12 June 2015
Keywords:
Pulmonary disease
Chronic obstructive
Hypertension
Pulmonary
Vascular capacitance
Vascular resistance
Pulmonary heart disease Article history:
Received 10 January 2015
Received in revised form
10 May 2015
Accepted 10 June 2015
Available online 12 June 2015 COPD is the second most common cause of pulmonary hypertension, and is a common complication of
severe COPD with significant implications for both quality of life and mortality. However, the use of a
rigid diagnostic threshold of a mean pulmonary arterial pressure (mPAP) of 25mHg when considering
the impact of the pulmonary vasculature on symptoms and disease is misleading. Even minimal exertion
causes oxygen desaturation and elevations in mPAP, with right ventricular hypertrophy and dilatation
present in patients with mild to moderate COPD with pressures below the threshold for diagnosis of
pulmonary hypertension. This has significant implications, with right ventricular dysfunction associated
with poorer exercise capability and increased mortality independent of pulmonary function tests. The compliance of the pulmonary artery (PA) is a key component in decoupling the right ventricle
from the pulmonary bed, allowing the right ventricle to work at maximum efficiency and protecting the
microcirculation from large pressure gradients. PA stiffness increases with the severity of COPD, and
correlates well with the presence of exercise induced pulmonary hypertension. A curvilinear relationship
exists between PA distensibility and mPAP and pulmonary vascular resistance (PVR) with marked loss of
distensibility before a rapid rise in mPAP and PVR occurs with resultant right ventricular failure. This
combination of features suggests PA stiffness as a promising biomarker for early detection of pulmonary
vascular disease, and to play a role in right ventricular failure in COPD. Early detection would open this
up as a potential therapeutic target before end stage arterial remodelling occurs. © 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license
(http://creativecommons.org/licenses/by/4.0/). Contents Contents Contents
1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a r t i c l e i n f o . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1381
1.1. Vascular changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1382
1.1.1. The right ventriculo-arterial model . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1382
1.1.2. Measuring pulmonary artery stiffness . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1384
1.1.3. Pulmonary arterial stiffness as a treatment target . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E-mail address: j.weirmccall@dundee.ac.uk (J.R. Weir-McCall). * Corresponding author. Division of Cardiovascular and diabetic medicine,
Ninewells Hospital, Dundee DD1 9SY, United Kingdom. http://dx.doi.org/10.1016/j.rmed.2015.06.005
0954-6111/© 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4 1.1. Vascular changes Vascular remodelling within COPD develops secondary to
intimal hyperplasia, with muscularisation of the arterioles and with
rather little change in the tunica media [21,22]. This hyperplasia is
induced by proliferation of smooth muscle cells and the deposition
of elastic and collagen fibres [23,24]. Originally hypoxia was held to
be the driving force behind this remodelling, however similar
arterial changes are present in smokers with and without COPD,
and arterial changes can be observed in mild COPD when hypoxia is
yet to develop [10,23,25]. The vascular remodelling in COPD is most
likely due to endothelial dysfunction and apoptosis which in turn
causes remodelling, capillary loss and small vessel thrombosis [26]. This is likely due to a combination of local inflammatory mediators
released from the endothelial dysfunction in the adjacent pulmo-
nary parenchyma, and direct effects from the inhaled cigarette
smoke on the vascular endothelium [24,26]. However an experi-
mental study by Ferrer et al. in guinea pig models shows hypoxia
still plays a contributory role, as a hypoxic environment after
cigarette exposure induced significantly greater changes in pul-
monary arterial pressures and pulmonary vascular remodelling
than cigarette smoking or hypoxia alone [17]. More recently the
pulmonary vascular sympathetic nerves which control the pul-
mopulmonary baroreceptor reflex originating at the pulmonary
trunk has been proposed to be of significant importance in the
development of pulmonary arterial hypertension [27]. This is evi-
denced by a rise in pulmonary arterial pressures and pulmonary
vascular resistance (PVR) induced by stretching the branch pul-
monary arteries [27e30]. In a single study by Chen et al. [31] on 21
patients
with
significant
pulmonary
hypertension
despite
maximum
medical
therapy,
intravascular
pulmonary
artery
denervation with cessation of medical therapy resulted in signifi-
cant improvement in pulmonary pressures and exercise capacity. However the role of this reflex in humans is still controversial and
has also yet to be evaluated in COPD induced pulmonary hyper-
tension [32]. 1.1.1.2. Pulmonary artery. There has been a great deal of recent
interest in systemic arterial stiffening in COPD due to the mecha-
nistically plausible link this provides between COPD and the excess
cardiovascular mortality associated with this condition [51]. In
comparison to the systemic arterial circulation there are several key
differences in the nature of the vascular tree between that of the
pulmonary and systemic circulation. 1. Introduction COPD is the second most common cause of pulmonary hyper-
tension after left sided heart disease [1]. Pulmonary hypertension J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 1382 no significant alterations in the pulmonary arterial or systemic
pressures following open ablation of the right heart in a dog model,
leading one of the authors to conclude that “a normal, contractile
right ventricular wall is not necessary for the maintenance of a
normal circulation” [33e35]. Despite this apparent swan song for a
significant role of the right ventricle, a paper in 1982 demonstrated
that the open pericardial model did not accurately represent the
functional haemodynamics of the heart [36]. Goldstein et al. demonstrated that when the right coronary artery was infarcted
within an intact pericardium there was a profound and rapid fall in
left ventricular pressures due to a fall in left ventricular (LV) pre-
load, equalisation of RV and LV pressures and elevated intra-
pericardial pressures which did not occur when they repeated their
experiment after pericardiectomy. Thus there is a marked ventric-
ular interdependence due to the shared pericardial compartment,
shared interventricular septum and the dependence of the left
heart preload on the output of the right [37e43]. comparison to other causes of pulmonary hypertension the pul-
monary pressure elevations are modest, however survival is poor
and correlates better with the pulmonary pressures and pulmonary
vascular resistance than with the severity of airflow obstruction
[5e8]. comparison to other causes of pulmonary hypertension the pul-
monary pressure elevations are modest, however survival is poor
and correlates better with the pulmonary pressures and pulmonary
vascular resistance than with the severity of airflow obstruction
[5e8]. Pulmonary hypertension is defined as a mean pulmonary arte-
rial pressure (mPAP) greater than or equal to 25 mmHg at rest [9]. However: Pulmonary vascular changes occur in mild COPD before
pulmonary hypertension occurs [10]; Right ventricular dysfunction
has been observed in normoxaemic COPD patients with normal
pulmonary arterial pressures [11]; and right ventricular hypertro-
phy is one of the earliest cardiac changes and is present even in
mild COPD [12e14]. Additionally in both mice and guinea pig
models, vascular remodelling occurs even before the onset of
emphysema [15e17]. 1. Introduction The notion of an elevated rest pressure as a
cut-off is also restrictive in evaluating the role of the pulmonary
arteries and right heart as even minimal exertion causes marked
elevations in mPAP and oxygen desaturation in COPD, with exercise
induced pulmonary hypertension present in 35%e58% of patients
with normoxaemia or mild hypoxia [5,18e20]. Recent work has shown the importance of the right heart in
health and disease. In health the right ventricle responds to exer-
cise, obesity and traditional cardiovascular risk factors in a manner
similar to, but independent of, the left ventricle [44e46]. A
decreased RV mass is associated with an increased risk of devel-
oping dyspnoea, and an increased RV mass is associated with an
increased risk of future congestive cardiac failure and cardiovas-
cular mortality in a healthy population [47,48]. In patients with
COPD, right ventricular remodelling is apparent even early in the
disease with right ventricular hypertrophy and diastolic dysfunc-
tion present in mild and moderate COPD with systolic impairment
occurring in severe COPD [12,13]. This has significant implications
with right ventricular dysfunction associated with poorer exercise
capabilities and increased mortality independent of pulmonary
function tests [49,50]. Therefore the pulmonary hypertension model, whereby right
ventricular failure occurs once pulmonary hypertension exists, is a
misnomer, and an alternate should be considered - the right
ventriculo-arterial model. 1.1. Vascular changes which showed the pulmonary arterial wall thickness to be related
to exercise pulmonary pressures rather than resting pulmonary
pressures, and to correlate highly with change in pressure from rest
to exercise [99]. The compliance of the pulmonary artery is a key component in
decoupling the right ventricle from the pulmonary bed, allowing
the right ventricle to work at maximum efficiency and protecting
the microcirculation from large pressure gradients [52,64]. Indeed
the stiffness of the pulmonary artery is a strong determinant of
right ventricular function [65], and increased stiffness causes distal
pulmonary arterial endothelial dysfunction and inflammation
[66,67]. Increased stiffness is independently associated with
reduced functional capacity [68], and higher mortality than the
pulmonary artery pressures or pulmonary vascular resistance
[69e73]. While vascular distensability is partly dependant on un-
derlying distending pressures [74,75], multiple studies have shown
the intrinsic stiffness of the arterial walls to be increased in pul-
monary hypertension independently of these [76e80]. 1.1.1.3. Right ventriculo-arterial model. Optimal ventriculo-arterial
coupling occurs when there is maximal transference of potential
energy from one elastic chamber to another, e.g. from the ventricle
to the artery. This is dependent on the elastance e the change in
pressure for a given change in volume e of the two chambers [81]. The ventricular elastance (EES) is a load independent measure of the
ventricular contractility combined with the modulating effects of
the geometric and structural properties of the ventricle [82]. It
relies on measuring the response of the ventricular pressure to
increasing volumes (see Fig. 1). The arterial elastance (EA) is an When the EA increases so that the EA:EES is consistently above
0.5 the ventricle adapts to the increased workload by inducing
hypertrophy [101]. This increases the EES thereby returning the
ventriculo-arterial system to its most energy efficient state, albeit at
a slightly higher energy consumption than previously [102e104]. As EA continues to rise the right ventricle eventually fails to adapt
with a resultant decrease in the EES and thereby a rapid increase in
EA/EES indicating uncoupling and reduced myocardial efficiency
[87,105]. Fig. 1. Calculation of EA:EES in the right ventricle and pulmonary artery. Ventricular
end systolic pressure-volume is linear and characterised by the slope Ees and is
generated by measuring pressure-volume loops under gradated preload and afterload
conditions. Reproduced from Wang et al., 2011 [100]. Ea ¼ Arterial elastance;
Ees ¼ Ventricular elastance. Fourie et al. 1.1. Vascular changes This energy is
considered wasted in terms of ventricular power output, but is
crucial in acting as a blood reservoir and maintaining steady flow
through the pulmonary tree [57]. As the pulmonary artery stiffens
the oscillatory component of the workload increases, although re-
mains as a constant of the total energy expenditure of the right
ventricle [58,59]. As the artery stiffens it also starts to dilate and
become less compliant, and the dilation of this in relation to the
aorta has been shown to be a useful clinical marker in COPD, with a
PA:aortic ratio>1 an independent predictor of future acute exac-
erbations, exercise capacity, pulmonary hypertension and mortality
[60e63]. At rest the ventricle and pulmonary artery are coupled so that
the right ventricle is working at optimal efficiency which occurs
when the stroke work is maximal with minimal oxygen con-
sumption [55,85e87]. This has been shown to occur when EA is half
EES [87e89]. In exercise this moves towards optimal coupling when
there is maximal cardiac efficiency which is where there is
maximum flow generation with minimal energy loss (EA ¼ EES)
[90e92]. Interestingly the optimal values for energy efficiency and
cardiac efficiency are similar throughout a variety of studied
mammals suggesting it has been conserved throughout mamma-
lian evolution [93,94]. During exercise, arterial stiffness has a pro-
gressively and intensity-dependant greater impact on EA than PVR
with the result that EA increases on exercise despite a fall in PVR in a
manner paralleling the rise in arterial stiffness [95e97]. A similar
relationship is present in pulmonary hypertension whereby pul-
monary stiffness has a 1.2e18 fold greater contribution to right
ventricular workload than PVR [74,76,54]. In the systemic circula-
tion, age and hypertension (two known factors associated with
increased arterial stiffness) result in an exaggerated increase in EA
during exercise increasing the pulsatile load on the heart and in
turn decreasing left ventricular efficiency [90,98]. This exaggerated
response secondary to arterial stiffening may explain the rapid rise
in mPAP observed in COPD during exercise despite a stable or
minimal rise in PVR [18]. Indeed in a study by Hilde et al., an in-
crease in pulmonary arterial stiffness was observed during exercise
with a greater contribution from this to changes in mPAP than PVR
[3]. Further indirect evidence comes from a study by Kubo et al. 1.1. Vascular changes The pulmonary artery is a low
pressure, high distensibility system which acts to transform the
highly pulsatile right ventricular output into the near steady flow at
the capillary level [52]. The distal pulmonary vascular bed is also
comprised of highly distensible vessels that, through distension
and recruitment of the microvascular bed, are able to accommodate
large increases in volume such as during exercise. This response is
the opposite of that experienced in the systemic circulation where
exercise induces vasoconstriction in the majority of the vascular
tree other than the vessels supplying the muscles. This inbuilt
reserve means that resting pulmonary arterial pressures will only
rise relatively late in a disease process when 60e70% of the bed is
obstructed [53]. The peripheral distensability of the vessels also
differs from the systemic circulation where only the proximal ar-
teries act as capacitance vessels, expanding and contracting with
each beat of the heart, while the peripheral vessels are responsible
for resistance. Due to the dual function of the vessels, there is an
inverse relationship between the resistance and capacitance of the
pulmonary circulation, with the product of the two (the resistance-
compliance time constant) remains the same in both health and
disease [54]. The pulmonary arteries and microcirculation have distinct ef-
fects on the right ventricular afterload which can be split into two
key components e the steady component required to drive and
maintain forward flow and the oscillatory component required to
overcome the pulsatile component of flow. The steady component 1.1.1. The right ventriculo-arterial model
1.1.1.1. Right ventricle. The right ventricle (RV) got off to an inaus-
picious start in the 1940s and 50s when a series of papers showed 1.1.1. The right ventriculo-arterial model 1.1.1.1. Right ventricle. The right ventricle (RV) got off to an inaus-
picious start in the 1940s and 50s when a series of papers showed J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 1383 index incorporating the elements affecting arterial load including
pulmonary vascular resistance, arterial compliance, characteristic
impedance, and systolic and diastolic time interval [83,84]. The EA/
EES calculates the extent of ventriculo-arterial coupling. is a function of peripheral vascular resistance whereas the oscilla-
tory component is a function both proximal and distal pulmonary
arterial stiffness [55]. Even in the healthy adult, 30% of the right
ventricular output is spent on generating pulsations (compared
with only 10% in the systemic circulation) [56]. 1.1.2. Measuring pulmonary artery stiffness The stiffness of the pulmonary artery can be quantified using a
variety of metrics, with these measuring stiffness locally, regionally
or systemically. Table 1 summarises the various methods of
measuring arterial stiffness and the common methods for acquiring
these. As can be seen from the table the majority of these require
knowledge of the arterial pressures to calculate the stiffness. In the
arterial circulation this is not a significant issue as the brachial
arterial pressures are readily obtained with a sphygmomanometer,
as mean BP and diastolic BP are relatively constant throughout the
large arteries [107]. However in the pulmonary arteries this pro-
vides a significant hurdle as an external measurement of the
pressures is not readily available. Despite numerous attempts to
quantify pulmonary arterial pressures and vascular resistance using Table 2
Summary of studies looking at invasive measurement of pulmonary arterial stiffness. Study
Population
No
Method
Finding Table 2
Summary of studies looking at invasive measurement of pulmonary arterial stiffness. Study
Population
No
Method
Finding
Milnor et al. [118]
Mixed PH
7
RHC
Increased impedance in PH
Nakayama et al. [119]
Mixed PH
62
RHC
Increased augmentation index and earlier inflection time in PH
Castelain et al. [77]
Mixed PH
14
RHC
Increased augmentation index and earlier inflection time in PH
Mahapatra et al. [70]
Mixed PH
104
RHC
Capacitance strongest predictor of mortality in PH. On multivariate analysis, capacitance
was the only statistically significant predictor of mortality. Hilde et al. [11] a
COPD
98
RHC
PA compliance greatly reduced in COPD Gold 2,3 and 4
Hilde at al [3] a
COPD
98
RHC
PA compliance fell with exercise in patients with and without PH, with similar elevations
in mPAP during exercise between the two groups. Muthurangu et al. [120]
IPAH
17
RHC with MRI
Inverse relationship between compliance and PVRI and mPAP. A fall in compliance was
evident in 7/17 following nitric oxide inhalation
Kuehne et al. [87]
Mixed
12
RHC with MRI
Increased elastance in PH, with resultant ventriculo-arterial uncoupling
Kopec et al. [121]
IPAH
26
RHC with IVUS
PWV high in IPAH (mean PWV 10 ms1) with excellent correlation with compliance
(b ¼ 0.81) and reasonable correlation with mPAP (b ¼ 0.48)
Lau et al., 2012 [79]
IPAH
8
RHC with IVUS
Increase in all measure of pulmonary arterial stiffness. Inverse curvilinear relationship
between mPAP and compliance and distensibility. 1.1. Vascular changes demonstrated that myocardial efficiency reached a
maximum when the EA reached 1mmHgml1 (corresponding to an
mPAP of 20 mmHg), while maximum stroke work corresponded to
an EA of 2 mmHgml1 (corresponding to a MPAP of 30e40 mmHg)
[52]. This corresponds with observations that at approximately
40 mmHg the pulmonary artery reaches it maximum dimension
and minimal distensability [78,105,106]. Consistent with this, Ste-
vens et al. demonstrated that in patients with pulmonary hyper-
tension there was a curvilinear relationship between RV function
and PA distensibility with marked loss of pulmonary artery
distensibility without commensurate loss of ventricular function
until only minimal distensibility remained when a rapid decom-
pensation of the right ventricle occurred [76]. A similar curvilinear
relationship has also been observed in COPD between pulmonary
arterial stiffness and peripheral vascular resistance, with a rapid
rise in PVR only occurring after significant stiffening of the pul-
monary arteries has occurred [11]. Pulmonary stiffness is increased
early in pulmonary hypertension development, and is increased
even in those with exercise induced pulmonary hypertension, both
in mixed pulmonary arterial hypertension studies and in COPD
[3,11,78,106]. This combination of features suggest pulmonary Fig. 1. Calculation of EA:EES in the right ventricle and pulmonary artery. Ventricular
end systolic pressure-volume is linear and characterised by the slope Ees and is
generated by measuring pressure-volume loops under gradated preload and afterload
conditions. Reproduced from Wang et al., 2011 [100]. Ea ¼ Arterial elastance;
Ees ¼ Ventricular elastance. J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 1384 1384 Table 1
Measures of arterial stiffness e Calculations, definitions and alternate means of assessment. 1.1. Vascular changes Calculation
Definition
Method of assessment
Local
Pulsatility (%)
(maxA e minA)/minA X 100
Relative change in lumen area during the cardiac cycle
MRI Echocardiography IVUS
Compliance (mm2/mmHg)
(maxA e minA)/PP
Absolute change in lumen area for a given change in pressure
RHC plus MRI/Echo/IVUS
Distensibility (%/mmHg)
[(maxA e minA)/PP X minA] X 100
Relative change in lumen area for a given change in pressure
RHC plus MRI/Echo/IVUS
Elastic modulus (mmHg)
PP X minA/(maxA e minA)
Pressure change driving a relative increase in lumen area
RHC plus MRI/Echo/IVUS
Stiffness index (N/A)
Ln(sPAP/dPAP)/[(maxA e minA)/minA]
Slope of the function between distending arterial
pressure and arterial distension
RHC plus MRI/Echo/IVUS
Young's elastic modulus
[3(1 þ minA/WCSA)]/Distensibility
Wall thickness for a given distensibility
RHC plus IVUS
Regional
Capacitance (mm3/mmHg)
SV/PP
Change in volume associated with a given change in pressure
RHC
PWV (ms1)
Dd/Dt (TT technique) DQ/DA
(Flow-area technique)
Speed of transmission of pressure wave
RHC MRI
Systemic
Elastance (mmHgml1)
EA/EES ¼ (ESP/SV)/(ESP/ESV-V0)
Change in pressure for a given change in volume
RHC with flow-volume loops
A ¼ area; d ¼ distance; dPAP ¼ diastolic pulmonary artery pressure; EA ¼ Arterial elastance; Echo ¼ echocardiography; EES ¼ Ventricular elastance; IVUS ¼ intravascular
ultrasound; maxA ¼ maximum cross sectional area; mina ¼ minimum cross sectional area; MRI ¼ magnetic resonance imaging; PP ¼ pulse pressure; PWV ¼ pulse wave
velocity; Q ¼ Flow over a single heart beat; RHC ¼ right heart catheter; sPAP ¼ systolic pulmonary artery pressure; SV ¼ stroke volume; t ¼ time; WCSA ¼ Wall cross sectional
area. Table 1 Table 1
Measures of arterial stiffness e Calculations, definitions and alternate means of assessment. Calculation
Definition Relative change in lumen area during the cardiac cycle
Absolute change in lumen area for a given change in pressure
Relative change in lumen area for a given change in pressure
Pressure change driving a relative increase in lumen area
Slope of the function between distending arterial
pressure and arterial distension
Wall thickness for a given distensibility Change in volume associated with a given change in pressure
RHC
Speed of transmission of pressure wave
RHC MRI RHC with flow-volume loops 1.1.2. Measuring pulmonary artery stiffness No change in PA stiffness following
6 months of bosentan therapy. Rodes-Cabau et al. [72]
IPAH
20
RHC with IVUS
Reduced pulsatility in PH, with reduced pulsatility predictive of future mortality. Epoprostenol infusion increased pulsatility by 53%. Lau et al., 2014 [122]
IPAH
5
RHC with IVUS
High PWV in IPAH (mean PWV 10.6 ms1) with the reflected backward compression
wave carrying 31% of the energy of the forward compression wave
COPD ¼ chronic obstructive pulmonary disease; CoV ¼ Coefficient of variation; EIPH ¼ Exercise induced pulmonary hypertension; FEV1 ¼ Forced expiratory volume in 1 s;
HV ¼ Healthy volunteers; IPAH ¼ Idiopathic pulmonary arterial hypertension; NYHA ¼ New York heart association; PASP ¼ Pulmonary artery systolic pressure; PH ¼ Pul-
monary hypertension; PWV ¼ pulse wave velocity; QA ¼ flow by area; TT ¼ transition time; 6MWT ¼ 6 min walking test. a Drawn from the same population. COPD ¼ chronic obstructive pulmonary disease; CoV ¼ Coefficient of variation; EIPH ¼ Exercise induced pulmonary hypertension; FEV1 ¼ Forced expiratory volume in 1 s;
HV ¼ Healthy volunteers; IPAH ¼ Idiopathic pulmonary arterial hypertension; NYHA ¼ New York heart association; PASP ¼ Pulmonary artery systolic pressure; PH ¼ Pul-
monary hypertension; PWV ¼ pulse wave velocity; QA ¼ flow by area; TT ¼ transition time; 6MWT ¼ 6 min walking test.
a Drawn from the same population. Change in pressure for a given change in volume A ¼ area; d ¼ distance; dPAP ¼ diastolic pulmonary artery pressure; EA ¼ Arterial elastance; Echo ¼ echocardiography; EES ¼ Ventricular elastance; IVUS ¼ intravascular
ultrasound; maxA ¼ maximum cross sectional area; mina ¼ minimum cross sectional area; MRI ¼ magnetic resonance imaging; PP ¼ pulse pressure; PWV ¼ pulse wave
velocity; Q ¼ Flow over a single heart beat; RHC ¼ right heart catheter; sPAP ¼ systolic pulmonary artery pressure; SV ¼ stroke volume; t ¼ time; WCSA ¼ Wall cross sectional
area. A ¼ area; d ¼ distance; dPAP ¼ diastolic pulmonary artery pressure; EA ¼ Arterial elastance; Echo ¼ echocardiography; EES ¼ Ventricular elastance; IVUS ¼ intravascular
ultrasound; maxA ¼ maximum cross sectional area; mina ¼ minimum cross sectional area; MRI ¼ magnetic resonance imaging; PP ¼ pulse pressure; PWV ¼ pulse wave
velocity; Q ¼ Flow over a single heart beat; RHC ¼ right heart catheter; sPAP ¼ systolic pulmonary artery pressure; SV ¼ stroke volume; t ¼ time; WCSA ¼ Wall cross sectional
area. echocardiography, CT and MRI, none of these have yet proven ac-
curate enough to negate the need for invasive measurement with
right heart catheterisation [108e115]. arterial stiffness as a promising biomarker for detection of early
disease and as a potential therapeutic target before end stage
arterial remodelling occurs with dire consequences for the right
ventricle. 1.1.2.1. Right heart catheterisation. Right heart catheterisation can
be used to calculate several metrics of arterial stiffness, including
capacitance, PWV and elastance, although the latter two require
dedicated catheters and equipment not routinely available in the
catheter laboratory. In addition, it can be combined with techniques
for the visualisation of the pulmonary artery cross sectional area
such as MRI or intravascular ultrasound, to derive a wide range of
metrics of arterial stiffness and ventriculo-arterial coupling. Table 2
summarises the studies looking at invasive assessment of pulmo-
nary arterial stiffness. Despite its utility, RHC has numerous disin-
centives including its invasive nature, cost, and exposure to ionising
radiation. While the combination of MRI and right heart catheter-
isation allows catheter guidance without exposure to ionising ra-
diation and simultaneous acquisition of anatomical and pressure
measurements, it does not negate the invasive or costly nature of Table 2 J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 1385 the study [116,117]. Of the measures of arterial stiffness, only pul-
satility and pulse wave velocity can be derived entirely non-
invasively. the study [116,117]. Of the measures of arterial stiffness, only pul-
satility and pulse wave velocity can be derived entirely non-
invasively. PWV is an established marker of arterial stiffness in the systemic
circulation with aortic pulse wave velocity shown to be associated
with future cardiovascular events, and to improve risk stratification
[134]. Transcutaneous waveform analysis methods are the gold
standard for aortic PWV measurement including pressure [135],
distension [136], and flow waveform [137]. However a trans-
cutaneous approach is not possible for the pulmonary arteries. While PWV can be measured invasively on right heart catheter-
isation [118], magnetic resonance imaging is a possible alternative
for non-invasive assessment of the pulmonary arteries using flow
waveforms [138]. It is performed by the acquisition of ECG gated
phase contrast sequences which measure flow the in proximal
main pulmonary artery and in the mid right or left pulmonary ar-
teries. A distance can be calculated between the two imaging
planes (Fig. 2), and the pulse wave timed from when it leaves the
main pulmonary artery until it arrives in branch pulmonary arteries
thus allowing a speed between the two points to be generated
[138]. MRI derived pulse wave velocity has been previously vali-
dated against both invasive and non-invasive techniques in the
aorta [139e141]. Several studies have shown this to be a feasible
and reproducible technique in the pulmonary arteries in healthy
volunteers [138,142], with the observed pulse wave velocity of
1.96e2.3 ms1 similar to that obtained invasively [118,129]. Pulse
wave velocity is raised in pulmonary hypertension on both invasive
and MRI assessment (See Table 2) [121,143]. To date there have been
no studies assessing pulmonary PWV in COPD. 1.1.2.2. Pulsatility. Multiple studies have shown reduced pulsatility
in pulmonary hypertension,
[68,69,80,106,123e125]. A
single
echocardiography study has shown reduced pulmonary artery
pulsatility in COPD, which correlated with right ventricular func-
tional parameters [126]. A single abstract using cardiac MR has also
described decreasing pulsatility with increasing severity of COPD
[127] (Table 3). 1.1.3. Pulmonary arterial stiffness as a treatment target 1.1.3. Pulmonary arterial stiffness as a treatment target Traditional treatments of pulmonary hypertension such as
phosphodiesterase inhibitors and endothelin-1 receptor antago-
nists have yielded disappointing results despite their success in
treating pulmonary hypertension, primarily due to worsening
ventilatory-perfusion
mismatches
[144e148]. However
these
treatments have all focussed on altering the peripheral vasculature. Table 3
Summary of studies looking at non-invasive measurement of pulmonary arterial stiffness. Study
Population
No
Method
Finding
Pulsatility
Bogren et al. [159] b
Mixed PH
4
MRI
Pulsatility 23% in HV and 8% in PH
Paz et al. [160]
HV
9
MRI
No difference in pulsatility between the main pulmonary artery, right pulmonary artery and
left pulmonary arteries. Gan et al. [69]
Suspected PH
70
MRI
Right pulmonary artery pulsatility showed a curvilinear relationship with mortality. Pulsatility <16%
Jardim et al. [125] b
IPAH
19
MRI
<10% pulsatility predicts non-responders to acute vasodilator testing with 100% sens and 56% spec. Sanz et al. [124] a
Suspected PH
59
MRI
Pulsatility 41% in non-PH and 17.4% in PH. No difference in pulsatility between different causes of PH. Sanz et al. [78] a
Suspected PH
94
MRI
No difference in pulsatility between group without PH and EIPH. Pulsatility <40% predicted
PH with 93% sens. and 63% spec. Kang et al. [68]
Mixed PH
35
MRI
Pulsatility correlated with 6MWT (R2 ¼ 0.61, p < 0.001)<20% pulsatility predicted poor function
(6MWT <400m) with 82% sens. and 94% spec. Stevens et al. [76] a
Suspected PH
124
MRI
Pulsatility correlates with right ventricular function
Stevens et al. [161]
Suspected PH
43
MRI
Pulsatility correlated with exercise capacity while RHC pressures did not. Swift et al. [106]
Suspected PH
134
MRI
Pulsatility elevated even in very mild elevations in PVR. Pulsatility predicted mortality. Revel et al. [162] b
Suspected PH
45
CT
Pulsatility of 16.5% predicted PH with sens 86% and spec 96%. Pasierski et al. [80]
Suspected PH
19
Echo
Pulsatility reduced in pulmonary hypertension with a linear relationship between pulsatility and PASP
Mahapatra et al. [71]
Suspected PH
54
Echo
Capacitance was the strongest predictor of mortality including invasive pressure measurements
Ertan et al. [126]
COPD
54
Echo
Right pulmonary artery pulsatility reduced in COPD patients, with significant differences between
NYHA functional classes. Liu et al. 1.1.3. Pulmonary arterial stiffness as a treatment target [127]
COPD
135
MRI
Pulsatility falls with increasing severity of COPD, with a positive association with %predicted FEV1
and inversely correlated with %emphysema
PWV
Peng et al. [142]
HV
17
MRI
PWV 1.96 ± 0.27 with high intra-scan and inter-scan reproducibility (5.5% and 10.9% respectively)
Bradlow et al. [138]
HV
10
MRI
No significant difference in PWV using the left or right pulmonary arteries. CoV 12% for both intra
and inter-observer assessment. Ibrahim et al. [143]
Heterogeneous
33
MRI
PWV raised in pulmonary hypertension. Comparable measurements between TT and QA methods. COPD ¼ chronic obstructive pulmonary disease; CoV ¼ Coefficient of variation; EIPH ¼ Exercise induced pulmonary hypertension; FEV1 ¼ Forced expiratory volume in 1 s;
HV ¼ Healthy volunteers; IPAH ¼ Idiopathic pulmonary arterial hypertension; NYHA ¼ New York heart association; PASP ¼ Pulmonary artery systolic pressure; PH ¼ Pul-
monary hypertension; PWV ¼ pulse wave velocity; QA ¼ flow by area; TT ¼ transition time; 6MWT ¼ 6 min walking test. a Drawn from the same population. b Different calculation for % pulsatility so not directly comparable to other studies. Table 3
Summary of studies looking at non-invasive measurement of pulmonary arterial stiffness. a Drawn from the same population.
b COPD ¼ chronic obstructive pulmonary disease; CoV ¼ Coefficient of variation; EIPH ¼ Exercise induced pulmonary hypertension; FEV1 ¼ Forced expiratory volume in 1 s;
HV ¼ Healthy volunteers; IPAH ¼ Idiopathic pulmonary arterial hypertension; NYHA ¼ New York heart association; PASP ¼ Pulmonary artery systolic pressure; PH ¼ Pul-
monary hypertension; PWV ¼ pulse wave velocity; QA ¼ flow by area; TT ¼ transition time; 6MWT ¼ 6 min walking test.
a Drawn from the same population Table 2 However, as discussed above, the pulmonary stiff-
ness is dependant not only upon distending pressures, but also
upon the intrinsic elasticity of the pulmonary vessel wall, thus a fall
in pulmonary pulsatility does not differentiate a fall in pulsatility
due to a fall in stroke volume or in distending pressures from that of
a rise in arterial stiffness. 1.1.2.3. Pulse wave velocity. Pulse wave velocity (PWV) is the speed
at which the pressure wave, generated by ventricular contraction, is
transmitted through the arterial tree. This is directly related to
vessel wall elasticity, with increased stiffness leading to increased
PWV [128]. A reflected pressure wave is present which is generated
when the pressure wave is reflected from the termination of the
arteries, with alterations in the smallest arteries increasing the size
of the reflected wave while large artery stiffening increases the
PWV [118]. In health this reflected wave only represents a tiny
fraction of the total output energy, and arrives in diastole and re-
duces
preload
of
the
subsequent
ventricular
contraction
[92,118,129,130]. However in pulmonary hypertension this can
reach 30% of the forward wave energy. with an increased PWV
resulting in the earlier arrival of the reflected wave during systole
thereby increasing the afterload of the heart and thus cardiac
workload [77,119,122,131e133]. 1.1.2.3. Pulse wave velocity. Pulse wave velocity (PWV) is the speed
at which the pressure wave, generated by ventricular contraction, is
transmitted through the arterial tree. This is directly related to
vessel wall elasticity, with increased stiffness leading to increased
PWV [128]. A reflected pressure wave is present which is generated
when the pressure wave is reflected from the termination of the
arteries, with alterations in the smallest arteries increasing the size
of the reflected wave while large artery stiffening increases the
PWV [118]. In health this reflected wave only represents a tiny
fraction of the total output energy, and arrives in diastole and re-
duces
preload
of
the
subsequent
ventricular
contraction
[92,118,129,130]. However in pulmonary hypertension this can
reach 30% of the forward wave energy. with an increased PWV
resulting in the earlier arrival of the reflected wave during systole
thereby increasing the afterload of the heart and thus cardiac
workload [77,119,122,131e133]. [1] A. Hyduk, J.B. Croft, C. Ayala, K. Zheng, Z.-J. Zheng, G.A. Mensah, Pulmonary
hypertension
surveillanceeUnited
States,
1980-2002,
MMWR
Surveill.
Summ. 54 (2005) 1e28. No difference in pulsatility between the main pulmonary artery, right pulmonary artery and
left pulmonary arteries. Its use in COPD has not currently been assessed. Long term oxygen therapy improves survival in severe COPD
and reduces pulmonary arterial pressures and vascular resistance
while improving cardiac output [6,183]. It has also been shown to
reduce peripheral arterial stiffness in COPD, however its effects on
the pulmonary arterial stiffness has not been evaluated [184]. Statins have been proposed as a route to reduce systemic arterial
stiffness [149], although a meta-analysis showed mixed response
with two studies showing improvement, one showing no change
and another showing progression of stiffening [150]. Statins in
guinea pigs and mice with emphysema have been shown to reverse
arterial
remodelling
and
halt
the
formation
of
emphysema
[151,152]. Indirect evidence of the benefits from statins in COPD
related pulmonary hypertension come from observational studies
showing lower pulmonary arterial pressures in patients on statins
prior to right heart catheterisation [153]. Furthermore a study on
pulmonary arterial stiffness showed that other than mPAP, the only
other factors independently associated with pulmonary arterial
stiffness was LDL cholesterol and BMI [121]. Statins are also asso-
ciated with increased exercise capacity and reduced mortality in
COPD [154e156], however due to the observational nature of these
studies the underpinning mechanisms are still in debate [157,158]. There are also several new classes of pharmaceutical agents
currently being designed which specifically target the processes
causing arterial stiffening. Currently these are all being targeted at
systemic vascular remodelling and are in the pre-clinical stages,
however should they prove successful at preventing or reversing
systemic arterial remodelling they would be promising agents for
evaluation in pulmonary remodelling. The most promising of these
are TGF-B1 antagonists, and drugs targeting the advanced glycation
end products (AGEs) [185]. TGF-B1 is responsible for extracellular
matrix remodelling and vascular fibrosis in chronic inflammatory
conditions, whilst AGE form irreversible cross-links with collagen
and elastin, reducing the elasticity of the arterial wall [185]. No difference in pulsatility between the main pulmonary artery, right pulmonary artery and
left pulmonary arteries. p
y
Right pulmonary artery pulsatility showed a curvilinear relationship with mortality. Pulsatility <16%
<10% pulsatility predicts non-responders to acute vasodilator testing with 100% sens and 56% spec. Pulsatility 41% in non-PH and 17.4% in PH. No difference in pulsatility between different causes of PH. No difference in pulsatility between group without PH and EIPH. Pulsatility <40% predicted
PH with 93% sens. and 63% spec. Pulsatility falls with increasing severity of COPD, with a positive association with %predicted FEV1
and inversely correlated with %emphysema J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 1386 Fig. 2. Measurement of PWV using MRI: The location of the two phase contrast planes are delineated on the left image with the distance between them measured. The two separate
waveforms are traced on the right relative to the triggering R-wave used to start image acquisition. The time between the waves arriving at the two consecutive locations can then
be measured allowing calculation of the speed of the pulse wave. Fig. 2. Measurement of PWV using MRI: The location of the two phase contrast planes are delineated on the left image with the distance between them measured. The two separate
waveforms are traced on the right relative to the triggering R-wave used to start image acquisition. The time between the waves arriving at the two consecutive locations can then
be measured allowing calculation of the speed of the pulse wave. with pulmonary hypertension without right heart failure to eval-
uate its efficacy in humans in modulating pulmonary vascular
remodelling [182]. Its use in COPD has not currently been assessed. Long term oxygen therapy improves survival in severe COPD
and reduces pulmonary arterial pressures and vascular resistance
while improving cardiac output [6,183]. It has also been shown to
reduce peripheral arterial stiffness in COPD, however its effects on
the pulmonary arterial stiffness has not been evaluated [184]. Given the greater impact of pulmonary stiffness on remodelling,
drugs to target and ameliorate this process may be a novel route to
target. Due to the relatively recent identification of the importance
of pulmonary arterial stiffness in right ventricular remodelling,
there have been relatively few studies targeted at its treatment. Despite this there are several studies suggestive of promising av-
enues for further exploration. with pulmonary hypertension without right heart failure to eval-
uate its efficacy in humans in modulating pulmonary vascular
remodelling [182]. 2. Conclusions Xanthine oxidase inhibitors have been shown to improve
endothelial function and arterial stiffness in the systemic circula-
tion [163e167]. They have also been shown to reduce endothelial
dysfunction induced by smoking and obstructive sleep apnoea
[168e170] e a condition similar to COPD in its effects of intermit-
tent hypoxia
[171]. Additionally allopurinol
inhibits
hypoxia
induced pulmonary vasoconstriction, pulmonary hypertension,
endothelial dysfunction and vascular remodelling in hypoxic ani-
mal models of pulmonary hypertension [172e176]. It has been
shown to reduce airway reactive nitrogen species production in
COPD, however its effects on smoking induced pulmonary vascular
damage has not been assessed [177]. Pulmonary arterial stiffness holds a crucial role in right ven-
tricular dysfunction in pulmonary hypertension, and demonstrates
promise as a useful biomarker in the development of this disease. Early work shows similar promise in COPD, and further work is
warranted to explore this promising new area. Acknowledgements: Funding, disclosures JRWM is supported by the Wellcome Trust through the Scottish
Translational Medicine and Therapeutics Initiative (Grant no. WT
085664) in the form of a Clinical Research Fellowship. The funder
had no role in study design, the collection, analysis, and interpre-
tation of data; in the writing of the manuscript; nor in the decision
to submit the manuscript for publication. As with xanthine oxidase inhibitors, mineralocorticoid receptor
antagonists have been shown to improve systemic endothelial
function and arterial stiffness [178e180]. In 2 separate mouse
models of pulmonary hypertension, mineralocorticoid receptor
antagonists prevented or reversed adverse pulmonary vascular
remodelling with improved pulmonary vascular resistance, pul-
monary artery pressure, and remodelling of the right ventricle
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sion induced by hypoxia, J. Appl. Physiol. 90 (2001) 1299e1306. [155] C.M.M. Lawes, S. Thornley, R. Young, R. Hopkins, R. Marshall, W.C. Chan, et
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national cohort study, Prim. Care Respir. J. 21 (2012) 35e40. [173] R.P. Jankov, C. Kantores, J. Pan, J. Belik, Contribution of xanthine oxidase-
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interleukin-6 as predictors of outcomes in patients with chronic obstructive
pulmonary disease receiving pravastatin, Am. J. Cardiol. 101 (2008) 530e535. [174] J.M. Dopp, N.R. Philippi, N.J. Marcus, E.B. Olson, C.E. Bird, J.J.M. Moran, et al.,
Xanthine oxidase inhibition attenuates endothelial dysfunction caused by
chronic intermittent hypoxia in rats, Respiration 82 (2011) 458e467. p
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al., Chronic obstructive pulmonary disease (COPD) is associated with pul-
monary artery stiffness - the MESA COPD study, J. Cardiovasc Magn. Reson
15 (2013) O62. [151] J.L.Wright,S. Zhou,O.Preobrazhenska, C.Marshall,D.D. Sin,I.Laher,etal.,Statin
reverses smoke-induced pulmonary hypertension and prevents emphysema
but not airway remodeling, Am. J. Respir. Crit. Care Med. 183 (2011) 50e58. J.R. Weir-McCall et al. / Respiratory Medicine 109 (2015) 1381e1390 Mahoney, et al.,
Aldosterone Inactivates the Endothelin-B Receptor via a Cysteinyl Thiol
Redox Switch to Decrease Pulmonary Endothelial Nitric Oxide Levels and
Modulate
Pulmonary
Arterial
Hypertension,
Circulation
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purinol treatment reduces arterial wave reflection in stroke survivors, Car-
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study of the effect of spironolactone therapy on exercise capacity and
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Supplemental oxygen increases arterial stiffness in chronic obstructive
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in conduit artery endothelial dysfunction in cigarette smokers, Am. J. Cardiol. 93 (2004) 664e668.
|
https://openalex.org/W4285827834
|
https://zenodo.org/record/3270699/files/9780359774524_frontcover_content_backcover.pdf
|
Russian
| null |
Эпигенетика почек = Kidneys epigenetics
|
Zenodo (CERN European Organization for Nuclear Research)
| 2,019
|
cc-by
| 36,725
|
Dolomatov S.I., Zukow W. Эпигенетика почек = Kidneys epigenetics. RSW. Radom, 144 p. ISBN 9780359774524. DOI
http://dx.doi.org/10.5281/zenodo.3270699 PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606 Dolomatov S.I., Zukow W. Эпигенетика почек = Kidneys epigenetics. RSW. Radom, 144 p. ISBN 9780359774524. DOI
http://dx.doi.org/10.5281/zenodo.3270699 PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606 Эпигенетика почек
Kidneys epigenetics Эпигенетика почек
Kidneys epigenetics Radom, 2019 Radomska Szkoła Wyższa w Radomiu, Radom, Poland С.И. Доломатов, В.А. Жуков
S.I. Dolomatov, W. Zukow Radom, 2019 Dolomatov S.I., Zukow W. Эпигенетика почек = Kidneys epigenetics. RSW. Radom, 144 p. ISBN 9780359774524. DOI
http://dx.doi.org/10.5281/zenodo.3270699 PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606 АННОТАЦИЯ Ц
В книге представлены сведения о роли эпигенетических механизмов в системе контроля
функции почек в норме и при патологии. Результаты анализа роли эпигенетического контроля экспрессии
генов транспортных и регуляторных белков почки в норме указывают, во-первых, на высокую
пластичность процессов изменения экспрессии генов. Во-вторых, иллюстрируют их способность
адекватно реагировать на изменения параметров гомеостатических функций почек, что, в свою очередь,
позволяет рассматривать данные процессы в качестве еще одного звена управления деятельностью
органа наряду с нейро-эндокринными и внутриорганными уровнями гуморального контроля водно-
солевого баланса организма. Приведены факты, подчеркивающие вовлеченность гуморальных факторов
системного действия и внутрипочечных систем гуморального контроля в процессы эпигенетической
перестройки экспрессии генов ренальной паренхимы в норме и при патологии. Также анализируется роль
факторов среды в регуляции экспрессии генов. Radom, 2019 Dolomatov S.I., Zukow W. Эпигенетика почек = Kidneys epigenetics. RSW. Radom, 144 p. ISBN 9780359774524. DOI
http://dx.doi.org/10.5281/zenodo.3270699 PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606 Reviewers:
dr hab. R. Muszkieta, prof. nadzw. (Poland)
dr hab. M. Napierała, prof. nadzw (Poland) ANNOTATION The book provides information about the role of epigenetic mechanisms in the system of monitoring
renal function in normal and pathological conditions. The results of the analysis of the role of epigenetic control
of gene expression of kidney transport and regulatory proteins normally indicate, firstly, the high plasticity of
gene expression change processes. Secondly, they illustrate their ability to adequately respond to changes in the
parameters of homeostatic functions of the kidneys, which, in turn, makes it possible to consider these processes
as another element in the management of organ activity along with neuro-endocrine and intraorgan levels of the
humoral control of the body’s water-salt balance. The facts that emphasize the involvement of humoral factors of
systemic action and intrarenal systems of humoral control in the processes of epigenetic rearrangement of the
expression of renal parenchyma genes in normal and pathological conditions are presented. The role of
environmental factors in the regulation of gene expression is also analyzed. Ключевые слова: почки, эпигенетика. Key words: kidneys, epigenetics. This monograph is published with Open Access. Open Access This monograph is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribu
medium, provided the original author(s) and source are credited. ttribution — You must attribute the work in the manner specified by the author or licensor (but not in any way that suggests that they endorse you or your use of the work). Noncomm
use this work for commercial purposes. Share Alike — If you alter, transform, or build upon this work, you may distribute the resulting work only under the same or similar lic Zawartość jest objęta licencją Creative Commons Uznanie autorstwa Użycie niekomercyjne Na tych samych warunkach 4.0
ISBN 9780359774524
DOI http://dx.doi.org/10.5281/zenodo.3270699
PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606
Radomska Szkoła Wyższa w Radomiu, Polska
ul. 1905 roku 26/28
26-600 Radom
Tel: 048 383 66 05
E-mail: med@rsw.edu.pl
144 p. Number of characters: 250 000 (with abstracts). Number of images: 4 x 1000 characters (lump sum) = 4 000 characters. Total: Number of characters: 254 000 (with abstracts, summaries and graphics) = 6,35 sheet publications. 144 p. Number of characters: 250 000 (with abstracts). Number of images: 4 x 1000 characters (lump sum) = 4 000 characters. Total: Number of characters: 254 000 (with abstracts, summaries and graphics) = 6,35 sheet publications. 2 СОДЕРЖАНИЕ СОДЕРЖАНИЕ ВВЕДЕНИЕ
7
ОБЩИЕ СВЕДЕНИЯ ОБ ЭПИГЕНЕТИКИХ МЕХАНИЗМАХ
8
INTRODUCTION
11
GENERAL INFORMATION ON EPIGENETIC MECHANISMS
13
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «ВВЕДЕНИЕ»
16
ГЛАВА 1. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ В СИСТЕМЕ
КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В НОРМЕ
21
1.1. ПЛАСТИЧНОСТЬ СИСТЕМ ЭПИГЕНЕТИЧЕСКОЙ
МОДУЛЯЦИИ
ЭКСПРЕССИИ
ГЕНОВ
РЕНАЛЬНОЙ
ПАРЕНХИМЫ
23
1.2. ЭНДОКРИННЫЕ ФАКТОРЫ РЕГУЛЯЦИИ ВОДНО-
СОЛЕВОГО БАЛАНСА ОРГАНИЗМА В СИСТЕМЕ
ЭПИГЕНЕТИЧЕСКИХ
МЕХАНИЗМОВ
КОНТРОЛЯ
ГОМЕОСТАЗА
27
1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП)
27
1.2.2. НАТРИЙУРЕТИЧЕСКИЕ ПЕПТИДЫ
30
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «ЭПИГЕНЕТИЧЕСКИЕ
МЕХАНИЗМЫ В СИСТЕМЕ КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В
НОРМЕ»
33 ВВЕДЕНИЕ
7
ОБЩИЕ СВЕДЕНИЯ ОБ ЭПИГЕНЕТИКИХ МЕХАНИЗМАХ
8
INTRODUCTION
11
GENERAL INFORMATION ON EPIGENETIC MECHANISMS
13
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «ВВЕДЕНИЕ»
16
ГЛАВА 1. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ В СИСТЕМЕ
КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В НОРМЕ
21
1.1. ПЛАСТИЧНОСТЬ СИСТЕМ ЭПИГЕНЕТИЧЕСКОЙ
МОДУЛЯЦИИ
ЭКСПРЕССИИ
ГЕНОВ
РЕНАЛЬНОЙ
ПАРЕНХИМЫ
23
1.2. ЭНДОКРИННЫЕ ФАКТОРЫ РЕГУЛЯЦИИ ВОДНО-
СОЛЕВОГО БАЛАНСА ОРГАНИЗМА В СИСТЕМЕ
ЭПИГЕНЕТИЧЕСКИХ
МЕХАНИЗМОВ
КОНТРОЛЯ
ГОМЕОСТАЗА
27
1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП)
27
1.2.2. НАТРИЙУРЕТИЧЕСКИЕ ПЕПТИДЫ
30
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «ЭПИГЕНЕТИЧЕСКИЕ
МЕХАНИЗМЫ В СИСТЕМЕ КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В
НОРМЕ»
33 3 3 ГЛАВА
2. НЕКОТОРЫЕ
ФАКТОРЫ
АКТИВАЦИИ
ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ
47
2.1. ИЗМЕНЕНИЕ ТЕМПЕРАТУРНОГО РЕЖИМА, КАК
ФАКТОР ИНДУКЦИИ ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ
49
2.2. ГИПОКСИЯ
50
2.3. ГИПЕРГЛИКЕМИЯ
52
2.4. ТЯЖЕЛЫЕ МЕТАЛЛЫ
54
2.5. ЭНДОКРИНОПАТИИ
57
2.6. ЭПИГЕНЕТИЧЕСКИЕ ПРОЦЕССЫ, ИНДУЦИРОВАННЫЕ
ПАТОГЕННЫМИ МИКРООРГАНИЗМАМИ
59
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «НЕКОТОРЫЕ ФАКТОРЫ
АКТИВАЦИИ ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ»
61
ГЛАВА 3. ФАКТОРЫ ВНУТРИОРГАННОЙ ГУМОРАЛЬНОЙ
РЕГУЛЯЦИИ ДЕЯТЕЛЬНОСТИ ПОЧЕК. ИХ МЕСТО И РОЛЬ В
ЭПИГЕНЕТИЧЕСКИХ
МЕХАНИЗМАХ
НАРУШЕНИЯ
ФУНКЦИОНАЛЬНОГО
СОСТОЯНИЯ
РЕНАЛЬНОЙ
ПАРЕНХИМЫ
69
3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС)
70
3.2. МИНЕРАЛОКОРТИКОИДЫ. 76
3.3. ТРАНСФОРМИРУЮЩИЙ ФАКТОР РОСТА-бета1
79
3.4. МОЛЕКУЛА ОКСИДА АЗОТА (NO)
83
3.5. ПАТОФИЗИОЛОГИЧЕСКОЕ
ЗНАЧЕНИЕ
ЭПИГЕНЕТИЧЕСКОЙ
ТРАНСФОРМАЦИИ
СИСТЕМЫ
ВНУТРИОРГАННОЙ
ГУМОРАЛЬНОЙ
РЕГУЛЯЦИИ
ДЕЯТЕЛЬНОСТИ ПОЧЕК
87
СПИСОК
ЛИТЕРАТУРЫ
К
ГЛАВЕ
«ФАКТОРЫ
ВНУТРИОРГАННОЙ
ГУМОРАЛЬНОЙ
РЕГУЛЯЦИИ
ДЕЯТЕЛЬНОСТИ ПОЧЕК. ИХ МЕСТО И РОЛЬ В
ЭПИГЕНЕТИЧЕСКИХ
МЕХАНИЗМАХ
НАРУШЕНИЯ
ФУНКЦИОНАЛЬНОГО
СОСТОЯНИЯ
РЕНАЛЬНОЙ
ПАРЕНХИМЫ»
89
ГЛАВА 4. БЕЛКИ РЕНИН-АНГИОТЕНЗИНОВОЙ СИСТЕМЫ
ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ
105
4.1. ДИАГНОСТИЧЕСКАЯ ЦЕННОСТЬ АНАЛИЗА
ЭКСПРЕССИИ
БЕЛКОВ-КОМПОНЕНТОВ
РАС
В
107
4 4 ОНКОЛОГИИ
4.1.1. Рецепторы к А-II
107
4.1.2. Ангиотензин-I-превращающий фермент (АСЕ-1)
108
4.1.3. Ангиотензин-I-превращающий фермент-2 (АСЕ-2) и
ось ACE2/Ang-(1–7)/MAS1
110
4.1.4. Ангиотензиноген
111
4.1.5. (Про)Ренин
112
4.2. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ, КАК ВОЗМОЖНЫЕ
РЕГУЛЯТОРЫ ЭКСПРЕССИИ ПРОТЕИНОВ-КОМПОНЕНТОВ
РАС ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ
114
4.3. ОНКОЛОГИЧЕСКИЕ АСПЕКТЫ ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА
116
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «БЕЛКИ РЕНИН-
АНГИОТЕНЗИНОВОЙ СИСТЕМЫ ПРИ ОНКОЛОГИЧЕСКИХ
ЗАБОЛЕВАНИЯХ»
120 ОНКОЛОГИИ
4.1.1. Рецепторы к А-II
107
4.1.2. Ангиотензин-I-превращающий фермент (АСЕ-1)
108
4.1.3. Ангиотензин-I-превращающий фермент-2 (АСЕ-2) и
ось ACE2/Ang-(1–7)/MAS1
110
4.1.4. Ангиотензиноген
111
4.1.5. (Про)Ренин
112
4.2. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ, КАК ВОЗМОЖНЫЕ
РЕГУЛЯТОРЫ ЭКСПРЕССИИ ПРОТЕИНОВ-КОМПОНЕНТОВ
РАС ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ
114
4.3. ОНКОЛОГИЧЕСКИЕ АСПЕКТЫ ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА
116
СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «БЕЛКИ РЕНИН-
АНГИОТЕНЗИНОВОЙ СИСТЕМЫ ПРИ ОНКОЛОГИЧЕСКИХ
ЗАБОЛЕВАНИЯХ»
120 5 5 6 6 ВВЕДЕНИЕ Заболевание почек является одной из наиболее актуальных
глобальных проблем современной медицины. Данные медицинской
статистики показывают неуклонный рост числа нефрологических
пациентов, нуждающихся в диализе и трансплантации органа (Reddy M.A,
Natarajan R., 2015; Uwaezuoke S.N. et al., 2016; Zununi Vahed S. et al., 2016). Дополняет драматичность картины тот факт, что данная тенденция
демонстрирует актуальность и в отношении детей, включая
новорожденных (Woroniecki R. et al., 2011; Uwaezuoke S.N. et al., 2016; Lee-
Son K., Jetton J.G., 2016). Проблема носит серьезный характер, но даже
привлечение к ее преодолению подходов генетики, основанных на законах
моногенного наследования Г. Менделя, не в полной мере решает
поставленную задачу. Выводы специалистов по медицинской генетике,
изучающих наследование патологических признаков, способствующих
повышению рисков возникновения почечной недостаточности,
сопровождаются констатацией практической значимости выяснения
эпигенетических механизмов поражения ткани почек (Köttgen A. et al.,
2010; Lee-Son K., Jetton J.G., 2016; Ma R.C.W., 2016). Действительно,
обзоры результатов экспериментальных и клинических исследований,
указывают на необходимость более глубокого развития научного
направления, связанного с эпигенетическими механизмами патогенеза
почечной недостаточности (Thomas M.C., 2016; Witasp A. et al., 2017). д
(
,
;
p
,
)
Вместе с тем, особенность деятельности почки заключается в том,
что различные сегменты ее структурно-функциональной единицы –
нефрона, обладают существенными отличиями между собой, связанными с
их транспортными возможностями, набором гуморальных факторов
регуляции их активности и физико-химическими параметрами среды
микроокружения. Действительно, гомеостатические функции разных
отделов нефрона, координируются достаточно сложной системой
гуморальных
факторов,
определяющих
физиологические
и
патофизиологические механизмы реакции почек на изменения параметров
водных бассейнов организма и внешних неблагоприятных воздействий. К
числу таких гуморальных факторов следует отнести ренин-ангиотензин-
альдостероновую систему (РААС) (Lee-Son K., Jetton J.G, 2016), оксид
азота (Shirodkar A.V., Marsden P.A., 2011), трансформирующий фактор
роста-бета (Shi M. et al., 2011) и т.д. Перечисленные гуморальные факторы
контроля органогенеза и гомеостатических функций почек требуют более
глубокого изучения, поскольку могут также являться медиаторами
структурно-функциональных
нарушений
ренальной
паренхимы,
связанных, в том числе и с эпигенетическими преобразованиями процессов 7 транскрипции и трансляции в условиях острой и хронической почечной
недостаточности. С этих позиций, актуальность эпигенетического подхода обусловлена
тем, что, во-первых, сведения об изменениях экспрессии генов,
осуществляющих контроль над биосинтезом и экспрессией протеинов
ренальной паренхимы, а также системных и внутрипочечных гуморальных
факторов регуляции гомеостатической функции почек могут быть
использованы в совершенствовании методов современной лабораторной
диагностики заболеваний почек (Kobori H. et al., 2008). Во-вторых,
выяснение эпигенетических механизмов патогенеза почечной
недостаточности открывает перспективу в разработке принципиально
новых фармакологических препаратов, в том числе, контролирующих
синтез ренальной паренхимой различных физиологически активных
молекул (Marumo T. ВВЕДЕНИЕ et al., 2008; Reddy M.A, Natarajan R., 2015). В-третьих,
выяснение эпигенетических механизмов прогрессирования почечной
недостаточности позволяет по-новому оценить спектр применения и
нефропротекторных свойств уже известных и широко применяемых
препаратов, действие которых основано на коррекции активности
гуморальных систем контроля гомеостатических функций органа (Reddy
M.A. et al., 2014; Hayashi K. et al., 2015). Таким образом, мы видим свою задачу в том, чтобы попытаться
интегрировать современные достижения молекулярной биологии и
биохимии в уже существующую систему научных представлений о
физиологических механизмах регуляции деятельности почек и
патофизиологических
процессах
патогенеза
становления
и
прогрессирования почечной недостаточности. Следовательно, цель нашей работы сводится к выяснению
нескольких вопросов. Во-первых, какова роль эпигенетической
трансформации хроматина и микро РНК в физиологических механизмах
регуляции деятельности почек? Во-вторых, какова роль эпигенетических
процессов нарушения баланса системного и внутрипочечного метаболизма
гуморальных факторов регуляции функции почек в становлении и
прогрессировании почечной недостаточности? 8 8 ОБЩИЕ СВЕДЕНИЯ ОБ ЭПИГЕНЕТИКИХ МЕХАНИЗМАХ Эпигенетика — современное научное направление, изучающее
механизмы регуляции экспрессии генов. Эпигенетические механизмы не
оказывают влияния на первичную структуру нуклеиновых кислот
(Beckerman P. et al., 2014) и реализуются процессами метилирования и
деметилирования ДНК (van der Wijst M.G.P. et al., 2015), РНК (Saletore Y. et
al., 2013) и посттрансляционным процессингом гистоновых белков (Voon
H.P.J., Wong L.H., 2016; Jamal A. et al., 2012). По нашему мнению, важно
отметить, что процессы синтеза и постранскрипционного процессинга
микро РНК находятся под жестким контролем энзимов (Treiber T. et al.,
2019; Michlewski G., Cáceres J.F., 201(). Помимо этого, микро РНК могут
выполнять важную функцию в регуляции биосинтеза белка на уровне
транскрипции или трансляции (Petrillo F. et al., 2017; Thomas M.J. et al.,
2018). По данным авторов цитируемых обзоров, изучение метаболизма
микро РНК в различных биологических средах организма способствует
разработке принципиально новых методов диагностики и лечения
заболеваний почек. Помимо этого, эпигенетические механизмы принимают
участие в адаптивных реакциях организма и популяции в ответ на
изменения факторов среды через модуляцию экспрессии генов (Zama A.M.,
Uzumcu M., 2010). Одним из наиболее изученных
эпигенетических механизмов
регуляции экспрессии генов является процесс метилирования ДНК. Данный процесс заключается в присоединении метиловых групп (CH3) к
одному из четырех видов нуклеотидов ДНК путем образования между
ними ковалентной связи, однако, порядок последовательности нуклеотидов
в цепи ДНК при этом не меняется (Lister R. et al., 2009;Woroniecki R. et al.,
2011). За присоединение метиловых групп к нуклеотидам отвечает один из
четырех изоферментов ядерных ДНК-метилтрансфераз (DNMT) – а именно
DNMT1, DNMT2, DNMT3a или DNMT3b (Reddy M.A., Natarajan R., 2015;
Efimova O.A. et al., 2012). DNMT1 распознает полуметилированную ДНК
(каждую из её цепей) во время репликации (Bechtel W. et al., 2010). DNMT3a и DNMT3b обеспечивают метилирование ДНК de novo, т. е. повторно, в новых сайтах. Функция DNMT2 до сих пор является
предметом дискуссии (Jamal A. et al., 2012). ДНК-метилтрансферазы обладают способностью встраивать особые
«метки» в нуклеиновые кислоты, что приводит к изменению экспрессии
данных генов (van der Wijst M.G.P. et al., 2015). Высказывается
предположение о том, что эпигенетические «метки» в цепи ДНК
блокируют процесс транскрипции более двух третей объема ДНК
(Ponnaluri V.K.C. et al., 2016). 9 Таким
образом,
клетки
организма
применяют
ковалентную
модификацию ДНК с целью регуляции экспрессии генов по так
называемому принципу “on/off” (Quarta C. et al., 2016). Метилирование ДНК в большинстве случаев происходит на
цитозиновом азотистом основании (С), располагающимся в паре с
гуанином (G), на так называемых CpG-участках, или CpG-кластерах
(Dwivedi R.S. et al., 2011). ОБЩИЕ СВЕДЕНИЯ ОБ ЭПИГЕНЕТИКИХ МЕХАНИЗМАХ В человеческом организме около 70-80% CpG-
динуклеотидов находятся в метилированном состоянии (Ziller M.J. et al.,
2013). Участки цепи ДНК, где плотность CpG-кластеров особенно высока,
называют CpG-островками (Zhang D. et al., 2009). Однако процесс
метилирования
ДНК
осуществляется
на
участках
с
пониженной
плотностью CpG-динуклеотидов (Ziller M.J. et al., 2013). p
у
(
)
Наряду с процессом метилирования ДНК большое значение в
эпигенетической
регуляции
экспрессии
генов
имеет
процесс
деметилирования (van der Wijst M.G.P. et al., 2015; Yefimova O.A. et al.,
2012). Деметилирование ДНК – процесс высвобождения ДНК от
метильных групп, который осуществляется при помощи специальных
ферментов демиталаз (Auclair G., Weber M., 2012). Кроме того, к эпигенетическим механизмам относят также
модификации белков-гистонов в результате процессов ацетилирования,
деацетилирования, фосфорилирования, убиквитинирования и др. (Ganai
S.A. et al., 2016; Araki Y., Mimura T., 2017;). Процессы ацетилирования и
деацетилирования возможны благодаря ряду специфических ферментов —
гистонацетилазам (ацетилтрансферазам, HAT) и гистондеацетилазам
(HDAC) (Gong F. et al., 2016). Фосфорилирование же происходит за счет
ферментов киназ (фосфотрансфераз) (Araki Y., Mimura T., 2017; Nathan D. et al., 2012). Ковалентная модификация гистонов, как правило, ослабляет
связь гистонового кора нуклеосомы и ДНК, что способствует доступности
молекулы ДНК для процесса транскрипции (Rossetto D. et al., 2012). Однако, это правило не носит универсальный характер. Например,
ацетилирование лизина в положении 27 гистона H3 (H3K27ac) ведёт к
усилению экспрессии генов. Сочетание ацетилированного лизина 14
(H3K14ac) и фосфорилированного серина 10 гистона H3 (H3S10ph) так же
говорит о повышенной экспрессии генов (Chen K.W., Chen L., 2017). Наряду с этим другие модификации, а именно ацетилирование лизина
H2AK5, H2BK20, H3K14, H4K5 и др. и фосфорилирование треонина H3T3
и серина H3S28 и H4S1 аналогично приводят к активированию гена (Araki
Y., Mimura T., 2017; Wang Z. et al., 2008; Rossetto D. et al., 2012). Деацетилирование гистонов, напротив, сопровождается инактивацией гена,
поскольку влечет за собой конденсацию ДНК и невозможность протекания
транскрипции (Ganai S.A. et al., 2016). 10 Посттранскрипционная экспрессия генов регулируется
таким
эпигенетическим механизмом как интерференция РНК. Интерференция
РНК — это процесс подавления процесса биосинтеза белка (транскрипция,
трансляция) при помощи микро РНК (Nabzdyk C.S. et al., 2017). При этом
микро РНК может оказывать непосредственное влияние на состояние
трансляции (Dwivedi R.S. et al., 2011; Cui J. et al., 2017). Собственно
процесс подавления экспрессии генов именуется сайленсингом (Cui J. et
al., 2017). МикроРНК — это класс коротких одноцепочечных некодирующих
РНК длиной в 21-24 нуклеотида (Dwivedi R.S. et al., 2011; Zhang D. et al.,
2009). За
производство
микро РНК
ответственны
эндогенные
некодирующие участки ДНК. INTRODUCTION Kidney disease is one of the most pressing global problems of modern
medicine. Medical statistics show a steady increase in the number of
nephrological patients in need of dialysis and organ transplantation (Reddy M.A,
Natarajan R., 2015; Uwaezuoke S.N. et al., 2016; Zununi Vahed S. et al., 2016). The drama of the picture complements the fact that this trend also shows
relevance for children, including newborns (Woroniecki R. et al., 2011;
Uwaezuoke S.N. et al., 2016; Lee-Son K., Jetton J.G., 2016). The problem is
serious, but even the attraction of genetics approaches based on the monogenic
inheritance laws of G. Mendel to its overcoming does not fully solve the
problem posed. The findings of specialists in medical genetics who study the
inheritance of pathological signs that increase the risk of renal failure are
accompanied by a statement of the practical importance of finding out the
epigenetic mechanisms of kidney tissue damage (Köttgen A. et al., 2010; Lee-
Son K., Jetton JG, 2016; Ma RCW, 2016). Indeed, reviews of the results of
experimental and clinical studies indicate the need for a deeper development of
the scientific direction related to the epigenetic mechanisms of the pathogenesis
of renal failure (Thomas M.C., 2016; Witasp A. et al., 2017). (
p
)
At the same time, the peculiarity of the kidney activity is that the various
segments of its structural and functional unit, the nephron, have significant
differences among themselves related to their transport capabilities, a set of
humoral factors regulating their activity and the physicochemical parameters of
the microenvironment. Indeed, the homeostatic functions of different parts of the
nephron are coordinated by a rather complex system of humoral factors that
determine the physiological and pathophysiological mechanisms of the reaction
of the kidneys to changes in the parameters of the body’s water basins and
external adverse effects. These humoral factors include the renin-angiotensin-
aldosterone system (RAAS) (Lee-Son K., Jetton JG, 2016), nitric oxide
(Shirodkar AV, Marsden PA, 2011), transforming growth factor-beta (Shi M. et
al., 2011), etc. The listed humoral factors controlling organogenesis and
homeostatic functions of the kidneys require further study, since they can also
mediate structural and functional disorders of the renal parenchyma, including
epigenetic transformations of transcription and translation in conditions of acute
and chronic renal failure. ОБЩИЕ СВЕДЕНИЯ ОБ ЭПИГЕНЕТИКИХ МЕХАНИЗМАХ Помимо трансляции микро РНК обладает
способностью подавлять экспрессию генов на стадии транскрипции
(Dwivedi R.S. et al., 2011). Все регуляторные эффекты микро РНК встроены в определенную
систему эпигенетического контроля функций данной популяции клеток
(Dwivedi R.S. et al., 2011). Анализируя современные данные литературы, можно предположить,
что эпигенетические механизмы являются важным элементом адаптации на
популяционном и организменном уровне, осуществляющим координацию
экспрессии генов адекватно изменениям факторов внешней среды. В
нашем сознании понятие «эпигенетические механизмы» чаще
ассоциируются с феноменом фетального программирования. Между тем,
их деятельность активно протекает и в постнатальный период онтогенеза. Сложно сказать в чем именно может иметь место несовершенство этой
формы адаптивного ответа, однако, ее реализация зачастую сопряжена с
активацией патогенетических механизмов различных систем органов. 11 GENERAL INFORMATION ON EPIGENETIC MECHANISMS Epigenetics is a modern scientific direction that studies the mechanisms of
regulation of gene expression. Epigenetic mechanisms do not affect the primary
structure of nucleic acids (Beckerman P. et al., 2014) and are implemented by
DNA methylation and demethylation processes (van der Wijst MGP et al.,
2015), RNA (Saletore Y. et al., 2013) and post-translational processing of
histone proteins (Voon HPJ, Wong LH, 2016; Jamal A. et al., 2012). In our
opinion, it is important to note that the processes of synthesis and post-
transcriptional processing of micro RNA are tightly controlled by enzymes
(Treiber T. et al., 2019; Michlewski G., Cáceres JF, 201 (). In addition, micro
RNA can perform an important function in the regulation of protein biosynthesis
at the level of transcription or translation (Petrillo F. et al., 2017; Thomas MJ et
al., 2018). According to the authors of the cited reviews, the study of the
metabolism of micro RNA in various biological media of the body contributes to
the development of fundamentally new diagnostic methods and kidney disease
treatment. of this, epigenetic mechanisms are involved in adaptive responses of
the organism and population in response to changes in environmental factors
through modulation of gene expression (Zama A.M., Uzumcu M., 2010). g
g
p
(
)
One of the most studied epigenetic mechanisms of regulation of gene
expression is the process of DNA methylation. This process involves the
addition of methyl groups (CH3) to one of four types of DNA nucleotides by
forming a covalent bond between them, however, the order of the sequence of
nucleotides in the DNA chain does not change (Lister R. et al., 2009;
Woroniecki R. et al., 2011). One of the four nuclear DNA methyltransferase
isoenzymes (DNMT), namely DNMT1, DNMT2, DNMT3a or DNMT3b, is
responsible for the addition of methyl groups to nucleotides (Reddy M.A.,
Natarajan R., 2015; Efimova O.A. et al., 2012). DNMT1 recognizes semi-
methylated DNA (each of its chains) during replication (Bechtel W. et al., 2010). DNMT3a and DNMT3b provide de novo methylation of DNA, i.e., repeatedly,
in new sites. The DNMT2 function is still under discussion (Jamal A. et al.,
2012). DNA methyltransferases have the ability to embed specific “tags” in
nucleic acids, which leads to changes in the expression of these genes (van der
Wijst M.G.P. et al., 2015). INTRODUCTION From this point of view, the relevance of the epigenetic approach is due to
the fact that, firstly, information on changes in the expression of genes that
control biosynthesis and expression of renal parenchyma proteins, as well as
systemic and intrarenal humoral factors regulating the homeostatic function of
the kidneys can be used to improve modern methods. laboratory diagnosis of
kidney disease (Kobori H. et al., 2008). Secondly, the clarification of the 12 epigenetic mechanisms of renal failure pathogenesis opens up the prospect of
developing fundamentally new pharmacological drugs, including those
controlling the synthesis of renal parenchyma of various physiologically active
molecules (Marumo T. et al., 2008; Reddy MA, Natarajan R., 2015) . Third,
clarifying the epigenetic mechanisms of progression of renal failure allows us to
re-evaluate the range of applications and the nephroprotective properties of
already known and widely used drugs, which are based on correcting the activity
of the humoral control systems of homeostatic organ functions (Reddy MA et
al., 2014; Hayashi K . et al., 2015). Thus, we see our task in trying to integrate modern advances in molecular
biology and biochemistry into the already existing system of scientific ideas
about the physiological mechanisms of regulating kidney activity and the
pathophysiological processes of the pathogenesis of renal failure and
progression. Consequently, the goal of our work comes down to clarifying a few
questions. First, what is the role of epigenetic transformation of chromatin and
micro RNA in the physiological mechanisms of regulation of kidney activity? Secondly, what is the role of the epigenetic processes of imbalance of the
systemic and intrarenal metabolism of humoral factors regulating the function of
the kidneys in the formation and progression of renal failure? 13 13 GENERAL INFORMATION ON EPIGENETIC MECHANISMS It is suggested that epigenetic “tags” in the DNA
chain block the transcription process for more than two thirds of the DNA
volume (Ponnaluri V.K.C. et al., 2016). Thus, the cells of an organism use covalent modification of DNA in order
to regulate gene expression according to the so-called “on / off” principle
(Quarta C. et al., 2016). 14 DNA methylation in most cases occurs on the cytosine nitrogen base (C),
which is paired with guanine (G), on the so-called CpG sites, or CpG clusters
(Dwivedi R.S. et al., 2011). In the human body, about 70-80% of CpG
dinucleotides are in the methylated state (Ziller M.J. et al., 2013). The portions
of the DNA chain where the density of CpG clusters is particularly high are
called CpG islands (Zhang D. et al., 2009). However, the process of DNA
methylation is carried out in areas with low density of CpG dinucleotides (Ziller
M.J. et al., 2013). ,
)
Along with the process of DNA methylation, the process of demethylation
is of great importance in the epigenetic regulation of gene expression (van der
Wijst M.G.P. et al., 2015; Yefimova O.A. et al., 2012). DNA demethylation is
the process of DNA release from methyl groups, which is carried out with the
help of special enzymes of demetallas (Auclair G., Weber M., 2012). In addition, modifications of histone proteins as a result of acetylation,
deacetylation, phosphorylation, ubiquitination, etc. are also considered
epigenetic mechanisms (Ganai S.A. et al., 2016; Araki Y., Mimura T., 2017;). Acetylation and deacetylation processes are possible due to a number of specific
enzymes - histone acetylases (acetyltransferases, HAT) and histone deacetylases
(HDAC) (Gong F. et al., 2016). Phosphorylation occurs at the expense of
enzymes of kinases (phosphotransferases) (Araki Y., Mimura T., 2017; Nathan
D. et al., 2012). Covalent modification of histones, as a rule, weakens the
connection between the histone core of the nucleosome and DNA, which
contributes to the availability of the DNA molecule for the transcription process
(Rossetto D. et al., 2012). Along with the process of DNA methylation, the process of demethylation
is of great importance in the epigenetic regulation of gene expression (van der
Wijst M.G.P. et al., 2015; Yefimova O.A. et al., 2012). DNA demethylation is
the process of DNA release from methyl groups, which is carried out with the
help of special enzymes of demetallas (Auclair G., Weber M., 2012). GENERAL INFORMATION ON EPIGENETIC MECHANISMS p
p
y
(
)
In addition, modifications of histone proteins as a result of acetylation,
deacetylation, phosphorylation, ubiquitination, etc. are also considered
epigenetic mechanisms (Ganai S.A. et al., 2016; Araki Y., Mimura T., 2017;). Acetylation and deacetylation processes are possible due to a number of specific
enzymes - histone acetylases (acetyltransferases, HAT) and histone deacetylases
(HDAC) (Gong F. et al., 2016). Phosphorylation occurs at the expense of
enzymes of kinases (phosphotransferases) (Araki Y., Mimura T., 2017; Nathan
D. et al., 2012). Covalent modification of histones, as a rule, weakens the
connection between the histone core of the nucleosome and DNA, which
contributes to the availability of the DNA molecule for the transcription process
(Rossetto D. et al., 2012). (
)
However, this rule is not universal. For example, acetylation of lysine at
position 27 of histone H3 (H3K27ac) leads to increased gene expression. The
combination of acetylated lysine 14 (H3K14ac) and phosphorylated serine 10
histone H3 (H3S10ph) also indicates increased gene expression (Chen K.W.,
Chen L., 2017). Along with this, other modifications, namely acetylation of
lysine H2AK5, H2BK20, H3K14, H4K5, etc., and threonine H3T3 and serine
H3S28 and H4S1 phosphorylation similarly lead to gene activation (Araki Y.,
Mimura T., 2017; Wang Z. et al ., 2008; Rossetto D. et al., 2012). On the
contrary, histone deacetylation is accompanied by gene inactivation, as it entails
DNA condensation and the impossibility of transcription (Ganai S.A. et al.,
2016). Post-transcriptional gene expression is regulated by such an epigenetic
mechanism as RNA interference. RNA interference is the process of suppressing
the process of protein biosynthesis (transcription, translation) using micro RNA
(Nabzdyk C.S. et al., 2017). At the same time, micro RNA can have a direct
impact on the state of translation (Dwivedi R.S. et al., 2011; Cui J. et al., 2017). 15 The actual process of suppressing gene expression is called silencing (Cui J. et
al., 2017). MicroRNA is a class of short single-stranded non-coding RNAs of 21-24
nucleotides in length (Dwivedi R.S. et al., 2011; Zhang D. et al., 2009). Endogenous noncoding regions of DNA are responsible for the production of
micro RNA. In addition to translation, micro RNA has the ability to suppress
gene expression at the transcription stage (Dwivedi R.S. et al., 2011). All regulatory effects of micro RNA are embedded in a specific system of
epigenetic control of the functions of a given cell population (Dwivedi R.S. GENERAL INFORMATION ON EPIGENETIC MECHANISMS et
al., 2011). Analyzing modern literature data, it can be assumed that epigenetic
mechanisms are an important element of adaptation at the population and
organism level, coordinating the expression of genes adequately to changes in
environmental factors. In our consciousness, the concept of “epigenetic
mechanisms” is more often associated with the phenomenon of fetal
programming. Meanwhile, their activity is actively proceeding in the postnatal
period of ontogenesis. It is difficult to say exactly what the imperfection of this
form of adaptive response can take place, however, its implementation often
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481. doi: 10.1038/nature12433 20 ГЛАВА 1. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ
В СИСТЕМЕ КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В НОРМЕ Эпигенетические системы управления экспрессии генов выполняют
принципиально важную функцию на разных этапах онтогенеза. Например,
у плода они координируют нормальное течение нефрогенеза. В зрелом
возрасте эпигенетические механизмы тесно вовлечены в систему контроля
гомеомтатических функций органа. Процессы снижения функции почек у
пожилых людей также тесно связаны с эпигенетическими механизмами. пожилых людей также тесно связаны с эпигенетическими механизмами. Оценивая
роль
эпигенетических
механизмов
в
процессах
органогенеза почки, необходимо отметить роль метилирования гистонов в
цитодифференцировке эмбриональных клеток (Adli M. et al., 2015). Наряду
с
этим,
подчеркивается
роль
баланса
активности
гистоновых
ацетилтрансфераз и деацетилаз в регуляции экспрессии генов на ранних
стадиях
органогенеза
почки
(Hilliard S.A, El-Dahr S.S., 2016). Высказывается мнение о том, что некоторые деацетилазы гистоновых
белков (HDAC1 and HDAC2) могут быть критически важны для процессов
развития канальцевого и сосудистого компонентов нефрона на ранних
стадиях онтогенеза почки (Liu H. et al., 2018). Наряду с этим, в литературе
имеются даные о том, что синтез некодидурющих РНК и реакции
ацетилирования гистонов выполняют важную роль в формировании юкста-
гломерулярного аппарата (ЮГА) в процессе нефрогенеза (Martini A.G.,
Danser A.H.J., 2017). С другой стороны, внутриорганная продукция
компонентов ренин-ангиотензиновой системы (РАС) на ранних этапах
онтогенеза
также критически важна
для
координации
гисто-
и
органогенеза. Установлено, что избыточное потребление хлорида натрия во
время беременности может нарушать эти процессы через изменения
активности внутриорганной экспрессии компонентов РАС и продукции
оксида азота в тканях плода (Stocher D.P. et al., 2018). Анализ роли
метилтрансфераз и деметилаз, а также гистон-ацетилтрансфераз и гистон-
деацетилаз в процессах нефрогенеза позволил выявить определенные
закономерности динамики активности данных групп ферментов по мере
формирования нефрона (Hilliard S.A., El-Dah S.S., 2016). Авторы
цитируемого обзора сопоставляют процессы нефрогенеза с топологией и
динамикой во времени активности систем ковалентной модификации
хроматина: остатков лизина в составе гистонов (H3K), остатков аргинина в
составе гистонов (H3R) и молекулы ДНК. Наряду с процессами
ковалентной модификации хроматина, механизмы транскрипции и
метаболизма некодирующих РНК также могут иметь принципиально
важное
значение
для
нормального
течения
морфогенеза
почки 21 млекопитающих (Ho J., Kreidberg J.A., 2012). В настоящее время роль
микро РНК в процессах органогенеза почки изучено достаточно подробно. В литературе имеются сведения о том, что некоторые семейства микро
РНК критически важны для морфогенеза сосудисто-клубочкового и
канальцевого отделов почки (Trionfini P., Benigni A., 2017). Возможно,
эпигенетические механизмы органогенеза почки находятся под контролем
гормонов системного действия материнского организма. В частности,
показано, что такой способностью может обладать мелатонин (Tain Y.-L. et
al., 2017). ГЛАВА 1. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ
В СИСТЕМЕ КОНТРОЛЯ ФУНКЦИИ ПОЧЕК В НОРМЕ По мнению авторов, мелатонин обладает способностью
контролировать не только формирование архитектуры нефрона, но и
регулировать уровень активности внутрипочечной системы NO-синтаз и
ренин-ангиотензиновой
системы
плода
через
интенсивность
метилирования ДНК и ацетилирования белков гистонов. Кроме того,
показано, что инсулин также обладает выраженным влиянием на состояние
эпигенетических механизмов в тканях почки человека (Lay A.C., Coward
R.J.M., 2018). В литературе приводятся данные о том, что гипометилирование
хроматина на уровне нейро-эндокринного звена контроля деятельности
почки — одна из причин увядания гомеостатической функции органа в
преклонном
возрасте
(Murgatroyd C. et al., 2010). Дальнейшие
исследования позволили установить важное значение роли метилирования
хроматина в возрастных изменениях системы контроля водно-солевого
баланса у млекопитающих (Greenwood M.P. et al., 2018). В литературе
уделяется внимание роли микро РНК и ковалентной модификации
хроматина в процессах возрастных нарушений функции почек человека
(Shiels P.G. et al., 2017). На основе анализа роли деацетилаз гистонов SIRT1
и SIRT3 в регуляции обменных процессов почки, делается вывод о том, что
данная группа ферментов обладает выраженным нефропротекторным
свойством, обеспечивая сдерживание процессов старения тканей органа
(Morigi M. et al., 2018). 22 1.1. ПЛАСТИЧНОСТЬ СИСТЕМ ЭПИГЕНЕТИЧЕСКОЙ
МОДУЛЯЦИИ ЭКСПРЕССИИ ГЕНОВ РЕНАЛЬНОЙ ПАРЕНХИМЫ Как уже отмечалось выше, в зрелом возрасте эпигенетические
механизмы сохраняют за собой важное место в регуляции функции почек,
в частности, адаптивных реакций ренальной паренхимы. Необходимо
подчеркнуть, что эпигенетические механизмы контроля биосинтеза белка
сохраняют
высокий
уровень
пластичности
в
зрелом
возрасте. Иллюстрируя высокие показатели пластичности обсуждаемых процессов,
можно упомянуть роль метилирования ДНК в формировании суточного
ритма поведенческой активности млекопитающих (Azzi A. et al., 2014). Следовательно, есть основания полагать, что молекулярные механизмы
регуляции экспрессии генов могут непосредственно координировать
адаптивные реакции ренальной паренхимы. Возможно, эпигенетические
механизмы, наряду с нейро-гуморальными системами контроля водно-
солевого обмена, принимают участие в регуляции гомеостатических
функций почек. В ряде публикаций указывается, что стимулом для молекулярных
механизмов управления экспрессии генов, как правило, является динамика
параметров констант водно-солевого баланса организма. Результаты более
ранних исследований показали, что метилтрансфераза гистонов Dot1a
непосредственно определяет альдостерон-зависимую транскрипцию гена
EnaC-альфа в дистальных отделах нефрона (Zhang D. et al., 2009). Согласно
данным литературы, состояние посттранскрипционного процессинга
предшественника
микроРНК
в
проксимальных
нефроцитах
может
выполнять ключевую роль в адаптации канальцевого эпителия к ишемии,
возможно, участвуя в патогенезе реперфузионного поражения S3-сегмента
(Wei Q. et al., 2010). Подчеркивается, что интенсивность репаративных
реакций ренальной паренхимы может контролироваться некодирующими
РНК и состоянием метилирования Н2А и Н3 гистонов (Chou Y.-H. et al.,
2017). В литературе имеются сведения о том, что у некоторых видов
млекопитающих в адаптивных реакциях почки на острые изменения
системных параметров водно-солевого обмена могут принимать участие
механизмы регуляции экспрессии генов (MacManes M.D., 2017). Наряду с
этим, в проксимальном сегменте нефрона объектом регуляторного влияния
эпигенетических механизмов являются гены субъединиц натрий\калиевой
АТФазы базолатеральной мембраны эпителия (Taub M., 2018). По мнению
автора
цитируемого
обзора,
сигналом
для
активации/инктивации
транскрипции указанных генов может служить концентрация натрия в
люминальной жидкости, а непосредственная реализация поступающих
сигналов определяется интенсивностью ацетилирования гистонов. Наряду 23 с этим, изменения внутриклеточной концентрации натрия в эпителии
проксимального сегмента нефрона и тонкой восходящей петли Генле также
может оказывать прямое влияние на состояние транскрипции генов
транспортных белков, экспрессируемых данной популяцией нефроцитов
(Gildea J.J. et al., 2018). Приводятся данные о том, что содержание натрия в
рационе питания оказывает влияние на экспрессию генов белков-
транспортеров натрия (ENaC и Na-Cl-котранспортер) в дистальном отделе
нефрона (Ivy J.R. et al., 2018). С другой стороны, показано, что
гипонатриевая диета стимулирует гипометилирование гена альдостерон-
синтазы через активацию РАС (Takeda Y. et al., 2018). Привлекает внимание
тот факт, что ядерные деацетилазы ренальной паренхимы (SIRT1,3,6,7)
обладают способностью регулировать экспрессию ряда белков в тканях
почки, имеющих фундаментальное значение для гомеостатических
функций органа (Morigi M. et al., 2018). 1.1. ПЛАСТИЧНОСТЬ СИСТЕМ ЭПИГЕНЕТИЧЕСКОЙ
МОДУЛЯЦИИ ЭКСПРЕССИИ ГЕНОВ РЕНАЛЬНОЙ ПАРЕНХИМЫ В частности, авторы обзора
сообщают, что SIRT1 непосредственно регулирует экспрессию альфа-
субъединицы эпителиального натриевого канала, эндотелиальной NO-
синтазы
и рецептора к ангиотензину-2 (AT1R)
в
подоцитах
и
гладкомышечных волокнах кровеносных сосудов почки. По данным
авторов SIRT3 участвует в регуляции обменных процессов в митохондриях,
обладает противовоспалительным и противосклерозирующим действием. Белок SIRT6 также необходим для сдерживания просклерозирующих
факторов. Следует отметить, что, наряду с ковалентной модификацией
хроматина, важная роль в
эпигенетическом
контроле ренального
транспорта веществ отводится некодирующим РНК (Hua J.X. et al., 2012). Показана важная роль микро РНК в регуляции транспорта натрия в
эпителии нефрона (Mladinov D. et al., 2013). Наряду с ионорегулирующей
функцией почек, установлено, что некодирующие РНК могут принимать
участие в управлении осморегулирующей функцией почек млекопитающих
(Huang W. et al., 2011; Luo Y. et al., 2014). Авторы цитируемых публикаций
указывают на роль микро РНК в регуляции экспрессии транспортных
белков
медуллярных
сегментов
нефрона
в
ответ
на
острый
гиперосмотический стимул. Следует отметить, что в норме экспрессия
некоторых типов микро РНК в корковом и мозговом слое почки имеет
четкие отличия (Chandrasekaran K. et al., 2012; Ichii O., Horino T., 2018). Приводится информация о непосредственном влиянии гиперосмотического
стимула на экспрессию строго определенных типов микро РНК во
внутренней медулле почки (Chandrasekaran K. et al., 2012). Вместе с тем,
авторы обращают внимание на тот факт, что состояние метаболизма микро
РНК в ренальной паренхиме может регулироваться гуморальными
факторами нейро-эндокринного звена контроля гомеостатических функций 24 почек. При этом микро РНК осуществляют контроль транспорта ионов не
только в почке, но и системные параметры ионного гомеостаза (Hua J.X. et
al., 2012). В ряде публикаций подчеркивается тезис о том, что микро РНК
могут осуществлять постоянную тонкую регуляцию обменных процессов в
ренальной паренхиме. Например, имеются сообщения о роли микро РНК в
регуляции обменных процессов в подоцитах, в зависимости от возможных
изменений
величины
гидростатического
давления
в
клубочке
и
химического состава ультрафильтрата (Trionfini P., Benigni A., 2017). Одним из наиболее перспективных направлений исследований роли
микро РНК в регуляции деятельности почки является анализ взаимосвязи
внутриорганного
метаболизма
микро РНК и их содержания в
биологических средах организма (Thomas M.J. et al., 2018). С точки зрения
практической медицины, ценность таких исследований обусловлена
необходимостью внедрения новых методов диагностики и терапии
заболеваний почек (Trionfini P., Benigni A., 2017; Thomas M.J. et al., 2018). заболеваний почек (Trionfini P., Benigni A., 2017; Thomas M.J. et al., 2018). Вместе с тем, значительное внимание уделяется роли гуморальных
систем контроля гомеостатических функций почек в регуляции эспрессии
генов в ренальной паренхиме (Hirohama D. et al., 2018; Lu C.C. 1.1. ПЛАСТИЧНОСТЬ СИСТЕМ ЭПИГЕНЕТИЧЕСКОЙ
МОДУЛЯЦИИ ЭКСПРЕССИИ ГЕНОВ РЕНАЛЬНОЙ ПАРЕНХИМЫ et al., 2018). Приводятся данные о молекулярных механизмах регуляции локальной
экспрессии белков-компонентов РАС (Martini A.G. et al., 2017; Lu C.C. et al.,
2018). Ранее была показана роль микро РНК в регуляции экспрессии генов
ренина (Sequeira-Lopez M.L.S. et al., 2010). В настоящее время установлена
роль метилирования ДНК, ацетилирования и метилирования гистонов
канальцевого эпителия в управлении экспрессией гена ангиотензиногена
(Marumo T. et al., 2015). Показано участие метилирования ДНК,
ковалентной модификации гистонов и метаболизма микро РНК в
экспрессии генов ренина в почке (Martini A.G., Danser A.H.J., 2017). С
другой стороны, выявлено значение ренин-ангиотензин-альдостероновой
системы в регуляции экспрессии генов транспортных белков канальцевого
отдела нефрона в ответ на изменение физиологических констант водно-
солевого баланса (Hirohama D. et al., 2018). Установлено, что ковалентная
модификация
гистонов
(метилирование
и
ацетилирование)
может
принимать
участие
в
контроле
экспрессии
гена
атриального
натрийуретического пептида (Hohl M. et al., 2013). Сообщается, что
эпигенетический
контроль
экспрессии
гена
атриального
натрийуретического
пептида
способствует
адаптивным
изменениям
продукции гормона (Sergeeva I.A. et al., 2016). При этом, атриальный
натрийуретический пептид также рассматривается в качестве индуктора
эпигенетических механизмов, реализуемых через специфические микро
РНК (Li Y. et al., 2016). Не меньший интерес привлекают сведения о
влиянии острого осмотического стимула на эпигенетические системы 25 контроля синтеза аргинин-вазопрессина - АВП (Hayashi M. et al., 2006;
Greenwood M.P. et al., 2016). Необходимо отметить, что половые
стероидные гормоны также могут оказывать влияние на экспрессию гена
АВП при участии эпигенетических механизмов (Augera C.J. et al., 2011). Поскольку канальцевые эффекты АВП реализуются при участии
специфических поробразующих белков — аквапоринов, в частности, при
участии аквапорина-2 (AQP2), привлекают интерес сведения о значении
эпигенетического контроля данного белка (Park E.-J., Kwon T.H.; 2015; Jung
H.J., Kwon T.-H.; 2016). 26 26 1.2. ЭНДОКРИННЫЕ ФАКТОРЫ РЕГУЛЯЦИИ ВОДНО-СОЛЕВОГО
БАЛАНСА ОРГАНИЗМА В СИСТЕМЕ ЭПИГЕНЕТИЧЕСКИХ
МЕХАНИЗМОВ КОНТРОЛЯ ГОМЕОСТАЗА Предполагая определенную роль эпигенетических механизмов в
регуляции гомеостатических функций почек и адаптивных изменениях
органа, по нашему мнению, необходимо проанализировать, во-первых,
информацию о роли эпигенетических механизмов в модуляции экспрессии
генов белковых гормонов-регуляторов водно-солевого обмена. Во-вторых,
свойства гуморальных факторов системного действия, как возможных
индукторов эпигенетической трансформации ренальной паренхимы. Общеизвестна роль аргинин-вазопрессина, как системного регулятора
осмотического гомеостаза, определяющего острую и точную реакцию
организма на изменение пищевого и внутривенного поступления жидкости
осмотически активных веществ (Bourque C.W., 2008; Thornton S.N.; 2010;
Greenwood M.P. et al., 2015; Park E.-J., Kwon T.-H., 2015). Физиологическая
роль ренин-ангиотензиновой системы определяется, как контролем
реабсорбции весьма значительного объема ультрафильтрата, растворенных
в нем натрия и калия, а также других жизненно важных компонентов
ультрафильтрата (Zhuo J.L., Li X.C., 2001; Kurtz A., 2012; Gomez R.A.,
Sequeira-Lopez M.L.S., 2018). Таким образом, Ангиотензин-II принимает
участие в регуляции показателей ионного, осмотического, волемического,
кислотно-основного гомеостаза, а также регулирует тонус кровеносных
сосудов. Атриальный (мозговой) натрий уретический пептид —
важнейший гуморальный регулятор волемического гомеостаза,
определяющий выведение натрия и жидкости на уровне дистального
отдела нефрона (Kuwahara К., Nakao К., 2010; Nakagawa Y. et al., 2019). 1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП) Результаты более ранних исследований позволили установить, что
изменения физиологических констант осмотического и волемического
гомеостаза оказывают влияние на уровни транскрипции гена аргинин
вазопрессина (АВП) (Kondo N. et al., 2004). Кроме того, авторами
цитируемой публикации была выявлена корреляция между концентрацией
катионов натрия во внеклеточной жидкости и уровнем экспрессии гена
аргинин вазопрессина. Было продемонстрировано также резкое усиление
транскрипции гена АВП под влиянием осмотического стимула (Hindmarch
C.C.T., Murphy D., 2010). Наряду с этим, было показано, что
гиперосмотический стимул усиливает транскрипцию ряда генов, белки
которых аккумулируются в задней доли гипофиза (Hindmarch C. et al., 27 2006). Выявлено, что активация транскрипции гена аргинин вазопрессина,
под влиянием осмотического воздействия, демонстрирует более
выраженную чувствительность к стимулу, в сравнении с другими
нейропептидами задней доли гипофиза (Yue C. et al., 2008). Сложность
вопроса в том, что к осмотическому стимулу чувствительны также гены
гипоталамо-гипофизарной оси, принимающие участие в регуляции
репродуктивной сферы (Qiu J. et al., 2007). В то же время, было
установлено, что осмотические нагрузки оказывают специфическое
влияние на экспрессию вполне определенной группы генов в
супраоптическом ядре крысы (Johnson K.R. et al., 2015). При этом,
необходимо отметить, что, вероятно, ген аргинин вазопрессина содержит
нуклеотидную последовательность в области промотора, обладающей
чувствительностью к изменениям показателей осмотического гомеостаза
(Ponzio T.A. et al., 2012). Авторами установлено отличие в первичной
последовательности нуклеотидов данного участка генов аргинин
вазопрессина и окситоцина. Далее, сопоставляя классическую схему
физиологического контроля осмотического гомеостаза и факты,
подтверждающие участие эпигенетических механизмов, опираясь на выше
изложенные результаты исследований, мы констатируем, что показатель
экспрессии гена аргинин вазопрессина обладает чувствительностью к
сдвигам осмоляльности внеклеточной жидкости организма. Вероятный
механизм влияния физико-химических условий внеклеточной жидкости
(концентрации хлорида натрия во внеклеточной жидкости) на состояние
транскрипции гена аргинин вазопрессина, в основном, подтвердили ранее
выполненные наблюдения (Kondo N. et al., 2004; Hindmarch C.C., Murphy
D., 2010). При этом отмечается, что АВП, помимо регуляции
осмотического гомеостаза, может отвечать за поведенческие реакции,
поэтому, с точки зрения авторов, нарушения осмотического гомеостаза
могут негативно отражаться на адаптивных поведенческих реакциях
(Mitchell N.C. et al., 2018). Показано, что экспрессия гена аргинин
вазопрессина демонстрирует высокий уровень пластичности, и что
интенсивность метилирования ДНК в области помотора гена гормона
может существенно изменяться в зависимости от состояния показателей
осмотического гомеостаза организма (Greenwood M.P. et al., 2016). Сообщается о видоспецифических молекулярных механизмах,
вовлеченных в индукцию транскрипции аргинин вазопрессина, на фоне
дегидратации организма (Stewart L. et al., 2011). 1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП) Высокий уровень
пластичности эпигенетических систем контроля биосинтеза аргинин
вазопрессина подтверждается тем фактом, что усиление транскрипции гена
гормона регистрируется в условиях острого гиперосмотического стимула
раствором хлорида натрия (Kawasaki M. et al., 2009). В настоящее время 2006). Выявлено, что активация транскрипции гена аргинин вазопрессина,
под влиянием осмотического воздействия, демонстрирует более
выраженную чувствительность к стимулу, в сравнении с другими
нейропептидами задней доли гипофиза (Yue C. et al., 2008). Сложность
вопроса в том, что к осмотическому стимулу чувствительны также гены
гипоталамо-гипофизарной оси, принимающие участие в регуляции
репродуктивной сферы (Qiu J. et al., 2007). В то же время, было
установлено, что осмотические нагрузки оказывают специфическое
влияние на экспрессию вполне определенной группы генов в
супраоптическом ядре крысы (Johnson K.R. et al., 2015). При этом,
необходимо отметить, что, вероятно, ген аргинин вазопрессина содержит
нуклеотидную последовательность в области промотора, обладающей
чувствительностью к изменениям показателей осмотического гомеостаза
(Ponzio T.A. et al., 2012). Авторами установлено отличие в первичной
последовательности нуклеотидов данного участка генов аргинин
вазопрессина и окситоцина. Далее, сопоставляя классическую схему
физиологического контроля осмотического гомеостаза и факты,
подтверждающие участие эпигенетических механизмов, опираясь на выше
изложенные результаты исследований, мы констатируем, что показатель
экспрессии гена аргинин вазопрессина обладает чувствительностью к
сдвигам осмоляльности внеклеточной жидкости организма. Вероятный
механизм влияния физико-химических условий внеклеточной жидкости
(концентрации хлорида натрия во внеклеточной жидкости) на состояние
транскрипции гена аргинин вазопрессина, в основном, подтвердили ранее
выполненные наблюдения (Kondo N. et al., 2004; Hindmarch C.C., Murphy
D., 2010). При этом отмечается, что АВП, помимо регуляции
осмотического гомеостаза, может отвечать за поведенческие реакции,
поэтому, с точки зрения авторов, нарушения осмотического гомеостаза
могут негативно отражаться на адаптивных поведенческих реакциях
(Mitchell N.C. et al., 2018). Показано, что экспрессия гена аргинин
вазопрессина демонстрирует высокий уровень пластичности, и что
интенсивность метилирования ДНК в области помотора гена гормона
может существенно изменяться в зависимости от состояния показателей
осмотического гомеостаза организма (Greenwood M.P. et al., 2016). Сообщается о видоспецифических молекулярных механизмах,
вовлеченных в индукцию транскрипции аргинин вазопрессина, на фоне
дегидратации организма (Stewart L. et al., 2011). Высокий уровень
пластичности эпигенетических систем контроля биосинтеза аргинин
вазопрессина подтверждается тем фактом, что усиление транскрипции гена
гормона регистрируется в условиях острого гиперосмотического стимула
раствором хлорида натрия (Kawasaki M. et al., 2009). В настоящее время 28 имеются данные и о том, какие энзиматические системы, отвечающие за
ковалентную трансформацию хроматина принимают участие в изменении
транскрипции гена аргинин вазопрессина (Archer T., 2015). Дальнейшие
исследования, проведенные научными сотрудниками групппы Murphy D. 1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП) показали, что к чувствительностью к осмотическому стимулу обладают
целый ряд генов (Caprin2), белки которых могут быть важны в
формировании адаптивного ответа супраоптических ядер гипоталамуса на
изменения осмотического гомеостаза организма (Loh S.-Y. et al., 2017). При том, что показана роль гена Caprin2 в механизмах стабилизации
матричной РНК аргинин вазопрессина (Konopacka A. et al., 2015). Высказанный тезис можно дополнить сведениями о том, что микро РНК
также принимают участие в эпигенетической модуляции активности
нейро-эндокринного контроля осмотического гомеостаза (Luo Y. et al.,
2014). В этом блоке анализа данных литературы необходимо выделить тот
факт, что аргинин вазопрессин может непосредственно контролировать
экспрессию транспортного белка
Na+,K+,2Cl-
котранспортера в
восходящей петле Генле нефрона (Konopacka A. et al., 2015). Однако,
непосредственно усиление экспрессии гена Na+,K+,2Cl-котранспортера по
влиянием аргинин вазопрессина, рассматривается в качестве
долговременной АВП- зависимой стимуляции белка (Knepper M.A. et al.,
2015). Вместе с тем, авторы обзора подчеркивают, что аргинин вазопрессин
может контролировать в дистальных сегментах нефрона экспрессию таких
транспортных белков, как: натрий-хлор котранспортирующий протеин,
переносчик мочевины, некоторые субъединицы эпителиального натриевого
канала порообразующих белков аквапоринов. Подчеркивается
актуальность данных механизмов в изучении патогенеза заболеваний почек
и сердечно-сосудистой системы (Qian Q., 2018). Также анализируется
АВП-зависимые системы внутриклеточной передачи сигнала (через
протеин киназы) в эпителии собирательных трубочек канальцевого отдела
нефрона, как звено индукции эпигенетического контроля экспрессии генов
транспортных белков (Sanghi A. et al., 2014). С другой стороны,
анализируется взаимосвязь различных изоформ аденилатциклаз и
протеинкиназ в системе регуляции генов транспортных белков в эпителии
собирательных трубочек (Roos K.P. et al., 2013). Завершая рассмотрение роли эпигенетических механизмов в
поддержании осмотического гомеостаза, необходимо отметить участие
АВП в долговременной стимуляции биосинтеза и экспрессии аквапорина-2
в эпителии собирательных трубочек канальцевого отдела нефрона (Wilson
J.L.L. et al., 2013). Были установлены механизмы активации транскрипции
гена аквапорина-2, включающие в себя механизмы внутриклеточной 29 передачи сигнала, а также
идентифицированы участки ДНК
предполагаемого связывания регулятора транскрипции (Yua M.-J. et al.,
2009). Проведен анализ метаболизма белка аквапорина-2 в эпителии
собирательных трубочек канальцевого отдела нефрона и роль АВП в
управлении транскрипции гена AQP2 (Jung H.J., Kwon T.H., 2016). Наряду
с этим, авторы цитируемого обзор указывают на роль микро РНК (miR-32
и miR-137) в процессах внутриклеточного метаболизма протеина
аквапорина-2. Оценивая роль эпигенетического контроля физиологических
функций собирательных трубочек (Xiao Z. et al., 2016), авторы приходят к
выводу, что баланс активности ядерных метилтрансфераз (Dot1lAC и
Dot1lf/f) в эпителии данного сегмента нефрона, может оказывать
существенное влияние на экспрессию белка аквапорин-2. 1.2.1. АРГИНИН-ВАЗОПРЕССИН (АВП) Поскольку
изменения в продукции и биологических эффектах аргинин вазопрессина
имеют отношение не только к регуляции водно-солевого гомеостаза, но и к
поведенческим реакциям человека, изучение эпигенетических процессов
контроля, например, рецепторов АВП также является объектом
междисциплинарных исследований (Bodden C. et al., 2017). 1.2.2. НАТРИЙУРЕТИЧЕСКИЕ ПЕПТИДЫ Интерес к эпигенетическим системам контроля АНП также в
некоторой степени обусловлен нейротропными эффектами гормона
(Frieling
H. et al.,
2008). Были достаточно подробно изучены
физиологически активные вещества, способные индуцировать
транскрипцию генов атриального (АНП), мозгового (БНП) и С-типа
натрийуретических пептидов, структура генов и самих натрийуретических
гормонов (Gardner D.G. et al., 2007; Kuwahara К., Nakao К., 2010; Ichiki T.,
Burnett J.C., 2017; Nakagawa Y. et al., 2019). Вместе с тем, имеются данные
о том, что АНП может синтезироваться эпителием канальцевого отдела
нефрона (Dong L. et al., 2016; Pandey K.N., 2018). При этом, в некоторых
обзорных публикациях высказывается тезис о важной практической
значимости исследований эпигенетического контроля экспрессии генов
натрийуретических пептидов (DiSalvo T.G., 2015; Man J. et al., 2018). Поскольку представляет интерес несколько аспектов данной проблемы:
эффективность использования параметров синтеза и секреции
натрийуретических пептидов в качестве диагностических маркеров ряда
актуальных нозологий, исследования собственно механизмов контроля
экспрессии генов этих гормонов и вовлеченность в этот процесс некоторых
гормонов и цитокинов, участвующих в патогенезе заболеваний сердечно-
сосудистой системы и почек: ангиотензин-2, трансформирующий фактор
роста-бета1, гормоны щитовидной железы (Sergeeva I.A., Christoffels V.M.,
2013). Вместе с тем, данные литературы подчеркивают значение уровня 30 экспрессии рецепторов натрийуретических пептидов в сердечно-
сосудистой системе и ренальной паренхиме для понимания
физиологических и патофизиологических эффектов гормонов (Pandey K.N.,
2011; Kumar P. et al., 2014). ;
,
)
Результаты экспериментальных исследований показали, что
гипертрофия кардиомиоцитов токсического генеза сопровождается
снижением продукции miR-133a на фоне усиления метилирования ДНК
метилтрансферазами ДНК DNMT1 и DNMT3b, а также дозозависимым
увеличением уровня м-РНК АНП и БНП (Huang L. et al., 2016). Снижение
уровня miR-133a в миокарде было обнаружено у лабораторных крыс,
подвергавшихся продолжительной инфузии ангиотензина-2 (Li Y. et al.,
2016). Вместе с тем, авторы сообщают, что предварительное введение
животным рекомбинантного АНП благоприятно сказывалось на динамике
miR-133a. Показано, что в условиях неишемической кардиомиопатии
наблюдается снижение экспрессии генов АНП и БНП в кардиомиоцитах на
фоне усиления метилирования остатка лизина H3 гистона нуклеосомы (Ito
E. et al., 2017). Наряду с этим, авторами публикации выявлена взаимосвязь
биосинтеза АНП в кардиомиоцитах и продукции (miR-133a) микро РНК. Результаты дальнейших наблюдений показали, что микро РНК-30 может
принимать участие в регуляции синтеза БНП (Nakagawa Y. et al., 2019). Важная роль в регуляторном эффекте гормона отводится его рецепторам. В
этом смысле, привлекают внимание сведения о том, что уровень
экспрессии рецептора популяции А к натрийуретическому пептиду,
наиболее физиологически активных рецепторов к АНП (БНП), негативно
коррелирует с параметрами энзиматической активности изоформы ДНК-
метилтрансферазы DNMT3B (Shen K. et al., 2017). 1.2.2. НАТРИЙУРЕТИЧЕСКИЕ ПЕПТИДЫ Наряду с этим,
высказывается мнение о ключевой роли деацетилаз гистоновых белков
(HDAC4) в регуляции экспрессии генов АНП И БНП в норме и при
патологии (Hohl M. et al., 2013). В результате экспериментальных
исследований установлено, что нарушения функционального состояния
миокарда сопровождаются изменениями продукции АНП и БНП на фоне
деметилирования H3K9 в промоторных областях генов данных белков и
умеренного повышения ацетилирования Н3 гистона (H3K27ac) (Sergeeva
I.A. et al., 2016). Тогда показатели ацетилирования H3K9 существенно не
были изменены. Ранее полученные сведения также подчеркивают роль
ацетилирования белков гистонов в регуляции экспрессии рецептора
популяции А к натрийуретическому пептиду в ренальной паренхиеме,
(Kumar P., Pandey K.N., 2009). По мнению авторы цитируемой публикации,
ацетилтрансфераза
белков
гистонов
(Р300),
при
участии
специализированной микро РНК, может регулировать экспрессию гена
гуанилил циклазы-А/рецептора-А натрийуретического пептида. В 31 дальнейших исследованиях авторами было показано, что эпигенетический
механизм регуляции экспрессии гуанилил циклазы-А/рецептора-А
натрийуретического пептида (Npr1) на основе баланса ацетилирования
гистонов (ацетилтрансфераза Р300 и деацетилазы гистонов HDAC1/2),
может выполнять важную роль в поддержании физиологических констант
волемического гомеостаза организма (Kumar P. et al., 2014). Вместе с тем,
авторами высказывается мысль о том, что величина осмотического
давления во неклеточной жидкости, уровень продукции ангиотензина-II и
витамин Д могут оказывать влияние на показатели экспрессии гена Npr1. Д
у
р
p
Подчеркивается, что амплификация генов (Nppa и Nppb), а также
рецепторов натрийуретических пептидов, препятствует повышению
кровяного давления, способствует усилению почечного кровотока,
увеличению скорости клубочковой фильтрации, ограничивает процессы
воспаления и фиброза в ренальной паренхме (Pandey K.N., 2018). Авторы
цитируемого обзора также указывают, что натрийуретические пептиды
обладают способностью сдерживать активность ренин-ангиотензин-
альдостероновой системы. Обращает на себя внимание тот факт, что
значительное количество публикаций по данной тематике отмечают
антагонизм физиологических эффектов натрийуретических пептидов и
трансформирующего фактора роста бета1 (ТФР бета1). Причина такого
внимания, по нашему мнению, объясняется фундаментальной ролью ТФР
бета1 в ряде патогенетических механизмов нарушения функций органов
сердечно-сосудистой системы и почек (Chen L. et al., 2018). В этом смысле,
уместно привести данные о том, что ТФР бета1 может оказывать влияние
на систему натрийуретических пептидов, подавляя транскрипцию гена
Npr1 - основного рецептора гормонов в мышечном слое стенки аорты
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PMID:26934489 45 46 ГЛАВА 2. НЕКОТОРЫЕ ФАКТОРЫ АКТИВАЦИИ
ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ Приступая к обсуждению данной темы, считаем необходимым
подчеркнуть, что, во-первых, в отличие от предыдущих разделов, при
рассмотрении большинства факторов, включая факторы внешней среды,
способных оказывать влияние на экспрессию генов, речь идет о
надорганизменном уровне, чаще всего, о популяции. Во-вторых, косвенно
затронуты вопросы участия эпигенетических механизмов в процессах
наследования приобретенных признаков и их роли в эволюционных
преобразований. Возможно, сочетание новых признаков, обусловленных не
только мутациями, но и эпигенетическими механизмами внутри
популяции, может оказать влияние на процессы микроэволюции. Необходимо отметить, что в современной литературе уделяется внимание
поставленным вопросам. В частности, указывается, что предполагаемые
глобальные изменения климатических условий учитывают актуальность
эпигенетических преобразований для динамики адаптивных изменений
популяций человека (Hu J.,
Barrett R.D.H., 2017). Поэтому,
экспериментальные данные, полученные в исследованиях на животных
позволяют, с одной стороны, расширить наши представления о роли
эпигенетических система контроля адаптивных реакций на изменения
факторов среды. С другой стороны, предположить возможность
закрепления этих адаптивных преобразований экспрессии генов в ряду
поколений. При этом, авторы цитируемого обзора, во-первых,
подчеркивают важное значение эпигенетических механизмов для
экологической пластичности различных видов животных. Во-вторых,
приводят конкретные примеры передачи в последующие поколения
эпигенетических изменений хроматина у некоторых видов
млекопитающих. Наряду с этим, привлекают внимание сведения об
устойчивых сочетаниях генов, выполняющих ведущую роль в
формировании экологической пластичности животных к изменениям,
например, температурного режима окружающей среды (Wollenberg Valero
K.C. et al., 2014). Заслуживает внимания и тот факт, что комбинация
данных генов в ряду позвоночных животных обладает достаточно высокой
эволюционной консервативностью. Поэтому, необходимо отметить, что в современной литературе
высказываются мнения о том, что эпигенетические преобразования,
сформированные в генотипе родительских особей, могут выполнять
принципиально важную функцию в эволюционном процессе, поскольку
могут передаваться потомству и играть существенную роль в адаптивных
реакциях потомства (Wang Y. et al., 2017). Авторы приводят ряд 47 аргументов, подтверждающих возможность наследования эпигенетических
трансформаций хроматина и у человека. Аналогичная точка зрения,
относительно возможности наследования в поколениях эпигенетической
модуляции экспрессии генов, обусловленной, в первую очередь,
ковалентной модификацией хроматина, высказывается и в последующих
публикациях (Norouzitallab P. et al., 2019). Вместе с тем, приведенные сведения о наследовании
эпигенетических модификаций генома, во-первых, не являются
общепризнанными. Во-вторых, возможный тип наследования
эпигенетических трансформаций также мало изучен. Тем не менее, мы
посчитали необходимым включить в обзор краткое упоминание об этих
аспектах эпигенетики, поскольку возможность их реализации существует. ГЛАВА 2. НЕКОТОРЫЕ ФАКТОРЫ АКТИВАЦИИ
ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ Следовательно, рассматривая роль эпигенетических механизмов в
адаптивных реакциях почки на факторы среды (не только внешних) в
масштабе популяций, уместно принять к сведению возможность
наследования эпигенетических перестроек в системе регуляции экспрессии
генов, равно, как и их возможное участие в эволюционных процессах. Также, на наш взгляд, необходимо учитывать интересы практической
медицины, особенно, если речь идет об участии эпигенетических
процессов в патофизиологических механизмах заболеваний почек. В данном разделе, в качестве тем для обсуждения нами выбраны
факторы внешней среды, в том числе и антропогенной природы. Наряду с
этим, некоторые факторы, связанные с устойчивыми нарушениями
физиологических констант организма также достаточно широко
распространены в популяциях человека и заслуживают рассмотрения. 48 2.1. ИЗМЕНЕНИЕ ТЕМПЕРАТУРНОГО РЕЖИМА, КАК ФАКТОР
ИНДУКЦИИ ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМОВ Даже принимая к сведению тот факт, что представители
биологического вида Homo sapiens sapiens проживают в искусственно
созданной среде и используют различные способы формирования
микроклимата своих жилищ, климатические факторы среды, в частности,
температура среды, до настоящего времени оказывает исключительно
важное влияние на человека (Cheshire W.P. Jr., 2016; Beker B.M. et al., 2018). Обсуждение вероятности стремительных изменений климатических
условий на планете Земля не входит в круг наших задач. Вместе стем, мы
исходим из того, что уже существующее разнообразие климатических
условий различных географических широт, а также межсезонные
флуктуации климатических условий можно рассматривать, как важный
стимул в изучении роли эпигенетических процессов в адаптации организма
к изменению температурного режима среды (Franks S.J., Hoffmann A.A.,
2012; Wollenberg Valero K.C. et al., 2014). В данном случае, температурный
режим рассматривается в качестве одного из основных факторов среды,
способных вызывать устойчивые эпигенетические изменения структуры
ДНК человека, которые, направлены на повышение адаптивных
возможностей популяции и, вероятно, могут передаваться по наследству
(Giuliani C. et al., 2015). Вместе с тем, авторы иллюстрируют адаптивный
характер эпигенетических изменений, адекватных геофизическим
условиям проживания данных популяций человека. Почки наземных позвоночных животных (амниот) выполняют
жизненно важную функцию поддержания постоянства внутренней среды
организма. При этом, с одной стороны, физиологические и
патофизиологические аспекты адаптации почек человека к изменению
температурного режима среды постоянно находятся в центре внимания
современной науки (Johnson R.J. et al., 2016; de Lorenzo A., Liaño F., 2017). С другой стороны, участие эпигенетических механизмов в этих процессах
требует более глубокого изучения. Тем не менее, установлено, что, в
частности, тепловой стресс оказывает мощное влияние на перестройку
метаболизма микро РНК в почках (Permenter M.G. et al., 2019). Вместе с
тем, авторы цитируемой публикации отмечают органоспецифический
характер изменений метаболизма микро РНК под влиянием повышения
температуры. Ранее было показано, что белки семейства аквапорины также
могут изменять свою экспрессию в почках и слюнных железах под
влиянием температурного фактора (как повышение, так и понижение
температуры) у позвоночных животных (Wollenberg Valero K.C. et al.,
2014). 49 2.2. ГИПОКСИЯ. Наряду с температурным фактором, одним из важнейших факторов
среды, способным оказать влияние на состояние эпигенетических
механизмов человека, является гипоксия (Giuliani C. et al., 2015). Гипоксическая гипоксия может оказывать влияние на структурные
показатели и функциональное состояние ренальной паренхимы через
систему HIFs-протеинов, контролируя экспрессию генов, белки которых
критически важны для регуляции деятельности почек (Poonit N.D. et al.,
2018). С другой стороны, гипоксия ренальной паренхимы различного
генеза рассматривается в качестве одного из базовых индукторов
эпигенетических механизмов трансформации гуморальных систем
контроля гомеостатических функций почек человека (Clarke N.E., Turner
A.J., 2012; Macconi D. et al., 2014). Известно, что, стимулируемый
гипоксией
HIF-1альфа, является одним из ведущих активаторов
эпигенетических механизмов (Perez-Perri J.I. et al., 2011). Являясь важным
звном в системе адаптации почки к гипоксии, HIF-1альфа может быть
непосредственно вовлечен в патогенетические механизмы хронизации и
прогрессирования почечной недостаточности (Shoji K. et al., 2014). Установлено, что HIFs-зависимое угнетение метилирования гистонов
(H3K9me3 и H3K27me3) может сопутствовать прогрессированию почечной
недостаточности (Nangaku M. et al., 2017). Сообщается, что эпигенетические механизмы активации ренин-
ангиотензиновой системы могут выполнять ключевую роль в хронизации и
прогрессирования почечной недостаточности (Chou Y.H. et al., 2017). Вместе с тем, показано, что HIF-1альфа на уровне транскрипции изменяет
баланс экспрессии компонентов РАС в направлении стимуляции
биосинтеза компонентов оси Ангиотензин-I-превращающий фермент
(АСЕ)/Ангиотензин-2/АТ1-рецепторы
против
угнетения
контура
отрицательной обратной связи РАС АСЕ-2/Ангиотензин-1-7/MASS1-
рецепторов (Clarke N.E., Turner A.J., 2012; Macconi D. et al., 2014). Помимо
того, что HIF-1альфа усиливает экспрессию АТ1-рецепторов и АСЕ, в
условиях гипоксии в почке наблюдается резкая активация АСЕ-
независимого пути образования Ангиотензин-I
в присутствии
индуцированного гипоксией лактат-химаза-зависимого механизма (Xie G. et al., 2017). В совокупности, индуцированное гипоксией смещение
баланса в пользу оси Ангиотензин-I-превращающего фермента
(АСЕ)/Ангиотензин-2/АТ1-рецептор
против
угнетения
контура
отрицательной обратной связи РАС АСЕ-2/Ангиотензин-1-7/MASS1
способствует активации воспаления, нарушению клеточного цикла клеток
ренальной паренхимы, состоянию энергетического обмена нефроцитов, а 50 также активации эпителиально-мезенхимальной трансформации (Macconi
D. et al., 2014; Chou Y.H. et al., 2017). Эпигенетические механизмы,
стимулированные гипоксией, выполняют важную роль в хронизации и
прогрессирования почечной недостаточности, индуцируя нарушение
функции подоцитов (Lin C.-L. et al., 2014) и мезангиума (Lu Z. et al., 2017). По мнению ряда исследователей, ключевым звеном в этом процессе
является поражение проксимального отдела нефрона (Matsusaka T. et al.,
2012; Kobori H. et al., 2013). Наряду с этим приводятся аргументы о том,
что стимулируемые семейством HIF-протеинов эпигенетический процессы
являются перспективным объектом фармакологических методов
сдерживания прогрессирующей почечной недостаточности (Shoji K. et al.,
2014). также активации эпителиально-мезенхимальной трансформации (Macconi
D. et al., 2014; Chou Y.H. et al., 2017). 2.2. ГИПОКСИЯ. Эпигенетические механизмы,
стимулированные гипоксией, выполняют важную роль в хронизации и
прогрессирования почечной недостаточности, индуцируя нарушение
функции подоцитов (Lin C.-L. et al., 2014) и мезангиума (Lu Z. et al., 2017). По мнению ряда исследователей, ключевым звеном в этом процессе
является поражение проксимального отдела нефрона (Matsusaka T. et al.,
2012; Kobori H. et al., 2013). Наряду с этим приводятся аргументы о том,
что стимулируемые семейством HIF-протеинов эпигенетический процессы
являются перспективным объектом фармакологических методов
сдерживания прогрессирующей почечной недостаточности (Shoji K. et al.,
2014). 51 51 2.3. ГИПЕРГЛИКЕМИЯ Гипергликемия,
в
подавляющем
большинстве
случаев,
рассматривается в качестве симптома, сопутствующего течению сахарного
диабета. Тем не менее, устойчиво повышенный уровень глюкозы во
внеклеточной жидкости выступает в качестве самостоятельного
патогенетического фактора ренальных дисфункций (Dounousi E. et al.,
2015), способного инициировать дальнейшее прогрессирование почечной
недостаточности при участии ковалентной трансформации хроматина
(Reddy M.A, Natarajan R., 2015; Lu Z. et al., 2017). Обсуждая роль
гипергликемии в эпигенетических механизмах перестройки функции
почки, необходимо отметить, что данному симптому сахарного диабета 2-
го типа сопутствует также изменения секреции инсулина, нарушения
обменных процессов, усиление продукции активных форм кислорода,
нарушение параметров системной и внутриорганной гемодинамики,
повышение уровня HIF-1 (Reddy M.A, Natarajan R., 2015). По мнению
цитируемых авторов, HIF-1 обладает способностью стимулировать
эпигенетические механизмы активации экспрессии ферментов деметилаз
гистонов. Высказывается мнение о том, что гипергликемия в значительной
степени ответственна за ряд характерных изменений систем передачи
внутриклеточного сигнала в целом ряде различных популяций клеток
почки, включая клетки канальцевого эптелия,
фибробласты,
эндотелиоциты, клетки мезангиума и подоциты (Reddy M.A, Natarajan R.,
2015). В обзоре указывается, что стимуляция фиброза тканей почки может
усиливаться TGF-β, индуцирующего повышение таких эпигенетических
меток, как miR-29, H3K9/14Ac, H3K9Ac, H3K4me1 и H3K4me3, на фоне
снижения
H3K9me3. Указанные изменения могут сопровождаться
усилением экспрессии гена AGT (ангиотензиногена) в проксимальных
нефроцитах, вызванной ингибированием DNMT и повышением активности
HDAC. С другой стороны, следует учитывая роль сопутствующих
сахарному диабету изменений гемодинамических параметров на
эпигенетические процессы. Известно, что устойчивое повышение
кровяного давления может способствовать повышению экспрессии гена
Асе (ангиотензин-превращающего фермента) в том числе и в почках через
повышение уровня меток H3KAc и H3K4me, на фоне снижения экспрессии
метки H3K9me2 (Liang M. et al., 2013; Reddy M.A, Natarajan R., 2015). Эпигенетические механизмы патогенеза и прогрессирования
гипертонической болезни рассмотрены в ряде обзорных публикаций (Friso
S. et al., 2013; Wise I.A., Charchar F.J., 2016). Авторами цитируемых работ
указан ряд генов, экспрессия которых тесно связана с течением
гипертонии, включая гены ренина, АСЕ, рецепторов ангиотензина-2 и 52 эндотелиальной NO-синтазы. Эпигенетическая перестройка экспрессии
генов системы NO-синтаз может индуцироваться гипоксией (Fish J.E. et al.,
2010) и гипергликемией (Advani A. et al., 2011; Schmidt Dellamea B. et al.,
2014). Показано, что ингибитор ферментов деацетилаз белков гистонов
vorinostat способствует снижению албуминурии, отложению коллагена IV
клетками мезангиума, а также оксидативный стресс в экспериментальной
модели сахарного диабета 1 типа (Advani A. et al., 2011). эндотелиальной NO-синтазы. Эпигенетическая перестройка экспрессии
генов системы NO-синтаз может индуцироваться гипоксией (Fish J.E. et al.,
2010) и гипергликемией (Advani A. et al., 2011; Schmidt Dellamea B. et al.,
2014). 2.3. ГИПЕРГЛИКЕМИЯ Показано, что ингибитор ферментов деацетилаз белков гистонов
vorinostat способствует снижению албуминурии, отложению коллагена IV
клетками мезангиума, а также оксидативный стресс в экспериментальной
модели сахарного диабета 1 типа (Advani A. et al., 2011). 53 53 2.4. ТЯЖЕЛЫЕ МЕТАЛЛЫ Широко известно нефротоксическое действие тяжелых металлов. Наряду с этим в литературе имеются отдельные сведения об их
эпигенетических эффектах (Ruiz-Hernandez A. et al., 2015). В частности,
авторами показано усиление метилирование ДНК в зависимости от
продолжительности экспозиции к кадмию, a также общая тенденция к
гипометилированию ДНК на фоне повышения свинца в крови. Относительно ртути, экспериментальные исследования свидетельствуют о
том, что ртуть может изменить характер метилирования ДНК. В
эмбриональных стволовых клетках крысы метилртуть уменьшала
пролиферацию нервных клеток, в связи с гипометилированием ДНК. Также авторы цитируемой публикации сообщают, что механизмы
индукции тяжелыми металлами эпигенетической перестройки ДНК
остаются крайне мало изучены. Поскольку высоко токсичные тяжелые
металлы (ртуть, кадмий и свинец) в организм человека поступают, как
правило, в следовых количествах не вызывая острого токсического
эффекта, представляет интерес анализ их влияния на изменение обменных
процессов в организме, эндокринных функций поджелудочной железы, в
патогенезе резистентности тканей к инсулину и избыточной массы тела
(Kuo C.-C. et al., 2013). (
)
Действительно, эпигенетические эффекты тяжелых металлов могут
быть индуцированы достаточно низкими уровнями поступления
ксенобиотиков, как правило, не превышающие санитарные нормы. Наиболее ранние публикации, посвященные данной тематике, содержат
информацию о том, что, например, тяжелый металл Со2+ может
стимулировать процессы транскрипции некоторых белков независимо от
внутриклеточной эндогенной продукции активных форм кислорода
(Salnikow K. et al., 2000). В дальнейшем были непосредственно указаны
индуцированные Со2+, HIF 1α-зависимые эпигенетические механизмы,
‐
связанные с ферментными системами метилирования ДНК и
ацетилирования гистонов (Maxwell P., Salnikow K., 2004). В современной
литературе роль эпигенетических механизмов в реализации токсических и
канцерогенных эффектов тяжелых металлов широко признана (Salnikow
K., Zhitkovich A., 2008; Chervona Y, Costa M., 2012; Brocato J., Costa M.,
2013). Также широко признана важность роли HIF 1α-зависимых
‐
эпигенетических механизмов, индуцируемых тяжелыми металлами
(Salnikow K. et al., 2008; Nagasawa H., 2011; Brocato J., Costa M., 2013;
Eskandani M. et al., 2017). Было также показано, что стимуляция кобальтом
отложений
белков внеклеточного матрикса, а также индукция
регуляторных пептидов VEGF и эритропоэтина связаны с HIF 1α
‐
(Tanaka 54 T. et al., 2005). По нашему мнению, научная новизна предлагаемого
подхода, состоит в том, что впервые было предложено теоретически
обоснованное эпигенетическими механизмами контроля экспрессии генов
объяснение патогенеза смертельно опасных онкологических заболеваний,
индуцированных тяжелыми металлами. При этом, патогенез этих
заболеваний не рассматривался, как результат прямого повреждения ДНК. Был разработан подход, основанный на малигнизирующих эффектах
тяжелых металлов, обусловленных специфической ковалентной
модификацией хроматина, изменяющей экспрессию генов (Salnikow K.,
Zhitkovich A., 2008; Salnikow K. et al., 2008). 2.4. ТЯЖЕЛЫЕ МЕТАЛЛЫ Продуктивность такого
подхода была подтверждена последующими результатами исследований
(Chervona Y, Costa M., 2012; Brocato J., Costa M., 2013). (
,
,
;
,
,
)
Результаты исследований in vitro на культуре малигнизированных
клеток показали, что присутствие в среде тяжелых металлов оказывает
существенное влияние на уровни HIF-1α в клетках, а также на состояние
экспрессии генов, идентифицированных, как гены факторов транскрипции,
маркеров дифференциации клеток, цитокинов и факторов роста,
протеинкиназ, супрессоров опухолей и онкогенов (Bae S. et al., 2012). По
данным авторов, им удалось выделить группу генов, чувствительных, в
частности, к ионам Со2+. Установлено также, что тяжелые металлы, через
процессы ацетилирования белков-гистонов, регулирует экспрессию гена
внеклеточной супероксид дисмутазы (Hattori S. et al., 2016). С другой
стороны, показано, что применение в эксперименте ингибитора деацетилаз
гистоновых белков (valproic acid),
способствует ослаблению
патофизиологических эффектов HIF-1α (Luo H.-M. et al., 2013; Kim Y.J. et
al., 2017). Наряду с этим, показано, что HIF-1α может регулировать не только
ковалентную модификацию хроматина, но и биосинтез малых
некодирующих РНК, способных определять биосинтез белка на уровне
транскрипции или трансляции (Kwak J. et al., 2018). Действительно, в
литературе имеются данные о том, что HIF могут оказывать влияние на
системы метаболизма некодирующих малых РНК (Ho J.J. et al., 2012;
Ibrahim A.A. et al., 2017). При этом, в исследованиях in vitro установлена
связь между присутствием в среде дихлорида кобальта, HIFs протеинами и
показателями экспрессии клетками микро РНК (Silakit R. et al., 2018). Приводятся данные о том, что HIF-зависимые механизмы, через систему
микро РНК принимают участие в регуляции экспрессии
провоспалительных цитокинов (Kwak J. et al., 2018). Механизмы индукции
Со2+ процессов воспаления занимают важное место в патогенезе
кобальтовой интоксикации, однако роль эпигенетических механизмов,
определяющих синтез (в том числе и микро РНК) провоспалительных 55 факторов белковой природы изучены пока не достаточно (Kumanto M. et
al., 2017). По нашему мнению, в литературе проведен достаточно детальный
анализ роли тяжелых металлов в индукции базовых механизмов
эпигенетической трансформации систем контроля экспрессии генов. В
тоже время, нельзя исключить определенных органоспецифических
особенностей их реализации. Например, в почках. При том, что ренальная
паренхима является одной из основных мишеней для данной группы
ксенобиотиков. 56 56 2.5. ЭНДОКРИНОПАТИИ Течение эндокринопатий связано с тем, что на состояние
эпигенетических механизмов может одновременно оказывать
существенное влияние несколько факторов. Пример такого
комбинированного влияния мы уже рассматривали, анализируя
эпигенетические эффекты гипергликемии. Вместе с тем, фактор
неадекватной секреции инсулина и изменение чувствительности тканей к
гормону не является второстепенным и может участвовать в
эпигенетических механизмах регуляции деятельности почки (Shiels P.G. et
al., 2017). Авторы цитируемого обзора рассматривают роль инсулина в
эпигенетической системе контроля деятельности почек в процессе
возрастных изменений функции органа. В этом смысле, представляют
интерес сведения о базовых эпигенетических механизмах, способных
детерминировать резистентность тканей к регуляторному воздействию
инсулина (Seok S. et al., 2018). Сложность точной оценки степени влияния
различных факторов (гипергликемия, изменение чувствительности тканей
к инсулину, оксидативный стресс и т.д) течения сахарного диабета второго
типа на перестройку экспрессии генов — вполне объективная проблема. Вместе с тем, в литературе имеются данные о том, что собственно
резистентность к инсулину может, через регуляцию метилирования
гистонов, принимать участие в патофизиологических механизмах
нарушении целостности слоя подоцитов, провоцируя усиление
альбуминурии и прогрессирование нефропатии (Lizotte F. et al., 2016). Действительно, ранее экспериментально было подтверждено участие
инсулина в регуляции экспрессии генов мыши и человека через систему
метилирования ДНК (Kuroda A. et al., 2009). Известно также, что альдостерон через систему метилирования
гистонов может непосредственно регулировать экспрессию гена альфа-
субъединицы эпителиального натриевого канала дистального отдела
нефрона αENAC (Kone B.C., 2013), а также эндотелина-1 (Welch A.K. et al.,
2016). Высказывается мнение, что понимание этих эпигенетических
механизмов альдостерона представляет интерес, как в лечении
гипертонической болезни, так и в борьбе с избыточным весом (Kawarazaki
W., Fujita T., 2016). В качестве потенциального индуктора эпигенетической
трансформации гуморальных систем контроля гомеостатических функций
почек можно упомянуть гормоны щитовидной железы. В литературе
имеются указания на регуляторные эффеты гормонов щитовидной железы,
рассматриваемых, как природные ингибиторы ацетилазы белков-гистонов
(Re A. et al., 2016). Также установлено, что эпигенетические эффекты
тироксина
стимуляции
деацетилазы гистонов-5
(HDAC5)
могут 57 реализовываться через путь передачи сигнала, сопряженный с интегрином
αvβ3/PKD/HDAC5 (Liu X. et al., 2014). В литературе представлены данные
и о том, что в условиях гипофункции щитовидной железы также
наблюдается закономерное изменение экспрессии некоторых генов через
механизм импринтинга (Hu Z. et al., 2014; Leow M.K., 2016). Следовательно, как гипо- так и гипертиреоз могут рассматриваться в
качестве потенциальных индукторов эпигенетической перестройки
гуморальных систем контроля деятельности почки. Поскольку широко
известен тот факт, что нарушение тиреоидного статуса организма
усиливает риск заболевания почек через активацию РАС (Kobori H. et al.,
1999). реализовываться через путь передачи сигнала, сопряженный с интегрином
αvβ3/PKD/HDAC5 (Liu X. 2.5. ЭНДОКРИНОПАТИИ et al., 2014). В литературе представлены данные
и о том, что в условиях гипофункции щитовидной железы также
наблюдается закономерное изменение экспрессии некоторых генов через
механизм импринтинга (Hu Z. et al., 2014; Leow M.K., 2016). Следовательно, как гипо- так и гипертиреоз могут рассматриваться в
качестве потенциальных индукторов эпигенетической перестройки
гуморальных систем контроля деятельности почки. Поскольку широко
известен тот факт, что нарушение тиреоидного статуса организма
усиливает риск заболевания почек через активацию РАС (Kobori H. et al.,
1999). реализовываться через путь передачи сигнала, сопряженный с интегрином
αvβ3/PKD/HDAC5 (Liu X. et al., 2014). В литературе представлены данные
и о том, что в условиях гипофункции щитовидной железы также
наблюдается закономерное изменение экспрессии некоторых генов через
механизм импринтинга (Hu Z. et al., 2014; Leow M.K., 2016). Следовательно, как гипо- так и гипертиреоз могут рассматриваться в
качестве потенциальных индукторов эпигенетической перестройки
гуморальных систем контроля деятельности почки. Поскольку широко
известен тот факт, что нарушение тиреоидного статуса организма
усиливает риск заболевания почек через активацию РАС (Kobori H. et al.,
1999). 58 58 2.6. ЭПИГЕНЕТИЧЕСКИЕ ПРОЦЕССЫ, ИНДУЦИРОВАННЫЕ
ПАТОГЕННЫМИ МИКРООРГАНИЗМАМИ Воспалительные реакции тканей почки человека в ответ на
инфекционные и неинфекционные заболевания, анализируются с учетом
их популяционных особенностей с позиций современных взглядов на
филогенез выделительной системы и принципы антропогенеза (Chevalier
R.L., 2017). В литературе подчеркивается роль эпигенетических
механизмов в эволюционных аспектах формирования адаптивных реакций
иммунной системы и тканей почки. При этом, особое внимание уделяется
механизмам иммунопатологии почки. В связи с этим в ряде публикаций
высказывается мнение о том, что у человека эпигенетическая перестройка
(метилирование и ацетилирование гистонов) клеток моноцитарного ряда,
направленная на регуляцию выработки провоспалительных факторов,
может сохранятся и передаваться дочерним клеткам, определяя
особенности течения заболевания (Venet F., Monneret G., 2018). По мнению
некоторых исследователей, эпигенетические изменения в иммунной
системе, вызванные хроническим воспалением и повышенным
окислительным стрессом, могут рассматриваться в качестве базового
патогенетического механизма патологии почек и могут приводить к
необратимым нарушениям ренальной паренхимы (Syed-Ahmed M.,
Narayanan M., 2019). Помимо этого, на основе результатов популяционных исследований
была проанализирована возможная роль микрофлоры организма человека в
эпигенетической перестройке иммунных реакций, связанных с риском
заболеваний почек (Uy N. et al., 2015). Дальнейшие исследования показали,
что состояние микрофлоры организма может оказывать влияние на риск
заболевания почек через эпигенетические механизмы перестройки
внутрипочечной РАС (Marques F.Z. et al., 2017). По мнению некоторых
авторов, нарушения функции почек могут быть тесно связаны с
нарушениями микрофлоры кишечника, поскольку данный показатель
оказывает влияние на состояние иммунитета кишечника таким образом,
что он больше не может поддерживать физиологический контроль
микробиоты (Syed-Ahmed M., Narayanan M., 2019). Авторы цитируемого
обзора рассматривают эпигенетическую активацию провоспалительных
реакций, возможно, за пределами почечной паренхимы, как мощный
индуктор патологических изменений органа. Аналогичную точку зрения
высказывают и другие авторы, обращая внимание на тот факт, что
эпигенетические механизмы могут выполнять определенную роль в
патогенезе прогрессирующей почечной недостаточности на фоне
нарушений микрофлоры кишечника (Lu C.C. et al., 2018). Наряду с эим, 59 привлекают внимание сведения о том, что метаболиты микрофлоры
кишечника могут оказывать влияние на состояние внутрипочечной ренин-
ангиотензиновой системы. Предполагая наличие патогенетических
механизмов активации внутрипочечных систем гуморального контроля
гомеостатических функций почек — ренин-ангиотензиновой системы. Показана актуальность эпигенетической индукции РАС и при вирусной
инвазии (Chandel N. et al., 2013). привлекают внимание сведения о том, что метаболиты микрофлоры
кишечника могут оказывать влияние на состояние внутрипочечной ренин-
ангиотензиновой системы.
Предполагая наличие патогенетических
механизмов активации внутрипочечных систем гуморального контроля
гомеостатических функций почек — ренин-ангиотензиновой системы.
Показана актуальность эпигенетической индукции РАС и при вирусной
инвазии (Chandel N. et al., 2013). привлекают внимание сведения о том, что метаболиты микрофлоры
кишечника могут оказывать влияние на состояние внутрипочечной ренин-
ангиотензиновой системы. Предполагая наличие патогенетических
механизмов активации внутрипочечных систем гуморального контроля
гомеостатических функций почек — ренин-ангиотензиновой системы. Показана актуальность эпигенетической индукции РАС и при вирусной
инвазии (Chandel N. et al., 2013). привлекают внимание сведения о том, что метаболиты микрофлоры
кишечника могут оказывать влияние на состояние внутрипочечной ренин-
ангиотензиновой системы. Предполагая наличие патогенетических
механизмов активации внутрипочечных систем гуморального контроля
гомеостатических функций почек — ренин-ангиотензиновой системы. Показана актуальность эпигенетической индукции РАС и при вирусной
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doi: 10.1189/jlb.0812383 68 ГЛАВА 3. ФАКТОРЫ ВНУТРИОРГАННОЙ ГУМОРАЛЬНОЙ
РЕГУЛЯЦИИ ДЕЯТЕЛЬНОСТИ ПОЧЕК. ИХ МЕСТО И РОЛЬ В
ЭПИГЕНЕТИЧЕСКИХ МЕХАНИЗМАХ НАРУШЕНИЯ
ФУНКЦИОНАЛЬНОГО СОСТОЯНИЯ РЕНАЛЬНОЙ ПАРЕНХИМЫ Обсуждая физиологические и патофизиологические аспекты
эпигенетического контроля экспрессии генов клеток ренальной паренхимы,
необходимо учитывать, что почки обладают автономной системой
продукции тканевых гормонов, с одной стороны, принимающих участие в
системе ауторегуляции гомеостатических функций и внутриорганного
кровотока. С другой стороны, способных индуцировать эпигенетическую
трансформацию экспрессии регуляторных и транспортных белков в
интересах системного контроля гомеостаза. Вместе с тем, данным
эпигенетическим системам контроля экспрессии генов отводится важная
патофизиологическая роль в индукции патогенеза почечной
недостаточности. Следовательно, еще одним объектом исследований
молекулярной биологии и генетики в изучении патогенеза заболеваний
почек являются принципиально новые фармакологические методы
сдерживания прогрессирования почечной недостаточности. 69 69 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) Для того чтобы более полно охарактеризовать результаты
эпигенетической перестройки локальной внутрипочечной РАС для
функции почек, мы позволим себе кратко упомянуть широко известную
схему функционирования внутрипочечной РАС. Согласно существующим
представлениям, утвердившимся в мировой литературе, в почке основным
местом синтеза ренина являются специализированные клетки ЮГА. Субстрат ренина – ангиотензиноген, синтезируется в печени. Основные
регуляторные эффекты ангиотензина-II, образующегося в результате
поступательной конверсии ангиотензиногена в ангиотензин-I при участии
ренина, а затем и в ангиотензин-II
при участии ангиотензин-
превращающего фермента-1 (АПФ-1), сосредоточены на уровне
проксимального сегмента нефрона и кровеносных сосудов почки, главным
образом, через АТ1-популяцию рецепторов. Благодаря этим эффектам
ангиотензин-II осуществляет контроль кровяного давления, волемического
гомеостаза, параметров ионного и кслотно-основного гомеостаза
организма, а также принимает участие в ауторегуляции почечного
кровотока. Некоторые авторы не исключают, что в норме ангиотензиноген
может в небольших количествах синтезироваться нефроцитами
проксимального отдела нефрона (Kobori H. et al., 2013). Вместе с тем,
результаты экспериментальных исследований указывают, что главным
источником ангиотензиногена в норме является печень (Matsusaka T. et al.,
2012). Помимо АПФ-1, в почке достаточно высокие уровни активности
АПФ-2, отвечающего за образование ангиотензина-1-7, отвечающего за
механизмы отрицательной обратной связи к ангиотензину-II, хотя, строго
говоря, ангиотензин-1-7 антагонистом октапептида не является. В указанной схеме преобразования ангиотензиногена в ангиотензин-
II регуляторным ферментом является ренин. При этом, более ранние
источники литературы указывали роль процесса ацитилирования гистонов
в контроле прогрессирования заболеваний почек, сердца, легких (Bush
E.W., McKinsey T.A., 2010). В обсуждении роли преобразований
экспрессии компонентов ренин-ангиотензиновой системы почки в
процессах патогенеза почечной недостаточности, высказывается мысль о
том, что индукцию экспрессии ренина/проренина в канальцевом эпителии
следует рассматривать в качестве одного из ключевых событий (Prieto M.C. et al., 2013). Рассматривается роль усиления экспрессии ренина в
канальцевом отделе нефрона в патогенезе фиброза почки и
гипертонической болезни (Prieto M.C. et al., 2013; Gonzalez A.A., Prieto
M.C., 2015). В литературе анализируются возможные молекулярные 70 механизмы индукции экспрессии гена ренина в почках, включая
механизмы экспансии ренин-секретирующих клеток за пределы юкста-
гломерулярного аппарата (Sequeira Lopez M.L., Gomez R.A., 2010; Kurtz A.,
2012). В настоящее время экспансия экспрессии гена ренина (за счет
рекрутирования новых, ранее не синтезировавших ренин клеток)
рассматривается, как результат, в основном, деятельности эпигенетических
механизмов (Gomez R.A., 2017). С другой стороны, эпигенетические
системы контроля экспрессии гена ренина сохраняют свою актуальность не
только для тканей почки, но и для процессов кроветворения,
иммунокомпетентных клеток и т. д. (Gomez R.A., Sequeira-Lopez M.L.S.,
2018). 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) механизмы индукции экспрессии гена ренина в почках, включая
механизмы экспансии ренин-секретирующих клеток за пределы юкста-
гломерулярного аппарата (Sequeira Lopez M.L., Gomez R.A., 2010; Kurtz A.,
2012). В настоящее время экспансия экспрессии гена ренина (за счет
рекрутирования новых, ранее не синтезировавших ренин клеток)
рассматривается, как результат, в основном, деятельности эпигенетических
механизмов (Gomez R.A., 2017). С другой стороны, эпигенетические
системы контроля экспрессии гена ренина сохраняют свою актуальность не
только для тканей почки, но и для процессов кроветворения,
иммунокомпетентных клеток и т. д. (Gomez R.A., Sequeira-Lopez M.L.S.,
2018). )
Необходимо отметить, что, по мнению ряда авторов, ангиотензин-II
следует рассматривать в качестве одного из основных факторов,
способствующих прогрессированию почечной недостаточности, через
нарушение внутриорганной гемодинамики, стимуляцию фиброза органа,
активацию провоспалительных факторов, ограничение клеточного цикла
канальцевого эпителия и нарушение обменных процессов в нефроцитах
(Kobori H. et al., 2013). Указывается, что по мере прогрессирования
почечной недостаточности концентрации ангиотензина-II в тканях почки
могут существенно повышаться, на фоне незначительных изменений
уровня октапептида в системном кровотоке (Matsusaka T. et al., 2012;
Kobori H. et al., 2013). Привлекает внимание тот факт, что существенному
увеличению внутриренальной продукции ангиотензина-II, на фоне
прогрессирования почечной недостаточности, сопутствует отчетливый
прирост биосинтеза белков-компонентов РАС: ангиотензиногена,
проренина, АПФ-1 и АТ1-рецепторов ангиотензина-II
(основной
популяции рецепторов, отвечающих за большинство физиологических и
патофизиологических эффектов ангиотензина-II), не только в
проксимальных нефроцитах, но и в атипичных очагах активности РАС -
эпителии дистальных отделов нефрона (Kobori H. et al., 2013). Авторы
цитируемой публикации детально не обсуждают возможную роль
эпигенетических механизмов в перестройке внутрипочечной РАС по мере
нарастания патологических изменений ренальной паренхимы. Тем не
менее, сама логика излагаемых фактов подводит к этому вопросу. Постараемся выяснить, насколько обосновано такое предположение. Действительно
дальнейшие
исследования
показали
что Действительно, дальнейшие исследования показали, что
экспрессия компонентов РАС может регулироваться эпигенетическими
механизмами на разных этапах онтогенеза (Tain Y.-L. et al., 2017; Tain Y.L.,
Hsu C.N., 2017, Witasp A. et al., 2017). При этом, эпигенетическая
модуляция экспрессии компонентов РАС рассматривается, в качестве
одного из ведущих патогенетических механизмов целого ряда опасных 71 заболеваний (Tain Y.-L. et al., 2017). В частности, показано, что
эпигенетические изменения критически важны для понимания перехода
острой почечной недостаточности в хроническую форму (Rodríguez-Romo
R. et al., 2015). В литературе мы встречаем данные о том, что в условиях
экспериментальной модели фетального программирования подтверждено
участие эпигенетических факторов в регуляции уровней экспрессии АТ1
рецепторов ангиотензина-II (Bogdarina I. et al., 2007; Wu L. et al., 2016). 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) Важным является тот факт, что эпигенетические механизмы,
усиливая синтез компонентов РАС, создают условия для активации
внутриклеточных (аутокринных) эффектов ангиотензина-II, что, по
мнению некоторых авторов, является базовым патогенетическим
механизмом РАС-зависимых повреждений тканей почек и сердца (De Mello
W.C., 2015). В качестве иллюстрации к высказанному мнению можно
привести данные о том, что ацетилирование гистонов 3 (H3Ac), а также их
триметилирование (H3K4me3) и диметилирование (H3K9me2) может
способствовать высвобождению промотора гена АПФ-1 в почечной
паренхиме, обеспечивая биосинтез фермента (Liang M. et al., 2013). С
одной стороны, в соответствии с классическим представлением о
деятельности РАС, АПФ-1 в нашем организме присутствует в избытке и не
является лимитирующим фактором в процессах образования ангиотензина-
II. Но если оценивать упомянутый факт с позиций формирования
полноценно функционирующей внутриклеточной РАС, то он приобретает
совершенно иное значение (Abadir P.M. et al., 2012; Ellis B. et al., 2012). Действительно, по данным литературы, повышение экспрессии в тканях
почки гена АПФ-1 является маркером неблагоприятного течения
диабетической нефропатии (Thomas M.C., 2016). В дополнение к сказанному, можно привести сообщение группы
исследователей, выявивших в условиях диабетической нефропатии
усиление внутриклеточной продукции ангиотензиногена в проксимальных
нефроцитах,
обусловленное
ацетилированием
(H3K9)
и
триметилированием (H3K4me3) белка гистона-3 (Marumo T. et al., 2015). По мнению авторов, выявленный эффект может в равной степени
свидетельствовать, как о повышении функциональной нагрузки на данный
сегмент нефрона, так и о включении патофизиологических механизмов,
индуцирующих повреждение данной популяции клеток канальцевого
эпителия. Мнение о том, что повышение экспрессии ангиотензиногена в
проксимальных нефроцитах может рассматриваться в качестве маркера
прогрессирования почечной недостаточности, высказывают и другие
авторы (O'Leary R. et al., 2016; Bourgeois C.T. et al., 2017). Патофизиологические и эпигенетические механизмы этого феномена 72 требуют более глубокого исследования. Однако, установлено, что на
процессы эпигенетического контроля синтеза ангиотензиногена
проксимальными нефроцитами могут оказывать влияние такие факторы,
как интерферон-гамма (Satou R. et al., 2013), IL-6 (O'Leary R. et al., 2016) и
половые стероидные гормоны (Bourgeois C.T. et al., 2017). Наряду с этим, ангиотензин-II также обладает способностью
модулировать состояние экспрессии белков в тканях почки, стимулируя
повышение экспрессии АТ1 рецепторов и трансформирующего фактора
роста-бета1, на фоне угнетения АПФ-2 (Macconi D. et al., 2014). Эпигенетические механизмы, инициируемые на стадии острой
почечной недостаточности, могут рассматриваться в качестве фактора,
создающего предпосылки прогрессирования почечной недостаточности,
формируя неблагоприятный прогноз течения заболевания (Beckerman P. et
al., 2014; Tang J., Zhuang S., 2015; Lee-Son K., Jetton J.G., 2016). Эпигенетические механизмы, инициируемые на стадии острой
почечной недостаточности, могут рассматриваться в качестве фактора,
создающего предпосылки прогрессирования почечной недостаточности,
формируя неблагоприятный прогноз течения заболевания (Beckerman P. 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) et
al., 2014; Tang J., Zhuang S., 2015; Lee-Son K., Jetton J.G., 2016). al., 2014; Tang J., Zhuang S., 2015; Lee Son K., Jetton J.G., 2016). В контексте обсуждаемой темы уместно напомнить, что
фармакологические ингибиторы РАС (ингибиторы АПФ-1, антагонисты
АТ1 рецепторов и ингибиторы ренина) довольно широко и успешно
применяются, в том числе, при решении проблемы сдерживания
прогрессирующей почечной недостаточности. Применение данной группы
препаратов способствует ослаблению протеинурии, предотвращает
поражение канальцевого эпителия, содействует ограничению воспаления и
фиброза почки (Macconi D. et al., 2014). Поэтому, вполне логичным
является вопрос о возможном участии блокаторов РАС в нормализации
изменений, индуцированных эпигенетической перестройкой хроматина. Установлено, что в условиях острой почечной недостаточности
токсического генеза, ренопротекторные свойства
антагониста АТ1
рецепторов (лозартана) обусловлены сдерживанием, в том числе,
эпигенетических механизмов, индуцирующих десквамацию подоцитов и
усиление протеинурии (Hayashi K. et al., 2015). В частности, авторами
выявлено, что лозартан влияет на состояние метилирования промотора
гена белка нефрина. По некоторым данным, в условиях экспериментальной
модели диабетической нефропатии, лозартан может оказывать умеренный
благоприятный эффект на состояние эпигенетических механизмов в тканях
почки крыс (Reddy M.A. et al., 2013). В дальнейшем, в условиях ранее
примененной экспериментальной модели, авторы показали, что лозартан
эффективно блокирует эпигенетические механизмы (через регуляцию
процессов ацетилирования H3K9/14Ac) экспрессии генов, ответственных
за стимуляцию синтеза ингибитора активатора плазминогена-1 (PAI-1) и
моноцитарного хемоаттрактанта протеина-1 (MCP-1), являющихся
важными медиаторами повреждения тканей почек (Reddy M.A. et al., 2014). На основании полученных данных авторы цитируемой публикации делают ,
;
g
,
g
,
;
,
,
)
В контексте обсуждаемой темы уместно напомнить, что
фармакологические ингибиторы РАС (ингибиторы АПФ-1, антагонисты
АТ1 рецепторов и ингибиторы ренина) довольно широко и успешно
применяются, в том числе, при решении проблемы сдерживания
прогрессирующей почечной недостаточности. Применение данной группы
препаратов способствует ослаблению протеинурии, предотвращает
поражение канальцевого эпителия, содействует ограничению воспаления и
фиброза почки (Macconi D. et al., 2014). Поэтому, вполне логичным
является вопрос о возможном участии блокаторов РАС в нормализации
изменений, индуцированных эпигенетической перестройкой хроматина. Установлено, что в условиях острой почечной недостаточности
токсического генеза, ренопротекторные свойства
антагониста АТ1
рецепторов (лозартана) обусловлены сдерживанием, в том числе,
эпигенетических механизмов, индуцирующих десквамацию подоцитов и
усиление протеинурии (Hayashi K. et al., 2015). В частности, авторами
выявлено, что лозартан влияет на состояние метилирования промотора
гена белка нефрина. По некоторым данным, в условиях экспериментальной
модели диабетической нефропатии, лозартан может оказывать умеренный
благоприятный эффект на состояние эпигенетических механизмов в тканях
почки крыс (Reddy M.A. et al., 2013). 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) В дальнейшем, в условиях ранее
примененной экспериментальной модели, авторы показали, что лозартан
эффективно блокирует эпигенетические механизмы (через регуляцию
процессов ацетилирования H3K9/14Ac) экспрессии генов, ответственных
за стимуляцию синтеза ингибитора активатора плазминогена-1 (PAI-1) и
моноцитарного хемоаттрактанта протеина-1 (MCP-1), являющихся
важными медиаторами повреждения тканей почек (Reddy M.A. et al., 2014). На основании полученных данных авторы цитируемой публикации делают 73 вывод о том, что наиболее эффективная фармакологическая терапия
почечной недостаточности должна базироваться на комбинированном
применении ингибиторов РАС и специфических модуляторов
эпигенетических механизмов. Аналогичную точку зрения высказываются и
другие авторы, предполагая, что к наиболее благоприятным
терапевтическим результатам может привести сочетанное назначение
нефрологическим пациентам ингибитора АПФ-1 и ингибитора
деацетилазы гистонов (HDACI) (Zhong Y. et al., 2013). Признавая
эффективность лозартана в ограничении метилирования гистонов
Harshman L.A. и Zepeda-Orozco D. (2016) видят перспективность
клинического использования в нефрологической практике препаратов,
относящихся к группе ингибиторов HDACI. Наряду с этим, высказывается
мнение о роли микро-РНК в эпигенетических механизмах активации
локальной РАС почек при хронической почечной недостаточности (Witasp
A. et al., 2017). В литературе высказывается мнение о том, что изучение
эпигенетических механизмов функционирования внутриклеточной РАС
является фундаментальным направлением современной медицинской
науки, призванное решать наиболее актуальные практические задачи в
области нефрологии и заболеваний сердечно-сосудистой системы (De
Mello W.C., 2017). )
Таким образом, проведенный анализ данных литературы показал,
что
эпигенетические
аспекты
перестройки
внутрипочечной
(внутриорганной) РАС принципиально важны для понимания
патофизиологических механизмов нарушения деятельности почек,
сопряженных с усилением внутриклеточной продукции ангиотензина-II. Во-первых, эпигенетическая модификация хроматинового комплекса
приводит к появлению новых атипичных очагов интенсивной продукции
ангиотензина-II в канальцевом эпителии проксимального и дистального
отдела нефрона. Во-вторых, самодостаточная (содержащая все основные
компоненты) внутриклеточная РАС канальцевого эпителия переключается
на аутокринный и паракринный механизмы, с одной стороны, ослабляет
свою роль в физиологической регуляции гомеостатических функций почек. С другой стороны, активация внутриклеточной РАС все более нацелена на
патофизиологические механизмы усиления повреждения ткани через
нарушения энергетического обмена клетки (De Mello W.C., 2017). Кроме
того, активируемые эпигенетическими механизмами гены белков-
компонентов РАС, через повышение продукции ангиотензина-II, запускают
новый виток каскадного усиления ковалентной модификации хроматина,
где в качестве индуктора эпигенетических преобразований, напрямую или
опосредовано выступает сам ангиотензин-II. Об этом убедительно 74 свидетельствует эффективность применения блокаторов РАС в отношении
эпигенетической трансформации хроматина клеток почки. В-третьих, в
доступной нам литературе имеются единичные косвенные данные,
позволяющие судить о том, насколько эффективно проникают внутрь
клеток (в том числе в эпителий канальца) фармакологические ингибиторы
РАС (Foster D.R. et al., 2009). 3.1. РЕНИН-АНГИОТЕНЗИНОВАЯ СИСТЕМА (РАС) При том, что существует очевидная
потенциальная возможность с помощью ингибиторов РАС оказывать
влияние на внутриклеточные эффекты ангиотензина-II, нацеленные на
регуляцию экспрессии генов (da Silva Novaes A. et al., 2018). При этом мы
можем только предполагать характер возможного терапевтического
действия селективных ингибиторов на внутриклеточную РАС. В-
четвертых, данное направление исследований способствует разработке
принципиально новых фармакологических препаратов, способствующих
более эффективному решению практических задач не только в нефрологии,
но и в борьбе с заболеваниями сердечно-сосудистой системы и в области
онкологии. 75 75 3.2. МИНЕРАЛОКОРТИКОИДЫ. Анализ фармакологических способов контроля метаболизма
минералокортикоидов вовлечен в довольно широкий круг задач, далеко
выходящий за пределы изучения патогенеза почечной недостаточности
(Zhang D. et al., 2009; Welch A.K. et al., 2016; Bavishi C. et al., 2016;
Kawarazaki W., Fujita T., 2016; Azzam Z.S. et al., 2017). Однако, роль
альдостерона в патогенезе заболеваний почек, по-прежнему занимает одно
из центральных мест (Currie G. et al., 2016). Строго говоря, альдостерон синтезируется вне почки. Тем не менее,
мы посчитали возможным рассмотрение эпигенетических эффектов,
связанных с его метаболизмом в контексте анализируемого вопроса,
поскольку
его
физиологические,
патофизиологические
и
фармакологические аспекты тесно связаны с функционированием
локальной РАС коры надпочечников и внутрипочечной РАС (Feraille E.,
Dizin E., 2016; Kawarazaki W., Fujita T., 2016; Nehme A., Zibara K., 2017). Возможно, такое объединение может иметь и более обоснованный
аргументы, однако, данный вопрос требует дополнительного изучения (De
Mello W.C., 2017). Тем не менее, уже известные факты, широко
применяемые в практической медицине (Bavishi C. et al., 2016; Currie G. et
al., 2016), дают нам право дополнить выше изложенные аргументы
сведениями о роли эпигенетических механизмов в патофизиологии
альдостерона и РААС. р
Позволим себе еще одно краткое замечание. В процессе филогенеза
появление у амниот минералокортикоидов произошло относительно
недавно – в связи с выходом позвоночных животных на сушу. В то время,
как у низших позвоночных (анамний) функцию минералокортикоидов
выполнял кортизол (Dolomatov S.I. et al., 2012). Вероятно, поэтому мы
наблюдаем интерференцию эффектов альдостерона и глюкокортикоидов на
процессы реабсорбции натрия в дистальном отделе нефрона человека
(Feraille E., Dizin E., 2016; Nehme A., Zibara K., 2017). В данном случае
упоминание о минералокортикоидной функции глюкокортикоидов следует
рассматривать, как попытку более полно оценить обсуждаемые процессы. р
р
у
у
р
Возможно, рассмотрение регуляторных эффектов альдостерона,
необходимо начать с того, что наиболее важными стимулами
интенсивности его секреции в коре надпочечников являются: повышение
содержания ионов калия во внеклеточной (внутрисосудистой) жидкости и
ангиотензина-II, образующийся в локальной (внутриорганной) РАС
надпочечников и почек (Feraille E., Dizin E., 2016; Kawarazaki W., Fujita T.,
2016; Nehme A., Zibara K., 2017). Поскольку стимулирующее действие
ангиотензина-II на уровень секреции альдостерона реализуется через АТ1- 76 популяцию рецепторов, уместно напомнить, что ранее была установлена
роль эпигенетических механизмов в управлении экспрессией АТ1
рецепторов, в том числе и в корковом веществе надпочечников (Bogdarina
I. et al., 2007; Liang M. et al., 2013). Кроме того, показано, что механизмы фетального
программирования, обусловленные даже непродолжительным повышением
кортизола в крови матери могут усиливать экспрессию их рецепторов у
плода (Liang M. et al., 2013; Tain Y.L., Hsu C.N., 2017). 3.2. МИНЕРАЛОКОРТИКОИДЫ. По мнению авторов
цитируемых публикаций, такой механизм может способствовать
неадекватной стимуляции реабсорбции натрия в зрелом возрасте, приводя
к системным нарушениям параметров гемодинамики. Кроме того, авторы
отмечают, что активация реабсорбции натрия в дистальном отделе нефрона
может осуществляться и за счет триметилирования of H3K36,
сопровождающегося подавлением экспрессии гена 11β-гидроксистероид
дегидрогеназы-2,
отвечающей
за
метаболический
клиренс
глюкокортикоидов. Необходимо подчеркнуть, что патофизиологические
механизмы альдостерона в почках непосредственно сопряжены со
стимуляцией фиброгенеза в тканях органа, повреждением подоцитов и
нарастанием протеинурии (Kawarazaki W., Fujita T., 2016). В современной литературе мы наблюдаем повышение интереса к
эпигенетическим механизмам перестройки работы почки, связанных с
изменением экспрессии транспортных систем натрия, калия и хлора в
различных сегментах нефрона (Tain Y.L., Hsu C.N., 2017). Одно из
центральных мест этого направления исследований прочно занимает
эпителиальный натриевый канал (ENaC) дистального отдела нефрона
(Duarte J.D. et al., 2012; Kone B.C., 2013; Yu Z. et al., 2013). В цитируемых
источниках сообщается, что альдостерон стимулирует транскрипцию гена
белка альфа-субъединицы EnaC (αENaC) через активацию фермента
глюкокортикоид-индуцируемую киназу-1, подавляющую активность Dot1a
(метилтрансферазу белков-гистонов H3K79), транскрипционного фактора
Af9 и гистоновой деацетилазы Sirt1, изменяя активность комплекса
Dot1/Af9. Кроме того, в литературе имеются данные о том, что
индуцированная альдостероном модификация хроматина может
способствовать усилению экспрессии гена эндотелина-1 в соединительных
трубочках внутренней медуллы (Welch A.K. et al., 2016). Поскольку рецепторам минералокортикоидов отводится важная
роль в реализации эпигенетических эффектов альдостерона, могут
представлять интерес данные о том, какова роль данной популяции
рецепторов в регуляции экспрессии генов, чувствительных к влиянию
альдостерона (Ueda K. et al., 2014). 77 Привлекают внимание сообщения о том, что эпигенетические
изменения в системе РААС могут принципиально нарушать механизмы
стимуляции секреции альдостерона в корковом веществе надпочечников,
ослабляя регуляторную роль внутриорганной РАС почек и надпочечников,
выводя на первые позиции совершенно иные факторы (например, лептин),
непосредственно не связанные с функциональным состоянием почек и не
привязанные к параметрам водно-солевого обмена (Kawarazaki W., Fujita
T., 2016). Таким образом, проведенный анализ данных литературы показал,
что эпигенетические механизмы перестройки метаболизма альдостерона
являются важным фактором в патогенезе ренальных дисфункций и
патологических нарушений системной гемодинамики. Установлено, что
эпигенетические механизмы затрагивают: систему регуляции метаболизма
неполовых стероидов; контролирующих экспрессию транспортных белков
дистального отдела нефрона; секрецию физиологически активных
пептидов в канальцевом отделе нефрона. Кроме того, есть основания
предполагать, что процессы регулирования секреции альдостерона также
могут подвергаться эпигенетическим изменениям, приводя к неадекватной
стимуляции продукции гормона. 3.2. МИНЕРАЛОКОРТИКОИДЫ. Возможно, совокупность выявленных
закономерностей позволяет некоторым авторам утверждать, что вызванная
эпигенетической перестройкой хроматина неограниченная активация РААС
и взаимное усиление патофизиологических эффектов ангиотензина-II и
альдостерона является одним из базовых патогенетических механизмов
хронических заболеваний почек и органов сердечно-сосудистой системы
(De Mello W.C., 2017). 78 78 3.3. ТРАНСФОРМИРУЮЩИЙ ФАКТОР РОСТА-бета1 Согласно данным литературы, трансформирующий фактор роста-
бета1 (ТФР-бета1) принадлежит к суперсемейству цитокинов, в состав
которых, помимо ТФР-бета, входит большое количество белков, например,
ВМР, в норме имеющих важное значение для цитодифференцировки
тканей и процессов заживления ран (Shi M. et al., 2011). В стимуляции
внутриренального синтеза ТФР-бета1важную роль играет ангиотензин-II
через АТ1 популяцию рецепторов (Reddy M.A. et al., 2014). Между тем,
авторы цитируемого источника отмечают, что антагонисты АТ1 рецепторов
и блокаторы АПФ-1 оказывают умеренное благоприятное воздействие на
процессы фиброза органа при хронической почечной недостаточности,
поскольку существуют РАС-независимые пути индукции ТФР-бета1. Известно, что ТФР-бета1 и ТФР-бета3 является ключевым фактором
стимуляции фиброгенеза ткани почки в условиях хронической почечной
недостаточности (Wing M.R. et al., 2013). Обнаружено, что патологические
нарушения почек в условиях экспериментальных моделей острой почечной
недостаточности сопровождаются достаточно быстрым приростом
продукции ТФР-бета1 в тканях почки, в том числе, благодаря активации
эпигенетических механизмов (Zager R.A. et al., 2011), нарушая нормальное
течение репаративных процессов в почке (Bonventre J.V., Yang L., 2011). В
экспериментальных условиях острой почечной недостаточности in vivo и в
моделировании острого токсического воздействия на культивируемые
проксимальные нефроциты было установлено, что стимуляция
метилирования Н3 (H3K4mе3) предшествует резкому повышению уровня
мРНК ТФР-бета1 в ткани (Zager R.A., Johnson A.C.M., 2010). Результаты
экспериментальных исследований подтверждают, что эпигенетическая
активация гена ТФР-бета1 происходит в условиях острой почечной
недостаточности, способствуя хронизации заболеваний почек (Sun G. et al.,
2014). Поскольку ТФР-бета1 может участвовать в метастазировании
злокачественных опухолей, является одним из основных индукторов
фиброза почек, печени, легких, кожи, проблеме клинического применения
анти-ТФР-бета терапии, основанной, в том числе, на эпигенетических
механизмах, уделяется значительное внимание, как наиболее
перспективному направлению в лечении целого ряда опасных заболеваний
(Zeisberg
M., Zeisberg E.M., 2015). В частности, анализируется
эффективность различных способов подавления патогенетических ТФР-
бета1-заисимых механизмов через селективное ингибирование популяции
II-типа рецепторов цитокина (Doi S. et al., 2011), применение
антисывороток ТФР-бета1 протеина (Zeisberg M., Zeisberg E.M., 2015), 79 использование селективных блокаторов активности деацетилаз гистоновых
белков (HDAC) (Guo W. et al., 2009). Хотя, по мнению некоторых авторов, в
качестве основной мишени специфических блокаторов деацетилаз
гистонов следует рассматривать фермент HDAC класса I, которая,
возможно, критически важна для стимуляции ТФР-бета1-зависимого
фиброза почек (Liu N. et al., 2013). Также, некоторыми авторами
высказывается мнение о целесообразности фармакологической коррекции
баланса активности феерментов ацетилтрансфераз гистонов (HATs) и
ферментов деацетилаз гистонов (HDACs) (Yuan H. et al., 2013). 3.3. ТРАНСФОРМИРУЮЩИЙ ФАКТОР РОСТА-бета1 Необходимо
отметить, что в литературе представлены обзоры, содержащие достаточно
глубокий и всесторонний анализ возможных системных терапевтических
эффектов ингибиторов энзиматической активности HDACs, нацеленных на
предотвращение фиброза внутренних органов, включая почки, а также
других модуляторов эпигенетических изменений в ренальной паренхиме
(Van Beneden K. et al., 2013; Tang J., Zhuang S., 2015). Приводятся аргументы в пользу терапевтической эффективности
ингибиторов метилирования в развитии
ТФР-бета1-зависимого
фиброгенеза почки (Bechtel W. et al., 2010). При этом, в качестве наиболее
актуальной мишени перспективных препаратов предлагается фермент
метилтрансфераза 7/9 (SET7/9), осуществляющая монометилирование
остатка лизина 4 белка-гистона H3 (H3K4me1) (Sasaki K. et al., 2016). (
) (
,
)
На том основании, что некоторые виды микроРНК (в частности, miR-
29b) обладают способностью подавлять некоторые просклерозирующие
эффекты ТФР-бета1, предполагается, данное направление также может
быть в перспективе применено для сдерживания прогрессирующей
почечной недостаточности (Wing M.R. et al., 2013). Установлено, что
некоторые микроРНК (микро РНК-21 и микро ТНК-192), могут
рассматриваться в качестве индукторов ТФР-бета1-зависимого
тубулоинтерстициального фиброза и гломерулосклероза (Liu R. et al.,
2015). Стимулированное ТФР-бета1 повышение транскрипции микро РНК-
192 подтверждено в опытах in vitro в культуре клеток (человека и мыши)
мезангиума, подоцитов, эндотелиоцитов и канальцевого эпителия (Kato M. et al., 2013). Авторам также удалось установить, что стимуляция ТФР-бета1
транскрипции микро РНК-192
зависит от нескольких участков
ацетилирования гистона Н3 (H3K9, H3K14 и H3K27). Кроме того, авторами
данной публикации высказывается мысль о том, что микро РНК-192
принадлежит особая роль в каскадном усилении просклерозирующих
эффектов ТФР-бета1 через активацию транскрипции микро РНК-200b и
микро РНК-200c, повышающих экспрессию генов коллагена-1альфа2
(COL1A2), КОЛЛАГЕНА-4АЛЬФА1(COL4A1) и самого ТФР-бета1 (TGF-β1). С
другой стороны, известно, что
ТФР-бета1 через Smad3-протеин, 80 стимулирует образование микро РНК-21, активирующей, в свою очередь,
экспрессию генов collagen I и fibronectin, а также способствующей
повышению уровня α-SMA в почке (Wing M.R. et al., 2013). Показано, что ТФР-бета1 через активацию фермента метилирования
гистонов H3K4-метилтрансферазы SET7/9, повышает экспрессию генов,
запускающих, процессы фиброгенеза в почке. Напротив, подавление
SET7/9 ингибирует экспрессию индуцируемых ТФР-бета1 генов фиброза
(Reddy M.A, Natarajan R., 2015; Dressler G.R., Patel S.R., 2015; Hilliard S.A.,
El-Dahr S.S., 2016). Возможно, медиаторами эффекта ТФР-бета1 в
отношении активности SET7/9 являются продукты реакции,
катализируемой ферментом 12/15-липоксигеназы (Yuan H. et al., 2016). Наряду с этим, сообщается о том, что ТФР-бета1-зависимая активация
фиброгенеза осуществляется через систему внутриклеточной передачи
сигнала Smad-протеинами (Reddy M.A, Natarajan R., 2015). Авторы
указывают, что, например, Smad2-протеин причастен к стимуляции
ацетилирования молекулы гистона Н3 (H3K9/14Ac). 3.3. ТРАНСФОРМИРУЮЩИЙ ФАКТОР РОСТА-бета1 Наряду с ранее названными эпигенетическими изменениями,
отмечается, что метилирование гистона Н3 (H3K9me2 и H3K9me3)
является важным механизмом в регуляции экспрессии генов коллагена-
1альфа1 (Col1α1) и ингибитора активатора плазминогена (PAI-1) (Reddy
M.A. et al., 2013; Sun G. et al., 2014). Одним из базовых патогенетических
механизмов тубулоинтерстициальных повреждений канальцевого отдела
нефрона является эпителиально-мезенхимальная трансформация,
маркером интенсивности которого служит экспрессия α-актина (αSMA). В
связи с этим представляет интерес сообщение о том, что в условиях
экспериментальной модели односторонней обструкции мочеточника у
мышей TGF-β1 не оказывал существенного влияния на состояние H3K9Ac
в проксимальных нефроцитах и миофибробластах. Наряду с этим, цитокин
приводил к перераспределению метки H3K9Me3 в хроматине ядра
фибробластов, что коррелировало с увеличением экспрессии α-SMA
(Hewitson T.D. et al., 2017). Таким образом, обзор литературы показал, что эпигенетические
эффекты TGF-β1 оказывают весьма значительное влияние на процессы
фиброгенеза в тканях почек, затрагивая, фактически, все известные
механизмы импринтинга: метилирование и ацетилирование гистоновых
белков, а также перестройку экспрессии некоторых специфических микро
РНК. Следует отметить, что эпигенетические механизмы, инициируемые
TGF-β1 в ренальной паренхиме, не только непосредственно участвуют в
реализации просклерозирующего эффекта цитокина, но и способствуют
резкому усилению TGF-β1-зависимых патогенетических механизмов
ремоделирования ренальной паренхимы. При этом, ингибирование TGF- 81 β1-зависимой модификации хроматина способствует сдерживанию
патологических изменений деятельности почек. Что, с одной стороны,
доказывает важную патогенетическую роль TGF-β1 в хронизации и
прогрессировании почечной недостаточности. С другой стороны, это
открывает новые перспективы использования селективных модуляторов
эпигенетических процессов в практической медицине, что подтверждается
сведениями о готовности их применения в доклинических испытаниях
(Van Beneden K. et al., 2013). β1-зависимой модификации хроматина способствует сдерживанию
патологических изменений деятельности почек. Что, с одной стороны,
доказывает важную патогенетическую роль TGF-β1 в хронизации и
прогрессировании почечной недостаточности. С другой стороны, это
открывает новые перспективы использования селективных модуляторов
эпигенетических процессов в практической медицине, что подтверждается
сведениями о готовности их применения в доклинических испытаниях
(Van Beneden K. et al., 2013). 82 82 3.4. МОЛЕКУЛА ОКСИДА АЗОТА (NO) По данным литературы, эпигенетические механизмы выполняют
очень важную функцию в регуляции аргинин-зависимого пути синтеза NO
в системе изоформ NO-синтаз: эндотелиальной (nNOS - NOS-1),
индуцибельной (iNOS - NOS-2), и нейрональной (eNOS - NOS-3). Некоторые авторы выделяют еще одну изоформу – митохондриальную
mtNOS. Имеются данные о том, что гипоксия, один из наиболее мощных
активаторов эпигенетической модификации хроматина, способствует
изменению экспрессии генов различных изоформ фермента NOS (Shirodkar
A.V., Marsden P.A., 2011). Согласно данным цитируемого обзора, ишемия
может сопровождаться репрессией гена eNOS в эндотелиоцитах, на фоне
активации транскрипции всех трех изоформ NOS в неоинтиме, включая
транскрипцию гена eNOS в мышечных волокнах стенки кровеносных
сосудов. Авторы отмечают, что добавление к культивируемым клеткам
гладкой мускулатуры сосудистой стенки ингибитора метилтрансферазы
ДНК (5-azacytidine), а также как ингибитора HDAC (Trichostatin A),
приводило к стимуляции транскрипцию гена eNOS в этих клетках, также,
способствуя увеличению мРНК eNOS. В исследованиях in vitro на культуре
проангиогенных клеток (early EPCs) и мезангиобластов было установлено,
что добавление в среду только 3-deazaneplanocin A (DZNep), ингибитора
триметилирования H3K27, не оказывало существенного влияния на
экспрессию гена eNOS, тогда, как сочетанное воздействие на клетки
DZNep и ингибитора гистоновой дезацетилазы Trichostatin A (TSA)
увеличивает экспрессию eNOS (Ohtani K. et al., 2011). Результаты
клинических наблюдений, подтверждая роль метилирования и
ацетилирования гистонов в регуляции экспресси гена eNOS, также
акцентируют внимание на процессах метилирования ДНК (Kheirandish-
Gozal L. et al., 2013). Возможно, анализ процесса метилирования
промотора гена eNOS представляет интерес в составлении прогноза рисков
патологических нарушений некоторых показателей минерального обмена
человека (Harvey N.C. et al., 2012). Эпигенетические механизмы контроля
экспрессия eNOS в эндотелии кровеносных сосудов почки критически
важны в процессе органогенеза, а также адаптации почки к гипоксии и
изменениям параметров внутрипочечной гемодинамики (Jamal A. et al.,
2012). По данным источника, эндотелиоциты могут не проявлять
чувствительность к действию цитокинов, стимулирующих экспрессию
iNOS, в том случае, если промотор этого гена обильно метилирован,
В норме в тканях почки преимущественно представлены nNOS
(NOS-1), в основном в области macula densa, а также eNOS (NOS-3) в В норме в тканях почки преимущественно представлены nNOS
(NOS-1), в основном в области macula densa, а также eNOS (NOS-3) в
эндотелиоцитах и в канальцевом эпителии. Известно, что NO участвует в 83 регуляции ренальной гемодинамики, канальцевого транспорта натрия,
регуляции величины скорости клубочковой фильтрации. Является важным
фактором контроля тубуло-гломерулярной обратной связи, регулятором
агрегатного состояния крови и процессов воспаления. Однако, динамика
изменения внутрипочечной продукции NO не всегда совпадает с уровнем
экспрессии генов NO-синтаз. 3.4. МОЛЕКУЛА ОКСИДА АЗОТА (NO) Так, как интенсивность внутрипочечного
синтеза NO, по мере прогрессирования почечной недостаточности, может
снижаться в результате поражения сосудистого русла, фиброза коркового
слоя почек, изменения метаболизма субстрата (L-Arginine), повышения
концентрации эндогенного блокатора NO-синтаз (асимметричного
диметиларгинина - ADMA) и доступности кофакторов NOS-синтазных
энзиматических комплексов. Установлено, что прогрессирование почечной недостаточности
сопровождается снижением внутрипочечной продукции NO, что
коррелирует с интенсивностью фиброгенеза в почке (Schmidt Dellamea B. et al., 2014). Вместе с тем, авторы отмечают роль некоторых биологически
активных веществ (инсулина, фактора некроза опухоли-альфа,
ангиотензина-II) в регуляции экспрессии генов NO-синтаз. Возможно,
стимулируемая инсулином избыточная экспрессия гена eNOS (NOS-3)
является одним из важнейших патогенетических механизмов
прогрессирования диабетической нефропатии, поскольку в условиях
экспериментальной модели, введение животным vorinostat (неселективного
ингибитора гистон-деацетилаз класса I и класса II понижало экспрессию
данного гена, что способствовало ограничению протеинурии о накопления
белков внеклеточного матрикса мезангиальными клетками (Advani A. et al.,
2011). Сообщается, что, во-первых, в условиях экспериментальной
патологии почек избыточная внутрипочечная продукция оксида азота
является важным патогенетическим фактором развития гломерулопаии. Во-
вторых, Trichostatin A (TSA) - ингибитор гистон-деацетилаз может
способствовать нормализации избыточной продукции NO, как клетками
мезангиума, так и индуцибельной iNOS, активируемой некоторыми
провоспалительными цитокинами (например, IL-1β) (Van Beneden K. et al.,
2013). )
Возможно, ингибиторы деацетилаз гистонов следует рассматривать в
качестве перспективной группы фармакологических препаратов,
позволяющих сдерживать целый ряд NO-зависимых патогенетических
механизмов прогрессирования почечной недостаточности: воспалительный
и
просклерозирующий
компоненты
тубуло-интерстициальных
повреждений, блокировать активацию фибробластов почки и апоптоз
канальцевого эпителия почки (Jamal A. et al., 2012). Кроме того,
ингибиторы деацетилаз гистонов, снижая в почке экспрессию генов iNOS и 84 eNOS, способствуют восстановлению функции почек на фоне ограничения
образования α-SMA, коллагена I, фибронектина, ТФР бета1, а также
ограничивают апоптоз в условиях диабетической нефропати (Khan S., Jena
G., 2014). Установлено, что гипоксия является одним из наиболее мощных
факторов, регулирующих экспрессию гена NOS3 эндотелиальной NO-
синтазы эндотелиоцитов через снижение и ацетилирования гистоновых
белков, и метилирования остатков лизина 4 в гистоне H3 (Fish J.E. et al.,
2010). Высказывается мнение о том, что, спровоцированное гипоксией
усиление экспрессии индуцибельной iNOS, также может оказывать
нефропротекторный эффект при реперфузионных повреждениях органа
(Bonventre J.V., Yang L., 2012). Тем не менее, продолжительная стимуляция
экспрессии iNOS при токсическом поражении почки рассматривается в
качестве неблагоприятного фактора, усугубляющего течение заболевания
(Sattarinezhad E. et al., 2017). Необходимо признать, что проблема
эпигенетической перестройки внутрипочечной системы оксида азота
достаточно многогранна, в частности, в литературе представлены работы,
посвященные анализу изменений баланса некоторых гуморальных
регуляторов деятельности почки (NO, ангиотензина-II, производных
арахидоновой кислоты) в условиях фетального программирования (Tain Y.-
L., Joles J.A., 2016). 3.4. МОЛЕКУЛА ОКСИДА АЗОТА (NO) Обсуждаемые вопросы постоянно находятся в поле
зрения современной науки, о чем свидетельствуют обзорные публикации,
содержащие сведения о фундаментальных эпигенетических механизмах
регуляции системы оксида азота (Vasudevan D. et al., 2016; Socco S. et al.,
2017). Однако, нельзя исключать возможности наличия
органоспецифических, в том числе в ренальной паренхиме, механизмов
контроля экспрессии различных изоформ NOS, в частности, iNOS и eNOS
(Cerkezkayabekir A. et al., 2017). Суммируя изложенные факты, можно сделать вывод о том, что
эпигенетическая трансформация системы оксида азота является важным
компонентом патогенетических механизмов нарушения деятельности
почек. Имеющиеся в литературе факты указывают, что инициирование
данной перестройки, например, гипоксией тканей или под воздействием
гормонов и цитокинов, может осуществляться на самых ранних этапах
заболевания почек. Кроме того, ренальная система NOS подвергается
радикальной модификации по мере прогрессирования почечной
недостаточности, что проявляется в уменьшении экспрессии nNOS в
корковом веществе почек, неуклонном снижении экспрессии eNOS в
эндотелиоцитах и появлении атипичной локализации eNOS в мышечном
слое сосудистой стенки, стимуляции экспрессии iNOS. у
у
р
С одной стороны, известно, что NOS почек (главным образом nNOS)
принимают участие в регуляции активности внутриорганной РАС, а 85 молекула NO – один из основных антагонистов ренотропных эффектов
ангиотензина-II на сосудистом и канальцевом уровне (Chappell M.C., 2012;
Thoonen R. et al., 2013). Следовательно, ослабление внутриорганных
регуляторных влияний конститутивных NO-синтаз может являться одной
из причин неограниченной активации внутрипочечной РАС и ТФР-бета в
условиях прогрессирования почечной недостаточности (Macconi D. et al.,
2014). Более того, по мнению некоторых авторов именно активацию
митохондриальной NOS допустимо рассматривать в ряду основных
патогенетических
механизмов
эпигенетической
активации
внутриклеточной РАС (De Mello W.C., 2017). С другой стороны, неадекватная активация экспрессии NOS (eNOS и iNOS)
на определенных этапах течения заболевания, в силу специфики физико-
химических свойств конечного продукта – молекулы оксида азота и
особенностей функционирования NO-синтазных комплексов, может
служить источником активных форм кислорода и азота, внося, таким
образом, существенный вклад в усиление патологических процессов
(Advani A. et al., 2011; Sattarinezhad E. et al., 2017; Tain Y.-L. et al., 2017). Помимо этого, обсуждение эпигенетических перестроек аргинин-
зависимого пути образования NO не исчерпывает всей темы метаболизма
оксида азота в почках в норме и при патологии. Известно, что существует
механизм ресинтеза молекулы NO, использующий в качестве субстрата
химически стабильные продукты окисления NO (нитриты, нитраты и др.) и
контролируемый такими белками, как гемоглобин или цитохром Р450. В
современных обзорах мы встречаем упоминание аргинин-независимого
синтеза NO в связи с эпигенетической модуляций системы оксида азота
(Vasudevan D. et al., 2016; Socco S. et al., 2017). 3.4. МОЛЕКУЛА ОКСИДА АЗОТА (NO) Однако, данные механизмы
имеют определенную специфику, характерную для обменных и
транспортных процессов, протекающих в тканях почек. молекула NO – один из основных антагонистов ренотропных эффектов
ангиотензина-II на сосудистом и канальцевом уровне (Chappell M.C., 2012;
Thoonen R. et al., 2013). Следовательно, ослабление внутриорганных
регуляторных влияний конститутивных NO-синтаз может являться одной
из причин неограниченной активации внутрипочечной РАС и ТФР-бета в
условиях прогрессирования почечной недостаточности (Macconi D. et al.,
2014). Более того, по мнению некоторых авторов именно активацию
митохондриальной NOS допустимо рассматривать в ряду основных
патогенетических
механизмов
эпигенетической
активации
внутриклеточной РАС (De Mello W.C., 2017). С другой стороны, неадекватная активация экспрессии NOS (eNOS и iNOS)
на определенных этапах течения заболевания, в силу специфики физико-
химических свойств конечного продукта – молекулы оксида азота и
особенностей функционирования NO-синтазных комплексов, может
служить источником активных форм кислорода и азота, внося, таким
образом, существенный вклад в усиление патологических процессов
(Advani A. et al., 2011; Sattarinezhad E. et al., 2017; Tain Y.-L. et al., 2017). молекула NO – один из основных антагонистов ренотропных эффектов
ангиотензина-II на сосудистом и канальцевом уровне (Chappell M.C., 2012;
Thoonen R. et al., 2013). Следовательно, ослабление внутриорганных
регуляторных влияний конститутивных NO-синтаз может являться одной
из причин неограниченной активации внутрипочечной РАС и ТФР-бета в
условиях прогрессирования почечной недостаточности (Macconi D. et al.,
2014). Более того, по мнению некоторых авторов именно активацию
митохондриальной NOS допустимо рассматривать в ряду основных
патогенетических
механизмов
эпигенетической
активации
внутриклеточной РАС (De Mello W.C., 2017). у р
(
,
)
С другой стороны, неадекватная активация экспрессии NOS (eNOS и iNOS)
на определенных этапах течения заболевания, в силу специфики физико-
химических свойств конечного продукта – молекулы оксида азота и
особенностей функционирования NO-синтазных комплексов, может
служить источником активных форм кислорода и азота, внося, таким
образом, существенный вклад в усиление патологических процессов
(Advani A. et al., 2011; Sattarinezhad E. et al., 2017; Tain Y.-L. et al., 2017). Помимо этого, обсуждение эпигенетических перестроек аргинин-
зависимого пути образования NO не исчерпывает всей темы метаболизма
оксида азота в почках в норме и при патологии. Известно, что существует
механизм ресинтеза молекулы NO, использующий в качестве субстрата
химически стабильные продукты окисления NO (нитриты, нитраты и др.) и
контролируемый такими белками, как гемоглобин или цитохром Р450. В
современных обзорах мы встречаем упоминание аргинин-независимого
синтеза NO в связи с эпигенетической модуляций системы оксида азота
(Vasudevan D. et al., 2016; Socco S. et al., 2017). Однако, данные механизмы
имеют определенную специфику, характерную для обменных и
транспортных процессов, протекающих в тканях почек. 86 3.5. ПАТОФИЗИОЛОГИЧЕСКОЕ ЗНАЧЕНИЕ ЭПИГЕНЕТИЧЕСКОЙ
ТРАНСФОРМАЦИИ СИСТЕМЫ ВНУТРИОРГАННОЙ
ГУМОРАЛЬНОЙ РЕГУЛЯЦИИ ДЕЯТЕЛЬНОСТИ ПОЧЕК Результаты проведенного обзора позволяют сделать вывод о том, что
эпигенетические механизмы вносят очень важный вклад в перестройку
гуморальных систем регуляции деятельности почек в условиях почечной
недостаточности, во многом способствуя прогрессирующему сокращению
популяции действующих нефронов, непосредственно создавая
предпосылки неблагоприятного течения заболевания. При этом, можно выделить несколько общих тенденций, характерных
для эпигенетической трансформации внутриренального синтеза и
метаболизма физиологически активных веществ. Во-первых,
формирование атипичных очагов их продукции, что наиболее явно
присутствует в процессах перестройки систем РАС и оксида азота. Во-
вторых, гуморальные факторы, осуществляющие в неповрежденной почке
координацию физиологических систем контроля гомеостатической
деятельности почек, по мере усиления некоторых эпигенетических
изменений, все более утрачивают функции регуляции гомеостаза и
переключаются на патофизиологический путь индукции ренальных
дисфункций и стимуляции прогрессирования почечной недостаточности. В-третьих, эпигенетические изменения, затрагивающие гены белков,
выполняющих ключевые функции в синтезе и метаболизме гуморальных
факторов регуляции функций почек, могут выполнять роль пускового
механизма становления и прогрессирования почечной недостаточности. В
дальнейшем, неограниченный синтез этих молекул белковой и небелковой
природы приводит к триггерному усилению процесса, в том числе, опять
же с привлечением эпигенетической перестройки хроматина. Следовательно, с одной стороны, гены белков, управляющих продукцией
гуморальных факторов регуляции функций почек, являются объектом
импринтинга. С другой стороны, стимулированная импринтингом
модуляция синтеза гуморальных факторов, на последующем витке
спирали, содействует дальнейшему углублению эпигенетической
модификации хроматина и усилению роста их образования. Наиболее
отчетливо указанная закономерность прослеживается в неограниченной
активации РААС и системы ТФР-бета почек. В-четвертых, анализируя
спровоцированную импринтингом трансформацию баланса ренотропных
регуляторных эффектов гуморальных факторов, можно сделать вывод о
том, что в этих условиях наблюдается безмерное нарастание
вазотонического, просклерозирующего и провоспалительного потенциала в
результате неограниченной активации РААС и системы ТФР-бета. На этом 87 фоне происходит неуклонное сокращение регуляторных возможностей
оппозиционного вектора контроля, представленного, в частности, системой
оксида азота, в первую очередь, конститутивными изоформами eNOS и
nNOS. В-пятых, раскрытие эпигенетических процессов в становлении и
прогрессировании нефропатий различного генеза не только способствует
созданию более прочной теоретической основы патогенеза почечной
недостаточности, но и открывает перспективы к разработке
принципиально новых фармакологических способов коррекции функции
почек. К сожалению, формат рукописи не позволяет уделить данной теме
того внимания, которое она безусловно заслуживает. Между тем, важный
практический интерес представляют результаты исследования роли
эпигенетических механизмов в модуляции систем аргинин-вазопрессина
(Murgatroyd C., 2014; Lesse A. et al., 2017), порообразующих белков
аквапоринов (Park E.-J., Kwon T.-H., 2015; MacManes M.D., 2017) и
атриального натрийуретического пептида (Sergeeva I.A. et al., 2014; 2016). Равно, как и ренотропные эпигенетические эффекты инсулина (Kumar S. et
al., 2016; Shiels P.G. 3.5. ПАТОФИЗИОЛОГИЧЕСКОЕ ЗНАЧЕНИЕ ЭПИГЕНЕТИЧЕСКОЙ
ТРАНСФОРМАЦИИ СИСТЕМЫ ВНУТРИОРГАННОЙ
ГУМОРАЛЬНОЙ РЕГУЛЯЦИИ ДЕЯТЕЛЬНОСТИ ПОЧЕК et al., 2017), факторов, индуцируемых гипоксией (HIFs
семейство протеинов) (Perez-Perri J.I. et al., 2011; Liu J. et al., 2017),
продуктов метаболизма арахидоновой кислоты (Yuan H. et al., 2016) и
тироксина (Liu X. et al., 2014; Re A. et al., 2016) могут иметь самое
непосредственное отношение к обсуждаемым вопросам. По нашему мнению, данная тема может представлять интерес для
понимания особенностей патофизиологических механизмов нарушения
функции почек. Поскольку, эпигенетическая модификация хроматина
играет принципиально важную роль в нарушении баланса
внутрипочечного метаболизма гуморальных факторов регуляции функции
почек. Средства фармакологической коррекции, разработанные в
соответствии с пониманием механизмов импринтинга в значительной
степени, могут способствовать в сдерживании хронизации и
прогрессирования почечной недостаточности. Напротив, отказ от более
современной стратегии сдерживания прогрессирующих нарушений
ренальной паренхимы, может иметь последствия в форме изменений
процессов синтеза гуморальных факторов под влиянием эпигенетических
механизмов, оказыващих дальнейшее влияние на ковалентную
модификацию хроматина, а также усиливающих патофизиологические
механизмы повреждения ренальной паренхимы. 88 СПИСОК
ЛИТЕРАТУРЫ
К
ГЛАВЕ
«ФАКТОРЫ
ВНУТРИОРГАННОЙ
ГУМОРАЛЬНОЙ
РЕГУЛЯЦИИ
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doi: 10.1007/s12020-015-0751-2 103 104 104 ГЛАВА 4. БЕЛКИ РЕНИН-АНГИОТЕНЗИНОВОЙ СИСТЕМЫ ПРИ
ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ По нашему мнению, обсуждение роли патофизиологических
механизмов контроля экспрессии генов белков, принимающих участие в
регуляции деятельности почек, предполагает использование в виде
иллюстрации какой-либо определенной модели патологии. В качестве
такой модели нами были выбраны процессы малигнизации и
метастазирования опухолей. С одной стороны, одним из важных маркеров
малигнизации клеток, есть изменения в процессах биосинтеза белков, в
норме не характерных для данной популяции клеток. С другой стороны,
онкологические заболевания, выбранные в качестве примера, на первый
взгляд, не имеют прямого отношения к системам контроля водно-солевого
гомеостаза. Тем не менее, они, во-первых, демонстрируют некоторые
специфические, нельзя сказать, что второстепенные, свойства протеинов,
объединяемых общим термином «компоненты РАС». Во-вторых,
описываемые закономерности изменения экспрессии белков —
«компонентов РАС» при онкологических заболеваниях дают повод оценить
широкий спектр функций данной группы протеинов. По данным литературы, компоненты ренин-ангиотензиновой
системы (РАС) могут принимать участие в процессах малигнизации
тканей, стимулировать рост и метастазирование опухолей (Regulska K. et
al., 2013; Gomez R.A., Sequeira-Lopez M.L.S., 2016; Pinter M., Jain R.K.,
2017;
Pinter M. et al., 2017). Более ранние исследования
продемонстрировали диагностическую ценность анализа экспрессии
компонентов РАС в онкологии (Rоmer F. K., 1981). Результаты
современных исследований подтверждают тезис о диагностической
ценности анализа экспрессии компонентов РАС, подчеркивая также их
значение в составлении прогноза течения заболевания и выборе способа
лечения злокачественных опухолей (Regulska K. et al., 2013; Tawinwung S. et al., 2015; Gomez R.A., Sequeira-Lopez M.L.S., 2016). Наряду с этим,
следует указать, что пептиды-компоненты РАС рассматривают в качестве
ключевых патогенетических механизмов роста и метастазирования
злокачественных опухолей, включая стимуляцию локальной продукции
ангиотензина-II (А-II), повышение экспрессии рецепторов к А-II,
изменение баланса экспрессии ангиотензин-I-превращающих ферментов
(АСЕ-1 и АСЕ-2) и уровень образования продуктов их реакции (А-II и А-1-
7 соответственно) (Regulska K. et al., 2013; Sobczuk P. et al., 2017; Sun H. et
al., 2017). Объектом внимания некоторых исследований является изучение
степени риска индукции канцерогенеза ингибиторами РАС (Connolly S. et
al., 2011; Azoulay L. et al., 2012; Yang Y. et al., 2015; Sobczuk P. et al., 2017). 105 Вместе с тем, патогенетические механизмы, индуцирующие увеличение
экспрессии белков-компонентов РАС в малигнизированных клетках, их
роль в процессах роста и метастазирования остаются мало изученными. 106 106 4.1.1. Рецепторы к А-II А-II оказывает свое влияние через АТ1 и АТ2 популяции рецепторов. Установлено, что в клетках астроцитомы человека частота выявления АТ1
рецепторов у пациентов с высокой степенью злокачественности опухоли
(степень III и IV) возрастает до 67% против 10% в группе с низким
уровнем злокачественности, что положительно коррелирует с
интенсивностью пролиферации клеток и плотностью неоангиогенеза
(Arrieta O. et al., 2008). В исследованиях на лабораторных животных,
привитых культурой клеток колоректального рака (CRC), было
установлено, что А-II через АТ1 и АТ2 рецепторы стимулирует миграцию
малигнизированных клеток и их метастазирование в печень (Nguyen L. et
al., 2016). Сообщается, что при некоторых онкологических заболеваниях
легких раковые клетки, демонстрирующие высокие уровни экспрессии
АТ1-рецепторов, обладают резистентностью к воздействию цитостатиков
(Cheng Q. et al., 2016). Анализ клинических наблюдений позволяет сделать
вывод о том, что повышение экспрессии АТ1 рецепторов
малигнизированными клетками свидетельствует о неблагоприятном
прогнозе
течения
заболевания,
обусловленном
стимуляцией
неоангиогенеза, роста и метастазирования опухоли (Keizman D. et al., 2011;
Sun H. et al., 2017). Подчеркивается, что активация плейотропных АТ1-
зависимых проонкогенных эффектов А-II может затрагивать в том числе
лимфоциты и связанные с опухолью макрофаги, что приводит к снижению
противоракового иммунитета, изменению продукции интерлейкинов и
провоспалительных цитокинов (Coulson R. et al., 2017; Pinter M., Jain R.K.,
2017). Значительный прирост АТ1 белка в трансформированных клетках
происходит за счет активации гена AGTR1 (Coulson R. et al., 2017). Возможно, стимуляция неоангиогенеза, реализуемая через АТ1 рецепторы,
является одним из универсальных патогенетических механизмов
прогрессирования опухолей различного генеза (Osumi H. et al., 2015; Pinter
M., Jain R.K., 2017). Приводятся данные о синэргических эффектах систем
рецепторов АТ1/А-II и АТ2/А-II в стимуляции неоангиогенеза (Ager E.I. et
al., 2011), а также усилении миграции клеток, воспаления формирование
внеклеточного матрикса через AT1 и AT2 рецепторы к А-II (Aydiner A. et
al., 2015). Показано, что изменения экспрессии АТ1 и АТ2 рецепторов
допустимо рассматривать в качестве маркеров малигнизации слизистой
желудка, индуцированной патогенными микроорганизмами, например
HELICOBACTER
PYLORI (Sugimoto M. et al., 2012), а также при 107 прогрессировании плоскоклеточного рака языка (Itinteang T. et al., 2016),
прогрессировании колоректального рака и оценке степени риска его
метастазирования (Kuniyasu H., 2012; Shimizu Y. et al., 2017), диагностике
онкологических заболеваний легких (Gallagher P.E. et al., 2011) и молочной
железы (Vinson G.P. et al., 2012). Уровень экспрессии рецепторов А-II
рассматривается в качестве прогностического критерия течения
плоскоклеточного рака пищевода (Li S.-H. et al., 2016) и светлоклеточного
рака почки (Dolley-Hitze T. et al., 2010). 4.1.1. Рецепторы к А-II Возможно, динамика изменения
экспрессии АТ1 И АТ2 может рассматриваться в качестве интегрального индикатора
чувствительности малигнизированной ткани к воздействию гуморальных
индукторов канцерогенеза (Rhodes D.R. et al., 2009; Vinson G.P. et al., 2012;
Sugimoto M. et al., 2012; Regulska K. et al., 2013; Pinter M., Jain R.K., 2017). В ряде обзорных публикаций достаточно подробно изложена оценка
результатов исследования особенностей экспрессии АТ1 и АТ2 рецепторов
А-II при различных онкологических заболеваниях, их диагностическая и
прогностическая ценность. Представлены аргументы с точки зрения их
роли в патогенезе заболеваний, прогрессировании и диссеминации
опухолей, а также перспективность клинического применения селективных
антагонистов рецепторов А-II в целях повышения эффективности
химиотерапии, иммунотерапии и ингибиторов неоангиогенеза в онкологии
(Vinson G.P. et al., 2012; Regulska K. et al., 2013; Wegman-Ostrosky T. et al.,
2015; Sobczuk P. et al., 2017; Pinter M., Jain R.K., 2017). 4.1.2. Ангиотензин-I-превращающий фермент (АСЕ-1). Ангиотензин-I-превращающий
фермент
(АСЕ-1),
карбоксипедиптидаза, один из ключевых факторов, осуществляющих
превращение ангиотензина-I
(А-1) в физиологически активный
ангиотензин-II
(А-II). Вместе с тем, при патологии, включая
онкологические заболевания, роль АСЕ-1 в образовании А-II может
изменяться за счет усиления вклада АСЕ-независимого пути конверсии А-1
в А-II в присутствии альфа-химазы и других пептидаз, формируя
резистентность опухолевых клеток к современным методам
противораковой терапии (Xie G. et al., 2017; Sobczuk P. et al., 2017). Широко известен и тот факт, что АСЕ-1, обладая относительно низкой
субстратной специфичностью, может участвовать не только в образовании
А-II, но и в метаболизме кининов, а также других физиологически
активных молекул, потенциально актуальных для процессов
канцерогенеза, роста и диссеминации опухолей (Regulska K. et al., 2013;
Sobczuk P. et al., 2017). Привлекают внимание сведения о том, что АСЕ-1, 108 помимо пептидазной активности, может непосредственно участвовать во
внутриклеточной передаче сигнала А-II, фактически являясь рецептором
октапептида (de Alvarenga E.C. et al., 2016). По мнению авторов
цитируемой публикации, механизм АСЕ-зависимой рецепции А-II может
выполнять важную роль в управлении миграции и пролиферации раковых
клеток. Следовательно, динамика изменения топологии и уровней
экспрессии АСЕ при онкологических заболеваниях может служить
маркером локализации проонкогенных эффектов
А-II
и других
гуморальных факторов, метаболизм которых связан с функциями
компонентов РАС. Например, при онкологических заболеваниях почек
наблюдается закономерное изменение активности и топологии экспрессии
белков АСЕ (Errarte P. et al., 2017; Sobczuk P. et al., 2017). (
,
;
,
)
В норме эпителий корковых сегментов канальцевого отдела нефрона,
в частности, эпителий проксимального отдела, демонстрирует высокие
показатели экспрессии АСЕ, который отсутствует в клетках
светлоклеточного рака почки (CCRCC) и выявляется только в кровеносных
сосудах опухоли (Errarte P. et al., 2017). Авторами показано, что уровень
экспрессии белка в опухоли и величина его энзиматической активности в
плазме крови могут служить маркером CCRCC агрессивности опухоли и
является индикатором выживаемости пациентов с CCRCC. С другой
стороны, перспектива применения широко известных ингибиторов АСЕ в
целях подавления неоангиогенеза в злокачественных новообразованиях
рассматривается в качестве одного из основных аргументов к применению
препаратов данной группы в онкологии (Shen J. et al., 2016). Показано, что
способствующее ускользанию от противоракового иммунитета
микроокужение опухолевых клеток мышей, может формироваться
макрофагами и связанными с опухолью фибробластами (Nakamura K. et al.,
2018). По мнению авторов, резко повышенный в макрофагах уровень
экспрессии АСЕ указывает на повышение интенсивности локальной
продукции физиологически активных веществ, вызывающих
иммуносупрессию: оксид азота, трансфомирующий фактор роста-бета1 и
PGE2. 4.1.2. Ангиотензин-I-превращающий фермент (АСЕ-1). В норме экспрессия АСЕ критически важна для формирования
специфического микроокружения в процессах цитодифференцировки на
стадии эмбрионального развития органа или в некоторых интенсивно
пролиферирующих тканях взрослого человека. Однако, чрезмерно высокий
уровень экспрессии не только ассоциируется с нарушениями гемопоэза, но
и рассматривается, как эффект ACE в онкогематологии (Haznedaroglu I.C.,
Malkan U.Y., 2016). Существенное повышение экспрессии АСЕ рака
гортани свидетельствует о неблагоприятном течении заболевания и
высоком риске метастазирования опухоли (Han C., Ge W., 2016). 109 Следовательно, изменение экспрессии АСЕ-1 наряду с изучением
полиморфизма гена АСЕ широко используется в современной онкологии в
качестве маркера тяжести течения заболевания и его прогноза (Regulska K. et al., 2013). Вместе с тем, уровень экспрессии АСЕ-1 клетками
злокачественных опухолей не всегда коррелирует с интенсивностью
локального продукции А-II, по причине усиления активности, например,
химазы, регулирующей АСЕ-независимый путь образования А-II (Xie G. et
al., 2017). Помимо этого, необходимо учитывать, что АСЕ непосредственно
принимает участие в регуляции иммунных реакций организма
(Haznedaroglu I.C., Malkan U.Y., 2016). 4.1.3. Ангиотензин-I-превращающий фермент-2 (АСЕ-2) и ось
ACE2/Ang-(1–7)/MAS1. АСЕ-2, гомолог АСЕ-1, отвечает за метаболический клиренс А-II,
используемого ферментом в качестве субстрата для синтеза ангиотензина-
1-7 (А-1-7). В свою очередь, А-1-7, осуществляя регуляторное влияние
через
МAS1-рецепторы,
оказывает
оппозиционное
действие
вазотоническим, провоспалительным и просклерозирующим эффектам А-
II (Clarke N.E., Turner A.J., 2012). Снижение уровня экспрессии АСЕ-2
клетками рака молочной железы рассматривается в качестве маркера
тяжелой формы течения заболевания с высоким риском метастазирования
(Yu С. et al., 2016). По мнению авторов, уровень экспрессии АСЕ-2
отражает степень влияния ACE2/Ang-(1–7)/MAS1 оси на ограничение
трансформации кальций-зависимых путей внутриклеточной передачи
сигнала, характерной для процесса малигнизации клеток. Показано, что
уровень экспрессии АСЕ-2 отрицательно коррелирует с интенсивностью
неоангиогенеза в некоторых опухолях легких и чувствительностью
раковых клеток к цитостатикам. Активность оси ACE2/Ang-(1-7)/MAS1
может угнетать секрецию VEGFа и подавлять активности матричных
металлопротеиназ MMP-2 и MMP-9, тем самым способствуя ограничению
неоангиогенеза, повышению чувствительности опухоли к цитостатикам и
снижению риска метестазирования (Feng Y. et al., 2011; Cheng Q. et al.,
2016). В ряде публикаций указывается, что гипоксия является признаком
солидных опухолей, подчеркивая, что условия гипоксии способствуют
усилению проонкогенного влияния АСЕ-1/А-II на фоне снижения эффектов
оси ACE-2/Ang-(1-7)/MAS1 (Fan L. et al., 2014). Авторами цитируемой
публикации показано, что in vitro в культуре клеток карциномы легких
Льюиса гипоксия способствует снижению экспрессии АСЕ-2 на фоне АСЕ-
1/А-II-зависимой индукции VEGFа. Приводятся аргументы в пользу
перспективности клинического использования А-1-7, как фактора 110 противораковой терапии опухоли груди, клетки которой не экспрессируют
рецепторы эстрогенов, рецепторы прогестерона и рецептора-2
эпидермального фактора роста (Luo Y. et al., 2015). Некоторые обзоры
также содержат позитивную оценку перспектив ACE-2/Ang-(1-7)/MAS1
оси в противораковой фармакологии (Regulska K. et al., 2013). Наряду с
этим, подчеркивается, что характер влияния ACE-2/Ang-(1-7)/MAS1 оси на
раковые клетки и прогрессирование опухоли может зависеть от
локализации опухоли (Wegman-Ostrosky T. et al., 2015; Haznedaroglu I.C.,
Malkan U.Y., 2016; Sobczuk P. et al., 2017). В частности, приводятся данные
о том, что А-1-7 стимулирует метастазирование почечно-клеточного рака
(RCC), индуцирует активацию генов провоспалительных факторов, в
целом способствуя прогрессированию заболевания (Sobczuk P. et al., 2017). Принимая к сведению изложенные факты, авторы склоняются к мнению о
том, что А-1-7 в отношении RCC обладает, скорее, пронкогенным
действием. 4.1.4. Ангиотензиноген. Ангиотензиноген (Agt) является универсальным предшественником
А-II и А-1-7. В норме Agt, главным образом, синтезируется в печени. При
онкологических заболеваниях, как правило, печень сохраняет роль
основного источника Agt (Vinson G.P. et al., 2012). Однако, представляют
интерес данные о диагностической ценности локальной продукции Agt,
как маркера канцерогенеза. Также привлекают внимание особенности
метаболизма Agt раковыми клетками. С одной стороны, локальная
продукция
Agt
рассматривается в качестве одного из наиболее
информативных маркеров активности опухолевого неоангиогенеза (Choi J.-
H. et al., 2014). С другой стороны, согласно данным цитируемой
публикации, доминирующим продуктом конверсии Agt в опухолевых
тканях является А-II. При этом, комбинированное влияние HIF-1-альфа и
А-II, на фоне более высокой продукции Agt, рассматривается в качестве
базового патогенетического механизма стимуляции ростовых факторов (в
частности, VEGFа), активирующих опухолевый неоангиогенез. Действительно, результаты клинических исследований показали, что, во-
первых, повышенная экспрессия гена Agt у пациентов с глиобластомой,
может расцениваться, как маркер резистентности опухоли к
противораковой терапии, направленной на угнетение опухолевого
неоангиогенеза (Urup T. et al., 2016). Во-вторых, более высокая экспрессия
гена Agt опухолевой тканью сопровождается усилением локальной
продукции А-II. Тем не менее, в литературе приводятся данные о том, что
также Agt обладает способностью угнетать неоангиогенез (Wegman- 111 Ostrosky T. et al., 2015). Обсуждая локальную продукцию Agt, необходимо
уточнить, что, по мнению некоторых авторов, стимуляция локальной
экспрессии компонентов РАС, включая Agt, рассматривается в качестве
центрального индуктора внутриклеточного каскада регуляторных белков,
определяющих процессы малигнизации клеток и метастазирования
(Sugimoto M. et al., 2012). Более того, опубликованные результаты не
исключают возможности активации и перестройки внутриклеточного
метаболизма компонентов РАС в раковых клетках (Blanco L.. et al., 2014). Что не противоречит мнению об универсальной патогенетической роли
активации внутриклеточной РАС, причастной также и к процессам
модуляции экспрессии генов (Ellis B. et al., 2012; De Mello W.C., 2015). Вместе с тем, указывается на тканеспецифические особенности экспрессии
Agt, как маркера риска онкологических заболеваний. Например, риск
возникновения рака легких ассоциируется со снижением продукции Agt
белка (Wang H. et., 2015). По мнению авторов, эпигенетические механизмы
снижения экспрессии гена Agt и точечные мутации гена Agt могут
рассматриваться в качестве факторов, усиливающих риск онкологических
заболеваний легких. Возможно, динамика локальной продукции Agt
протеина и его уровни в плазме крови могут по-разному формировать
прогноз течения метастазов колоректального рака (Martin P. et al., 2014). Авторами установлено, что повышение уровня Agt протеина в сыворотке
крови достоверно ассоциировалось с худшей общей выживаемостью, а
эпителиальная экспрессия Agt достоверно ассоциировалась с улучшенной
выживаемостью без прогрессирования заболевания. 4.1.4. Ангиотензиноген. Еще один аспект
диагностической ценности локальной продукции
Agt
протеина
опухолевыми тканями может быть обусловлен закономерным изменением
экспрессии гена Agt по мере течения заболевания (Vinson G.P. et al., 2012). 4.1.5. (Про)Ренин. В последнее время молекула (про)ренина и ее рецепторы привлекает
все большее внимание не только, в качестве регуляторного фермента РАС,
но и как важный элемент механизмов контроля онтогенеза, заживления ран
и патогенеза ряда заболеваний (Gomez R.A., Sequeira-Lopez M.L.S., 2016). В некоторых обзорах, посвященных анализу патогенетической роли РАС в
онкологических заболеваниях, встречаются сведения о важном влиянии
ренина на процессы малигнизации клеток и прогрессирования опухоли
(Vinson G.P. et al., 2012; Sugimoto M. et al., 2012). В исследованиях in vitro
установлено, что ренин может оказывать стимулирующее влияние на рост
культуры клеток почечно-клеточного рака (Hu J. et al., 2015). Экспрессия
ренина может рассматриваться, как маркер нормального созревания
предшественников клеток крови или их малигнизации (Haznedaroglu I.C., 112 Malkan U.Y., 2016). Авторы цитируемого обзора подчеркивают, что
экспрессия ренина была обнаружена в клетках острого миелоидного
лейкоза, в клетках хронического миелоидного лейкоза и острого
лимфолейкоза. Высказывается мнение о том, что стволовые клетки
костного мозга, которые экспрессируют ренин являться источником
лимфобластного лейкоза (Belyea B.C. et al., 2014). По данным литературы,
экспрессия гена ренина в процессе нормального и малигнизированного
емопоэза может регулироваться эпигенетическими механизмами (Belyea
B.C. t al., 2014; Haznedaroglu I.C., Malkan U.Y., 2016). В контексте
обсуждаемой емы уместно напомнить, что сложно функционирующая,
относительно мало зученная, система рецепторов к (про)ренину имеет
отношение не только к АС, но и к регуляции экспрессии генов белков-
индукторов процессов оспаления и фиброза тканей (Nguyen G., 2011). Дальнейшие исследования одтвердили РАС-независимые эффекты
системы рецепторов к (про)ренину, родемонстрировав их роль в регуляции
фундаментальных механизмов онтроля гомеостаза клетки (Müller D.N. et
al., 2012). Также было установлено, то уровни в плазме крови рецепторов
к (про)ренину ((P)RR) в группе нкологических пациентов были резко
повышены (Shibayama Y. et al., 2015). На сновании анализа динамики
экспрессии (P)RR в клетках на различной стадии алигнизации авторами
цитируемой публикации делается вывод о том, что P)RR могут быть тесно
вовлечены в процессы онкогенез в поджелудочной елезе. Результаты
изучения in vitro экспрессии PRR в культивируемых клетках лиомы
человека позволяютс сделать вывод о том, что этот рецептор может ыть
как прогностическим маркером, так и мишенью в лечении заболевания
Kouchi M. et al., 2017). Приводятся данные о том, что изменение
экспрессии P)RR в процессе гемопоэза может рассматриваться в качестве
перспективного аркера диагностики в онкогематологии (Haznedaroglu I.C.,
Malkan U.Y., 2016). 113 4.2. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ, КАК ВОЗМОЖНЫЕ
РЕГУЛЯТОРЫ ЭКСПРЕССИИ ПРОТЕИНОВ-КОМПОНЕНТОВ РАС
ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ Представленная выше краткая информация о динамике экспрессии
протеинов-компонентов РАС в опухолевых тканях свидетельствует о том,
что, во-первых, этот показатель может существенно повышаться в тканях,
для которых в норме не характерны высокие уровни экспрессии данной
группы протеинов (Sugimoto M. et al., 2012; Shibayama Y. et al., 2015; Han
C., Ge W., 2016; Itinteang T. et al., 2016; Yue Z. et al., 2016). И напротив, при
некоторых онкологических заболеваниях клетки постепенно утрачивают
присущую им в норме способность экспрессировать белки РАС (Errarte P. et al., 2017; Sobczuk P. et al., 2017). Во-вторых, отмечается закономерное
изменение экспрессии и топологии белков РАС в опухолевых тканях в
зависимости от стадии течения и степени тяжести заболевания (Vinson G.P. et al., 2012; Haznedaroglu I.C., Malkan U.Y., 2016; Kouchi M. et al., 2017). В
ряде публикаций приводятся доказательства ведущей роли
эпигенетических механизмов в изменении синтеза белков, способных
стимулировать процессы малигнизации, воспаления, фиброза и
метастазирования (Tsai
Y.-P., Wu K.-J., 2012; Tan W. et al., 2014; Harb-De la
Rosa A.et al., 2015; Cheng Y. et al., 2016; Haznedaroglu I.C., Malkan U.Y.,
2016; Semenza G.L., 2016). При этом, уделяется внимание эпигенетической
перестройке экспрессии генов протеинов-компонентов РАС в процессах
малигнизации и роста раковых клеток (Tsai
Y.-P., Wu K.-J., 2012; Han C.-D.,
Ge W.-S., 2016; Haznedaroglu I.C., Malkan U.Y., 2016; ie G. et al., 2017). Эпигенетическая перестройка экспрессии компонентов РАС тносительно
новое и мало изученное направление в онкологии. В норме, л ние
пигенетических механизмов на динамику экспрессии генов белков АС про
еживается на ранних стадиях гисто- и органогенеза, а также в нтенсивно
про ферирующих тканях (Belyea B.C. et al., 2014; Haznedaroglu .C., Malkan
U.Y , 016). Сообщается, что одним из универсальных индукторов
кспрессии ген в елков-компонентов РАС, по мере прогрессирования
локачественных нов бразований, может являться HIF-1АЛЬФА (Tsai
Y.-P.,
Wu .-J., 2012; Choi J.-H. et l., 2014; Xie G. et al., 2017). Установлено, что
ряд акторов, соп тствующих ечению сахарного диабета, также может
оказывть лияние на экс рессию генов елков РАС, усиливая риск
онкологических аболеваний (Ya g X. et al., 2012; eddy M.A. et al., 2012;
Reddy M.A., Natarajan ., 2015; Weg an-Ostrosky T. et al., 015). ВОЗМОЖНО,
HIF-1АЛЬФА
ЕПОСРЕДСТВЕННО ПРИ ИМАЕТ УЧАСТИЕ В
ЕГУЛЯЦИИ
ЭКСПРЕССИИ НГИОТЕНЗИНОГЕНА (Agt) (Choi J.- . et al., 2014). В свою
чередь, локальная родукция Agt критически важна для усиления 114 образования -II, тимулирующего опухолевый неоангиогенез и мет
стазирование через Т1-рецепторы. Сообщается, что антагонист АТ1-
рецепторов олмесартан ожет ерез активацию синтеза микроРНК-205 инг
бировать экспрессию VEGF-а аковыми клетками (Yue Z. 4.2. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ, КАК ВОЗМОЖНЫЕ
РЕГУЛЯТОРЫ ЭКСПРЕССИИ ПРОТЕИНОВ-КОМПОНЕНТОВ РАС
ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ et al., 2016). В дан
ом случае А-II рассматривается ачестве регулятора процессов тра
скрипции. Действительно, спериментальные исследования показывают,
что А-II может усиливать дукцию провоспалительных цитокинов (IFNγ,
TNF ) и матричных ме аллопротеиназ (MMP2, MMP9), стимулируя адгезию
раковых клеток к эндотелиальным клеткам, а также активируя их
трансэндотелиальную миграцию и миграцию опухолевых клеток через
белки внеклеточного матрикса (Rodrigues-Ferreira S., et al. 2012). Наряду с
этим, высказывается мнение об универсальности эпигенетической
перестройки экспрессии компонентов РАС в патогенезе, в том числе, и
онкологических заболеваний (Kemp J.R. et al., 2014; Reddy M.A., Natarajan
R., 2015). С другой стороны, приводятся сведения об эпигенетических
эффектах А-1-7, направленных на ограничение подвижности раковых
клеток и их способности к метастазированию (de Oliveira da Silva B. et al.,
2016). С этой точки зрения особое значение приобретают данные о
способности АСЕ-1 участвовать в механизмах внутриклеточной передачи
сигнала А-II (de Alvarenga E.C. et al., 2016). Не менее актуальной является
информация о важности системы (про)ренин — рецепторы к (про)ренину в
управлении экспрессии генов, независимо от состояния активности РАС
(Nguyen G., 2011; Müller D.N. et al., 2012). Результаты дальнейших
исследований подтверждают тезис о том, что система (про)ренин —
рецепторы к (про)ренину могут выполнять важную функцию в патогенезе
и течении онкологических заболеваний (Shibayama Y. et al., 2015; Wang C. et al., 2016; Kaneko K. et al., 2017). образования -II, тимулирующего опухолевый неоангиогенез и мет
стазирование через Т1-рецепторы. Сообщается, что антагонист АТ1-
рецепторов олмесартан ожет ерез активацию синтеза микроРНК-205 инг
бировать экспрессию VEGF-а аковыми клетками (Yue Z. et al., 2016). В дан
ом случае А-II рассматривается ачестве регулятора процессов тра
скрипции. Действительно, спериментальные исследования показывают,
что А-II может усиливать дукцию провоспалительных цитокинов (IFNγ,
TNF ) и матричных ме аллопротеиназ (MMP2, MMP9), стимулируя адгезию
раковых клеток к эндотелиальным клеткам, а также активируя их
трансэндотелиальную миграцию и миграцию опухолевых клеток через
белки внеклеточного матрикса (Rodrigues-Ferreira S., et al. 2012). Наряду с
этим, высказывается мнение об универсальности эпигенетической
перестройки экспрессии компонентов РАС в патогенезе, в том числе, и
онкологических заболеваний (Kemp J.R. et al., 2014; Reddy M.A., Natarajan
R., 2015). С другой стороны, приводятся сведения об эпигенетических
эффектах А-1-7, направленных на ограничение подвижности раковых
клеток и их способности к метастазированию (de Oliveira da Silva B. et al.,
2016). С этой точки зрения особое значение приобретают данные о
способности АСЕ-1 участвовать в механизмах внутриклеточной передачи
сигнала А-II (de Alvarenga E.C. et al., 2016). 4.2. ЭПИГЕНЕТИЧЕСКИЕ МЕХАНИЗМЫ, КАК ВОЗМОЖНЫЕ
РЕГУЛЯТОРЫ ЭКСПРЕССИИ ПРОТЕИНОВ-КОМПОНЕНТОВ РАС
ПРИ ОНКОЛОГИЧЕСКИХ ЗАБОЛЕВАНИЯХ Не менее актуальной является
информация о важности системы (про)ренин — рецепторы к (про)ренину в
управлении экспрессии генов, независимо от состояния активности РАС
(Nguyen G., 2011; Müller D.N. et al., 2012). Результаты дальнейших
исследований подтверждают тезис о том, что система (про)ренин —
рецепторы к (про)ренину могут выполнять важную функцию в патогенезе
и течении онкологических заболеваний (Shibayama Y. et al., 2015; Wang C. et al., 2016; Kaneko K. et al., 2017). 115 4.3.
ОНКОЛОГИЧЕСКИЕ
АСПЕКТЫ
ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА Само понятие «локальная РАС» формировалось, как представление
об элементе внутриорганного гуморального комплекса контроля
гомеостатических функций данного органа. На примере локальной РАС
почек эту мысль можно проиллюстрировать следующим примером. В
норме, адекватным стимулом активации внутриренальной РАС есть два
различных механизма, обусловленных внутриорганными изменениями
кровяного давления и интенсивности транспорта хлорида натрия в области
macula densa. В результате активации системы происходит усиление
секреции клетками юкста-гломерулярного аппарата регуляторного
фермента РАС — ренина и повышение продукции А-II. Основные
ренотропные физиологические эффекты А-II реализуются, в основном, в
отношении параметров внутрипочечной гемодинамики, процессов
фильтрации и канальцевого транспорта веществ (г.о. натрия) на уровне
проксимального отдела нефрона. В том числе через инициацию эффектов,
контролирующих транскрипцию транспортных и регуляторных белков в
проксимальных нефроцитах (Li X.C. et al., 2012; Satou R., Gonzalez-
Villalobos R.A., 2012). В физиологических условиях, индукция секреции
ренина (Sparks M.A. et al., 2014) и А-II-зависимая стимуляция
транскрипции транспортных белков канальцевого эпителия (Shao W. et al.,
2013) адекватны текущему состоянию водно-солевого баланса организма. Например, вызванная гипонатриевой диетой физиологическая стимуляция
РАС не сопряжена с повышением ренальных потерь Agt или с развитием
повреждений ренальной паренхимы (Shao W. et al., 2013). Следовательно,
конечным результатом деятельности сложного комплекса гуморальных
регуляторов внутрипочечной системы контроля гомеостаза является
поддержание стабильных параметров кровяного давления, ионного
гомеостаза, кислотно-основного равновесия, постоянства объема
внеклеточной жидкости организма (Satou R., Gonzalez-Villalobos R.A.,
2012; Zhuo J.L. et al., 2013; Sparks M.A. et al., 2014; Ferrão F.M. et al., 2014). При этом, топология основной массы ренин-секретирующих клеток
почки в области ЮГА, скорее всего, имеет принципиально важное
значение. Поскольку данная популяция клеток непосредственно получает
информацию о параметрах гемодинамики и о состоянии транспорта ионов
хлора и натрия в дистальных извитых сегментах нефрона. Напротив,
появление эктопических очагов секреции, например, ренина за пределами
ЮГА (в канальцевом эпителии, клетках мезангиума) лишает клетки-
продуценты адекватной стимуляции. По нашему мнению, такие события, в 116 совокупности с локализацией продукции всех белков-компонентов РАС в
одной клетке может создавать предпосылки для неограниченной активации
сформировавшихся эктопических очагов РАС, нацеленных на инициацию и
каскадное усиление патологических изменений в ренальной паренхиме. у
р
р
С другой стороны, гипоксия, оксидативный стресс, высокий уровень
глюкозы в крови и другие неблагоприятные факторы способны
индуцировать эпигенетические механизмы активации фиброза и
воспаления ренальной паренхимы, в том числе и изменений топологии
компонентов РАС через механизмы контроля экспрессии генов (Macconi
D.et al., 2014; Reddy M.A., Natarajan R. , 2015, Nangaku M. et al., 2017). 4.3.
ОНКОЛОГИЧЕСКИЕ
АСПЕКТЫ
ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА et al., 2013; Pinter M., Jain R.K., 2017;
Sobczuk P. et al., 2017). Также, как в патогенезе и прогрессировании
почечной недостаточности, компоненты РАС раковых клеток не причастны
к выполнению гомеостатических функций. Следовательно, по нашему
мнению, локальной РАС не являются. С точки зрения интересов
практической медицины речь идет о целесообразности использования
блокаторов РАС в лечении онкологических заболеваний. По нашему
мнению, в этом вопросе можно выделить несколько аспектов. С одной
стороны, усиление экспрессии эктопических очагов РАС клетками
злокачественной опухоли, на первый взгляд, дает основание ожидать
результативности использования ингибиторов АСЕ и антагонистов
рецепторов А-II
в лечении онкологических заболеваний. В
действительности, монотерапия блокаторами РАС онкологических
заболеваний
может
демонстрировать
достаточно
умеренный
терапевтический результат. Сообщается о путях усиления противораковых
эффектов ингибиторов РАС в комплексе с мероприятиями иммунотерапии
и химиотерапии (Pinter M., Jain R.K., 2017). Наряду с этим, указывается на
потенциальные риски, связанные с использованием определенных
блокаторов РАС в лечении конкретных онкологических заболеваний
(Sobczuk P. et al., 2017), вплоть до полной нецелесообразности их которого носят оппозиционный характер по отношению к А II. В результате, в отличии от физиологических условий, патологическое
изменение топологии и уровня экспрессии компонентов РАС приводит:
1.К появлению эктопических очагов РАС, в т.ч. усилению внутриклеточной
РАС канальцевого эпителия.2.К образованию эктопических очагов РАС
(ренин, ангиотензиноген) способствующих ускользанию пусковых
механизмов активации РАС от базовых егуляторных стимулов параметров
водно-солевого баланса организма и гемодинамики. 3.К изменению
вектора регуляторных эффектов эктопических очагов РАС, которые больше
не направлены на поддержание гомеостаза. Сформировавшиеся
эктопические очаги экспрессии компонентов РАС обеспечивает
дальнейшую неограниченную индукцию фиброза, воспаления и
гипертрофии клеток ренальной паренхимы. 4.К изменению
внутриклеточных систем передачи сигнала (Satou R., Gonzalez-Villalobos
R.A., 2012) и баланса регуляторного влияния А-II и А-1-7, в сторону
усиления активности АСЕ-1 и альфа-химазы на фоне снижения экспрессии
АСЕ-2 (Sparks M.A. et al., 2014).Отметим, что подобные изменения
происходят и в результате перестройки экспрессии компонентов РАС в
клетках злокачественных опухолей, индуцирующих малигнизацию клеток,
фиброз и воспаление ткани, неоангиогенез, метастазирование и
иммуносупрессию (Regulska K. et al., 2013; Pinter M., Jain R.K., 2017;
Sobczuk P. et al., 2017). Также, как в патогенезе и прогрессировании
почечной недостаточности, компоненты РАС раковых клеток не причастны
к выполнению гомеостатических функций. Следовательно, по нашему
мнению, локальной РАС не являются. С точки зрения интересов
практической медицины речь идет о целесообразности использования
блокаторов РАС в лечении онкологических заболеваний. По нашему
мнению, в этом вопросе можно выделить несколько аспектов. 4.3.
ОНКОЛОГИЧЕСКИЕ
АСПЕКТЫ
ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА Одним из важных результатов указанных событий есть усиление секреции
ренина в паренхиме мозгового слоя почки (Zhuo
J.L., 2011), в
гладкомышечных клетках артериол, мезангиальных клетках и в
интерстиции через эпигенетические механизмы контроля экспрессии генов
(Sparks M.A. et al., 2014; De Mello W.C., 2015). Ренин — регуляторный
фермент РАС, детерминирующий интенсивность дальнейшей продукции
А-II. Накопление активных форм кислорода (ROS) в почечной ткани
способствует усилению экскреции Agt почками, усиливая цепь обратной
связи между активацией ROS и дальнейшей активацией ROS синтеза Agt
(Nguyen M.T.X. et al., 2015). Показано, что инфузия животным А-II
существенно активирует биосинтез Agt в проксимальных нефроцитах,
приводя к дальнейшему росту канальцевой продукции А-II и повышению
его патогенетического влияния (Ramkumar N. et al., 2016). В целом,
аккумуляция компонентов РАС в проксимальных нефроцитах, усиление
внутриклеточной продукции А-II
и его эффектов на процессы
транскрипции, функции митохондрий, усиление продукции ROS — один из
базовых патогенетических механизмов нарушений гомеостатических
функций почек (Navar L.G. et al., 2011; Ellis B. et al., 2012; Li X.C., Zhuo
J.L., 2016). В качестве диагностического критерия патологической
трансформации внутрипочечной РАС рекомендуется использование
ренальной экскреции ангиотензиногена (Kobori H.et al., 2002; Navar L.G. et
al., 2011; Alge J.L. et al., 2013). Таким образом, патологическая трансформация внутрипочечной РАС
осуществляется: 1.Под контролем эпигенетических механизмов, изменяющих процессы
транскрипции, в том числе белков-компонентов РАС. 2.В результате формирования эктопических очагов биосинтеза ключевых
белков-компонентов РАС. 3.В результате усиления внутриклеточной продукции А-II и усиления
влияния октапептида на транскрипцию белков. 117 4.Ослаблением экспрессии АСЕ-2, снижением продукции А-1-7, эффекты
которого носят оппозиционный характер по отношению к А-II. которого носят оппозиционный характер по отношению к А II. В результате, в отличии от физиологических условий, патологическое
изменение топологии и уровня экспрессии компонентов РАС приводит:
1.К появлению эктопических очагов РАС, в т.ч. усилению внутриклеточной
РАС канальцевого эпителия.2.К образованию эктопических очагов РАС
(ренин, ангиотензиноген) способствующих ускользанию пусковых
механизмов активации РАС от базовых егуляторных стимулов параметров
водно-солевого баланса организма и гемодинамики. 3.К изменению
вектора регуляторных эффектов эктопических очагов РАС, которые больше
не направлены на поддержание гомеостаза. Сформировавшиеся
эктопические очаги экспрессии компонентов РАС обеспечивает
дальнейшую неограниченную индукцию фиброза, воспаления и
гипертрофии клеток ренальной паренхимы. 4.К изменению
внутриклеточных систем передачи сигнала (Satou R., Gonzalez-Villalobos
R.A., 2012) и баланса регуляторного влияния А-II и А-1-7, в сторону
усиления активности АСЕ-1 и альфа-химазы на фоне снижения экспрессии
АСЕ-2 (Sparks M.A. et al., 2014).Отметим, что подобные изменения
происходят и в результате перестройки экспрессии компонентов РАС в
клетках злокачественных опухолей, индуцирующих малигнизацию клеток,
фиброз и воспаление ткани, неоангиогенез, метастазирование и
иммуносупрессию (Regulska K. 4.3.
ОНКОЛОГИЧЕСКИЕ
АСПЕКТЫ
ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА С одной
стороны, усиление экспрессии эктопических очагов РАС клетками
злокачественной опухоли, на первый взгляд, дает основание ожидать
результативности использования ингибиторов АСЕ и антагонистов
рецепторов А-II
в лечении онкологических заболеваний. В
действительности, монотерапия блокаторами РАС онкологических
заболеваний
может
демонстрировать
достаточно
умеренный
терапевтический результат. Сообщается о путях усиления противораковых
эффектов ингибиторов РАС в комплексе с мероприятиями иммунотерапии
и химиотерапии (Pinter M., Jain R.K., 2017). Наряду с этим, указывается на
потенциальные риски, связанные с использованием определенных
блокаторов РАС в лечении конкретных онкологических заболеваний
(Sobczuk P. et al., 2017), вплоть до полной нецелесообразности их 118 применения (Sørensen G.V. et al., 2013; Chae Y.K. et al., 2014; Nakai Y. et. al.,
2016). С другой стороны, на основе анализа природы экспрессии
эктопических очагов РАС, предполагаются качественно иные способы их
фармакологической коррекции, основанные на модулировании
эпигенетических механизмов подавления активности эктопической РАС
(Zhong Y. et al., 2013; Reddy M.A. et al., 2014). Высказывается мнение об
универсальной роли эпигенетических механизмов в патогенезе
формирования эктопической РАС при онкологических и неонкологических
заболеваниях (Kemp J.R. et al., 2014; Tang J., Zhuang S., 2015; Reddy M.A.,
Natarajan R., 2015). Анализируются перспективные способы лечения
онкологических заболеваний, полностью базирующиеся на управлении
эпигенетическими механизмами при помощи синтетических microRNA
(Tan W. et al., 2014; Felipe A.V. et al., 2014). Новые перспективы
использования селективных модуляторов эпигенетических процессов в
практической медицине, представляющих интерес и для онкологии,
подтверждаются сведениями о готовности применения данной группы
фармакологических препаратов (ингибиторы деацетилаз) в доклинических
испытаниях (Van Beneden K. et al., 2013). Таким образом, проведенный
обзор литературы показал, что изменение экспрессии компонентов РАС
тесно связано с патогенезом малигнизации клеток, прогрессированием
раковых опухолей, а также стимуляцией процессов метастазирования. Сведения о состоянии экспрессии белков компонентов РАС способствуют
пониманию механизмов канцерогенеза и диссеминации клеток опухоли. Эти данные позволяют использовать качественные и количественные
параметры экспрессии компонентов РАС в качестве маркеров тяжести
течения онкологического заболевания. Тесная вовлеченность компонентов
РАС вканцерогенез послужила основой для использования ингибиторов
РАС (ингибиторов АСЕ-1 и антагонистов рецепторов А-II) в терапии
онкологических заболеваний. Вместе с тем, анализ причин изменения
экспрессии белков РАС в клетках опухоли позволил выявить, что весьма
значимая функция в этих процессах принадлежит эпигенетическим
механизмам регуляции экспрессии генов. Благодаря исследованиям
состояния эпигенетических механизмов при онкологических заболеваниях
были разработаны принципиально новые методы их коррекции,
основанные на применении селективных регуляторов систем ковалентной
модификации белков-гистонов (например, ингибитор деацетилаз) и
технология синтеза микро РНК. Имеющиеся в литературе данные о
фармакологических свойствах указанных препаратов позволяют
предположить их перспективность в эффективном лечении онкологических
заболеваний применения (Sørensen G.V. 4.3.
ОНКОЛОГИЧЕСКИЕ
АСПЕКТЫ
ЭКСПРЕССИИ
КОМПОНЕНТОВ
РАС
И
ЛОКАЛЬНАЯ
РЕНИН-
АНГИОТЕНЗИНОВАЯ СИСТЕМА et al., 2013; Chae Y.K. et al., 2014; Nakai Y. et. al.,
2016). С другой стороны, на основе анализа природы экспрессии
эктопических очагов РАС, предполагаются качественно иные способы их
фармакологической коррекции, основанные на модулировании
эпигенетических механизмов подавления активности эктопической РАС
(Zhong Y. et al., 2013; Reddy M.A. et al., 2014). Высказывается мнение об
универсальной роли эпигенетических механизмов в патогенезе
формирования эктопической РАС при онкологических и неонкологических
заболеваниях (Kemp J.R. et al., 2014; Tang J., Zhuang S., 2015; Reddy M.A.,
Natarajan R., 2015). Анализируются перспективные способы лечения
онкологических заболеваний, полностью базирующиеся на управлении
эпигенетическими механизмами при помощи синтетических microRNA
(Tan W. et al., 2014; Felipe A.V. et al., 2014). Новые перспективы
использования селективных модуляторов эпигенетических процессов в
практической медицине, представляющих интерес и для онкологии,
подтверждаются сведениями о готовности применения данной группы
фармакологических препаратов (ингибиторы деацетилаз) в доклинических
испытаниях (Van Beneden K. et al., 2013). Таким образом, проведенный
обзор литературы показал, что изменение экспрессии компонентов РАС
тесно связано с патогенезом малигнизации клеток, прогрессированием
раковых опухолей, а также стимуляцией процессов метастазирования. Сведения о состоянии экспрессии белков компонентов РАС способствуют
пониманию механизмов канцерогенеза и диссеминации клеток опухоли. Эти данные позволяют использовать качественные и количественные
параметры экспрессии компонентов РАС в качестве маркеров тяжести
течения онкологического заболевания. Тесная вовлеченность компонентов
РАС вканцерогенез послужила основой для использования ингибиторов
РАС (ингибиторов АСЕ-1 и антагонистов рецепторов А-II) в терапии
онкологических заболеваний. Вместе с тем, анализ причин изменения
экспрессии белков РАС в клетках опухоли позволил выявить, что весьма
значимая функция в этих процессах принадлежит эпигенетическим
механизмам регуляции экспрессии генов. Благодаря исследованиям
состояния эпигенетических механизмов при онкологических заболеваниях
были разработаны принципиально новые методы их коррекции,
основанные на применении селективных регуляторов систем ковалентной
модификации белков-гистонов (например, ингибитор деацетилаз) и
технология синтеза микро РНК. Имеющиеся в литературе данные о
фармакологических свойствах указанных препаратов позволяют
предположить их перспективность в эффективном лечении онкологических
заболеваний. 119 СПИСОК ЛИТЕРАТУРЫ К ГЛАВЕ «БЕЛКИ РЕНИН-
АНГИОТЕНЗИНОВОЙ СИСТЕМЫ ПРИ ОНКОЛОГИЧЕСКИХ
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doi:10.1186/1755-1536-6-1 131 131 132 132 133 133 134 134 135 135 136 136 137 137 138 138 139 139 140 140 141 141 142 142 143 143 ISBN 9780359774524
DOI http://dx.doi.org/10.5281/zenodo.3270699
PBN Poland https://pbn.nauka.gov.pl/sedno-webapp/works/917606
Radomska Szkoła Wyższa w Radomiu, Polska
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26-600 Radom
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144 p. Number of characters: 250 000 (with abstracts). Number of images: 4 x 1000 characters (lump sum) = 4 000 characters. Total: Number of characters: 254 000 (with abstracts, summaries and graphics) = 6,35 sheet publications. 144 p. Number of characters: 250 000 (with abstracts). Number of images: 4 x 1000 characters (lump sum) = 4 000 characters. Total: Number of characters: 254 000 (with abstracts, summaries and graphics) = 6,35 sheet publications. ISBN 9780359774524 144
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Modern blast furnace ironmaking technology: potentials to meet the demand of high hot metal production and lower energy consumption
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Metallurgical & Materials Engineering
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Metallurgical and Materials Engineering
Association of Metallurgical Engineers of Serbia AMES Metallurgical and Materials Engineering
Association of Metallurgical Engineers of Serbia AMES Review paper
https://doi.org/10.30544/414 * Corresponding author: Elsayed Mousa, mousa71@yahoo.com MODERN BLAST FURNACE IRONMAKING TECHNOLOGY:
POTENTIALS TO MEET THE DEMAND OF HIGH HOT METAL
PRODUCTION AND LOWER ENERGY CONSUMPTION E.A. Mousa 1,2 * 1Researcher Scientist at Process Metallurgy Department, Swerim AB,
SE-971 25 Luleå, Box 812 Sweden. 2Associated Professor at Central Metallurgical Research and Development Institute
(CMRDI), Helwan, P.O. Box 87 Egypt. Received 04.04.2019
Accepted 20.06.2019 Received 04.04.2019
Accepted 20.06.2019 Abstract Iron and steel making is one of the most intense energy consuming in the industrial
sectors. The intensive utilization of fossil carbon in the ironmaking blast furnace (BF) is
related directly to CO2 emission and global warming. Lowering the energy consumption
and CO2 emission from BF comes on the top priorities from both economic and
environmental aspects. The BF has undergone tremendous modifications and
development to increase production and improve the overall efficiency. Both
technological development and scientific research drive one another to reach optimum
operation conditions, which are very close to the ideal conditions; however, further
development is still required to meet the stringent environmental regulations. The present
article provides a comprehensive review of recent research and development which were
carried out in modern blast furnace to increase the productivity meanwhile reduce the
energy consumption and CO2 emission to meet the demand of steel market and the
environmental protection. The recent technological and metallurgical improvements in
the BF are intensively discussed including: (i) modifications of BF design, top charging
and measuring system, (ii) upgrading of conventional top charging burden and alternative
agglomerates, (iii) developing of tuyeres injection system and injected materials, and (iv)
potentials of waste heat recovery and usage. These topics are reviewed and discussed in
some details to elucidate the potential of recent progress in BF technology in saving the
energy consumption and lowering CO2 emission. In this paper, the major research and
development which have been carried out in ironmaking BF technology are reviewed
with an overview of the future prospects. Keywords: blast furnace; CO2 emission; energy saving; top charging; tuyeres
injection; waste heat recovery. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 70 Introduction The steel industry is considered as a pillar of the economy in many of advance and
development countries. It is one of the most important sectors which have a high impact
on global economic growth. More than 6.0 million people around the world work for this
vital sector, with an estimated turn offer of 900 billion USD per year [1]. In recent years,
steel production has sharply increased to reach about 1.7 billion tons in 2017 [2]. Moreover, steel production and usage are expected to increase to reach 2.8 billion tons to
meet the future needs of a growing population in 2050 [3]. On the other hand, steel
manufacturing is one of the most significant energy- and carbon- consuming sectors. The
global energy consumption in steelmaking is estimated to be about 20% of the annual
industrial energy requirements. The fossil carbon is the primary source of heat and
reducing agents in iron and steel making and consequently represents a major contribution
to the global anthropogenic CO2 emission. According to the International Energy
Agency, the iron and steel industry accounts for approximately 7-9% of total world CO2
emissions [4]. The CO2 emission from iron and steelmaking reached to about 2.3 billion
tons in 2007 and it is expected to reach more than 3.0 billion tons in 2050 [3]. Tackling
CO2 emission is one of the most critical tasks facing the world today because further
pollutions and emissions will threaten our health, communities, economy, and
international security. Nowadays, the reduction of specific energy consumption and CO2
emission are coming on the top priorities of iron and steelmaking due to the dynamic
growth of energy prices as well as the commitment of governments to decrease CO2
emissions according to Paris agreement which entered into force on 4 November 2016 [5,
6]. Several ambitious actions were set all over the world to combat carbon footprint. In
Europe, for instance, 20-20-20 target aims at 20% decrease in greenhouse gas emissions
compared to 1990 levels, 20% share of renewable energy and 20% reduction of primary
energy consumption by 2020 [7]. Besides, CO2 Breakthrough Program was launched by
the American steel industry aims to dramatically reduce CO2 emission from iron and
steelmaking processes [8]. Contents 1. Introduction
71
2. Development in BF Design and Measuring Systems
72
3. Development in Top Charging Materials
75
3.1 Active Nut Coke
76
3.2 Cold Bonded Briquettes and Pre-reduced Agglomerates
77
3.3 Renewable Biomass
79
4. Development in Tuyeres Injections
81
4.1 Pulverized Coal Injection (PCI)
81
4.2 Oil and Natural Gas Injection
85
4.3 Off-Gases Injection
86
4.4 Waste Plastics and Biomass Injection
88
5. Potentials of Waste Heat Recovery (WHR)
89
5.1 Analysis of Waste Heat in Steel Plant
89
5.2 Benefits and Challenges of WHR
90
5.3 WHR from BF Slag
92
Acknowledgment
98
References
98 71 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … Introduction Although the steel industry is energy and carbon-intensive, it is essential to
mention that steel industry represents the core of green economy as the sectors and
technologies which drive the green economy such as wind energy, low-carbon transport,
clean energy and light vehicles, fuel-efficient infrastructure and recycling facilities are all
dependent on steel products. Furthermore, steel is the most recycled material in the world;
with over 650 Mt (million ton) recycled annually and over 97% of the raw materials used
for steelmaking were converted to steel products and by-products [1]. Once the steel is
produced it becomes a permanent resource for society as long as it is recovered at the end
of each product life cycle [9]. The combination of strength, recyclability, availability,
versatility, and affordability makes the steel as a unique material for many applications. The steel industry has recently undergone tremendous modification to reduce energy
consumption and CO2 emission, but further decreasing is still required. The ironmaking
process is the highest CO2 emission part in steel production route due to the intensive
utilization of fossil fuels for heating, melting, and reduction of iron ores. The most
common ironmaking process used worldwide is the BF which produces nowadays about
70% of total world steel production. Although many alternative ironmaking technologies
were developed, the BF is still the most cost-effective and highly productive process for
hot metal and probably remains so for the next decades. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 72 This article gives insights on the major modifications which have been done in
ironmaking BF in terms of technical and scientific aspects. The recent development in
the BF technology and its potentials in lowering the energy consumption and CO2
emission will be thoroughly discussed and the research gaps will be filled by providing
thoughts and recommendation that contribute in solving the challenges. The discussion
will focus on the development in BF design and measuring systems, developing top
charging materials, the progress of tuyeres injection and the recent potentials in waste
heat recovery from BF slag. This article gives insights on the major modifications which
have been done in BF in terms of scientific and technical aspects. Development in BF Design and Measuring Systems The BF was created in a long way of experiences through which tremendous
modifications and improvements were conducted to reach the current status. Intensive
work was done to increase BF productivity in order to meet the high demand for steel. The working volume of the BF has increased from less than 100 m3 to more than 5000
m3 [10]. The largest BFs at present are about 80 m in total height, with an inner height of
about 33-34 m and a maximum internal diameter of about 16-17 m and have an internal
volume of about 4500-5500 m3. A furnace of this size produces approximately 10,000-
13,000 tons of hot metal a day with annual production more than 4.0 Mt. In 2016 POSCO
Steelworks in South Korea has started hot metal production form the world`s largest BF
with an internal volume ~6000 m3 and annual production of about 5.65 Mt [10, 11]. It
was reported that BFs with an internal volume in the range of 3000-5000 m3 seems
optimal for BF-performance [12-16]. This means the BF-performance is more correlated
to specific productivity which measures the efficiency in terms of ton hot metal
(tHM)/m2/day [17]. The highest specific productivity corresponds to the working volume
of about 2700-3700 m3 with maximum specific productivity ~2.5 t/m3/d. The minimum
coke rate corresponds to the working volume of about 3000-3800 m3. The minimum coke
consumption rate is around 330 kg/tHM and the average is around 363 kg/tHM [13]. The furnace size accounts for only 50% of BF improvements in productivity while many
other parameters are responsible for increasing productivity as can be seen in Fig. 1 [18]. Developing of charged burden, furnace design, injection technologies, and process
control helps in the reduction of coke consumption. For instance, the BF charging system
was developed to sustain a good distribution of feeding materials inside the furnace and
consequently improve the gas streaming and production rate. The introduction of a bell-
less top charging system in the early 1970s instead of the two-bell system was a milestone
for modern charging system of BF. This system allows easier burden distribution through
a rotating chute and consequently uniform distribution of voidage and particle size of
charged materials inside the furnace. E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 73 Fig. 1. Advances in the blast furnace to reduce the coke consumption [18]. Fig. 1. Development in BF Design and Measuring Systems Advances in the blast furnace to reduce the coke consumption [18]. Parallel with the development of the top charging system, the hot stoves system at
the lower part of the shaft has undergone tremendous modification as can be seen in Fig. 2 [19]. About 1400 m3/tHM hot blast is blown into the modern BF at the temperature of
1100-1200 °C. The hot blast reacts with coke and pulverized coal in the BF raceway to
generate reducing gases (CO and H2) meanwhile increasing the raceway adiabatic flame
temperature (RAFT) to ~2100-2300 °C. Fig. 1. Advances in the blast furnace to reduce the coke consumption [18]. Parallel with the development of the top charging system, the hot stoves system at
the lower part of the shaft has undergone tremendous modification as can be seen in Fig. 2 [19]. About 1400 m3/tHM hot blast is blown into the modern BF at the temperature of
1100-1200 °C. The hot blast reacts with coke and pulverized coal in the BF raceway to
generate reducing gases (CO and H2) meanwhile increasing the raceway adiabatic flame
temperature (RAFT) to ~2100-2300 °C. Fig. 2. Schematic diagram of blast furnace and auxiliary hot blast stoves [19]. Fig 2 Schematic diagram of blast furnace and auxiliary hot blast stoves [19] Fig. 2. Schematic diagram of blast furnace and auxiliary hot blast stoves [19]. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 74 Preparation of hot blast begins by heating the stoves using gas fuel mixture of coke
oven gas (COG), natural gas (NG) and BF gas (BFG) [20-22]. The hot stove consists of
two parts work as combustion and heat exchanger for fuel gases and hot blast. The hot
blast can be enriched with oxygen to improve the combustion efficiency at the BF
raceway. Moreover, the enrichment of combustion air with oxygen reduces the nitrogen
content and consequently increases the RAFT. As oxygen enrichment increases, the coke
consumption in BF decreases as can be seen in Fig. 3 [21]. Therefore, as the blast
temperature and oxygen enrichment increase, the BF efficiency increases, meanwhile the
coke consumption and CO2 emission decrease. Fig. 3. Correlation between oxygen level in the blast and coke rate [21]. Besides the development of the upper and lower part of BF, the measuring systems
were significantly modified. Development in BF Design and Measuring Systems In the past, the BF was considered as “black box” due to the
less available information on the inner status and inner phenomena. During the last 30
years, many studies and experimental work based on sophisticated measuring devices and
techniques, as can be seen in Fig. 4, were developed and helped the ironmakers to
understand many of complex inner phenomena and to work on improving the furnace
efficiency [23]. Fig. 3. Correlation between oxygen level in the blast and coke rate [21]. Fig. 3. Correlation between oxygen level in the blast and coke rate [21]. Besides the development of the upper and lower part of BF, the measuring systems
were significantly modified. In the past, the BF was considered as “black box” due to the
less available information on the inner status and inner phenomena. During the last 30
years, many studies and experimental work based on sophisticated measuring devices and
techniques, as can be seen in Fig. 4, were developed and helped the ironmakers to
understand many of complex inner phenomena and to work on improving the furnace
efficiency [23]. E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 75 Fig. 4. Development in blast furnace measuring systems [21]. Fig. 4. Development in blast furnace measuring systems [21]. Recently measuring devices (vertical probe and tuyeres probe) were designed to
assist the development of BF modeling. The vertical probe gives feedback on the gas
temperature and gas composition from the charging level to the cohesive zone. On the
other hand, the tuyeres probe gives feedback on gas distribution in the raceway zone. Utilization of such sophisticated devices and measuring units has provided a clear vision
of the inner status of BF. Consequently, further R & D were conducted to improve the
BF-performance. Some developments were able to reach a full scale implementation
(such as pulverized coal injection, charging nut coke, and cold-bond briquettes) in modern
BF while others (such as self-reduced agglomerates, biomass for top charging and
injection, top gas recycling, recycling of dust and sludge, and minimizing dust emissions)
are still under investigation in pilot and lab scale. Also, specific attention is being paid
nowadays on the recovery of waste heat in the integrated steel plant using waste heat
recovery unit. These themes will be discussed in detail in the next sections. Development in Top Charging Materials As indicated in the previous section, the BF in old design was quite small, and it
was mainly based on top charging lump iron ores and charcoal. Later, in large and modern
BF, charcoal and lump ores were replaced by more efficient coke, sinter, and pellets. Modern BFs favor high strength coke and high iron content sinter and pellets. The high-
quality coke, sinter, and pellets increased BF efficiency and productivity. Even with these
improvements, the economic and environmental restrictions represented in high energy
cost and fossil CO2 emission have enforced the ironmakers to continue further
developments on top charging materials. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 76 Active Nut Coke Modern BF requires large coke size (40-60 mm or 40-80 mm) with low reactivity
to reduce the coke consumption in the upper part of BF shaft and to maintain good
permeability in the lower part of the shaft [24]. Also, the size distribution should be
narrow to maintain permeable coke structure in the BF [25]. The screening of
metallurgical coke to get the proper size for BF resulted in a generation of under-sieve
small coke namely “nut coke”. Due to the relatively high price of coke in addition to
pollutions emissions during its production, there are several attempts to exploit the under-
sieve coke or nut coke in the BF. The attempts were started by charging nut coke with
different ratios (5-30%) within the coke layers. This charging mode demonstrated a
negative impact on BF productivity (-0.9-6.5%) due to the non-uniform distribution and
disturbance of ascending gases in the shaft [26]. Studies were conducted on charging nut
coke within iron burden layers, as shown in Fig. 5 [26-34]. This charging mode
demonstrated a positive impact on the iron burden permeability, reducibility, and
reducing gas efficiency. Fig. 5. Comparison between the conventional charging method and coke mixed
charging method [35]. Fig. 5. Comparison between the conventional charging method and coke mixed
charging method [35]. One of the critical issues that should be considered to get smooth operation is the
uniform mixing of nut coke in the ore layer to maintain proper gas distribution and
maintain the required permeability especially in the cohesive zone [26]. Mixing of nut
coke within the ore bed is very effective in the enhancement of carbon solution loss
reaction from active coke while protecting the large size coke from gasification and
disintegration [35, 36]. This participates in lowering the temperature of the thermal
reserve zone (TRZT) and consequently decreases the total coke consumption in the BF. Fig. 6 shows the relation between TRZT and coke rate consumption. E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 77 Fig. 6. Reducing agents rate and thermal reserve zone temperature [35]. Fig. 6. Reducing agents rate and thermal reserve zone temperature [35]. It can be seen that lowering TRZT from 1000 °C to 750 °C can reduce the reducing
agents consumption by ~50 kg/tHM due to the shifting of Femet/FeO equilibrium towards
lower CO/CO2 [35]. Active Nut Coke Nowadays, the majority of BFs in Europe, Japan, and others use nut
coke at a different rate from 10–20 to 70–100 kg/tHM and more [37]. Although the
charging of nut coke can contribute to saving of lump coke in BF and reducing the coke
losses, it is still a type of fossil fuel that generates fossil CO2 emissions. Cold Bonded Briquettes and Pre-reduced Agglomerates q
gg
The integrated steel plant involves a series of interconnected units such as cake
making, sintering, BFs, basic oxygen furnaces, continuous casting, hot and cooled rolling. Each process generates a type of residues which are not suitable for further direct
processing to the final product without pre-treatment. Such secondary materials include
flue dust, filter dust, sludge, desulphurized scrap, Linz–Donawitz (LD) dust, coke fines,
and mill scales. These materials contain a valuable amount of iron, lime, and carbon but
it has a wide range of particle size, moisture content and chemical composition which
make it unstable for direct recycling without sorting, preparation, and agglomeration to
fulfill the specific requirements for charging back into BF. Intensive work was done to
return generated by-products as substitutes to raw materials in the steel making process. Some of the steel plants recycled these materials during sintering of iron ore while other
companies have a briquetting plant which can efficiently convert such secondary
materials into cold bonded briquettes instead of the landfill. Cement is usually used as a
binder to give the proper mechanical strength of top charged briquettes into the BF [38-
40]. Nowadays, cold bonded briquettes are charged at the rate of 100-120 kg/tHM in some
of BFs (e.g., SSAB in Sweden and Finland). These BFs are among the most efficient in
the world in terms of CO2 efficiency [41]. Feasibility of recycling of high-zinc fraction
of upgraded BF sludge to the hot metal (HM) in deS plant and BOF was recently studied
[42]. The BF sludge was incorporated in self-reducing cold-bonded briquettes and pellets. Charging of developed briquettes or pellets into the hot metal showed endothermic
reactions within the briquettes, which decreased the heating and melting rate in case of Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 78 briquettes. Smaller size pellets were completely reduced within 8 minutes of contact with
HM but still suffer from melt-in difficulties. On the other hand, the industrial trials
demonstrated possible recycling of 10.9 kg of briquettes (containing 40% of BOF sludge)
per tHM to the steel shop. briquettes. Smaller size pellets were completely reduced within 8 minutes of contact with
HM but still suffer from melt-in difficulties. On the other hand, the industrial trials
demonstrated possible recycling of 10.9 kg of briquettes (containing 40% of BOF sludge)
per tHM to the steel shop. Cold Bonded Briquettes and Pre-reduced Agglomerates p
p
Beside the efficient utilization of by-products and waste fines, potentials were
conducted on partial substitution of cement with biomass lignin from the pulp and paper
industry [42, 43]. The replacement of cement with biomass lignin up to 25% has
demonstrated good mechanical strength and enhanced the reducibility of the briquettes. The replacement of cement with lignin has increased the total carbon content while the
sulfur content did not significantly change in the briquettes. Novel iron ore carbon
composites were suggested to enhance the reduction rate of iron ore in the conventional
BF and also alternative ironmaking processes (e.g., rotary hearth furnace and rotary kiln
furnace) [45, 46]. Such composites are very attractive to the aspects of energy saving and
environmental protection. The studies carried out on iron ore-carbon composite have
demonstrated higher reduction rate and lower coke consumption compared to the ordinary
burden materials [47-52]. It was demonstrated that the starting temperature of carbon
solution-loss reaction (Boudouard reaction) decreased as the degree of contact between
iron ore and carbonaceous material increased [52, 53]. The iron ore carbon composites
are considered as a type of micro-reactors [54]. These composites are highly reactive due
to the close contact between Fe2O3 and carbon particles, which enhances the reduction
process [55, 56]. The CO2 generated from the reduction of iron oxide (e.g., FeO + CO =
Fe + CO2) and CO generated from the Boudouard reaction can be exchanged quickly in
vacancies inside such type of micro-reactors. The trials in the large BF using cement-
bonded self-reducing pellets (SRP) were demonstrated with a maximum rate of 54 kg-
SRP/tHM [57]. Utilization of pre-reduced agglomerate (PRA) has recently been
developed instead of conventional sinter to reduce the coke consumption and CO2
emissions in the BF. The new process is based on a partial reduction of sinter during the
sintering process to produce PRA as can be seen in Fig. 7 [58]. Fig. 7. Comparison of conventional and new agglomeration process [94]. Fig. 7. Comparison of conventional and new agglomeration process [94]. E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 79 It was reported that charging of PRA with a reduction degree of about 70% could
reduce the CO2 emission from BF by ~10%, as shown in Fig. 8 [58]. Cold Bonded Briquettes and Pre-reduced Agglomerates Other work was
directed to use non-coking coal with high ash and high volatile matter content for pre-
reduction of iron ore nuggets [59]. As the metallic iron content in pre-reduced nuggets
increases, the coke consumption and CO2 emission from BFs decrease. Fig. 8. Relationship between the reduction degree of PRA and CO2 emission [94]. Fig. 8. Relationship between the reduction degree of PRA and CO2 emission [94]. Both cold bonded briquettes and self-reduced pellets have the potential to enhance
the recycling rate, and it can efficiently participate in charging of pre-treated biomass to
the BF. However, the deterioration of the mechanical strength by adding the biomass, the
higher need for cement and the disintegration of such agglomerates by increasing the
temperature in the BF shaft are still the main challenges limited its positive impact. Our
recent research has discussed the effect of charging briquettes containing torrefied
sawdust on the BF efficiency and process stability. The charging of bio-briquettes (39-64
kg/tHM) was demonstrated a positive impact on reducing coke consumption and CO2
emission [60]. Renewable Biomass Increasing attention was recently paid on using renewable biomass as a source of
heating and reducing agent instead of fossil fuels to mitigate the fossil CO2 emission in
ironmaking [61]. Besides the environmental benefits related to the photosynthesis effect,
the use of biomass and especially products of their thermal treatment will provide
technological and economic advantages due to their unique properties represented in:
very low sulphur content, high ratio of carbon to ash, relatively few and unreactive
inorganic impurities as well as stable pore structure and high specific surface area [62-
64]. On the other hand, the presence of harmful elements in some types of biomass must
be controlled because it could negatively affect the hot metal quality and production
operation [65]. Low density and high affinity to absorb moisture from the atmosphere are
also problematical for biomass utilization in the iron and steel industry. The results carried
out using life cycle assessment methodology indicated that the use of charcoal in the
integrated steelmaking route has the potential to reduce the greenhouse gas (GHG)
footprint of steel by 0.69–1.21 t CO2/t steel without any credits, and by 0.91–1.61 t CO2/t
steel with including by-product credits as given in Table 1 [66-68]. There is a strong Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 80 potential for biomass to be used in the production of bio-coke, bio-sinter, bio-composites,
replacement of nut coke, and replacement of pulverized coal in BF [69, 70]. It was
reported that substitution of 20% of coke in BF with biomass could reduce the CO2
emission by 300 kg/tHM, which represents a 15% reduction in the total CO2 emission. Table 1. Typical carbon addition rates in steelmaking and expected reduction in
CO2 emissions [67]. Application and
replaced carbon
source
Typical
addition,
kg/tHM
Charcoal
substitution
rate, %
Charcoal
amount,
kg/tHM
Net emission reduction
t-CO2/t-
Crude
steel
% of CO2
Emissions
Sintering solid fuel
76.5-102
50-100
38.3-102
0.12-0.32
5-15
Cokemaking
480-560
2-10
9.6-56
0.02-0.11
1-5
BF tuyere fuel
injection (PC)
150-200
0-100
0-200
0.41-0.55
19-25
BF nut coke
replacement
45
50-100
22.5-45
0.08-0.16
3-7
BF carbon/ore
briquette
10-12
0-100
0-12
0.06-0.12
3-5
Steelmaking
recarburiser
0.25
0-100
0-0.25
0.001
0.04
Totals
761.75-
919.25
0-100
70.4-415.25 0.69-1.25
31.57 Table 1. Typical carbon addition rates in steelmaking and expected reduction in
CO2 emissions [67]. Renewable Biomass The top charged bio-agglomerates (bio-coke, bio-sinter, and bio-composite)
should be able to sustain the requirements of mechanical strength for BF; otherwise, it
will deteriorate the process efficiency [61]. Also, any change in the BF input materials
will directly affect the mechanism of mass and heat transfer, temperature profile and gas
distribution inside the BF from raceway zone to the upper shaft. Intensive investigations
were carried out to estimate the influence of bio-materials on the BF process. It was
reported that charcoal could replace up to 20% of coke in the BF [71-73]. Charging of
20 kg/tHM of charcoal could reduce the coke consumption by 30 kg/tHM due to
decreasing of the TRZT [74, 75]. The porous structure of bio-sinter was demonstrated a
higher reduction rate compared to that of conventional sinter [76]. Although the nut coke
in the iron ore burden layers could be entirely replaced by small size charcoal, the low
density and low strength of charcoal could lead to a segregation and dust generation in
the BF shaft. Iron ore-biomass composite is another option to reduce the energy consumption
and CO2 emission in the BF. These composites could enhance the reduction of iron oxide
with closely adjacent carbon [52]. Recently, great attention was paid to investigate the
influence of iron ore-carbon composite on the BF operation [77-81]. Hematite is
completely reduced with woody biomass at temperature ≥ 1000 °C [82]. The reduction
rate increased with temperature which attributed not only to the endothermic nature of
the reduction reaction but also due to the faster decomposition of hydrocarbons (e.g., 81 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … CH4) into fresh reducing gases (CO and H2). A full reduction of iron ore to metallic iron
was achieved by 30 wt.% of sawdust [83]. Although the biomass seems like an efficient
reducing agent for iron ore in lab-scale tests, the practical application in large BF is still
limited. The main reason is the low crushing strength of such iron ore-biomass
composites. The weak strength of iron ore-biomass composite compared to iron ore-coal
composite is attributed to the absence of coal thermal plasticity. The thermal plasticity of
coal provides the proper mechanical strength of iron ore-coal composites. Also, the
volatile matter of biomass is often escaped at relatively low temperature before starting
the reduction of iron oxides. Development in Tuyeres Injections Besides the development of the top charging system and top charging materials,
tremendous modifications were done in the lower part of the BF shaft. Maximizing the
auxiliary fuels injection into the BF becomes one of the vital options to lower the
operating cost and maximize productivity. Usage of cheaper fuels and H2 rich materials
for partial substitution of metallurgical coke can be effectively reduced the CO2 emission. The next sub-sections will thoroughly discuss the tuyere injection of different auxiliaries’
materials into the BF. Renewable Biomass For this reason, a special design of iron ore-biomass
composite is still required to enhance the utilization of volatile biomass matter in the
reduction of iron oxides. Further development and modifications are still required to enhance the application
of iron ore-biomass composites in the modern BF. In order to overcome the obstacle of
biomass top charging, the tuyeres injection provides a flexible option for biomass
utilization in the large modern BF. The next section will discuss the development of
tuyeres injections. Pulverized Coal Injection (PCI) PCI is one of the essential methods to enhance BF profitability. The majority of
BFs all over the world apply PC due to the relatively lower cost of coal compared to other
fuels beside the beneficial effect on the BF efficiency. Injection of PC into the BF
provides various economic and operational benefits [84]. These benefits are including (i)
lower consumption of expensive coke, (ii) replacing high-rank expensive coal with low
grade cheaper coals, (iii) longer life period of coke oven, (iv) higher BF productivity, (v)
higher flexibility in BF operation, (vi) improving the hot metal quality, and (vii) lower
CO2 emissions. Recently, stable BF operation with PCI rates of around 200 kg/tHM and
coke rate below 300 kg/tHM was successfully maintained. The replacement ratio of PC
for coke is about 1.0 at the injection rate up to 200 kg/tHM [85]. At higher PCI (>200
kg/tHM), there is a reduction in the incremental replacement ratio of PC for coke. In
contrast with other fuels which could be injected through tuyeres, PC has the minimum
negative impact on the raceway flame temperature and therefore, it has the highest
possible injection rate than other fuels. The combustion of PC in the raceway is a complex
process that requires high control of various parameters. Efficient PCI is obtained by
convenient coal treatment, including the control of moisture content and grain size. The
proper mixing of PC with the hot blast, uniform distribution of PC across all tuyeres, low
N2 input into the BF and maximizing retention time of coal in the tuyeres are necessary
to achieve high combustion efficiency. The injection of coal through the tuyeres is
certainly not a new technology, but it was recently developed to sustain an optimum Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 82 condition. The enriched oxygen hot blast (1100-1250 °C) is injected into the blowpipe at
velocity 100-200 m/s. The coal/gas mixture (e.g., 150 kg coal/tHM) is injected by lance
into the blowpipe at a rate equal of 150 g coal/m3 of the blast. The combustion of the PC takes place in three main steps. In the first step, the
volatile matters of coal are released and combusted with the surrounding oxygen. This
volatiles generates different gases including N2, CO2, H2O and small portions of CO, H2,
and CxHy. Pulverized Coal Injection (PCI) In the second step, the combustion of residual char takes place, which has a
slower rate than the combustion of volatiles. In the third step, ash is released after the
complete combustion of char [86, 87]. Fig. 9 illustrates a schematic view of the tuyeres
and different reaction zones in the raceway during PC injection [88]. The raceway
consists of: (i) PC de-volatilization zone, (ii) oxidation zone and (iii) solution loss reaction
zone. Fig. 9. The schematic diagram for PC reaction in raceway [87, 88]. Fig. 9. The schematic diagram for PC reaction in raceway [87, 88]. E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 83 The concentration of oxygen is sharply decreasing in the oxidation zone due to its
reaction with the carbon of coke and coal to produce CO2 as given in Eq. 1. As the oxygen
concentration decreases, the solution loss reaction by CO2 or steam takes place, as given
in Eqs. 2 and 3, and the concentration of CO increases while CO2 decreases. Therefore,
the reaction rate of PC is dependent on the oxygen content in the hot blast. The PC
injection rate, blast condition (temperature, pressure, moisture, and oxygen content), coal
properties (carbon content, ash content, VM content, particle size), lance construction and
properties of top charged coke are important factors affecting the combustion efficiency
of coal in the raceway. y
p
g
in Eqs. 2 and 3, and the concentration of CO increases while CO2 decreases. Therefore,
the reaction rate of PC is dependent on the oxygen content in the hot blast. The PC
injection rate, blast condition (temperature, pressure, moisture, and oxygen content), coal
properties (carbon content, ash content, VM content, particle size), lance construction and
properties of top charged coke are important factors affecting the combustion efficiency
of coal in the raceway. 𝐶(𝑠) + 𝑂2(𝑔) = 𝐶𝑂2(𝑔); ∆𝐻𝑜= −393.5 𝑘𝐽𝑚𝑜𝑙
⁄
1
𝐶(𝑠) + 𝐶𝑂2(𝑔) = 2𝐶𝑂(𝑔); ∆𝐻𝑜= +172.467 𝑘𝐽𝑚𝑜𝑙
⁄
2
𝐶(𝑠) + 𝐻2𝑂(𝑔) = 𝐶𝑂(𝑔) + 𝐻2(𝑔); ∆𝐻𝑜= +131.294 𝑘𝐽𝑚𝑜𝑙
⁄
3 𝐶(𝑠) + 𝑂2(𝑔) = 𝐶𝑂2(𝑔); ∆𝐻𝑜= −393.5 𝑘𝐽𝑚𝑜𝑙
⁄ 1 2 𝐶(𝑠) + 𝐻2𝑂(𝑔) = 𝐶𝑂(𝑔) + 𝐻2(𝑔); ∆𝐻𝑜= +131.294 𝑘𝐽𝑚𝑜𝑙
⁄ 3 The Raceway Adiabatic Flame Temperature (RAFT) is defined as the temperature
that the raceway gas reaches as soon as all C, O2, and H2O are converted to CO and H2. Pulverized Coal Injection (PCI) The RAFT is normally in the range of 1900-2300 °C based on PCI rate, PC quality, blast
temperature, blast moisture, and oxygen enrichment. The RAFT is theoretically
calculated from Eq. 4 [89] based on mass and heat balance. With increasing PCI, O2
enrichment is an important factor that affects the RAFT [89]. The higher PCI rate, the
higher O2 enrichment is required to sustain the heat balance in the upper and lower part
of the furnace. Too low RAFT results in very low top gas temperature, which will delay
the drying of the cold top-charged burden and consequently make the upper shaft less
efficient. On the other hand, too high O2 enrichment in the blast is accompanied by
relatively high RAFT, which will result in an erratic descend of the burden. Therefore,
the oxygen content in the hot blast has to be optimized very carefully to reach the proper
RAFT and shaft efficiency. 𝑅𝐴𝐹𝑇= 1489 + 0.82 ∙𝐵𝑇−5.705 ∙𝐵𝑀+ 52.778 ∙𝑂𝐸−18.1 ∙
𝐶𝑜𝑎𝑙
𝑊𝐶∙100 − 𝑅𝐴𝐹𝑇= 1489 + 0.82 ∙𝐵𝑇−5.705 ∙𝐵𝑀+ 52.778 ∙𝑂𝐸−18.1 ∙
𝐶𝑜𝑎𝑙
𝑊𝐶∙100 −
43.01 ∙
𝑂𝑖𝑙
𝑊𝐶∙100 −27.9 ∙
𝑇𝑎𝑟
𝑊𝐶∙100 − 50.66 ∙
𝑁𝐺
𝑊𝐶∙100
4 43.01 ∙
𝑂𝑖𝑙
𝑊𝐶∙100 −27.9 ∙
𝑇𝑎𝑟
𝑊𝐶∙100 − 50.66 ∙
𝑁𝐺
𝑊𝐶∙100 4 Where; BT: Blast Temperature, °C; BM: Blast moisture, g/m3 STP dry blast; OE:
Oxygen enrichment (%O2-21); Oil: dry oil injection rate, kg/tHM; Tar: dry tar injection
rate kg/tHM; Coal: dry coal injection rate, kg/tHM; NG: Natural gas injection rate,
kg/tHM; WC: Wind consumption, m3/tHM. The transit time of the injected coal particles in the raceway is in range of 20-30
ms [90]. Within such limited time, it is crucial to assure that most of the coal is gasified
before escaping the raceway region; otherwise, it will be swept into the coke bed and
consequently deteriorates the permeability of active coke zone [90, 91]. Fig. 10
summarizes the various problems which could be occurred due to the high injection rate
of PC [87, 88]. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 84 Fig. 10. Expected technical issues and countermeasure at a high rate of PCI into the
blast furnace [87]. Fig. 10. Expected technical issues and countermeasure at a high rate of PCI into the
blast furnace [87]. Oil and Natural Gas Injection j
The injection of oil and natural gas (NG) into the BF were firstly implemented
before PC, but the energy crisis during the 1970s resulted in more attention to PCI. Nowadays, the countries which are rich with oil and NG are still injecting these fuels into
the BF to reduce coke consumption. The injection of 1.0 ton of oil or NG can replace 1.2
tons of coke [94, 95]. The average consumption of NG is 70-100 m3/tHM (~50-70
kg/tHM) but often exceeds to 150-170 m3/tHM (~105-120 kg/tHM). The operation of
BFs using NG and oxygen-enriched blast is called composite blast technology. With oxy-
oil technology, the amount of injected oil could reach a level of 180 m3/tHM (130
kg/tHM). Oil and NG are decomposed and combusted in the BF raceway. The
decomposition and combustion of hydrocarbons generate CO/CO2 and H2/H2O. The CO2
and H2O are in-situ react with the carbon of coke and PC or with hydrocarbons of NG to
finally generate CO and H2. The participation of hydrogen in the reduction of iron oxide
leads to a decrease of CO2 emissions. The heavy oil and NG have a higher percentage of
hydrocarbons compared to coal/coke and thereby the endothermic effect of these
hydrocarbons results in a higher drop in RAFT [10]. It was estimated that the RAFT
would be decreased by 350-450 °C per 100 m3/tHM of NG injection, 300-350 °C per 100
kg/tHM of heavy oil injection, 200-250 °C per 100 m3/tHM of coke oven gas injection,
80-120 °C of low volatile coal injection and 150-220 °C of high volatile coal injection
[10]. Recent studies indicated that the rate of NG injection into the BF could be increased
significantly by injecting part of the NG in the shaft of the furnace. This will reduce the
endothermic effect of NG injection on the RAFT and the total oxygen consumption would
be slightly lower compared with the tuyere-only injection [96]. A mathematical model
based on the heat balance in the bottom segment of the BF was developed to calculate the
maximum injected amount of NG (V𝑁𝐺
𝑚𝑎𝑥) to attain the minimum value of the theoretical
flame temperature, minimum possible coke consumption (𝐶min ) and the minimum direct
reduction (rd𝑚𝑖𝑛) [97, 98]. Pulverized Coal Injection (PCI) The fine char particles tend to block the bed voids and consequently disturb the
gas flow and increase the pressure drop inside the furnace, reduce the permeability of
liquids and gases in the cohesive zone and reduce the shaft efficiency and the furnace
productivity [92, 93]. Coals with higher volatile matter (VM) content exhibit superior
combustibility in raceway due to the higher reactivity, however coals with very high VM
contain less carbon and hence the combustion efficiency in the raceway will be reduced
[11]. Moreover, at high PCI, sulfur and phosphorous should be kept as low as possible to
avoid the negative effect on the hot metal quality. The PCI is considered one of the recent
significant modifications in the BF technology which provided good opportunity to
reduce the load on cokemaking, utilize poor quality coal resources, and to reduce the
overall costs. One of the main challenges is the decreasing of coke-to-coal replacement
ratio by increasing the PCI rate higher than 200 kg/tHM. Further research and
development on PCI technologies and understanding the behavior of PC at high injection
rate (200-300 kg/tHM) to the BF are still needed. 85 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … Oil and Natural Gas Injection It was reported that the minimum coke consumption will be
obtained when the coke carbon which is used as a source of heat is equal to the coke
carbon which is consumed in the reduction processes [97]. Based on the developed model,
rd𝑚𝑖𝑛, 𝐶min and V𝑁𝐺
𝑚𝑎𝑥 can be theoretically calculated from Eqs. 5-7; respectively. 𝑟𝑑𝑚𝑖𝑛=
1
𝜂̅𝑐𝑜−𝑛 (2
𝜂̅𝐻2
𝜂̅𝐶𝑂−
𝑄̅𝐶𝐻4
𝑄̅𝐶∅
+1)− 𝐶𝑖
ℎ
1
𝜂̅𝑐𝑜+ ∆𝐻𝑇1
𝑄̅𝐶∅
+1
𝐶𝑚𝑖𝑛=
1−𝑟𝑑𝑚𝑖𝑛
𝜂̅𝑐𝑜
−2𝑛
𝑉𝑁𝐺
𝑚𝑎𝑥=
22.4𝐹𝑒𝑒
𝑀𝐹𝑒(2 1
𝜂𝐶𝑂−𝐶𝑖
ℎ)
2+
𝜂̅𝐻2
𝜂̅𝑐𝑜−
𝑄𝐶𝐻4
𝑄𝐶∅
+1
[𝑚3(𝐶𝐻4)𝑡−1𝐻𝑀−1] 𝑟𝑑𝑚𝑖𝑛=
1
𝜂̅𝑐𝑜−𝑛 (2
𝜂̅𝐻2
𝜂̅𝐶𝑂−
𝑄̅𝐶𝐻4
𝑄̅𝐶∅
+1)− 𝐶𝑖
ℎ
1
𝜂̅𝑐𝑜+ ∆𝐻𝑇1
𝑄̅𝐶∅
+1
𝐶𝑚𝑖𝑛=
1−𝑟𝑑𝑚𝑖𝑛
𝜂̅𝑐𝑜
−2𝑛
𝑉𝑁𝐺
𝑚𝑎𝑥=
22.4𝐹𝑒𝑒
𝑀𝐹𝑒(2 1
𝜂𝐶𝑂−𝐶𝑖
ℎ)
2+
𝜂̅𝐻2
𝜂̅𝑐𝑜−
𝑄𝐶𝐻4
𝑄𝐶∅
+1
[𝑚3(𝐶𝐻4)𝑡−1𝐻𝑀 𝑟𝑑𝑚𝑖𝑛=
1
𝜂̅𝑐𝑜−𝑛 (2
𝜂̅𝐻2
𝜂̅𝐶𝑂−
𝑄̅𝐶𝐻4
𝑄̅𝐶∅
+1)− 𝐶𝑖
ℎ
1
𝜂̅𝑐𝑜+ ∆𝐻𝑇1
𝑄̅𝐶∅
+1
5
𝐶𝑚𝑖𝑛=
1−𝑟𝑑𝑚𝑖𝑛
𝜂̅𝑐𝑜
−2𝑛
6
𝑉𝑁𝐺
𝑚𝑎𝑥=
22.4𝐹𝑒𝑒
𝑀𝐹𝑒(2 1
𝜂𝐶𝑂−𝐶𝑖
ℎ)
2+
𝜂̅𝐻2
𝜂̅𝑐𝑜−
𝑄𝐶𝐻4
𝑄𝐶∅
+1
[𝑚3(𝐶𝐻4)𝑡−1𝐻𝑀−1]
7 𝑟𝑑𝑚𝑖𝑛=
1
𝜂̅𝑐𝑜−𝑛 (2
𝜂̅𝐻2
𝜂̅𝐶𝑂−
𝑄̅𝐶𝐻4
𝑄̅𝐶∅
+1)− 𝐶𝑖
ℎ
1
𝜂̅𝑐𝑜+ ∆𝐻𝑇1
𝑄̅𝐶∅
+1 5 𝑤ℎ𝑒𝑟𝑒, 𝜂̅𝑐𝑜: 𝑑𝑒𝑔𝑟𝑒𝑒 𝑜𝑓 𝐶𝑂 𝑢𝑡𝑖𝑙𝑖𝑧𝑎𝑡𝑖𝑜𝑛 𝑓𝑜𝑟 𝐹𝑒𝑂 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛; 𝜂̅𝐻2: 𝑑𝑒𝑔𝑟𝑒𝑒 𝑜𝑓 𝐻2 𝑢𝑡𝑖𝑙𝑖𝑧𝑎𝑡𝑖𝑜𝑛
𝑓𝑜𝑟 𝐹𝑒𝑂 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛; 𝑄̅𝐶𝐻4: ℎ𝑒𝑎𝑡 𝑔𝑖𝑣𝑒𝑛 𝑏𝑦 𝑡ℎ𝑒 𝑐𝑜𝑚𝑏𝑢𝑠𝑡𝑖𝑜𝑛 𝑜𝑓 1 𝑚𝑜𝑙𝑒 𝑜𝑓 𝐶𝐻4 𝑎𝑡 𝑡ℎ𝑒𝑟𝑚𝑎𝑙
𝑟𝑒𝑠𝑒𝑟𝑣𝑒 𝑧𝑜𝑛𝑒 𝑖𝑛 𝑡ℎ𝑒 𝑓𝑢𝑟𝑛𝑎𝑐𝑒 (900 ℃); 𝑄̅𝐶∅: ℎ𝑒𝑎𝑡 𝑔𝑖𝑣𝑒𝑛 𝑏𝑦 𝑏𝑢𝑟𝑛𝑖𝑛𝑔 𝑜𝑓 𝑐𝑜𝑘𝑒 𝑐𝑎𝑟𝑏𝑜𝑛 𝑎𝑡
𝑡ℎ𝑒𝑟𝑚𝑎𝑙 𝑟𝑒𝑠𝑒𝑟𝑣𝑒 𝑧𝑜𝑛𝑒 𝑖𝑛 𝑡ℎ𝑒 𝑓𝑢𝑟𝑛𝑎𝑐𝑒 (900 ℃); ∆𝐻𝑇1: 𝑒𝑛𝑡ℎ𝑎𝑙𝑝𝑦 𝑜𝑓 𝑑𝑖𝑟𝑒𝑐𝑡 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛
𝑜𝑓 𝑖𝑟𝑜𝑛 𝑜𝑥𝑖𝑑𝑒; 𝑀𝐹𝑒: 𝑚𝑜𝑙𝑎𝑟 𝑚𝑎𝑠𝑠 𝑜𝑓 𝐹𝑒; 𝐹𝑒𝑒: 𝑖𝑟𝑜𝑛 𝑐𝑜𝑛𝑡𝑒𝑛𝑡 𝑖𝑛 ℎ𝑜𝑡 𝑚𝑒𝑡𝑎𝑙. 𝑤ℎ𝑒𝑟𝑒, 𝜂̅𝑐𝑜: 𝑑𝑒𝑔𝑟𝑒𝑒 𝑜𝑓 𝐶𝑂 𝑢𝑡𝑖𝑙𝑖𝑧𝑎𝑡𝑖𝑜𝑛 𝑓𝑜𝑟 𝐹𝑒𝑂 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛; 𝜂̅𝐻2: 𝑑𝑒𝑔𝑟𝑒𝑒 𝑜𝑓 𝐻2 𝑢𝑡𝑖𝑙𝑖𝑧𝑎𝑡𝑖𝑜𝑛
𝑓𝑜𝑟 𝐹𝑒𝑂 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛; 𝑄̅𝐶𝐻4: ℎ𝑒𝑎𝑡 𝑔𝑖𝑣𝑒𝑛 𝑏𝑦 𝑡ℎ𝑒 𝑐𝑜𝑚𝑏𝑢𝑠𝑡𝑖𝑜𝑛 𝑜𝑓 1 𝑚𝑜𝑙𝑒 𝑜𝑓 𝐶𝐻4 𝑎𝑡 𝑡ℎ𝑒𝑟𝑚𝑎𝑙
𝑟𝑒𝑠𝑒𝑟𝑣𝑒 𝑧𝑜𝑛𝑒 𝑖𝑛 𝑡ℎ𝑒 𝑓𝑢𝑟𝑛𝑎𝑐𝑒 (900 ℃); 𝑄̅𝐶∅: ℎ𝑒𝑎𝑡 𝑔𝑖𝑣𝑒𝑛 𝑏𝑦 𝑏𝑢𝑟𝑛𝑖𝑛𝑔 𝑜𝑓 𝑐𝑜𝑘𝑒 𝑐𝑎𝑟𝑏𝑜𝑛 𝑎𝑡
𝑡ℎ𝑒𝑟𝑚𝑎𝑙 𝑟𝑒𝑠𝑒𝑟𝑣𝑒 𝑧𝑜𝑛𝑒 𝑖𝑛 𝑡ℎ𝑒 𝑓𝑢𝑟𝑛𝑎𝑐𝑒 (900 ℃); ∆𝐻𝑇1: 𝑒𝑛𝑡ℎ𝑎𝑙𝑝𝑦 𝑜𝑓 𝑑𝑖𝑟𝑒𝑐𝑡 𝑟𝑒𝑑𝑢𝑐𝑡𝑖𝑜𝑛
𝑜𝑓 𝑖𝑟𝑜𝑛 𝑜𝑥𝑖𝑑𝑒; 𝑀𝐹𝑒: 𝑚𝑜𝑙𝑎𝑟 𝑚𝑎𝑠𝑠 𝑜𝑓 𝐹𝑒; 𝐹𝑒𝑒: 𝑖𝑟𝑜𝑛 𝑐𝑜𝑛𝑡𝑒𝑛𝑡 𝑖𝑛 ℎ𝑜𝑡 𝑚𝑒𝑡𝑎𝑙. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 86 Off-Gases Injection Efficient utilization of integrated steel plant top-gases becomes necessary to
achieve profitability to steel mill operation from both economic and environmental
aspects. The major fuel gases that could be recovered in the integrated steelworks are
including BF gas (BFG), coke oven gas (COG), and basic oxygen furnace gas (BOFG). The chemical compositions of these gases are given in Table 2 [99]. p
g
g
The off-gas from the BF is the lowest from heat content and the highest from the generated
amount in an integrated steel plant. Top Gas Recycling BF (TGR-BF) technology was
studied in one of the largest European project, namely Ultra Low CO2 Steel (ULCOS)
[100, 101]. The four breakthrough technologies identified were: (i) Top Gas Recycling
BF (TGR-BF) with CO2 Capture and Storage (CCS); (ii) HIsarna Smelting Reduction
with CCS; (iii) Ultra Low CO2 direct Reduction (ULCORED) with CCS; and (iv)
Electrolysis. TGR-BF aimed at lowering the usage of fossil carbon (coke) via re-usage of
the reducing agents (CO and H2) after the removal of the CO2 from the BF top gas. In
order to reach the 50% CO2 reduction, the application of CO2 capture and storage (CCS)
technology is necessary. Heat and mass balance models were used to identify the carbon
consumption at different hot and cold TGR as given in Fig. 11 [102]. Table 2. Typical chemical composition of top-gases in the integrated steel mill [99]. Table 2. Typical chemical composition of top-gases in the integrated steel mill [99]. Type of
gas
H2
CO
CH4
C2H6
CO2
N2
MJ/Nm3
Nm3/t
vol.%
COG
63
5.8
22
2.9
2.0
4
17.9
410-560
BOFG
-
55
-
-
15
30
8.95
50-100
BFG
2.6
20.3
-
-
24.3
52.8
3.37
1414-1500
Fig. 11. Heat and mass balance model for TGR-BF at [102]: (a) TGR hot at the stack
and cold at main tuyeres, b) TGR hot at main tuyeres, c) TGR hot at the stack and main
tuyeres. Fig. 11. Heat and mass balance model for TGR-BF at [102]: (a) TGR hot at the stack
and cold at main tuyeres, b) TGR hot at main tuyeres, c) TGR hot at the stack and main
tuyeres. 87 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … The highest carbon saving (25.1%) is achieved by hot TGR at stack and tuyeres
injection. Off-Gases Injection Trials were conducted at LKAB s’ Experimental BF (EBF) in Luleå, Sweden
[103, 104]. The EBF was modified and a gas separation plant based on VPSA technology
was built near the EBF. Although the tests at the EBF are considered successful, the
industrialization of the ULCOS-BF technology requires an additional scale-up step. The
COG has the largest net calorific value in the range of 16.4-18 MJ/Nm3 (STP) compared
to that of either BOFG (~8.8 MJ/Nm3) or BFG (3.0-3.7 MJ/Nm3) [105, 106]. The specific
amount of generated coke oven gas is in the range of 410-560 Nm3/t of coke while the
amount of BOFG is in the range of 50-100 Nm3/t of steel in the suppressed combustion
system, and it could reach 500-1000 Nm3/t of steel in the open combustion system. The
annual worldwide metallurgical coke production is more than 500 Mt which is produced
from about 650 Mt of coking coals [107]. The cokemaking process is accompanied by
more than 310 billion Nm3 of COG [108]. The COG is currently used in the heating of
BF stoves, ignition furnaces in sintering plant, heating furnace in rolling mills and electric
power generation in power plant [105, 106]. p
g
p
p
The estimations which were carried out on optimizing the energy consumption in
the integrated iron and steel works indicated that the utilization of COG for power
generation is not always the optimal credits [105]. The composition of COG, which is
rich with hydrogen, is attracting much attention for its utilization in the reduction
processes [109-112]. The flexibility of COG utilization in the integrated steel plant for
DRI production through the addition of Midrex process is expected to be also an efficient
method that can introduce many benefits to the integrated steel mill [106]. The merging
of the “traditional” ironmaking BF with the “alternative” direct and smelting reduction
processes is an innovative approach to achieve the economic and environmental targets
of higher production and lower emissions. The recent studies which were carried out to
evaluate the reduction potential of original coke oven gas (OCOG) and reformed coke
oven gas (RCOG) for direct reduction of iron ore pellets compared to reformed natural
gas (RNG) indicated the high efficiency of these gases for DRI production [113-116]. Waste Plastics and Biomass Injection Recycling and utilization of waste plastics have received increasing interest in
recent decades. The majority of this material is currently being landfilled or incinerated,
which causes environmental problems. The combustion of waste plastics generates
hazardous pollutants, such as dioxins/furans, as well as undesirable carbon dioxide. The
waste plastics can be efficiently utilized in BFs as an alternative reducing agent and as a
source of heat to reduce the coke consumption. As the plastics contain hydrogen and
carbon it can also be provided additional benefits similar to oil and NG injection into the
BF. Due to the difficulty of waste plastics crushing, coarse particles were preferred to
inject into the furnace; however, the fine particles are better in combustion in the raceway. Therefore, a technology for fine pulverization of waste plastics was developed and
implemented in a full-scale industrial BF in Japan. Since the collected waste plastics are
heterogeneous mixture form different types, it is recommended to conduct heat treatment
at 200 °C before its injection into the BF. The pre-treatment will generate a homogenous
pulverized waste plastic mixture [123]. Moreover, the pre-treatment of waste plastics will
perform de-chlorination for waste plastics which contain chlorine such as polyvinyl
chloride (PVC) and hence avoid the corrosive effect on the BF tuyeres and the refractory
materials in hearth lining. The theoretical limit of waste plastics injection is estimated to
be 70 kg/tHM while the higher injection rate will result in problems similar to that
obtained with the relatively high PCI. Every ton of plastics used in the BF can replace
750 kg of coke [124]. Injection of waste plastics into the BF could reduce the CO2
emissions by 30% due to the higher H2 content [125]. On the other hand, the cost of waste
plastic collection and treatment is still representing one of the main challenges for its
implementation in ironmaking. Efficient and effective systems are required to secure a
sustainable and reliable supply of waste plastic into the BF. pp y
p
Renewable biomass represents one of the vital options to mitigate CO2 emissions in the
steel industry. The recent analysis of the long-term potential for renewable energy in
industrial applications suggested that up to 21% of total energy use in the manufacturing
industry in 2050 could be covered from renewable origin materials [126]. Off-Gases Injection Optimization of the reforming process of coke oven gas is crucial to avoid the severe
crack of pellets in the case of reduction with RCOG and the carbon deposition in the case
of OCOG. The reduction potential of COG with/without mixing with converter gas
(BOFG) was studied for natural lump ore at 800-1050 °C [116, 117]. It was reported that
the highest reduction rate was achieved with RCOG followed by OCOG. The reduction
rate was decreased in the order of RCOG>OCOG>RNG>OCOG-BOFG>RCOG-BOFG
at 900-1050 °C. The injection of COG into the BF through tuyeres influences the raceway
conditions and iron ore reduction. The combustion of COG hydrocarbons in the front of
tuyeres by blast oxygen results in the generation of carbon monoxide and hydrogen gases,
which increase the potential of reducing gas on account of inert N2 [118]. The theoretical
calculation and commercial trails, which carried out on the replacement of NG with coke
oven gas in BF showed lower coke consumption and higher hot metal production [119]. The high efficiency of COG is because it contains 3.5-4.0 times less hydrocarbons
compared to that of NG [10]. This improves the combustion in the tuyere hearth, activate
coke column, and increase gases utilization in the furnace. The injection of COG into the
BF is practiced in some countries with different injection rates from about 30 to 280
m3/tHM [120, 121]. It was reported that the maximum level of COG injection into the BF
is 100 kg COG/tHM according to the thermochemical conditions while the replacement
ratio is 0.98 ton of coke/t of COG [122]. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 88 Waste Plastics and Biomass Injection This will
compensate for about 50 exajoules (1018 J) per year (EJ/year) from the total energy (230
EJ/year) used in the industrial sectors. Utilization of renewable biomass in the industrial
sector has the potential to reduce GHG emissions by 10% in 2050 [127]. As the top
charging of biomass into the modern BF still suffers from some problems which are
related to the lower mechanical strength and the high volatile matter compared to coke,
the tuyeres injection provides a flexible option for biomass utilization. In this case the
mechanical strength is not required; however, the combustion behavior of biomass is a
very important factor affecting the RAFT. A static heat and mass balance model was
applied to investigate the substitution rate of biomass products (charcoal, torrefied
material, and wood pellets) for PCI [128, 129]. It was found that 166.7 kg/tHM charcoal
could fully replace 155 kg/tHM PCI. The torrefied material and wood pellets could be
partially replaced PCI by 22.80% and 20% respectively [129]. The reactive structure of
charcoal improves the combustion process in raceway to become comparable to that of
coal [130]. It was reported that the maximum injection rate of charcoal would be in the
range of 200-220 kg/tHM which could reduce the net CO2 emissions by 40% [131]. Besides the environmental benefits, the injection of charcoal will result in a lower amount
of slag and higher production rate due to the low sulfur and ash content and higher content
of CaO compared to PCI [132]. Although the investigations and the commercial trails E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … 89 conducted on biomass injection into the BF demonstrated many benefits, its application
is not practiced yet in the large modern BFs. The practical limitations could be
summarized in four main points [131]: (i) less calorific value of biomass products
compared to coal which required efficient pre-treatment and pyrolysis, (ii) difficulties in
biomass injection at high rate due to the porous nature and the low density which required
an optimization for the injection process, (iii) wider particle size distribution of biomass
after grinding which required an efficient sieving to get the proper particle size for
injection and (iv) higher alkalis in biomass products which should be controlled and
minimized before utilization to avoid its negative impact on the refractory materials. Waste Plastics and Biomass Injection Also,
the higher cost of biomass products compared to that of fossil fuels represents another
challenge for its implementation. The availability and characteristics of different sources
of biomass and the suitable pretreatment technologies for their upgrading are discussed
and evaluated elsewhere [132]. Potentials of Waste Heat Recovery (WHR) Within the steel industry, 20-60% of the primary energy is transformed into waste
heat. These waste heats are carried in different sources, such as flue gas, water/liquid
streams and hot solid at different temperature levels. Some amounts of waste heat can be
utilized within the process, or power plant, while the rest is flared or lost. New
technologies are under development in order to economically maximize the recovery of
waste heat recovery. Analysis of Waste Heat in Steel Plant In some of the globally integrated steel plants, with total production around 60 Mt
of steel, the waste heat is varied from about 20% up to 60% as can be seen in Fig. 12(a)
[133]. The waste heat comes from heat conduction, convection, and radiation of hot
surfaces, heated products, and by-products. The waste heat carriers are mainly hot slag from BF and BOF, hot flue gases from
EAF, BOF, reheating furnaces and heat treatment furnaces, hot air from annealing furnace
and hot water from reheating furnace and annealing furnace as shown in Fig. 12(b) [133]. The highest temperature streams are demonstrated by BOF gas, EAF gas, and BF slag. It
can be seen that the heat in the iron and steel industry is mainly stored in hot products,
molten slag, and waste gas. About 55% of energy is stored in the hot products while
around 45% is lost as waste heat with solid by-products and exhaust gases [134]. Nowadays, three main sources of waste heat are commonly recovered in steel plants for
steam production; COG, BOF gas, and EAF gas. The produced steam can be used for
different internal usage in the plant and it can also be used for power generation for an
internal and external user. On the other hand, the waste heat recovery from reheating
furnaces and annealing furnaces are more challenging. Production of different grades of
steel is required to control and adjust the parameters of reheating and annealing to achieve
specific mechanical properties of steel. The fluctuations of production rate and product
type beside the relatively low temperature of streams are required type of highly flexible
heat recovery unit to adapt with these variations; otherwise, it will harm the operation
stability. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 90 Fig. 12. (a) Total energy used and waste heat in integrated steel plants [133];
(b) Typical temperatures and flow rates of waste heat sources in integrated
steel plant [134]. Fig. 12. (a) Total energy used and waste heat in integrated steel plants [133];
(b) Typical temperatures and flow rates of waste heat sources in integrated
steel plant [134]. Benefits and Challenges of WHR Waste heat recovery provides an attractive opportunity for getting energy from
waste heat, which is usually lost to the environment without any useful work. The basic
idea behind waste heat recovery is to try to recover maximum amounts of heat in the plant
and to reuse it as much as possible, instead of just flaring it into the surrounding
atmosphere. There is a great scope to recover waste heat from various industries by
installing waste heat recovery technologies. Although various technologies are
commercially available for waste heat recovery, the practical recovery of waste heat is
still low and it needs extensive research and development. The waste heat management
system is based on the new principal of Reduce, Recycle, and Recovery (RRR) [135,
136]. The process starts with minimizing the waste heat by upgrading the process
equipment and increasing the system efficiency. The released heat can be internally
recycled and reused in different purposes such as preheating furnaces and materials. Such
recycling process of waste heat is considered the most efficient method due to the lower
investment cost and higher energy efficiency. The last step is the recovery of extra heat
and its utilization either in steam production to achieve some mechanical work or power
generation to produce electricity. 91 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … Ways of recovery and reusing heat locally are varied from site to site. The
recovered heat can be used in preheating of combustion air or feed water in industrial
boilers and consequently reduce the energy consumption. Alternately, the heat exchanger
could be used to transfer heat from combustion exhaust gases to hot air which can be used
for drying oven or wet materials and subsequently reduce the energy consumption. Based
on this concept, many benefits could be achieved form efficient recovery and utilization
of waste heat. The benefits include a reduction in an operating cost, production cost, fossil
energy consumption, and waste emissions. On the other hand, the main barriers for the
implementation of waste heat recovery are represented mainly in long payback periods,
lack of viable end users, material constraints (e.g., corrosive or condensable materials in
the stream), and specific constraints (product control, inaccessibility and limited space)
[137, 138]. Benefits and Challenges of WHR Other barriers are related to the handling process of high-temperature gas
streams; for instance, loss of sensible heat due to the installation of scrubber for cleaning
of exhaust gas to meet the environmental regulation without connection to heat recovery
unit. Many factors are controlling the feasibility of waste heat recovery from which heat
quantity/quality, the chemical composition of hot streams, economy scale, accessibility,
and transportability [139, 140]. The recent analysis was conducted on the role of technology innovation in the
enhancement of energy efficiency and reduction of CO2 emissions in [141]. There are a
number of Best Available Technologies (BATs) which can be applied in many plants at
different production units and enable a significant reduction in energy and CO2 emissions. BATs have demonstrated different relevance of energy saving, as shown in Fig. 13. Fig. 13. Potentials of energy saving using BATs in the iron and steel industry [141]. The potential of energy saving at each unit will be achieved if BATs are applied at
possible sites. State-of-art power plant exhibited the highest potential for energy saving,
which could reach to 200 PJ. This can be attributed to the relatively low efficiency of the
currently applied heat conversion technologies at many sites. The average efficiency of 13
l
f
A
h
d
l
d
[141] Fig. 13. Potentials of energy saving using BATs in the iron and steel industry [141]. 13. Potentials of energy saving using BATs in the iron and steel industry [141]. The potential of energy saving at each unit will be achieved if BATs are applied at
possible sites. State-of-art power plant exhibited the highest potential for energy saving,
which could reach to 200 PJ. This can be attributed to the relatively low efficiency of the
currently applied heat conversion technologies at many sites. The average efficiency of Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 92 power generation from steam is about 32%, which is below the best practice and it could
be improved by the installation of efficient steam boiler and turbine technologies. Coke
dry quenching (CDQ) represents the second highest potential with energy saving around
85 PJ. In many cases, the hot coke at 1100 °C is cooled down by wet quenching which
results in loss of sensible heat. Benefits and Challenges of WHR The dry quenching using an inert gas such as nitrogen
enables the recovery of sensible heat which can be used for the production of high-
pressure steam or electricity. Other potentials of energy saving could be achieved through
efficient recovery and utilization of BOF gas, installation of continuous casting instead
of ingot casting, pre-heating of scrap before charging to EAF gas using hot off-gas,
sensible heat recovery from hot sinter and optimization of sinter/pellet ratio to reach
50/50. In BF the energy recovery is possible through heat recovery from hot stoves,
application of top gas pressure recovery turbine (TRT) and heat recovery from slag. Heat
recovery from BF stoves can be achieved through the implementation of recuperative,
steam production by a heat pump or drying of pulverized coal. Many of BFs are not
operate at high pressure, which makes the heat recovery from top gas somewhat difficult
and uneconomical. The gases leaving the top of the furnace are approximately 0.25-2.5
bar and have a temperature of around 200°C. This pressure and temperature are
comparatively quite low and make the investment in heat recovery quite difficult. Top
gas pressure recovery is suitable only for large modern BFs and high-temperature gases. On the other hand, waste heat recovery from BF slag represents the next frontier to
enhance energy efficiency and reduce emissions in the steel industry. WHR from BF Slag WHR from BF Slag WHR from BF Slag The global annual generation of BF slag is approximately 400 Mt. Slag is tapped
at 1450-1500 °C with heat content ~1.6-1.8 GJ/ton slag. Other slags come from
steelmaking BOF and EAF as can be seen in Table 3 [142]. Table 3. Types and characteristics of slags [142]. Table 3. Types and characteristics of slags [142]. Process
Average
temperature,
°C
Average
slag rate,
kg/tls
Estimated
slag
production
in 2008, Mt
Slag
enthalpy
GJ/t
Heat
value,
TWh/a
Hot metal
(BF)
1450
275
236
1.6
105
Steel
(BOF)
1300
126
112
1.3
40
Steel
(EAF)
1550
169
69
1.8
35 The waste heat recovery from slags could play a vital role in achieving the goals
of energy saving and the reduction of CO2 emission in the iron and steel industry. Slag
has a wide range of applications as it can be used in Portland cement, ceramics, concrete,
asphalt, roadbed materials, and construction aggregate. The efficiency of slag in these
different applications is mainly based on its cooling conditions and applied the cooling
process. Currently, wet granulation by water is the most common method used for quick
cooling of slag to produce a glassy product, namely ground granulated BF slag (GGBFS), 93 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … which is used to improve slag cement quality. In the wet granulation method, 1.0-1.2 ton
of water is required to granulate 1.0 ton of slag. The rapid cooling of molten slag in water
prevents the crystallization process of slag and breaks the slag into small particles. The
glassy phase in ground granulated BF slag can reach 95% using wet granulation method. Although the wet granulation works properly for glassy slag production, it seems
inefficient from economic and environmental aspects. Most of the sensible heat of slag is
lost and a large quantity of low-temperature steam generates which can be only used as
heat supply during the winter time in some countries. It causes environmental pollution
from the generated SO2 and H2S, also energy lost in the drying process of wet slag. Up to
now, there is no technology applied on an industrial scale to recover and utilize the waste
heat of molten slag all over the world. Dry cooling with energy recovery is recently
supposed to granulate slag and recover the waste heat [143]. Fig. WHR from BF Slag 14 illustrates a schematic
diagram of the traditional wet and innovative dry granulation of slag [143, 144]. The
granulated slag by fast dry cooling does not need any further drying, which could cut
30 kg CO2/t slag and reduce the annual CO2 emission by 6.3 Mt/a compared to wet
granulation [143]. There is a wide range of dry granulation and heat recovery techniques
which can be classified into physical and chemical methods. Fig. 14. The schematic diagram for wet and dry granulation of blast furnace
slag [143, 144]. Fig. 14. The schematic diagram for wet and dry granulation of blast furnace
slag [143, 144]. In WHR from BF slag using physical methods, a high-speed rotating cup (RCA or
RMCA) is used for the granulation as shown in Fig. 15 [143, 144]. High-temperature
liquid slag is directly poured into the high-speed rotating cup which enforced the slag to
eject radially under the influence of centrifugal force. The hot granules (1-6 mm) bounce
off the vessel walls and fall into the bed. The diameter and speed of the rotating cup have
to be optimized to reach an adequate level of slag granulation. The bed kept in motion by
injecting cooled air down the bed. Heat exchange takes place between the hot granules
and cooled air. The hot air is collecting at the top of the vessel and passes to a waste heat
recovery system to produce superheated steam. It was estimated that 120 t slag/h could
generate 29 ton of steam for a steam turbine connected to a generator to produce about
6.5 MWel [143]. In a successful commercial trial, about 59% of heat is recovered by RCA
and slag particles were cooled down to 250 °C [144]. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 94 Fig. 15. (a) Rotary cylinder atomizer (RCA) with heat recovery [143],
b) Rotary multi-nozzle cup atomizer (RMCA) [144]. Fig. 15. (a) Rotary cylinder atomizer (RCA) with heat recovery [143],
b) Rotary multi-nozzle cup atomizer (RMCA) [144]. Various techniques were applied for dry granulation of slag based on the
mechanical crushing method [145, 146]. Some of these techniques (e.g., solid slag
impingement, mechanical stirring) are commercialized for a short term and it could not
continue due to the low quality of granulated slag, for instance, non-uniform size. In one
of the recently developed dry granulation method, as seen in Fig. WHR from BF Slag 16, the liquid slag is
poured into the mold and small cold steel spheres are added eventually over the liquid
slag surface. The sphere balls penetrate the liquid steel bath and act as a coolant for hot
slag. The temperature of the buffer, from slag and steel balls, becomes around 650 °C,
which allows efficient heat recovery based on counter-current heat flow between the
descending hot buffer and ascending cold air. The outcome hot air at ~600 °C can be used
in pre-heating combustion air or to operate the steam turbine. The ingot consists of steel
balls and slag is dropped down on steel plate to disintegrate into granulated slag particles
and steel spheres. The glassy phase in the granulated slag is about 95%, which meets the
requirements of the cement industry. Fig. 16. Paul Wurth dry granulation mixing method [146]. Chemical methods can also be efficiently used in WHR from molten slag as many
of chemical reactions are endothermic and required heat to proceed. The sensible waste Fig. 16. Paul Wurth dry granulation mixing method [146]. Fig. 16. Paul Wurth dry granulation mixing method [146]. Chemical methods can also be efficiently used in WHR from molten slag as many
of chemical reactions are endothermic and required heat to proceed. The sensible waste Chemical methods can also be efficiently used in WHR from molten slag as many
of chemical reactions are endothermic and required heat to proceed. The sensible waste 95 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … E. A. Mousa - Modern Blast Furnace Ironmaking Technolog heat of slag can supply the required heat. Reforming of NG by water vapor or carbon
dioxide to generate CO and H2 is an endothermic process. Molten slag can be used for
methane reforming [147, 148]. In this case, the hot slag will not only act as a heat carrier
for methane reforming but also as a reaction catalyst. Streams of CH4 and H2O/CO2 are
applied to cool down the granulated liquid slag and intensive heat exchange takes place. The sensible heat from slag promoted the endothermic reaction of methane reforming and
generates H2 and CO, which means that the sensible heat is transformed into chemical
heat stored in the product fuel gases. WHR from BF Slag The generated hot gases are cooled down and go to
the methanation reactor in which the reversible reaction takes place to give CH4 and H2O,
which can be once more recycled for slag cooling. The sensible heat in molten slag can
be exploited in pyrolysis or endothermic gasification reaction of coal/biomass, as shown
in Fig. 17(a) [148]. It can be also used in pyrolysis/gasification of municipal solid wastes
such as electronic board as shown in Fig. 17(b) [144, 149]. The solid wastes are crushed
into small pieces and fed into shaft bed. Rotary multi-nozzle cup atomizer is used for
granulation of slag particles. The granulated slag particles at ~900 °C are fed into shaft
containing the crushed waste solid materials. The gasification takes place to develop CO,
H2, and CH4. Fig. 17. Utilization of hot slag in (a) Pyrolysis of biomass [148],
(b) Pyrolysis of solid waste [144, 149]. Fig. 17. Utilization of hot slag in (a) Pyrolysis of biomass [148],
(b) Pyrolysis of solid waste [144, 149]. Fig. 17. Utilization of hot slag in (a) Pyrolysis of biomass [148],
(b) Pyrolysis of solid waste [144, 149]. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 96 Table 4 summaries the developed processes for waste heat recovery form BF slag
and illustrate the capacity and rate of heat recovery [150]. Table 4 summaries the developed processes for waste heat recovery form BF slag
and illustrate the capacity and rate of heat recovery [150]. Table 4 summaries the developed processes for waste heat recovery form BF slag
and illustrate the capacity and rate of heat recovery [150]. Table 4. Developed processes for waste heat recovery from slags [150]. Table 4. Developed processes for waste heat recovery from slags [150]. Developer/
Process
name
Slag
type
&
temp. Summary and Conclusions The present article has thoroughly discussed the recent modifications and
developments which were recently conducted in the BF technology to reduce energy consumption and CO2 emission and can be summarized in the following points: (1) The top charging system was developed in large modern BF to assure a proper
distribution of feeding materials and gas streaming inside the furnace. Developing new burden distribution system is expected in the future, which will
be able to reach an optimum packed structure in the BF maintaining the shaft
permeability in low coke rate. The developed measuring systems and devices
were able to give a clear vision on the inner status of the BF and describe many
of unclear phenomena. Developing new burden distribution system is expected in the future, which will
be able to reach an optimum packed structure in the BF maintaining the shaft
permeability in low coke rate. The developed measuring systems and devices
were able to give a clear vision on the inner status of the BF and describe many
of unclear phenomena. (2) The conventional top charged burden including sinter, pellets and metallurgical
coke was reached a high level of quality makes BF works close to the ideal
conditions. Successful trails were implemented on charging active nut coke
inside the iron ore burden layers which accompanied by lower coke consumption
and higher productivity of hot metal. The top charging of in-plant fines in the
form of cold bonded briquettes or pre-reduced agglomerates into the BF has
provided good solutions to enhance the recirculation of residues, save virgin
resources, reduce the landfill materials, and reduce the energy consumption. The
partial replacement of conventional agglomerates and fossil fuels with novel top
charging biomass-iron ore composites were able to reduce the thermal reserve
zone temperature and consequently the coke consumption and fossil CO2
emission. The main challenge is still represented in the relatively low mechanical
strength of the novel composites and bio-agglomerates compared to the
conventional agglomerates. (3) The injection of pulverized coal through tuyeres was able to reduce the load on
cokemaking, utilize low quality coal resources, and reducing the overall costs;
however, further development in PCI technology is still needed to keep the coal
replacement ratio for coke high at injection rate higher than 200 kg/tHM. WHR from BF Slag Industrial
test
scale, t/h
Process
function and
characteristics
Recovered
media
Heat
recovery,
%
Exergy
recovery,
%
Slag
product
1
NKK,
Mitsubishi
heavy
industry
BOF
slag,
1600
80
Air-
atomization,
heat
exchanger
generates
steam
500
o
C air
and 270
kPa
saturated
steam
Hot air:
39.95%,
steam:
41.4,
total
81.35
39.9
Easy to be
weathering
dusted
2
Sumitomo
metal
BF
slag,
1400
50
RCA, FB,
packed bed,
heat exchange
600
o
C air
55
38.9
Fine
aggregate
for
concrete
3
MEROTEC
BF
slag,
1450
40
FB, heat
exchanger
recover steam
400 kPa
saturated
steam
65
38.8
Un-known
4
NKK
BF
slag,
1400
N.A
Drum
quenching,
recovering
heat
250
o
C
high
boiling
point
organic
liquid
40
16.3
Vitreous
ratio
>95%
5
NSC
BF
slag,
1400
7
Molten slag
poured into
mold, heat
exchange
mold wall
Hot water
60-70
<12
Fine
aggregate
for
concrete
6
NSC
BF
slag,
1450
100
Air-
atomization,
first heat
exchange by
the wind
tunnel,
second heat
exchange by
the multi-
fluid bed
510
o
C
air (1
st
exchange),
650oC air
(2
nd
exchange)
1
st
: 47.8
2
nd
: 14.8
Total
62.6
41.5
Cement
material
7
Conventional
water
granulation
BF
slag,
1450
N.A
High-pressure
water
granulation, 5
t water/ t slag
80
o
C hot
water
73
9.5
Cement
material
8
CDQ gas-
solid heat
exchange
process
Red
hot
coke,
1000
N.A. Coke packed
heat exchange
800
o
C
nitrogen
80
73
N.A. 97 E. A. Mousa - Modern Blast Furnace Ironmaking Technology: Potentials to Meet … (5) The recovery of waste heat represents the next frontier to enhance energy
efficiency and reduce emissions in the steel industry. The sensible heat of BF
slag can be used in the production of superheated steam for power generation.
The sensible heat can be used in the reforming of natural gas or pyrolysis of coal,
biomass, and waste plastics. The long payback period of investment cost
represents the main challenges for its practical implementation. Summary and Conclusions Injection of pre-treated biomass on account of PC has significant potential in
reducing the fossil CO2 emission from the BF, but technical and economic
constraints are still main issues to be addressed in future work. (4) The generated off-gases in the integrated steel plant can be used more efficient
if it is injected into the BF instead of its flaring in the atmosphere. The BF off-
gas can be recycled after separation of CO2 while coke oven gas and converter
gas can be directly injected into the BF to reduce the fossil fuels and CO2
emission. The waste plastics can be efficiently injected into the BF as an
alternative reducing agent to fossil fuels; however, the collection system and pre-
treatment methods are still representing the main challenges for its efficient
implementation. (5) The recovery of waste heat represents the next frontier to enhance energy
efficiency and reduce emissions in the steel industry. The sensible heat of BF
slag can be used in the production of superheated steam for power generation. The sensible heat can be used in the reforming of natural gas or pyrolysis of coal,
biomass, and waste plastics. The long payback period of investment cost
represents the main challenges for its practical implementation. Metall. Mater. Eng. Vol 25 (2) 2019 p. 69-104 98 Acknowledgment The author acknowledges the support provided for this work from EIT
RawMaterials for CIRRUS Project and the support from Vinnova and Energy Agency in
Sweden supporting the research projects OsMet (project no. 2017-01327), InnoBind
(project no. 2017-05449), and Bioagglomerat (project no. 39150-1). The paper is a
contribution from CAMM, Centre of Advanced Mining and Metallurgy, at the Luleå
University of Technology. The author wish to thank PRISMA Center at Swerim for the
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https://www1.eere.energy.gov/manufacturing/intensiveprocesses/pdfs/waste_heat_r
ecovery.pdf. Accessed 14 December 2016. https://www1.eere.energy.gov/manufacturing/intensiveprocesses/pdfs/waste_heat_r Creative Commons License This work is licensed under a Creative Commons Attribution 4.0 International License. This work is licensed under a Creative Commons Attribution 4.0 International License.
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DOI: 10.19139/soic-2310-5070-915 DOI: 10.19139/soic-2310-5070-915 DOI: 10.19139/soic-2310-5070-915 STATISTICS, OPTIMIZATION AND INFORMATION COMPUTING
Stat., Optim. Inf. Comput., Vol. 8, September 2020, pp 668–683.
Published online in International Academic Press (www.IAPress.org) STATISTICS, OPTIMIZATION AND INFORMATION COMPUTING
Stat., Optim. Inf. Comput., Vol. 8, September 2020, pp 668–683. Published online in International Academic Press (www.IAPress.org) STATISTICS, OPTIMIZATION AND INFORMATION COMPUTING
Stat., Optim. Inf. Comput., Vol. 8, September 2020, pp 668–683. Published online in International Academic Press (www.IAPress.org) CQ-free optimality conditions and strong dual formulations for a special
conic optimization problem Olga Kostyukova 1, Tatiana Tchemisova 2,∗ 1Institute of Mathematics, National Academy of Sciences of Belarus, Minsk, Belarus
2Center for Research and Development in Mathematics and Applications, University of Aveiro, Aveiro, Portugal Abstract
In this paper, we consider a special class of conic optimization problems, consisting of set-semidefinite (or K-
semidefinite) programming problems, where the set K is a polyhedral convex cone. For these problems, we introduce the
concept of immobile indices and study the properties of the set of normalized immobile indices and the feasible set. This
study provides the main result of the paper, which is to formulate and prove the new first-order optimality conditions in
the form of a criterion. The optimality conditions are explicit and do not use any constraint qualifications. For the case of a
linear cost function, we reformulate the K-semidefinite problem in a regularized form and construct its dual. We show that
the pair of the primal and dual regularized problems satisfies the strong duality relation which means that the duality gap is
vanishing. Keywords Conic optimization, set-semidefinite optimization, optimality conditions, normalized immobile index set,
regularized dual problem, strong duality AMS 2010 subject classifications 90C25, 90C30, 90C34 ∗Correspondence to: Tatiana Tchemisova (Email: tatiana@ua.pt). Center for Research and Development in Mathematics and Applications,
Department of Mathematics, University of Aveiro, Campus Universit´ario Santiago, 3810-193, Aveiro, Portugal. ∗Correspondence to: Tatiana Tchemisova (Email: tatiana@ua.pt). Center for Research and Development in Mathematics and Applications,
Department of Mathematics, University of Aveiro, Campus Universit´ario Santiago, 3810-193, Aveiro, Portugal.
ISSN 2310-5070 (online) ISSN 2311-004X (print)
C
i ht c⃝2020 I t
ti
l A
d
i P 1. Introduction Convex optimization is a subfield of mathematical optimization where the problems of minimizing convex
functions over convex sets are solved. Convex optimization has applications in a wide range of disciplines, such
as automatic control systems, estimation and signal processing, communications and networks, electronic circuit
design, data analysis and modeling, statistics, optimal experimental design, and structural optimization, where the
approximation concept has proven to be efficient ( see [7, 8] and the references therein). With recent advancements
in computing and optimization algorithms, convex programming is indispensable in emerging methods of data
mining, analytics, machine learning, as well as management, economics, and finance (e.g.[23, 27, 31, 32]). Conic optimization studies problems of minimizing a convex function over the intersection of an affine subspace
and a convex cone. Conic models are efficiently used in dynamic processes, biomedical and chemical engineering,
biology, credit risk optimization, etc. (see [3, 32, 35], and others). According to F. Glineur ([13]), conic optimization
is an elegant framework for convex optimization, due to the fact that conic formulations present various advantages
over the traditional formulations: the standard primal-dual pair of conic problems does feature a great deal of
symmetry, the dual problem and the associated weak and strong duality properties can be derived in a ”seamless
manner” once the dual cone has been computed, and the conic formulations are helpful when designing and
analyzing numerical algorithms. ISSN 2310-5070 (online) ISSN 2311-004X (print)
Copyright c⃝2020 International Academic Press 669 O. KOSTYUKOVA AND T. TCHEMISOVA In this paper, we consider a special class of conic optimization problems, consisting of set-semidefinite (or K-
semidefinite) programming problems, where the set K is a polyhedral convex cone. The interest in such problems
is due to the fact that they have a wide range of applications in such important areas as systems and management
theory, robust and combinatorial optimization. Among the most important subclasses of conic problems under
consideration, there are semidefinite programming (SDP) and copositive problems which have many applications
in robust optimization, graph theory, computational biology, systems and control theory, sensor network location,
and data analysis, among others [2, 9, 22, 24]. Semidefinite and copositive programming are recognized in
combinatorial optimization as valuable techniques for modeling and obtaining bounds on solutions of NP - hard
problems (for the references see [5, 10, 14, 33]). 1. Introduction New numerical scheme based on semidefinite programming to
compute approximately and sometimes exactly the Nash equilibria, the min–max of any finite game, and the values
and the optimal strategies of any polynomial 2-player zero-sum game are presented in [21, 34] et. al. One of the main difficulties faced in solving conic optimization problems is that many of the problems in
this class are characterized by a violation of regularity (and/or so-called CQ – constraint qualifications). This
makes it impossible to apply the known results from the general theory of optimization, entails a violation of strict
duality relations which in turn leads to significant difficulties in justifying and implementing numerical methods for
solving such problems. In this regard, it is relevant to study the problems of conic optimization without additional
assumptions about regularity. p
g
y
The purpose of this work is to formulate and prove new optimality conditions which do not use any CQs (are
CQ-free) and obtain new dual formulations which satisfy the strong duality relations and therefore may form the
basis of efficient numerical methods. The paper is organized as follows. Section 1 hosts Introduction. The problem’s settings and basic notations are
introduced in section 2. The new optimality conditions for a set-semidefinite problem with a polyhedral set are
formulated in section 3. In section 4, we explore the properties of the set of immobile indices and the feasible set
of the problem under study and use them to prove, in section 5, the main result of the paper, which is the new
first-order CQ-free optimality conditions for the set-semidefinite programming (Theorem 1). In section 6, for a
linear case, we deduce a regularized pair of primal and dual K-semidefinite problems and show that for this pair
the strong duality property holds. The final section 7 contains some conclusions. minimize c(x), minimize c(x),
subject to x ∈Rn, t⊤A(x)t ≥0 ∀t ∈K := {t ∈Rp : At = 0, Bt ≥0},
(3) (3) subject to x ∈Rn, t⊤A(x)t ≥0 ∀t ∈K := {t ∈Rp : At = 0, Bt ≥0},
(3) where A(x) :=
n∑
k=1
Akxk + A0, Ak ∈S(p), k = 0, 1, ..., n, A ∈Rm0×p, B ∈Rm∗×p, are given matrices. Problem (3) is a generalization of semidefinite and copositive programming problems since in the case K = Rp,
it is a problem of SDP, and in the case K = Rp
+, a copositive problem. The main purpose of this paper is to prove the optimality conditions and to obtain strong duality results for
the set-semidefinite problem (3) that do not require any regularity conditions (constraint qualifications or, shortly,
CQ). We will use here the approach developed in our previous papers (see e.g. [16, 18, 20]) for different classes of
convex problems. 670
Some prop
is convex for
Let A : Rn
is called a set
case of conic
In this pap
c : Rn →R
optimization
where A(x) :
Problem (3
it is a problem
The main
the set-semid
CQ). We will
convex probl
2.1. Some ot
It is evident t
On the other
the form
where the de
M ⊂Rp is a
Suppose th
c(x), x ∈Rn
with given m
Problem (4
Let us show
above, can be 670 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION Some properties of the cone of K-semidefinite matrices are studied in [12]. Notice, in particular, that this cone
is convex for any set K. Some properties of the cone of K-semidefinite matrices are studied in [12]. Notice, in particular, that this cone
is convex for any set K. y
Let A : Rn →S(p) and c : Rn →R be some given continuous maps. A problem minimize c(x),
subject to x ∈Rn, A(x) ∈CK,
(2) minimize c(x),
subject to x ∈Rn, A(x) ∈CK,
(2) (2) is called a set-semidefinite (or K-semidefinite) optimization problem. This problem can be considered as a particular
case of conic optimization problems. In this paper, we consider problem (2) under assumptions that the map A : Rn →S(p) is affine, the function
c : Rn →R is convex and the set K ⊂Rp is a polyhedral convex cone. Namely, we consider the following
optimization problem: minimize c(x), 2. Problem’s settings and basic notation In what follows, we will use the following notation. Given integers p and k, let Rp and Rk×p denote the set of
all real-valued p - vectors and k × p - matrices, respectively. Denote by Rp
+⊂Rp the subset of all p - vectors with
non-negative components and by S(p) the space of symmetric p × p matrices with the trace inner product: A • B := trace(AB), for A, B ∈S(p). A • B := trace(AB), for A, B ∈S(p). For an arbitrary nonempty set K ⊂Rp, consider the set et K ⊂Rp, consider the set
CK := {D ∈S(p) : t⊤Dt ≥0
∀t ∈K},
(1) CK := {D ∈S(p) : t⊤Dt ≥0
∀t ∈K},
(1) (1) called a cone of K - semidefinite (set-semidefinite) matrices, or simply, the K - semidefinite cone, see e.g. [11, 12]. It worth mentioning that the cone CK is a generalization of a well-known and well-studied in literature cone of
positive semidefinite matrices S+(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp}, S+(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp}, and a less studied cone of copositive matrices and a less studied cone of copositive matrices COP(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp
+},
ences therein. Indeed, for K = Rp, we have CK = S+(p) and for K = Rp
+, we have COP(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp
+}, COP(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp
+}, COP(p) := {D ∈S(p) : t⊤Dt ≥0 ∀t ∈Rp
+}, see e.g. [1, 4, 5, 6] and the references therein. Indeed, for K = Rp, we have CK = S+(p) and for K = Rp
+, we have
CK = COP(p). Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 670
OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION 2.1. Some other formulations It is evident that problem (3) can be considered as a special case of Semi-Infinite Programming (SIP) problems [7]. On the other hand, some more general than (3) problems can be reduced to it. In fact, consider a SIP problem in
the form minimize c(x),
subject to x ∈Rn, f(x, t) ≥0 ∀t ∈M,
(4) (4) minimize c(x),
subject to x ∈Rn, f(x, t) ≥0 ∀t ∈M, where the decision variable x = (x1, ..., xn) is an n−vector, the index variable t = (t1, . . . , tp) is a p-vector, and
M ⊂Rp is a given index set. Suppose that in (4), the constraint function f(x, t) is linear w.r.t. x and linear- quadratic w.r.t. t, the cost function
c(x), x ∈Rn, is convex, and the index set M is a convex polyhedron: M := {t ∈Rp : At = a, Bt ≥b}, M := {t ∈Rp : At = a, Bt ≥b},
(5) (5) with given matrices A ∈Rm0×p, B ∈Rm∗×p and vectors a ∈Rm0, b ∈Rm∗. Problem (4) with the index set in the form (5) is a special case of polinomial SIP problems. Problem (4) with the index set in the form (5) is a special case of polinomial S Let us show that problem (4) with the index set in the form (5) and with the constraints function f(x, t) described
above, can be reduced to problem (3). p
Indeed, it is evident that problem (4) can be rewritten as follows: Indeed, it is evident that problem (4) can be rewritten as follows: minimize c(x),
subject to x ∈Rn, t⊤¯
A(x)t + 2t⊤(Dx + d0) + q⊤x + q0 ≥0 ∀t ∈M,
(6) (6) subject to x ∈Rn, t⊤¯
A(x)t + 2t⊤(Dx + d0) + q⊤x + q0 ≥0 ∀t ∈M,
(6) where the matrix function ¯
A(x) has the form ¯
A(x) :=
n
∑
k=1
¯Akxk + ¯A0, and the problem’s data is set:
matrices ¯Ak ∈S(p), k = 0, 1, . . . , n, D ∈Rp×n, vectors d0 ∈Rp, q ∈Rn, and the number q0 ∈R. where the matrix function ¯
A(x) has the form ¯
A(x) :=
∑
k=1
¯Akxk + ¯A0, and the problem’s data is set:
matrices ¯Ak ∈S(p), k = 0, 1, . . . , n, D ∈Rp×n, vectors d0 ∈Rp, q ∈Rn, and the number q0 ∈R. 2.1. Some other formulations k
1
s ¯Ak ∈S(p), k = 0, 1, . . . , n, D ∈Rp×n, vectors d0 ∈Rp, q ∈Rn, and the number q0 ∈R. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 671 O. KOSTYUKOVA AND T. TCHEMISOVA Let X1 be the set of feasible solutions of problem (6): Let X1 be the set of feasible solutions of problem (6): Let X1 be the set of feasible solutions of problem (6): Let X1 be the set of feasible solutions of problem (6): X1 = {x ∈Rn : t⊤¯
A(x)t + 2t⊤(Dx + d0) + q⊤x + q0 ≥0 ∀t ∈M}. Denote by d·k ∈Rp, k = 1, . . . , n, the columns of the matrix D ∈Rp×n: D = (d·1 . . . d·n), and let
q = (q1, . . . , qn)⊤. Introduce the following notation: Bk :=
(
qk
d⊤
·k
d·k
¯Ak
)
∈S(p + 1), k = 0, 1, ..., n; B(x) :=
n
∑
k=1
Bkxk + B0, Bk :=
(
qk
d⊤
·k
d·k
¯Ak
)
∈S(p + 1), k = 0, 1, ..., n; B(x) :=
n
∑
k=1
Bkxk + B0,
¯τ := (τ0, τ ⊤)⊤, where τ0 ∈R, τ = (τ1, ..., τp)⊤∈Rp,
K := {¯τ ∈Rp+1 : (−a, A)¯τ = 0, (−b, B)¯τ ≥0, τ0 ≥0}, ¯τ := (τ0, τ ⊤)⊤, where τ0 ∈R, τ = (τ1, ..., τp)⊤∈Rp,
K := {¯τ ∈Rp+1 : (−a, A)¯τ = 0, (−b, B)¯τ ≥0, τ0 ≥0}, K := {¯τ ∈Rp+1 : (−a, A)¯τ = 0, (−b, B)¯τ ≥0, τ0 ≥0}, and consider the set and consider the set X2 := {x ∈Rn : ¯τ ⊤B(x)¯τ ≥0 ∀¯τ ∈K}. (7) X2 := {x ∈Rn : ¯τ ⊤B(x)¯τ ≥0 ∀¯τ ∈K}. (7) One can easily prove the following lemma. Lemma 1
Gi
h f
ibl
X
f
bl
(6)
d h
X d fi
d i (7) h
li
X
X h ld Lemma 1 Lemma 1
Given the feasible set X1 of problem (6) and the set X2 defined in (7), the equality X1 = X2 holds. en the feasible set X1 of problem (6) and the set X2 defined in (7), the equality X1 = X2 holds. 2.1. Some other formulations Note that the set X2 can be considered as the feasible set of the problem in the form (3) with function A(x)
replaced by B(x) and the index set K substituted by K. It follows from Lemma 1, that the study of the special SIP problem in the form (6) with the index set (5) is
equivalent to the study of the conic (set-semidefinite) problem in the form (3). OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION It will be shown (see Lemma 3 below) that the set Tim is either empty or a union of a finite number of convex
bounded polyhedra. Then, evidently, the set conv Tim is either empty or a convex polyhedron. Here conv D stays
for the convex hull of a set D. In what follows, we will use the vertices of the set conv Tim. Denote the set of these vertices b (9) {t∗(j) ∈Tim, j ∈J, |J| < ∞}. In the case Tim = ∅, we set J = ∅. In the case Tim = ∅, we set J = ∅. such that (U + V ) • Ak = ∂c(x0)
∂xk
, k = 1, ..., n; (U + V ) • A(x0) = 0,
(11) (11) where matrices U and V are defined as follows: where matrices U and V are defined as follows: U :=
∑
i∈I
τ(i)(τ(i))⊤, V :=
∑
j∈J
(λ(j)(t∗(j))⊤+ t∗(j)(λ(j))⊤),
(12) U :=
∑
i∈I
τ(i)(τ(i))⊤, V :=
∑
j∈J
(λ(j)(t∗(j))⊤+ t∗(j)(λ(j))⊤), (12) and t∗(j), j ∈J, are vertices of the polyhedron conv Tim. and t∗(j), j ∈J, are vertices of the polyhedron conv Tim. and t∗(j), j ∈J, are vertices of the polyhedron conv Tim. We will prove this theorem in section 5. We will prove this theorem in section 5. For applications, it may be useful to reformulate Theorem 1 in a different way. For applications, it may be useful to reformulate Theorem 1 in a different way. For applications, it may be useful to reformulate Theorem 1 in a different way. For applications, it may be useful to reformulate Theorem 1 in a different way. Set ¯m := |J| and denote by Rp× ¯m(K) the set of p × ¯m matrices Λ = (λ(j), j ∈J) with λ(j) ∈K, j ∈J. Having introduced the matrix T := (t∗(j), j ∈J) ∈Rp× ¯m and the sets of matrices Set ¯m := |J| and denote by Rp× ¯m(K) the set of p × ¯m matrices Λ = (λ(j), j ∈J) with λ(j) ∈K, j ∈J. Having introduced the matrix T := (t∗(j), j ∈J) ∈Rp× ¯m and the sets of matrices V(K) := {V ∈S(p) : V = ΛT ⊤+ T Λ⊤, Λ ∈Rp× ¯m(K)},
(13)
(CK)∗:= conv {tt⊤: t ∈K}, (13) we can reformulate Theorem 1 as follows. 2.2. Normalized set of immobile indices In accordance with the generally accepted definition (see, e.g., [7, 13]), problem (3) satisfies the Slater condition
if for some ¯x ∈Rn it holds: t⊤A(¯x)t > 0 ∀t ∈K \ {0}. Notice that from the very beginning, we do not make
assumptions that the constraints of problem (3) satisfy the Slater condition or some other conditions. p
p
y
Since the set K is a cone, problem (3) has an unbounded index set. It is easy to show that this problem is
equivalent to the following SIP problem with a compact index set: minimize c(x),
subject to x ∈Rn, t⊤A(x)t ≥0 ∀t ∈T := {t ∈K : ||t||∞= 1},
(8) minimize c(x),
subject to x ∈Rn, t⊤A(x)t ≥0 ∀t ∈T := {t ∈K : ||t||∞= 1},
(8) (8) where ||t||∞= max
k∈P |tk|. The index set T is normalized in this problem. where ||t||∞= max
k∈P |tk|. The index set T is normalized in this problem. In our papers [15, 16, 19], and others, we introduced for convex SIP problems the concept of immobile indices
(the indices of constraints which are active for all feasible solutions) and showed that these indices play a special
role in formulating the optimality conditions which do not need the fulfillment of the Slater condition. Here, we
will show how our approach can be applied to the conic problem under consideration. Denote by X the set of feasible solutions in the SIP problem (8) and in the equivalent K- semidefinite problem
(3):
⊤ the set of feasible solutions in the SIP problem (8) and in the equivalent K- semidefinite problem Denote by X the set of feasible solutions in the SIP problem (8) and in the equivalent K- sem Denote by X the set of feasible solutions in the SIP problem (8) X = {x ∈Rn : t⊤A(x)t ≥0 ∀t ∈T}, X = {x ∈Rn : t⊤A(x)t ≥0 ∀t ∈T}, and by Tim the set of (normalized) immobile indices in problem (8): and by Tim the set of (normalized) immobile indices in problem (8): Tim := {t ∈T : t⊤A(x)t = 0 ∀x ∈X}. It can be proved (see [15]) that the Slater condition for problem (8) is equivalent to the condition Tim = ∅. It can be proved (see [15]) that the Slater condition for problem (8) is equivalent to the condition Tim = ∅. Stat., Optim. Inf. Comput. Vol. 3. CQ-free optimality criteria The main aim of this paper is to prove necessary and sufficient optimality conditions for the conic optimization
problem (3) and the equivalent problem (8). In this section, we formulate two theorems which have the form of
criteria. These theorems do not demand that the feasible set of the problem satisfies any constraints qualifications
and therefore, we refer to them as to CQ-free optimality conditions. The main concepts that allow us to formulate
and prove these optimality conditions are the set of immobile indices and the vertices of its convex hull. Theorem 1
A vector x0 ∈X is an optimal solution of the set-semidefinite problem (3) (and the equivalent SIP problem (8)) iff
there exist vectors Theorem 1
A vector x0 ∈X is an optimal solution of the set-semidefinite problem (3) (and the equivalent SIP problem (8)) iff
there exist vectors or x0 ∈X is an optimal solution of the set-semidefinite problem (3) (and the equivalent SIP prob
xist vectors there exist vectors (10) λ(j) ∈K, j ∈J; τ(i) ∈K, i ∈I, |I| ≤n,
(10) 2.2. Normalized set of immobile indices 8, September 2020 672 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION 4. The properties of problem (8) To prove Theorem 1 we need some auxiliary results which will be considered in this section. we can reformulate Theorem 1 as follows. Then αA(¯x −h/α) = ˜D −¯S and αt⊤A(¯x −h/α)t = t⊤˜Dt −t⊤¯St > 0 ∀t ∈K, t ̸= 0. It follows from these relations that, for problem (3), the Slater condition holds true with (¯x −h/α) ∈X. Suppose that α > 0. Then αA(¯x −h/α) = ˜D −¯S and αt⊤A(¯x −h/α)t = t⊤˜Dt −t⊤¯St > 0 ∀t ∈K, t ̸= 0. It follows from these relations that, for problem (3), the Slater condition holds true with (¯x −h/α) ∈X. ,
p
( ),
(
/ ) ∈
Now suppose that α = 0 in (16). Then
n∑
k=1
Akhk = ¯S and A(¯x −h) = A(¯x) −¯S. Hence
⊤
⊤
⊤¯ Now suppose that α = 0 in (16). Then
n∑
k=1
Akhk = ¯S and A(¯x −h) = A(¯x) −¯S. Hence t⊤A(¯x −h)t = t⊤A(¯x)t −t⊤¯St > 0 ∀t ∈K, t ̸= 0. This implies that for problem (3), the Slater condition holds true with (¯x −h) ∈X. The main contribution of this paper consists of new optimality criteria formulated in Theorems 1 and 2. These
criteria are CQ-free since they do not need any additional assumptions, whether it is the KRZ condition, or the
Slater condition or any other CQ. This means that the scope of these optimality conditions is much wider than of
other known conditions. This result was possible due to the application of the approach based on the concept of
immobility. In section 6, we will show how the obtained results can be used to produce strong dual formulations
for problem (3) with a linear cost function. we can reformulate Theorem 1 as follows. where ¯x ∈X, Ξ := {Z ∈S(p) : Z =
n∑
k=1
Akhk, ∀hk ∈R, k = 1, ..., n}, and coneD stays for the conic hull of a
set D. where ¯x ∈X, Ξ := {Z ∈S(p) : Z =
n∑
k=1
Akhk, ∀hk ∈R, k = 1, ..., n}, and coneD stays for the conic hull of a
set D. D. et us prove that, for an affine matrix function A(x), condition (15) implies the Slater condition. Let us prove that, for an affine matrix function A(x), condition (15) implies the Slater condition. Suppose that condition (15) fulfills and let ¯S ∈S(p) be such that t⊤¯St < 0 for all t ∈Rp, t ̸= 0. It follows from
(15) that there exist h = (hk ∈R, k = 1, ..., n)⊤, αi ≥0, Di ∈CK, i ∈I, such that p
( )
p
Suppose that condition (15) fulfills and let ¯S ∈S(p) be such that t⊤¯St < 0 for all t ∈Rp, t ̸= 0. It follows from
(15) that there exist h = (hk ∈R, k = 1, ..., n)⊤, αi ≥0, Di ∈CK, i ∈I, such that n
∑
k=1
Akhk +
∑
i∈I
αi(Di −A(¯x)) = ¯S ⇔
n
∑
k=1
Akhk + ( ˜D −αA(¯x)) = ¯S
(16)
with ˜D =
∑
i∈I
αiDi ∈CK, α =
∑
i∈I
αi ≥0. n
∑
k=1
Akhk +
∑
i∈I
αi(Di −A(¯x)) = ¯S ⇔
n
∑
k=1
Akhk + ( ˜D −αA(¯x)) = ¯S
(16) (16) with ˜D =
∑
i∈I
αiDi ∈CK, α =
∑
i∈I
αi ≥0. ∈
∈
Suppose that α > 0. Then αA(¯x −h/α) = ˜D −¯S and αt⊤A(¯x −h/α)t = t⊤˜Dt −t⊤¯St > 0 ∀t ∈K, t ̸= 0. It follows from these relations that, for problem (3), the Slater condition holds true with (¯x −h/α) ∈X. Now suppose that α = 0 in (16). Then
n∑
k=1
Akhk = ¯S and A(¯x −h) = A(¯x) −¯S. Hence Suppose that α > 0. Then αA(¯x −h/α) = ˜D −¯S and αt⊤A(¯x −h/α)t = t⊤˜Dt −t⊤¯St > 0 ∀t ∈K, t ̸= 0. It follows from these relations that, for problem (3), the Slater condition holds true with (¯x −h/α) ∈X. Now suppose that α
0 in (16) Then
n∑A h
¯S and A(¯x
h)
A(¯x)
¯S Hence Suppose that α > 0. we can reformulate Theorem 1 as follows. Theorem 2 Theorem 2
A vector x0 ∈X is an optimal solution of problem (3) iff there exist matrices U 0 ∈(CK)∗and V 0 ∈V(K) such
that A vector x0 ∈X is an optimal solution of problem (3) iff there exist matrices U 0 ∈(CK)∗a
that (U 0 + V 0) • Ak = ∂c(x0)
∂xk
, k = 1, 2, ..., n; (U 0 + V 0) • A(x0) = 0. (14 (14) At the end of this section, we would like to make several remarks. R
k 1 At the end of this section, we would like to make several remarks. At the end of this section, we would like to make several remarks. It can be shown that the cone (CK)∗is dual to the cone CK. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 O. KOSTYUKOVA AND T. TCHEMISOVA 673 Remark 2 Theorem 1 can be easily generalized for the case when the set K is an union of the finite num It was mentioned above that the conic problem (3) can be considered as a special case of SIP problems. Therefore,
to study the optimality in this problem, one can use the optimality conditions known from the theory of SIP. Notice
that almost all known optimality conditions for SIP problems require the fulfillment of some additional conditions. As a rule, these conditions are either of the Slater type (see [7, 26, 28, 29] and the references therein) or include
some weaker assumptions (see, e.g. [15, 16, 19]). It worth to be mentioned that there is a small number of publications in the literature, where optimality conditions
for set-semidefinite problems are formulated. These optimality conditions were also obtained under assumptions
that in the case of problem (3), are equivalent to the Slater condition. In [12], for a set-semidefinite problem in Banach and finite dimensional spaces, optimality conditions in a non-
degenerate form were proved under a so-called Kurcyusz-Robinson-Zowe (KRZ) regularity condition (see (16)
from [12]). For the problem in the form (3), this condition takes the form Ξ + cone(CK −{A(¯x)}) = S(p),
(15) (15) Ξ + cone(CK −{A(¯x)}) = S(p),
(15)
where ¯x ∈X, Ξ := {Z ∈S(p) : Z =
n∑
k=1
Akhk, ∀hk ∈R, k = 1, ..., n}, and coneD stays for the conic hull of a
set D. Consider a subspace of Rp in the form Consider a subspace of Rp in the form H: = {t ∈Rp : At = 0, Bt = 0}
(18) (18) and denote by bs ∈Rp, s ∈S, |S| ≤p, the vectors of an orthogonal basis of this subspace. C
id
th
t and denote by bs ∈Rp, s ∈S, |S| ≤p, the vectors of an orthogonal basis of this subspace. Consider the set y
Consider the set H∗:= {t ∈Rp : At = 0, Bt ≥0, t⊤bs = 0, s ∈S}. (19) H∗:= {t ∈Rp : At = 0, Bt ≥0, t⊤bs = 0, s ∈S}. (19) By construction, H∗is a polyhedral convex cone. Let us show that H∗is a pointed cone, i.e. By construction, H∗is a polyhedral convex cone. Let us show that H∗is a pointed cone, i.e. t ̸= 0, t ∈H∗
⇒
−t ̸∈H∗. t ̸= 0, t ∈H∗
⇒
−t ̸∈H∗. In fact, suppose that, on the contrary, there exists ¯t ̸= 0 such that ¯t ∈H∗and −¯t ∈H∗. Then A¯t = 0, B¯t = 0,
and, consequently, ¯t ∈H. Thus we can conclude that ¯t = ∑
s∈S
βsbs for some scalars βs, s ∈S. As ¯t ∈H∗, the
following equalities should be satisfied: 0 = ¯t⊤bi =
∑
s∈S
βsb⊤
s bi = βi, i ∈S. These equalities contradict the assumption that 0 ̸= ¯t = ∑
s∈S
βsbs. The obtained contradiction proves that the convex
polyhedral cone H∗is pointed. Denote by as, s ∈S∗, |S∗| < ∞, the extreme directions in the cone H∗. Recall that by the definition, the extreme
directions in a polyhedral cone are its faces of dimension one. The following lemma formulates the known result about representation of polyhedral cones. Lemma 2 The cone K defined in (17), can be written as follows: K = {t ∈Rp : t =
∑
s∈S
βsbs +
∑
s∈S∗
αsas, αs ≥0, s ∈S∗}. Note that the formula above is well consistent with the well-known fact that any nonempty convex set C admits
representation in the form C = linC + (C ∩(linC)⊥), where linC is the largest subspace contained in the recession
cone of C, the so-called lineality space of C (see e.g. [25], p.65). Notice that for the polyhedral cone K in the form
(17), it holds linK = H and K ∩(linK)⊥= H∗, where H and H∗are defined in (18),(19). 4.1. A parametric representation of the cone K. First, consider the cone introduced in the previous section and used to identify the index sets of the equivalent
problems (3) and (8): the previous section and used to identify the index sets of the equivalent K = {t ∈Rp : At = 0, Bt ≥0}. (17) (17) Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION 674 Consider a subspace of Rp in the form Lemma 3 Given convex SIP problem (8), the immobile index set Tim is either empty or can be represented as a union of a
finite number of convex closed bounded polyhedra. Given convex SIP problem (8), the immobile index set Tim is either empty or can be represented as a union of a
finite number of convex closed bounded polyhedra. Proof. If Tim = ∅, the lemma is evident. Suppose that Tim ̸= ∅. Consider any t ∈Tim ⊂K. It follows from Lemma 2 that t admits the representation Proof. If Tim = ∅, the lemma is evident. Proof. If Tim
∅, the lemma is evident. Suppose that Tim ̸= ∅. Consider any t ∈Tim ⊂K. It follows from Lemma 2 that t admits the representation im
,
Suppose that Tim ̸= ∅. Consider any t ∈Tim ⊂K. It follows from Lemma 2 that t admits the representation Tim ̸= ∅. Consider any t ∈Tim ⊂K. It follows from Lemma 2 that t admits the representation t =
∑
s∈S
βsbs +
∑
s∈S∗
αsas, αs ≥0, s ∈ t =
∑
s∈S
βsbs +
∑
s∈S∗
αsas, αs ≥0, s ∈S∗. For t ∈Tim, denote S+
∗(t) := {s ∈S∗: αs > 0}. S+
∗(t) := {s ∈S∗: αs > 0}. Set Z := {Z ⊂S∗: ∃t ∈Tim, Z = S+
∗(t)}. Z := {Z ⊂S∗: ∃t ∈Tim, Z = S+
∗(t)}. From the definition of the set Z, it follows that it is finite: |Z| ≤2|S∗|, where 2|S∗| < ∞is the number of all subsets
of the finite set S∗. For any Z ∈Z, consider the corresponding set ider the corresponding set eT(Z) := cl{t ∈Tim : S+
∗(t) = Z}, eT(Z) := cl{t ∈Tim : S+
∗(t) = Z}, where cl(D) stays for the closure of a set D ⊂Rp. (
)
It is evident that It is evident that It is evident that
Tim =
∪
Z∈Z
eT(Z). It is evident that
Tim =
∪
Z∈Z
eT(Z). (22) Tim =
∪
Z∈Z
eT(Z). (22) Tim =
∪
Z∈Z
eT(Z). Tim =
∪
Z∈Z
eT(Z). (22) By construction, for any ¯t ∈eT(Z), we have ¯t ∈Tim, S+
∗(¯t) ⊂Z, and By construction, for any ¯t ∈eT(Z), we have ¯t ∈Tim, S+
∗(¯t) ⊂Z, and By construction, for any ¯t ∈eT(Z), we have ¯t ∈Tim, S+
∗(¯t) ⊂Z, and ¯t = lim
ω→∞t(ω),
where t(ω) ∈Tim, S+
∗(t(ω)) = Z. 4.2. The properties of the set of immobile indices of problem (8) Here, as above, we consider a general case when the set of immobile indices of problem (8) may be nonempty. Proposition 1 Here, as above, we consider a general case when the set of immobile indices of problem (8) may be nonempty. p
Suppose that for problem (8) it is satisfied: X ̸= ∅and Tim ̸= ∅. Then for any x ∈X and any t ∈Tim ⊂K, the
following relations hold true:
⊤
⊤ a⊤
s A(x)t ≥0, s ∈S∗; b⊤
s A(x)t = 0, s ∈S,
(20 a⊤
s A(x)t ≥0, s ∈S∗; b⊤
s A(x)t = 0, s ∈S, (20) where {bs ∈Rp, s ∈S, |S| ≤p} is the orthogonal basis of the subspace H defined in (18) and {as, s ∈S∗,
|S∗| < ∞} is the set of extreme directions in the pointed cone H∗defined in (19). Proof. Suppose that, on the contrary, there exist ¯x ∈X and ¯t ∈Tim, such that one of two situations occurs: Proof. Suppose that, on the contrary, there exist ¯x ∈X and ¯t ∈Tim, such that one of two situations occurs:
1 ∃
S
h h
⊤A( )¯t
0
2 ∃
S
h h
b⊤A( )¯t ̸
0 1. ∃s0 ∈S∗such that a⊤
s0A(¯x)¯t < 0 or 2. ∃s0 ∈S such that b⊤
s0A(¯x)¯t ̸= 0. Let us assume, first, that situation 1 has happened. By construction, the vector as0 is a recession direction for ¯t in
the cone K: t(θ) := ¯t + θas0 ∈K
∀θ ≥0. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 675 O. KOSTYUKOVA AND T. TCHEMISOVA Let us compute (t(θ))⊤A(¯x)t(θ) = 2θa⊤
s0A(¯x)¯t + θ2a⊤
s0A(¯x)as0. Taking into account that a⊤
s0A(¯x)¯t < 0, one
can conclude from the latter equality that there exists ¯θ > 0 such that (t(¯θ))⊤A(¯x)t(¯θ) < 0. The latter inequality
implies the inequality Let us compute (t(θ))⊤A(¯x)t(θ) = 2θa⊤
s0A(¯x)¯t + θ2a⊤
s0A(¯x)as0. Taking into account that a⊤
s0A(¯x)¯t < 0, one
can conclude from the latter equality that there exists ¯θ > 0 such that (t(¯θ))⊤A(¯x)t(¯θ) < 0. The latter inequality
implies the inequality et⊤A(¯x)et < 0, where et := t(¯θ)/||t(¯θ)||∞∈T,
(21) et⊤A(¯x)et < 0, where et := t(¯θ)/||t(¯θ)||∞∈T, (21) that contradicts the condition ¯x ∈X. that contradicts the condition ¯x ∈X. Now suppose that the situation 2 has occurred. Set γ := b⊤
s0A(¯x)¯t. By construction, the direction −γbs0 is
feasible for ¯t in K: t(θ) := ¯t −θγbs0 ∈K
∀θ ≥0. By simple computations, we get Now suppose that the situation 2 has occurred. Set γ := b⊤
s0A(¯x)¯t. By construction, the direction −γbs0 is
feasible for ¯t in K: t(θ) := ¯t −θγbs0 ∈K
∀θ ≥0. By simple computations, we get (t(θ))⊤A(¯x)t(θ) = −2θγb⊤
s0A(¯x)¯t + θ2γ2b⊤
s0A(¯x)bs0 = −2θγ2 + θ2γ2b⊤
s0A(¯x)bs0, and taking into account the inequality γ ̸= 0, one can conclude that there exists ¯θ > 0 such that t⊤(¯θ)A(¯x)t(¯θ) < 0. But this inequality implies relations (21) and hence contradicts the condition ¯x ∈X. The proposition is proved. 2 and taking into account the inequality γ ̸= 0, one can conclude that there exists ¯θ > 0 such that t⊤(¯θ)A(¯x)t(¯θ) < 0. But this inequality implies relations (21) and hence contradicts the condition ¯x ∈X. The proposition is proved. 2
Lemma 3 But this inequality implies relations (21) and hence contradicts the condition x ∈X. The proposition is proved. 2
Lemma 3 Lemma 3 Lemma 3 (23) ¯t = lim
ω→∞t(ω),
where t(ω) ∈Tim, S+
∗(t(ω)) = Z. (23) Let us show that for any ¯t ∈eT(Z), we have Let us show that for any ¯t ∈eT(Z), we have a⊤
s A(x)¯t = 0 ∀s ∈Z ∀x ∈X. (24) a⊤
s A(x)¯t = 0 ∀s ∈Z ∀x ∈X. (24) Suppose the contrary. Then for some ¯t ∈eT(Z), taking into account Proposition 1, there exist s0 ∈Z and ¯x ∈X,
such that a⊤
s0A(¯x)¯t > 0. From the latter inequality and relations (23), we conclude that there exists t(ω) ∈Tim
such that S+
∗(t(ω)) = Z and Suppose the contrary. Then for some ¯t ∈eT(Z), taking into account Proposition 1, there exist s0 ∈Z and ¯x ∈X,
such that a⊤
s0A(¯x)¯t > 0. From the latter inequality and relations (23), we conclude that there exists t(ω) ∈Tim
such that S+
∗(t(ω)) = Z and unt Pro
(23), a⊤
s0A(¯x)t(ω) > 0. a⊤
s0A(¯x)t(ω) > 0. (25) (25) Since S+
∗(t(ω)) = Z and s0 ∈Z, the direction −as0 is feasible for t(ω) in K, i.e. there exists θ0 > 0 such that
t(θ) := t(ω) −θas0 ∈K
∀θ ∈[0, θ0]. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION 676 Compute (t(θ))⊤A(¯x)t(θ) = −2θa⊤
s0A(¯x)t(ω) + θ2a⊤
s0A(¯x)as0. From the latter equality and inequality (25), it follows that there exists 0 < ¯θ ≤θ0 such that (t(θ))⊤A(¯x)t(¯θ) < 0. But this inequality implies relations (21) that contradict the assumption ¯x ∈X. Hence equalities (24) are proved. N
l
h
h Now let us show that bt ⊤A(x)¯t = 0 ∀bt ∈eT(Z), ∀¯t ∈eT(Z), ∀x ∈X. (26) (26) In fact, any vector bt ∈eT(Z) can be written as follows: In fact, any vector bt ∈eT(Z) can be written as follows: bt =
∑
s∈S
βsbs +
∑
s∈Z
αsas. bt =
∑
s∈S
βsbs +
∑
s∈Z
αsas. bt =
∑
s∈S
βsbs +
∑
s∈Z
αsas. Taking into account this representation, equalities (24) and relations (20), we obtain bt ⊤A(x)¯t =
(∑
s∈S
βsbs +
∑
s∈Z
αsas
)⊤
A(x)¯t = 0 ∀x ∈X. Consequently, we have proved equalities (26). Given Z ∈Z, let 0 < r = dim span( eT(Z)) ≤p, being here span(D) the linear subspace generated by D ⊂Rp. Let {t1, . . . , tr} ⊂eT(Z) be a basis of span( eT(Z)). Denote Consequently, we have proved equalities (26). Given Z ∈Z, let 0 < r = dim span( eT(Z)) ≤p, being here span(D) the linear subspace generated by D ⊂Rp. Let {t1, . . . , tr} ⊂eT(Z) be a basis of span( eT(Z)). Denote bT(Z) := {t ∈Rp : t =
∑
i∈I
tiγi, Bt ≥0, ||t||∞= 1}, where I := {1, . . . , r}. It is easy to see that the set bT(Z) is a union of a finite number of convex bounded polyhedra. Suppose that t ∈eT(Z). Hence, by construction, Bt ≥0, ||t||∞= 1 and t admits a representation t = ∑
i∈I
tiγi. b( )
e( )
b( ) It is easy to see that the set bT(Z) is a union of a finite number of convex bounded polyhedra. Suppose that t ∈eT(Z). Hence, by construction, Bt ≥0, ||t||∞= 1 and t admits a representation t = ∑
i∈I
tiγi. Hence t ∈bT(Z) and eT(Z) ⊂bT(Z) Consequently Suppose that t ∈T(Z). Hence, by construction, Bt ≥0, ||t||∞
1 and t admits a representation t
∑
i∈I
t γi. Hence t ∈bT(Z) and eT(Z) ⊂bT(Z). Consequently Hence t ∈bT(Z) and eT(Z) ⊂bT(Z). Consequently ∪
Z∈Z
eT(Z) = Tim ⊂
∪
Z∈Z
bT(Z). (27) (27) Now consider any t ∈bT(Z). Denote Denote X := {x ∈Rn : a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J}. (29) (29) Evidently, X ⊂X. Proposition 2 Proposition 2
Consider the SIP problem (8) with the set of immobile indices Tim. Let the set X be defined in (29). Then the
following relations hold true: (30) t⊤A(x)t ≥0 ∀t ∈convTim, ∀x ∈X. (30)
Proof. Let t ∈convTim. Then t = ∑
j∈J
γjt∗(j), ∑
j∈J
γj = 1, γj ≥0, j ∈J. Hence for any x, it holds
t⊤A(x)t =
(∑
γjt∗(j)
)⊤
A(x)
(∑
γjt∗(j)
)
=
∑∑
γiγj(t∗(i))⊤A(x)t∗(j). (31) t⊤A(x)t =
(∑
j∈J
γjt∗(j)
)⊤
A(x)
(∑
j∈J
γjt∗(j)
)
=
∑
i∈J
∑
j∈J
γiγj(t∗(i))⊤A(x)t∗(j). (31) (31) Let us show that for any i ∈J, j ∈J, (t∗(i))⊤A(x)t∗(j) ≥0, ∀x ∈X. (t∗(i))⊤A(x)t∗(j) ≥0, ∀x ∈X. (32) (32) In fact, since the vector t∗(i) belongs to K, it admits a representation In fact, since the vector t∗(i) belongs to K, it admits a representation tor t∗(i) belongs to K, it admits a representation t∗(i) =
∑
s∈S
βsibs +
∑
s∈S∗
αsias,
where αsi ≥0, s ∈S∗. Hence, taking into account (29), we get Hence, taking into account (29), we get (t∗(i))⊤A(x)t∗(j) =
(∑
s∈S
βsibs +
∑
s∈S∗
αsias
)⊤
A(x)t∗(j) =
∑
s∈S∗
αsia⊤
s A(x)t∗(j) ≥0, ∀x ∈X. Inequalities (30) follow from (31) and (32). The proposition is proved. 2 Inequalities (30) follow from (31) and (32). The proposition is proved. 2 Compute By construction, we have At = 0, Bt ≥0, ||t||∞= 1, and, consequently, t ∈T. Since ti ∈eT(Z), i ∈I, it follows from (26) that Now consider any t ∈bT(Z). By construction, we have At = 0, Bt ≥0, ||t||∞= 1, and, consequently, t ∈T. Since ti ∈eT(Z), i ∈I, it follows from (26) that (ti)⊤A(x)tj = 0, i ∈I, j ∈I, ∀x ∈X. (ti)⊤A(x)tj = 0, i ∈I, j ∈I, ∀x ∈X. (ti)⊤A(x)tj = 0, i ∈I, j ∈I, ∀x ∈X. Taking into account these equalities, for any x ∈X, we get t⊤A(x)t =
∑
i∈I
∑
j∈I
γiγj(ti)⊤A(x)tj = 0. This implies that t ∈Tim and hence bT(Z) ⊂Tim. Consequently, ∪
Z∈Z
bT(Z) ⊂Tim. This inclusion together with
(27) imply This implies that t ∈Tim and hence bT(Z) ⊂Tim. Consequently, ∪
Z∈Z
bT(Z) ⊂Tim. This inclusion together with
(27) imply (27) imply
∪
Z∈Z
bT(Z) = Tim. ∪
Z∈Z
bT(Z) = Tim. ∪
Z∈Z
bT(Z) = Tim. Taking into account that |Z| < ∞and the fact that each set bT(Z) is a union of a finite number of convex bounded
polyhedra, we conclude from the latter equality that the statement of the lemma is proved. 2 Consider the set {t∗(j), j ∈J} of vertices of the polyhedron conv Tim. Recall that in the case Tim = ∅, we have
conv Tim = ∅and J = ∅. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 O. KOSTYUKOVA AND T. TCHEMISOVA
677 O. KOSTYUKOVA AND T. TCHEMISOVA O. KOSTYUKOVA AND T. TCHEMISOVA
677 677 Suppose that Tim ̸= ∅. Since t∗(j) ∈Tim for any j ∈J, it follows from Proposition 1 that the following relations
hold true:
a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J, ∀x ∈X. (28) Suppose that Tim ̸= ∅. Since t∗(j) ∈Tim for any j ∈J, it follows from Proposition 1 that the following relations
hold true:
a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J, ∀x ∈X. (28) a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J, ∀x ∈X. (28 (28) Proof. It is evident that X ⊂X(ε) for all ε > 0. Since the sets {1, . . . , p} and {1, . . . , m∗} are finite, there exists a sequence εk > 0, k = 1, 2, ... and sets P ±, N 0
such that lim
k→∞εk = 0, P ±(εk) = P ±, and N 0(εk) = N 0 for sufficiently large k > 0. To simplify the designation,
without loss of generality, we can consider that where tk(ε) denotes the k-th component of the vector t(ε) and B⊤
k stays for the k-th raw of the matrix B ∈Rm∗×p. Since the sets {1, . . . , p} and {1, . . . , m∗} are finite, there exists a sequence εk > 0, k = 1, 2, ... and sets P ±, N 0
such that lim
k→∞εk = 0, P ±(εk) = P ±, and N 0(εk) = N 0 for sufficiently large k > 0. To simplify the designation,
without loss of generality, we can consider that k→∞
without loss of generality, we can consider that P ±(ε) = P ± and N 0(ε) = N 0 for sufficiently small ε > 0. (37) (37) Given a sufficiently small ε > 0, consider vector l(ε) := t(ε) −t∗. It is easy to see that due to the convexity of the
set eT, we have t∗+ λ(t(ε) −t∗) = t∗+ λl(ε) ∈eT for all λ ∈[0, 1]. Hence l(ε) is a feasible direction for t∗in eT. Since by construction, Al(ε) = 0, then, according to definition, the direction l(ε) will be feasible for t(ε) in
the set eT if there exists θ0 > 0 such that −1 ≤tk(ε) + θlk(ε) ≤1, k = 1, ..., p, and B(t(ε) + θl(ε)) ≥0, for all
θ ∈[0, θ0]. Hence the direction l(ε) is feasible for t(ε) in the set eT if Given a sufficiently small ε > 0, consider vector l(ε) := t(ε) −t∗. It is easy to see that due to the convexity of the
set eT, we have t∗+ λ(t(ε) −t∗) = t∗+ λl(ε) ∈eT for all λ ∈[0, 1]. Hence l(ε) is a feasible direction for t∗in eT. Given a sufficiently small ε > 0, consider vector l(ε) := t(ε) −t∗. It is easy to see that due to the convexity of the
set eT, we have t∗+ λ(t(ε) −t∗) = t∗+ λl(ε) ∈eT for all λ ∈[0, 1]. Proof. It is evident that X ⊂X(ε) for all ε > 0. uch that X(ε0) ⊂X. Suppose the contrary. Then for all ε > 0 there exists ( )
hat there exists ε0 > 0 such that X(ε0) ⊂X. Suppose the contrary. Then for all ε > 0 there exis ( )
Let us show that there exists ε0 > 0 such that X(ε0) ⊂X. Suppose the contrary. Then for all ε > 0 there exists
x(ε) ∈X(ε) such that
(t(ε))⊤A(x(ε))t(ε) < 0,
(35)
⊤ ( )
Let us show that there exists ε0 > 0 such that X(ε0) ⊂X. Suppose the contrary. Then for all ε > 0 there exists
x(ε) ∈X(ε) such that
(t(ε))⊤A( (ε))t(ε) < 0
(35) ( )
hat there exists ε0 > 0 such that X(ε0) ⊂X. Suppose the contrary. Then for all ε > 0 there exis
h that (t(ε))⊤A(x(ε))t(ε) < 0,
(35) (35) where t(ε) ∈arg{min
t
t⊤A(x(ε))t, s.t. t ∈T}. where t(ε) ∈arg{min
t
t⊤A(x(ε))t, s.t. t ∈T}. t
It is easy to prove that t(ε) is also an optimal solution of the problem t
It is easy to prove that t(ε) is also an optimal solution of the problem minimize t⊤A(x(ε))t, subject to t ∈eT,
(36) minimize t⊤A(x(ε))t, subject to t ∈eT, minimize t⊤A(x(ε))t, subject to t ∈eT,
(36) (36) where eT := {t ∈K : ∥t ∥∞≤1}. Notice that T ⊂eT, and the set eT is convex. By construction (see Proposition 2
and definition (34)), it holds where eT := {t ∈K : ∥t ∥∞≤1}. Notice that T ⊂eT, and the set eT is convex. By construction (see Proposition 2
and definition (34)), it holds where eT := {t ∈K : ∥t ∥∞≤1}. Notice that T ⊂eT, and the set eT is convex. By construction (see Proposition 2
and definition (34)), it holds t⊤A(x(ε))t ≥0, ∀t ∈T(ε) ∪conv Tim. t⊤A(x(ε))t ≥0, ∀t ∈T(ε) ∪conv Tim. Hence we can conclude that t(ε) ∈d
T(ε) \ conv Tim and there exists a limiting point t∗of the sequence t(ε), ε →0
such that t∗∈conv Tim ⊂T. To simplify notation without loss of generality suppose that t∗= lim
ε→0 t(ε). Set P ±(ε) := {k ∈{1, . . . , p} : tk(ε) = ±1}, N 0(ε) := {k ∈{1, . . . , m∗} : B⊤
k t(ε) = 0}, where tk(ε) denotes the k-th component of the vector t(ε) and B⊤
k stays for the k-th raw of the matrix B ∈Rm∗×p. 4.3. The properties of the feasible set of problem (8) In this section, we will obtain some properties of the feasible set of problem (8). These properties are closely
connected with the properties of the elements of the convex hull of the set of normalized immobile indices in this
problem. D
t Denote T(ε) := {t ∈T, ρ(t, conv Tim) ≥ε}, d
T(ε) := {t ∈T, ρ(t, conv Tim) ≤ε},
(33 (33) X(ε) := {x ∈X : t⊤A(x)t ≥0 ∀t ∈T(ε)}, X(ε) := {x ∈X : t⊤A(x)t ≥0 ∀t ∈T(ε)},
(34) (34) where ε > 0, the set X is defined in (29), the distance between a vector l and a set B in Rp is defined as
ρ(l, B) = min
τ∈B ||l −τ||, and ||a|| stays for a norm of vector a ∈Rp. Lemma 4
Consider the SIP problem (8) with the set of immobile indices Tim and the feasible set X. There exists ε0 > 0 such
that X(ε0) = X, where the set X(ε) is defined in (34). Lemma 4 Consider the SIP problem (8) with the set of immobile indices Tim and the feasible set X. There exists ε0 > 0 such
that X(ε0) = X, where the set X(ε) is defined in (34). Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 678 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION Proof. It is evident that X ⊂X(ε) for all ε > 0. Proof. It is evident that X ⊂X(ε) for all ε > 0. TCHEMISOVA
679 O. KOSTYUKOVA AND T. TCHEMISOVA O. KOSTYUKOVA AND T. TCHEMISOVA
679 679 As σ∗∈(0, σ0), one can conclude from (38) that 2aσ∗+ 2b = 2a + 2b = 0. Then −a = b and we get the following
equivalent equalities: As σ∗∈(0, σ0), one can conclude from (38) that 2aσ∗+ 2b = 2a + 2b = 0. Then −a = b and we get the following
equivalent equalities: −(l(ε))⊤A(x(ε))t∗= (l(ε))⊤A(x(ε))l(ε)
⇐⇒
(t(ε))⊤A(x(ε))t∗= (t(ε))⊤A(x(ε))t(ε). (39) (39) By construction, t∗∈conv Tim. Therefore t∗= ∑
j∈J
γjt∗(j),
∑
j∈J
γj = 1, γj ≥0, j ∈J. By construction, t∗∈conv Tim. Therefore t∗= ∑
j∈J
γjt∗(j),
∑
j∈J
γj = 1, γj ≥0, j ∈J. Since t(ε) ∈eT ⊂K, then t(ε) = ∑
s∈S
βsbs + ∑
s∈S∗
αsas, where αs ≥0, s ∈S∗. Consequently, taking into
account the inclusion x(ε) ∈X(ε) ⊂X, where X is defined in (29), we have Since t(ε) ∈eT ⊂K, then t(ε) = ∑
s∈S
βsbs + ∑
s∈S∗
αsas, where αs ≥0, s ∈S∗. Consequently, taking into
account the inclusion x(ε) ∈X(ε) ⊂X, where X is defined in (29), we have (t(ε))⊤A(x(ε))t∗=
( ∑
s∈S
βsbs + ∑
s∈S∗
αsas
)⊤
A(x(ε)) ∑
j∈J
γjt∗(j)
= ∑
s∈S∗
∑
j∈J
αsγja⊤
s A(x(ε))t∗(j) ≥0. The latter inequality and inequality (35) contradict the equality (39). The lemma is proved. 2 The next lemma can be proved using the same reasoning scheme as in the proof of Lemma 3 from [20]. Lemma 5 The next lemma can be proved using the same reasoning scheme as in the proof of Lemma 3 from [20]. Lemma 5
Given SIP problem (8) with the set of immobile indices Tim, for any ε > 0 there exists a vector x(ε) ∈X such that
t⊤A(x(ε))t > 0 ∀t ∈T(ε),
(40) (40) where the sets X and T(ε) are defined in (29), (33). where the sets X and T(ε) are defined in (29), (33). where the sets X and T(ε) are defined in (29), (33). Proof. It is evident that X ⊂X(ε) for all ε > 0. Hence l(ε) is a feasible direction for t∗in eT. Since by construction, Al(ε) = 0, then, according to definition, the direction l(ε) will be feasible for t(ε) in
the set eT if there exists θ0 > 0 such that −1 ≤tk(ε) + θlk(ε) ≤1, k = 1, ..., p, and B(t(ε) + θl(ε)) ≥0, for all
θ ∈[0, θ0]. Hence the direction l(ε) is feasible for t(ε) in the set eT if ( ( )
)
( )
[ , ]
( )
Since by construction, Al(ε) = 0, then, according to definition, the direction l(ε) will be feasible for t(ε) in
the set eT if there exists θ0 > 0 such that −1 ≤tk(ε) + θlk(ε) ≤1, k = 1, ..., p, and B(t(ε) + θl(ε)) ≥0, for all
θ ∈[0, θ0]. Hence the direction l(ε) is feasible for t(ε) in the set eT if lk(ε) ≤0 for k ∈P +; lk(ε) ≥0 for k ∈P −; B⊤
k l(ε) ≥0 for k ∈N 0. The relations above are fulfilled since it follows from (37) that lk(ε) := tk(ε) −t∗
k = 0, k ∈P + ∪P −and
B⊤
k l(ε) = B⊤
k (tk(ε) −t∗
k) = 0 for k ∈N 0. Consequently, the direction l(ε) is feasible for both vectors, t∗and
t(ε), in the set eT and, hence, The relations above are fulfilled since it follows from (37) that lk(ε) := tk(ε) −t∗
k = 0, k ∈P + ∪P −and
B⊤
k l(ε) = B⊤
k (tk(ε) −t∗
k) = 0 for k ∈N 0. Consequently, the direction l(ε) is feasible for both vectors, t∗and
t(ε), in the set eT and, hence, ¯t(σ) := t∗+ σl(ε) = t∗+ σ(t(ε) −t∗) ∈eT, ∀σ ∈[0, σ0], where σ0 > 1. For a fixed ε > 0 that satisfies the assumptions above, define the function ed ε > 0 that satisfies the assumptions above, define the function (σ) := (¯t(σ))⊤A(x(ε))¯t(σ) = (t∗)⊤A(x(ε))t∗+ 2σ(l(ε))⊤A(x(ε))t∗+ σ2(l(ε))⊤A(x(ε))l(ε) = aσ2 + 2bσ + c, σ ∈[0, σ0], where c := (t∗)⊤A(x(ε))t∗, b := (l(ε))⊤A(x(ε))t∗, a := (l(ε))⊤A(x(ε))l(ε). By construction, for σ∗= 1 we have ¯t(σ∗) = t(ε). Then w(σ∗) = (t(ε))⊤A(x(ε))t(ε), and w(σ∗) is the optimal
value of the cost function of the problem (36). Hence w(σ∗) =
min
σ∈[0,σ0] w(σ) =
min
σ∈[0,σ0](aσ2 + 2bσ + c). (38) (38) Stat., Optim. Inf. Comput. Vol. 8, September 2020 KOSTYUKOVA AND T. 5. The proof of Theorem 1 We will prove here Theorem 1 for problem (8). Taking into account that problems (3) and (8) are equivalent, the
theorem will be valid also for the K-semidefinite problem (3). Necessity. Let x0 ∈X be an optimal solution of the problem. Choose ε0 > 0 as in Lemma 4 and consider a
perturbed SIP problem minimize c(x),
subject to x ∈X(ε0), 8, September 2020 OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION 680 such that the following relations hold: such that the following relations hold: such that the following relations hold: −∂c(x0)
∂xk
+ ∑
i∈I
y(i)(t(i))⊤Akt(i) + ∑
j∈J
( ∑
s∈S∗
αsjas + ∑
s∈S
βsjbs
)⊤
Akt∗(j) = 0, k = 1, ..., n;
(43)
(t(i))⊤A(x0)t(i) = 0, i ∈I; αsja⊤
s A(x0)t∗(j) = 0, s ∈S∗, j ∈J. (44) (43) (44) Denote τ(i) :=
√
y(i) t(i), i ∈I, λ(j) := 1
2
( ∑
s∈S∗
αsjas + ∑
s∈S
βsjbs
)
∈K, j ∈J. It is easy to see that relations
b
i
i
h f
i h
i
d
d fi
d i
h
i
i
d Denote τ(i) :=
√
y(i) t(i), i ∈I, λ(j) := 1
2
( ∑
s∈S∗
αsjas + ∑
s∈S
βsjbs
)
∈K, j ∈J. It is easy to see that relations
(43), (44) can be written in the form (11) with matrices U and V defined in (12). The necessity is proved. Denote τ(i) :
√
y(i) t(i), i ∈I, λ(j) :
2
( ∑
s∈S∗
αsjas + ∑
s∈S
βsjbs
)
∈K, j ∈J. It is easy to see that relations
(43), (44) can be written in the form (11) with matrices U and V defined in (12). The necessity is proved. ∗
(43), (44) can be written in the form (11) with matrices U and V defined in (12). The necessity is proved. Sufficiency. Suppose that for x0 ∈X, there exist vectors (10) such that for matrices (12) the relations (11) hold
true. Then it is easy to show that the vector x0 is an optimal solution in the following convex problem with linear
constraints: minimize c(x),
(CP) :
subject to x ∈Rn, (τ(i))⊤A(x)τ(i) ≥0, i ∈I,
a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J. It is evident that the feasible set X of problem (8) belongs to the set of feasible solutions of the problem (CP). Hence the optimality of x0 ∈X in (CP) implies the optimality of x0 in the original problem (8). The theorem is
proved. 2 It is evident that the feasible set X of problem (8) belongs to the set of feasible solutions of the problem (CP). minimize c(x),
subject to x ∈X(ε0), minimize c(x),
subject to x ∈X(ε0), minimize c(x),
subject to x ∈X(ε0), where the set X(ε) is defined in (34). This problem can be rewritten as follows: ε) is defined in (34). This problem can be rewritten as follows: minimize c(x),
subject to x ∈Rn, t⊤A(x)t ≥0 ∀t ∈T(ε0),
a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J. (41) minimize c(x), (41) o x ∈Rn, t⊤A(x)t ≥0 ∀t ∈T(ε0),
a⊤A(x)t∗(j) ≥0 s ∈S ; b⊤A(x)t∗(j) = 0 s ∈S j ∈J
(41) a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J. a⊤
s A(x)t∗(j) ≥0, s ∈S∗; b⊤
s A(x)t∗(j) = 0, s ∈S, j ∈J. According to Lemma 4, the set of feasible solutions of problem (41) coincides with the set of feasible solutions of
the original problem (8). Hence vector x0 is an optimal solution of the perturbed problem (41) as well. Problem (41) has the following important properties: Problem (41) has the following important properties: • the index set T(ε0) is compact, • the index set T(ε0) is compact, • the index set T(ε0) is compact, (
)
p
aints satisfy the Slater type condition (see Lemma 5). • the constraints satisfy the Slater type condition (see Lemma 5). • the constraints satisfy the Slater type condition (see Lemma 5). • the constraints satisfy the Slater type condition (see Lemma 5). Hence, applying the classic optimality conditions (see, for example, Theorem 5.107 in [7]) for the optimal
solution x0 of problem (41), we can state that there exist numbers and vectors Hence, applying the classic optimality conditions (see, for example, Theorem 5.107 in [7]) for the optimal
solution x0 of problem (41), we can state that there exist numbers and vectors y(i) > 0, t(i) ∈T(ε0), i ∈I, |I| ≤n; αsj ≥0, s ∈S∗, βsj ∈R, s ∈S, j ∈J,
(42) y(i) > 0, t(i) ∈T(ε0), i ∈I, |I| ≤n; αsj ≥0, s ∈S∗, βsj ∈R, s ∈S, j ∈J, y(i) > 0, t(i) ∈T(ε0), i ∈I, |I| ≤n; αsj ≥0, s ∈S∗, βsj ∈R, s ∈S, j ∈J,
(42) (42) Stat., Optim. Inf. Comput. Vol. minimize c(x),
subject to x ∈X(ε0), Hence the optimality of x0 ∈X in (CP) implies the optimality of x0 in the original problem (8). The theorem is
proved. 2 Stat., Optim. Inf. Comput.
Vol. 8, September 2020 6. Dual formulations for the linear K- semidefinite problem (3): standard Lagrangian and regularized dual
problems These cones are explicitly generated by the finite set {t∗(j), j ∈J} of vertices of the polyhedron conv Tim, where
Tim is the normalized immobile index set in problem (45). p
One can show that for any j ∈J, the cone CK(j) is convex and closed and the corresponding dual cone has the
form (CK(j))∗= {t(t∗(j))⊤+ t∗(j)t⊤: t ∈K}. Denote K := (
∩
j∈J
CK(j)) ∩CK. Evidently, K ⊂CK, where the cone CK is defined in (1). It can be shown (see e.g. [4, 6]) that the dual cone to K
has the form
(
) Evidently, K ⊂CK, where the cone CK is defined in (1). It can be shown (see e.g. [4, 6]) that the dual cone to K
has the form CK is defined in (1). It can be shown (see e.g. [4, 6]) that the dual cone to K
K∗= cl
(∑
j∈J
(CK(j))∗+ (CK)∗)
. K∗= cl
(∑
j∈J
(CK(j))∗+ (CK)∗)
. 6. Dual formulations for the linear K- semidefinite problem (3): standard Lagrangian and regularized dual
problems In this section, we will suppose that the cost function of the special conic problem (3) is linear and discuss some
new dual formulations for this problem. In our papers [17] and [20], we derived regularized primal and dual formulations for linear copositive problems. These formulations are explicit and guarantee the strong duality. In this section, in a similar way, we will derive
new strong dual formulations for the K-semidefinite problem (3). To obtain these results, we will use Theorem 2. Consider a K-semidefinite problem with linear cost function in the form midefinite problem with linear cost function in the form minimize c⊤x
subject to A(x) ∈CK, x ∈Rn,
(45) (45) re c = (c1, c2, . . . , cn), function A(x) and the polyhedral set K are defined as in (3). First of all, let us formulate for problem (45), the (standard) Lagrangian dual problem. According to [24], it has
the form: maximize(−W • A0),
subject to −ck + W • Ak = 0, k = 1, 2, ..., n; W ∈(CK)∗,
(46) maximize(−W • A0), (46) = conv {ll⊤: l ∈K} is the dual cone to the cone CK. where the cone (CK)∗= conv {ll⊤: l ∈K} is the dual cone to the cone CK. (
)
{
}
It is a known fact (see e.g.[24]) that if the constraints of problem (45) satisfy the Slater condition, then there is
no gap between the optimal values of this problem and its Lagrangian dual. It is a known fact (see e.g.[24]) that if the constraints of problem (45) satisfy the Slater condition, then there is
no gap between the optimal values of this problem and its Lagrangian dual. If the constraints of problem (45) do not satisfy the Slater condition, then the positive gap is possible even in the
case when the primal problem has an optimal solution. Consider the following cones: CK(j) := {D ∈S(p) : t⊤Dt∗(j) ≥0 ∀t ∈K}, j ∈J. CK(j) := {D ∈S(p) : t⊤Dt∗(j) ≥0 ∀t ∈K}, j ∈J. Stat., Optim. Inf. Comput. Vol. 8, September 2020 Stat., Optim. Inf. Comput. Vol. 8, September 2020 STYUKOVA AND T. TCHEMISOVA
681 O. KOSTYUKOVA AND T. TCHEMISOVA
681 O. KOSTYUKOVA AND T. TCHEMISOVA 681 These cones are explicitly generated by the finite set {t∗(j), j ∈J} of vertices of the polyhedron conv Tim, where
Tim is the normalized immobile index set in problem (45). Hence e
K∗= cl{W ∈S(p) : W = U + V, U ∈(CK)∗, V ∈V(p)},
(47)
V( ) i d fi
d i (13) K∗= cl{W ∈S(p) : W = U + V, U ∈(CK)∗, V ∈V(p)},
(47) (47) where V(p) is defined in (13). ( )
Taking into account Proposition 1, one can show that the K-semidefinite problem (45) can be rewritten in the
conic form Taking into account Proposition 1, one can show that the K-semidefinite problem (45) can be rewritten in the
conic form minimize c⊤x, subject to x ∈Rn, A(x) ∈K ⊂CK. (48) minimize c⊤x, subject to x ∈Rn, A(x) ∈K ⊂CK. (48) (48) Let us refer to problem (48) as a regularized primal (set-semidefinite) problem. The dual problem to (48) (the
regularized dual problem) has the form maximize(−W • A0),
subject to −ck + W • Ak = 0 ∀k = 1, 2, ..., n, W ∈K∗. (49) maximize(−W • A0),
(49 (49) (
)
subject to −ck + W • Ak = 0 ∀k = 1, 2, ..., n, W ∈K∗. (49) It is easy to show that, given any feasible solution x of problem (48) and any feasible solution W of the dual
problem (49), the following inequality (weak duality) is satisfied: c⊤x ≥−W • A0. c⊤x ≥−W • A0. Moreover, taking into account Theorem 2 and representation (47), we can conclude that for any optimal solution
x0 of problem (48), there exists a feasible solution W 0 = U 0 + V 0, U 0 ∈(CK)∗, V 0 ∈V(p), of the dual problem
(49) such that the strong duality property holds: c⊤x0 = −W 0 • A0. (50) (50) This conclusion implies the following proposition. Proposition 3 7. Conclusions The main contribution of the paper consists in the formulation of new CQ-free optimality conditions and strong
duality results for set-semidefinite programming problems. To obtain these results, we used the approach, suggested
in our previous papers for convex SIP, SDP, and copositive problems, and based on the concept of immobile indices. The main conclusions we can draw from the results of the paper, are the following. The main contribution of the paper consists in the formulation of new CQ-free optimality conditions and strong
duality results for set-semidefinite programming problems. To obtain these results, we used the approach, suggested
in our previous papers for convex SIP, SDP, and copositive problems, and based on the concept of immobile indices. p
p p
p
p
p
The main conclusions we can draw from the results of the paper, are the following. p
p p
p
p
The main conclusions we can draw from the results of the paper, are the followin • The concept of immobile indices being applied to set-semidefinite optimization permits one to obtain new
optimality conditions. • The concept of immobile indices being applied to set-semidefinite optimization permits one to obtain new
optimality conditions. • The set of immobile indices has a special structure and its study is important. Notice that the optimality
conditions formulated in Theorems 1 and 2 do not use the proper normalized immobile indices but are
formulated with the help of a finite number of vertices of the convex hull of the normalized immobile index
set. In the future, we plan to obtain CQ-free optimality conditions and strong duality results without explicit
use of there vertices as well. • The optimality conditions and strong duality relations obtained in the paper for the special set-semidefinite
problems may be used to develop numerical methods for solving these problems. • The optimality conditions and strong duality relations obtained in the paper for the special set-semidefinite
problems may be used to develop numerical methods for solving these problems. OPTIMALITY CONDITIONS AND STRONG DUALITY FOR A SPECIAL CONIC OPTIMIZATION In [24, 30], a CQ-free duality theory for conic optimization was developed in terms of the so-called minimal
cone. Being quite general, this theory has one disadvantage in terms of its application, namely, it is very abstract. To the best of our knowledge, for problem (45), there are no other explicit CQ-free dual formulations satisfying
strong duality relations. In [24, 30], a CQ-free duality theory for conic optimization was developed in terms of the so-called minimal
cone. Being quite general, this theory has one disadvantage in terms of its application, namely, it is very abstract. g q
g
,
y
g
pp
,
y,
y
To the best of our knowledge, for problem (45), there are no other explicit CQ-free dual formulations satisfying
strong duality relations. g
y
We plan to dedicate a special paper to the detailed comparison of the new conic formulations obtained using the
notion of the immobile indices with that from [24, 30]. Acknowledgements This work was partially supported by state research program “Convergence”(Republic Belarus), Task 1.3.01, by
Portuguese funds through CIDMA - Center for Research and Development in Mathematics and Applications, and
FCT - Portuguese Foundation for Science and Technology, within the project UID/MAT/04106/2019. g
gy
p
j
The authors are grateful to the anonymous reviewers for careful reading of the paper and for their constructive
comments and advices. 10.
D¨ur M. Copositive Programming – a Survey, In: Diehl M, Glineur F, Jarlebring E, Michielis W., editors. Recent advances in
Optimization and its applications in Engineering; Springer-Verlag Berlin Heidelberg X1: 535 p., 2010. Proposition 3 Proposition 3 p
Suppose that the K-semidefinite problem (45) has an optimal solution. Then
ti
l
l ti
f th
l i
d d
l
bl
(49)
i t
d e that the K-semidefinite problem (45) has an optimal solution. Then p
Suppose that the K-semidefinite problem (45) has an optimal solution. Then • an optimal solution of the regularized dual problem (49) exists, and • an optimal solution of the regularized dual problem (49) exists, and • there is no gap between the optimal values of problem (45) (equivalently, the regularized primal K-
semidefinite problem (48)), and its regularized dual problem (49). • there is no gap between the optimal values of problem (45) (equivalently, the regularized primal K-
semidefinite problem (48)), and its regularized dual problem (49). From this proposition, it follows that the strong duality is guaranteed for the primal problem (45) and its
regularized dual problem (49), while, as it was mentioned above, for the pair constituted by the primal problem
(45) and its (standard) Lagrangian dual problem (46), the strong duality may fail. The main contribution of this section is to obtain for the set-semidefinite problem (45) a regularized dual one
(49) satisfying the strong duality conditions without CQs or any other assumptions. The dual problem (49) has an
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constraints in the primal problem. Stat., Optim. Inf. Comput. Vol. 8, September 2020 682 9.
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Application of non-traditional raw materials in the production of low-humidity bakery products
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* Corresponding author: kireevagos@mail.ru © The Authors, published by EDP Sciences. This is an open access article distributed under the terms of the Creative Commons
Attribution License 4.0 (https://creativecommons.org/licenses/by/4.0/). Application of non-traditional raw materials in
the production of low-humidity bakery products
Olga Kireeva1*, Tatiana Lazareva1, Nikita Murlenkov1, Natalya Berezina1,
Marina Yarkina1, Sergey Zhuchkov1, Vladimir Kryukov1, Oksana Safronova2, Elena
Anatol'yevna Kuznetsova2, and Elena Aleksandrovna Kuznetsova2 1Orel State Agrarian University named after N.V. Parakhin, Generala Rodina str. 69, Orel, 302019
Russia 2Orel State University named after I.S. Turgenev, Komsomol’skaya str. 95, Orel, 302026, Russia Abstract. Recently, the use of non-traditional plant components as a
functional ingredient in the development of fortified products for healthy
and therapeutic nutrition has been very relevant. An analysis was carried out
of the influence of the type of functional additive from brown algae and
blackcurrant pomace powder on the organoleptic and physico-chemical
properties of bakery products of reduced importance - wafer bread. A
decrease in dough density, an increase in humidity and wetness of finished
products was established when functional ingredients were added to the
wafer bread recipe. An increase in the acidity of the experimental samples
of enriched wafer breads was noted in comparison with the control sample. An increase in the acidity of the experimental samples of enriched wafer
breads was noted in comparison with the control sample with a slight change
in their organoleptic characteristics. A high content of iodine and dietary
fiber in new types of bakery products with reduced humidity has been
established. E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 https://doi.org/10.1051/e3sconf/202448602011 1 Introduction Recently, the development of recipes for enriched bakery products has been very promising,
because bread and bakery products are products of everyday demand and are present daily in
the diet of the vast majority of consumers. One of the non-traditional plant components
increasingly used in the development of new bakery products for healthy and therapeutic
nutrition is brown algae [1-3]. Brown seaweed is a valuable source of biologically active substances that have preventive
and therapeutic effects. In terms of the content of macro- and micronutrients, brown algae
are significantly superior to terrestrial plants, and are also the only source of a number of
polysaccharides (alginates, carrageenan, laminarin, fucoidan), which have bacteriostatic,
bactericidal, anti-inflammatory, immunomodulatory, antioxidant and wound-healing effects
[4-6]. The polysaccharide components of algae help correct lipid metabolism, reduce body
weight, have a pronounced antidiabetic effect, have antihypertensive and antitumor effects, E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 https://doi.org/10.1051/e3sconf/202448602011 can prevent the development of oxidative stress in living organisms or reduce its intensity,
etc. [7-11]. can prevent the development of oxidative stress in living organisms or reduce its intensity,
etc. [7-11]. Domestic and foreign studies of the biological activity of both individual compounds
isolated from brown algae and drugs based on them confirm the medicinal properties and
preventive effects and explain the widespread use of brown algae in the development of
medicines and biologically active food additives [6, 11-13 ]. Brown algae have found application in the creation of healthy and functional food
products as a source of bioactive food components, including iodine, which help prevent
iodine deficiency conditions [14-20]. Numerous scientific studies on the development of
technologies for processing brown algae, the development of recipes and technologies for
creating food products enriched with bioactive nutrients from algae, especially based on mass
consumption products (dairy, meat products, bakery products, etc.) confirm the relevance of
work in this direction. The prospects for the use of resource-saving, waste-free technologies in the food industry
also explain the recent increased interest in the use of by-products of juice production
(vegetable and berry pomace) in the formulations of functional food products in order to
enrich the latter with biologically active substances (mineral elements, dietary fiber) [21, 22]. The use of juice production by-products in the development of new food products makes it
possible to increase the economic efficiency of processing enterprises by converting
production waste into food ingredients. 1 Introduction The development of food products using renewable natural resources (brown algae), as
well as waste from the production of juices and drinks (berry pomace), makes it possible to
increase the nutritional and biological value of new products, the use of which in the daily
diet will help to compensate for the deficiency of macro- and micronutrients caused by
insufficient or an unbalanced diet, as well as excessive consumption of refined foods. When developing recipes for food products enriched with non-traditional functional
ingredients, it is advisable to study the influence of functional additives on the
physicochemical and organoleptic characteristics of the finished product to avoid a decrease
in its quality in comparison with the traditional one [2, 23, 24]. In this regard, the purpose of
the study was to study the effect of a type of brown algae additive in combination with
blackcurrant pomace powder on the organoleptic and physicochemical properties of low-
humidity bakery products (wafer bread) for further optimization of the product formulation. E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 Before kneading the dough, wheat flour was first sifted through a sieve. Powders from
brown algae were obtained by grinding crushed fucus and crushed kelp (TU 03.11.63-005-
41669896-2019, manufacturer Arkhangelsk Algae Plant LLC) to a particle size of no more
than 0.3 mm. Powder from blackcurrant pomace was obtained by grinding pre-dried
blackcurrant pomace remaining after extracting the juice by pressing to a fine state. In the recipes for waffle breads, peeled rye baking flour and premium-grade wheat baking
flour were used in a ratio of 2:1. Salt, sugar and sunflower oil were added to the recipe in
amounts of 1.5%, 3% and 8.3% by weight of flour, respectively. When producing
experimental samples of wafer bread, powders from brown algae and blackcurrant pomace
were added to the recipe in an amount of 1% of each component by weight of flour. The moisture content of the dough was determined by drying a sample of the semi-
finished dough product (4–5 g) in an oven at a temperature of 105°C to constant weight
(about 4–5 hours). In this case, drying was completed when the difference between the last
two weighings did not exceed 0.01 g. Dough density was calculated as the ratio of dough mass to volume, expressed in g/cm3
(measurements were carried out at a temperature of 20ºC). Determination of the moisture
content of wafer breads was carried out according to GOST 8494-96, acidity - according to
GOST 5670-96, the content of the mass fraction of fat was determined according to GOST
5668-68, the mass fraction of sugar - according to GOST 5672-68, wetness was studied
according to GOST 10114-80. The fiber content was determined using a Fibertek 1020 device
(Foss), the iodine content was determined by the titrimetric method according to MUK
4.1.1106–02. 2 Materials and methods At the Innovative Research Testing Center for Collective Use of the Orel State Agrarian
University, a recipe for enriched wafer bread containing brown algae powder as a functional
ingredient was developed and patented [25]. Previously, studies were carried out on the
chemical composition and nutritional value of enriched wafer breads when Fucus vesiculosus
powder was used in the recipe as an iodine-containing additive [26]. Kelp is known to contain
a significantly higher amount of iodine [6], and blackcurrant pomace powder is a source of
dietary fiber, macro- and microelements. In this connection, to analyze the effect of the type
of brown algae additive in combination with black currant pomace powder on the quality
indicators of bakery products with reduced humidity, fucus and kelp powders were used as
functional ingredients in this study. Thus, the objects of the study were: y p
y
p p
functional ingredients in this study. Thus, the objects of the study were:
rye-wheat waffle bread (control);
rye-wheat waffle bread (control);
rye-wheat waffle bread with the addition of fucus algae powder and blackcurrant
pomace powder (sample No. 1);
rye-wheat waffle bread with the addition of fucus algae powder and blackcurrant
pomace powder (sample No. 1);
rye-wheat waffle bread with the addition of kelp algae powder and blackcurrant
pomace powder (sample No. 2). 2 https://doi.org/10.1051/e3sconf/202448602011 E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 3 Results and discussion The results of the study of the moisture content of the test of control and experimental
samples of rye-wheat waffle bread showed that the test humidity of all the studied samples
was the same within the margin of error, the difference was less than 1% (Figure 1). The
density of the rye-wheat bread dough when using the addition of brown algae powder
decreased by 1.74% compared to the control sample and 4.35% when adding fucus and kelp
additives to the recipe, respectively. a)
b)
Fig. 1. Density (a) and humidity of dough (b) samples of waffle loaves. a) b) b) a) Fig. 1. Density (a) and humidity of dough (b) samples of waffle loaves. The results of the study of the effect of the type of brown algae additive and blackcurrant
powder on the physico-chemical parameters of finished waffle rye-wheat loaves are
presented in Table 1. 3 https://doi.org/10.1051/e3sconf/202448602011 E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 Table 1. Physico-chemical parameters of rye-wheat waffle bread. Indicators
Control sample
Sample No. 1
Sample No. 2
Moisture content, %
5.19±0.02
6.52±0.04
6.24±0.06
Acidity, degrees
2.30±0.10
3.97±0.08
3.56±0.06
Mass fraction of sugar in terms of
dry matter (weight), %
4.02±0.10
4.24±0.14
4.19±0.12
Mass fraction of fat in terms of
dry matter (weight), %
7.57±0.19
7.93±0.21
7.55±0.22 Table 1. Physico-chemical parameters of rye-wheat waffle bread. When adding brown algae powder (regardless of the type of algae) and blackcurrant
pomace powder to the recipe for rye-wheat wafer bread, an increase in the moisture content
of the finished products was noted in comparison with the control, which is probably due to
the swelling of hydrocolloids contained in functional additives and having high moisture-
holding capacity. This prevents the free removal of moisture during baking and leads to an
increase in the moisture content of the finished product. The acidity of the experimental
samples of wafer breads increases by 1.67 degrees compared to the control sample. and by
1.26 degrees. when using kelp and fucus powder in the formulation, respectively. The results
obtained are due to the high acidity of blackcurrant pomace powder due to the high content
of organic acids in blackcurrant berries and their processed products. 3 Results and discussion In terms of sugar and fat content, the experimental samples of rye-wheat waffle bread did
not differ significantly from the control sample, which showed that the presence of the
selected amount in the recipe, as well as the type of functional additive, did not influence the
content of these macronutrients. The use of non-traditional herbal additives in the recipes of low-humidity bakery products
should not negatively affect the sensory characteristics of the finished product, and therefore,
an organoleptic evaluation of samples of rye-wheat waffle bread was carried out (Table 2). Table 2. Organoleptic evaluation of rye-wheat wafer breads. Indicator
name
Characteristic
Control
Sample No. 1
Sample No. 2
Physical appearance
form
The breads are flat, thin, round or slightly oval, without cracks
surface
Rough with a
characteristic relief
from baking, without
swelling or inclusions
Rough with a
characteristic relief
from baking, without
swelling, with small
dark inclusions
Rough with a
characteristic relief
from baking, without
swelling with small
light inclusions
color
Light gray to light
brown
From light gray to light
brown with a pinkish
tint
From light gray to light
brown with a pinkish
tint
fragility
Products are fragile, slightly breakable
sectional view
Baked, dry, evenly baked
taste
Characteristic of this
type of products,
without extraneous taste
Characteristic of this
type of product, with a
slight taste of algae
Characteristic of this
type of product, with a
slight taste of algae
smell
Characteristic for this
type of product, without
foreign odor
Characteristic of this
type of product with a
slight aroma of algae
Characteristic of this
type of product with a
slight aroma of algae
The results of organoleptic evaluation of samples of rye-wheat waffle loaves showed the
presence of small dark inclusions, as well as the presence of a slightly palpable taste and
aroma characteristic of algae in experimental samples of waffle loaves. When used in the Table 2. Organoleptic evaluation of rye-wheat wafer breads. The results of organoleptic evaluation of samples of rye-wheat waffle loaves showed the
presence of small dark inclusions, as well as the presence of a slightly palpable taste and
aroma characteristic of algae in experimental samples of waffle loaves. When used in the The results of organoleptic evaluation of samples of rye-wheat waffle loaves showed the
presence of small dark inclusions, as well as the presence of a slightly palpable taste and
aroma characteristic of algae in experimental samples of waffle loaves. 3 Results and discussion When used in the 4 4 E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 https://doi.org/10.1051/e3sconf/202448602011 formulation of waffle loaves, a functional additive of black currant pomace powder allows
you to give the experimental samples a pinkish hue. formulation of waffle loaves, a functional additive of black currant pomace powder allows
you to give the experimental samples a pinkish hue. One of the properties of waffle loaves, indirectly characterizing their porosity and density
and causing a delicate consistency and crunch when chewing the product, is the amount of
absorbed moisture. The amount of absorbed moisture is not a regulated indicator, however,
in order to study the effect of the type of functional additives introduced on the quality of the
finished product, it was considered advisable to determine the change in the wetness of wafer
loaves, since the wetness indirectly reflects the taste characteristics of the finished product. The results of the study are presented in Figure 2. Fig. 2. Changing the amount of absorbed moisture of waffle loaves when adding functional additives. . 2. Changing the amount of absorbed moisture of waffle loaves when adding functional additives. The introduction of functional additives in the form of brown algae powder and black
currant pomace powder into the formulation of waffle loaves contributed to an increase in
the absorbed moisture of the prototypes compared to the control by 56% when using fucus
powder and by 74% when using kelp powder. The data obtained suggested an increase in
porosity and obtaining a looser texture of the prototypes with functional additives in
comparison with the control sample of waffle loaves. Along with a decrease in the density of the dough and an increase in the moisture
absorbency of the finished products, it can be concluded that the introduction of functional
additives from brown algae and powder from black currant pomace contributed to obtaining
a finished product with a more delicate consistency. Studies of the content of dietary fiber in the final product showed an increase in the fiber
content up to 7.1% in experimental samples of waffle loaves in comparison with the control
(3.17%). There was a significant increase in the mass fraction of iodine in the experimental
samples to 180.9 mcg/100g and 70.5 mcg/100g in samples with fucus and kelp powder,
respectively, compared with the control sample of waffle loaves (2.36 mcg/100g). 3 Results and discussion With a
daily intake rate of iodine from 120 to 150 mcg and dietary fiber from 20 to 25 g (for the
adult population), the data obtained indicate the functional properties of new waffle loaves,
and a new type of bakery products with reduced humidity is a source of iodine and dietary
fiber in the diet. E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 The work was carried out at the expense of the federal budget within the framework of the state task
"Development of biologically active food additives based on fruit, vegetable and medicinal plant raw
materials" (FEEF-2023-0016, registration number 1023053100014-0-2.11.1). 4 Conclusion The analysis of the effect of functional additives on the organoleptic and physico-chemical
properties of wafer rye-wheat loaves was carried out. It has been found that when brown
algae powders are added to the formulation in combination with black currant pomace
powder, the density of the dough decreases, the humidity and wetness of the finished products
increases. An increase in the acidity of the experimental samples of waffle loaves was noted
in comparison with the control sample. The type of functional additive in the amount used
does not significantly affect the content of the mass fraction of fat and sugar in the finished
product. There was a slight change in the organoleptic parameters of wafer rye-wheat loaves
when adding functional additives. A high content of iodine and dietary fiber in new types of
bakery products with reduced humidity has been established. 5 https://doi.org/10.1051/e3sconf/202448602011 E3S Web of Conferences 486, 02011 (2024)
AGRITECH-IX 2023 References 1. E.S. Smertina, L.N. Fedyanina, V.A. Lyakh, T.V. Chadova, A.G. Vershinina, Res. J. Pharm. Biol. Chem. Sci. 7(2), 989-997 (2016) 2. M.H. Kodzokova, J.M. Kunasheva, New Tech. 4, 28-33 (2018) 3. G.S. Sagdeeva, R.I. Aisina, Int. Res. J. 12(102), 173-176 (2020) 4. K. Ajisaka, T. Yokoyama, K. Matsuo, J. Appl. Glyosci. 63(2), 31-37 (2016) 5. N.N. Besednova, T.A. Kuznetsova, T.S. Zaporozhets, S.P. Kryzhanovekiy, A.K. Gaz,
E.Yu. Dobryakov, T.N. Zvyagintseva, Antibiot. Chemother. 65(1-2), 44-53 (2020) 6. A.V. Podkorytova, A. N. Roshchina, Proc. VNIRO 186(4), 156-172 (2021) 7. C. Wan-Loy, P. Siew-Moi, Mar Drugs 14(12), 222 (2016) 8. N.N. Besednova, S.P. Kryzhanovsky, T.N. Zvyagintseva, E.V. Persiyanova,
I.A. Korneeva, Antibiot. Chemother. 64(3-4), 58-69 (2019) 9. N.N. Besednova, T.A. Kuznetsova, T.S. Zaporozhets, S.P. Kryzhanovsky, L.G. Guseva,
T.N. Zvyagintseva, Antibiot. Chemother. 65(7-8), 42-51 (2020) 10. P.P. Sharma, V. Baskaran, Algal Res. 54 (2021) 11. T.P.T. Truong, T.M. Tran, T.X.T. Dai, Ch.L. Tran, J. Tradit. Complem. Med. 13(4), 408-
416 (2023) 12. S.Yu. Kalinchenko, A.S. Smykalova, L.O. Vorslov, Quest. Diet. 9(1), 25-32 (2019) 13. E.V. Semenova, A.S. Bilimenko, V.V. Chebotok, Mod. Probl. Sci. Educ. 5, 118 (2019) 14. M.A. Varzugina, R.N. Makarchuk, A.S. Yavorsky, O.A. Nikolaenko, L.K. Kuranova,
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C. Soccol, Food Chem. Adv. 2, (2023) 20. T.A. Pavlova, Cheesemaking buttermaking 1, 45-47 (2023) 21. G.V. Ivanova, E.O. Nikulina, O.Y. Kolman, A.N. Ivanova, IOP Conference Series: Earth
and Environmental Science 315 (2019) 22. L.G. Ermosh, K.A. Fadeev, N.V. Prisukhina, Polzunovsky Bull. 4, 20-26 (2021) 23. A.P. Krekhnova, O.A. Voroshilova, Natural resources, their current state, protection,
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Factors that provide protection against intimate partner physical violence among married adolescents in Bangladesh
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Carolina Digital Repository (University of North Carolina at Chapel Hill)
| 2,023
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cc-by
| 11,091
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TYPE Original Research
PUBLISHED 03 April 2023
DOI 10.3389/fpubh.2023.1125056 TYPE Original Research
PUBLISHED 03 April 2023
DOI 10.3389/fpubh.2023.1125056 CITATION Rahman M, Jamil K, Nahar Q, Chakraborty N,
Haider MM and Khan S (2023) Factors that
provide protection against intimate partner
physical violence among married adolescents
in Bangladesh. Front. Public Health 11:1125056. doi: 10.3389/fpubh.2023.1125056 Objectives: We examined factors associated with IPPV experienced by married
adolescents ages 15–19 and tested four hypotheses: (1) adolescent girls married
to relatively older husbands, (2) adolescents living in extended families with parents
or parents-in-law, (3) adolescents who are minimally controlled by husbands, and
(4) adolescents who have a child after marriage are protective of IPPV. COPYRIGHT
© 2023 Rahman, Jamil, Nahar, Chakraborty,
Haider and Khan. This is an open-access article
distributed under the terms of the Creative
Commons Attribution License (CC BY). The use,
distribution or reproduction in other forums is
permitted, provided the original author(s) and
the copyright owner(s) are credited and that
the original publication in this journal is cited, in
accordance with accepted academic practice. No use, distribution or reproduction is
permitted which does not comply with these
terms. Methods: We analyzed IPPV data from 1,846 married girls ages 15–19 obtained
from a national adolescent survey conducted in 2019–20. IPPV is defined as
the respondent having physical violence perpetrated by her husband at least
once in the last 12 months. We implemented logistic regression models to test
our hypotheses. Results: Sixteen percent of married adolescent girls experienced IPPV. Girls living
with parents-in-law or parents had adjusted odds ratio (AOR) of 0.56 (p < 0.001) of
IPPV compared to those girls who lived with husband alone. Girls with husbands
ages 21–25 years and 26 years or older had AORs of 0.45 (p < 0.001) and 0.33 (p
< 0.001) of IPPV compared to those girls with their husband ages 20 and younger. Married adolescent girls who did not own a mobile phone (an indicator of spousal
power dynamics) had an AOR of 1.39 (p < 0.05) compared to those girls who had
a phone. IPPV risk increases with an increased duration of marriage for those with
no living children (p < 0.001) but not for those with at least one living child; the
risk was higher among those who had a child within the 1st year of marriage than
those who had not yet had a child. At a duration of 4 years and longer, IPPV risk
was higher among those with no living children than those with children. intimate partner violence, intimate partner physical violence, adolescence marriage, living
arrangement, spousal control OPEN ACCESS OPEN ACCESS
EDITED BY
Omid Dadras,
University of Bergen, Norway
REVIEWED BY
Natarajan Gopalan,
Central University of Tamil Nadu, India
Kalinga Tudor Silva,
University of Peradeniya, Sri Lanka
*CORRESPONDENCE
Mizanur Rahman
rahmanm@email.unc.edu
SPECIALTY SECTION
This article was submitted to
Life-Course Epidemiology and Social
Inequalities in Health,
a section of the journal
Frontiers in Public Health
RECEIVED 15 December 2022
ACCEPTED 24 February 2023
PUBLISHED 03 April 2023 OPEN ACCESS
EDITED BY
Omid Dadras,
University of Bergen, Norway
REVIEWED BY
Natarajan Gopalan,
Central University of Tamil Nadu, India
Kalinga Tudor Silva,
University of Peradeniya, Sri Lanka
*CORRESPONDENCE
Mizanur Rahman
rahmanm@email.unc.edu
SPECIALTY SECTION
This article was submitted to
Life-Course Epidemiology and Social
Inequalities in Health,
a section of the journal
Frontiers in Public Health
RECEIVED 15 December 2022
ACCEPTED 24 February 2023
PUBLISHED 03 April 2023 Mizanur Rahman1*, Kanta Jamil2, Quamrun Nahar3,
Nitai Chakraborty1, M. Moinuddin Haider1,4 and Shusmita Khan1 1Data for Impact (D4I), Carolina Population Center, University of North Carolina at Chapel Hill, Chapel
Hill, NC, United States, 2IAP Research Inc., Dayton, OH, United States, 3Maternal and Child Health
Division (MCHD), International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka,
Bangladesh, 4Health Systems and Population Studies Division (HSPSD), International Centre for
Diarrhoeal Disease Research, Bangladesh (icddr,b), Dhaka, Bangladesh Background: Intimate partner violence (IPV), and especially intimate partner
physical violence (IPPV), perpetrated by husbands, and within adolescence
marriage are pervasive in Bangladesh. Younger women are more vulnerable
to IPPV. Introduction • A husband’s controlling attitude, his sense of superiority over
his wife, or his perceived and proclaimed legitimacy over his
wife and related behavior (21, 25–27). Four types of intimate partner violence (IPV)—physical
violence, sexual violence, stalking, and psychological aggression—
are identified by the Centers for Disease Control and Prevention
(1). IPV is one of the most common forms of violence against
women and includes physical, sexual, and emotional abuse and
controlling behaviors by an intimate partner (2). IPV is pervasive
across countries, especially in those with traditional economies
(2). Such violence has short- and long-term negative effects on
women’s health and wellbeing with varying degrees and magnitude
(1). For example, morbidity and mortality of under-five children
were significantly higher among women who experienced IPV than
those women who did not (3, 4) and major depressive disorder was
significantly higher among married adolescent IPPV victims than
those who did not face IPPV (5). • A wife’s lack of interest in sex or refusing sex at a time upon
the husband’s demand (28). • An argument between a wife and her husband. • If a wife questions her husband in day-to-day matters. • If a wife leaves the home without letting her husband or other
family member know (28–30). • If a wife does not meet her husband’s expectations in
household chores (i.e., negligence of children or meals are not
properly prepared) (7, 15). • Not conforming to wearing a veil or other expected
behavior (29). • The inability to become pregnant (28). Other family members such as in-laws (usually mother- or/and
sisters-in-law) can also instigate IPV (7). However, the role of
in-laws in IPV has likely minimized over time with increased
education and women’s greater participation in income earning
(27). In Bangladesh, married women agreed that wife-beating
is justified for the following reasons: argument with husband,
neglecting children, going outside home without permission of
husband, refusing sex when asked by husband, and burning of
food (31). Bangladesh is a country with a moderate to high level of IPV
(2, 6). Younger women are more vulnerable to IPV (2, 7–11),
and the country is among the top three with high incidence of
adolescent marriages (Statcompiler). Six in ten women are married
by age 18 and about three in ten women begin their childbearing in
their teenage period (12). CITATION Discussion: Findings related to those living with parents-in-law or parents, girls
married to relatively older boys/men, having the ability to communicate with
outside world, and having a child are protective of IPPV in Bangladesh are new,
to our knowledge. Strictly adhering to the law that requires men waiting until
the age of 21 to marry can reduce married girls’ risk of IPPV. Raising girls’ legal
marriage age can minimize adolescents’ IPPV and other health risks associated
with adolescent childbearing. intimate partner violence, intimate partner physical violence, adolescence marriage, living
arrangement, spousal control Frontiers in Public Health 01 frontiersin.org Rahman et al. Rahman et al. 10.3389/fpubh.2023.1125056 Marriage partner selection Marriage is typically arranged by parents or grandparents,
family members, and relatives (32). No or limited chance
exists for potential marital partners of knowing each other
although this norm is changing, especially in urban areas. Partner selection by the spouses, if any, usually occurs among
relatively older people, especially those with more education. Partners are now given a chance to formally meet for a few
hours immediately prior to the wedding, if desired. However,
there are few cases of adolescent marriage that take place
following love affair, initially without the knowledge of parents
and family members and eventually family accept the couples
as married. Conceptual framework In this paper, the term “intimate partner” refers to husband
and wife; living together as intimate partner is uncommon
and not typically allowed as a societal norm in Bangladesh. At the outset, we review the reported reasons behind IPV in
Bangladesh. We then review various aspects of marriage formation
processes, demography of intimate partners, living arrangement
after marriage, and expectations of the newlywed female partner
from the husband and other household members. We then develop
hypotheses on how some selected factors can lead to intimate
partner violence. A schematic diagram is shown to show the
associations between factors and IPPV (Figure 1). Most couples receive gifts from the bride’s family in cash,
in kind, or both. Negotiated gifts from bride’s family to the
groom’s family is known as a dowry and is a common
practice, especially within low-income households. The dowry
is given at the time of wedding or before, but the payment
can be deferred partially or in full for months or years after
the wedding. Introduction Child marriage or marriage before age 18
has been positively associated with IPV in Bangladesh, India, Nepal,
and Pakistan (13–17). In this paper, we examine the factors associated with IPPV
experienced by married adolescents ages 15–19. We use data
from a nationally representative survey of adolescents conducted
in 2019–20. We concentrate on factors involving household
living arrangement, partner (husband-wife) demography, partner
control, and onset of childbearing after marriage. All these factors
are likely to play important roles at the onset of their life-time
partnership development immediately after marriage. Frontiers in Public Health frontiersin.org Living arrangement after marriage The bride moves to the household of the groom after
marriage, but in rare cases the couples in Bangladesh begin
their
family
life
as
a
standalone,
nuclear
family. Some
siblings living in the groom’s home may be married and
have children. There may be grandparents there too, along
with
grandparents’
other
married
or
unmarried
children
(married children may have their children), extending the
household structure further. More than one in four families
is a three-generation family according to the recent BDHS
2017–18 (12). Partner demography Various reasons from more serious to day-to-day routine issues
are reported as reasons behind the physical violence that are
perpetrated by the husband against his wife. Lack of or inability
of dowry payment (15, 18–23) and related familial conflict (24)
are more serious and important factors. Other common causes of
IPV include: As mentioned above, teenage marriage is the cultural norm
in Bangladesh, especially in rural areas, but it is still common
in urban areas. Husbands are, on average, almost 8 years older
than their wives (12, 32). According to the 2017–18 BDHS, 32
and 46% married women ages 15–49 had husbands 10 or more Frontiers in Public Health 02 frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 FIGURE 1
Association of spousal/intimate partners’ (IP) demographic characteristics, household living arrangement, childbearing, and partner control with IPV. FIGURE 1
Association of spousal/intimate partners’ (IP) demographic characteristics, household living arrangement, childbearing, and partner control with IPV. father-in-law or her own parents. Those who live with in-laws may
also have sister- and/or brother in-laws in the house. years older and 5–9 years older, respectively. Only 20% had
husbands who were less than 5 years older and less than 1% had
husbands their same age or younger. Data shows that the age gap
between partners is gradually declining over time (Statcompiler). According to the 2019–20 BAHWS, age distribution of husbands
of married adolescents was as follows: less than 10% were 20 years
or younger, 46% were 21–25 years old, and 44% were older than
25 (12). The wife’s expected role in the household Household chores, cooking, cleaning, caring for children, and
caring for older adults (if the household has such person[s]) are all
carried out by the wife in Bangladesh. A husband rarely participates
in these activities. The expectation is that these activities will be
completed efficiently and timely. Additionally, a wife is expected
to adhere to the advice and/or directions of the in-laws if she lives
with them. Frontiers in Public Health Living arrangement A wife may acquire empowerment from her own personality
asset (e.g., negotiation) or wealth which may help lessen the
controlling strength of the husband. For example, if the wife
earns money through a job, she may also participate in
microfinance or receive remittances from parents or relatives. This empowerment of the wife, however, may be negatively or
positively associated with IPV. Women who work for money
were less likely to experience IPV in Bangladesh (9, 20). Such
earning is a source of empowerment which helps reduce the
burden of IPV (36). Hadi (38, 39) maintained that women’s
productive roles through participation in credit programs and
making financial contributions to their families not only improved
women’s positions in their households but also significantly
reduced domestic violence. In Pakistan, support from family members was negatively
associated with IPV (34). Married adolescent girls living with
her parent(s) are likely to benefit from a parent moderating or
intervening in an event that could lead to IPV. Schuler et al. (27) observed that with increased economic collaboration between
husbands and wives, the power and importance of mothers-in-
law is fast diminishing. Education is becoming more common
among the current generation of brides and as husbands become
more dependent on their wives’ economic contributions, more
egalitarian attitudes are growing. However, women’s participation in microfinance for earning
was not associated with higher level of IPV (8). Women who
meaningfully contributed to the family income through earnings
were more likely to be experiencing violence (18). Women who
were more equal with their husbands in their family relationships
by participating in decision making increased their exposure to
IPV by membership in microfinance programs (37). Women who
make decisions about household purchases were more likely to
experience physical abuse by their husbands (9, 40). Social isolation is a risk factor of IPV across societies (35). Living in nuclear households may increase social isolation as
those who live in extended households are more likely to receive
social/familial support to avoid IPV. Stoffet al. (35) found in
Bangladesh that women who maintain natal family contact at least
six times a year were more likely protected from psychological
IPV. When they contact their natal family more than ten times
a year, she is ten times more protected from the likelihood that
she will be a victim of sexual IPV. Living arrangement IPV incidence can vary within household living arrangements. The following scenarios may be considered in terms of the link
between the living arrangement and IPV: In the Bangladesh patriarchal society, husbands have a
controlling attitude toward their wives. In fact, daughters grow up
under the “guardianship” of the father and through a marriage
the guardianship is transferred to the husband. Wives are exposed
to the risk of IPV through this controlling mechanism (21). A
husband’s controlling behavior is a strong predictor for domestic
violence in India (37). 1. When an argument or other situation arises within a nuclear
family (where only the spouses live), there is no mediator who
can help to calm or pacify the episode and/or intervene. can help to calm or pacify the episode and/or intervene Partner control may be influenced by two behaviors—wife-
husband connectedness and wife having a power of some kind. Wife-husband connectedness may be a condition under which the
wife is less likely to be controlled by the husband if the partners are
connected. A wife may be classified as connected if she feels that
she enjoys spending time with husband and she talks with husband
about very personal things always or most of the time. Therefore,
connectedness may be treated as a proxy for controlling the wife by
the husband. Negotiating as a power/skill by the wife can lessen the
controlling power of the husband. 2. If a couple lives with other family member(s) and an argument
or an incidence that may lead to physical violence occurs, it
can worsen if one or more members seem to support either of
the spouses. In-laws, especially, mothers-in-law, can worsen
the situation when spouses argue and this can increase the risk
of IPV. 3. In contrast, there may be no incidence of IPV if a family
member mediates the argument between the spouses. Considering these scenarios, living in an extended family may
be protective or harmful in terms of IPV. However, as we will see
below, ideological shifts around women’s roles are occurring. These
are, over time, associated with social and economic transformations
that lower the risk of IPV. A recent study indicates that mothers-in-
law are now more tolerant about daughters-in-law, especially for
those daughters-in-law who participate in income earning (33). It
is therefore possible that mothers-in-law can help ease the situation
and minimize the chance of IPV. Partner’s age Some brides temporarily live with their parents, a trend
increasing with the international migration of males who work
in the Middle East and other countries. One in four married
adolescents ages 15–19 reported that husbands live elsewhere
(12). In the case of a migrant husband, the bride is likely to
continue to live with her parents and her husband will visit
her once every year or two. In a few cases, a groom moves
to his in-laws’ household, meaning the bride will live with
her parents. The male partner (a husband in Bangladesh) is mainly the
perpetrator of IPV, and his age is an important factor associated
with that IPV. Partners of adolescents (ages 15–19) are likely
to be young men (30). Youths or adolescents are likely to be
impulsive in their behavior and thus are more likely to be prone
to violent behavior. Relatively younger males (or youth males)
may react more aggressively to situations than their relatively older
counterparts merely because of the age. The United Nations defines
youths as those ages 15–24. In the 2019–20 BAHWS, one in three married adolescent girls
reported that she lives only with her husband, and not with any
parents-in-law or parents (30). Married adolescent girls who live
in non-nuclear households live with her husband’s mother- and/or Thus, we hypothesize that adolescent girls who are married to
youth males are more likely to encounter more violent attitudes and
behavior than those who are married to males ages 25 and older. 03 frontiersin.org 10.3389/fpubh.2023.1125056 Rahman et al. 10.3389/fpubh.2023.1125056 Frontiers in Public Health Childbearing or demonstrating the proof of
fecundity Bangladesh is an early-childbearing society, as indicated above,
three in ten women begin childbearing during adolescent ages. Parents in laws, family members, and relatives encourage the
newlywed couples to have a child early after marriage at whichever
age the marriage takes place. It is the social norm, there is even a
pressure for newlyweds in favor of early childbearing and proving
their ability of childbearing (41). Young male adults working
abroad is common in Bangladesh, and Gipson and Hindin (41)
observed in the communities with male migration observed that
having a child may be one way to cement the bond between
the husband and wife before he leaves for the country for
work, meaning the encouragement of childbearing immediately
after marriage. Many young newly married girls/women fear that they may
become childless or may not be able to bear a child with passage of
time which is bolstered by their natal and marital family members
(32). The family wants to see that the wife has given birth to a child. This has probably been derived from the perception that exists in
subsistence and agricultural economies that children are beneficial
to family earning and that children can join the workforce at an
early age. It is also expected that a son will be born and will reach
adulthood before the father reaches old age or dies so the son can
take of the family. All target adolescents in the selected households were included
in the survey. However, if there were more than one adolescent
in a household, the module that has questions on violence was
not implemented. This was done on a consideration that multiple
sample adolescents from the same household may feel embarrassed
of the reportable incident(s) that occurred in the past 12 months
with a fear that other respondent(s) would know about it. Only
3.3% of sample households had multiple adolescents. For our
analysis we excluded 200 married adolescents whose husband
resided outside home and did not visit home in past 12 months. Pressures come from in-laws to have a child immediately after
marriage in India (42). The decision to have a child after marriage
is largely influenced by in-laws and husbands—a woman’s choice
of low importance in Bangladesh (43). Living arrangement Instrumental social support
can also be negatively associated with psychological, sexual, and
physical violence (35). This is when a bride has someone outside
the home but in the village who will be willing to provide
financial or other support if needed. It is also possible that
community members can intervene when IPV occurs (36). Social
support acted as a protective buffer against spousal physical
violence (22). A relatively new idea of women’s exit options from abusive
marriages such as separation of marriage or divorce, which is
more becoming common and recognized by the community, as
indicated by Schuler and Nazneen (36). This is also a source of
women’s empowerment. Schuler et al. (33) observed that men’s
attitude of controlling their wives are changing and commented:
Men’s growing acceptance of egalitarian gender norms and their
self-reported decreased engagement in IPV are driven largely by
pragmatic self-interest: their desire to improve their economic status
and fear of negative consequences of IPV. Another vital source of empowerment of a married girl may
be her ability to communicate with people outside home seeking
advice, suggestions, or even help in crisis. Access to a mobile
phone, for example, may serve this purpose, and such access may
be realized through her own earnings or remittances from natal In this study, we hypothesize that a married adolescent girl
living with some family members, especially parent(s)-in-law or
parent(s), experiences a different risk of IPV than one living alone
with her husband. Frontiers in Public Health 04 frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 family or relatives or from her husband. This is also a source of
empowerment which may help protect a wife from IPV perpetrated
by her husband. may support family members’ view of early childbearing and
demand the wife to have a child, and if there is disagreement the
husband may become violent. In India, IPV was associated with the
intention of delaying the first birth (45). We hypothesize that spousal control is associated with IPV, i.e., a
husband who fully controls his wife is more likely to practice IPV than
a husband who does not control or minimally controls. The effect of
control is minimized when (a) husband and wife are connected, i.e.,
spouses have a sense of togetherness, and (b) wife is empowered with
her ability to communicate with people outside home owing to some
resources owned by her. Data Data for this study come from the nationally representative
Bangladesh Adolescent Health and Wellbeing Survey 2019–20
(30) [The survey details are given in NIPORT (30), including the
questionnaires.]. The age limit for adolescents in this survey was
15–19 years. The survey was based on a two-stage stratified sample
of households, which involved sampling of primary sampling units
(PSU), and sampling of households. At the first stage, PSUs were
randomly selected from each stratum according to probability
proportional to size of the number of households. PSUs were
randomly and equally divided into Type 1 and Type 2 PSUs,
and information on physical violence was collected from Type 2
PSUs. At the second stage, 67,093 households were selected from
where 7,800 unmarried female adolescents, 4,926 ever-married
female adolescents (2,904 in Type 1 and 2,022 in Type 2), and
5,523 unmarried male adolescents were selected for interview. The
response rate for married adolescent sample was 97.2%. Data were
collected on paper and pencil through face-to-face interviews at the
home of the respondents by using a structured questionnaire. Our
sample come from Type 2 PSUs and thus Type 2 questionnaire, and
the respondents were 2,022 married adolescents. Frontiers in Public Health Childbearing or demonstrating the proof of
fecundity Delaying the first birth after
marriage can cause rumors of infertility, bring shame upon the
family, and in some cases lead the husband’s family to seek another
wife for their son (44). Additionally, social stigma for childless
women, emigration of husbands, and the belief that using modern
contraceptives prior to the birth of the first child results in infertility
also inhibits couples from delaying their first pregnancy. It should
be mentioned that there has been a rapid demographic transition
in Bangladesh wherein fertility declined from over 6.0 births per
woman to 2.3 births and life expectancy has declined from about 50
years to over 70 years in the last 40 years, the perceived benefit of
early childbearing seems to remain in the older generation. Our analysis considers appropriate sampling weights calculated
based on the complex sampling design of the BAHWS 2019–20,
and our sample consisted of 1,822 (weighted n = 1,846) married
adolescent girls ages 15–19. Study variables Table 1 shows the variables and measurements of factors and
outcome used in this analysis. Living arrangement Therefore, we hypothesize that delaying of childbearing is
associated with IPV among adolescents in Bangladesh. Intimate partner physical violence The young newly married women, e.g., adolescents ages 15–19
want to delay their childbearing and the family members encourage
or sometimes pressurize newly married ones to have a child earliest
possible time. While the women may pursue the delay, her spouse The 2019–20 BAHWS had five questions on the nature of
physical violence experienced by married adolescents, see the first
row in Table 1. If such violence was perpetrated by her husband at 05 frontiersin.org frontiersin.org 10.3389/fpubh.2023.1125056 Rahman et al. TABLE 1 Indicators and measurement of factors and outcome. least once in the past 12 months of the survey is treated as IPPV. Answering Yes to at least one of the questions is defined as an
outcome of IPPV and coded as 1, and otherwise as 0. d ca o s a d
easu e
e
o
ac o s a d ou co
e
Factor/outcome
Indicator
Intimate partner physical
violence (IPPV)
Experience
of
physical
violence
perpetrated by the husband or husband
along with other family member
• Those who reported having experienced
physical violence at least once in the 12
months preceding the survey, i.e.:
• Slapped, pushed, or pulled hair
• Punched, thrown something at her,
or hit with a stick or something heavy
• Kicked, dragged, or beat up
• Tried to choke her or burn her on
purpose with something hot (fire,
object, acid)
• Threatened or attacked her with a knife,
gun, or any other weapon
Socioeconomic
• Years of schooling (≤5, 6–9, 10+)
• Household asset quintile (Bottom 40%,
Middle 20%, and Upper 40%)∗
• Residence (Urban, Rural)
• Region of the country (Western
[Rangpur, Rajshahi, and Khulna
Divisions], Central [Mymensingh,
Dhaka, and Barishal Divisions], Eastern
[Sylhet and Chattogram Divisions]). Attitude toward gender roles
• Egalitarian if disagree to each of the four
statements: (a) It is important that sons
have more education than daughters; (b)
outdoor sports are only for boys not
for girls; (c) household chores are for
women only, not for men, even if the
woman works outside the house; and (d)
women should not be allowed to work
outside of home. • Inegalitarian, otherwise. Spousal control We use two variables to capture the effect of spousal control:
respondent’s connectedness with her husband and respondent
having her own mobile phone. On the married girl’s connectedness
with her husband, we assume that a married girl who has a feeling
of connectedness with her husband, she is less likely to be fully
controlled by her husband. A wife possessing a phone and using it is
likely due to indicate two characteristics: (1) she can communicate
with people outside home and (2) she has power, economic or
otherwise, acquired either by her own earning, through inheritance
of resources from relatives, or through negotiation with the
husband. Such characteristics can soften or minimize a husband’s
controlling attitude and/or actual control over his wife. Living arrangements We consider three categories of living arrangements: (a) with
husband (and children, if any), (b) with husband and parents-in-
law or parents and children if any, and (c) husband lives elsewhere
but wife lives with parents-in-law or parents and children, if any. In
group (c), there may be person(s) of other type of relationship(s). The category (a) here is treated as the reference category in the
logistics regression. Spousal demographic characteristics Husband’s age is categorized as ≤20, 21–25, and 26+ and
≤20 category is treated as the reference category in the logistic
regression. Duration of marriage is divided in to three groups: ≤1
year, 2–3 years, and 4+ years and the 1st year of marriage is treated
as reference category. Intimate partner physical violence Attitude toward gender
responsibilities and
spousal/IP power dynamics
• Egalitarian if disagree to each of the
three statements: (e) Looking after the
household and kids is the responsibility
of women only; (f) a woman should
always listen to her husband even if she
disagrees; and (g) a husband has the
right to physically assault or beat his wife
if she does not listen to him. • Inegalitarian, otherwise. Spousal/IP demographic
characteristics
• Husband’s age (≤20, 21–25, and 26+)
• Duration of marriage (0–1 year, 2–3
years, and 4+ years)
Household living
arrangement
Living arrangement
• With husband and children (if any)
• With husband, parents in laws or
parents, and children (if any)
• Husband lives elsewhere but she lives
with (a) parents in laws or (b) parents,
or (c) lives alone. (There may be
child[ren] if there are any). In group
(c), there may be person(s) of other type
of relationship(s). Spousal/IP connectedness
Connectedness with husband
• Connected (wife enjoys spending time
with husband and talks with husband
about very personal things most of the
time or always)
• Weakly connected, otherwise
Wife’s ability of
communicating with people
outside home
• Having her own mobile phone (yes, no)
Childbearing
• Having living child(ren) (0, 1+)
∗Household asset quintile is constructed by the household ownership of a set of assets as
reported by the respondent or observed by the interviewer at the survey. Frontiers in Public Health Childbearing The childbearing indicator is dichotomized as having at least
one living child at the time of survey (coded as 1) and no living
child is coded as zero. We
also
include
the
following
variables
(see
Table 1
for definitions): • Years of schooling in three categories—≤5 years (reference
category), 6–9 years, and 10+ years. • Attitude
toward
gender
roles—Inegalitarian
(reference
category) and egalitarian. • Attitude
toward
gender
responsibilities
and
power
dynamics in the family—Inegalitarian (reference category)
and egalitarian. • Household asset quintiles—Bottom 40% (reference category),
middle 20%, and upper 40%. • Region of the country—Western (reference category), Central,
and Eastern. ∗Household asset quintile is constructed by the household ownership of a set of assets as
reported by the respondent or observed by the interviewer at the survey. • Residence—Urban (reference category) and Rural. frontiersin.org Frontiers in Public Health 06 frontiersin.org Rahman et al. Rahman et al. 10.3389/fpubh.2023.1125056 Prevalence and pattern of IPPV: Bivariate
findings It is possible that IPPV is influenced by husband’s education,
husband’s smoking or drinking habits, or respondent’s parental
education but the BAHWS 2019–20 did not collect information
on
those,
neither
did
we
have
adolescents’
information
on religion. Table 2 (Column 4) shows the prevalence of IPPV according to
their characteristics. Sixteen percent of married girls reported that
they experienced IPPV at least once in the past 12 months. IPPV
was similar for rural and urban areas; it was slightly lower, not
significantly, in the Eastern region than other regions; about 20%
girls in the bottom 40% households experienced IPPV compared to
12% in the upper 40% households, the difference is significant (p
< 0.001). IPPV substantially decreases with education, from 22%
among girls with no or below 6 years of schooling to 17% among
girls with 6–9 years of schooling to only 7% among those with 10 or
more years of schooling (p < 0.001). Sample characteristics Table 2 shows the distribution of 1,846 married adolescent girls
according to categories of factors considered in the analysis. About
one in four is an urban resident; about two in five are each from
Western and Central regions; and about two in five from the upper
40% and about one in three from the bottom 40%. About one in
four had no or less than 6 years of schooling, about half had 6
to 9 years of schooling, and about one in four had ten or more
years of schooling. About two in five married girls had egalitarian
attitudes toward gender roles (see Table 1) and about one in three
had egalitarian attitudes toward gender responsibilities and spousal
power dynamics. IPPV risk is moderately associated with girls’ “connectedness”
with their husbands (p < 0.05)−14 vs. 18% for the connected and
weakly connected groups, respectively. Not having a mobile phone
exposes girls to IPPV as it was 19% among those who did not
have a phone compared to 13% among those who had a phone
(p < 0.001). IPPV rate was 21% among married girls without a living child
compared to only 12% among those who had at least one living
child (P < 0.001). About one in four married girls lived alone with husband
(including own children if any), about two in three lived in
extended family with husband and at least one parent or parent-
in-law, and about one in ten lived without husband but with at least
one parent or parent-in-law. Results IPPV risk significantly decreases from 26% among those whose
husband was 20 years old or younger to 17% among those whose
husband were 21–25 years old to only 12% among those with
husband of 26 years or older (p < 0.001). The risk increases with the
duration of marriage (11% among those with 0–1 year of duration
vs. about 20% among those with higher duration [p < 0.001]). Statistical analysis We estimate the effects of various variables on IPPV in
logistic regression model. The independent variables included
in the regression model to represent the factors indicated
in
the
conceptual
framework. We
did
not
use
stepwise
procedure to include or exclude the variables. We checked
for multicollinearity between independent variables and none
of the tests required to exclude any independent variables. For example, the mean Variance Inflation Factor (VIF) was
1.3 (and maximum limit was 1.76) which is much less than
10, the acceptable limit. We used STATA version 17. There
were only seven records (0.38%) with missing values in two
variables, and the STATA exclude those records that has a
missing value. Girls with inegalitarian attitude toward gender roles had an
IPPV rate of 18% compared to 13% among egalitarian (p < 0.05). IPPV differential in terms of attitude toward gender responsibilities
and spousal power dynamics was similar between the inegalitarian
and egalitarian groups (17 vs. 14%, not significant). IPPV is strongly associated with living arrangement (p <
0.001); 25% girls living with husband alone (i.e., nuclear family)
experienced IPPV compared to 14% among those who lived with
husband and parents-in-law, or parents. Those girls who lived
without her husband (i.e., her husband lived elsewhere but visited
home sometimes) but with in-laws or parents have significantly
lower IPPV rate of 9% than other groups. IPPV risks: Multivariate findings We
modeled
IPPV
risk
associated
with
factors
under
consideration by using logistic regression (Table 3). We considered
contextual
and
socioeconomic
factors
such
as
rural-urban
residence,
geographical
region,
household
asset
quintiles,
girls’ education, and two variables capturing girls’ attitudes
toward gender roles. The model includes spousal demographic
characteristics (husband age and marital duration), spousal
connectedness and the girl’s ownership of a mobile phone,
household living arrangement, and a childbearing (measured
as having at least one living child) indicator. We included the
interaction term “living children x marriage duration.” Adjusted
odds ratios (AOR) are shown in Table 3. While the girls were ages 15–19, 9, 46, and 44% of their
husband were 20 or below, 21–25, and 26 and higher years of age,
respectively (Table 2). Nearly half (48%) were married for less than
2 years, one third (33%) for 2–3 years, and about one in five (18%)
were married for 4 or more years. Slightly less than half of married girls were “connected” with
their husband, measured by the indicator “enjoy spending time
with husband and can talk with husband about very personal things
most of the time or always” (Tables 1, 2). Also, half of the girls had
their own mobile phone. Regarding childbearing, 44% of married girls had at least one
surviving child (Table 2). 07 frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 of sample of currently married adolescents ages 15–19 who are exposed to IPPV, Bangladesh Adolescents Health and Wellbeing LE 2 Description of sample of currently married adolescents ages 15–19 who are exposed to IPPV, Bangladesh Adolescents Health
ey 2019–20. Survey 2019 20. IPPV risks: Multivariate findings Sample size
(n)
% Sample
% Experienced IPPV at
least once in last 12
months
Chi-square
P value
Total
1,846
100.0
16.0
–
Residence
0.65
0.45
Urban
440
24.9
14.7
Rural
1,405
76.1
16.4
Region
4.04
0.18
Western
777
42.1
16.8
Central
726
39.3
16.7
Eastern
343
18.6
12.3
Household asset quintile
16.92
0.0007∗∗∗
Bottom 40%
803
43.5
19.9
Middle 20%
408
22.1
14.1
Upper 40%
635
34.4
12.1
Years of schooling
37.86
0.0000∗∗∗
0–5 years
428
23.1
22.4
6–9 years
974
52.8
17.0
10+ years
444
24.1
7.4
Attitude toward
gender roles
7.69
0.017∗
Inegalitarian
1,135
61.6
17.9
Egalitarian
706
38.4
13.0
Attitude toward gender
responsibilities and spousal
power dynamics
1.38
0.286
Inegalitarian
1,256
68.2
16.6
Egalitarian
586
31.8
14.4
Household living arrangement
35.72
0.000∗∗∗
With husband (and children, if any)
460
24.9
24.5
With husband, parents-in-law, parents
(and children, if any)
1,176
63.7
13.8
Husband lives elsewhere but she lives
with parents in laws, parents, or lives
alone (and children, if any)
209
11.3
9.0
Husband’s age
22.04
0.000∗∗∗
≤20 years
172
9.3
26.1
21–25 years
856
46.4
17.3
26+ years
817
44.2
12.4
Marriage duration
30.88
0.000∗∗∗
0–1 year
892
48.3
11.1
2–3 years
617
33.4
21.1
4+ years
337
18.3
19.5
(Continued) 08 Frontiers in Public Health frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 TABLE 2 (Continued) Sample size
(n)
% Sample
% Experienced IPPV at
least once in last 12
months
Chi-square
P value
Connected with husband
6.15
0.012∗
Connected
844
45.7
13.7
Weakly connected
1,001
54.2
17.9
Owning a phone
12.06
0.000∗∗∗
Having a mobile phone
916
49.6
12.9
Having no mobile phone
930
50.4
18.9
Childbearing
25.02
0.000∗∗∗
Having no living children
1,039
56.3
12.2
Having at least 1 child
805
43.6
20.8
∗p < 0.05; ∗∗∗p < 0.001. counterpart without a child. Having a living child is associated
with marriage duration which is associated with IPPV as shown
in bivariate findings (Table 2). We include an interaction between
marriage duration and the childbearing indicator in Table 3. Efects of other factors Rural-urban residence and region were not significantly
associated with IPPV in the bivariate analysis (Table 2) nor in the
multivariate regression (Table 3). Household asset quintiles and
girls’ years of schooling were significantly associated with IPPV
in bivariate analysis but only years of schooling is significantly
associated with IPPV in the multivariate regression (p < 0.01). IPPV risk is significantly lower among girls who have 10 or more
years of schooling than others. Among the two factors influencing
attitudes toward gender roles, none were significantly associated
with IPPV according to the regression models. Spousal control Spousal connectedness is not associated with IPPV as indicated
by the non-significant AOR related to the category “weakly
connected,” although it was significant in the bivariate association. Owning a mobile phone is significantly associated with the risk
of experiencing IPPV as AOR of not owning one is 1.39 and is
statistically significant (p < 0.05). Living arrangements • IPPV risk for girls with a child is about similar for the years of
marriage duration. Table 3 shows that adolescent girls who lived with their parents-
in-law or parents have an AOR of 0.56 (p < 0.01), meaning that the
odds of their IPPV are 44% lower than those girls who live with her
husband alone or within a nuclear family. Thus, living with in-laws
or parents provide a protective effect on IPPV, and a support for
our hypothesis. Girls who live with in-laws or parents, but without
their husbands, have even lower odds of IPPV, i.e., 56% ([1.00–0.44]
× 100) lower than girls who live with alone with their husbands or
in nuclear families. These lower odds reflect the effect of infrequent
exposure to IPPV as the husband lived elsewhere and visited the
wife and family occasionally. We found that the longer girls wait to have a child the higher
the risk of IPPV, and thus, our hypothesis is supported. Partner’s age AOR of IPPV is 0.45 and 0.33, respectively, for girls whose
husband were ages 21–25 and 26 or more years old compared to
girls with husband younger than 21 (p < 0.001) (Table 3). These
findings demonstrate that girls with husbands ages 21–25 and 26
or higher have 55% ([1.00–0.45] × 100) and 67% ([1.00–0.33] ×
100) lower odds of IPV than their counterpart girls with husbands
younger than 21. Thus, we find that adolescent girls marrying
relatively older husband provide a IPPV protective effect, and our
hypothesis is supported. Significant interaction occurs between having a child and the
duration of marriage (Table 3). Figure 2 shows three distinct effects: • IPPV risk for girls who did not a have living child increases
from 11% during year 0–1 to 17% during years 2–3 to 29%
after 4 or more years of marriage. • IPPV risk is higher among those who have a child immediately
after marriage (0–1 year) than those who have not yet have a
child (18 vs. 11%). Frontiers in Public Health frontiersin.org Childbearing We analyzed intimate partner physical violence data from
1,846 married adolescent girls ages 15–19 based on the nationally
representative adolescents survey conducted in 2019–20. We tested Our measure of childbearing is whether a girl has at least one
living child (Table 2). Our hypothesis is that a girl who survives
having a child is less likely to experience IPPV risk than her 09 frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 TABLE 3 Binary logistic regression-based unadjusted odds ratios (OR) and adjusted odds ratios (AOR) of IPPV among currently married adolescents ages
15–19, Bangladesh Adolescents Health and Wellbeing Survey 2019–20. TABLE 3 Binary logistic regression-based unadjusted odds ratios (OR) and adjusted odds ratios (AOR) of IPPV among currently married adolescents ages
15–19, Bangladesh Adolescents Health and Wellbeing Survey 2019–20. TABLE 3 Binary logistic regression-based unadjusted odds ratios (OR) and adjusted odds ratios (AOR) of IPPV among currently ma
15–19, Bangladesh Adolescents Health and Wellbeing Survey 2019–20. TABLE 3 Binary logistic regression-based unadjusted odds ratios (OR) and adjusted odds ratios (AOR) of IPPV among currently married adolescents ages
15–19, Bangladesh Adolescents Health and Wellbeing Survey 2019–20. TABLE 3 Binary logistic regression based unadjusted odds ratios (OR) and adjusted odds ratios (AOR) of IPPV among currently married adolescents ages
15–19, Bangladesh Adolescents Health and Wellbeing Survey 2019–20. Childbearing Some young girls work in the non-agricultural sector (e.g.,
garment factories) and their husbands may accompany her to live
in city areas. The proportion of this nuclear living arrangement is
probably increasing because of the growing economic activities of
the non-agricultural sector, and thus, likely increases IPPV risk for
young married women. four hypotheses—adolescent girls married to husbands ages 25
and older, adolescents living in extended families with parents-
in-law or parents, adolescents who are minimally controlled by
their husbands, and adolescents who have a child immediately after
marriage are protective of IPPV in Bangladesh. We find support
in favor of each of the hypotheses. The findings are new, to our
knowledge, except for that on the association between spousal
control and IPPV. The finding that girls married to husbands beyond the age of
youthhood, i.e., 25 years or older, are protective of IPPV is like that
of Adebowale (46) in Nigeria which indicates that women married
to relatively older husband (measured by age difference between
husband and wife) have lower likelihood of IPV in general and
IPPV and emotional violence, but not true for sexual violence. Our
finding implies that strictly adhering to the Bangladesh law that
requires men marrying at 21 or later can reduce married girls’ risk
of IPPV. Increase in males’ ages at marriage will benefit adolescent
girls from the risk of IPV as well as through their reduced likelihood
of adolescence marriage, and men should be encouraged to marry
girls who are 20 or older or at least 18 years old girls as required
by law. A husband’s control of his wife is a source of IPPV, but this
control is moderated or lessened when his wife has some power. This can mean having a phone that enables her to communicate
with the outside world. Owning a phone may also be related to her
economic power, helping reduce the burden of IPPV. We argue
that ownership of a mobile phone is possible through a wife’s
income earnings or through remittance obtained from relatives
or even from her husband. However, the literature suggests, as
we show in our conceptual framework, that income earning can
have both negative and positive effect on IPV in Bangladesh
(18, 36, 38–40). The negative association of having a phone and
IPPV is likely to work through social isolation and IPV hypothesis
as well as instrumental social support and IPV hypothesis (35). Childbearing Bivariate
Multivariate
Factors
OR
95% confidence
interval
AOR
95% confidence
interval
Residence
Urban
1.00
1.00
Rural
1.13
(0.80, 1.61)
1.04
(0.72, 1.50)
Region
Western
1.00
1.00
Central
0.99
(0.75, 1.32)
1.11
(0.82, 1.51)
Eastern
0.70
(0.44, 1.10)
0.83
(0.53, 1.30)
Household asset quintile
Bottom 40%
1.00
1.00
Middle 20%
0.66∗
(0.47, 0.94)
0.81
(0.55, 1.18)
Upper 40%
0.56∗∗∗
(0.41, 0.77)
0.82
(0.57, 1.19)
Years of schooling
0–5 years
1.00
1.00
6–9 years
0.71∗
(0.52, 0.97)
0.86
(0.62., 1.20)
10+ years
0.28∗∗∗
(0.18, 0.43)
0.47∗∗
(0.29, 0.77)
Attitude toward gender roles
Inegalitarian
1.00
1.00
Egalitarian
0.68∗∗
(0.51, 0.91)
0.83
(0.59, 1.18)
Attitude toward gender
responsibilities and spousal
power dynamics
Inegalitarian
1.00
1.00
Egalitarian
0.85
(0.63, 1.14)
1.27
(0.88, 1.83)
Husband’s age
≤20 years
1.00
1.00
21–25 years
0.59∗
(0.39, 0.90)
0.45∗∗∗
(0.29, 0.70)
26+ years
0.40∗∗∗
(0.26, 0.62)
0.33∗∗∗
(0.20, 0.52)
Marriage duration
0–1 year
1.00
1.00
2–3 years
2.15∗∗∗
(1.58, 2.92)
1.75∗
(1.10, 2.80)
4+ years
1.95∗∗∗
(1.35, 2.81)
3.72∗∗
(1.65, 8.37)
Connectedness with husband
Connected
1.00
1.00
Weakly connected
1.38∗
(1.05, 1.82)
1.30
(0.98, 1.72)
Owning a mobile phone
Having a mobile phone
1.00
1.00
Having no mobile phone
1.57∗∗
(1.19, 2.06)
1.39∗
(1.03, 1.89)
Household living arrangement
With husband (and children, if any)
1.00
1.00
(Continued) 10 10 Frontiers in Public Health frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 TABLE 3 (Continued) Bivariate
Multivariate
Factors
OR
95% confidence
interval
AOR
95% confidence
interval
With husband, parents-in-law, parents (and children,
if any)
0.50∗∗∗
(0.37, 0.67)
0.56∗∗
(0.42, 0.78)
Husband lives elsewhere but she lives with parents in
laws, parents, or lives alone (and children, if any)
0.31∗∗∗
(0.18, 0.53)
0.44∗∗
(0.25, 0.79)
Childbearing
Having no living children
1.00
1.00
Having at least 1 child
1.90∗∗∗
(1.45, 2.49)
1.90∗
(1.07, 3.39)
Living children × Marriage
duration
Having a child × Marriage duration (0–1 year)
1.00
Having a child × Marriage duration (2–3 years)
0.77
(0.37, 1.60)
Having a child × Marriage duration (4+ years)
0.26∗∗
(0.09, 0.70)
∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. or parent(s), and 11% with parents-in-law or parent(s) but the
husband lives elsewhere (usually for work). One reason young
couples live in a nuclear family is when one of the spouses’ (mostly
the husband’s) workplace is distant from the husband’s natal
home. Frontiers in Public Health Childbearing In many countries
it is required that young people complete 12 years of schooling
(i.e., high school graduation) and by that time they are usually
18 years old or over. Providing girls with high school or higher
education also helps nation building and thus facilitating social and
economic development. Girls from families that cannot afford this
may be given scholarships. Cost-benefit analysis will likely show
this is a noble option, and future research should be conducted in
this area. marriage, we reiterate that an increase in age at marriage will
minimize or even eliminate the chance of adolescent childbearing
and thus reduce maternal and child health risks and the risk of
IPV. However, there is only a slow increase in age at marriage—
it has increased from 15.3 years in 1993 to 17.5 years in 2017,
roughly a year per decade. A germane question is how can we
accelerate the pace of increasing age at marriage? An obvious
solution to this is keeping girls in school for 12 years of education. Although there has been a praiseworthy improvement in education
in Bangladesh, only 34%, or one in three, married women aged 20–
24 had 10 or more years of schooling in 2017–18. In many countries
it is required that young people complete 12 years of schooling
(i.e., high school graduation) and by that time they are usually
18 years old or over. Providing girls with high school or higher
education also helps nation building and thus facilitating social and
economic development. Girls from families that cannot afford this
may be given scholarships. Cost-benefit analysis will likely show
this is a noble option, and future research should be conducted in
this area. FIGURE 2
Predicted probability of IPPV based on model interaction between
marriage duration and having a child. FIGURE 2
Predicted probability of IPPV based on model interaction between
marriage duration and having a child. We did not find any significant association between spousal
connectedness and IPPV. Our argument was that connectedness
between spouses would reduce the feeling of husband’s control
over his wife and thus lower IPPV. One reason why we did not
find significant association between connectedness and IPPV may
be that the indicator we used is not an appropriate measure of
connectedness. Childbearing It is also possible that IPPV itself has a negative
effect on connectedness in that a wife experiencing IPPV does not
feel close to her husband. Without closeness, a wife may not share
personal ideas with her husband, and thusly there is no significant
association between connectedness and IPPV. An important limitation of this study is that we analyzed cross-
sectional IPPV data of quantitative nature from an adolescent
health and wellbeing survey which did not have more in-depth
information on issues related to IPPV. We only identified factors
affecting IPPV but do not know the exact mechanisms of how
these factors caused IPPV. We exercise caution that we measure
association but not cause-and-effect relationships between various
factors and IPPV. Further research particularly of qualitative nature
can help better understand such mechanisms of IPPV among
married adolescents. There is dearth of qualitative studies of IPV
in Bangladesh, e.g., 6 in his review mentioned that there were 17
quantitative studies, 11 jointly quantitative and qualitative, and
only 3 qualitative studies. Childbearing is strongly associated with IPPV, specifically in
that IPPV continues to increase with marriage duration if the
wife does not have a living child. In contrast, having a living
child puts a married girl at a moderate risk of IPPV in the early
years of marriage but it does not increase with marriage duration. This finding is consistent with the common belief that the family,
including the in-laws, and the society in general expect that a child
should be born immediately after marriage as observed by previous
researchers (41, 42). If that does not happen the married girl is at
the risk for IPPV. To conclude, married adolescent girls living with parents-in-
law or parents, girls married to relatively older boys/men, having
a power of communicating with outside world, and having a child
immediately after marriage are protective of IPPV in Bangladesh. Strictly adhering to the law that requires men to marry at 21 or older
can reduce married girls’ risk of IPPV. Raising girls’ age at marriage
can minimize adolescents’ IPPV and other health risks associated
with adolescent childbearing. This
finding
is
disturbing
because
childbearing
during
adolescence is harmful for both maternal and infant/child health. Healthcare providers recommend avoiding having a birth before
age 20. Adolescent girls are in a quandary and those who desire to
follow the health advice intend to use contraception. Data availability statement Publicly available datasets were analyzed in this study. This
data
can
be
found
here:
https://dataverse.unc.edu/
dataset.xhtml;jsessionid=2041ade5046836520c1117daa52b? persistentId=doi%3A10.15139%2FS3%2FDVEI9A&version=&
q=&fileTypeGroupFacet=%22Tabular+Data%22&fileAccess=&
fileTag=&fileSortField=&fileSortOrder=. We are in a dilemma; should we recommend delaying
childbearing after age 20 as recommended by public health
practitioners? If we do, the adherent girls will be at increased risk
of IPPV. Also, a significant portion of married adolescent girls who
had a birth reported that they wanted to have the birth later, as
shown in our companion study (5). Adolescent girls who wanted to
have the birth later had 1.30 times higher risk of major depressive
disorder than those who wanted the birth earlier. Childbearing But pressures
come from in-laws and family against contraceptive use at this time
of her life, and, in some cases the husband joins the family against
contraceptive use. This situation may lead to violence against
the girl. Frontiers in Public Health Childbearing The social isolation hypothesis asserts that those women who
communicate with natal or other kins or relatives more frequently
are less likely to experience IPV and the instrumental social
support hypothesis claims that those women who have someone
in the same or neighboring communities willing to provide
support are less likely to face IPV. In Bangladesh, instrumental
support is available even at the local government level through
the newly enacted Domestic Violence (Prevention and Protection)
Act 2020 (7). Phone calls can be a vehicle of communication
to satisfy these hypotheses and thus likely reduce IPPV. IPPV
is likely to decline over time with economic growth, which is
occurring rapidly in Bangladesh, that benefits women in owning
mobile phone. We find that married adolescents living with parents-in-law
or parent(s) helps reduce IPPV. Various studies and anecdotes
indicate that living with in-laws, especially mother-in-law, is a
risk of IPPV, but Schuler et al. (33) observed that there has
been ideological change among in-laws wherein they believe
that daughters-in-law need to be treated with dignity and
respect. This is because of social and economic development and
their daughter-in-law’s increased economic role in families. Our
finding is consistent with the notion of changed attitudes toward
compromised behavior of husbands in terms of IPPV. As
Bangladesh
is
in
transition
from
a
traditional
to
contemporary economy, it is still expected that newlywed
couples will begin their lives in extended family settings and then
move toward nuclear ones. Our findings show that 25% married
adolescents live in nuclear families, 64% with parents-in-law 11 frontiersin.org Rahman et al. 10.3389/fpubh.2023.1125056 10.3389/fpubh.2023.1125056 marriage, we reiterate that an increase in age at marriage will
minimize or even eliminate the chance of adolescent childbearing
and thus reduce maternal and child health risks and the risk of
IPV. However, there is only a slow increase in age at marriage—
it has increased from 15.3 years in 1993 to 17.5 years in 2017,
roughly a year per decade. A germane question is how can we
accelerate the pace of increasing age at marriage? An obvious
solution to this is keeping girls in school for 12 years of education. Although there has been a praiseworthy improvement in education
in Bangladesh, only 34%, or one in three, married women aged 20–
24 had 10 or more years of schooling in 2017–18. Conflict of interest The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be
construed as a potential conflict of interest. References and
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Gynecol Obstetr. (2010) 110:35–9. doi: 10.1016/j.ijgo.2010.01.022 7. Khan AR. Ethics statement The studies involving human participants were reviewed and
approved by icddr,b ERC. Written informed consent to participate
in this study was provided by the participants’ legal guardian/next
of kin. We will still recommend waiting to have children after the
age of 20. Following our recommendation of raising the age at 12 frontiersin.org Rahman et al. Rahman et al. 10.3389/fpubh.2023.1125056 Funding This publication was produced with the support of the United
States Agency for International Development (USAID) under
the terms of USAID’s Research for Decision Makers (RDM)
Activity cooperative agreement no. AID-388-A-17-00006 and of
Data for Impact (D4I) associate award no. 7200AA18LA00008. Data source for this publication is the Bangladesh Health and
Wellbeing Survey 2019-20 (BAHWS 2019-20) funded by the
US Agency for International Development (USAID) through the
MEASURE Evaluation Project, Data for Impact (D4I) Project and
Research for Decision-Makers (RDM) Activity (GR: 154601546). Additional funding was provided by the Foreign, Commonwealth
and Development Office (FCDO) and by the Government of
Bangladesh. The BAHWS 2019-20 was implemented under the
authority of the National Institute of Population Research and
Training (NIPORT) of the Government of the People’s Republic
of Bangladesh. The BAHWS 2019-20 data were collected and
processed by Human Development Research Center (HDRC). Acknowledgments MR and KJ conceptualized the study and designed the analysis. NC and MMH performed the statistical analysis. MR drafted
the manuscript. SK provided support in literature review and in
writing part of the discussion. KJ, MMH, QN, and SK reviewed the
manuscript. All authors reviewed, contributed to finalization of the
manuscript, and approved the final version of the article. The authors are grateful to Anadil Alam from icddr,b who
facilitated the research team’s activities and Kaiser Mahmud
who helped in the initial data analysis. Nurul Alam from
icddr,b and Md. Mahabubur Rahman from icddr, b/D4I provided
helpful comments. The authors would also like to thank Sian
Curtis and Gabriela Escudero from D4I for their careful review
of the publication and Katie Kosma from the Knowledge
Management (KM) team of D4I for providing editing support of
the manuscript. Publisher’s note All
claims
expressed
in
this
article
are
solely
those
of the authors and do not necessarily represent those of
their
affiliated
organizations,
or
those
of
the
publisher,
the editors and the reviewers. Any product that may be
evaluated in this article, or claim that may be made by
its
manufacturer,
is
not
guaranteed
or
endorsed
by
the
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EHR Foundation Models Improve Robustness in the Presence of Temporal Distribution Shift
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TITLE: EHR Foundation Models Improve Robustness in the Presence of Temporal Distribution
Shift TITLE: EHR Foundation Models Improve Robustness in the Presence of Temporal Distribution
Shift reprint reports new research that has not been certified by peer review and should not be used to guide clinical practice .
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medRxiv preprint TITLE: EHR Foundation Models Improve Robustness in the Presence of Temporal Distribution
Shift
AUTHORS: 1Lin Lawrence Guo PhD, 2Ethan Steinberg, 2Scott Lanyon Fleming, 3Jose Posada
1Joshua Lemmon, 2Stephen R Pfohl PhD, 2Nigam Shah MD, PHD, *2Jason Fries PhD, *1,4Lillia
Sung MD, PhD
*co-senior authors
AFFILIATIONS:
1Program in Child Health Evaluative Sciences, The Hospital for Sick Children, Toronto, ON,
Canada
2Stanford Center for Biomedical Informatics Research, Stanford University, Palo Alto, CA
3Universidad del Norte, Colombia
5Division of Haematology/Oncology, The Hospital for Sick Children, Toronto, ON
ADDRESS FOR CORRESPONDANCE:
Lillian Sung MD, PhD
Division of Haematology/Oncology
The Hospital for Sick Children,
555 University Avenue, Toronto, Ontario, M5G1X8, Canada
Telephone: 416-813-5287
Fax: 416-813-5979
Email: Lillian.sung@sickkids.ca
KEY WORDS: electronic health records, foundation model, model robustness, machine
learning, self-supervised learning
WORD COUNT: Abstract 250; Text 3,702; Tables 0; Figures 6; AFFILIATIONS: WORD COUNT: Abstract 250; Text 3,702; Tables 0; Figures 6; . CC-BY 4.0 International license
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medRxiv preprint ABSTRACT Background: Temporal distribution shift negatively impacts the performance of clinical Background: Temporal distribution shift negatively impacts the performance of clinical
prediction models over time. Pretraining foundation models using self-supervised learning on
electronic health records (EHR) may be effective in acquiring informative global patterns that
can improve the robustness of task-specific models. Objective: To evaluate the utility of EHR foundation models in improving the in-distribution (ID)
and out-of-distribution (OOD) performance of clinical prediction models. Methods: The cohort consisted of adult inpatients admitted between 2009-2021. Gated
recurrent unit (GRU)- and transformer (TRANS)-based foundation models were pretrained on
EHR of patients admitted between 2009-2012 and were subsequently used to construct patient
representations (CLMBR). These representations were used to learn logistic regression models
(CLMBRGRU and CLMBRTRANS) to predict hospital mortality, long length of stay, 30-day
readmission, and ICU admission. We compared CLMBRGRU and CLMBRTRANS with baseline
logistic regression models learned on count-based representations (count-LR) and end-to-end
(ETE) GRU and transformer models in ID (2009-2012) and OOD (2013-2021) year groups. Performance was measured using area-under-the-receiver-operating-characteristic curve, area-
under-the-precision-recall curve, and absolute calibration error. Results: Models trained on CLMBR generally showed better discrimination relative to count-LR
in both ID and OOD year groups. In addition, they often matched or were better than their ETE
counterparts. Finally, foundation models’ performance in the self-supervised learning task
tracked closely with the ID and OOD performance of the downstream models. Conclusions: These results suggest that pretraining foundation models on electronic health
records is a useful approach for developing clinical prediction models that perform well in the
presence of temporal distribution shift. . CC-BY 4.0 International license
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medRxiv preprint INTRODUCTION (which was not certified by peer review)
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medRxiv preprint downstream models have demonstrated performance benefits compared to standard baselines
including count-based models, especially when the number of patient records was small. downstream models have demonstrated performance benefits compared to standard baselines
including count-based models, especially when the number of patient records was small. In this study, we evaluated the utility of CLMBR in mitigating the impact of temporal
distribution shift on model performance. We hypothesized that the global patterns embedded in
CLMBR can be adapted into models that perform better than count-based representations in
out-of-distribution (OOD) years. In addition, we characterized the robustness of popular
architectures used in clinical settings, namely gated recurrent unit (GRU)[14] and transformers
(TRANS)[15, 16]. Further, to understand the contribution of CLMBR, we evaluated both GRU
and TRANS end-to-end (ETE) models. Therefore, the objectives were to compare the in-
distribution (ID) and OOD performance of CLMBRGRU, CLMBRTRANS, ETEGRU and ETETRANS
compared to models trained using count-based representations. Data Source We used data from the STAnford medicine Research data Repository (STARR)[17]. Data in STARR are routinely collected in the EHR of Stanford Medical Center, comprised of
Stanford Health Care (primarily adult-directed care) and Lucile Packard Children’s Hospital
(primarily pediatric-directed care). These data are mapped to the Observational Medical
Outcomes Partnership Common Data Model (OMOP-CDM), which facilitates multi-center
observational research[18, 19]. The resulting dataset was named STARR-OMOP. This study
used de-identified data in which dates were jittered by up to 30 days but were accurate within a
patient timeline. Because of de-identification, the requirement for Institutional Review Board
approval was waived by Stanford Medical Center. INTRODUCTION The large increase in the adoption of electronic health records (EHR) has enabled the
use of machine learning to develop highly performant clinical prediction models that have the
potential to improve the care of patients[1]. However, the non-stationary healthcare environment
can bring about changes in the data distribution between model development and
deployment[2], which can degrade the model’s performance over time[3] and consequently its
clinical utility[4]. In this study, we explored temporal distribution shift alongside the suitability of
foundation models[5] – deep neural networks trained on large-scale unlabeled data using self-
supervised learning – and whether they can be adapted via transfer learning to improve the
robustness of clinical prediction models in the presence of temporal distribution shift. The cause of temporal distribution shift in clinical medicine is often subtle[6] and the
extent of its impact on model performance is heterogeneous across tasks[3, 7-9]. Nonetheless,
the consequence of the impact on patient care and physician’s trust can be severe. An example
is the widely implemented Epic sepsis model developed on data collected between 2013-2015
that performed below expectation when evaluated at Michigan Medicine on data collected
between 2018-2019 and resulted in a large number of spurious alerts[4]. Recent approaches that mitigate the impact of temporal distribution shift on model
performance in clinical medicine largely rely on model monitoring and updating policies that do
not leverage the entire patient population available[10]. In addition, proactive approaches using
domain generalization and adaptation have shown little to no success[3]. To date, few studies have explored learning contextualized patient representations at
scale. Findings from domains outside of clinical medicine suggest significant performance[11]
and robustness[12, 13] benefits to pretraining foundation models. In this study, we adopt
CLMBR,[14] an EHR foundation model pretrained on patient timelines comprised of sequential
structured medical codes using autoregressive sequence modeling as the self-supervised
learning task. Transfer of the structure learned by CLMBR from the entire patient population to . CC-BY 4.0 International license
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is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Outcomes We defined four clinical outcomes. Hospital mortality was defined as a patient death
occurring during the hospital stay. Long length of stay (LOS) (long LOS) was defined as a
hospital admission of seven or more days. Readmission in 30 days (30-day readmission) was
defined as a readmission to an inpatient unit within 30 days after discharge. Intensive care unit
(ICU) admission was defined as a patient transfer to the intensive care unit during the hospital
admission. Each outcome was considered as a binary classification task where the prediction
time (also the index time) was set as 11:59PM on the day of admission for the hospital mortality,
long LOS and ICU admission tasks, and 11:59PM on the day of discharge for the 30-day
readmission prediction task. For the 30-day readmission task, we removed patients who were
re-admitted on the day of discharge, and for the ICU admission task, we removed patients
transferred on the day of admission since these events would have occurred before prediction
time. Cohort . CC-BY 4.0 International license
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medRxiv preprint We included adult patients over the age of 18 with admissions to the inpatient unit
between EHR inception (2009) and August 22, 2021. Admissions to the inpatient unit were
either direct or transfers from the emergency department. Encounters with clinic visit alone and
encounters in which patient death or discharges occurred on the same day of admission were
excluded. For patients with multiple admissions, one was randomly selected so that each
patient was only represented once in the dataset. Patient Representations EHR data corresponding to a particular patient can be treated as a sequence of days
that is ordered by time, d1 … dN, where each day consists of a set of events represented by
medical codes such as diagnoses, lab tests, procedures, and medication administrations or
prescriptions as examples. In this study, we considered two approaches to construct patient . CC-BY 4.0 International license
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medRxiv preprint representations over the patient timelines as illustrated in Figure 1: count-based representations
and CLMBR. representations over the patient timelines as illustrated in Figure 1: count-based representations
and CLMBR. representations over the patient timelines as illustrated in Figure 1: count-based representations
and CLMBR. Count based representations: The count-based representations were constructed using an
open-source count-based featurizer[20] which follows standard practices for patient count-
based featurization.[1, 21] This approach constructed patient representations as binary features
based on counts of both unique OMOP CDM concepts and derived elements recorded prior to
the time of prediction. The feature set consisted of demographic and clinical features. Demographic features included sex at birth, race, ethnicity, and age at admission discretized
into five-year intervals. Clinical features were constructed as the concatenation of the results of
a time-dependent extraction procedure applied independently to data elements recorded in time
bins defined relative to the time of prediction. The time bins were as follows: 24 hours prior, 1-7
days prior, 8-30 days prior, and 31 days-any time prior. The time-dependent extraction
procedure identified all unique OMOP CDM concepts from the following OMOP CDM tables:
condition occurrence (diagnosis codes), drug exposure (administration or prescription of
medications), procedure occurrence, measurement (includes laboratory tests), device exposure
(exposure to implantable objects, medical equipment, supplies, and instruments) and
observation (non-standardized tests or clinical observations). Continuous measurement results
were represented as binary indicators for abnormal results for each measurement on the basis
of whether the result was above or below the reference range. Clinical language model-based representations – CLMBR: The core idea behind CLMBR is that
if a sequence model is able to predict sets of medical codes over a patient timeline, then that
model may have discovered informative global patterns that can be re-used in various other
clinical prediction tasks. Note that the term “language model” in CLMBR merely reflects the . CC-BY 4.0 International license
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this version posted April 19, 2022. representations over the patient timelines as illustrated in Figure 1: count-based representations
and CLMBR. CC-BY 4.0 International license
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medRxiv preprint via a sigmoid transformation of the dot product between the code embedding and the patient
representation. To deal with the computational complexity of the matrix product induced by the
large code space, we used a variant of the hierarchical softmax optimization[24] in which we
replaced the softmax transformations with sigmoid transformations. The hierarchical structures
of the code space were the same as the ones used for ontology extension (e.g., the hierarchical
structure in the ICD10 vocabulary for ICD10 codes). We used the binary cross entropy loss as
the loss function during training. Once the sequence models were trained, we used them to construct representations
(the output of the linear layer) for each patient in the cohort to be used by downstream models
for clinical prediction tasks (CLMBRGRU and CLMBRTRANS). For hospital mortality, long LOS, and
ICU admissions, patient representations were obtained up until the day of admission, whereas
for 30-day readmission patient representations were obtained up until the day of discharge. representations over the patient timelines as illustrated in Figure 1: count-based representations
and CLMBR. ;
https://doi.org/10.1101/2022.04.15.22273900
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medRxiv preprint similarity in the computations involved between sequence modeling of medical codes and
language modeling, and therefore does not indicate natural language processing of any kind. similarity in the computations involved between sequence modeling of medical codes and
language modeling, and therefore does not indicate natural language processing of any kind. First, we mapped clinical codes for labs, medications, diagnoses, and procedures to a
finite vocabulary of discrete symbols. This vocabulary was then mapped into a clinical ontology
to reduce sparsity and used to construct patient sequences for the CLMBR encoder. The
medical codes were obtained from the same OMOP CDM tables as used for count-based
representations except for the observation table. The Unified Medical Language System
(UMLS)[22] was used to extend each medical code to the set of parents in its native ontology
when applicable (ICD10 for diagnoses, CPT or MTHH for procedures, and ATC for
medications). For instance, the occurrence of the ICD10 code “H61.23” for the diagnosis of
impacted cerumen, bilateral, resulted in two additional parent codes, namely “H61.2” (impacted
cerumen) and “H61” (other disorders of external ear). We chose GRU and transformer as the architectures for our sequence models as they
have each demonstrated success in the sequence modeling of medical codes in the EHR[14,
18, 19, 23]. To construct patient representations, sets of codes for each day in the patient
timeline were first passed through the embedding bag layer of the networks, which computes
the mean code embedding for each day. Next, each mean embedding was concatenated with a
vector that captured time information including the patient’s age on that day, the time delta from
the previous day, whether that day was the first day of the sequence, and the log transform of
the age and time delta. Patient representations were then computed by feeding the
concatenated vectors into the GRU or transformer (see Supplementary Methods for details on
architecture), followed by a linear layer with output size equal to the number of dimensions of
the patient representation, which was set to 800 in this study. To predict the set of codes for a given day, di, the patient representation from the
previous day, di-1, was used. We formulated the set prediction problem as a series of
independent binary classification problems, where the probability of a given code was computed . Experimental Setup First, we established baseline model performance for each of the four clinical prediction
tasks and investigated whether model performance degraded over time as a result of temporal
distribution shift. We trained logistic regression models on count-based representations
constructed for patients admitted between 2009-2012 (count-LR) and evaluated the models on
all years from 2009-2021. The years on which the models were trained (2009-2012) constituted
the ID years and the subsequent years (2013-2021) constituted the OOD years for the baseline
experiment. We also included oracle models that were trained and evaluated on each of the
OOD years for comparison. Next, we compared ID and OOD performance for four different representation
construction and modeling approaches relative to count-LR, namely CLMBRGRU, CLMBRTRANS,
ETEGRU, and ETETRANS. For this experiment, ID years were 2009-2012 and the OOD years were
2013-2016 and 2017-2021. For statistical comparisons, we focused on comparing CLMBRGRU . CC-BY 4.0 International license
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medRxiv preprint and CLMBRTRANS vs. count-LR, CLMBRGRU and CLMBRTRANS vs. their respective ETE
architecture, and CLMBRGRU vs. CLMBRTRANS. To gain insight into relative performance, we
subtracted the model’s OOD performance in 2017-2021 by its ID performance in 2009-2012 for
the same five representation construction and modeling approaches. To limit multiple testing,
describing relative OOD performance and OOD statistical comparisons only focused on 2017-
2021 and not 2013-2016. We also examined the contribution of CLMBR to downstream model performance by
examining the Pearson correlation between each sequence model’s pretraining performance
and the downstream logistic regression performance in each clinical prediction task. Performance was measured using binary cross-entropy loss. was measured using binary cross-entropy loss. As a sensitivity analysis, we trained and compared task-specific models on count-based
representations and CLMBR using light gradient-boosted machines (LightGBM) instead of
logistic regression. Experimental Setup In addition, to aid clinical interpretation of the changes in performance
between count-LR and CLMBR-based models, we quantified the numbers of decisions that
would have been affected if the CLMBR-based models were used instead of count-LR for tasks
in which performance degraded over time. Specifically, we selected the better performing
CLMBR model (CLMBRGRU vs CLMBRTRANS) and calculated the proportion of patients that would
have been classified correctly and incorrectly with CLMBR-based models instead of count-LR
across various risk thresholds. Model development: The cohort was divided into training (70%), validation (15%), and test
(15%) sets with random sampling stratified by the year in which the admission occurred. We
extracted count-based representations and CLMBR for each patient admission. For count-
based representations, we additionally pruned features with less than 25 observations in the
training set. We then pruned the same features from the validation and test sets. To tune GRU
and transformer for CLMBR, we performed grid search over the hyperparameter settings for . CC-BY 4.0 International license
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medRxiv preprint each architecture separately. For GRU, the hyperparameters consisted of learning rate, L2
regularization strength and dropout rate. For transformer, we additionally tuned the number of
transformer layers and the rate for code dropout. Supplementary Methods detail the
hyperparameter grid and the selected hyperparameter settings. We trained GRUs and
transformers on the timelines of 80% of the patients in the training set of 2009-2012 (29026
patients, ~1.1 million patient days, and ~41.5 million medical codes), and selected
hyperparameter settings based on model performance in the left out 20% of the training set. Then, using the selected GRU and transformer, we constructed CLMBRGRU and CLMBRTRANS for
each patient in the cohort. After computing the representations, we trained logistic regression with L2-regularization
and LightGBM models on count-based representations and CLMBR for each clinical outcome in
the training set of 2009-2012. Experimental Setup Hyperparameter tuning was done on L2 strength, which ranged
from 10-6 to 102 in increments of powers of 10. We selected hyperparameter values based on
the model’s binary cross entropy loss in the validation set of 2009-2012. The ETE models were
trained for each clinical outcome separately on the training set of 2009-2012, and
hyperparameter tuning was conducted using the same grid as CLMBRGRU and CLMBRTRANS for
fair comparisons. We selected hyperparameter settings for ETE models based on model
performance in the validation set of 2009-2012. Oracle models for each OOD year (2013-2021)
as comparisons for count-LR were trained on count-based representations in the training set of
the OOD year, and hyperparameters were selected based on performance in the validation set
of the OOD year. Supplementary Methods provide details on the selected hyperparameter
setting for logistic regression and ETE models for each clinical prediction task. CLMBR and ETE models were implemented using Pytorch[25] and were trained on two
Nvidia V100 GPUs. We used the Sci-kit Learn’s[26] implementation of logistic regression. Analyses were implemented in Python 3.8[27]. . CC-BY 4.0 International license
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medRxiv preprint Model evaluation: We evaluated each model’s discrimination performance in the test sets using
the area-under-the-receiver-operating-characteristic curve (AUROC) and the calibrated area-
under-the-precision-recall curve (AUPRC)[28]. The calibrated AUPRC computes the precision
using a reference outcome prevalence, here set as the prevalence in the ID year group 2009-
2012. Thus, the calibrated AUPRC is invariant to change in outcome prevalence in OOD years
and allows us to better interpret its variation over time. We used the absolute calibration error
(ACE)[20] as a measure of calibration. ACE is similar to the integrated calibration index[29] but
applies a logistic regression estimator to the logit of the predicted probability outputs rather than
locally weighted least squares and is thus more computationally efficient. Experimental Setup Statistical Analysis: For each metric, we computed the median and 95% confidence interval (CI)
of the distribution over performance in the test set obtained from 1000 bootstrap samples. To
compare models, we computed the 95% CI of the differences between a pair of models over
1000 bootstrap samples. Statistical significance was defined as comparisons where the 95% CI
did not cross 0. RESULTS Supplementary Table 1 presents cohort characteristics for each year and outcome
prevalence. Figure 2 shows the impact of temporal distribution shift on performance (AUROC,
AUPRC, and ACE) of count-LR trained on admissions from 2009-2012. Model degradation
occurred in the OOD years (2013-2021) for long LOS and ICU admission prediction tasks, with
larger degradations observed in 2017-2021. Figure 3 shows the relative performance of CLMBR-based and ETE models compared
to count-LR in ID (2009-2012) and OOD (2013-2016 and 2017-2021) year groups (see
Supplementary Tables 2 and 3 for statistical results of pre-specified comparisons). First, and CLMBRTRANS outperformed count-LR in discrimination performance in both ID . CC-BY 4.0 International license
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medRxiv preprint and OOD year groups across all tasks except for 30-day readmission. In terms of calibration, ID
improvement was observed for 30-day readmission for CLMBRTRANS vs. count-LR but other
comparisons between CLMBRGRU or CLMBRTRANS vs. count-LR did not show significant
differences. In contrast, OOD (2017-2021) calibration results showed deterioration in hospital
mortality and 30 day readmission tasks. Second, in general, the ID performance of CLMBRGRU
and CLMBRTRANS were similar to or better than ETEGRU and ETETRANS respectively. However,
differences in OOD 2017-2021 performance varied across tasks: CLMBR models generally
performed better in predicting ICU admission, and the ETE models generally performed better in
predicting long LOS. Third, CLMBRGRU outperformed CLMBRTRANS in discrimination
performance in tasks other than 30-day readmission prediction for ID and for OOD hospital
mortality and long LOS tasks. Figure 4 shows relative OOD performance compared to ID performance across
representation construction and modeling approaches (see Supplementary Table 4 for statistical
results). Comparison of CLMBRGRU or CLMBRTRANS vs. count-LR showed heterogeneous
results. In terms of relative discrimination, both CLMBRGRU and CLMBRTRANS had significantly
better relative AUROC vs. count-LR for long LOS but were similar for other tasks. RESULTS In terms of
relative calibration, CLMBRGRU and CLMBRTRANS were significantly worse than count-LR for
hospital mortality and 30-day readmission tasks. Other comparisons of relative performance
were heterogeneous. Figure 5 plots the average binary cross entropy loss of GRU sequence models (trained
using various hyperparameter settings) in the pretraining validation set against the average
binary cross entropy loss of their downstream logistic regression models in each task and year
group (see Supplementary Figure 1 for the same analysis conducted on CLMBRTRANS). The
performance of GRU sequence models had high correlations (Pearson correlation coefficient)
with the ID and OOD performance of their downstream logistic regression models in all tasks
except for 30-day readmission. . CC-BY 4.0 International license
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medRxiv preprint Figure 6 plots the proportion of patients re-classified differently using CLMBRGRU instead
of count-LR in long LOS and ICU admission. There were generally more correct re-
classifications than incorrect re-classifications across risk thresholds and year groups for long
LOS. For ICU admission, there were more correct re-classifications in lower thresholds. The sensitivity analysis where we trained and compared task-specific models on count-
based representations and CLMBR using LightGBM instead of logistic regression showed
qualitatively similar findings (Supplementary Experiment). DISCUSSION We observed count-LR models resulted in large performance degradation over time for
some tasks, namely long LOS and ICU admission. Models trained on CLMBR generally
displayed better discrimination relative to count-LR in ID and OOD year groups but could result
in worse OOD calibration. In addition, models trained on CLMBR often matched and were
sometimes even better than their ETE counterparts. Finally, in general, CLMBRGRU performed
better than CLMBRTRANS, and its performance in the autoregressive sequence modeling task
tracked closely with the ID and OOD performance of the downstream models for the majority of
tasks considered. Large-scale self-supervised pretraining takes place less frequently and enables machine
learning practitioners to focus on rapid adaptation of these foundation models to downstream
tasks. This research paper contributes more evidence that this approach brings not only
performance benefits over traditional count-based models, but robustness benefits in the
presence of temporal distribution shift. These benefits decrease the need for model retraining
and preserves the clinical utility of models deployed into practice. The strong relationship
between sequence modeling performance and the performance of downstream clinical
prediction tasks suggests that the observed favorable ID and OOD performance can be directly . CC-BY 4.0 International license
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medRxiv preprint attributed to the self-supervised pretraining. Whether the potential for worse calibration is
clinically meaningful will depend on the specific use case. attributed to the self-supervised pretraining. Whether the potential for worse calibration is
clinically meaningful will depend on the specific use case. The transformer architecture was more difficult to tune and performed worse than the
GRU in both CLMBR-based models and ETE models. This may be explained by the smaller
training set size used in this study, which totaled to ~1.1 million patient days and ~41.5 million
medical codes across ~29 thousand patients. of using CLMBR-based models instead of models trained on count-based representations. of using CLMBR-based models instead of models trained on count-based representations. However, this study is limited as we only used a single dataset with a limited number of tasks. Performance of CLMBR may differ in other settings and with other asks. Another limitation is
that we lack insight into scenarios in which CLMBR may be more or less helpful. In conclusion, models trained on CLMBR generally displayed better discrimination
relative to count-LR in both ID and OOD year groups. Models trained on CLMBR often matched
or were better than their ETE counterparts. Finally, autoregressive sequence modeling
performance tracked closely with the ID and OOD performance of the downstream models. These results suggest that pretraining EHR foundation models is a useful approach for
developing clinical prediction models that perform well ID as well as OOD. Acknowledgments Not applicable. Ethics approval and consent to participate This study used de-identified data and so the requirement for Institutional Review Board
approval and participant informed consent were waived by Stanford Medical Center. Data availability statement The Stanford Medicine Research Data Repository is not made publicly available. The code for
all analyses is open-source and available at https://github.com/som-
shahlab/temp_ds_shift_robustness DISCUSSION In comparison, state of the art transformer-based
natural language processing models leveraged much larger volumes of data, for example,
~3,300 times larger (137 billion tokens) for BERT[11], and ~53,000 times larger (2.2 trillion
tokens) for RoBERTa[30]. We examined two attributes of OOD performance, namely absolute performance and
relative (to ID) performance. It is notable that while CLMBR resulted in generally better absolute
discrimination than count-LR, improvement in relative discrimination was more modest, with
improvement only being observed for long LOS. It is likely that absolute performance is more
meaningful to clinicians since better relative performance does not necessarily indicate better
absolute performance[31]. However, decision makers may be more concerned about using a
model that does not perform as well as that originally promised (relative performance). It is for
that reason that we choose to report both aspects. Despite the reasonable performance of CLMBR-based models in OOD year groups, they
are not immune to the impact of distribution shift as seen in predicting ICU admissions and long
LOS. Increasing parameter size and training set size, which have demonstrated benefits in
other modalities[30, 32], could provide additional improvements to robustness. In addition,
structure and certain types of invariances could be incorporated during pretraining or at the
adaptation stage using metadata[33], regularization[34], or contrastive learning[35]. The strengths of this study include the evaluation of a novel approach to self-supervised
representation learning on electronic health records, namely CLMBR, in both ID and OOD
settings. Another strength is the adoption of interpretable metrics to evaluate the clinical impact . CC-BY 4.0 International license
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medRxiv preprint Funding No external funding was received for the study. Competing interests The authors declare that they have no competing interests. . CC-BY 4.0 International license
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medRxiv preprint Authors’ contributions LLG, JF, and LS conceptualized and designed the study with input from all authors. L.L.G. performed all experiments. ES and SRP contributed to the codebase. LLG, JF, and LS analyzed
and interpreted results with input from all authors. LLG wrote the manuscript. All authors revised
and commented on the manuscript. All authors read and approved the final manuscript. . CC-BY 4.0 International license
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medRxiv preprint Figure 1. An overview of the two approaches of constructing patient representations
used in this study. The purple box in the construction of count-based representations
represents the reference range comparison and binary feature construction procedures
for a specific time-bin. The construction of CLMBR illustrates the self-supervised
pretraining stage, hence the inclusion of the self-supervised learning objective. The
construction of CLMBR for the purpose of transfer learning (e.g., for predicting hospital
mortality) does not include the self-supervised learning objective. Abb
i
i
CLMBR
li i
l l
d l b
d
i Abbreviations: CLMBR: clinical language model-based representations. . CC-BY 4.0 International license
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medRxiv preprint Figure 2. The impact of temporal distribution shift on the performance (AUROC,
AUPRC, and ACE) of logistic regression models trained on count-based
representations. Shaded regions indicate time windows in which performance in out-of-
distribution years (2013-2021) is worse (red) or better (green) than performance in the
in-distribution year group (2009-2012). REFERENCES Oracle models were trained and evaluated on
each of the out-of-distribution years. Error bars indicate 95% confidence interval
obtained from 1000 bootstrap iterations. p
Abbreviations: AUROC: area under the receiver operating characteristics curve;
AUPRC: area under the precision recall curve; ACE: absolute calibration error; LOS:
length of stay; ICU: intensive care unit. . CC-BY 4.0 International license
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medRxiv preprint . CC-BY 4.0 International license
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medRxiv preprint Figures 3. Change in performance (AUROC, AUPRC, and ACE) across representation
construction and modeling approaches relative to count-based logistic regression
models (Count-LR) in 2009-2012 (09-12), 2013-2016 (13-16), and 2017-2021 (17-21). Performance of count-LR was subtracted to obtain relative performance. Green regions
indicate the range of values that is better with respect to count-LR, and red regions
indicate the range of values that is worse. Error bars indicate 95% confidence interval
obtained from 1000 bootstrap iterations. Supplementary Tables 2 and 3 detail the
statistical results. s Abbreviations: AUROC: area under the receiver operating characteristics curve;
AUPRC: area under the precision recall curve; ACE: absolute calibration error; LOS:
length of stay; ICU: intensive care unit; CLMBR: clinical language model-based
representation; GRU: gated recurrent unit; Trans: transformer; LR: logistic regression;
ETE: end-to-end. . CC-BY 4.0 International license
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is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)
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medRxiv preprint Figure 4. Relative out-of-distribution (OOD) performance (AUROC, AUPRC, and ACE)
across representation construction and modeling approaches. REFERENCES Each relative
performance was obtained by subtracting the model’s OOD performance in 2017-2021
(17-21) by its in-distribution (ID) performance in 2009-2012 (09-12) represented by the
solid line. Colored regions indicate the range of values that is worse (red) or better
(green) with respect to the relative OOD performance of logistic regression models
trained on count-based representations. Error bars indicate 95% confidence interval
obtained from 1000 bootstrap iterations. Supplementary Table 5 details the statistical
results. Abbreviations: AUROC: area under the receiver operating characteristics curve;
AUPRC: area under the precision recall curve; ACE: absolute calibration error; LOS:
length of stay; ICU: intensive care unit; CLMBR: clinical language model-based
representation; GRU: gated recurrent unit; Trans: transformer; LR: logistic regression;
ETE: end-to-end. . CC-BY 4.0 International license
It is made available under a
is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)
The copyright holder for this preprint
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medRxiv preprint . CC-BY 4.0 International license
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medRxiv preprint Figure 5. Correlation between the GRU sequence model’s validation performance and
the performance of the downstream logistic regression models in each clinical prediction
tasks. Performance for both the sequence model and the logistic regression model were
measured using binary cross entropy loss. Each point in the scatter plot represents the
GRU’s performance in the validation set and its downstream logistic regression model’s
performance in the test set. Each GRU sequence model was pretrained using a
different hyperparameter setting from the hyperparameter grid. Shaded error envelope
represents the 95% confidence interval around the regression line. Abbreviations: GRU: gated recurrent unit; LOS: length of stay; ICU: intensive care unit. on
re
’s p
g
Abbreviations: GRU: gated recurrent unit; LOS: length of stay; ICU: intensive care unit. . CC-BY 4.0 International license
It is made available under a
is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)
The copyright holder for this preprint
this version posted April 19, 2022. ;
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doi:
medRxiv preprint . REFERENCES CC-BY 4.0 International license
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is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)
The copyright holder for this preprint
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medRxiv preprint . CC-BY 4.0 International license
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medRxiv preprint Figure 6. The proportion of patients correctly and incorrectly re-classified by
CLMBRGRU. Green bars indicate the proportion of patients that were incorrectly
classified by count-LR but were correctly re-classified by CLMBRGRU. Orange bars
indicate the proportion of patients that were correctly classified by count-LR but were
incorrectly re-classified by CLMBRGRU. Together, these represent the percentage of
decisions that would have been affected if CLMBRGRU models were used instead of
count-LR. Abbreviations: count-LR: logistic regression models trained on count-based
representations; CLMBRGRU: logistic regression models trained on gated recurrent unit-
based CLMBR. CLMBR: clinical language model-based representations; LOS: length of
stay; ICU: intensive care unit. of
|
https://openalex.org/W3164849643
|
https://ris.utwente.nl/ws/files/254859456/Wijnhoven_2021_Search_engine_gender_bias.pdf
|
English
| null |
Search Engine Gender Bias
|
Frontiers in big data
| 2,021
|
cc-by
| 11,664
|
Search Engine Gender Bias This article discusses possible search engine page rank biases as a consequence of
search engine profile information. After describing search engine biases, their causes, and
their ethical implications, we present data about the Google search engine (GSE) and
DuckDuckGo (DDG) for which only the first uses profile data for the production of page
ranks. We analyze 408 search engine screen prints of 102 volunteers (53 male and 49
female) on queries for job search and political participation. For job searches via GSE, we
find a bias toward stereotypically “female” jobs for women but also for men, although the
bias is significantly stronger for women. For political participation, the bias of GSE is toward
more powerful positions. Contrary to our hypothesis, this bias is even stronger for women
than for men. Our analysis of DDG does not give statistically significant page rank
differences for male and female users. We, therefore, conclude that GSE’s personal
profiling is not reinforcing a gender stereotype. Although no gender differences in page
ranks was found for DDG, DDG usage in general gave a bias toward “male-dominant”
vacancies for both men and women. We, therefore, believe that search engine page ranks
are not biased by profile ranking algorithms, but that page rank biases may be caused by
many other factors in the search engine’s value chain. We propose ten search engine bias
factors with virtue ethical implications for further research. INTRODUCTION Search engines enable their users to receive relevant search results in the enormous mass of data on
the Web. The most used and best known search engine is the Google search engine, named GSE in
the rest of this article. By collecting data about its users, GSE creates personal profiles of its users for
targeted advertising service (Thurman and Schifferes, 2012). With the same process, GSE also
personalizes search outcomes and page ranks for optimizing a search outcomes relevance experience
for its users (Rogers 2013; De Corniere and Taylor, 2014). Personal profile processing, as part of
search query processing, may result in a high use value experience but also can keep important less
relevant information away from its user (Helberger et al., 2018). This “filter bubble effect” (Pariser
2011) also comes with benefits, as people lack the capacity to process all the zettabytes of information
available on the Internet. This also implies that people with different profiles who enter the same
query into the GSE receive a different list of relevant Web pages. This ability of semantic search
enables GSE to differentiate the meaning of ambiguous terms in a query. For example, while a lawyer
would refer to the keyword “code” as a set of rules or the law, an information technology specialist
may be more interested in “code” referring to software, and thus a profile aware search engine will
serve both differently. This subjective relevance increasing process may, in a subtle way, result in
influencing users to certain viewpoints and opinions (Epstein and Robertson 2015), which is a
process also named nudging (Gal et al., 2020). *Correspondence:
Fons Wijnhoven
a.b.j.m.wijnhoven@utwente.nl Specialty section:
This article was submitted to
Data Mining and Management,
a section of the journal
Frontiers in Big Data Received: 27 October 2020
Accepted: 04 May 2021
Published: 26 May 2021 Edited by:
Mascha Kurpicz-Briki,
Bern University of Applied Sciences,
Switzerland Reviewed by:
Jahna Otterbacher,
Open University of Cyprus, Cyprus
Jason Millar,
University of Ottawa, Canada Reviewed by:
Jahna Otterbacher,
Open University of Cyprus, Cyprus
Jason Millar,
University of Ottawa, Canada
*Correspondence:
Fons Wijnhoven
a.b.j.m.wijnhoven@utwente.nl ORIGINAL RESEARCH
published: 26 May 2021
doi: 10.3389/fdata.2021.622106 Keywords: personalization, Google, DuckDuckGo, filter bubble, gender bias, job search, political participation
search BACKGROUND, THEORY, AND
HYPOTHESES Personalized search results are customized search results based
on the digital profile of an individual user. When using a personal
profile aware search engine such as GSE or Bing, search results
will differ depending on who is supplying the query to the search
engine. This profile is supposed to represent the interests of the
individual user (Harvey et al., 2013). GSE’s updated privacy
policy of 2012 states that information is being shared on all
sorts of Google tailors, which include Gmail, YouTube, and
Google Maps for further profiling of the search engine user
(Voss 2014). Also, search location indicators (Hannak et al.,
2013) and browsing histories can combine with user attributes
to produce personalized outcomes (Goel et al., 2012). A
manipulation effect occurs if the search engine can influence
users’ attitudes, beliefs, or decisions based on the order of the
search engine results. A manipulation effect may result from
search results that are biased toward one view. By providing one-
sided search results, GSE could either reinforce an existing
opinion or it may turn an undecided person to be nudged
toward one opinion direction (Epstein and Robertson 2015;
Wijnhoven and Brinkhuis 2015). There is a substantial volume of research outcomes regarding
the difference in the use of the Internet between men and women
(Colley and Maltby 2008; Yom-Tov and Elad, 2019), but there is
little knowledge on what the role of the medium is in this story. Hargittai and Shafer (2006) suggest that the supply-side of
content—due to its structure and presentation—is in itself
male-biased. This would imply that gender bias of search
engine
page
ranks
is
not
necessarily
the
result
of
personalization but could be the result of biased supplies of
information on the Internet as well. Unfortunately, it is
difficult to identify what GSE knows about you, we only know
that personal profile processing is a key part of GSE’s proprietary
way of delivering relevant content to its user (Bozdag 2013;
Rogers 2013). Such a personal profiling algorithm is lacking in
privacy-aware search engine DuckDuckGo (DDG), and thus
comparing page ranks of male and female users of both using
GSE and DDG for the same queries would be able to determine if
the search engine is the cause of page rank bias or if alternative
reasons exist for page rank bias. Citation: Wijnhoven F and van Haren J (2021)
Search Engine Gender Bias. Front. Big Data 4:622106. doi: 10.3389/fdata.2021.622106 Wijnhoven F and van Haren J (2021)
Search Engine Gender Bias. Front. Big Data 4:622106. doi: 10.3389/fdata.2021.622106 May 2021 | Volume 4 | Article 622106 1 Frontiers in Big Data | www.frontiersin.org Wijnhoven and van Haren Search Engine Bias A common nudge in computer environments is the setting of
defaults, which are preset courses of action that take effect if
nothing is specified by the decision maker (Cronqvist et al., 2018). But nudging can also take more elaborate and sinister forms. For
example, in 2017, it became known that advertisers on Facebook
could target advertisements at teenagers during moments of
psychological vulnerability. This kind of nudging can also be
employed by organizations through the use of algorithmic
systems to encourage “appropriate” behaviors. For instance,
Kaptein et al. (2015) have shown that emails and Web site
usage nudges can move people to more healthy behavior. But
these
practices
can
also
be
manipulative
and
ethically
questionable when they are aimed at covertly subverting
people’s
decision-making
capacity
by
exploiting
their
psychological, cognitive, or emotional vulnerabilities to change
their beliefs, thoughts, or behaviors. Such manipulative practices
limit people’s capacity for voluntary action and learning. Nudging
may not only be unethical from a utilitarian perspective (Tavani
2012), that is, the diminishing value of what is received for the
search engine user, but also from a virtue ethics perspective as it
disallows the user to identify the source of bias, learn from that,
and reduces the freedom of voluntary action of what to believe or
not (Gal et al., 2020). Following Gal et al. (2020), we choose for a
virtue ethics approach for the analysis of search engine bias. Virtue ethics, based on the philosophical work of Aristotle,
highlights personal characteristics in determining the ethical
nature of individuals and their actions. Virtue ethics focus on
the virtuous agent rather than on the right actions to reach certain
values (i.e., the utilitarian approach) or on what anyone should do
given certain rules (i.e., the deontological approach to ethics). reproduce, or reinforce biases with virtue ethical implications
for other social segments. BACKGROUND, THEORY, AND
HYPOTHESES For knowing if this bias has virtue
ethical consequence, we will take queries that may have important
consequences for women to learn and develop themselves. The ranking of search results is especially important
because users tend to pay more attention to the results that
rank higher on the search results page (Bozdag 2013). The
actual impact of search engine and page rank biases is also
named as the search engine manipulation effect. A slight
manipulation of a page rank can have an impact on the
outcome of elections by providing search results that are in
favor of one candidate (Epstein and Robertson, 2015). This
eventually causes a user to be less likely to discover new topics
on the Web or to see documents with different views on a topic
that contradicts his or her values (Wilson et al., 2012; Burger
et al., 2016). The purpose of this study was to gain further
insights into possible search engine bias toward one-sided
search results with ethical implications. This leads to the research question: To what extent does the
Google search engine creates a biased page rank for its users with
negative virtual ethical consequences? To answer this question, this article focuses on search engine
gender bias and its virtue ethical implications for women. Besides
the commitment of the authors to equal opportunities of women
in our society, gender characteristics are one of the most common
and easily traced search engine profile features (Kosinski et al.,
2013). A search engine’s gender bias is the influence of gender
features in a search engine profile on gender bias of a search
engine’s outcomes and page rank (Lopes et al., 2016). The insight
from
a
gender-bias
study
can
produce
generalizable
understandings
on
how
search
engines
may
produce, This section first goes into more details about causes and
consequences of search engine bias. Then it will set out the
concerns regarding gender inequalities and our hypotheses. Frontiers in Big Data | www.frontiersin.org Jobs According to Scherer (2004), the labor market entry and a
successful transition from school to work are of crucial
importance for subsequent career chances, it is a good start to
look at the possibly different opportunities here for men and
women. Defloor et al. (2015) found that the quality of a person’s
first job is largely dependent on personal effort, although
circumstances, for example, gender do have a considerable
influence on these efforts. This would, therefore, suggest that a
difference of jobs between men and women is due to a difference
in effort spent between men and women when searching for their
first job. Equality of opportunity besides efforts is also highly
debated in relation to gender. Equality of opportunity means that
outcomes experienced by a population depend only on factors for
which persons can be considered to be responsible (Roemer and
Trannoy 2015). As people are increasingly searching for jobs
online (Nikolaou 2014), gender indicators in personal profiles
may disturb the equality of finding the same job opportunities on
the Internet, although Chen et al. (2018) did not find evidence for
gender bias in recruiting site algorithms. As Pariser (2011)
pointed out, the filter bubble not only reflects a person’s
identity but it also selectively shows the related options a
person has. This may be positive, because some options may
be less relevant for people with a specific profile, for example, jobs
for university alumni have less relevance for those with higher
vocational education. In defense of personal profile processing,
one may see this as the effective working of relevance. Alternatively,
however,
personal
profile
processing
search
engines can replicate and reinforce current gender stereotypes
on the job market. The existence of these stereotypical outcomes
as a consequence of the usage of user profile information has been
identified before by Otterbacher et al. (2017), but one may ask the
question if a privacy-aware search engine like DDG will be free of
such bias. Table 1 gives a list of various occupations classified as
male-dominated, mixed, or female-dominated based on data
from 12 EU countries. Page rank bias has ethical implications, because search engine
outcomes may impact people’s opinions, choices, and actions,
and search engines may be intentionally used by stakeholders to
influence society by attempts of censorship and search engine
optimization (Fu and Karan 2015; Baye et al., 2016). Jobs The
influence of search engine should not be understated and thus
identifying possible page rank biases is important from an ethical
perspective (Flaxman et al., 2016; Haim et al., 2018), and its page
rank outcomes can nudge people toward certain opinions and
decision (Epstein and Robertson 2015) and reinforce possible
stereotypical views on people and gender (Kay et al., 2015;
Otterbacher et al., 2017). Recent experiences have shown that
GSE sometimes intentionally nudges its users toward Web pages
that are commercially more interesting from Google’s own
perspective (Blanckenburg 2018). Google has been punished
for this by the European Committee and corrections have
been implemented, but also for nonshopping purposes, search
biases can easily happen because of a lack of transparency and the
proprietary nature of GSE services (van Drunen et al., 2019). The
virtue ethical implications of search engines can be very diverse. For example, it may result in a lack of capability of voters to judge
and choose among election candidates (Epstein et al., 2017), or
the reinforcement of racist views and behavior (Noble 2018), and
the lack of critical thinking (Heersmink 2018). We choose for a
focus on virtue ethics from a gender perspective, that is, the
abilities of women to emancipate from stereotypical roles and
achieve gender equality on the workplace and society. Furthermore, regarding working sectors, women make up
almost 80% of those employed in health and social work, over
70% of those employed in education, and over 60% of those
working in retailing. In contrast, only 8% of those employed in
construction and 14% of those in land transport are women. This
pattern is visible throughout all EU member states. We thus
expect the following from a profile-aware search engine: Gender Inequalities q
According to the United Nations Development Program (UNDP,
United Nations Europe and Central Asia, 2018), there are still
significant inequalities between women and men in Europe and
Central Asia, particularly when it comes to jobs and income,
political participation, and the distribution of unpaid domestic
and care work. Similarly, gender stereotypes are prevalent,
hindering women’s access to opportunities. Furthermore, men
are more likely to gain promotion to top management positions
and prestigious leadership roles than women. Moreover, women
are more likely to have insecure jobs, no contract or regular salary,
or part-time jobs (UNDP, United Nations Europe and Central
Asia, 2018). Search engine outcomes, like any message or piece of
information, can reproduce or reinforce views on and the reality
of gender inequalities related to jobs and leadership roles in
society (Otterbacher et al., 2017; Chen et al., 2018). We, therefore, H1: Women are mainly shown vacancies for female-dominant
jobs in their GSE search results. H2: Men are mainly shown vacancies for male-dominant jobs
in their GSE search results. Causes and Virtue Ethical Consequences of
Search Engine Bias Search engine bias includes a selective presentation of Web
documents to a search engine user. This selectivity can be May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 2 Wijnhoven and van Haren Search Engine Bias will go in more detail on jobs and leadership roles in society as
two gender-specific virtue ethical topics. will go in more detail on jobs and leadership roles in society as
two gender-specific virtue ethical topics. selectivity of sources, content, views, and page ranks (Bozdag
2013). The causes for these biases can be explicit by censorship
that disables users to access certain sources, content, viewpoints,
and algorithms, that is, will make it difficult for users to find and
see certain content, or the outcome of optimization actions of
content suppliers, that is, through search optimization. Finally,
search engine bias may also be the result of account and behavior
information that is delivered to the search engine, and by which
the search aims at increasing its service level may increase in the
perception of the user. If this user profile information is used for
page ranking, it increases the probability that users will access less
diverse sources, content, and viewpoints, which is called the filter
bubble (Pariser 2011; Flaxman et al., 2016). Political Participation Cabeza-García et al. (2018) found differences between men and
women regarding political participation. They found evidence
that participation in bodies of power and institutions is
predominantly
masculine,
whereas
voluntary
associations,
organizations, and “informal” community politics tend to be May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 3 Search Engine Bias Wijnhoven and van Haren TABLE 1 | Gender occupation stereotypes (Bettio and Verashchagina 2009). Occupation
Number of countries where occupation is dominantly. . . Male
Mixed
Female
Armed forces
10
0
0
Legislators, senior officials, and managers
7
5
0
Corporate managers
8
4
0
Managers of small enterprises
5
7
0
Physical, mathematical, and engineering science professionals
12
0
0
Life science and health professionals
0
7
5
Teaching professionals
0
2
10
Other professionals
0
11
1
Physical and engineering science associate professionals
11
1
0
Life science and health associate professionals
0
0
12
Teaching associate professionals
0
2
8
Other associate professionals
0
10
2
Office clerks
0
0
12
Customer services clerks
0
0
12
Personal and protective services workers
0
2
10
Models, salespersons, and demonstrators
0
1
11
Skilled agricultural and fishery workers
8
4
0
Extraction and building trades workers
12
0
0
Metal, machinery, and related trades workers
12
0
0
Precision, handicraft, craft printing, and related trades workers
5
7
0
Stationary plant and related operators
11
1
0
Machine operators and assemblers
5
6
1
Drivers and mobile plant operators
12
0
0
Sales and services elementary occupations
0
0
12
Agricultural, fishery, and related laborers
3
6
2
Laborers in mining, construction, manufacturing, and transport
12
0
0 As explained above, women are more likely to engage in
certain political practices while men in other, which could
possibly be reflected and reinforced through personalization
techniques used by GSE. We believe that this difference
between men and women in politics can be interpreted as
involvement of women in less powerful political activities and
involvement of men in more powerful activities, consistent with
previously identified stereotypes of competent men and warm
women as found on an analysis of Internet images (Otterbacher
et al., 2017). This leads to the following hypotheses: led by women. Moreover, the number of women as member of
governments was only 16% in 127 countries, and not one of the
women was in a top parliamentary management post. Political Participation Furthermore,
the
women
who
had
a
post
in
local,
municipal, or national government occupied posts with a
more social and cultural nature and thus with less political
importance. Brandtzaeg (2017) studied the difference between men and
women in expressions of civic engagement on Facebook and
found that millennial women more often engage with posts
related to children and the environment when compared to
men of the same age. Moreover, it was found that patterns in
the offline world regarding the differences among men and
women in civic engagement were reproduced and reinforced
rather than equalized on Facebook. H3:
Women
are
mainly
shown
political
involvement
opportunities for positions of low power and influence in their
GSE results. H4:
Men
are
mainly
shown
political
involvement
opportunities for positions of high power and influence in
their GSE results. Pfanzelt and Spies (2019) studied the difference between
young
German
men
and
women
regarding
political
participation. They
distinguished
between
three
types
of
political participation, namely, institutional, noninstitutional,
and expressive. The first one covers activities that mainly
address the state via participation in elections or actively
running for or holding office. Noninstitutional participation
refers
to
protest
activities
like
boycotts
and
expressive
participation includes giving voice to political aims and
intentions of citizens. The Internet is a main medium for
expressive participation. They found that young men are more
likely to participate in institutional and expressive forms, while
young women tend toward noninstitutional, protest-oriented
activities. Participants’ Actions p
We received 144 respondents; however, not all of these
respondents could be used due to certain requirements set for
this research. One of these requirements is related to cookies. Participants were asked when they removed their cookies for the
last time. A cookie is a small piece of data sent from the Web site
and stored on the user’s browser that is sent back to the Web site
every time the user returns (Coey and Bailey 2016). The
information stored on the cookies is used to create a user’s
profile by GSE. This has led to the decision of including all
participants who removed their cookies two weeks ago or later as
the frequent use of the Internet and the instant installation of
cookies gives GSE an impression of the users’ profiles in these
2 weeks. Other requirements include the use of a personal
computer or laptop instead of a phone, being 18 or older, and
the use of GSE as the main search engine. This left 102 suitable
participants for this study, 53 males and 49 females. All of the
participants are students following a higher vocational education,
bachelor, or master. The age ranges from 18 to 29 years, with
most participants in the range of 20–23 years. The three most
occurring nationalities are Dutch (60.8%), Mexican (8.8%), and
German (7.8%). This section describes the methodology for observing and
analyzing if people’s search results are biased. DuckDuckGo DuckDuckGo (DDG) is said to not gather personal data from its
users for its page ranks (Hannak et al., 2013). Instead, DDG bases
their search results on expert advice. This means that they focus
only on the search term and its semantics and what documents an
expert would recommend to access (Jansen and Spink 2006;
Jatwani et al., 2020). Thus, considering the method used by
DDG, it is expected that search results for every individual
user will be nearly the same. This leads to the following
hypotheses: H5: There is no gender bias in the vacancies shown by
DuckDuckGo to men and women. May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 4 Wijnhoven and van Haren Search Engine Bias H6: There is no gender bias in the political involvement
opportunities shown on DuckDuckGo to men and women. outcomes, one of them who is also the co-author of this article. Furthermore, it has been shared in the WhatsApp groups of
these two students. Research Design DDG and GSE are the units of analysis in this study. In total,
144 students participated in this study. All participants
needed to be at least 18 years of age to assure that they are
legally allowed to accept the terms of this research study and
that their data are being used for research purposes. Participants should use GSE as their main search engine
and should have also not removed their browser cookies too
recently so that a rich personal profile could be created by
GSE. Participants have been excluded from the research if
they have removed their browser cookies in the last 2 weeks. Furthermore,
individuals
need
to
participate
in
the
experiment on their own personal laptops or computers to
be suitable for the study. Two search queries have been used that address issues
that
can
cause
people
to
have
different
page
ranks. The
selected
search
queries
are
about
highly
different gender virtue issue to avoid possible carry over
effects, that is, one search query influencing another one in
the same study. Analysis To analyze the collected data, a coding scheme is designed for
determining if a query result is biased toward one side. The data
analysis in this research is a directed content analysis because this
research has started with an explanation of theories related to the
research question and from this the coding scheme has followed
(Hsieh and Shannon 2005). Only the first four delivered screen
shots of organic search results have been analyzed, as the fifth and
the sixth result were not always visible on all screenshots. This
makes a total of 408 screen shots to be coded. The points given to
a screenshot range from −6, “only male-biased search results are
shown,” to 6, “only female-biased search results are shown.” The
first two results have been given extra weight and score −2 for
male-biased and 2 for female-biased, as according to (Epstein
et al., 2017) people generally believe that the top results are
presented first. The third and the fourth result are given either −1,
for male-biased, or 1 for female-biased. This way, this study
measures page ranking bias. If there is no bias present, zero points
are given. For example, when the first three search results were
determined to be female-biased and the fourth was determined
male-biased, the screenshot would receive a score of 2 + 2 + 1 −
1 4. The detailed coding scheme is given in Supplementary
Appendix Tables S1, S2. Frontiers in Big Data | www.frontiersin.org May 2021 | Volume 4 | Article 622106 Data Collection (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
6.286
96
0.000
1.361
0.93
1.79
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
7.757
0.006
−1.875
95
0.064
−0.802
0.428
−1.652
0.047
Not assumed
−1.845
81.795
0.069
−0.802
0.435
−1.667
0.063
One sample test including only male participants
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
3.951
50
0.000
0.980
0.48
1.48
Wijnhoven and van Haren
Search Engine Bias Search Engine Bias Wijnhoven and van Haren distributed and therefore for all four variables (“Jobs Google,”
“Politics
Google,”
“Jobs
DDG,”
and
“Politics
DDG”),
a
Shapiro–Wilk test has been conducted to test for normality. The Shapiro–Wilk test uses the null hypothesis that the
population is normally distributed, which is rejected whenever
the p-value falls below a certain alpha value, in our case when the
p-value is lower than 0.05. For all four variables, the null
hypothesis of normal distribution is rejected as the p-value is
TABLE 2 | SPSS output for “Job openings near me Google.”
Sample statistics for “Job openings near me Google”
Mean
Std. deviation
Std. error mean
N
97
1.36
2.132
0.216
Male
51
0.98
1.772
0.248
Female
46
1.78
2.421
0.357
One-sample test
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
6.286
96
0.000
1.361
0.93
1.79
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
7.757
0.006
−1.875
95
0.064
−0.802
0.428
−1.652
0.047
Not assumed
−1.845
81.795
0.069
−0.802
0.435
−1.667
0.063
One sample test including only male participants
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
3.951
50
0.000
0.980
0.48
1.48
TABLE 3 | SPSS output for “How to become involved in politics Google.”
Sample statistics for “How to become involved in politics Google”
Mean
Std. deviation
Std. error mean
N
93
−1.89
2.656
0.275
Male
46
−1.48
2.689
0.397
Female
47
−2.30
2.587
0.377
One-sample test
T
df
Sig. Data Collection Regarding the gender issues, participants of the survey are
asked to type in the two search queries into GSE and the same
two into DDG. These two search queries are “job openings
near me” and “how to become involved in politics.” These
two search queries are based on the literature about the
gender gap in occupations and political participation as
described in Background, Theory, and Hypotheses section
before. To be able to measure the construct of bias, the study consisted
of five parts. The first part asked questions to determine the
suitability of a participant. The second part asked general
questions regarding demographics and behavior on GSE of
the respondent, for example, age, educational level, and the
language mostly used to conduct searches. The third and the
fourth parts asked the participants to upload screenshots of
their first six organic search results appearing, when searching
with “job openings near me” and “how to become involved in
politics.” These organic search outcomes are the nonpaid
search outcomes of the search engine, normally found
below the paid search outcomes (i.e., advertisements) of the
search engine. The third part of the study asked this for GSE,
while the fourth part asked to do this for DDG. The fifth part is
the analysis of the results. After the data have been coded, it has been statistically
analyzed. Independent-samples t tests have been chosen for
comparing the statistical significance of mean for the two GSE
and DDG male and female users (sub)samples. One-sample t tests
are conducted for determining the differences of the means
relative to neutral values for the GSE and DDG page rank
outcomes. Both
tests
require
the
data
to
be
normally To gather enough participants, the link of the answer form
has been shared on the personal Facebook page of two of the
author’s students who also participated in the coding of the May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 5 TABLE 2 | SPSS output for “Job openings near me Google.”
Sample statistics for “Job openings near me Google”
Mean
Std. deviation
Std. error mean
N
97
1.36
2.132
0.216
Male
51
0.98
1.772
0.248
Female
46
1.78
2.421
0.357
One-sample test
T
Df
Sig. Data Collection (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
−2.816
93
0.006
−0.255
−0.44
−0.08
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
0.006
0.939
−0.055
92
0.956
−0.010
0.183
−0.373
0.353
Not assumed
−0.055
91.232
0.956
−0.010
0.182
−0.372
0.352 TABLE 4 | Detailed SPSS output for “Job openings near me DDG.” below 0.05. Hence, a further analysis of Q-Q plots is used to check
the normality of the data distribution by a visual inspection of the
line that the data make with the theoretically ideal normal
distribution. These plots show straight upward lines, thus
accepting the data for further analysis (Jaccard and Becker
2002) (data are available on request). below 0.05. Hence, a further analysis of Q-Q plots is used to check
the normality of the data distribution by a visual inspection of the
line that the data make with the theoretically ideal normal
distribution. These plots show straight upward lines, thus
accepting the data for further analysis (Jaccard and Becker
2002) (data are available on request). below 0.05. Hence, a further analysis of Q-Q plots is used to check
the normality of the data distribution by a visual inspection of the
line that the data make with the theoretically ideal normal
distribution. These plots show straight upward lines, thus
accepting the data for further analysis (Jaccard and Becker
2002) (data are available on request). May 2021 | Volume 4 | Article 622106 Data Collection (2-tailed)
Mean difference
95% confidence interval of the difference
−6.872
92
0.000
−1.892
Lower −2.44
Upper −1.35
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of
difference
Equal variance
F
Sig. T
df
Sig. (2-tailed)
Mean difference
Std. error, difference
Lower
Upper
Assumed
2.421
0.123
1.498
91
0.138
0.820
0.547
−0.267
1.906
Not assumed
1.497
90.667
0.138
0.820
0.547
−0.268
1.907
Frontiers in Big Data | www.frontiersin.org
May 2021 | Volume 4 | Article 622106
6 Levene’s test for equality of variances T 1.48 population is normally distributed, which is rejected whenever
the p-value falls below a certain alpha value, in our case when the
p-value is lower than 0.05. For all four variables, the null
hypothesis of normal distribution is rejected as the p-value is distributed and therefore for all four variables (“Jobs Google,”
“Politics
Google,”
“Jobs
DDG,”
and
“Politics
DDG”),
a
Shapiro–Wilk test has been conducted to test for normality. The Shapiro–Wilk test uses the null hypothesis that the TABLE 3 | SPSS output for “How to become involved in politics Google.”
Sample statistics for “How to become involved in politics Google”
Mean
Std. deviation
Std. error mean
N
93
−1.89
2.656
0.275
Male
46
−1.48
2.689
0.397
Female
47
−2.30
2.587
0.377
One-sample test
T
df
Sig. (2-tailed)
Mean difference
95% confidence interval of the difference
−6.872
92
0.000
−1.892
Lower −2.44
Upper −1.35
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of
difference
Equal variance
F
Sig. T
df
Sig. (2-tailed)
Mean difference
Std. error, difference
Lower
Upper
Assumed
2.421
0.123
1.498
91
0.138
0.820
0.547
−0.267
1.906
Not assumed
1.497
90.667
0.138
0.820
0.547
−0.268
1.907
Frontiers in Big Data | www.frontiersin.org
May 2021 | Volume 4 | Article 622106
6 TABLE 3 | SPSS output for “How to become involved in politics Google.” Levene’s test for equality of variances 6 Search Engine Bias Wijnhoven and van Haren below 0.05. Hence, a further analysis of Q-Q plots is used to check
the normality of the data distribution by a visual inspection of the
line that the data make with the theoretically ideal normal
distribution. These plots show straight upward lines, thus
accepting the data for further analysis (Jaccard and Becker
2002) (data are available on request). Data Collection Reliability
The two persons who helped in the data collection independently
performed the coding of the screenshots after discussing
the coding scheme of Supplementary Appendix Tables S1, S2
with the first author. For testing the interrater reliability of the
coding, we used Krippendorff’s alpha (Hayes and Krippendorff
TABLE 4 | Detailed SPSS output for “Job openings near me DDG.”
One-sample statistics for “Job openings near me DDG”
N
Mean
Std. Deviation
Std. Error mean
N
94
−0.26
0.879
0.091
Male
50
−0.26
0.899
0.127
Female
44
−0.25
0.866
0.131
One-sample test
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
−2.816
93
0.006
−0.255
−0.44
−0.08
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
0.006
0.939
−0.055
92
0.956
−0.010
0.183
−0.373
0.353
Not assumed
−0.055
91.232
0.956
−0.010
0.182
−0.372
0.352
TABLE 5 | Detailed SPSS output for “How to become involved in politics DDG.”
One-sample statistics “How to become involved in politics” DDG
N
Mean
Std. deviation
Std. error mean
N
91
0.03
0.314
0.033
Male
48
0.00
0.000
0.000
Female
43
0.07
0.457
0.070
One-sample test
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
1.000
90
0.320
0.033
−0.03
0.10
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
4.692
0.033
−1.057
89
0.293
−0.070
0.066
−0.201
0.061
Not assumed
−1 000
42 000
0 323
−0 70
0 070
−0 211
0 071
Wijnhoven and van Haren
Search Engine Bias b l
f
h
l
f
l
d
h
k
R li bili
TABLE 4 | Detailed SPSS output for “Job openings near me DDG.”
One-sample statistics for “Job openings near me DDG”
N
Mean
Std. Deviation
Std. Error mean
N
94
−0.26
0.879
0.091
Male
50
−0.26
0.899
0.127
Female
44
−0.25
0.866
0.131
One-sample test
T
Df
Sig. Hypothesis H1: Women are mainly shown vacancies for female related jobs in their GSE search
results H1: Women are mainly shown vacancies for female related jobs in their GSE search
results
H2: Men are mainly shown vacancies for male related jobs in their GSE search results
H3: Women are mainly shown political involvement opportunities for positions of low
power and influence in their GSE results
H4: Men are mainly shown political involvement opportunities for positions of high
power and influence in their GSE results
H5: There is no gender bias in the vacancies shown on DDG to men and women True. The bias toward female-oriented jobs is 1.36. This bias is significantly larger than
for men Not true. Men also receive a bias (0.98) toward female-oriented jobs
Not true. On the contrary women are presented more powerful opportunities than
men, but the difference between men and women is not statistically significant
True True, no significant difference between men and women, but a bias toward male-
dominated vacancies for both men and women
T
i
ifi
diff
b
d H6: There is no gender bias in the political involvement opportunities shown on
DuckDuckGo to men and women True, no significant difference between men and women 2007). Krippendorff’s alpha is a statistical measure that displays
the extent of agreement between coders on the values given to a
variable on the basis of a coding scheme. This test is suitable for
ratio data which we collected for this study. With the scores
assigned to the screenshots a meaning is given to the data
ranging from −6 (fully male-biased) to 6 (fully female-
biased), the intervals are of equal distance and there is an
absolute zero point as assigning a zero means that there is
no bias. Before checking for reliability, the results of coding for
GSE and DDG have been combined per variable. This has been
done because the same coding scheme has been used for “Jobs
Google” and “Jobs DDG” as well as for “Politics Google” and
“Politics DDG.” For both categories, Krippendorff’s alpha were
higher than 0.67, namely 0.7682 for “Jobs” and 0.7725 for
“Politics,” which means that the coding schemes for both
political participation and jobs were sufficiently reliable
(Hayes and Krippendorff 2007). indeed significantly different than zero (one-sample t test
3.951, df 50, p < 0.000). RESULTS We present the results for the GSE and DDG jobs and political
participation queries per hypothesis in the following subsections. Hypothesis This means that they are shown one-
sided views on the search query “job openings near me”;
however, it was not in the expected direction. Therefore, the
second hypothesis is not supported. The SPSS output for these
tests is given in Table 2. Political Participation GSE (H3 and H4) Starting with a one-sample t test to determine if there is a bias
to be found in the search results for the query “how to become
involved in politics,” the test gives that the results differ
significantly from zero (one-sample t test −6.872, df 92,
p < 0.000). The general bias is in the direction of political
positions with high power and influence, as the mean is −1.89. Hypothesis 3 states that women would have significantly
higher scores than men and that women would have a
positive score. This hypothesis is rejected because women
have a mean score −2.30, even higher than men who have
an average score of −1.48. Clearly, men have a negative score,
as expected, and hypothesis 4 is not rejected, but the t test has
no
significant
difference
between
men
and
women
(independent-samples t test 1.498, df 91, p 0.069). The SPSS output is given in Table 3. Jobs DuckDuckGo (H5) (
)
A one-sample t test is used to test if there is no gender bias in the
search results of DDG, thus the average score should not differ
significantly from zero. The one-sample t test shows a significant
difference from zero, which means that there is a general bias in
the search results (one-sample t test −2.816, df 93, p 0.006). When looking at the average score of this sample, it shows that
the direction of this bias is toward male-dominated vacancies, as
the mean is −0.26. The test does not show significantly different
results between men and women and therefore supports
hypothesis 5 (independent-samples t test −0.055, df 92,
p 0.478). The SPSS output is given in Table 4. A one-sample t test has been conducted to test if there is a bias
toward either side. The results differ significantly from zero
(one-sample t test 6.286, df 96, p < 0.000), meaning that the
search results for “job openings near me” are biased toward
one side as zero means neutral results. This bias is in the
direction of female-dominated jobs as the average score of the
screenshots is 1.36. With the coding scheme used in this study, the hypothesis is
that women will have a significantly higher score than men. An
independent-samples t test supports this first hypothesis:
“women are mainly shown vacancies for female-related jobs
in their GSE search results” (independent-samples t test
−1.845, df 81.795, p 0.0345). Surprisingly, the mean score
of men is 0.98, which also implies that on average men were
shown
more
female-dominated
jobs
instead
of
male-
dominated jobs. Before being able to say with evidence that
the second hypothesis is not supported, a one-sample t test
only for men has been conducted. The results from men are Reliability
Th t y
The two persons who helped in the data collection independently
performed the coding of the screenshots after discussing
the coding scheme of Supplementary Appendix Tables S1, S2
with the first author. For testing the interrater reliability of the
coding, we used Krippendorff’s alpha (Hayes and Krippendorff TABLE 5 | Detailed SPSS output for “How to become involved in politics DDG.”
One-sample statistics “How to become involved in politics” DDG
N
Mean
Std. deviation
Std. error mean
N
91
0.03
0.314
0.033
Male
48
0.00
0.000
0.000
Female
43
0.07
0.457
0.070
One-sample test
T
Df
Sig. (2-tailed)
Mean difference
95% confidence interval of the
difference
Lower
Upper
1.000
90
0.320
0.033
−0.03
0.10
Independent-samples test
Levene’s test for equality of variances
t test for equality of means
95% confidence
interval of the
difference
Equal variance
F
Sig. t
Df
Sig. (2-tailed)
Mean difference
Std. error difference
Lower
Upper
Assumed
4.692
0.033
−1.057
89
0.293
−0.070
0.066
−0.201
0.061
Not assumed
−1.000
42.000
0.323
−0.70
0.070
−0.211
0.071
Frontiers in Big Data | www.frontiersin.org
May 2021 | Volume 4 | Article 622106
7 TABLE 5 | Detailed SPSS output for “How to become involved in politics DDG.” Levene’s test for equality of variances May 2021 | Volume 4 | Article 622106 7 Search Engine Bias Wijnhoven and van Haren Frontiers in Big Data | www.frontiersin.org DISCUSSION This study tried to answer the question to what extent GSE biases its
users by showing one-sided search results with possible virtue ethical
gender implications. Overall, GSE gives a bias, but not necessarily
against a stronger female participation in more powerful jobs and
political participation opportunities. As far as GSE could nudge people
toward certain choices and behavior, it also can nudge people toward
virtues by showing opportunities of emancipation. A nonprofile aware
search engine like DDG may be nonbiased from a profile, in our case,
gender perspective, but is, therefore, not free from virtue ethical
implications. Our study shows that DDG gives male-dominant job
outcomes, and this may have virtue ethical implications if this results in
a larger attractiveness or social value for such jobs, and thus an
undervaluation of female-dominant jobs. We do not want to say
that DDG bears any responsibility for this outcome, because profile-
free search engines may just give a representation of a biased society or
information market. Therefore, it is important that Internet users know
of the existence of such a bias and the risk that it can bring (Epstein
et al., 2017). Epstein et al. (2017) state that a bias awareness tool may
help search engine users to decide for themselves what they want to
accept as valid, credible, or just, but developing a critical view on search
engine outcomes is often difficult (Lazer et al., 2018; Flanagin et al.,
2014). Nobias.com iscurrently developingasa bias-alertingtool.Search
engine services could also develop dialectic search outcomes, with
which we mean a search engine that gives contrasting search outcomes
and alerts users to information sources with alternative views on a
subject (Wijnhoven and Brinkhuis 2015). Regarding the political participation results for Google, the
result of the tests does not support the hypothesis that women
would be given higher page ranks for less powerful activities. The
general bias toward more powerful positions could support the
view that the supply side of content on the Web—due to its
structure and presentation—is in itself male-biased (Hargittai and
Shafer 2006). DDG shows different patterns as for jobs, there is a
slight but significant bias to more male-oriented job outcomes
and for political participation no bias toward more or less
powerful activities exists. For both queries, the differences
between men and women are not significant, as expected. Political Participation DDG (H6) Hypothesis 6 states the absence of DDG gender bias in its search
results for men and women regarding political participation. The
scores given to the screenshots give no significant difference from
zero (one-sample t test 1.000, df 90, p 0.320). An
independent-samples t test of the average scores of men and May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 8 Wijnhoven and van Haren Search Engine Bias is needed for more clearly concluding about its implications. We also have not separated participants in those who search
via a GSE account (i.e., explicit profile) and those who do not. Because GSE will likely know more about the first group, the
filter bubble effect may be greater for them than for those who
search without an account. A third limitation is the queries we
use. The number of possible queries is of course unlimited and
as researchers we needed to focus. We selected only two topics
that are highly relevant for students searching for a social
position and have been highly debated in the literature on
women equality. We leave it to the readers to redo our study
for other topics. Finally, our study is limited to only two search
engines. The difference between GSE and DDG, however, is
theoretically and practically relevant and found back in the
evidence of this study. However, we cannot generalize over all
search engines with personalization or not. Comparative
studies
of
search
engines
remain
important
from
the
perspective of knowing their biases and how these biases
could be canceled out via searches with multiple search
engines (Jansen and Spink 2006; Nguyen et al., 2012). women gives no significant difference between the results shown
to men and women (independent-samples t test −1.000, df
42.000, p 0.1615), which supports hypothesis 6. The SPSS
output is given in Table 5. DISCUSSION For
DDG, the bias found is thus independent of the gender of its users
and only for job opportunities. This bias thus is not caused by
profile processing but possibly by indexing algorithms and the
effects of supply and demand of content. Conclusion We summarize our conclusions regarding the hypotheses in
Table 6. In this study, we find that men have higher page ranks for
female-dominated jobs as GSE query outcomes. This is not in line
with our expectations. One explanation for this is that GSE applied
student (our sample) profile data and with that the page ranked
flexible jobs higher. Student employment includes any form of paid
work during the academic year or during summer, but especially
flexible jobs (Baert et al., 2016). As in this study, flexible work has
been seen as an aspect of a female job following the UNDP, United
Nations Europe and Central Asia, 2018; this may give male users
higher scores for female-stereotyped jobs. Moreover, as this study
has been conducted around the time that people were searching for
summer jobs, GSE may even generate more outcomes toward
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Female Personal Perspectives. Comput. Hum. Behav. 24 (5), 2005–2013. doi:10. 1016/j.chb.2007.09.002 Strategy 25 (1), 6–31. doi:10.1111/jems.12141 Bettio, F., and Verashchagina, A. (2009). Gender Segregation in the Labour Market:
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(3), 211–224. doi:10.1108/DPRG-05-2017-0020 Bozdag, E. (2013). Bias in Algorithmic Filtering and Personalization. Ethics Inf. Technol. 15 (3), 209–227. doi:10.1007/s10676-013-9321-6 Defloor, B., Van Ootegem, L., and Verhofstadt, E. (2015). A Good or Bad
Transition from School to Work: Who Is Responsible? Int. J. Manpower 36
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(1), 103–125. doi:10.1177/0894439315605806 Epstein, R., Robertson, R. E., Lazer, D., and Wilson, C. (2017). Limitations Our study has a few important limitations that need further
research. One limitation is our sample. Although the sample
size as a whole and for the subsamples of men and women is
above 40 and allows the tests we did, the sample is restricted to
students from Western countries and even with mainly Dutch
and German origins. Samples including other age-groups,
different occupations, and different regions could result in
alternative insights. For example, the differences between men
and women in Asian societies are stated to be larger than in
Western societies (Hofstede et al., 2010). A second limitation is
our operationalization of GSE’s profile. GSE is not transparent
to us regarding the content of the profiles they use, but search
profiles include more than gender indicators alone (Pan et al.,
2007; Scheitle 2011; Kosinski et al., 2013). Consequently, the
actual bias is not only a filter on gender. A further study of
other profile characteristics and their search engine bias effects GSE and DDG both can deliver biased search results, but no
evidence in our study is found of a filter bubble, and thus we need a
more complete overview of possible sources of search engine bias. For
such an analysis, one may go for a full analysis of the search engine
value chain (Andersen 2018), starting with publishing biases on the
Internet, that is, different volumes of certain content available on the
Internet and thus a higher chance of being retrieved. Search engines May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 9 Wijnhoven and van Haren Search Engine Bias DATA AVAILABILITY STATEMENT Other biases further on the road to the
information end consumer are biases in query composition based
on the user’s world view (language and interests). Possible query
support tools could help users avoiding unwanted biases or to be
aware of their search biases. The algorithm by which a search engine
matches queries with documents is the search matching bias. When
applying personal profiling, profile biases may include the search
engine’s understanding of the user’s subjective perceived need, but
search engines also may have classification biases related to their
understanding of documents and actual needs. Finally, also browsers
may present search outcomes in different ways and users may
interpret outcomes in different ways. To summarize, we therefore
identify the following search engine biases: 1) publishing biases, 2)
crawling and indexing biases, 3) query biases, 4) search support biases,
5) algorithm biases, 6) profile biases, 7) content matching biases, 8)
content classification biases, 9) browser presentation biases, and 10)
interpretation biases. This study only uncovered some part of possible
profile biases and more work is to be done to gain a fuller
understanding of search engine biases. Search engine firms may
have a tendency to optimize profile biases for creating a highest
subjective satisfaction of their users, thus optimizing utility ethics, but
this can be at the cost of virtue ethics, that is, understanding and
opportunities of voluntary acting (Stucke and Ezrachi 2016; Gal et al.,
2020). In contrast, search engine services should be free from the need
to optimize user subjective satisfaction, and instead search engine
services should be developed that can be free from commercial or
political funding, serving the users’ virtues of personal skills, wisdom,
and voluntary action (Fuchs 2011; Zuboff 2019). The raw data supporting the conclusions of this article will be
made available by the authors, without undue reservation. ETHICS STATEMENT The studies involving human participants were reviewed and
approved by the University of Twente, faculty of Behavioral,
Management
and
Social
Sciences. The
patients/participants
provided their written informed consent to participate in this study. ACKNOWLEDGMENTS We thanks Rebecca Schaefer for her support in the data collection
and data coding. AUTHOR CONTRIBUTIONS FW designed the study, analyzed the data, wrote the article and
communicated with reviewers and the publisher. JH co-designed
the study, collected the data and did a first raw analysis. SUPPLEMENTARY MATERIAL The Supplementary Material for this article can be found online at:
https://www.frontiersin.org/articles/10.3389/fdata.2021.622106/
full#supplementary-material DATA AVAILABILITY STATEMENT use crawlers to index this content in which they necessarily have to be
selective because of the excessive size of the Internet. In this selection
process, crawlers are, therefore, biased to certain criteriaand the setting
of these selection criteria may help avoiding an unethical bias in the
search engine’s index. Other biases further on the road to the
information end consumer are biases in query composition based
on the user’s world view (language and interests). Possible query
support tools could help users avoiding unwanted biases or to be
aware of their search biases. The algorithm by which a search engine
matches queries with documents is the search matching bias. When
applying personal profiling, profile biases may include the search
engine’s understanding of the user’s subjective perceived need, but
search engines also may have classification biases related to their
understanding of documents and actual needs. Finally, also browsers
may present search outcomes in different ways and users may
interpret outcomes in different ways. To summarize, we therefore
identify the following search engine biases: 1) publishing biases, 2)
crawling and indexing biases, 3) query biases, 4) search support biases,
5) algorithm biases, 6) profile biases, 7) content matching biases, 8)
content classification biases, 9) browser presentation biases, and 10)
interpretation biases. This study only uncovered some part of possible
profile biases and more work is to be done to gain a fuller
understanding of search engine biases. Search engine firms may
have a tendency to optimize profile biases for creating a highest
subjective satisfaction of their users, thus optimizing utility ethics, but
this can be at the cost of virtue ethics, that is, understanding and
opportunities of voluntary acting (Stucke and Ezrachi 2016; Gal et al.,
2020). In contrast, search engine services should be free from the need
to optimize user subjective satisfaction, and instead search engine
services should be developed that can be free from commercial or
political funding, serving the users’ virtues of personal skills, wisdom,
and voluntary action (Fuchs 2011; Zuboff 2019). use crawlers to index this content in which they necessarily have to be
selective because of the excessive size of the Internet. In this selection
process, crawlers are, therefore, biased to certain criteriaand the setting
of these selection criteria may help avoiding an unethical bias in the
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abstract, or link of the search result that relates to a
part-time job, insecure contract, and/or the following
female dominated occupations: • When there is a vacancy mentioned in either the header,
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absence of any commercial or financial relationships that could be construed as a
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Greenhill, F. M., et al. (2018). The Science of Fake News. Science 359 (6380),
1094–1096. doi:10.1126/science.aao2998 Copyright © 2021 Wijnhoven and van Haren. This is an open-access article
distributed under the terms of the Creative Commons Attribution License (CC
BY). The use, distribution or reproduction in other forums is permitted, provided the
original author(s) and the copyright owner(s) are credited and that the original
publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with
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On Computer Science & Software Engineering - C3S2E ’16. Editors E. Desai. New York, New York, USA: ACM Press, 131–132. May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 11 Search Engine Bias Wijnhoven and van Haren of search result First result mentions male dominated occupation(s)
2nd result mentions male dominated occupation(s)
3rd result mentions male dominated occupation(s)
4th result mentions male dominated occupation(s)
First result mentions female dominated occupation(s)
2nd result mentions female dominated occupation(s)
3rd result mentions female dominated occupation(s)
4th result mentions female dominated occupation(s)
When no occupations are mentioned in the results When is it one-sided for men? When is it one-sided for men? • When there is a vacancy mentioned in either the header,
abstract, or link of the search result that relates to a
full-time job, and/or the following male dominated
occupations SpargWhen is it one-sided for men? • Armed forces • Armed forces • Legislators, senior officials, and managers
• Corporate managers • When the search result mention in either the header,
abstract, or link, relate to high power and influence
positions regarding politics like:
• Regarding economics
• Foreign and internal affairs
• Defense and justice
• Direct involvement (e.g., run for office, get in touch with
politicians)
• As well as if somewhere it mentions explicitly male/man etc. • Physical,
mathematical,
and
engineering
science
professionals • Physical and engineering science associate professionals
• Skilled agricultural and fishery workers
• Extraction and building trades workers
• Metal, machinery, and related trades workers
• Stationary-plant and related operators
• Drivers and mobile plant operators
• Laborers in mining, construction, manufacturing, and • Laborers in mining, construction, manufacturing, and
transport Content of search result
Code
First result mentions high power and influence position(s)
-2
2nd result mentions high power and influence position(s)
-2
3rd result mentions high power and influence position(s)
-1
4th result mentions high power and influence position(s)
-1
First result mentions low power and influence position(s)
2
2nd result mentions low power and influence position(s)
2
3rd result mentions low power and influence position(s)
1
4th result mentions low power and influence position(s)
1
No particular position(s) of power and influence are mentioned
0 APPENDIX 2: CODING SCHEME FOR
POLITICAL PARTICIPATION. • Customer services clerks • Personal and protective services (e.g., security) workers
• Models, salespersons, and demonstrators When is it one-sided for women? • Sales and services elementary occupations • When the search result mention in either the header,
abstract, or link, relate to examples of low power and
influence positions regarding politics, like: • Furthermore, when it is explicitly mentioned that the
vacancy is for a woman than it is also showing a one-
sided result. • Social and cultural activities Content of search result • Furthermore, when it is explicitly mentioned that the
vacancy is for a man than it is also showing a one-sided
result. ! If a part time or full-time job is mentioned in the results
as well as occupations, occupations are determining the
bias,
however
if
for
example
two
female
dominated
occupations are mentioned and one male + full-time then
neutral! ! If the occupation(s) does not relate to some in the above lists
than it’s neutral! ! If the occupation(s) does not relate to some in the above lists
than it’s neutral! Coding system: ! When there are more occupations mentioned (for
example two relate to male and two to female), then also
neutral (0)! ! If the position(s) does not relate to some in the above lists than it’s
neutral! ! When there are more positions mentioned (for example two
relate to male and two to female), then also neutral (0)! ! When the screenshot cannot be used, use code 999! Coding system: ! When the screenshot cannot be used, use code 999! Coding system: May 2021 | Volume 4 | Article 622106 Frontiers in Big Data | www.frontiersin.org 12
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Necessity of oral health intervention in schizophrenic patients – A review
|
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Review Article Open Access Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP.
www.nepjol.info/index.php/NJE Abstract: Individuals with mental illness often cannot perform day to day activities due to a psychiatric or emotional
disorder. Schizophrenia is one such psychiatric disorder characterized by worsening self-care ability with
progressing mental illness. This disease may potentially deteriorate oral health by affecting the subject's ability
to perform oral hygiene measures. Literature on oral disease manifestations in schizophrenia is limited. Lack of
desire for oral health care as well as generally poor awareness of oral health issues in these patients, compounded
further by side effects of medications, may complicate dental management in schizophrenic patients. The present
review explores clinical features and possible factors associated with oral health status among those with
Schizophrenia. Keyword: Oral health; Schizophrenia; Treatment awareness. Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP. www.nepjol.info/index.php/NJE
Correspondence: Dr. Pratibha PK, Head and Professor, Department of Periodontology, Manipal College of
dental sciences, Manipal, Karnataka, India
Email: bg_pratibha@yahoo.co.in
Received 1 November 2016/Revised 28 December 2016/Accepted 30 December 2016
Citation:
Gupta S, Pratibha PK, Gupta R. Necessity of oral health intervention in schizophrenic patients – A review. Nepal J
Epidemiol. 2016;6(4); 605-612. This work is licensed under a Creative Commons Attribution 4.0 International License. Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP. www.nepjol.info/index.php/NJE
Correspondence: Dr. Pratibha PK, Head and Professor, Department of Periodontology, Manipal College of
dental sciences, Manipal, Karnataka, India
Email: bg_pratibha@yahoo.co.in
Received 1 November 2016/Revised 28 December 2016/Accepted 30 December 2016
Citation:
Gupta S, Pratibha PK, Gupta R. Necessity of oral health intervention in schizophrenic patients – A review. Nepal J
Epidemiol. 2016;6(4); 605-612. This work is licensed under a Creative Commons Attribution 4.0 International License. Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP. www.nepjol.info/index.php/NJE
Correspondence: Dr. Pratibha PK, Head and Professor, Department of Periodontology, Manipal College of
dental sciences, Manipal, Karnataka, India
Email: bg_pratibha@yahoo.co.in
Received 1 November 2016/Revised 28 December 2016/Accepted 30 December 2016
Citation:
Gupta S, Pratibha PK, Gupta R. Necessity of oral health intervention in schizophrenic patients – A review. Nepal J
Epidemiol. 2016;6(4); 605-612. This work is licensed under a Creative Commons Attribution 4.0 International License. Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP. www.nepjol.info/index.php/NJE
Correspondence: Dr. Pratibha PK, Head and Professor, Department of Periodontology, Manipal College of
dental sciences, Manipal, Karnataka, India
Email: bg_pratibha@yahoo.co.in
Received 1 November 2016/Revised 28 December 2016/Accepted 30 December 2016
Citation:
Gupta S, Pratibha PK, Gupta R. Necessity of oral health intervention in schizophrenic patients – A review. Nepal Journal of Epidemiology
R
i
A ti l
eISSN 2091-0800 eISSN 2091-0800 Open Access Open Access Nepal Journal of Epidemiology
eISSN 2091-0800 Introduction Diseases affecting the oral cavity are frequently neglected or
under diagnosed; having a great impact on an individual’s
health in the long term [1]. Therefore, maintenance of oral
health becomes imperative for one’s general wellbeing. Moreover, a healthy dentition is increasingly being perceived as
important for self-image and confidence in affairs of routine life
[2]. Poor oral health status in schizophrenics has been attributed to
factors like poverty, less resources, imbalance of mental status,
medications and most of all the effect of systemic diseases like
diabetes, hypertension, osteoporosis which reduce the self-care
ability of the patients [12]. A study in institutionalized patients
found the oral hygiene status to be poorer in schizophrenic
patients [13]. It has also been suggested that oral health
deteriorates with increasing age, length of hospitalization and
has been found to be worse amongst inpatients as compared to
outpatients [14]. Oro systemic health interrelationship becomes ever more
relevant in cases of mental illness. Oral health care is of
particular concern in mentally challenged and institutionalized
patients, as poor oral hygiene not only socially excludes these
patients but also predisposes them to various other systemic
health problems like respiratory infections and cardiac diseases. Insufficient institutionalized care by caretakers augments the
problem. Measures to institute plaque control in conditions such
as Down’s syndrome / retardation in children are still
manageable to an extent through concerted efforts on the part of
the parents, nurses and dental professionals. However, certain
psychiatric disorders such as schizophrenia warrant special
attention as the health condition is relatively difficult to identify,
diagnose and manage; compounding the dental problems
further. Higher rates of caries and multiple missing teeth compared to
healthy subjects, has been reported in patients with mental
illness [15]. A survey was conducted on 565 institutionalized
patients, out of which 62% were diagnosed with schizophrenia. Higher DMFT scores were observed in female patients and
those with dementia. It was concluded from the survey that
decayed-missing-filled teeth (DMFT) score increased with age
[16]. Socioeconomic status is another factor that poses a risk for
schizophrenia development. Demanding household and harsh
community environments may be responsible for poor access to
social services like health care and education, often leading
them to social exclusion [17]. Hence, those in the lower socio
economic classes may show many more teeth extracted,
decayed, and filled, than those from a higher class[18]. Abstract: Nepal J
Epidemiol. 2016;6(4); 605-612. This work is licensed under a Creative Commons Attribution 4.0 International License. ; ( );
k is licensed under a Creative Commons Attribution 4.0 International License. Copyright © 2016 CEA& INEA. Published online by NepJOL-INASP. www.nepjol.info/index.php/NJE 605 Genetics is considered to be a major reason, amongst many
etiological factors causing schizophrenia. Rare genetic
mutations and other early influences, such as gestational and
birth complications are also thought to be involved in the causal
pathways for schizophrenia [8]. Changes in levels of
neurotransmitters, dopamine and serotonin, in the brain has
been suggested to be a probable mechanism for this condition
[9]. Social isolation, hostile family atmosphere and drug abuse
may be other reasons predisposing to this condition [10, 11]. Introduction Bizarre behaviour resulting in self-mutilation in the form of
autoextractions [24], glossectomy [25] and excoriation of
gingival tissues with sharp fingernails [26] has also been
reported. behavior assessment in 372 psychiatric inpatients showed that
these patients have less impetus to visit a dentist. They brushed
their teeth for shorter periods and did not realize that oral health
may have an influence on their general health condition [34]. Attitude of dental professionals and limited knowledge of
causes and effects of oral diseases in this special group of
patients, are other issues resulting in delay in providing
treatment. Instances of low tolerance on the part of the dental
staff in dealing with patients with low compliance, and
unwillingness to treat such patients has also been reported [36]. Due consideration to these factors by dental professionals and
modification of treatment plans to specifically cater to these
patients will motivate them to accept dental treatment and also
reduce the number of dental visits [37]. Over 60% of schizophrenics are found to be heavy smokers
[27]. In an institutional study, the smoking prevalence was more
in psychiatric outpatients as compared to local or national
population-based samples (52% versus 30% and 33%). Factors
like age, sex, marital status, socioeconomic status, alcohol use,
coffee use, or institutionalization of the psychiatric patients had
no bearing on the increased prevalence rate. Schizophrenics
(88%), Maniacs (70%) and severely ill patients were found to
be the heavy abusers [28]. The reasons for such high frequency
of heavy smoking prevalent in schizophrenic patients is thought
to be partially related to its enhanced stimulant effect. Smoking
induces release of dopamine in the brain, stimulating its activity
and inhibiting its degradation [29]. While managing patients with this chronic illness, a dentist
should take utmost care of the oral hygiene maintenance. A
thorough history regarding the patient’s oral hygiene practices,
tobacco consumption, parafunctional and other abusive habits,
medications as well as attitude of the patient’s family towards
oral health should be taken into consideration. The patient
should be dealt with empathetically and treated like any other
patient. Good communication and rapport should be built to
avoid any discomfort to the patient. Priority should be given to
counseling of the care takers, oral prophylaxis, restoration of
decayed teeth, oral rehabilitation of edentulous patients and any
emergency treatment possible. Cigarette smoking in these patients may predispose them to
periodontal disease and oral cancer. Introduction A cross-sectional survey on
250 schizophrenic patients investigated the relationship
between periodontal disease and schizophrenia. An increased
gingival and plaque index, higher probing depths in chronic
schizophrenic patients (P < 0.001) pointed towards poor
periodontal conditions in these patients [30]. Smoking is one of
the contributory factors to poor periodontal health, which is
characterized
by
increased
pocket
depth,
periodontal
attachment loss leading to increased incidence of tooth loss
[29]. The prevalence of salivary P. gingivalis in schizophrenic
patients has been reported to many folds higher when compared
to non-psychiatric controls. In one study, a quantitative relation
was established between severity of schizophrenia and P. gingivalis cells [31]. Patient’s physician should be consulted for recall and further
maintenance of the patient’s oral health. Oral health education
programs can be organized for institutionalized patients to
interact with patients and educate them about good oral hygiene
practices. This would also meet their dental care needs and
provide easy access to these patients. The effective use of tooth
brushes, non-alcohol mouth washes can be introduced in the
daily regime of the patient with the coordination of the patient’s
care taker. Moreover, dentists can take up a more active role in
helping these patients with smoking cessation [38]. The chronicity of the illness has been attributed to negative
symptoms, which are potentially devastating to oral health as
they impair a patient's desire to maintain a good oral hygiene.14
Poor self-care, neglected oral care, low perception of dental
treatment needs, and poor diet may further contribute to oral
neglect in such patients. Therefore routine dental care becomes
a challenging task for the patient, caretaker, and the physician. Tremors have been observed as side effects of antipsychotic
medications [32]. A cross sectional study correlated the
association between tremors and poorer dental status. Tremors
interfere with day to day fine motor skills of the patient,
impairing one’s tooth brushing, hence again leading to poor oral
health [33]. The chronicity of the illness has been attributed to negative
symptoms, which are potentially devastating to oral health as
they impair a patient's desire to maintain a good oral hygiene.14
Poor self-care, neglected oral care, low perception of dental
treatment needs, and poor diet may further contribute to oral
neglect in such patients. Therefore routine dental care becomes
a challenging task for the patient, caretaker, and the physician. In patients with periodontal problems, routine oral prophylaxis
can be performed. Introduction Disturbances in thoughts, behavioral changes, impaired
cognitive functions which affect a person’s ability to work or
communicate socially and incapability of self-care characterize
schizophrenia [3].It is often associated with positive and
negative symptoms. Hallucinations, conversing voices with or
about the patient, and paranoid delusions are few of the positive
symptoms whereas ‘negative’ symptoms are flattened affect,
loss of a sense of pleasure leading to social withdrawal,
isolating the patient who eventually loses the will or drive to
perform routine activities [4]. Presently, the diagnostic criteria
put forward by the World Health Organization and the
American Psychiatric Association are widely used for the
diagnosis of schizophrenia. Few signs and symptoms
considered essential for confirming the diagnosis include
symptom duration of 6 months, delusions, hallucinations,
disorganized speech or behavior, affective flattening, alogia,
and avolition, social and occupational dysfunctions [5]. These
features make it difficult for the affected individual to carry out
daily activities and maintain personal hygiene. The incidence
rate of schizophrenia has been reported to be 7 per thousand of
the adult population, mostly in the age group of 15-35 years. Due to the chronic nature of the condition, it has a high
prevalence of about 24 million people worldwide [6]. An
estimated 85 percent of older adults with schizophrenia develop
their illness before age of 45 years, mostly in their second or
third decade of life [7]. These patients generally are prone to adverse orofacial effects
such as xerostomia, sialorrhea and oral dyskinesia caused by
psychotropic medications [19,3,20]. The first generation anti-
psychotics
(haloperidol,
chlorpromazine,
thioridazine,
trifluoperazine), used for schizophrenia treatment, are known to
cause hypo-salivation by blocking the parasympathetic
stimulation of the salivary glands, putting the patients at a high
caries risk, particularly root caries [21]. Due to xerostomia
various other lesions like candidiasis, glossitis, generalized
stomatitis set in, affecting speech and swallowing or leading to
poor denture tolerance, instability of dentures or denture
trauma. Dyskinesia and dystonia seen in these patients affect
motor activities producing abnormal jaw movements, giving
rise to presentations such as tongue protrusion, retraction and
facial grimacing [22]. Fear and anxiety in these patients may manifest as premature
tooth wear [23]. Most of the schizophrenic patients have some 606 kind of Para functional habit like nail biting, lip biting etc. Table I: Oral health status in schizophrenics Authors
Study design
Parameters
recorded
Sample size
Results
Conclusion
EltasA, Kartalc
S, EltasSD, Du
ndar
S, Uslu
MO(2013)
(40)
Cross-sectional study
Group A- medications
causing xerostomia
Group B- medications
causing sialorrhea
Plaque index (PI),
Bleeding
on
probing
(BoP),
Probing
pocket
depth (PPD) and
Clinical
attachment levels
(CAL),
DMFT
Group A-20
Group B- 20
PI and BOP higher in
group A
PPD,
CAL,
DMFT
similar in both groups
Higher
risk
of
periodontal disease
in patients with
schizophrenia
irrespective of the
medication. Persson
K, Axtelius
B, Soderfeldt
B, Ostman M
(2009)
Descriptive Population
surveys of outpatient
under psychiatric care. DMFT and oral
hygiene
correlated
with
age and no. of
antipsychotic
drugs
113
Oral health neglected in
men more than women
Regular
dental
check-ups should
be encouraged for
patients
under
psychiatric care
ArnaizA, Zuma
rragaM, Díez-
Altuna
I, Uriarte
JJ, Moro
J, Perez-
Ansorena MA
(2011)
Evaluate the oral health
of
a
group
of
schizophrenic
outpatients and a control
group
without
psychiatric illness. DMFT, CPITN,
PANSS index,
DM
index,
CPITN
higher
in
schizophrenics. Filled
teeth were lower than
controls. PANSS
negative
score
correlated
with
oral
health variables
Oral
health
in
psychiatric
patients is poorer
than
others
irrespective
of
habits and age. Introduction Surgical procedures like root planing and flap
surgeries are not a contraindication and can be performed with
precautions in patients who are stable and under medication. Health care professionals should be trained and updated
regarding the oral diseases, side effects of medications, and
possibility of dental treatment for these chronically ill patients. Schizophrenic patients should not be refused treatment on
grounds of inability to maintain oral hygiene. Tremors have been observed as side effects of antipsychotic
medications [32]. A cross sectional study correlated the
association between tremors and poorer dental status. Tremors
interfere with day to day fine motor skills of the patient,
impairing one’s tooth brushing, hence again leading to poor oral
health [33]. The reasons for infrequent dental visits in schizophrenics and
patients with mental illness have been observed as lack of drive
to maintain self-care, due to poor general health, inability to
meet cost of treatment and low priority for dental check-up,
unless, in case of emergency[34,35]. Dental care associated 607 Table I: Oral health status in schizophrenics Table I: Oral health status in schizophrenics Authors 608 Chu KY, Yang
NP, Chou P,
Chiu, Hsien J,
Chi LY (2010)
(14)
Cross-sectional survey
DMFT,
community
periodontal index
in schizophrenic
patients DMFT
index
was
14.3%;
5% were edentulous,
39.4% had (community
periodontal index ≥3)
Aging men with lower
educational levels and a
longer
stay
in
institutions were likely
to have lower filling rate
of the DMFT index
Long-term
care
institutions
that
care for inpatients
with schizophrenia
should
exert greater
efforts
in
providing
dental
care
Univariate
general
linear model older age,
smoking,
tremor
burden,
and
less
frequent tooth brushing
were associated with a
greater DMFT score. Schizophrenia
patients who do
not regularly brush
their teeth or who
exhibit tremor, it
may be advisable
for caregivers to
encourage
and 1103 (14) Tani H, Uchida
H, Suzuki T,
Shibuya
Y,
Shimanuki
H,
Watanabe
K(2012)
(35)
Cross-Sectional study
DMFT score: age,
sex,
smoking
status,
daily
intake of sweets,
dry
mouth,
frequency of daily
tooth
brushing,
tremor,
523 patients
in age group
of
40-60
years Tani H, Uchida
H, Suzuki T,
Shibuya
Y,
Shimanuki
H,
Watanabe
K(2012)
(35) Schizophrenia
patients who do
not regularly brush
their teeth or who
exhibit tremor, it
may be advisable
for caregivers to
encourage
and
help
them
to
perform
tooth
brushing
more
frequently. McCreadie
RG, Stevens
H, Henderson
J, Hall
D, McCaul
R, Filik
R
(2004)
(21)
Descriptive study
Self-report
questionnaire
428 people
with
schizophreni
a
in
six
different
areas of the
UK
More of the younger
patients were edentate
(3-39% vs. 1-20%) and
fewer had more than 20
teeth (70% vs. 83%). Fewer patients cleaned
their teeth daily; this
group
had
more
negative symptoms. Dental health of
people
with
schizophrenia
is
poor. Community
mental
health
teams
should
encourage them to
attend
their
community dentist
regularly. 609 4. Schultz SH, North SW, Shields CG. Schizophrenia: A
review. Am Fam Physician. 2007 Jun; 75 (12):1821-9. PMid:17619525 Hence, it becomes imperative on the part of the patient’s family
and other health care professionals to assimilate knowledge on
this medical condition and its management and also keep
themselves abreast with newer advances in the field. The
traditional view of assuming oral health care as secondary to
other aspects of systemic health must give way to the positive
impact of improved oral health care on general health. 5. American Psychiatric Association. Diagnostic and 5. American Psychiatric Association. Diagnostic and
Statistic Manual of Mental Disorders, 4th ed. Washington
DC: American Psychiatric Association, 1994. 5. American Psychiatric Association. Diagnostic and
Statistic Manual of Mental Disorders, 4th ed. Washington Statistic Manual of Mental Disorders, 4th ed. Washington Statistic Manual of Mental Disorders, 4th ed. Washington
DC: American Psychiatric Association, 1994. DC: American Psychiatric Association, 1994. 6. World
Health
Organisation
Health
Topics:
Schizophrenia [online]2013 [ cited 2011 July 10 ]
Available
from:
URL: To conclude, some of the clinical features of this illness are not
evident and therefore the dental practitioner must understand
the more common presentations and management considering
that poor dental condition in schizophrenics may be associated
with an increased risk of systemic comorbidity as well. Physicians have to work in coordination with dental
practitioners to maintain good oral health. Therefore,
knowledge and awareness of different aspects of this disease
will help dental professionals in diagnosis and planning of
better treatment strategies for this specific population. http://www.who.int/mental_health/management/schizophr
enia/en/WHO2013. 7. Jeste DV, Lanquette NM, Vahia IV. Schizophrenia and
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Psychiatry. 4th ed. p
g
j
PMid:17626963 PMCid:PMC1914490 Acknowledgements: Authors would like to acknowledge Dr. Nishanth (Assistant Professor, Department of Psychiatry) for
his support and help. 11. World Fellowship for Schizophrenia and Allied
Disorders [online] [accessed 2011 July 10]. Available
from:
http://www.world- Nishanth (Assistant Professor, Department of Psychiatry) for
his support and help. Conflict of interest: Conflict of interest: The authors hereby announce that they have no conflict of
interest arising from the study. p
g
p
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with
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Four Plasma Glucose and Insulin Responses to a 75 g OGTT in Healthy Young Japanese Women
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Journal of diabetes research
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cc-by
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Hindawi
Journal of Diabetes Research
Volume 2018, Article ID 5742497, 7 pages
https://doi.org/10.1155/2018/5742497 Hindawi
Journal of Diabetes Research
Volume 2018, Article ID 5742497, 7 pages
https://doi.org/10.1155/2018/5742497 Hindawi
Journal of Diabetes Research
Volume 2018, Article ID 5742497, 7 pages
https://doi.org/10.1155/2018/5742497 Kei Takahashi
,1 Hidetaka Nakamura,1 Hiroshi Sato,1 Hideto Matsuda,2 Kazuo Takada,1
and Tomiko Tsuji1 1Department of Health and Nutrition, Faculty of Health and Human Life, Nagoya Bunri University, 365, Maeda, Inazawa-Cho,
Inazawa City, Aichi, Japan Correspondence should be addressed to Kei Takahashi; takahashi.kei@nagoya-bunri.ac.jp Received 17 July 2017; Revised 12 October 2017; Accepted 24 October 2017; Published 30 January 2018 Academic Editor: Ed Randell Copyright © 2018 Kei Takahashi et al. This is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The incidence of diabetes has been gradually increasing, not only in middle-aged individuals but also in young individuals. However, insulin and glucose patterns have not been investigated in apparently healthy young individuals, as they are typically
grouped as controls. In this study, we investigated and classified glucose and insulin patterns in healthy young women. Sixty-
two nonobese women without metabolic disease were recruited. The subjects underwent a 75 g oral glucose tolerance test
(OGTT), physical measurements, and a biochemical examination. Two subjects displayed impaired glucose tolerance. The 62
subjects were categorized into four patterns by plasma glucose and insulin peak time during OGTT: normal type (n = 39),
insulin-late type (n = 11), insulin- and glucose-late type (n = 7), and insulin-very late type (n = 5). OGTT glucose and insulin
levels at all time points, insulinogenic index, HOMA-IR, and glucose area under the curve (AUC) significantly differed among
the four groups. However, insulin AUC did not significantly differ. We did not detect significant differences in body condition
or biochemical measurements. Our study demonstrated that some healthy young individuals might have delayed insulin
secretion by OGTT. Early detection of altered glucose metabolism might be helpful to improve lifestyle choices and prevent
progression to diabetes. 1. Introduction insulin resistant [7]. These reports indicate that postprandial
plasma glucose and insulin curves are important. Chronic diabetes mellitus is characterized by hyperglycemia,
which promotes oxidative stress and leads to microangiopa-
thy, such as retinopathy, nephropathy and neuropathy, and
macroangiopathy, such as ischemic heart disease and cere-
brovascular disease [1, 2]. Recently, it has been reported that
not only fasting plasma glucose levels but also postchallenge
glucose spikes are problematic in patients with diabetes [3, 4]. For example, cardiovascular disease risk in patients with
hyperglycemia 120 min after a meal was higher than in
patients with fasting hyperglycemia. Controlling postpran-
dial hyperglycemia is beneficial for prevention and reduction
of cardiovascular risks in patients with diabetes [5, 6]. Simi-
larly, patients with type 2 diabetes display delayed early insu-
lin secretion compared with that in healthy adults and are The incidence of diabetes in younger individuals is
gradually increasing [8]. However, there are few studies
investigating healthy young subjects. In Japanese universities,
most college students do not undergo medical examinations
and are not aware of their carbohydrate metabolic condi-
tions. In adults, fasting glucose levels are assessed during a
medical examination, but fasting insulin secretion and post-
challenge glucose levels are not. The Funagata Study [5], a
large Japanese epidemiology study, investigated subjects over
40 years of age but did not include younger individuals. Fur-
thermore, this study did not assess postchallenge insulin
levels. Therefore, we believed that it was necessary to assess
carbohydrate metabolism and prediabetes status in young
healthy individuals. Journal of Diabetes Research 2 Mitsui et al. [9] investigated the characteristics of diabetes
with isolated fasting hyperglycemia (DM/IFH), with isolated
postchallenge hyperglycemia (DM/IPH), and with both fast-
ing and postchallenge hyperglycemia (DM/FPH) in Japanese
men. Furthermore, in this study, the DM/IFH and DM/IPH
groups were subdivided into two groups and compared. In contrast, young healthy individuals were used as a con-
trol group and were not divided into subgroups or analyzed
in detail. In several studies investigating diabetes using 75 g
oral glucose tolerance tests (OGTTs) [7, 10, 11], healthy
individuals were the standard control group with which
individuals with diabetes and prediabetes were compared. However, type 2 diabetes in Asia is an increasing epidemic,
particularly in individuals at a relatively young age and
with a low BMI [8, 12]. Furthermore, Fukushima et al. 3. Results Two of 62 subjects (3.2%) displayed elevated 120 min
plasma glucose levels (160 and 187 mg/dL) following oral
glucose administration, despite fasting glucose levels below
110 mg/dL, and were, therefore, diagnosed with IGT. This
62 nondiabetic subjects were categorized into four groups
based on plasma insulin and glucose peak times, which
were the times with the highest insulin and glucose con-
centrations, respectively, during an OGTT (Figure 1). The four groups were as follows: the peak insulin and glu-
cose times both occurred at 30 min (normal type; n = 39
(62.9%), Figure 1(a)); insulin and glucose peak times were
60 min and 30 min, respectively (insulin- (I-) late type; n =
11 (17.7%), Figure 1(b)); both peak times occurred at
60 min (insulin- and glucose- (I&G-) late type; n = 7
(11.3%), Figure 1(c)); and insulin and glucose peak times
were 120 min and 30 min, respectively (I-very late type; n =
5 (8.1%), Figure 1(d)). In each group, insulin and plasma glu-
cose concentrations at the peak time were higher than those
at baseline (0 min). Sixty-two female college students were recruited for
this study from Nagoya Bunri University in Japan after
receiving an explanation of the study. Inclusion criteria for
the study were apparently healthy young adults between 19
and 23 years of age with no history of lifestyle-related disease
(e.g., diabetes, hypertension, or lipid abnormalities). g
y
The subjects received both physical and blood examina-
tions. Body weight, bone mass, lean body mass, and percent
body fat were measured by body composition analyzer
InBody 720 (InBody Japan Inc., Tokyo), and body mass
index (BMI) was calculated. Subjects underwent a 75 g
OGTT according to the recommendations of the World
Health Organization [14]. Venous blood samples were
obtained directly before (0 min) and during the OGTT (30,
60, and 120 min) for determination of plasma glucose and
insulin concentrations. Blood parameters were measured by
FALCO Biosystems Ltd. (Kyoto, Japan). Plasma glucose
and insulin concentrations were measured by the hexoki-
nase
glucose-6-phosphate
dehydrogenase
method
and
radioimmunoassay, respectively [15, 16]. Parameters related
to protein and lipid metabolism, liver and kidney function,
and blood characteristics were measured in blood collected
following an overnight fast to confirm the absence of dis-
ease. Furthermore, to evaluate carbohydrate metabolism, Table 1 displays the ages and body conditions based on
each OGTT response type. There were no significant
between-type differences. 2. Materials and Methods Ethical approval was obtained from the Research Ethics
Committee of the Nagoya Bunri University, and the study
was conducted according to the principles expressed in the
Declaration of Helsinki. Subjects were informed about the
experimental procedures and purpose of the study prior to
giving written consent. 1. Introduction [13] reported that impaired early-phase insulin secretion
played a very important role in the deterioration from nor-
mal glucose tolerance to impaired glucose tolerance (IGT) in
Japanese middle-aged individuals. Therefore, we thought it
necessary to show detailed insulin and glucose patterns in
healthy young women during a 75 g OGTT. we calculated homeostasis model assessment-insulin resis-
tance (HOMA-IR), homeostasis model assessment for β-cell
function (HOMA-β) [17], insulinogenic index (II) [18],
quantitative insulin sensitivity check index (QUICKI) [19],
and area under the curve of glucose (AUC-G) and insulin
(AUC-I). HOMA-IR, HOMA-β, II, and QUICKI were calcu-
lated using the following formulas: insulin0 ×glucose0/405,
insulin0 ×360/(glucose0 −63),
Δinsulin30/Δglucose30,
and
1/[log(insulin0) + log(glucose0)], respectively, with insulin
expressed in μU/mL and glucose in mg/dL. The diagnostic
criteria for diabetes from the Japanese Diabetes Society
[20] and the World Health Organization [14] were used. The normal type in criteria for diabetes was evaluated at
less than 110 mg/dL of fasting plasma glucose levels and
140 mg/dL of 120 min glucose levels. IGT was evaluated
at less than 110 mg/dL of fasting plasma glucose levels
and from 140 mg/dL to 199 mg/dL of 120 min glucose
levels. The diabetes was evaluated with fasting glucose
of 126 mg/dL and over or 120 min glucose of 200 mg/dL
and over. Glucose and insulin response types were determined
based on the time of peak insulin and plasma glucose con-
centrations during a 75g OGTT [21–24], and each param-
eter was compared. Values are expressed as means ±SD. All data analyses were conducted using Prism (version
5.01; obtained from GraphPad Software, San Diego, CA,
USA), and p values<0.05 were considered statistically sig-
nificant. For the statistical evaluation, the significance of
the differences in mean values between the four types were
tested by one-way ANOVA followed by Dunn’s multiple
comparison tests. In this study, we assessed prediabetes and diabetes by 75 g
OGTTs in healthy undergraduates in Japan and investigated
varying insulin and plasma glucose responses. In addition,
we compared physical and biochemical parameters in the dif-
ferent insulin and glucose response groups. 3. Results 0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
A
B
A
A
A
B
C
200
0
50
100
150
200
C
(a) A
A
B
B
A
B
C
A,B
0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(b) Immunoreactive insulin (휇U/mL) Plasma glucose (mg/dL) (a)
A
A
A,B
B
A
A,B
B
B
0 min
Plasma glucose (mg/dL)
30 min
l
(휇
)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(c) (a) (b) (b)
A
A,B
A,B
B
A
B
A,B
A,B
0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(d) Plasma glucose (mg/dL) (c) (d) Figure 1: Relationship between plasma glucose levels and insulin reactivity during a 75 g oral glucose tolerance test. Squares: insulin levels. Open circles: plasma glucose levels. (a) Normal type (n = 39). (b) Insulin-late type (n = 11). (c) Insulin- and glucose-late type (n = 7). (d)
Insulin-very late type (n = 5). Values are means ± SD. Values for each index with different superscript letters are statistically different
at p < 0 05 by comparing different time points of each patterns, as determined by Dunn’s multiple comparison test. Table 1: Body condition of healthy women by insulin and glucose response type. Type
Normal
Insulin-late
Insulin- & glucose-late
Insulin-very late
Number
39 (62.9%)
11 (17.7%)
7 (11.3%)
5 (8.1%)
Age (years)
20.6 ± 0.9
20.4 ± 1.0
19.9 ± 1.1
20.7 ± 0.5
Body weight (kg)
52.3 ± 6.9
53.1 ± 6.8
52.1 ± 6.2
53.3 ± 2.2
Height (cm)
158.0 ± 4.5
157.7 ± 5.1
160.9 ± 8.1
158.3 ± 1.7
Body mass index (kg/m2)
20.9 ± 2.3
21.4 ± 1.0
20.1 ± 1.4
21.3 ± 1.0
Bone mass (kg)
2.23 ± 0.21
2.25 ± 0.11
2.18 ± 0.31
2.23 ± 0.11
Lean body mass (kg)
37.2 ± 3.3
37.8 ± 1.9
36.4 ± 5.6
38.2 ± 1.6
Percent body fat (%)
27.0 ± 4.8
27.5 ± 4.5
28.6 ± 4.7
29.6 ± 3.7
Values are means ± SD. There were no significant differences. Table 1: Body condition of healthy women by insulin and glucose response type. 3. Results than those in the other groups. II values in the normal type
group was higher than those in the other type groups. AUC-G in the I&G-late and I-very late type groups were
higher than that in the normal type group, but there were
no significant differences in AUC-I among the four groups. four groups. The 60 min plasma glucose concentrations in
the I-late and I&G-late type groups were significantly higher
than those in the normal type group, but there were no differ-
ences in 120 min glucose levels. In I-very late type group, the
30 min, 60 min, and 120 min glucose levels were significantly
higher than those in normal type. In contrast, 0 min and
30 min immunoreactive insulin levels in the I&G-late type
group were lower than those in the normal and I-late type
groups. The 60 min insulin concentrations in the I-late type
group were higher than those in the normal type group. And the 120 min insulin in I-very late group was higher than
those in the normal. HOMA-IR and HOMA-β in the I&G-
late type group was lower while QUICKI values were higher g
g
g
p
Additional biochemical parameters are displayed in
Table 3. There were no significant differences in any param-
eter among types. 3. Results The comparisons of indexes of glucose metabolism in
each group are summarized in Table 2. There were no signif-
icant differences in plasma glucose levels at 0 min among the Journal of Diabetes Research 3 0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
A
B
A
A
A
B
C
200
0
50
100
150
200
C
(a)
A
A
B
B
A
B
C
A,B
0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(b)
A
A
A,B
B
A
A,B
B
B
0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(c)
A
A,B
A,B
B
A
B
A,B
A,B
0 min
Plasma glucose (mg/dL)
30 min
Immunoreactive insulin (휇U/mL)
60 min
120 min
0
50
100
150
200
0
50
100
150
200
(d)
Figure 1: Relationship between plasma glucose levels and insulin reactivity during a 75 g oral glucose tolerance test. Squares: insulin levels. Open circles: plasma glucose levels. (a) Normal type (n = 39). (b) Insulin-late type (n = 11). (c) Insulin- and glucose-late type (n = 7). (d)
Insulin-very late type (n = 5). Values are means ± SD. Values for each index with different superscript letters are statistically different
at p < 0 05 by comparing different time points of each patterns, as determined by Dunn’s multiple comparison test. 4. Discussion This study was performed in young healthy women without
lifestyle-related diseases or other diseases. They underwent 4 Journal of Diabetes Research Table 2: Glucose metabolism of women by insulin and glucose response type. Type
Normal
Insulin-late
Insulin- & glucose-late
Insulin-very late
Number
39
11
7
5
Plasma glucose (mg/dL)
0 min
83.4 ± 6.1
83.5 ± 4.5
83.0 ± 5.7
86.0 ± 3.0
30 min
117.0 ± 22.2a
133.2 ± 10.9a,b
133.1 ± 12.0a,b
147.4 ± 22.1b
60 min
89.2 ± 22.2a
112.1 ± 14.5b
142.0 ± 15.6b
139.0 ± 29.4b
120 min
89.3 ± 14.8a
88.8 ± 13.5a,b
92.0 ± 9.4a,b
114.7 ± 24.8b
Immunoreactive insulin (μU/mL)
0 min
6.2 ± 2.7a
5.7 ± 1.9a
3.2 ± 0.8b
5.7 ± 1.4a,b
30 min
74.1 ± 33.4a
47.2 ± 18.6a
38.1 ± 11.1b
44.5 ± 24.0a,b
60 min
42.3 ± 23.7a
65.9 ± 26.0b
53.5 ± 19.2a,b
45.0 ± 12.9a,b
120 min
36.1 ± 21.5a
40.6 ± 22.1a,b
35.6 ± 15.6a,b
68.0 ± 14.5b
HbA1c (%)
5.18 ± 0.27
5.12 ± 0.22
5.05 ± 0.20
5.10 ± 0.17
HOMA-IR
1.32 ± 0.63a
1.23 ± 0.43a
0.66 ± 0.18b
1.20 ± 0.26a,b
Insulinogenic index
2.50 ± 2.14a
1.10 ± 0.94b
0.76 ± 0.38b
0.59 ± 0.25b
HOMA-β
115 ± 47a
108 ± 44a,b
65 ± 33b
92 ± 35a,b
QUICKI
0.38 ± 0.03a
0.38 ± 0.03a
0.42 ± 0.02b
0.38 ± 0.02a,b
AUC-glucose (hr·mg/dL)
191 ± 31a
216 ± 21a,b
240 ± 11b
253 ± 53b
AUC-insulin (hr.μU/mL)
91.9 ± 44.8
94.7 ± 38.5
77.8 ± 27.3
88.0 ± 29.8
Values are means ± SD. HOMA-IR: homeostasis model assessment-insulin resistance; HOMA-β: homeostasis model assessment for β-cell function; QUICKI:
quantitative insulin sensitivity check index; AUC: area under the curve. The values for each index with different superscript letters are statistically different at
p < 0 05 by comparing different patterns of each time, as determined by Dunn’s multiple comparison test. Table 2: Glucose metabolism of women by insulin and glucose response type. Values are means ± SD. HOMA-IR: homeostasis model assessment-insulin resistance; HOMA-β: homeostasis model assessment for β-cell function; QUICKI:
quantitative insulin sensitivity check index; AUC: area under the curve. The values for each index with different superscript letters are statistically different at
p < 0 05 by comparing different patterns of each time, as determined by Dunn’s multiple comparison test. Values are means ± SD. There were no significant differences. 4. Discussion Insulin
secretion during the early stage was low, and glucose levels
remained elevated above 110 mg/dL at 120 min post-glucose
administration, despite maintained insulin secretion between
30 and 120 min (Table 2 and Figure 1(d)). This phenomenon
was caused by a secretion delay and not insulin resistance,
because HOMA-IR was not abnormal and AUC-I values
did not significantly differ from those in the normal type
group. Tanabe et al.’s study [24] in healthy people averaging
60 years old showed that the insulin peak at 120 min was
25 out of 680 (3.7%). Though we had 3 people (5.0%) in
our I-very late type group without IGT subjects, it was
more than that proportion. This phenomenon might be
consistent with the gradually increasing number of young
individuals with diabetes [8]. In this study, we were unable to investigate physical
activity or food requirements. However, our subjects were
studying nutritional science to become dietitians and might,
therefore, have better exercise and dietary habits than general
college students. The number of subjects in each group were
not equal, and the statistical difference might be hard to con-
firm, especially for the I-very late type group. However, it was
important to reveal patterns prior to the appearance of a car-
bohydrate metabolic disorder in young healthy women,
because it might facilitate diabetes prevention. We did not identify subjects with abnormal fasting
plasma glucose levels or other blood characteristics but did
identify two subjects with IGT and four distinct carbohydrate
patterns in healthy subjects by OGTT. In the two subjects
with IGT, BMIs were below 25 kg/m2 and the insulin peak
time was 120 min after glucose administration. The other
biochemical parameters were normal, and they did not have
a family medical history of diabetes in a relative within the
second degree of relationship. g
p
Relative to the normal group, insulin secretion was
delayed in the I-late and I&G-late groups and even further
delayed in the I-very late group. And the number of subjects
with normal response patterns was greater while that with
abnormal patterns was smaller. However, this might be
because this study was conducted only with young healthy
women. If we did it with a larger number of subjects
including men middle aged and older, this ratio might
have changed. 4. Discussion Table 3: Biochemical status of woman by insulin and glucose response type. Type
Normal
Insulin-late
Insulin- & glucose-late
Insulin-very late
Number
39
11
7
5
Total protein (g/dL)
7.67 ± 0.40
7.50 ± 0.45
7.28 ± 0.25
7.30 ± 0.43
Albumin (g/dL)
4.70 ± 0.25
4.79 ± 0.16
4.80 ± 0.22
4.63 ± 0.35
Albumin/globulin ratio
1.73 ± 0.25
1.84 ± 0.36
2.00 ± 0.32
1.75 ± 0.21
AST (U/L)
17.3 ± 3.2
17.6 ± 3.7
20.2 ± 6.1
15.5 ± 2.4
ALT (U/L)
13.3 ± 4.9
12.6 ± 5.8
14.8 ± 8.1
14.0 ± 6.8
γ-GTP (U/L)
17.7 ± 8.8
16.1 ± 4.9
16.2 ± 4.4
20.0 ± 8.4
LDL-C (mg/dL)
91.7 ± 21.3
90.0 ± 19.8
86.7 ± 13.8
105.3 ± 19.4
Triglyceride (mg/dL)
57.8 ± 22.7
43.0 ± 9.8
48.2 ± 16.1
68.5 ± 48.8
HDL-C (mg/dL)
68.9 ± 14.9
76.2 ± 7.8
73.3 ± 14.9
66.8 ± 13.7
Blood uric nitrogen (mg/dL)
11.7 ± 2.5
12.3 ± 4.5
10.3 ± 2.1
16.0 ± 6.7
Serum creatinine (mg/dL)
0.57 ± 0.08
0.58 ± 0.05
0.59 ± 0.08
0.55 ± 0.08
Serum uric acid (mg/dL)
4.18 ± 0.76
4.17 ± 0.74
4.18 ± 1.05
4.98 ± 0.87
White blood cell count (×102/μL)
60.7 ± 15.5
57.5 ± 11.7
55.1 ± 11.6
60.0 ± 14.3
Red blood cell count (×104/μL)
446 ± 28
451 ± 36
459 ± 27
444 ± 19
Hemoglobin (g/dL)
13.3 ± 0.9
13.3 ± 1.3
12.6 ± 2.7
13.3 ± 0.9
Hematocrit (%)
40.9 ± 2.2
40.1 ± 3.7
40.3 ± 6.5
40.6 ± 2.3
MCV (fL)
91.9 ± 3.5
90.6 ± 4.0
87.4 ± 11.4
91.4 ± 3.4
MCH (pg)
29.8 ± 1.8
29.4 ± 1.7
27.3 ± 5.3
30.0 ± 1.4
MCHC (%)
32.4 ± 1.2
32.5 ± 0.9
30.9 ± 2.7
32.8 ± 1.1
Platelet count (×104/μL)
26.6 ± 5.1
26.7 ± 6.0
24.7 ± 8.0
26.4 ± 4.2
V l
± SD Th
i
ifi
t diff Table 3: Biochemical status of woman by insulin and glucose response type. Journal of Diabetes Research 5 a 75g OGTT and were categorized in four groups based on
peak times of plasma insulin and glucose. There were several
studies categorized with insulin secretion time. Hayakawa
[21] and Kai et al. [22] divided the two insulin types into type
with peak time from 30min to 60min (normal) and those
with peaks at 120 min. 4. Discussion Sasaki et al. [23] and Tanabe et al. [24] divided the three insulin types into type with peak time
at 30 min, 60min, and 120 min, and we used this classifica-
tion in this study. However, they divided the types without
considering the peak time of plasma glucose levels. In this
study, we divided the types considering not only insulin
peak time but also plasma glucose peak, and as a result
we could divide them into four types. There were no subjects
with fever or menstruation on the day of the experiment. Even if there is a missing declaration, insulin secretion is
more susceptible to pregnancy and aging than menstruation
[25], so we think that it does not have a significant influence
on grouping of subjects. normal, because the increase in insulin secretion from 30 to
60 min resulted in a decrease in glucose levels. In addition,
insulin secretory capacity was normal, because HOMA-IR,
HOMA-β, AUC-G, and AUC-I, which are indices of carbo-
hydrate metabolism, were not significantly different from
the normal type group (Table 2). The third group was the
I&G-late type group (Figure 1(c)), characterized by peak
insulin and glucose times at 60min. This group displayed
not only an insulin secretory delay but also a secretion short-
age during fasting and the early stages of the OGTT. For
example, immunoreactive insulin levels and HOMA-IR
values, which were calculated using fasting glucose and insu-
lin levels, were significantly lower than those in the control
and I-late type groups (Table 2). II values, which were calcu-
lated using glucose and insulin levels at fasting and 30 min
after glucose administration, were lower than those in the
normal groups, because early insulin secretion after glucose
administration was the lowest. However, although the
60 min insulin level was the peak value in the I&G-late group,
it did not differ significantly from that in the normal type
group. AUC-I values in the I&G-late type group did not differ
from the other groups. These results suggest that fasting and
early insulin secretion were lower than those in the normal
group, but total secretion was comparable. The final group
was the I-very late type group, characterized by a peak insulin
time at 120 min and peak glucose time at 30min. This insulin
and glucose pattern was similar to subjects with IGT. 5. Conclusions We revealed that insulin secretion and plasma glucose pat-
terns could be categorized into four groups, despite all sub-
jects being healthy young women, because postchallenge
and fasting insulin secretion varied among individuals. The
first group was the normal type group, in which insulin and
glucose levels 30 min after glucose administration during an
OGTT were highest and then gradually decreased. The
second type displayed delayed postchallenge insulin secre-
tion. However, other indices of carbohydrate metabolism
(e.g., HOMA-IR, HOMA-β, AUC-I, and AUC-G) were
not significantly different. The third group displayed delayed
and reduced insulin secretion. In this group, fasting insulin
levels were low, and the lack of additional insulin secretion
caused a delay in peak plasma glucose levels. The fourth
group displayed very late postchallenge insulin secretion,
and this group contained IGT subjects. The delayed secretion
caused persistent postprandial hyperglycemia until 120 min
after glucose administration. Even in some healthy young
women, insulin secretion was delayed after oral glucose
administration, and they might, therefore, need improve-
ments in their lifestyle habits. Confirmation of insulin
secretion and plasma glucose patterns in young healthy
individuals might be helpful for early type 2 diabetes preven-
tion. Future studies should compare lifestyle habits among [6] DECODE Study Group and On behalf of the European
Diabetes Epidemiology Group, “Glucose tolerance and car-
diovascular mortality: comparison of fasting and 2-hour
diagnostic criteria,” Archives of Internal Medicine, vol. 161,
pp. 397–405, 2001. [7] C. Weyer, C. Bogardus, D. M. Mott, and R. E. Pratly, “The
natural history of insulin secretory dysfunction and insulin
resistance in the pathogenesis of type 2 diabetes mellitus,”
The Journal of Clinical Investigation, vol. 104, no. 6, pp. 787–
794, 1999. [8] J. C. N. Chan, V. Malik, W. Jia et al., “Diabetes in Asia: epide-
miology, risk factors, and pathophysiology,” JAMA, vol. 301,
no. 20, pp. 2129–2140, 2009. [9] R. Mitsui, M. Fukushima, Y. Nishi et al., “Factors responsible
for deteriorating glucose tolerance in newly diagnosed type 2
diabetes in Japanese men,” Metabolism, vol. 55, no. 1,
pp. 53–58, 2006. [10] C. Moris, C. O’Grada, M. Ryan et al., “Identification of differ-
ential responses to an oral glucose tolerance test in healthy
adults,” PLoS ONE, vol. 8, no. 8, article e72890, 2013. [11] H. K. Yang, J. H. Lee, I. Y. 4. Discussion In the normal type group (Figure 1(a)), insulin
secretion increased until 30 min after glucose administration,
at which time, plasma glucose and insulin levels gradually
decreased. Therefore, only glucose values at 30min, which
also corresponded to the insulin peak time, were significantly
higher than 0 min glucose values. Glucose levels at 60 and
120 min declined, returning to concentrations comparable
to those at baseline. Compared with that of the normal group,
the I-late and I&G-late type groups displayed intermedi-
ate impairment of carbohydrate metabolism. The subjects
in these groups displayed delayed insulin secretion follow-
ing glucose administration. In the I-late type group
(Figure 1(b)), in which the peak time for insulin was
60 min and glucose was 30min, insulin sensitivity might be In this study, we did not detect differences in AUC-I
levels between the four groups but did find differences in
AUC-G levels and peak times. This result suggests that differ-
ences in carbohydrate metabolism in healthy young women
were caused by different insulin secretion times and not
secretory capacity or insulin resistance. However, the mech-
anisms underlying differences in secretion times remain
unclear. For example, these differences might be caused by
genetic background, obesity, exercise habits, skipping break-
fast, having a midnight meal, and speed eating [12, 26–29]. However, in this study, there were subjects with BMIs above
25 kg/m2 in the normal and I-late type groups but not in the
I&G-late and I-very late type groups. In each group, there
were subjects with a family medical history of diabetes in a
relative within the second degree of relationship, and this
did not differ between groups (data not shown). Biochemical
parameters, including HOMA-IR and II, were normal. We
did not investigate the other factors in this study. Kikuoka 6 Journal of Diabetes Research the groups and conduct a follow-up survey to identify sub-
jects that develop IGT. the groups and conduct a follow-up survey to identify sub-
jects that develop IGT. et al. demonstrated that 13.1% of subjects with impaired fast-
ing glucose progressed to type 2 diabetes within 5 years [30]. Future studies will investigate lifestyle habits and conduct a
follow-up survey of the four insulin and plasma glucose
response groups. References [1] J. E. Gerich, “Clinical significance, pathogenesis, and manage-
ment of postprandial hyperglycemia,” Archives of Internal
Medicine, vol. 163, no. 11, pp. 1306–1316, 2003. [2] M. Brownlee, “The pathobiology of diabetic complications:
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pp. 920–924, 1999. Acknowledgments The funding of this investigation was provided in part by
Nagoya Bunri University. Conflicts of Interest Normal indices of carbohydrate metabolism (e.g., fasting
plasma glucose, HOMA-IR, and II) did not allow us to clas-
sify the subjects into four groups. For example, if we classified
only with peak insulin times, we might not have noticed the
significant differences in HOMA-IR, QUICKI, and immuno-
reactive insulin levels between the I-late type and I&G-late
type groups. If we classified subjects only according to peak
glucose times, we might not have noticed the maintenance
of hyperglycemia between 30 and 120 min in the I-very
late type group. We revealed these differences by using
both peak glucose and insulin times. In our study, about
37% of subjects had delayed insulin secretion. In a typical
study, these individuals might be categorized into the con-
trol group. In our 60 subjects, excluding two IGT subjects,
the average plasma glucose levels were 83.6 ±5.6, 123.3 ±
21.3, 100.8 ± 27.0, and 90.8 ± 15.2mg/dL at 0, 30, 60, and
120 min post-glucose administration, respectively. The aver-
age insulin levels were 5.8 ± 2.5, 68.1 ± 42.4, 52.4 ±37.5, and
41.6 ±30.8 μU/mL, respectively. These values were similar
to normal type values in this study, and thus, individuals
with delayed insulin secretion, who might be at risk for
diabetes, would have been overlooked in the healthy control
group. We believe that an understanding of one’s own carbo-
hydrate metabolism is important for type 2 diabetes preven-
tion. OGTTs may allow us to assess glucose and insulin
response types. The authors have no conflicts of interest directly relevant to
the content of this article. 5. Conclusions Choi et al., “The insulin resistance
but not the insulin secretion parameters have changed in the
Korean population during the last decade,” Diabetes & Metab-
olism Journal, vol. 39, no. 2, pp. 117–125, 2015. Journal of Diabetes Research 7 characteristics and the comparison of lifestyle and socioeco-
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“Comparison of the effect of abdominal obesity on new onset
of type 2 diabetes in a general Japanese. Elderly population
with that in a non-elderly population-the Tanno and Sobetsu
Study (in Japanese),” Nippon Ronen Igakkai Zasshi Japanese
Journal of Geriatrics, vol. 48, pp. 71–77, 2011. [27] T. Fukui, S. Yoshitaka, Y. Yamomoto, Y. Ayada, and
T. Yasuda, “Dose lifestyle really affect insulin resistance? (in Japanese),” Ningen Dock, vol. 22, no. 1, pp. 51–58, 2007. [13] M. Fukushima, H. Suzuki, and Y. Seino, “Insulin secretion
capacity in the development from normal glucose tolerance
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for progression from IFG to type 2 diabetes - follow-up
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betes Society, vol. 47, no. 9, pp. 723–726, 2004. [15] J. L. B. Richard and C. M. Donna, “Evaluation of glucose
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Clinical Chemistry, vol. 20, no. 5, pp. 586–590, 1974. [16] J. D. M. Albano, R. P. Ekins, G. Maritz, and R. C. 5. Conclusions Tuner, “A
sensitive, precise radioimmunoassay of serum insulin relying
on charcoal separation of bound and free hormone moieties,”
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genic index in secondary diabetes,” Hormone and Metabolic
Research, vol. 7, no. 2, pp. 107–115, 1975. [19] A. Katz, S. S. Nambi, K. Mather et al., “Quantitative insulin
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(in Japanese),” Japanese Journal of Gastroenterology, vol. 67,
no. 6, pp. 457–468, 1970. [22] Y. Kai, R. Abe, S. Katono et al., “Plasma insulin response in
the cases with a borderline 50g oral glucose tolerance test
(in Japanese),” Journal of the Japan Diabetes Society, vol. 17,
no. 1, pp. 16–22, 1974. [23] A. Sasaki, K. Matsumiya, M. Arao, K. Hasegawa, and
N. Horiuchi, “Evaluation of insulin response in subjects with
normal and mildly impaired glucose tolerance (in Japanese),”
Journal of the Japan Diabes Society, vol. 21, no. 11, pp. 983–
992, 1978. [24] M. Tanabe, Y. Arase, H. Tsuji et al., “Insulin secretory response
to oral glucose in subjects with normal glucose intolerance
(in Japanese),” Ningen Dock, vol. 26, no. 4, pp. 601–606, 2011. [25] T. Okuyama and Y. Terauchi, “Blood glucose: gender-specific
medicine in diabetes mellitus (in Japanese),” Journal of the
Japan Diabetes Society, vol. 56, no. 8, pp. 522–524, 2013. [26] Y. Azami, A. Ikota, K. Ito et al., “A multi-center study of young
adults
with
type
2
diabetes
mellitus:
their
clinical
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MENINGKATKAN KESEJAHTERAAN SUBJEKTIF PASIEN GAGAL GINJAL KRONIS MELALUI TERAPI KOGNITIF PERILAKU RELIGIUS
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JIP/JIP (Jurnal Intervensi Psikologi)
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Elda Trialisa Putri
Qurotul Uyun
Rr. Indahria Sulistyarini
Fakultas Psikologi dan Ilmu Sosial Budaya Universitas Islam Indonesia, Yogyakarta
Email: eldatrialisa@gmail.com Elda Trialisa Putri
Qurotul Uyun
Rr. Indahria Sulistyarini
Fakultas Psikologi dan Ilmu Sosial Budaya Universitas Islam Indonesia, Yogyakarta
Email: eldatrialisa@gmail.com ABSTRACT The purpose of this study is to examine the effectiveness of religious cognitive behavioral therapy in
improving subjective well-being of chronic kidney disease patients. The hypothesis of the study was that
there is a significant difference in the subjective well-being between experimental and control groups. Four
chronic kidney disease patients with age between 22-45 years old, Muslim, and suffering chronic kidney
disease more than 6 months participated this study. The design of this experiment was a quasi experiment
with pretest-posttest control group design and follow up. Data were collected with satisfaction with Life
Scale (SWLS) from Diener, Emmons, Larsen and Griffin’s Scale (1985) and Positive Affect and Negative
Affect Schedule (PANAS) from Watson, Clark, and Tellegen’s scale (1988). Independent sample t-test was
conducted to analyze the data. The result showed that there was a difference life satisfaction between the
experimental and control group (t=4, 287, p=0, 005 (p<0,01). Also, there were difference on an
affection score between the experimental and control group (t=4, 407, p=0, 005 (p<0,01). Keywords: religious cognitive behavioral therapy, subjective well-being, life satisfaction, affection, chronic
kidney disease kidney disease ABSTRAK Penelitian ini bertujuan untuk mengetahui efektivitas terapi kognitif perilakuan religius untuk meningkatkan
kesejahteraan subjektif pasien gagal ginjal kronis yang menjalani hemodialisis. Subjek dalam penelitian ini
adalah pasien gagal ginjal kronis yang menjalani hemodialisis lebih dari 6 bulan, laki-laki maupun
perempuan, beragama Islam, berusia antara 22-45 tahun (N kontrol=4, N eksperimen=4). Desain
penelitian ini adalah kuasi eksperimen, dengan pretest-posttest control group design. Penelitian ini
menggunakan analisis independent sample t-test. Hasil menunjukkan terdapat perbedaan yang signifikan
pada nilai kepuasan hidup dengan t=4,287, p=0,005 (p<0,01) antara kelompok eksperimen dan
kelompok kontrol. Pada skala afeksi terdapat perbedaan yang signifikan antara kelompok kontrol dan
eksperimen yaitu t=4,407, p=0,005 (p<0,01). Berdasarkan hasil analisis terlihat bahwa terapi kognitif
perilakuan religius mampu meningkatkan kesejahteraan subjektif pasien gagal ginjal kronis. ata Kunci: Terapi kognitif perilakuan religius, kesejahteraan subjektif, kepuasan hidup, afeksi. Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 89 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Penyakit ginjal adalah kelainan
yang mengenai organ ginjal yang timbul
akibat berbagai faktor, misalnya infeksi,
tumor, kelainan bawaan, penyakit meta-
bolik atau degeneratif, dan lain-lain
(Kementrian
Kesehatan
RI,
2013). Kelainan tersebut dapat memengaruhi
struktur dan fungsi ginjal dengan tingkat
keparahan yang berbeda-beda. Pasien
mungkin merasa nyeri, mengalami gang-
guan berkemih, dan lain-lain. Terkadang
pasien penyakit ginjal tidak merasakan
gejala sama sekali. Pada keadaan terbu-
ruk, pasien dapat terancam nyawanya
jika tidak menjalani hemodialisis (cuci
darah) berkala atau transplantasi ginjal
untuk menggantikan organ ginjalnya
yang telah rusak parah. Di Indonesia,
penyakit
ginjal
yang
cukup
sering
dijumpai antara lain adalah penyakit
gagal ginjal dan batu ginjal. Prevalensi gagal ginjal kronis di
Indonesia -berdasarkan diagnosis dokter-
sebesar 0,2%. Prevalensi tertinggi di
Sulawesi Tengah sebesar 0,5 %, diikuti
Aceh, Gorontalo, dan Sulawesi Utara
masing-masing 0,4 %. Sementara Nusa
Tenggara
Timur,
Sulawesi
Selatan,
Lampung, Jawa Barat, Jawa Tengah, DI
Yogyakarta, dan Jawa Timur masing–
masing 0,3 %. Prevalensi penyakit gagal
ginjal kronis yang didiagnosis dokter
meningkat seiring dengan bertambahnya
umur, meningkat tajam pada kelompok
umur 35-44 tahun (0,3%), diikuti umur
45-54 tahun (0,4%), dan umur 55-74
tahun (0,5%), tertinggi pada kelompok
umur ≥75 tahun (0,6%). Prevalensi pada
laki-laki (0,3%) lebih tinggi dari perem-
puan (0,2%), prevalensi lebih tinggi pada
masyarakat perdesaan (0,3%), tidak ber-
sekolah (0,4%), pekerjaan wiraswasta,
petani/nelayan/buruh (0,3%), dan kuintil
indeks kepemilikan terbawah dan mene-
ngah bawah masing-masing 0,3% (Ke-
mentrian Kesehatan RI, 2013). Gagal Ginjal Kronik (GGK) adalah
suatu sindrom klinis yang disebabkan
penurunan fungsi ginjal yang bersifat
menahun, berlangsung progresif dan
cukup lanjut, serta bersifat persisten dan
irreversibel (Mansjoer, 2000). GGK di-
definisikan sebagai gagal ginjal kronis
jika pernah didiagnosis menderita penya-
kit gagal ginjal kronis (minimal sakit
selama 3 bulan berturut-turut) oleh dok-
ter. Peningkatan jumlah pasien gagal
ginjal kronik tidak diikuti dengan pena-
nganan fisik dan psikis yang lebih baik
sehingga banyak pasien gagal ginjal
kronik kondisinya memburuk, bahkan
menyebabkan kematian (Safitri, 2013). Hal ini juga sejalan dengan hasil wawan-
cara yang dilakukan oleh peneliti se- 90 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 90 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……… Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi … belumnya kepada pasien gagal ginjal
kronik bahwa proses dialisis harus
dialami pasien selama hidupnya. Dialisis
dilakukan 2 kali seminggu selama paling
sedikit 4 atau 5 jam per terapi (Sadif,
2013). Terapi hemodialisis pada umum-
nya akan menimbulkan stres fisik seperti
kelelahan, sakit kepala dan keringat di-
ngin akibat tekanan darah yang menurun. Efek ini juga memengaruhi kondisi
psikologis pasien di mana pasien gagal
ginjal kronik mengalami gangguan dalam
proses berpikir dan konsentrasi serta
gangguan dalam hubungan sosial. menarik diri dari lingkungan sekitar
sehingga kualitas hidup dari pasien GGK
juga terlihat menurun. Permasalahan
psikologis
yang
dialami pasien GGK sudah ditunjukkan
dari sejak pertama kali pasien divonis. Kecemasan dan ketakutan adalah reaksi
umum terhadap stress penyakit. Perasaan
hilang kendali, bersalah dan frustrasi juga
turut berperan dalam reaksi emosional
pasien. Seren mengungkapkan bahwa
kecemasan ber-pengaruh kepada kondisi
fisik, kualitas hidup yang buruk, dan
perasaan
ketidak-berdayaan
(Foreze,
Martin, Patton, Zadeh & Kopple, 2010). Penelitian ini juga menemukan bahwa
kecemasan mening-kat pada saat pasien
akan mulai me-masuki rutinitas dialisis. Hal ini me-nyebabkan penderita menjadi
semakin terpuruk dan memengaruhi
kondisi fisik-nya. Kesejahteraan subjektif
yang rendah ditunjukkan dengan afek
positif dan kualitas hidup yang rendah
pula sementara afek negatifnya yang
tinggi. Afek negatif yang tinggi inilah
yang turut memengaruhi kondisi fisik
pasien GGK. Penelitian yang dilakukan Safitri
dan Sadif (2013) menemukan bahwa
terdapat enam gejala yang muncul pada
pasien GGK, yaitu kemarahan karena
penyakitnya
telah
membuat
dirinya
menderita, keputusasaan, ketidakberda-
yaan, merasa lelah menjalani hemodia-
lisis, merasa lebih baik bila ada du-
kungan keluarga dan pasrah pada Tuhan
yang memberi kekuatan untuk meng-
hadapi penyakitnya. Gejala-gejala yang
muncul ini menunjukkan ketidaksiapan
pada diri pasien GGK telah mengalami
penyakit kronis, sehingga mengalami
perubahan hidup yang drastis. Hal ini
memengaruhi
kesejahteraan
subjektif
pada pasien GGK di mana terdapat
perubahan dalam hidup seperti merasa
putus asa, merasa tidak berguna, dan Penelitian yang terkait dengan
terapi yang digunakan untuk mening-
katkan
kesejahteraan
subjektif
pada
pasien GGK adalah penelitian Cahyareni
(2014),
tentang
efektivitas
pelatihan
kebersyukuran
untuk
meningkatkan | 91 91 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Elda Trialisa Putri, Qurotul Uyun, & Rr. 92 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi … Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi … dalam kehidupan personalnya. Menetap-
nya perubahan positif menjadi keung-
gulan CBT-Religius. Religiusitas melalui
kemampuan penyesuaian diri memiliki
keterkaitan dengan subjective well being
pada usia akhir dewasa madya. Keter-
kaitan ini menunjukkan bahwa kehi-
dupan religi seseorang akan mengarah-
kan
seseorang
pada
pengembangan
kemampuan penyesuaian dirinya, sehing-
ga dapat tercapai kondisi subjective well
being (Hida, Suroso, & Muhid, 2013). Berdasarkan hal tersebut peneliti
melakukan terapi kognitif perilakuan re-
ligius guna meningkatkan ke-sejahteraan
subjektifpada pasien gagal ginjal kronik
yang melakukan hemo-dialisis. Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……… Indahria Sulistyarini yang difokuskan secara langsung untuk
mengubah sisi kognitif, dengan asumsi
bahwa hal tersebut akan diikuti oleh
perubahan perilaku, serta sebaliknya
intervensi yang difokuskan untuk meng-
ubah perilaku, dengan asumsi akan
diikuti dengan perubahan kognisi (Sundel
& Sundel, 2005). Koreksi terhadap
kesalahan konstruk pikir dapat meng-
arahkan pada peningkatan kondisi klinis
seseorang. Hal ini terlihat dari sisi
kognitif subjek yang seringkali menya-
lahkan dirinya karena sakit yang dimiliki
subjek sehingga subjek menjadi lebih
sensitif dan mudah tersinggung. subjective well being pada pasien gagal
ginjal. Penelitian ini melibatkan 16 orang
pasien GGK yang menjalani hemodialisis
dengan usia berkisar antara 20-35 tahun
yang memiliki skor SWLS sedang atau
rendah dan afek negatif yang sedang atau
tinggi. Hasil dari intervensi yang telah
dilakukan adalah pelatihan kebersyu-
kuran mempunyai pengaruh terhadap
peningkatan subjective well-being (SWB)
pada pasien gagal ginjal. Penelitian lain
yang digunakan pada pasien GGK adalah
tentang Gratitude Cognitive Behavior
Therapy (G-CBT) untuk meningkatkan
penerimaan diri pada pasien gagal ginjal
kronik yang menjalani hemodialisis oleh
Safitri (2013). Hasil dari penelitian ini
yaitu
G-CBT
memberikan
pengaruh
terhadap peningkatan penerimaan diri
pada
pasien
GGK
yang
menjalani
hemodialisis. Pendekatan kognitif perilakuan di-
percaya memiliki hasil yang lebih opti-
mal bila didukung pendekatan religius. Menurut Ancok dan Suroso (2011),
religiusitas merupakan perilaku ritual
(ibadah) yang dilakukan individu serta
aktivitas lainnya yang didorong kekuatan
supranatural
guna
mendekatkan
diri
kepada Allah SWT termasuk didalamnya
aktivitas yang tampak maupun yang tidak
tampak (di dalam hati). Semakin klien
merasakan
adanya
peningkatan
dan
keyakinan beragama, perubahan positif
yang dirasakan cenderung lebih menetap
(McCullough & Larson, 1999). Penelitian
Utami (2012) mendapatkan bahwa reli-
giositas memiliki pengaruh terhadap
kesejahteraan subjektif pada mahasiswa Berdasarkan beberapa penelitian di
atas, pasien GGK sudah menerima
beberapa macam terapi untuk dapat
meningkatkan
kondisi
pasien
GGK. Pasien GGK membutuhkan keadaran dari
dalam diri untuk dapat mengubah pola
pikir
(cognitive)
serta
mengarahkan
emosi yang lebih positif sehingga ter-
bentuk perilaku yang dapat meningkat-
kan kesejahteraan subjektif. Terapi kognitif perilakuan menca-
kup dua tipe intervensi, yaitu intervensi 92 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Desain Penelitian Adapun
bentuk
penelitian
ini
adalah kuasi eksperimen dengan meng-
gunakan rancangan pretest-posttest con-
trol group design. Rancangan pretest-
posttest control group design adalah
metode eksperimen yang berusaha untuk
membandingkan efek suatu perlakuan
terhadap variabel tergantung yang diuji
dengan cara membandingkan keadaan
variabel
tergantung
pada
kelompok
eksperimen setelah dikenai perlakuan
dengan kelompok kontrol yang tidak
dikenai perlakuan (Azwar, 2012). Keuntungan adanya penambahan
nilai
religius
dalam
terapi
kognitif
perilakuan untuk meningkatkan kesejah-
teraan subjektif antara lain: klien menjadi
lebih mudah menerima terapi, lebih
termotivasi untuk berpartisipasi dalam
terapi, meningkatkan efektivitas terapi
dalam meningkatkan kesejahteraan sub-
jektif dan membantu mempertahankan
kesejahteraan subjektif pada pasien GGK. Tabel 1. Rancangan eksperimen yang digunakan dalam penelitian ini
Kelompok
Prates
Perlakuan
Pascates
Tindaklanjut
KE
O1
X
O2
O3
KK
O1
-X
O2
O3
Keterangan :
KE
: Kelompokeksperimen
O3
: Tindaklanjut
KK
: Kelompokkontrol
X
: Perlakuan
O1
: Pengukuran prates
-X
: Tanpa perlakuan
O2
: Pengukuran pascates
Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 | 93 Tabel 1. Rancangan eksperimen yang digunakan dalam penelitian ini Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini negatif terdiri atas 20 kata yang berkaitan
dengan emosi dan atau perasaan. Skala
ini dibuat oleh Watson, Clark & Tellegen
(1988) yang digunakan American Psycho-
logical
Association
(APA). Sebelum
digunakan oleh peneliti, instrumen ini
telah diuji cobakan di dalam penelitian
yang dilakukan oleh Gatari kepada 50
responden yang berasal dari Jawa. Se-
telah data diperoleh kemudian dilakukan
uji reliabilitas. Hasilnya menunjukkan
instrumen ini reliable dengan koefisien
alpha sebesar 0,91 (Gatari, 2008). Pengumpulan Data Pengumpulan data berikutnya dila-
kukan dengan menggunakan skala kese-
jahteraan subjektif. Pertama: The Satis-
faction with Life Scale (SWLS). Skala ini
disusun berdasarkan teori Diener, dkk
(1985) yang terdiri atas 5 item dengan
skor yang bergerak dari 1-7 (sangat tidak
setuju
hingga
sangat
setuju). Pada
penelitian sebelumnya, instrumen ini
sudah pernah diujicobakan kepada 50
orang Jawa. Setiap item diukur validitas
dan reliabilitasnya dengan bantuan SPSS. Teknik yang digunakan adalah Cronbach-
Alpha. Item ini memiliki item-total
correlation minimal 0,3 dan koefisien
alpha instrumen ini sebesar 0,83 (Gatari,
2008). Subjek Penelitian Subjek penelitian adalah penderita
gagal ginjal kronis yang menjalani hemo-
dialisis dengan kriteria sebagai berikut:
beragama Islam, berjenis kelamin laki-
laki/ perempuan, usia 22-45 tahun, pen-
didikan terakhir adalah SMA, menjalani
proses hemodialisis > 6 bulan, mem-
punyai skor rendah sampai sedang pada
skala kepuasan hidup, bersedia secara
sukarela mengikuti penelitian. 94 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Mengakhiri terapi 76 Service Solution (IBM SPSS) 22.0 for
windows. Jika data penelitian diterima
melalui uji asumsi (normal dan homo-
gen), maka pengujian hipotesis dalam
penelitian ini dilakukan dengan analisis
parametrik dengan teknik Independent
Sample t-test digunakan untuk menguji
perbedaan antara kelom-pok eksperimen
dan kelompok kontrol setelah diberikan
terapi kognitif perilakuan religius. Keenam: Pengambilan data pasca-
tes dilakukan setelah terapi selesai dilak-
sanakan. Selanjutnya pengambilan data
tindak lanjut dilakukan dua minggu
setelah terapi. Ketujuh: Tahapan akhir
adalah
melakukan
analisis
secara
keseluruhan Teknik Analisis Data Metode analisis data yang diguna-
kan dalam penelitian ini adalah analisis
data kuantitatif dan kualitatif. Analisis
data kuantitatif dalam penelitian ini
menggunakan
program
International
Business Machine Statistical Product and Prosedur Intervensi Prosedur dalam penelitian ini meli-
puti beberapa tahapan penelitian, yaitu:
Pertama: Persiapan penelitian meliputi
analisis kebutuhan terkait permasalahan
dalam penelitian (wawancara dan studi
pustaka) serta pengurusan perizinan. Kedua: Penyusunan modul terapi kognitif
perilakuan religius. Ketiga: Pengambilan
data
prates
pada
subjek
penelitian
(kelompok eksperimen dan kelompok
kontrol) dilakukan untuk mendapatkan
gambaran mengenai kesejahteraan sub-
jektif pada pasien gagal ginjal kronis
yang menjalani hemodialisis. Keempat: Subjek penelitian (kelom-
pok eksperimen) diberi terapi kognitif
perilakuan religius dalam tiga kali per-
temuan. Terapi diberikan oleh psikolog Kedua: Skala PANAS (Positive and
Negative Afect Scale). Skala ini dimak-
sudkan untuk mengukur afek positif dan 94 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 94 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……….. yang berpengalaman memberikan inter-
vensi yang terkait dengan keislaman. Tabel 2. Kegiatan Terapi Kognitif Perilakuan Religius
Pertemuan
Kegiatan
Tujuan
Waktu
I
1. Pembukaan:
Perkenalan antara terapis
dan klien
2. Menangkap pikiran
3. Latihan memonitor dan
mengenali aktivitas
4. Psikoedukasi tentang
keterkaitan pikiran,
perasaan, perilaku dan
religius
5. Relaksasi pelepasan
religius: kepasrahan
Membangun rapport
Memberikan penjelasan pada klien
mengenai terapi
Memotivasi klien
Melatih klien mengenali pikiran
irasional
Melatih klien memonitor aktivitas
keseharian serta mengenali dan
merencanakan aktivitas yang dapat
dilakukan
dalam
meningkatkan
aktivitas
Melatih klien mengenali keterkaitan
antara pikiran, perasaan, perilaku
dan religius
Melatih klien menggunakan konsep-
konsep
ajaran
agama
untuk
merelaksasikan diri
90 menit
II
1. Presentasi diri
2. Latihan menguji dan
memodifikasi pikiran
negatif berdasarkan ajaran
agama
3. Aplikasi niat dalam
beraktivitas
Evaluasi tugas rumah dan
Membangun konteks religius dalam
terapi
Melatih klien mengenali, menguji
dan mengganti pikiran irasional
dengan pikiran yang lebih tepat
menurut keyakinan agama klien
Melatih
klien
menerapkan
niat
dalam aktivitas sehari-hari
75 menit Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini 4. Relaksasi dengan isyarat
religius : doa
Melatih klien menggunakan konsep-
konsep
ajaran
agama
untuk
merelaksasikan diri
III
1. Presentasi diri
2. Melatih rasa syukur
3. Pernyataan diri dengan
keyakinan agama
4. Praktik Kebersyukuran:
Sujud Syukur
5. Terminasi
Evaluasi
tugas
rumah
dan
Membangun konteks religius dalam
terapi
Melatih klien mensyukuri nikmat
dalam aktivitas keseharian
Melatih
klien
mengatasi
situasi
menekan dengan pernyataan diri
yang mendekatkan dengan Tuhan
Melatih klien menggunakan konsep
kebersyukuran dengan sujud syukur
Mengakhiri terapi
75 menit 5. Terminasi HASIL PENELITIAN Deskripsi Subjek Penelitian Deskripsi Subjek Penelitian 96 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi … Tabel 3. Deskripsi Kepuasan Hidup (SWLS) Kelompok Eksperimen
Subjek
JK
Prates
Pascates
Tindak
Lanjut
Gained
Score
(Pra-
Pasca)
Gained
Score
(Pra-
Tindak
Lanjut)
ES
L
17
18
21
1
4
DS
L
13
13
16
0
3
R
P
15
17
21
2
6
AD
P
13
19
22
6
9 Tabel 3. Deskripsi Kepuasan Hidup (SWLS) Kelompok Eksperimen pada nilai kepuasan hidup. Peningkatan
terlihat dari gained score pra-pasca dan
pra-tindak lanjut. Berdasarkan tabel 3, semua par-
tisipan kelompok eksperimen dalam
penelitian ini mengalami peningkatan Tabel 4. Deskripsi Data Penelitian Skala Afek (PANAS) Kelompok Eksperimen
Subjek
JK
Prates
Pascates
Tindak
Lanjut
Gained
Score
(Pra-
Pasca)
Gained
Score
(Pra-
Tindak
Lanjut)
ES
L
-1
14
16
15
17
DS
L
-3
0
2
3
5
R
P
-19
-12
5
7
24
AD
P
-2
8
9
10
11 abel 4. Deskripsi Data Penelitian Skala Afek (PANAS) Kelompok Eksperimen peningkatan pada nilai afeksi. Hal ini
didasarkan gained score pra-pasca dan
pra tindak lanjut keempat subjek. peningkatan pada nilai afeksi. Hal ini
didasarkan gained score pra-pasca dan
pra tindak lanjut keempat subjek. Berdasarkan tabel 4, semua partisi-
pan kelompok eksperimen yang terdiri
atas 4 orang subjek, 2 subjek laki-laki
dan 2 subjek perempuan mengalami Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 97 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Tabel 5. Deskripsi Kepuasan Hidup (SWLS) Kelompok Kontrol
Subjek
JK
Prates
Pascates
Tindak
Lanjut
Gained
Score
(Pra-
Pasca)
Gained
Score (Pra-
Tindak
Lanjut)
VR
P
14
12
15
-2
1
L
L
18
16
16
-2
-2
S
L
14
12
13
-2
-1
E
P
20
13
17
-7
-3 Tabel 5. Deskripsi Kepuasan Hidup (SWLS) Kelompok Kontrol pascates. Kemudian pada gained score
pra-tindak lanjut terlihat satu orang
subjek yang mengalami peningkatan
skor, dan tiga orang subjek memiliki
penurunan skor kepuasan hidup di-
bandingkan dengan prates. Berdasarkan tabel 5, semua partisi-
pan kelompok kontrol yang berjumlah 4
orang subjek yang terdiri atas 2 orang
laki-laki dan 2 orang perempuan meng-
alami penurunan skor kepuasan hidup. Hal ini didasarkan pada gained score pra- Tabel 6. HASIL PENELITIAN Deskripsi Afek (PANAS) Kelompok Kontrol
Subjek
JK
Prates
Pascates
Tindak
Lanjut
Gained
Score
(Pra-
Pasca)
Gained
Score
(Pra-
Tindak
Lanjut)
VR
P
-11
-10
-13
1
-2
L
L
-3
-5
-9
-2
-6
S
L
-5
-8
-8
-3
-3
E
P
-6
-9
-12
-3
-6 Tabel 6. Deskripsi Afek (PANAS) Kelompok Kontrol Tabel 6. Deskripsi Afek (PANAS) Kelompok Kontrol
Subjek
JK
Prates
Pascates
Tindak
Lanjut
Gained
Score
(Pra-
Pasca)
Gained
Score
(Pra-
Tindak
Lanjut)
VR
P
-11
-10
-13
1
-2
L
L
-3
-5
-9
-2
-6
S
L
-5
-8
-8
-3
-3
E
P
-6
-9
-12
-3
-6 penurunan afeksi. Pada gained score pra-
tindak lanjut terlihat keempat orang
subjek mengalami mengalami penurunan
afeksi. Berdasarkan tabel 6, pada gained
score pra-pascates terlihat bahwa 1 orang
subjek mengalami peningkatan afeksi
dan
tiga
orang
subjek
mengalami 8 | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 98 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ………. Hasil Uji Asumsi
Tabel 7. Uji Normalitas Skala Kepuasan Hidup (SWLS)
Kelompok
Perlakuan
P
Keterangan
Eksperimen dan Kontrol
Prates
0,067;
p>0,05
Normal Berdasarkan tabel 7 diketahui hasil
sebaran skor kepuasan hidup pada pene-
litian ini terdistribusi secara normal. Tabel 8. Uji Normalitas Skala Afek (PANAS)
Kelompok
Perlakuan
P
Keterangan
Eksperimen dan Kontrol
Prates
0,099;
p>0,05
Normal Pada tabel 8 diketahui bahwa hasil
sebaran skala afeksi (PANAS) berdistri-
busi normal (p>0,05). Tabel 9. Uji Homogenitas Data Penelitian Skala Kepuasan Hidup (SWLS)
Kelompok
Levene Statistic
p
Keterangan
Prates Eksperimen dan
Kontrol
3,000
0,134;
p>0,05
Normal Tabel 9. Uji Homogenitas Data Penelitian Skala Kepuasan Hidup (SWLS) dapat
dikatakan
bahwa
tidak
ada
perbedaan varians data pada saat prates
kelompok eksperimen dan kelompok
kontrol pada subjek penelitian. Pada tabel 9 hasil analisis pada
skala kepuasan hidup (SWLS) subjek
penelitian menunjukkan nilai Levene
statistic
3,000
dengan
p=0,134
(p>0,05). Berdasarkan kaidah, maka Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 99 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini Tabel 10. Uji Homogenitas Data Penelitian Skala Afek (PANAS)
Kelompok
Levene Statistic
p
Keterangan
Prates Eksperimen dan
Kontrol
2,808
0,145;
p>0,05
Normal Tabel 10. Uji Homogenitas Data Penelitian Skala Afek (PANAS) kaidah, maka dapat dikatakan bahwa
tidak ada perbedaan varians data pada
saat prates kelompok eksperimen dan
kelompok kontrol pada subjek penelitian. Pada tabel 10 hasil analisis pada
skala afek (PANAS) subjek penelitian
menunjukkan nilai Levene statistic 2,808
dengan p=0,145 (p>0,05). Berdasarkan Hasil Uji Hipotesis Tabel 11. Data Perbandingan Hasil Uji Hipotesis Skala Kepuasan Hidup
Perhitungan
t
Sig. (p)
Kesimpulan
Selisih skor Prates-Pascates
3,031
0,023
Signifikan
Selisih skor Pascates-Tindak Lanjut
4,287
0,005
Signifikan artinya terdapat perbedaan kepuasan
hidup pada kelompok eksperimen dan
kelompok kontrol setelah dua minggu
dari pemberian intervensi berakhir (tin-
dak lanjut). Hasil menunjukkan bahwa
terdapat perbedaan kepuasan hidup yang
signifikan setelah diberikan terapi kogni-
tif perilakuan religius. Dengan demikian
terapi kognitif perilakuan religius ber-
pengaruh terhadap peningkatan kepuas-
an hidup para pasien gagal ginjal kronis. Hasil uji independent sample t-test
skala kepuasan hidup pada kelompok
eksperimen dan kelompok kontrol pada
saat prates dan pascates adalah t=3,031,
=0,023 (p<0,05) artinya terdapat perbe-
daan kepuasan hidup pada kelompok
eksperimen
dan
kelompok
kontrol. Selanjutnya hasil uji independent sample
t-test
skala
kepuasan
hidup
pada
kelompok eksperimen dan kelompok
kontrol pada saat prates dan tindak lanjut
adalah t=4,287, p=0,005 (p<0,01) Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 100 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……….. Tabel 12. Data Perbandingan Hasil Uji Hipotesis Skala Afek (PANAS)
Perhitungan
t
Sig. (p)
Kesimpulan
Selisih skor Prates-Pascates
3,888
0,008
Signifikan
Selisih skor Pascates-Tindak Lanjut
4,407
0,005
Signifikan perilakuan religius terhadap peningkatan
kesejahteraan subjektif pada pasien gagal
ginjal kronis yang menjalani hemo-
dialisis. Hasil yang diperoleh dalam
penelitian ini menunjukkan bahwa terapi
kognitif perilakuan religius berpengaruh
dalam peningkatan kesejahteraan sub-
jektif pada pasien gagal ginjal kronis
yang menjalani hemodialisis. Hasil pene-
litian menunjukkan bahwa terjadi perbe-
daan rerata gain score kesejahteraan
subjektif setelah diberi perlakuan berupa
terapi kognitif perilakuan religius pada
kelompok eksperimen dan kelompok
kontrol yang tidak diberikan perlakuan
pada pasien gagal ginjal kronis yang
menjalani hemodialisis. Hasil uji independent sample t-test
skala Afek pada kelompok eksperimen
dan kelompok kontrol pada saat prates
dan pascates adalah t=3,888, p=0,008
(p<0,01) artinya terdapat perbedaan
nilai afek pada kelompok eksperimen
dan kelompok kontrol sebelum dan
sesudah
pemberian
terapi
kognitif
perilakuan religius. Selanjutnya hasil uji
independent sample t-test skala afek
pada kelompok eksperimen dan kelom-
pok kontrol pada saat prates dan tindak
lanjut
adalah
t=4,407,
p=0,005
(p<0,01) artinya terdapat perbedaan
nilai afek pada kelompok eksperimen
dan kelompok kontrol setelah dua
minggu dari pemberian intervensi ber-
akhir (tindak lanjut). Hasil menunjukkan
bahwa terapi kognitif perilakuan religius
memiliki
pengaruh
yang
signifikan
terhadap nilai afeksi pasien gagal ginjal
kronis
hingga
masa
tindak
lanjut
berakhir. Hasil Uji Hipotesis Pada penelitian ini, kesejahteraan
subjektif terdiri atas 2 aspek yaitu
kepuasan hidup yang diukur dengan
skala kepuasan hidup atau Satisfaction
with Life Scale (SWLS) dan aspek afeksi
yang diukur dengan Positive Affect and
Negative
Affect
Schedule
(PANAS). Berdasarkan analisis data yang dilakukan
pada kesejahteraan subjektif pasien gagal
ginjal kronis yang menjalani hemodialisis
menunjukkan bahwa terdapat perbedaan Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 PEMBAHASAN Penelitian
ini
bertujuan
untuk
mengetahui pengaruh terapi kognitif 101 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini kesejahteraan subjektif antara kelompok
eksperimen yang diberikan perlakuan
terapi kognitif perilakuan religius dan
kelompok kontrol yang tidak diberikan
perlakuan. tindak lanjut) didapat nilai t=4,407,
p=0,005 (p<0,01). Hal tersebut menun-
jukkan bahwa ada perbedaan yang
signifikan pada tingkat afeksi pasien gagal
ginjal kronis antara kelompok ekspe-
rimen dan kelompok kontrol pada saat
dilakukannya tindak lanjut. Berdasarkan hasil analisis uji beda
independent sample t-test pada perubah-
an nilai kepuasan hidup diketahui bahwa
terdapat perubahan signifikan pada saat
sebelum dilaksanakan terapi kognitif
perilakuan religius dan setelah terapi
(pra-pasca) antara kelompok eksperimen
dan kelompok kontrol dengan t=3,031,
p=0,023 (p<0,05). Selanjutnya pada
saat sebelum pelaksanaan terapi kognitif
perilakuan religius dan pada saat tindak
lanjut (pra-tindak lanjut) didapat nilai
t=4,287,
p=0,005
(p<0,01). Hal
tersebut
menunjukkan
bahwa
ada
perbedaan yang signifikan pada tingkat
kepuasan hidup pasien gagal ginjal
kronis antara kelompok eksperimen dan
kelompok kontrol. Hasil analisis di atas menunjukkan
bahwa pada pasien gagal ginjal kronis
yang menjalani hemodialisis dan men-
dapat perlakuan terapi kognitif perilaku-
an religius mengalami peningkatan ke-
sejahteraan subjektif dilihat dari dua
aspek yaitu kepuasan hidup dan aspek
afeksi dibandingkan dengan pasien gagal
ginjal kronis yang tidak mendapatkan
terapi kognitif perilakuan religius. Hal ini
juga didukung oleh hasil-hasil penelitian
Wang dan Chen (2012) bahwa terapi
kognitif perilaku efektif dapat mening-
katkan kualitas hidup pasien gagal ginjal
kronis yang menjalani hemodialisis dan
juga dapat meningkatkan kualitas tidur
juga
menurunkan
kecemasan
dan
depresi. Pada skala afeksi hasil analisis uji
beda independent sample t-test pada
perubahan nilai afeksi kelompok ekspe-
rimen dan kelompok kontrol pada saat
sebelum dilaksanakan terapi kognitif
perilakuan religius dan setelah terapi
(pra-pasca) yakni t=3,888, p=0,008
(p<0,01). Selanjutnya pada saat sebelum
pelaksanaan terapi kognitif perilakuan
religius dan pada saat tindak lanjut (pra- Berdasarkan hasil analisis data
kualitatif, yaitu dari hasil observasi dan
wawancara selama proses terapi menun-
jukkan bahwa terapi kognitif perilakuan
religius secara signifikan berpengaruh
dalam meningkatkan kesejahteraan sub-
jektif pada pasien gagal ginjal kronis
yang
menjalani
hemodialisis. terapi Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 102 | 102 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ………. Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……….. kognitif perilakuan religius memberi ban-
tuan kepada pasien untuk dapat meng-
ubah pikiran atau keyakinan negatif,
irasional dan mengalami penyimpangan
menjadi
lebih
positif
dan
rasional
sehingga secara bertahap akan merubah
perilaku menjadi lebih sehat dan normal
(Hepple, 2004). bahwa individu yang menuliskan hal-hal
yang disyukuri memiliki tingkat kese-
jahteraan subjektif yang lebih tinggi. Pada dasarnya religiusitas memiliki
pengaruh terhadap kesehatan mental dan
fisik dimana religiusitas dapat membantu
coping stress, kelelahan dan kematian
pada individu (Pledmont, 2009). Terapi
perilaku dirancang untuk meningkatkan
aktivitas sehingga memunculkan perilaku
yang mampu membuat subjek puas dan
bahagia serta ditujukan dalam kerangka
keyakinan beragama yang diharapkan
akan memunculkan perilaku yang sesuai
dengan yang diniatkan. Hal ini sejalan
dengan penelitian sebelumnya dimana
keyakinan beragama dan praktek beraga-
ma dapat meningkatkan kualitas hidup
dan kesejahteraan subjektif pada maha-
siswa Muslim (Abdel-Khalek, 2010). Teknik perilaku dalam sesi apikasi niat
dalam beraktivitas, relaksasi religius dan
praktiksujud syukur membantu klien
secara lebih mudah dalam pemahaman
materi yang disampaikan. Hal ini juga
berdampak pada menurunnya ketegang-
an
yang
dirasakan
subjek
dalam
menghadapi sakit gagal ginjal kronis
yang dideritanya. Selain itu, peneliti menggunakan
teknik kebersyukuran, serta pernyataan
diri dengan keyakinan agama dalam
proses restrukturisasi kognitif. Hal ini
sejalan dengan kondisi subjek penelitian
yang banyak memiliki pikiran negatif
sehingga memengaruhi subjek dalam
kehidupan sehari-harinya yang lebih
banyak memunculkan afek negatif. Selain
itu, pikiran negatif yang membuat subjek
penelitian menjadi cemas, khawatir,
malu dan lain sebagainya yang berdam-
pak pula pada proses pengobatan subjek. Setelah menyadari bahwa pikiran subjek
tersebut adalah negatif dan memperoleh
pikiran alternatif yang lebih positif,
subjek saat ini mampu meningkatkan
kepuasan hidup dan afek positifnya untuk
lebih semangat dan optimis dalam
menjalani kehidupan. Pada sesi melatih
rasa
syukur,
subjek
diminta
untuk
menuliskan hal-hal yang disyukuri dan
bagaimana cara untuk mensyukurinya. Hal ini sesuai dengan penelitian Emmons
dan McCullough (2003) menemukan Desain penelitian ini menggunakan
aktivitas-aktivitas pembelajaran melalui
pengalaman (experiental learning) yakni
pembelajaran
melalui
pengalaman, Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 103 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini proses pembelajaran menjadi efektif
karena individu mendapatkan stimulasi
yang berulang melalui berbagai indera,
baik penglihatan, pendengaran, penge-
capan, penciuman dan perabaan (John-
son dan Johnson, 2009). Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ………. Pada penelitian
kali ini subjek diberikan tugas rumah
berupa mengontrol aktivitas serta me-
maknainya dan mengidentifikasi pikiran
otomatis yang muncul lalu digantikan
dengan pemikiran alternatif yang lebih
positif yang berguna untuk restukturisasi
kognitif pada subjek dan memaknai hal-
hal yang dapat disyukuri. teraan subjektif pada pasien sakit kronis. Krisnawati (2015) dan Rahmanita (2016)
menjelaskan dalam penelitiannya bahwa
pelatihan kebersyukuran memiliki pe-
ngaruh terhadap kesejahteraan subjektif
pada penderita diabetes mellitus tipe 2
dan pasien hipertensi. Kebersyukuran
merupakan
salah
satu
bagian
dari
religiusitas yang digunakan di sesi terapi
kognitif
perilakuan
religius
dalam
penelitian ini. Keberhasilan terapi kognitif peri-
lakuan religius ini didukung oleh antu-
siasme, motivasi dan penerimaan yang
baik dari subjek. Hal ini dapat dilihat dari
intensitas kehadiran subjek, peran aktif
dalam berpendapat dan memberikan
umpan balik. Hal tersebut juga didukung
berdasarkan hasil evaluasi terapi kognitif
perilakuan
religius
dimana
subjek
mendapat tambahan informasi, wawasan
dengan seluruh informasi yang diberikan
sehingga lebih termotivasi untuk men-
jalankan pengobatan dan pola hidup
yang lebih sesuai dengan anjuran dokter. Para subjek terdorong untuk mem-
biasakan diri mengutamakan pikiran dan
perasaan positif dengan selalu bersyukur
dan pasrah pada setiap kondisi apapun. Berdasarkan hasil data kualitatif
diketahui
pula,
bahwa
peningkatan
kesejahteraan subjektif subjek berbeda-
beda. Hal tersebut terlihat dari pening-
katan nilai kepuasan hidup dan afeksi
yang
berbeda
dari
setiap
subjek. Perbedaan tersebut dipengaruhi oleh
perbedaan dari setiap subjek dalam
memaknai dan mengaplikasikan materi
terapi yang telah diberikan. Ada subjek
yang mempraktekkan materi yang telah
disampaikan secara keseluruhan dan ada
juga subjek yang baru mempraktek-
kannya sebagian, selain itu faktor dari
dukungan sekitar subjek juga meme-
ngaruhi hasil dari penelitian ini. Berdasarkan penjelasan di atas, da-
pat disimpulkan bahwa hipotesis dalam
penelitian ini terpenuhi. Pertama, hasil
penelitian
ini
membuktikan
bahwa Hasil dari penelitian ini juga
mendukung penelitian-penelitian sebe-
lumnya mengenai peningkatan kesejah- | Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 104 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……… terdapat peningkatan nilai kepuasan
hidup pada pasien gagal ginjal yang
menjalani hemodialisis yang mendapat-
kan terapi kognitif perilakuan religius
dibandingkan dengan pasien gagal ginjal
kronis yang tidak mendapatkan per-
lakuan. Kedua, terdapat peningkatan
pada nilai afeksi pada pasien gagal ginjal
kronisyang diberikan terapi kognitif peri-
lakuan religius dibandingkan dengan
pasien gagal ginjal kronis yang tidak
mendapatkan perlakuan. Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ………. Ketiga, terdapat
perbedaan nilai kesejahteraan subjektif
pada pasien gagal ginjal kronis yang
diberikan
terapi
kognitif
perilakuan
religius dibandingkan dengan kelompok
pasien gagal ginjal kronis yang tidak
mendapatkan perlakuan. komunikasi yang aktif dan lebih terbuka. Sedangkan pada saat penentuan subjek
penelitian peneliti menentukan berdasar-
kan kategorisasi kesejahteraan subjektif
dan kesediaan subjek untuk mengikuti
proses terapi. Simpulan Berdasarkan hasil penelitian yang
telah dilakukan, maka dapat disimpulkan
bahwa terapi kognitif perilakuan religius
dapat meningkatkan kesejahteraan sub-
jektif pada pasien gagal ginjal kronis
yang menjalani hemodialisis. Kelompok
yang mendapat terapi kognitif perilakuan
religius lebih tinggi kesejahteraan sub-
jektifnya dibandingan kelompok yang
tidak
mendapatkan
terapi
kognitif
perilakuan religius. Penelitian ini juga memiliki bebe-
rapa kekurangan, seperti jumlah per-
temuan yang masih dirasa kurang oleh
subjek penelitian karena waktu untuk
diskusi perlu ditambahkan lebih lama
lagi. Dalam hal mengontrol subjek
penelitian untuk mengikuti aturan dalam
terapi kognitif perilakuan religius secara
benar juga dirasa sulit karena perubahan
sikap biasanya digerakkan oleh emosi
dengan cara yang positif dan atau negatif
(Wulandari, 2015). Selain itu, bentuk
intervensi yang dilakukan secara ber-
kelompok lebih efektif diberikan kepada
individu yang memiliki kemampuan Saran Penelitian ini telah dilakukan se-
maksimal mungkin, namun tidak menu-
tup kemungkinan masih terdapat keku-
rangan. Oleh karena itu, peneliti mem-
berikan saran sebagai berikut. Pertama:
Saran bagi subjek penelitian. Diharapkan
dapat menerapkan, membiasakan dan
meningkatkan hal-hal yang telah didapat-
kan
dalam
proses
terapi,
seperti
penerapan modifikasi pikiran negatif dan Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 105 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Elda Trialisa Putri, Qurotul Uyun, & Rr. Indahria Sulistyarini mengenali aktivitas. Pada sesi tersebut,
beberapa subjek masih kesulitan untuk
modifikasii pikiran negatifnya menjadi
pikiran alternatif yang lebih positif dan
berlandaskan
nilai
agama. Sehingga
disarankan kepada seluruh subjek untuk
dapat lebih memiliki pemikiran yang
lebih
positif
dan
dapat
memaknai
aktivitas yang dilakukan berlandaskan
nilai agama sehingga subjek dapat
menjalankan
kehidupan
yang
lebih
positif dan optimis. Ketiga: Saran bagi pihak rumah
sakit. Manfaat dalam penelitian ini juga
dapat menjadi masukan bagi setiap pusat
layanan kesehatan bahwa terapi kognitif
perilakuan religius dapat meningkatkan
kesejahteraan subjektif pada pasien gagal
ginjal. Dengan demikian, diharapkan
pihak Rumah Sakit dapat memberikan
kegiatan berkelanjutan bagi para pasien
gagal ginjal lainnya. DAFTAR PUSTAKA Kedua: Saran bagi penelitian selan-
jutnya. Peneliti yang ingin mela-kukan
penelitian serupa, diharapkan (a) dapat
meneliti
terapi
kognitif
perilakuan
religius secara individu untuk melihat
perkembangan subjek dari hari ke hari
dan lebih dapat mengontrol tugas rumah
yang diberikan. (b) Peneliti berharap
untuk terapi kognitif perilakuan religus
berikutnya juga dapat menambahkan
banyaknya jumlah pertemuan sehingga
kesempatan para subjek untuk berdiskusi
jauh lebih besar. (c) Peneliti selanjutnya
juga dapat meneliti kefektifitasan terapi
kognitif perilakuan religius terhadap
pasien penyakit-penyakit lainnya serta
dapat digunakan untuk dapat mening-
katkan variabel lain yang berhubungan
dengan kehidupan pasien sakit kronis
seperti penerimaan diri, kecemasan dan
juga resiliensi. Abdel-Khalek, A.M. (2010). Quality of
life, subjective well-being, and
religiousity
in
muslim
college
students. Quality of Life Research,
19, 1133-1143. Ancok, D & Suroso, F.N. (2011). Psikologi Islami. Yogyakarta: Pus-
taka Pelajar. Azwar. S. (2012). Penyusunan Skala
Psikologi; Edisi 2. Yogyakarta: Pus-
taka Pelajar. Cahyareni, L. (2014). Efektivitas Pelatihan
Kebersyukuran
dengan
Metode
Dzikir untuk Meningkatkan Subjec-
tive Well Being pada Pasien Gagal
Ginjal. Tesis (tidak diterbitkan). Yogyakarta: Fakultas Psikologi dan
Ilmu Sosial Budaya Universitas
Islam Indonesia. Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 Jurnal Intervensi Psikologi Vol. 8 No. 1 Juni 2016 106 Meningkatkan Kesejahteraan Subjektif Pasien Gagal Ginjal Kronis Melalui Terapi ……….. Krisnawati, E. (2015). Efektivitas Pela-
tihan Kebersyukuran untuk Me-
ningkatkan Kesejahteraan Subjektif
pada Penderita Diabetes Mellitus
Tipe II. Tesis (tidak diterbitkan). Yogyakarta: Fakultas Psikologi dan
Ilmu Sosial Budaya Universitas
Islam Indonesia Emmons,
R.A.,&
McCullough,
M.E. (2003). Counting blessings versus
burdends: an experimental investi-
gation of gratitude and subjective
well being in daily life. Journal of
Personality and Social Psychology,
84 (2),377-389. Foreze, Martin, Patton, Zadeh and
Kopple, (2010) Mansjoer, A. 2000. Kapita Selekta Kedok-
teran. Jakarta: Media Aesculapius. Hepple, J. (2004). Psychoterapies with
older people: an oveview. Advance
in Psychiatric Treatment, 10, 371-
377 McCullough, M.E & Larson, D. B. (1999). Prayer, In W, R. Miller (Ed),
Intergrating Spirituality into Treat-
ment; Resourcesfor Practitioners. Washington,
D. C:
American
Psychological Association Hida,S.,
Suroso,
&
Muhid,A.(2013). Hubungan
antara
Religiusitas
dengan Subjective Well Being
melalui Kemampuan Penyesuaian
diri pada Akhir Dewasa Madya. academia.edu
http://www.academia.edu/4472682
/Hubungan_antara_Religiusitas_de
ngan_Subjective_Well_Being_mela
lui_Kemampuan_Penyesuaian_diri
_pada_akhir_dewasa_madya
(diunduh 3 Mei 2015) Pledmont, R.L. (2009). The Contribution
of Religiousness and Spirituality to
Subjective Well-being and Satis-
faction with Life(Chapter 5). In
Souza, M., Francis, L.J., O’Higgins-
Norman, J., & Scott, D.G (Edited). DAFTAR PUSTAKA International Handbook of Edu-
cation for Spirituality, Care and
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89-105.New
York:
Springer Johnson, D.W & Johnson, F.P. (2009). Joining Together: Group Therapy
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Jersey: Person Education Inc Rahmanita, A. (2016). Efektivitas Pelatih-
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katkan
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pada Penderita Penyakit Hiper-
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(tidak
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Relaksasi Zikir untuk Menurunkan
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menjalani
Hemodialisis di RSUD X. Tesis
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Seed testing of foliar-fertilised red clover crops after various periods of storage
|
Notulae botanicae Horti Agrobotanici Cluj-Napoca
| 2,020
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cc-by
| 6,493
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Tomic D et al. (2020)
Notulae Botanicae Horti Agrobotanici Cluj-Napoca 48(1):284-293
DOI:10.15835/nbha48111814
Research Article
Seed testing of foliar-fertilised red clover crops
after various periods of storage
Dalibor TOMIĆ1*, Vladeta STEVOVIĆ1, Dragan ĐUROVIĆ1, Rade
STANISAVLJEVIĆ2, Milomirka MADIĆ1, Mirjana PETROVIĆ3,
Đorđe LAZAREVIĆ3, Jasmina KNEŽEVIĆ4
1University of Kragujevac, Faculty of Agronomy, Cara Dušana 34, 32000 Čačak, Serbia; dalibort@kg.ac.rs (*corresponding author)
2Institute for Plant Protection and Environment, Teodora Drajzera 9, 11 000 Belgrade, Serbia; stanisavljevicrade@gmail.com
3Institute of Forage Crops, +38137251 Globoder, Kruševac, Serbia; mirjana.petrovic@ikbks.com
4Univerzity of Priština, Faculty of Agriculture, Kopaonička bb, 38219, Lešak, Serbia; jasmina.kneska@gmail.com
Abstract
AcademicPres
Notulae Botanicae Horti
Cluj-Napoca
Agrobotanici Tomic D et al. (2020)
Notulae Botanicae Horti Agrobotanici Cluj-Napoca 48(1):284-293
DOI:10.15835/nbha48111814
Research Article AcademicPres AcademicPres 1University of Kragujevac, Faculty of Agronomy, Cara Dušana 34, 32000 Čačak, Serbia; dalibort@kg.ac.rs (*corresponding author)
2Institute for Plant Protection and Environment, Teodora Drajzera 9, 11 000 Belgrade, Serbia; stanisavljevicrade@gmail.com
3Institute of Forage Crops, +38137251 Globoder, Kruševac, Serbia; mirjana.petrovic@ikbks.com
4Univerzity of Priština, Faculty of Agriculture, Kopaonička bb, 38219, Lešak, Serbia; jasmina.kneska@gmail.com Abstract The aim of the present study was to analyse the effect of foliar fertilisation of red clover (Trifolium
pratense L.) crops with cobalt, boron and phosphorus-potassium, which could significantly increased seed yield,
on seed quality after different periods of storage. The experiment involving four red clover cultivars and four
foliar treatments was laid out in a randomised block design with four replications and with a plot size of 5 m2. Red clover cultivars ‘K-37’, ‘K-39’, ‘Una’ and ‘Viola’ were planted at a seeding rate of 18 kg ha-1. Foliar
fertilisation with cobalt, boron and phosphorus-potassium was carried out during intensive growth (at BBCH
34-36) in the second year of cultivation. After all storage periods, red clover cultivars differed in root length,
seedling mass and hard seededness under all foliar fertilisation treatments. In all storage periods, foliar
treatments did not affect germination, stem length, seedling mass and percentage of hard seed in any cultivar
(no significant cultivar/foliar treatment interaction). At 20 months of storage, there were no significant
differences between foliar treatments and the unfertilised control in any of the studied traits. Foliar nutrients
were applied prior to the emergence of generative organs; therefore, the absence of their positive effect on seed
development can be associated with their overall short-term effect. The relationship between seed quality and
the content of mineral nutrients in the seed after their foliar application at different stages of crop development
should be a topic for future research. Keywords: foliar fertilisation; red clover; seed quality parameters Received: 14 Feb 2020. Received in revised form: 15 Mar 2020. Accepted: 19 Mar 2020. Published online: 31 Mar 2020. Introduction Important indicators of legume seed quality are germination, hard seededness and seedling growth
parameters. Germination and seedling emergence are critical growth stages in plant development, which
determine crop density, growth rate and final yield (Desai et al., 1997). Seedling growth parameters are
important seed quality components which affect the success of crop establishment, growth and productivity of Received: 14 Feb 2020. Received in revised form: 15 Mar 2020. Accepted: 19 Mar 2020. Published online: 31 Mar 2020. Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 285 plants (Sawan et al., 2009). Dormancy is the inability of a viable seed to germinate immediately after harvest,
even under favourable conditions (Copeland and McDonald, 1995; Aydin and Uzun, 2001). Hard seededness
is a special type of dormancy found only in legumes, and in species of the genus Trifolium it is mainly the
consequence of seed coat impermeability to water (Desai et al., 1997; Ates, 2011). With aging, the seed loses
hard seededness (Asci et al., 2011). Depending on plant species, the release of hard seededness may take from a
few days to several years (Baskin and Baskin, 2004). y
y
(
)
Outside their optimal range, abiotic and biotic factors can have a negative effect on seed germination
and seedling emergence, especially under stressful conditions e.g. acid soils, insufficiently cultivated top soil,
low temperatures, excess or lack of water, pathogenic soil microorganisms, seed damage by pests and chemicals
(Sawan et al., 2009). Proper mineral nutrition, in particular macro- and micronutrients necessary for the
formation of generative organs, seed setting and filling, may have a positive effect on yield and seed quality. In
Southeast Europe, red clover seed crops are commonly established on acid soils, where certain macro- and
micronutrients are less available to plants. As determined by Tomić et al. (2011; 2014; 2015), on such soils,
foliar fertilisation of red clover crops with cobalt, boron, phosphorus and potassium can positively affect seed
yield. Cobalt has an important role in a number of physiological processes (Mathur et al., 2006; Jayakumar et
al., 2007; Jayakumar and Jallel, 2009). In addition, cobalt is the central atom in the porphyrin structure of
cobalamin coenzyme and is essential for the nodulation and development of bacteroids (Collins and Kinsela,
2011). Introduction Given the low mobility of Co in the plant (Austenfeld, 1979) and its faster movement from
aboveground parts to the root rather than in the opposite direction (Danilova et al., 1970; Palit et al., 1994),
foliar treatment is assumed to lead to a more efficient and optimal status of cobalt in the root and in the nodules,
in comparison with soil application. Boron is actively involved in cell division, flowering, fruit ripening,
nitrogen and carbohydrate metabolism, plant resistance to diseases, and in many other reactions in which it
acts as a catalyst (Goldbach et al., 2001; Farooq et al., 2011). Accordingly, its deficiency causes a significant
decline in the seed yield of many plant species. High concentrations of aluminium ions in acidic soils limit
boron uptake (Matsumoto, 2000; Yau, 2000). Boron is relatively immobile in plants, and thus its availability is
essential at all stages of growth, especially during seed development (Waqar et al., 2012). The authors also
emphasise that research for improving crop yields must move beyond applications of B based on general
recommendations, and that deriving methods to predict site-specific deficiencies (e.g. soil or plant tests) are
essential since the potential for B toxicity is large and the difference between deficiency and toxicity is very
narrow. Phosphorus and potassium are necessary for the growth of both vegetative and generative organs in
leguminous plants (Hégh-Jensen et al., 2001; Abbasi et al., 2016). Phosphorous is easily fixed by clay minerals
that are abundant in acidic soils, including various iron oxides and kaolinite, hence rendering it unavailable for
root uptake (Zheng, 2010). In this research, it was assumed that foliar treatments with cobalt, boron, phosphorus and potassium on
an acidic soil would increase seed yield and seed quality of red clover cultivars differing in total biomass
production. Their effect on seed yield was analysed in earlier studies (Tomić et al., 2011; 2014; 2015). The
objective of this study was to evaluate major quality characteristics, i.e. germination, seedling growth parameters
and hard seededness of the seed obtained from foliar-fertilised red clover crops, after various periods of storage. Soil properties A field experiment was established at a trial field of the Veterinary Extension Service in Čačak, Serbia
(43°54'39.06" N, 20°19'10.21" E, 246 m a.s.l.) in 2009-2011. The trial was established on a leached vertisol
acidic in reaction (pH H2O 4.8), which contained 3.2% organic matter. The soil was well supplied with Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 286 phosphorus (22.1 mg 100 g-1) and potassium (30.0 mg 100 g-1 soil), had a moderate content of cobalt (27.2 mg
kg-1 soil), low levels of calcium (0.01% CaCO3) and boron (0.06 mg kg-1 soil), and a relatively high
concentration of aluminium ions (12.0 mg kg-1 soil). The preceding crop was natural meadow. In autumn, in
the year before sowing, 45 kg ha-1 N, 45 kg ha-1 P2O5 and 45 kg ha-1 K2O (NPK 15:15:15) were incorporated
into the soil through primary tillage. Measurements Analyses were performed in three periods i.e. one, eight and twenty months after seed harvest. In
Southeast Europe, red clover seed is harvested mostly from the second cut, in late July and early August. In this
region, red clover sowing in autumn is performed from the end of August to late September, when the seed is
approximately one month old. Sowing in the spring of the following year begins after eight-month storage of
seeds from the previous year. Twelve months later (at the time of the following spring sowing), the seed is about
twenty months old. In these periods, four replicates of 100 seeds per treatment and field replication were
germinated on filter paper, according to the ISTA Rules (ISTA, 2011). Seeds were germinated at 20 °C in dark
germination cabinets. To overcome hardseededness, seeds were subjected to precooling at 3 °C for 7 days (Uzun
and Aydin, 2004). Final germination and hardseededness were determined 10 days later by identifying the
number of normally germinated seeds and number of hard seeds, respectively, and expressing them as
percentages. Measurement of seedling growth parameters (primary root length, shoot length and fresh seedling
biomass, i.e. root and shoot) was done after final germination. Seedling length was measured by a ruler and fresh
seedling biomass was determined using an analytical balance (Shimadzu, Japan). At the end, the sum of total
root length, total shoot length and total seedling mass per replication were calculated. Considering large
differences in germination percentage among seeds after one, eight and twenty months of storage, the values of
primary root length, shoot length and fresh seedling biomass for both years and treatments (Table 1) are
expressed as mean values per seedling. Experimental design The experiment, involving treatments as combinations of four red clover cultivars and four foliar
fertilisation variants (unfertilised, cobalt, boron and phosphorus-potassium), was laid out in a randomised
block design with four replications and with a plot size of 5 m2. Four red clover cultivars, differing in total
biomass production and leaf proportion, namely ‘K-17’, ‘K-39’ (Institute of Forage Crops, Kruševac, Serbia),
‘Una’ (Institute of Field and Vegetable Crops, Novi Sad, Serbia) and ‘Viola’ (a Polish cultivar) were planted in
two consecutive years (2009 and 2010, on 28 and 30 March, respectively) at a row spacing of 0.2 m and a
seeding rate of 18 kg ha-1. Foliar treatments included: unfertilised control; cobalt applied as Co(NO3)2 at a
concentration of 0.033g L-1 with a water rate of 1000 L ha-1 in two applications, one at the stage of intensive
growth, i.e. stage 34-36 of the BBCH scale (Meier, 2001) during the first cut and two applications during the
second cut in the second year of cultivation - the first at BBCH 34-36; active vegetative growth, and the second
at BBCH 51-55; just before flowering; boron (Boron-feed, Haifa, Israel) applied at a concentration of 0.1%,
with a water rate of 1000 L ha-1, at the stage of intensive growth during the second cut; and phosphorus
combined with potassium (P52K34, Haifa, Israel), applied at a concentration of 1% with a water rate of 1000 L
ha-1, at the same growth stage as boron. Boron and phosphorus fertilisers were applied at the manufacturer's
recommended concentrations, while the application rate of cobalt was used according to Vranný (1978). Seeds were collected at full maturity (12% seed moisture content) in July 2010 and July 2011 in the
second year of cultivation. Harvested seeds were cleaned manually, placed into paper bags and stored at room
temperature (15-20 °C during summer, and 5-15 °C in winter) and at a relative humidity below 50%. Results Averaged across years, cultivars and foliar treatments, significantly higher seed germination, resulting
from the significantly lower number of hard seeds, was recorded in seeds after eight and twenty months of
storage, compared with seeds after one-month storage (Table 1). After one month of storage, the proportion of hard seeds was 52.8% on average for all cultivars, this
being the main reason for the very low germination percentage (24.8%). After eight months’ storage, germination percentage was increased to 89.6% and the proportion of hard
seeds was reduced to 1.6%. After eight months, seedling growth parameters didn’t change at seeds that were
stored for one month (P>0.05) (Table 1). At the same time, the proportion of dead seeds and abnormal
seedlings was reduced by more than twice. Germination percentage and seedling growth parameters were not
significantly affected by the prolongation of the storage period, i.e. there were no differences between eight-
month and twenty-month storage periods. However, the proportion of hard seeds was reduced more than
twice, although the difference was not significant. For each storage period, there were also differences among cultivars in germination percentage (except
for one-month storage), root length, stem length (only for one-month storage), seedling mass and the number
of hard seeds (Table 2). These differences among cultivars were consistent across foliar fertilisation treatments
(the absence of the cultivar/fertilisation interaction) in all storage periods. Foliar fertilisation with cobalt in this experiment had no effect on germination percentage (Table 2),
regardless of storage period. Seeds from foliar cobalt treatment had a significantly greater seedling root length
compared to the unfertilised crop after one-month storage; eight-month-old seeds had significantly lower
values, whereas no significant differences were observed in seeds after twenty months of storage. Foliar-applied boron did not affect germination percentage, regardless of the length of the storage period
(Table 2). Germination percentage was significantly higher after boron treatment than after cobalt treatment
in eight-month-old seeds and phosphorus-potassium treatment in seeds after twenty-month storage. Seedling
root length was significantly greater after foliar boron application than in the unfertilised control only after one
month of storage. Significantly greater seedling mass was observed following boron treatment compared to
cobalt treatment and the unfertilised control after eight months of storage and compared to phosphorus-
potassium treatment after twenty months of storage. Foliar fertilisation with phosphorus-potassium had no significant effect on germination percentage and
seedling growth parameters in any storage period (Table 2). Statistical analysis The obtained results were subjected to a two-factor analysis of variance (cultivars and foliar fertilisation
as fixed effects) using SPSS (SPSS 1995). Given that there were no significant differences between years, as well Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 287 as significant interactions between year and other factors, the analysis of variance was performed using average
values. To correct for normality, the statistical analysis was performed on √arcsine transformed values for fnal
germination and on √x+1 for hard seededness. Te signifcance of diferences between storage periods was
tested by t-test, and between cultivars and foliar fertilisation treatments by LSD test. Results Positive correlations were observed among germination percentage, stem length, root length and
seedling mass, while, expectedly, the proportion of hard seeds was negatively correlated with all other
characteristics (Table 3). Table 1. Mean values of germination percentage, stem length, fresh seedling mass and dormant seed of red
clover depending on storage period (average for years, cultivars and foliar treatments)
Storage
period
Germination
(%)
Stem length
(mm)
Root length
(mm)
Seedling mass
(mg)
Hard seed
(%)
Dead
seed/abnormal
seedling (%)
1 month
24.8±1.45 b
36.3±2.67 a
37.2±2.76 a
27.7±2.47 a
52.8±1.86 a
22.4±1.26 a
8 months
89.6±0.85 a
35.3±0.72 a
35.2±1.05 a
25.6±0.67 a
1.58±0.33 b
8.8±0.53 b Table 1. Mean values of germination percentage, stem length, fresh seedling mass and dormant seed of red
clover depending on storage period (average for years, cultivars and foliar treatments) Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293
288
20 months
88.4±0.96 a
38.7±1.27 a
35.7±1.29 a
25.9±0.45 a
0.67±0.18 b
10.9±0.57 b
Data represent mean ± s.d. Mean values in each storage period followed by the same letter are not significantly diferent
(P > 0.05). Table 2. Results Mean values of seed quality traits of red clover cultivars (‘K-39’, ‘K-17’, ‘Una’, ‘Viola’) depending
on foliar fertilisation (unfertilised, cobalt – Co, boron – B, phosphorus-potassium - PK) in different storage
periods (n=48)
Storage
period
Cultivar/
Foliar
fertilisation
Germination
(%)
Stem length
(cm)
Root length
(cm)
Seedling mass
(g)
Hard seed
(%)
One
month
‘К-39’
28.8±3.66
58.5±8.63 a
53.8±10.16 a
0.441±0.06 a
49.2±5.08 b
‘К-17’
22.8±2.57
39.3±5.15 b
46.1±6.15 ab
0.321±0.04 b
52.5±3.88 ab
‘Una’
23.0±2.28
38.8±6.05 b
49.1±5.37 ab
0.314±0.03 b
57.2±2.01 a
‘Viola’
24.5±2.91
44.1±5.30 ab
36.5±4.22 b
0.298±0.04 b
52.3±3.33 ab
P1
ns
*
*
**
*
Unfertilised
23.2±2.61
45.6±8.63
34.2±2.28 b
0.326±0.04
54.3±3.72 ab
Co
27.0±2.54
49.9±5.70
54.5±6.35 a
0.362±0.04
51.5±3.19 ab
B
27.0±3.78
45.5±7.28
59.2±9.86 a
0.389±0.06
47.0±3.66 b
PK
22.0±2.52
39.5±4.87
37.7±4.26 b
0.299±0.03
58.3±3.91 a
P
ns
ns
***
ns
*
Eight
months
‘К-39’
92.83±1.64 а
157.4±5.51
173.4±7.71 а
1.258±0.05 а
1.33±0.62 аb
‘К-17’
88.17±1.88 b
161.5±3.87
146.0±7.25 b
1.157±0.08 a
0.33±0.22 b
‘Una’
90.17±1.78 ab
164.7±9.87
170.1±9.33 a
0.989±0.04 b
2.00±0.69 a
‘Viola’
87.17±1.14 b
149.7±4.61
142.1±10.60 b
1.184±0.06 a
2.67±0.79 a
P
*
ns
**
**
*
Unfertilised
89.67±1.67 ab
150.4±5.50
166.0±9.15 a
1.082±0.06 bc
1.17±0.57
Co
86.01±1.83 b
159.2±4.91
142.3±9.48 b
1.019±0.03 c
2.50±0.89
B
92.52±1.56 a
159.0±3.41
158.4±9.39 ab
1.285±0.08 a
1.00±0.46
PK
90.17±1.38 ab
164.8±10.0
164.8±9.39 ab
1.202±0.06 ab
1.67±0.59
P
*
ns
*
**
ns
Twenty
months
‘К-39’
91.5±1.44 a
164.8±12.8
173.4±7.71 a
1.086±0.03 c
0.5±0.26 ab
‘К-17’
91.17±2.17 a
175.6±14.7
144.2±7.21 ab
1.191±0.04 b
0.17±0.17 b
‘Una’
87.83±1.45 ab
173.6±9.48
157.3±15.8 ab
1.284±0.05 a
0.67±0.28 ab
‘Viola’
83.17±1.69 b
180.6±8.42
142.1±10.6 b
1.021±0.04 c
1.33±0.57 a
P
**
ns
*
***
*
Unfertilised
88.17±2.33 ab
163.5±9.16
151.6±15.45
1.12±0.03 ab
1.17±0.57
Co
89.83±1.66 a
189.3±13.9
142.4±9.48
1.174±0.04 ab
0.67±0.28
B
91.0±1.77 a
166.9±12.4
158.4±9.39
1.185±0.07 a
0.17±0.17
PK
84.67±1.56 b
174.9±9.22
164.8±9.39
1.103±0.04 b
0.67±0.28
P
*
ns
ns
*
ns
Data represent mean ± s.d. Mean values in each storage period for cultivars and foliar fertilisation treatments followed 288 288
20 months
88.4±0.96 a
38.7±1.27 a
35.7±1.29 a
25.9±0.45 a
0.67±0.18 b
10.9±0.57 b
Data represent mean ± s.d. Mean values in each storage period followed by the same letter are not significantly diferent
(P > 0.05). Table 2. Data represent mean ± s.d. Mean values in each storage period for cultivars and foliar fertilisation treatments followed
by the same letter are not significantly diferent (P > 0.05). 1 P indicates the significance of the cultivar and foliar
fertilisation efects in the analysis of variance. ns – non-significant, * - P < 0.05, ** - P < 0.01, *** - P < 0.001 Results Mean values of seed quality traits of red clover cultivars (‘K-39’, ‘K-17’, ‘Una’, ‘Viola’) depending
on foliar fertilisation (unfertilised, cobalt – Co, boron – B, phosphorus-potassium - PK) in different storage
i d (
48) Table 3. Coefficients of correlation among germination rate, stem length, root length and seedling mass
(n
144) Table 3. Coefficients of correlation among germination rate, stem length, root length and seedling mass
(n = 144)
Traits
Stem length
(cm)
Root length
(cm)
Seedlings mass
(g)
Hard seed
(%)
Germination (%)
0.90*
0.87*
0.94*
-0.96*
Stem length (cm)
0.76*
0.85*
-0.88*
Root length (cm)
0.80*
-0.84* Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 289 Discussion Hard seededness is a common trait in species of the genera Medicago and Trifolium. Some ecotypes of
species belonging to these genera, immediately after harvest, may have hard seededness at a rate of up to 100%
(James et al., 1970; Crawford et al., 1989; Aydin and Uzun, 2001; Can et al., 2009). Colgecen et al. (2008)
found the germination percentage of 13.5% immediately after harvest in red clover seeds from natural
populations. The higher germination percentage in this research may be due to the agro-environmental
conditions of the experimental location, cultivar or, possibly, seed chilling before the germination test was
performed. Smýkal et al. (2014) observed that, under natural conditions, in temperate-zone species, there are two
steps in breaking dormancy by low winter temperatures. During the first step, low winter temperatures make
seeds sensitive to alternating temperatures, and during the second step, these alternating regimes occurring in
early spring cause the sensitive seeds to become water-permeable. Seeds can only respond to alternating spring
temperatures if they become sensitive during the winter (Baskin and Baskin, 2014). Accordingly, the authors
pointed out that several artificial techniques are used to break physical dormancy in seeds, such as dry storage
i.e. low temperatures. In the present experiment, during the winter, seeds were kept at low temperatures (from +5 to +15 ºC);
along with ageing, this chilling treatment reduced hard seededness. Consequently, in eight- and twenty-month-
old seeds, there was an increase in germination percentage and seedling growth parameters, as well as a
significant reduction in the proportion of hard seeds, especially in twenty-month-old seeds. Balouch and
Modarres (2006) reported that the proportion of hard seeds in red clover decreased and, consequently,
germination percentage increased with ageing and also after chilling treatment. Differences among cultivars in germination percentage, root length, seedling mass and the number of
hard seeds in any storage period can be attributed to the high genetic variability of red clover as a species (Steiner
et al., 1995; Helgadottir, 1996; Zielinska et al., 2012). Acharya et al. (1999) reported that the percentage of
hard seededness in alfalfa largely depends on cultivar. Also, when studying the influence of different pre-sowing
treatments on dormancy breakdown, germination and vigour of red clover seeds, Simić et al. (2018) observed
significant differences in the proportion of hard seeds among red clover cultivars. Discussion In the future, as more emphasis is placed on seed quality, the need for the classification of cultivars based
on seed quality traits will increase (Zielinska et al., 2012). As determined by Zeid (2001) and Jayakumar et al. (2007), cobalt content in soil up to 50 mg kg-1 was
progressively increased germination percentage, while higher concentrations negatively affected this trait. The
foliar application of cobalt in this experiment had different effects on seedling root length in seeds after
different periods of storage. Germination percentage in all foliar fertilisation treatments did not significantly differ from the
unfertilised control. This result is in agreement with the findings of Stoltz and Wallenhammar (2013), who,
on a soil with a very low content of boron (<1 mg kg-1), found a smaller (although non-significant) proportion
of ungerminated red clover seeds under soil/foliar boron treatment compared to the control. The authors also
emphasised that higher boron concentrations in seeds increased the percentage of normal seedlings and reduced
the proportion of hard seeds. On the contrary, Ćwintal et al. (2010) on a soil with a very low content of boron
(1.1 mg kg-1) reported a positive effect of foliar boron fertilisation on the germination percentage and
germination energy of red clover seeds. Also, foliar boron application favoured the seed germination and
viability of alfalfa (Medicago sativa L.) seeds on the calcareous loam soil (Dordas, 2006). As indicated by Dos Tomic D et al. (2020). Not Bot Horti Agrobo 48(1):284-293 290 Santos et al. (2004), boron deficiency leads to necrosis of the meristematic tissue and reduces root system
growth in alfalfa. Numerous studies have confirmed the positive effect of boron, applied as a solution for seed
treatment, on the proportion of hard seeds, germination percentage and seedling growth parameters in many
plants (Rerkasem et al., 1997; Farooq et al., 2011; Mirshekari, 2012; Rehman et al., 2012). However, the use of
boron at high concentrations may exhibit a toxic effect (Bonilla et al., 2004; Farooq et al., 2011). In the present
research (except the second cobalt treatment), foliar treatments were applied at BBCH 34-36. The period from
this stage to the beginning of flowering (at BBCH 60-62) under given agro-environmental conditions,
depending on the sum of active temperatures, is 14–25 days. Overall, the positive effect of foliar-applied
mineral nutrients is time-limited, lasting generally up to 15 days (Brown and Shelp, 1997). Conclusions Seeds of all cultivars had low germination percentages after one month of storage, due to the high
proportion of hard seeds. In all foliar fertilisation treatments, red clover cultivars differed in hard seededness,
root length and seedling mass at all storage periods, but their differences were inconsistent. In any storage
period, foliar treatments did not affect germination, stem length, seedling mass and percentage of hard seed in
any cultivar (no significant cultivar/foliar treatment interaction). At 20 months of storage, there were no
significant differences between foliar treatments and the unfertilised control in any of the studied traits. Foliar
nutrients were applied prior to the emergence of generative organs; therefore, the absence of their positive effect
on seed development can be associated with their overall short-term effect. Further research should focus on
the relationship between seed quality and the content of macro- and micronutrients in the seed under different
crop nutrient management practices, especially their foliar application at different stages of plant growth. Discussion The period from
fertilisation to physiological maturity (at BBCH 85) in red clover is 25 days on average (Angsumalee et al.,
2016). This may explain the absence of significant effect of foliar boron fertilisation on seed quality traits. The positive effect of phosphorus and potassium, applied as a solution for seed treatment before
germination, on germination percentage and subsequent stages of plant development, as well as on total yield
in many plant species, has been shown in numerous reports (Kalita et al., 2002; Yang et al., 2004; Zeļonka et
al., 2005; Ali et al., 2008). In this study, foliar fertilisation with phosphorus-potassium could not have a
significant effect on germination percentage and seedling growth parameters in any storage period. As with
boron, the absence of the positive effect of these nutrients on seed quality can be associated with their earlier
application. The positive effect of foliar-applied cobalt, which was also supplied immediately before flowering,
on red clover seed yield has been reported by Tomić et al. (2014). However, the available literature provides no
data on the effect of cobalt on seed quality traits after its foliar application. Acknowledgements This work is part of the research project Ref. No. TR-31016 funded by the Ministry of Education,
Science and Technological Development, Republic of Serbia. Conflict of Interests This work is part of the research project Ref. No. TR-31016 funded by the Ministry of Education
Science and Technological Development, Republic of Serbia.
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copyright/to retain publishing rights without restriction. The journal offers free, immediate, and unrestricted access to peer-reviewed research and scholarly work. Users are
allowed to read, download, copy, distribute, print, search, or link to the full texts of the articles, or use them for any
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License - Articles published in Notulae Botanicae Horti Agrobotanici Cluj-Napoca are Open-Access,
distributed under the terms and conditions of the Creative Commons Attribution (CC BY 4.0) License.
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106:183-184. Zielinska M, Zapotoczny P, Białobrzewski I, Zuk-Golaszewska K, Markowski M (2012). Engineering properties of red clover
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(Trifolium pratense L.) seeds. Industrial Crops and Products 37:69-75. © Articles by the authors; UASVM, Cluj-Napoca, Romania. The journal allows the author(s) to hold the
copyright/to retain publishing rights without restriction.
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Fast Augmented Reality Authoring: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations
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Fast Augmented Reality Authoring: Fast Creation
of AR Step-by-Step Procedures for
Maintenance Operations Fast Augmented Reality Authoring: Fast Creation
of AR Step-by-Step Procedures for
Maintenance Operations RICCARDO PALMARINI1, IÑIGO FERNÁNDEZ DEL AMO1, DEDY ARIANSYAH1,
SAMIR KHAN1, (Member, IEEE), JOHN AHMET ERKOYUNCU
1,
AND RAJKUMAR ROY
2 1Cranfield Manufacturing, School of Aerospace, Transport and Manufacturing, Cranfield University, MK43 0AL Cranfield, U.K. 2School of Mathematics, Computer Science and Engineering, City, University of London, EC1V 0HB London, U.K. Corresponding author: John Ahmet Erkoyuncu (j.a.erkoyuncu@cranfield.ac.uk) is work was supported in part by the High-Speed Sustainable Manufacturing Institute (HSSMI), and in part by the Engineering a
ysical Sciences Research Council (EPSRC) by the Project ‘‘Digi-TOP’’ under Grant EP/R032718/1. This work involved human subjects or animals in its research. Approval of all ethical and experimental procedures and protocols was
granted by Cranfield University. ABSTRACT Augmented Reality (AR) has shown great potential for improving human performance in
Maintenance, Repair, and Overhaul (MRO) operations. Whilst most studies are currently being carried out at
an academic level, the research is still in its infancy due to limitations in three main aspects: limited hardware
capabilities, the robustness of object recognition, and content-related issues. This article focuses on the last
point, by proposing a new geometry-based method for creating a step-by-step AR procedure for maintenance
activities. The Fast Augmented Reality Authoring (FARA) method assumes that AR can recognise and track
all the objects in a maintenance environment when CAD models are available, to knowledge transfer to a
non-expert maintainer. The novelty here lies in the fact that FARA is a human-centric method for authoring
animation-based procedures with minimal programming skills and the manual effort required. FARA has
been demonstrated, as a software unit, in an AR system composed of commercially available solutions
and tested with over 30 participants. The results show an average time saving of 34.7% (min 24.7%; max
55.3%) and an error reduction of 68.6% when compared to the utilisation of traditional hard-copy manuals. Comparisons are also drawn from performances of similar AR applications to illustrate the benefits of
procedures created utilising FARA. INDEX TERMS Augmented reality, authoring, content, digital engineering, maintenance. his work is licensed under a Creative Commons Attribution 4.0 License. For more information, see https://creativecommons.org/licenses/by/4.0/ Received 19 December 2022, accepted 30 December 2022, date of publication 24 January 2023, date of current version 27 January 2023. Received 19 December 2022, accepted 30 December 2022, date of publication 24 January 2023, date of current version 27 January 2023. Digital Object Identifier 10.1109/ACCESS.2023.3235871 VOLUME 11, 2023
This work is licensed under a Creative Commons Attribution 4.0 License. For more information, see https://creativecommons I. INTRODUCTION range of fields: medical applications, marketing, entertain-
ment, education, maintenance, and manufacturing [3], [4]. This article focuses on Maintenance, Repair, and overhaul
(MRO) operations. MRO operations impact the lifecycle cost
of industrial equipment [5]. The increasing complexity and
automation of industrial machinery require new technolo-
gies for ensuring reliability and productivity through MRO
operations. In the aviation field, MRO operations costs can
reach 80% of the entire aircraft lifecycle [6]. MRO operations
strongly rely on maintenance technicians’ expertise [7]. The
latter can affect both the errors and completion time involved AUGMENTED Reality (AR) is an innovative technology that
aims to enhance the human perception of reality by providing
digitally created content in the real context [1]. Another
definition has been provided by Azuma [2] who stated that
an AR system should have three characteristics: Combine real
and virtual, 2) Interactive real-time, and 3) Registered in 3-D. AR applications have been developed and tested in a wide The associate editor coordinating the review of this manuscript and
approving it for publication was Yu Liu
. 8407 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations and expensive [15]. The contents are now implemented in
AR as ‘‘standalone’’ programs by programmers [16]. Com-
prehensive reviews of AR authoring for maintenance appli-
cations have been done [10]. The remaining paragraphs
highlight some of the existing efforts for content creation and
identify the crucial gap of AR authoring for industrial imple-
mentation to be addressed in this work. The main knowledge
gap is that current authoring environments require com-
prehensive programming and graphical expertise. The
most common tools for authoring AR content comprise plug-
ins, software development kits (SDK), and graphical pro-
gramming languages. Among these, it is worth mentioning
Unity, Unreal, Panda3D, ArToolkit, Vuforia, and Max/MPS. in the MRO operation thus influencing the MRO cost. AR can
help reduce errors and completion time by allowing easy
access to MRO information which today belongs mostly
to the expert maintainers’ memory [8]. Even though the
advantages (time savings and error reductions) of AR in
maintenance have been proven by academics, the technology
still lacks the robustness and flexibility to become of com-
mon use. I. INTRODUCTION Among the main research topics, it is possible to
find [9], [10], [11]: • Tracking and recognition of performances • Hardware (head-mounted displays) capabilitie • Contents-related issues The last one comprises difficulties in creating and manag-
ing content for AR applications. The traditional process of
creating content (a.k.a. authoring) for AR requires different
professionals: programmer, animator, CAD modeller, and
AR developer. More innovative authoring solutions which
provide a friendly user interface and content adaptation have
also been proposed ([12], [13], [14]). However, they still
require a lot of human effort and have limited flexibility. This
article contributes to this gap by implementing an AR within
a maintenance application. For authoring ‘‘AR step-by-step’’
procedures to guide a non-expert technician in carrying out a
maintenance task ‘‘Step-by-step AR instructions’’ or ‘‘proce-
dures’’ is a common terminology which refers to the action
of gradually providing a set of information at each step of
an MRO operation. The data considered in this project is
visual (3D animations). The method developed and validated
has been named FARA: Fast Augmented Reality Authoring. The novelty of this work lies in the approach that FARA
is a humancentric method for authoring animation-based
procedures with minimal programming skills and manual
effort required when learning to use the software features
for creating step-by-step AR instructions. It takes inspi-
ration from Fast Programming Robots that enable users to
teach robots by demonstration. Similarly, FARA allows users
to teach an AR system how to overlay the virtual content into
a real environment without programming. Nowadays, only a few have attempted to ease and de-skill
the authoring process. Shim et al proposed an interactive
features-based AR authoring tool [17]. This allows users to
rotate, move, enlarge merge and occlude virtual objects the
virtual objects visualized over a 2D printed marker. The men-
tioned transformations are done through marker interaction
and gesture interaction. Similarly, Yang et al proposed an
authoring tool that takes advantage of a mobile device to inter-
act with the virtual objects visualized through an HMD [18]. Both approaches do not require any programming skills and
are not time-consuming for the content creator. Still, the
solution does not allow the creation of animation, which is
powerful in the maintenance environment [6], [19].1 Csurka
et al proposed SUGAR, as an easy-to-use AR editor that does
not require programming skills [20]. 1It must be noted that step-by-step animations might not always be
required. I. INTRODUCTION However, part of the
creation of the content must be done through the SUGAR
editor. The latter requires the content creator to input the
picture of the working environments and manually create or
import the virtual objects and animation that users want to
over-impose on the real environment at each step of the main-
tenance procedure. Even though the advantages compared
with the traditional authoring methods have been proven,
it is the authors’ opinion that most methods are still time-
consuming. Zhu et al proposed an on-site authoring tool that
allows maintenance technicians to change or create informa-
tion instances related to maintenance procedures [21]. This
means that only text information can be created and edited. Other authoring solutions have been used or proposed in
the literature [22], [23], [24], [25], [26], [27]. Using these
authoring solutions requires some knowledge and a manual
process in understanding how to use the software features
such as creating and positioning the 3D animation on top of
the real component. This article is structured as follows. The research back-
ground and motivation are provided in Sections I and II. This
is followed by a description of FARA: how it works and its
structure and the detailed methodology for FARA’s valida-
tion. This includes the quantitative test design (Section IV-A),
the case study utilised (Section IV-B), and FARA’s implemen-
tation in an AR system for testing purposes (Section IV-C). Analysis and results are reported in Section IV-C. Finally, the
discussion of the results and the conclusions are presented in
Sections V and VI. Taking advantage of the valuable contributions of previous
studies, the aim of simplifying the authoring process of AR
procedures should remain and will likely help to implement
AR in the industry. The availability of AR authoring tools that
can be operated by non-programmers and non-AR experts is
essential for the success of AR technology in both the main-
tenance field and other areas [15]. The approach proposed A. HOW FARAIS WORKS: A PRACTICAL EXAMPLE The FARAIS shown in this example has been developed using
commercial hardware and the open-source limited versions
of Vuforia and Unity 3D software. These will be described
in Section IV-C. The procedure selected is the assembly of
a mock-up designed and utilised for testing purposes. It will
be described in Section IV-B. First, consider the maintenance
environment shown in Fig 1. It includes the maintainer (Nr.1),
the product to be maintained (Nr.2) and FARAIS (Nr.3). There are two scenarios: 1) involving an expert technician,
and 2) involving a non-expert technician. in this article is an authoring technique that automates the
creation and positioning of 3D animation content overlaid on
the physical components. III. FAST AUGMENTED REALITY AUTHORING Fast Augmented Reality Authoring (FARA) aims to over-
come the contents-related issues previously described and
therefore ease the implementation of AR in the industry. Its name implies ‘‘fast programming’’ when implemented
in an existing AR system for maintenance. In this context,
‘‘fast programming’’ indicates ‘‘fast AR contents-creation for
maintenance procedures’’. From now on, the authors will
refer to any AR system for maintenance implemented with
FARA as FARAIS: FARA Implemented System. FARAIS is
a tool for easy knowledge transfer from experts to non-expert
technicians within procedural operations (e.g., dis/assembly,
repair, inspections). It will allow the expert (user confident
with the maintenance procedures) to ‘‘record’’ the MROs
and the non-expert to access the MROs in a ‘‘step-by-step’’
format. Ideally, a FARAIS would be suitable for any oper-
ation involving humans, e.g., both preventive and corrective
maintenance. FARA is based on two assumptions. The first
one is that current object recognition and tracking issues [10]
will be solved by providing reliable and real-time tools that
can identify objects independently from the light condition
and background noise. The second hypothesis is that CAD
models are available for the components involved in any
maintenance procedure. II. RESEARCH BACKGROUND AND MOTIVATION II. RESEARCH BACKGROUND AND MOTIVATION This research focuses on the creation of content for AR
(authoring) for maintenance applications which are known as
one of the main problems that prevent AR to become main-
stream. The simple and easy creation of AR content is not
currently available. The authoring process is time-consuming 8408 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 1. Maintenance environment simulation for testing purposes. The
environment includes the technician (Nr 1), the product to be maintained
(Nr 2) and FARAIS (Nr 3). FIGURE 2. 2D graphical representation of ‘‘transform’’. The transformed
vector of object 2 is (1x, 1y, 1α). FIGURE 1. Maintenance environment simulation for testing purposes. The
environment includes the technician (Nr 1), the product to be maintained
(Nr 2) and FARAIS (Nr 3). FIGURE 2. 2D graphical representation of ‘‘transform’’. The transformed
vector of object 2 is (1x, 1y, 1α). FIGURE 2. 2D graphical representation of ‘‘transform’’. The transformed
vector of object 2 is (1x, 1y, 1α). FIGURE 1. Maintenance environment simulation for testing purposes. The
environment includes the technician (Nr 1), the product to be maintained
(Nr 2) and FARAIS (Nr 3). B. FARA METHOD FIGURE 3. AR step-by-step procedure animation example. FARA is a method that, integrated with an AR system, forms
what in this article has been named FARAIS. On one hand,
the AR system can recognise the environment by perform-
ing object recognition and tracking and overlaying virtual
objects on the real environment following pre-programmed
rules (e.g., overlying the virtual object over the real one by
overlapping the corners). On the other hand, it provides the
maintainer with the ability to produce the virtual overlay rule
by collecting the data from the MRO. This formalizes the AR
step-by-step procedures. The FARA method is schematized
in Fig 4. The figure is divided into three main squared areas:
FARAIS, AR system, and FARA. This division is meant to
show that the union of an AR System and the FARA method
becomes FARAIS. In simple words, FARAIS consists of an
AR system (hardware and software) which utilises FARA to
record and display AR step-by-step procedures. The inputs
(arrows in) required by FARA are the AR system outputs
(arrows out) reported at the top of Fig 4: and orientations are stored as ‘‘transform’’. A transform is
a vector which consists of the linear and angular distances
between an object and an anchor object. The third one refers
to the capability of an AR system to overlay a virtual object
on a real one following a predetermined rule. The basic rules
of alignment and scale, as well as the rendering information,
are called, in this article, the basic overlay rule. Once the
procedure is completed, the expert technician can quit the
‘‘record mode’’ through the UI. FARAIS will automatically
build the AR step-by-step ‘‘Procedure 1’’. The ‘‘How’’ is
explained in Section IV. The maintainer’s effort in creating
the AR step-by-step procedure is low as the only duty is to
press the record button and perform the maintenance proce-
dure as usual. It is worth mentioning that video recording is
performed by FARAIS. and orientations are stored as ‘‘transform’’. A transform is
a vector which consists of the linear and angular distances
between an object and an anchor object. The third one refers
to the capability of an AR system to overlay a virtual object
on a real one following a predetermined rule. The basic rules
of alignment and scale, as well as the rendering information,
are called, in this article, the basic overlay rule. 1) SCENARIO 1 – EXPERT TECHNICIAN In this scenario, FARAIS ‘‘captures’’ the expert technician’s
knowledge. The expert has to carry out a maintenance pro-
cedure that they are confident with, on the product shown
in Fig 1 (Nr.2). He accesses FARAIS through the hardware
provided. Here, it is a ‘‘head-mounted video-see-through dis-
play’’. Before starting the procedure, he will select ‘‘record
mode’’ to ‘‘capture’’ the procedure and select a name: e.g.,
‘‘Procedure 1’’. Once started the procedure, FARAIS will
simultaneously perform three actions: • Recognise and track the real objects in the Field of View
(FOV) of the technician • Store the transforms of the real objects in the table
dedicated to ‘‘Procedure 1’’ • Overlay the virtual objects over the real ones utilising
the ‘‘basic overlay rule’’ available on a database (DB) The first action comprises tracking the position and orien-
tations of the objects. The second one means that the positions The first action comprises tracking the position and orien-
tations of the objects. The second one means that the positions VOLUME 11, 2023 8409 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 3. AR step-by-step procedure animation example. The latter consists of animating the virtual objects on the
real ones suggesting the positions and orientations that the
real objects must reach at each step. An example is reported
in Fig 3. First, FARAIS recognises the objects and overlays the
corresponding virtual objects utilising the basic overlay rule
available on the DB (Fig 3a). Then, the virtual object animates
detaching from the real object and moving to the target posi-
tion and orientation as suggested by the selected procedure
(Fig 3b, 3c, and 3d). It is worth noting that FARAIS also takes
into account user interaction. For instance, if the technician
moved the real object in the opposite direction, the animation
would start from the position of the real object and make a
new trajectory to get to the final position. It will be done
by getting first closer to the recorded trajectory, and then
continuing with the recorded animation. When the technician
puts the real object in the target position, the virtual object
will become green (Fig 3e). FARAIS will then move to the
next step of the procedure showing the next animation. The
screen shows the message ‘‘Procedure Completed’’ when the
procedure is completed. B. FARA METHOD Once the
procedure is completed, the expert technician can quit the
‘‘record mode’’ through the UI. FARAIS will automatically
build the AR step-by-step ‘‘Procedure 1’’. The ‘‘How’’ is
explained in Section IV. The maintainer’s effort in creating
the AR step-by-step procedure is low as the only duty is to
press the record button and perform the maintenance proce-
dure as usual. It is worth mentioning that video recording is
performed by FARAIS. 2) SCENARIO 2 – NON-EXPERT TECHNICIAN • The object recognition and tracking data In this scenario, FARAIS suggests a ‘‘step-by-step’’ AR pro-
cedure to a non-expert technician. The non-expert operator
must do a maintenance procedure that he has not done before
on the product. He/she accesses FARAIS through the hard-
ware provided. In this case, it is a head-mounted video-see-
through display. Before starting the procedure, he will select
‘‘play mode’’ and input the procedure name he wants to
perform (e.g., ‘‘Procedure 1’’). At this point FARAIS will: • Virtual object overlay basic rule • The User Kind (UK) • The Procedure Number (PN) The first input consists of the geometrical transforms in space
related to the objects in the environment. Usually, an AR
system can recognise an object and track it by estimating
its pose: the relative position and orientation of an object in
space concerning the camera. These can be translated into
a transform relative to an anchor object in the scene. The
second input consists of the basic information for overlaying
the virtual object over the real object. The third input indicates
the experience level of the operator utilising the AR system. For this study FARA only considers two levels of users: • Recognise and track the real objects in the Field-Of-
View (FOV) of the technician • Overlay the virtual objects over the real ones following
the basic overlay rule • Find ‘‘Procedure 1’’ in its DB • Show the step-by-step AR ‘‘Procedure 1’’ 8410 8410 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 4. FARA method. The arrows in and out represent the inputs and outputs of each process. The dotted lines
refer to the pre-defined input that do not depend on any decisional choices. The firm lines are driven by decisional
choices among alternative options. FIGURE 4. FARA method. The arrows in and out represent the inputs and outputs of each process. The dotted lines
refer to the pre-defined input that do not depend on any decisional choices. The firm lines are driven by decisional
choices among alternative options. 1) PROCESSING DATAi Data processing applied by FARA in process 2∗shown in Fig TABLE 1. INclination questionnaire. TABLE 1. INclination questionnaire. clarifying that these processes are automated by selecting the
threshold of time and distance required for both the smooth-
ing and the step identification. The step information is then
stored and, together with the tracking data, represents the AR
step-by-step procedure. 2) SHOW AR PROCEDURE OVERLAY RULE This process aims to create and show the step-by-step AR
procedure created to the non-expert operator. The process for
its creation takes three inputs: • Live stream of object recognition and tracking data • Transforms table was corrected and updated with the
steps created in the process ‘‘2’’ • Virtual object basic overlay rule The first one is provided in the same format as the one
stored: transforms of the objects involved in the maintenance
procedure. This will be compared every second with the
first transform of the AR step-by-step procedure built-in
process ‘‘2’’ Fig 4. If they differ, the AR step-by-step overlay
rule will be created by gradually positioning the virtual object
on the target position. An example of the animation produced
is shown in Fig. 3. When the real object transform reaches
the correct transform, the correct transform becomes the next
row of the table, and another animation will be shown. When
all the transforms in the table related to the first step are
completed, process ‘‘3’’ will move to the next step until the
procedure is completed. IV. TEST DESIGN AND METHODOLOGY 1) PROCESSING DATAi Experienced (E) and Non-Experienced (NE). The last one is
an ‘‘id’’ used for identifying the maintenance procedure that
is going to be carried out by the maintainer. It is relevant to
note that potentially, all of them can be identified without
input from the operator. Having all these 4 inputs, FARA
will first check the procedure ‘‘id’’ and then the user expe-
rience level. Only in two cases, FARA will proceed. More
specifically, if the procedure ‘‘id’’ is not already available on
the DB and the user is experienced, FARA will go through
the processes ‘‘1∗’’, and ‘‘2∗’’ in Fig 4. This is the scenario
described in the practical example in Section III-A1. On the
other hand, if the procedure is already available on DB and
the user is non-expert, FARA will go through the process
‘‘3∗’’. This is the scenario described in Section III-A2. In the
other two possible combinations, ‘‘new procedure/non-expert
user’’ and ‘‘available procedure/expert user’’, FARA will
not go through any process. While process 1∗is commonly
utilised for software development, processes 2∗, and 3∗,
in Fig. 4, have been designed for FARA. This process modifies the data acquired by the AR system
and temporarily stored at process 1∗in Fig 4. The ‘‘raw’’
data is acquired in real-time. Whilst progressively filling
the rows of a 2D table as time advances, each row needs
object recognition and tracking information related to one of
the objects in the environment at each time, ti. FARA is a
geometrical-based method. Hence, the information utilised is
(x, y, z) positions and (α, β, γ ) rotations. As stated before,
these together build the transformed vector (x, y, z, α, β, γ ). An example of the data collected for one object within one
MRO is reported in Fig 5a. For simplicity, rotations are not
shown. The data acquired is then smoothed (Fig 5b). The
data acquisition will have different errors due to the object
recognition and tracking system. These have to be deleted or
modified to store the correct information. In this example,
due to the dimensions (distances and time) of the case, the
author used exponential smoothing, applying a threshold of
40mm and 2 seconds. It means that any transformation in 8411 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 5. IV. TEST DESIGN AND METHODOLOGY FARA has been described in Section II. Its advantages
in terms of time-saving and low-effort requirements for
creating AR step-by-step procedures have been described
(Section III-A1). It is essential to validate if FARAIS’ step-
by-step procedures created with the method described in
Section III-B are as valuable as the contents created using
traditional methods. If so, it would be clear that FARA could
provide a step forward to ease AR implementation in Industry
and partially solve content-related issues by providing an
intuitive tool for creating AR content. The approach taken by
the authors to validate FARA’s method consists of two steps: FIGURE 5. Data processing applied by FARA in process 2∗shown in Fig 4. space smaller than 40 mm that lasted for less than 2 seconds
has been deleted since it is not considered a movement, but
a tracking error. The threshold has been selected arbitrarily
based on the author’s experience in this case. The process
can potentially be automated. Once the table is corrected,
process 2 will divide the transforms into groups to identify
the procedure steps. For splitting the steps, FARA considers
that each step is completed when the transforms of the objects
in the environment do not change for a predefined amount of
time. In this case, the minimum amount of time considered
is 2 seconds. For instance, in Fig 5c, it is possible to see that
‘‘Step 3’’ has been identified between the non-variation of
(x, y, z) after ‘‘Step2’’ until the re-stabilization of (x, y, z) that
follows the variation of y from 220mm to 10mm. It is worth • Quantification of the average time and errors improve-
ments of a maintenance procedure carried out by using a
FARAIS versus the same maintenance procedure carried
out by using a hard-copy manuali FARAIS versus the same maintenance procedure carried
out by using a hard-copy manuali • Comparison of the results of the quantification with the
average time and error improvements of a maintenance
procedure carried out using a traditional AR system 8412 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations study is the dis/assembly maintenance of a hydraulic valve In
the selected case study, the authors designed and 3D-printed
a ‘‘maze’’ which has similar translations and rotations of its
component. A. THE QUANTITATIVE TEST This section describes the test carried out for quantifying the
time/error improvements within a maintenance procedure due
to the utilisation of FARAIS versus a hard-copy manual. The
quantitative test methodology is described in Fig 7. Starting
from the top, the participant is asked to answer a short Likert
scale inclination questionnaire in Table 1. q
A higher average score for the first three or the last
three questions corresponds to a more AR or maintenance-
oriented profile. Based on the results, the participant will
be assigned to one between the Contents Creator (CC) and
Contents Tester (CT) groups. The first is for maintenance-
oriented profiles and the second is for AR-oriented ones. In the first case, the CC is then asked to carry out different
MRO operations using a hard-copy manual. He is given an
initial time to read the manual and become confident with
the objects and then start the procedure. The observer will
measure the time and errors by filling out a pre-designed form
that lists the errors inserted in the mockup. These will provide
the ‘‘dataset1’’ regarding the hard-copy manual supported
maintenance procedure. At this point, the CC will be allowed
any amount of time until he becomes an expert in performing
the maintenance procedure. No data is collected in this phase. The CC can now utilise FARAIS described in Section III-A1
and create an AR procedure. Each CC had to create four tasks
in which each task was taken and mixed up with each task
from different CCs. In total, there were four tasks from four
different CCs to be used by each CT. If identified as CT, the
participant is firstly inducted about the AR application and
how it works and then is asked to perform the same MRO
operation carried out by CCs. During the latter, the observer
will collect the time and error data which will provide the
FARA-support maintenance procedure ‘‘dataset2’’. In both
datasets, the time has been recorded in seconds and the
number of errors has been stored. It is worth mentioning that
the MRO operations and the product to be maintained have
been designed ad-hoc. Explicit errors have been inserted in
the mock-up design for ensuring the objectivity of the data
collection. ‘‘dataset1’’ and ‘‘dataset2’’ will then be compared
to quantitatively extract eventual improvements in terms of
time-saving and number of errors. A. THE QUANTITATIVE TEST It has been done through
the inferential statistical analysis reported in Section V-A. The quantified results will then be compared with the ones
found in the literature related to traditional AR systems for
supporting maintenance. Results are reported qualitatively in
Section V-B. FIGURE 6. Mock-up designed for testing purposes. FIGURE 6. Mock-up designed for testing purposes. IV. TEST DESIGN AND METHODOLOGY The mock-up CAD model is shown in Fig 6c
and the 3D printed version is shown in Fig 6d. The assembly
consists of three components: the basement, the board, and
the top. The basement has a planar maze that has to be com-
pleted by sliding (right, left, up, and down) the bottom part
of the board. Three errors have been inserted in the planar to
ease the collection of the data during the test. Once completed
the assembly of the board into the base, the top component
will be assembled into the board. It has a cylindrical maze
that has to be completed by rotating (CW or ACW) and
sliding (up and down) the top component on the top side of
the board component. Also in this, the authors inserted three
dis/assembly errors for testing purposes. These consist of the
misplacement of the assembly components concerning each
other in three different phases of the assembly. For easing
Vuforia’s object recognition, the surfaces of the objects have
been enriched with colour. versus the same maintenance procedure carried out
using a hard-copy manuali While the latter can be found in literature, the first one has
been calculated utilising the test described in the following
Section IV-A. C. FARA IMPLEMENTATION This section describes how the FARA method has been imple-
mented as a software unit in an AR system. This has been
done for validating the FARA method within this study, but
its implementation can be different in terms of Hardware and
Software for other research or industrial purposes. V. ANALYSIS AND RESULTS To understand whether there is a statistically significant dif-
ference between the means of timely completion of the main-
tenance tasks performed using FARAIS support vs hard-copy
manual, the author decided to carry out the one-way ANOVA
test. The two different supports, as described in section IV-A
are 1) Hard-copy manual instructions, and 2) FARAIS. The
maintenance procedures tested are: This section reports the analysis and test results described in
Section IV. Firstly, the results of the quantification test are
reported in Section V-A. Then the results of the comparison
with the current AR system’s performances in maintenance
are shown. B. THE MAINTENANCE CASE STUDY FOR
TESTING PURPOSES This section describes the hardware of the AR system
where FARA has been implemented and become a FARAIS. These are shown in Fig 1. FARA requires hardware with In this section, the case study utilised for testing purposes is
described. The real case maintenance scenario chosen in this 8413 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 7. Schematic representation of the validation test. input/output capabilities. FARAIS has to collect data from the
proposed environment and transfer the processed information
to the operator through the output device. In this specific
case, an RGB camera and touchscreen are used as the input
device and a display as the output device; both are installed
on most of the commercial mobiles available, i.e., the Sam-
sung Galaxy S8. Moreover, the display has been fitted on
a headset developed by XVENO. It has to be mentioned
that, even though the software has been designed for these
specific input/output characteristics, FARA’s method could
be utilised with different sensors and devices. For instance,
rather than capturing the current environment with an RGB
camera, it could use a depth camera and infrared cameras. The input device could consist of a laptop or a head-mounted
display (HMD). As an example, Microsoft Hololens or Epson
Moverio HMDs could be used to meet these characteristics. As an input device, it is not unexpected that depth sensors
will be soon utilised for more efficient object recognition and
tracking. In the same way, see-through displays will soon
be preferred to the video see-through display utilised in this
example. Finally, a virtual server has been set up utilising
XAMPP. 2) SOFTWARE The AR system software and FARA software unit have
been developed together as a tablet/mobile-based application
which mainly carries out three duties/units: FIGURE 7. Schematic representation of the validation test. • Hardware control complete and included the execution of an average of 3 ran-
domly chosen tasks per participant. The data collected has
been stored in compliance with Cranfield’s research ethics
policy. The following subsections report the statistical anal-
ysis and the results for the dependent variables ‘‘completion
time’’ and ‘‘the number of errors’’ affected by the utilisation
of the FARAIS support vs. hard-copy manual support. • Data processing and storage unit • Provide a responsive user interface (UI) It has been developed utilising Unity3d as a game engine and
Vuforia SDK and Android SDK. Moreover, a local virtual
server with an SQL DB has been set up to, not only provide
storage for the information collected but also process them
offline easing the workload of the mobile device. A. QUANTITATIVE TEST TABLE 3. Homoscedasticity and normality test results for the number of
errors dataset collected in the test. Normality and homoscedasticity are
verified only for the overall task. TABLE 3. Homoscedasticity and normality test results for the number of
errors dataset collected in the test. Normality and homoscedasticity are
verified only for the overall task. FIGURE 8. Overall task completion time with hard-copy manual and
FARAIS support. TABLE 4. Quantitative test results summary. TABLE 4. Quantitative test results summary. completion time described in the previous section. In per-
forming the normality tests of the error datasets, it has been
found that the data collected regarding the errors performed in
the single tasks are not normally distributed because these did
not pass the normality test. As shown in Table 3, the p-values
calculated through the Shapiro-Wilk test for the errors related
to Task1, Task2, Task 3, and Task 4, are smaller than 0.05. Thus, applying the ANOVA test to single-task datasets has not
been possible. On a positive note, the overall task error dataset
resulted to be both normally distributed and homogeneous in
terms of variance. FIGURE 8. Overall task completion time with hard-copy manual and
FARAIS support. the ‘‘assume of normality’’ for relatively big-sized samples,
in this study case it is required to validate the normality. Since the sample is smaller than 50, the Shapiro-Wilk test
is carried out and each task sample results are normal since
all the p values are greater than 0.05 as shown in Table 2. The homoscedasticity of the sample has been validated by
applying the Levene test. Also, in this case, the p values
resulted are greater than 0.05 hence the samples have the
same variance. Due to this above, the ANOVA test has been performed
only for the overall task errors. The analysis of variance
showed that the effect of the support method on the overall
task number of errors is significant, F (1,28) = 30.919,
p ≤0.05 (95% confidence). Utilising FARAIS decreased the
number of errors of the overall task by 68.6% (1.53 vs 4.87)
compared to the hard-copy manual support. No sample has been removed from the dataset after these
tests. The analysis of variance showed that the effect of
the support method on the overall task completion time
is significant, F (1,28) = 32.013, p ≤0.05 (95% confi-
dence). B. QUANTITATIVE COMPARISON OF RESULTS This section reports the comparison between AR systems
reported in the literature and FARAIS in which the valida-
tion results are presented in Section V-A. The quantitative
performance results of FARAIS for time and error reductions
are summarized in Table 4. The results have been collected
by applying the empirical test methodology described in
Section IV and reported in detail in Section IV-C. These
can be qualitatively compared with the ones found in the
literature. The latter reports the results of the literature studies
which compared the utilisation of AR systems for supporting
maintenance (designed and tested within their projects) vs the
utilisation of hard-copy manual supports. A. QUANTITATIVE TEST Utilising FARAIS improves the completion time of
the overall task by 34.7% (501s vs 768s) compared to the
hard-copy manual support. Similarly, each task separately
showed improvements in time completion. More specifically:
Task1 - F (1,28)=39.793, p ≤0.05 - 55.3% (79s vs 177s). A. QUANTITATIVE TEST • Task 1: Assembly of the Board into the Basement This section reports the results of the quantification test which
aims to quantify the improvements due to the utilisation of
FARAIS vs hard-copy manuals as support for carrying out
MRO operations. A total of 30 participants (18m/12f) took
part in the study. These include students, staff, and industrial
personnel. The average age was 28.8 (20, 36, SD = 4.28). The
participant’s background has been assessed mainly through
the inclination questionnaire. None of the participants was
working in maintenance. Because of the first inclination ques-
tionnaire, half of them have been asked to perform the test as
CC and half as CT. Each test took from 35 to 65 minutes to • Task 2: Assembly of the Top on the Board • Task 3: Disassembly of the Top from the Board • Task 4: Disassembly of the Board from the Basement • Overall Task: The complete assembly and disassembly
of the product. (Task1 + Task2 + Task3 + Task4) The number of tasks and their characteristics has been
chosen based on the case study described in Section IV-B
and the authors’ experience. To apply ANOVA to a sam-
ple, the normality and homoscedasticity of the latter must
be validated. Even though it is generally correct to make 8414 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations TABLE 2. Homoscedasticity and normality test results for the completion
time dataset collected in the test. Both are validated. FIGURE 8. Overall task completion time with hard-copy manual and
FARAIS support. TABLE 3. Homoscedasticity and normality test results for the number of
errors dataset collected in the test. Normality and homoscedasticity are
verified only for the overall task. TABLE 4. Quantitative test results summary. completion time described in the previous section. In per
forming the normality tests of the error datasets, it has been
found that the data collected regarding the errors performed in
the single tasks are not normally distributed because these did
not pass the normality test. As shown in Table 3, the p-value
calculated through the Shapiro-Wilk test for the errors related TABLE 2. Homoscedasticity and normality test results for the completion
time dataset collected in the test. Both are validated. TABLE 2. Homoscedasticity and normality test results for the completion
time dataset collected in the test. Both are validated. 2) NUMBER OF ERRORS The approach utilised for analyzing the number of errors
collected is the same as the one utilised for analysing the 8415 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 9. Task 1 completion time with hard-copy manual and FARAIS
support. It has to be mentioned that not all the studies which propose
an AR application for maintenance report in detail the time
and error reductions (AR vs. hard-copy manual) as well as
the insight of the methodology and testing material utilised
for collecting the data and analysing it. For instance, some
authors clearly explain the methodology utilised for vali-
dating a large-screen AR application [28]. In the study, the
independent variables have been the instruction mode (paper
vs. AR) and the task. Measures have been carried out on
completion time and errors. Still, the reader does not have
access to the complete paper instructions or AR step-by-
step procedure. What kind of information has been provided
to participants to perform the maintenance task with the
paper instructions? Was there a lot to read which could have
affected the completion time? Were the instructions clear
or this could lead to misinterpretations? How were the AR
instructions displayed? Similarly, others have outlined a vali-
dation methodology with four instruction modes utilised [29]:
printed manual, LCD instructions, HMD AR and spatially
registered AR. Still, not having access to the actual instruc-
tions makes it difficult to understand whether their structure
and contents could have affected the test results. Therefore,
it would not be accurate to quantitatively compare the liter-
ature outcomes with this study’s results which might have
used different user-AR interfaces and printed manual levels of
detail. For this reason, the authors of this study decided to do
a qualitative comparison of the results against others similarly
published research in the literature. More specifically, the
qualitative comparison is made with the eight studies which
the authors considered to have more similarities in terms of
field of application and validation procedure. FIGURE 9. Task 1 completion time with hard-copy manual and FARAIS
support. FIGURE 10. Task 2 completion time with hard-copy manual and FARAIS
support. TABLE 5. Maintenance performance improvements in terms of time and
errors reductions found in the literature. TABLE 5. Maintenance performance improvements in terms of time and
errors reductions found in the literature. FIGURE 10. 2) NUMBER OF ERRORS The setup is pro-
gressively being reduced as the advancement in AR technolo-
gies are advancing. It must be mentioned that one limitation
of FARA and in general of AR systems will remain the
recognition of similar and symmetrical objects which have a
different internal composition (e.g., two spheres with differ-
ent weights but the same radius). Furthermore, registration
also poses an important issue in AR, particularly in handling
occlusion between real and virtual objects. In an AR envi-
ronment, improper occlusion can hinder process efficiency
rather than enhance it. The utilisation of multiple cameras
could be a potential workaround for this issue. The second
hypothesis can be considered generally true for the industrial
environment. FARA has been described as a method and its
implementation in an AR system has been named FARAIS. method but an algorithm (FARA method) that automates the
authoring process from the tracking data. The setup is pro-
gressively being reduced as the advancement in AR technolo-
gies are advancing. It must be mentioned that one limitation
of FARA and in general of AR systems will remain the
recognition of similar and symmetrical objects which have a
different internal composition (e.g., two spheres with differ-
ent weights but the same radius). Furthermore, registration
also poses an important issue in AR, particularly in handling
occlusion between real and virtual objects. In an AR envi-
ronment, improper occlusion can hinder process efficiency
rather than enhance it. The utilisation of multiple cameras
could be a potential workaround for this issue. The second
hypothesis can be considered generally true for the industrial
environment. FARA has been described as a method and its
implementation in an AR system has been named FARAIS. The intent of having two acronyms (FARA and FARAIS) is
to emphasize the difference between the method (FARA) and
its actual practical utilisation once implemented in an already
existing AR system (FARAIS: FARA Implemented System). Even if different FARAIS can be proposed (Section IV-C1),
the authors believe that this would not negatively affect the
result of the test reported in this article. The FARAIS pro-
posed in this project has been developed to comply with the
minimum requirements in terms of performance and user
interface because this project aimed to validate FARA and
not the AR system. 2) NUMBER OF ERRORS This method was tested by having main-
tainers perform the task in front of an AR system (therefore
among the other things capable of recognizing the objects)
to create a step-by-step procedure which can be understood
by untrained users. Ideally, if all the maintainers would use
a FARAIS, lots of step-by-step procedures could be created
without additional programming.i FIGURE 12. Task 4 completion time with hard-copy manual and FARAIS
support. 2) NUMBER OF ERRORS Task 2 completion time with hard-copy manual and FARAIS
support. similarities with the one proposed in this study since both
concern mechanical components with axial and rotating
movements involved. Ong et al [4] and Sanna et al [32],
had reported time reductions from 36%. The tasks involved
in their studies were notebook disassembly and inspection
operation. These hardly relate to the mockup utilised in this
work and are shown in Section IV-B. FARAIS error reduc-
tion calculated is around 70% (Table 4). The average time
reduction in Table 5 is about 72%. Not all the studies which
observed and reported the time reductions also observed or
reported the errors reductions. The systematic observation
of the errors occurring in a maintenance operation requires
a methodology which lists in detail how and when an error
should be detected. In this study the author designed in
the mock-up design, three dis/assembly errors explained in
Section IV-B. By qualitatively comparing the performance in Table 5,
it is possible to understand that the performance of FARAIS
is close to other reported AR systems in the literature. The
time reduction calculated for FARAIS is close to the average
time reduction shown in previous studies and listed in Table 5. More specifically in this study, the average time reduction
is around 36% (Table 4). The average time reduction of
referenced studies (Table 5) is 35.5%. It is interesting to
note that Fiorentino et al observed a time reduction close
to the one observed in this study [28]. It might be because
the assembly task utilised for testing purposes presents 8416 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 11. Task 3 completion time with hard-copy manual and FARAIS
support. FIGURE 13. Overall task number of errors with hard-copy manual and
FARAIS support. FIGURE 11. Task 3 completion time with hard-copy manual and FARAIS
support. FIGURE 13. Overall task number of errors with hard-copy manual and
FARAIS support. FIGURE 11. Task 3 completion time with hard-copy manual and FARAIS
support. FIGURE 13. Overall task number of errors with hard-copy manual and
FARAIS support. FIGURE 11. Task 3 completion time with hard-copy manual and FARAIS
support. FIGURE 12. Task 4 completion time with hard-copy manual and FARAIS
support. method but an algorithm (FARA method) that automates the
authoring process from the tracking data. VI. DISCUSSION The authors’ intent in developing FARA is to provide a
method for allowing technicians to create AR-based main-
tenance step-by-step procedures while performing the task
and with minimum effort. FARA is developed based on two
assumptions: 1) To have robust and reliable object tracking
and recognition; and 2) When CAD models are available. The
authors believe that both these assumptions will be validated
in the near future. The current research effort is working on
object recognition and tracking solutions which, through the
utilisation of new sensors and technology (depth cameras,
point cloud, etc.) will overcome current lighting, occlusions,
and background noise issues. As AR technology gets more
matured over time so does the tracking system and the use
of 3D models for visualization purposes. Therefore, there
will be no additional hardware for implementing the FARA To validate FARA, the authors quantified its per-
formance and compared it with the literature findings. 8417 8417 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations FIGURE 14. Schematic representation of the FARA method implemented with Intelligent Learning. FIGURE 14. Schematic representation of the FARA method implemented with Intelligent Learning. The average error reduction (FARAIS vs hard-copy manual
support) is 68.6%. It has not been possible to calculate the
error reductions of the single tasks because the hypothesis of
normality and homoscedasticity were not verified. It has to
be considered, for future studies, the need of implementing
more variance in terms of errors collected. This could be
achieved by testing longer maintenance tasks or artificially
creating more tricks for the study participants. It must be
mentioned that the case study utilised for validation is limited
to an assembly and disassembly procedure and does not
validate FARP in other maintenance operations. However,
the application of the FARP method for repair, inspections,
and overhaul operations seems feasible and needs further
investigation. The FARA method can be improved to detect
assembly/disassembly errors to inform the AR user regarding
the correct procedure (e.g., misalignments). In addition, this The average error reduction (FARAIS vs hard-copy manual
support) is 68.6%. It has not been possible to calculate the
error reductions of the single tasks because the hypothesis of
normality and homoscedasticity were not verified. It has to
be considered, for future studies, the need of implementing
more variance in terms of errors collected. VI. DISCUSSION It should be compared with the time required
for learning how to use other authoring tools available. The
time required for creating AR procedures, on the other side,
should be quantified by collecting the time required when
utilising FARA and when utilising other authoring tools but
all applied to the same case study. Figure 14 shows the
planned expansion of the method. Comparing this with the
current schematic representation of FARA, it is possible to
see one new process: Machine Learning (ML) for improving
AR step-by-step procedures. Furthermore, AR should not be
limited to animation overlay only. The approach needs to be
formalized into a software package which can easily integrate
with existing AR authoring tools to enrich the informative-
ness of AR procedures. For example, automated arrows to
show the direction and rotation, text boxes for enriching each
step with descriptions, pdf links for opening manuals, and
vocal indications to support the operation. The idea should be
to utilise the data collected during any maintenance procedure
for automatically creating and/or enhancing the AR step-by-
step procedures. By the application of intelligent learning,
FARA could potentially, classify the MRO operation steps
and propose a different solution to a similar maintenance
problem, without the need for expert training. Finally, future
works should remove the assumptions made at the outset
of this study and more validation should be performed on
various maintenance scenarios. The methodology for comparing FARA’s performance
results is not as strict as the quantifying one. This is due
to that not all the studies about AR application in mainte-
nance have as results a quantified time saving and error rate
number. Based on the author’s experience, the results of the
comparison test can still be considered satisfying. It must
be said that, for more effective validation of FARA as an
authoring method, it should have been directly compared with
the authoring methods developed by other research centres
and applied to the same case study. Unfortunately, this pro-
cess would have required, not only access to the conceptual
authoring methods but also access to the actual tool that
different research centres have utilised for the validation. This
includes software and hardware. This approach for validating
FARA seemed impractical and not suitable at this stage of the
study. VI. DISCUSSION This could be
achieved by testing longer maintenance tasks or artificially
creating more tricks for the study participants. It must be
mentioned that the case study utilised for validation is limited
to an assembly and disassembly procedure and does not
validate FARP in other maintenance operations. However,
the application of the FARP method for repair, inspections,
and overhaul operations seems feasible and needs further
investigation. The FARA method can be improved to detect
assembly/disassembly errors to inform the AR user regarding
the correct procedure (e.g., misalignments). In addition, this The methodology for quantifying the time and error reduc-
tion has been explained. It is worth mentioning that
similar methodologies have been utilised by other stud-
ies [19], [28], [29]. For these reasons, even though for rel-
atively small samples a non-parametric approach such as
the Friedman test would be recommended, the authors still
preferred to assume normality and homoscedasticity based
on previous results. The average time reduction in perform-
ing the test’s maintenance tasks in Section V-A1 (FARAIS
vs hard-copy manual support) is 35.8%. The biggest time
reduction has been observed in Task 1, followed by Task 3,
Task 2, and Task 4. The authors believe this is because
the participants who had to perform Task 1 with the hard-
copy manual needed to take confidence in the manual itself. On the other side, the participants performing Task 1 with
FARAIS intuitively followed the instructions on the screen. 8418 VOLUME 11, 2023 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations method is currently more suited to maintenance applications
because the mechanical part is more likely to have its CAD
model available. sufficient to run the application and the transformations in
space of the objects in the FOV will be recorded and utilised
to automatically build the AR maintenance procedure. This
can then be accessed by a non- expert user to carry out the
same operation. Future studies should aim for more testing
that represents the actual industrial case study to test the
usability of the FARA method. The objective of the study
should expand validation and take more advantage of the
information recorded. The validation also needs to consider
both the learning time and the time required for creating an
AR step-by-step procedure. The first consists of the time a
maintainer needs to gain confidence with FARA and create
AR procedures. Data supporting this study are openly available from (CORD)
at DOI: 10.17862/cranfield.rd.21858771. Data supporting this study are openly available from (CORD)
at DOI: 10.17862/cranfield.rd.21858771. Data supporting this study are openly available from (CORD)
at DOI: 10.17862/cranfield.rd.21858771. VI. DISCUSSION Therefore, to have a comparative evaluation of the AR
system across different studies with traditional methods, there
is a necessity in creating a standard for the components to be
assembled (e.g., a mockup) and the assembly manuals for a
particular task that will be compared against an AR solution. The authors have included materials in this article that are
used to test this approach such as paper-based manuals that
describes the task being tested and 3D models. In this way,
it is possible to have a fair comparison by replicating the
task and comparing the efficacy of FARA implementation
with other types of authoring systems. Nevertheless, since
FARA implementation enables ‘‘task recording’’ for building
step-by-step instruction, it could be hypothesized that there
is much more time saving gained in comparison to any other
platforms that require time spent in programming or develop-
ing AR step-by-step instruction. VII. CONCLUSION FARA is a geometry-based method for authoring AR content
for maintenance. It is based on two assumptions: 1) machines
can recognise the objects in a working environment, and
2) CAD models are available for all the real objects involved
in the maintenance procedure. FARAIS is intrinsically not
time-consuming and not tedious since it does not require
the maintainer to do anything, but only perform the main-
tenance task. At the same time, it can provide a similar
amount of time and error reductions as other AR systems
for maintenance. For these reasons, the authors consider
FARA as a step forward in the development of authoring
solutions for AR (for maintenance but not only). It does
not require any programming skills to be operated. FARA
has been presented and its validation has been reported in
this article. The proposed method can be applied to any
AR system and has to be implemented as a software unit. To the best of the authors’ knowledge (literature and expe-
riences) it is the only method that allows users to create
AR step-by-step procedures with minimum effort. It will be RICCARDO PALMARINI RICCARDO PALMARINI received the M.Sc. degree in aircraft vehicle design from Cranfield
University and the Ph.D. degree from Cranfield
University, in 2019. He is currently working as
an Instrument and Automation Engineer at Tech-
nipFMC. His Ph.D. project focused on developing
an augmented reality (AR) based on visualization
system to support with the delivery of maintenance
and manufacturing. [13] G. Klinker, O. Creighton, A. H. Dutoit, R. Kobylinski, C. Vilsmeier, and
B. Brugge, ‘‘Augmented maintenance of powerplants: A prototyping case
study of a mobile AR system,’’ in Proc. IEEE ACM Int. Symp. Augmented
Reality, Oct. 2001, pp. 124–133. [14] A. Nee, S. Ong, G. Chryssolouris, and D. Mourtzis, ‘‘Augmented reality
applications in design and manufacturing,’’ CIRP Ann., vol. 61, no. 2,
pp. 657–679, 2012. [15] C. Knopfle, J. Weidenhausen, L. Chauvigne, and I. Stock, ‘‘Template based
authoring for AR based service scenarios,’’ in Proc. IEEE Virtual Reality
Conf. (VR), Mar. 2005, pp. 237–240. [16] J. Choi, Y. Kim, M. Lee, G. J. Kim, Y. Nam, and Y. Kwon, ‘‘K-MART:
Authoring tool for mixed reality contents,’’ in Proc. IEEE Int. Symp. Mixed
Augmented Reality, Oct. 2010, pp. 219–220. IÑIGO FERNÁNDEZ DEL AMO received the
degree in industrial engineering from the Poly-
technic University of Madrid (UPM), in 2016, the
degree in manufacturing consultancy from Cran-
field University, in 2016, and the Ph.D. degree
from Cranfield University. He received the schol-
arship from Babcock International for his Ph.D. degree. His primary interest is to study innovative
software methods to capture maintenance experts’
knowledge and reduce the cost of AR’s industrial
implementation. Besides his Ph.D. research, he has co-supervised various
M.Sc. thesis and group projects that involve the implementation of AR and
digital twins in the industrial workplace using SW technologies. His overall
aim is to enable the transformation of operative roles into a source of valuable
information for through-life engineering services using innovative visualisa-
tion and data-structuring technologies. His research interests include the use
of augmented reality (AR) and semantic web (SW) for knowledge capture in
maintenance diagnosis. He won the 2017 Cranfield Vice-Chancellor’s Prize
for being the most outstanding M.Sc. student. [17] J. Shim, M. Kong, Y. Yang, J. Seo, and T.-D. Han, ‘‘Interactive features
based augmented reality authoring tool,’’ in Proc. IEEE Int. Conf. Consum. Electron. (ICCE), Jan. 2014, pp. 47–50. [18] Y. Yang, J. Shim, S. Chae, and T.-D. REFERENCES [1] P. Milgram and F. Kishino, ‘‘A taxonomy of mixed reality visual displays,’’
IEICE Trans. Inf. Syst., vol. 77, no. 12, pp. 1321–1329, Dec. 1994. [2] R. T. Azuma, ‘‘A survey of augmented reality,’’ Presence, Teleoperators
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aware maintenance support for augmented reality assistance and syn-
chronous multi-user collaboration,’’ Proc. Cirp, vol. 59, pp. 18–22,
Jan. 2017. 8419 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations [6] F. Lamberti, F. Manuri, A. Sanna, G. Paravati, P. Pezzolla, and
P. Montuschi, ‘‘Challenges, opportunities, and future trends of emerg-
ing techniques for augmented reality-based maintenance,’’ IEEE Trans. Emerg. Topics Comput., vol. 2, no. 4, pp. 411–421, Dec. 2014. [27] G. Westerfield, A. Mitrovic, and M. Billinghurst, ‘‘Intelligent augmented
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‘‘Augmented reality on large screen for interactive maintenance instruc-
tions,’’ Comput. Ind., vol. 65, no. 2, pp. 270–278, Feb. 2014. [7] U. Neumann and A. Majoros, ‘‘Cognitive, performance, and systems
issues for augmented reality applications in manufacturing and main-
tenance,’’ in Proc. IEEE Virtual Reality Annu. Int. Symp., Mar. 1998,
pp. 4–11. [29] A. Tang, C. Owen, F. Biocca, and W. Mou, ‘‘Experimental evaluation of
augmented reality in object assembly task,’’ in Proc. Int. Symp. Mixed
Augmented Reality, 2002, pp. 265–266.i [8] S. Takata, F. Kirnura, F. J. Van Houten, E. Westkamper, M. Shpitalni,
D. Ceglarek, and J. Lee, ‘‘Maintenance: Changing role in life cycle man-
agement,’’ CIRP Ann., vol. 53, no. 2, pp. 643–655, 2004. [30] S. Lee and Ö. Akin, ‘‘Augmented reality-based computational fieldwork
support for equipment operations and maintenance,’’ Autom. Construct.,
vol. 20, no. 4, pp. 338–352, 2011. [9] M. Hincapie, A. Caponio, H. Rios, and E. G. Mendivil, ‘‘An introduction to
augmented reality with applications in aeronautical maintenance,’’ in Proc. 13th Int. Conf. Trans. Opt. Netw., Jun. 2011, pp. REFERENCES 1–4. [31] H. Ramirez, E. G. Mendivil, P. R. Flores, and M. C. Gonzalez, ‘‘Authoring
software for augmented reality applications for the use of maintenance and
training process,’’ Proc. Comput. Sci., vol. 25, pp. 189–193, Jan. 2013. [10] R. Palmarini, J. A. Erkoyuncu, R. Roy, and H. Torabmostaedi, ‘‘A sys-
tematic review of augmented reality applications in maintenance,’’ Robot. Comput.-Integr. Manuf., vol. 49, pp. 215–228, Feb. 2018. [32] A. Sanna, F. Manuri, F. Lamberti, G. Paravati, and P. Pezzolla, ‘‘Using
handheld devices to sup port augmented reality-based maintenance and
assembly tasks,’’ in Proc. IEEE Int. Conf. Consum. Electron. (ICCE),
Jan. 2015, pp. 178–179.i [11] F. Zhou, H. B.-L. Duh, and M. Billinghurst, ‘‘Trends in augmented reality
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documentation for maintenance and repair,’’ IEEE Trans. Vis. Comput. Graphics, vol. 17, no. 10, pp. 1355–1368, Oct. 2011. [12] T. Jung, M. D. Gross, and E. Y.-L. Do, ‘‘Annotating and sketching on 3D
web models,’’ in Proc. 7th Int. Conf. Intell. User Interfaces, Jan. 2002,
pp. 95–102. RICCARDO PALMARINI Han, ‘‘Interactive augmented reality
authoring system using mobile device as input method,’’ in Proc. IEEE Int. Conf. Syst., Man, Cybern. (SMC), Oct. 2016, pp. 001429–001432. [19] S. J. Henderson and S. K. Feiner, ‘‘Augmented reality in the psychomotor
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system to assist the maintenance operators,’’ Int. J. Interact. Des. Manuf. (IJIDeM), vol. 8, no. 4, pp. 293–304, Nov. 2014. [22] H. Alvarez, I. Aguinaga, and D. Borro, ‘‘Providing guidance for mainte-
nance operations using automatic markerless augmented reality system,’’
in Proc. 10th IEEE Int. Symp. Mixed Augmented Reality, Oct. 2011,
pp. 181–190. DEDY ARIANSYAH DEDY ARIANSYAH worked previously as a
Research Fellow at Cranfield University, under
the EPSRC-funded project DigiTOP, where his
research focused on how immersive technologies
like augmented reality can improve industrial pro-
ductivity and digital twin development to sup-
port decision-making. He is currently involved
in the AI Research Team, where he focuses on
the embedded AI in immersive technologies and
knowledge capture to improve task productivity,
increase the efficiency of knowledge transfer, and data-driven process opti-
mization. His Ph.D. degree was focused on the virtual reality driving
simulator for driver behaviour analysis, where he developed and integrated
physiological data and driving performance to study different human cogni-
tive operations when interacting with driver assistance systems. [23] T. V. Nguyen, B. Mirza, D. Tan, and J. Sepulveda, ‘‘ASMIM: Aug-
mented reality authoring system for mobile interactive manuals,’’ in
Proc. 12th Int. Conf. Ubiquitous Inf. Manag. Commun., Jan. 2018,
pp. 1–6. [24] T. V. Nguyen, D. Tan, B. Mirza, and J. Sepulveda, ‘‘MARIM: Mobile
augmented reality for interactive manuals,’’ in Proc. 24th ACM Int. Conf. Multimedia, Oct. 2016, pp. 689–690. [25] M. E. C. Santos, A. Chen, T. Taketomi, G. Yamamoto, J. Miyazaki,
and H. Kato, ‘‘Augmented reality learning experiences: Survey of proto-
type design and evaluation,’’ IEEE Trans. Learn. Technol., vol. 7, no. 1,
pp. 38–56, Jan. 2014. [26] J. Wang, Y. Feng, C. Zeng, and S. Li, ‘‘An augmented reality based
system for remote collaborative maintenance instruction of complex prod-
ucts,’’ in Proc. IEEE Int. Conf. Autom. Sci. Eng. (CASE), Aug. 2014,
pp. 309–314. 8420 VOLUME 11, 2023 R. Palmarini et al.: FARA: Fast Creation of AR Step-by-Step Procedures for Maintenance Operations SAMIR KHAN (Member, IEEE) received the
B.Eng. and Ph.D. degrees from Loughborough
University. He is currently a Senior Lecturer
often collaborating with Rolls-Royce, Jaguar Land
Rover, BAE Systems, and MoD. His research
interests include intelligent monitoring, AI-based
control, perception, and planning in the context
of autonomous systems and service robots, with a
focus on the embedded hardware implementation
of machine learning techniques for anomaly detec-
tion and prognostics and system health management, development of the IoT
technologies, digital twins for maintenance decision making, and intelligent
monitoring of intermittent failures and false alarms in electronic systems. He is a Chartered Engineer and a member of IET. RAJKUMAR
ROY
received the B.Eng. and
M.Eng. degrees in production engineering from
Jadavpur University, India, and the Ph.D. DEDY ARIANSYAH degree
in computing from the University of Plymouth,
U.K. He started his career as an Engineer at Tata
Motors; pioneered research in Through-life Engi-
neering Services (TES) with Rolls-Royce, BAE
Systems, Bombardier Transportation, the Min-
istry of Defence, and Babcock International; and
established an internationally known TES Centre. He joined the City, University of London from Cranfield University, where
he was the Director of Manufacturing. His cost engineering and obsolescence
research has transformed contemporary understanding of the engineering
effort required to design, make and support high-value products, resulting in
tools used by BAE Systems, Airbus, the Ministry of Defence, Rolls-Royce,
and Ford Motor Company. He is the Founding Editor-in-Chief of the Applied
Soft Computing journal (Elsevier); and a fellow of CIRP (International
Academy for Production Engineers), the Institute of Engineering Designers
(IED), and the Higher Education Academy (HEA). JOHN AHMET ERKOYUNCU received the Ph.D. degree in uncertainty modeling for maintenance
from Cranfield University, in 2011. He is currently
the Head of the Digital Engineering and Manu-
facturing Centre. He is active with Innovate U.K. and EPSRC-funded projects around research top-
ics: digital twins, augmented reality, digitization
(degradation assessment), and simulation of com-
plex manufacturing and maintenance procedures. He is a Chartered Engineer, an Associate Member
of CIRP, and the Co-Chair of the Through-life Engineering Services Council. 8421 VOLUME 11, 2023 Cranfield University
CERES Research Repository https://dspace.lib.cranfield.ac.uk/ School of Aerospace, Transport and Manufacturing (SATM)
Staff publications (S School of Aerospace, Transport and Manufacturing (SATM) Staff publications (SATM) Palmarini R, Fernandez del Amo I, Ariansyah D, et al., (2023) Fast augmented reality authoring:
fast creation of AR step-by-step procedures for maintenance operations. IEEE Access, Volume
11, 2023, pp. 8407-8421 https://doi.org/10.1109/ACCESS.2023.3235871
Downloaded from CERES Research Repository, Cranfield University Fast augmented reality authoring: fast
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2023-01-24
Attribution 4.0 International Palmarini, Riccardo
2023-01-24
Attribution 4.0 International Palmarini, Riccardo Attribution 4.0 International Palmarini R, Fernandez del Amo I, Ariansyah D, et al., (2023) Fast augmented reality authoring:
fast creation of AR step-by-step procedures for maintenance operations. IEEE Access, Volume
11, 2023, pp. 8407-8421
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https://openalex.org/W2146174092
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https://academicjournals.org/journal/AJB/article-full-text-pdf/C7DA92843982.pdf
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English
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Growth of Scenedesmus dimorphus in different algal media and pH profile due to secreted metabolites
|
African journal of biotechnology
| 2,014
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cc-by
| 3,949
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Vol. 13(16), pp. 1714-1720, 16 April, 2014
DOI: 10.5897/AJB2013.13455
Article Number: C7DA92843982
ISSN 1684-5315
Copyright © 2014
Author(s) retain the copyright of this article
http://www.academicjournals.org/AJB
African Journal of Biotechnology Vol. 13(16), pp. 1714-1720, 16 April, 2014
DOI: 10.5897/AJB2013.13455
Article Number: C7DA92843982
ISSN 1684-5315
Copyright © 2014
Author(s) retain the copyright of this article
http://www.academicjournals.org/AJB
African Journal of Biotechnology Vol. 13(16), pp. 1714-1720, 16 April, 2014
DOI: 10.5897/AJB2013.13455
Article Number: C7DA92843982
ISSN 1684-5315
Copyright © 2014
Author(s) retain the copyright of this article
http://www.academicjournals.org/AJB
African Journal of Biotechnology African Journal of Biotechnology Received 11 November, 2013; Accepted 14 March, 2014 In this study investigation was made to evaluate the effects of different algal media components to get
optimized cell count of Scenedesmus dimorphus. Five different fresh water algal media such as Bold’s
Basal Medium (BBM), M4N medium, BG-11 medium, N-8 medium and M-8 medium were used for
culturing S. dimorphus in flask culture. A set of environmental factors including light, temperature, air
flow rate and nutritional components was standardized to obtain the highest productivity of 0.1406 g/L
with specific growth rate of 0.10483/day. This study designates the bold basal medium as advantageous
one for S. dimorphus and also reveals that production of metabolites by the same algal strain depends
mostly on the nature of constituents of media and might have different influence on the pH. Key words: Scenedesmus dimorphus, bold basal medium, algal growth. Full Length Research Paper 1Department of Chemical and Process Engineering. Faculty of Engineering and Built Environment, Universiti
Kebangsaan Malaysia (UKM), 43600 Bangi, Selangor, Malaysia. 2School of Biosciences and Biotechnology, Faculty of Science and Technology, University Kebangsaan Malaysia, UKM
43600 Bangi, Selangor, Malaysia. 3 partment of Chemical and Process Engineering. Faculty of Engineering and Built Environm
Kebangsaan Malaysia (UKM), 43600 Bangi, Selangor, Malaysia. g
y
(
),
g ,
g
,
y
School of Biosciences and Biotechnology, Faculty of Science and Technology, University Kebangsaan Malaysia, UKM
43600 Bangi, Selangor, Malaysia. 3 g ,
g
,
y
entre for Energy Systems, National University of Sciences and Technology, Sector H-12, Is g ,
g
,
y
ems, National University of Sciences and Technology, Sector H-12, Islamabad Pakistan. *Corresponding author. E-mail: dr.ehsan@ces.nust.edu.pk. Tel: +92-51-9085 5275. Fax: +92-51-9085 5272.
Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0
International License INTRODUCTION To meet the existing energy and environmental issues,
renewable energy has been designated as a sustainable
solution (Amin, 2009; Hallenbeck and Benemann, 2002). It has been documented that more than 80% of the
energetic resources has been utilized to achieve the
existing
status
of
advancement
on
this
planet
(Huesemann, 2006). Algae as a source of oil/fuel has
been documented as preferred on oil producing terrestrial crops. In addition to oil/fuel, algae are also well known for
producing polyunsaturated fatty acids, which have been
used as feed for fish and other animals (Harlioğlu, 2012;
Narejo and Rahmatullah, 2010; Spolaore et al., 2006). Microalgae as a source of biofuel/renewable energy have
been documented for significant environmental and
commercial importance. Microalgae are not only sources
of fuel, food for humans and animals, but are also the *Corresponding author. E-mail: dr.ehsan@ces.nust.edu.pk. Tel: +92-51-9085 5275. Fax: +92-51-9085 5272. Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0
International License 1715 Al-Shatri et al. Al-Shatri et al. Al-Shatri et al. sources of a wide range of chemical compounds used in
industry, food technology, and pharmaceuticals as well
(Bruton et al., 2009). Photosynthetic microalgae are
potential candidates for utilizing excessive amount of
CO2, since these organisms are capable of fixing CO2 to
produce energy and chemical compound upon exposure
to sunlight (Neenan et al., 1986). Microalgae have high
growth rates and tolerance for varying environmental
conditions. Microalgae can be grown in arid and in semi-
arid regions with poor soil quality where woody or
herbaceous crops cannot be grown. Saline water from
aquifers or the ocean can be used for growing micro-
algae. Such water has few competing uses and cannot
be used for agriculture, forestry, or as potable water. The
yield of biomass per hectare from microalgae is three to
fivefold greater than the yield from typical crop plants
(Bischoff and Bold, 1963). Effect of different media on S. dimorphus growth rate Cultures were subjected to five different media of different chemical
compositions, BBM medium, M4N medium, GB-11 medium, N-8
medium and M-8 medium. All the media were in broth form, growth
was monitored through optical density (OD) and cell count using
microscope. Simultaneously, five batch photobioreactor (Scott
Duran bottles) having 2 L capacity containing 900 mL of each
medium and 100 mL S. dimorphus were subjected to evaluate the
effect of different media on S. dimorphus. All media in the duran
bottles were pre sterilized in autoclave at 121°C for 20 min before
inoculation. The cultures were incubated at room temperature 30 ±
2°C under continuous light illuminated with cool fluorescent lamp
(the light intensity was 1.3 ± 0.05 KLux). High quality chemical
constituents were used for preparation of media with maximum
accuracy (0.0001 g) in weighing by using Electronic balance,
Precise, XT220A. Observations of algae growth was carried out
daily in all media. The initial cell concentration was 5.3 x 106
cell/mL. To avoid settling, and for accelerating the growth process,
air supply with constant volumetric flow rate 1.5 L/min (1.5 vvm)
was used. Growth was followed through optical density during 15
days which was recorded by using spectrophotometer at
wavelength of 730 nm. Algae strain S. dimorphus is in the Chlorophyta family and was provided by
Algaetech International Sdn Bhd Malaysia on agar Petri plates. Algae are cultivated first by transferring to test tube in BBM media. The test tubes are placed in a well-lit window with temperature 28
to 30°C until the medium turns green, signaling adequate algae
growth. They were transferred to the batch PBR (Duran bottle 2 L
size), where a larger volume (900 ml) of medium was used for
higher biomass. The test tubes are placed in a well-lit window with temperature 28
to 30°C until the medium turns green, signaling adequate algae
growth. They were transferred to the batch PBR (Duran bottle 2 L
size), where a larger volume (900 ml) of medium was used for
higher biomass. Growth of algal strains in batch system The batch system, shown in Figure 1 was used for studying culture
performance. In this system, 2 L Duran bottles were used as batch
reactors and sealed with cap stoppers with two stainless tubes
through which air was fed, exhausted and screwed by plastic cover. Air flow through polyvinylchloride pipes connected with 0.2 µm
membrane filters and connected between stainless steel tubes and
air pumps. The air feed tube was kept immersed inside the growth
container and angled at the bottom of the container to allow mixing,
to prevent sedimentation of the algae, to ensure that all cells of the
population are equally exposed to the light and nutrients, and to
improve gas exchange between the culture medium and the air. Two cool white fluorescent lamps were employed as the light
source of growth with an average light intensity of 1.3 ± 0.05 KLux. The volumetric flow rate of air was 1.5 L/min. Bioreactor
temperature was monitored at ambient laboratory conditions as 30
± 2°C. (
,
)
Microalgae cultivation is gaining importance for its
application in fuel and feed sectors in the world. A range
of documented algal media can be used to cultivate
specific algal strain for maximum growth and strategies
can be designed to obtain the targeted products
optimally. A number of algal media to culture freshwater
algae have been reported, but the productivity is different
from strain to strain. This study focuses on using five
different
media
to
culture
fresh
water
algae
Scenedesmus dimorphus. These media include Bold’s
Basal Medium (BBM), M4N medium, BG-11 medium, N-8
medium and M-8 medium (Eliasson et al., 1999;
http://web.biosci.utexas.edu/utex/mediaDetail.aspx?medi
aID=26). The source of carbon in all is carbon dioxide for
photoautotrophic growth except BG-11 medium. The aim
of this study is to determine the best medium for S. dimorphus in the batch photo bioreactors. Here, the study
was designed to investigate the most suitable media
under optimized influencing factors for the significant
growth of S. dimorphus at lab scale. At the same time a
critical investigation was made on nature of metabolites
production from same strain but in different media with
dissimilar nutrients or concentration of nutrients to
influence the pH, a newly designed flocculant was also
used to harvest the biomass for maximum biomass
recovery. Specific growth rate of S. dimorphus All the chemicals used were of analytical grade unless pointed out
clearly. Bold’s Basal Medium (BBM) was prepared using distilled
water, and the pH was adjusted (6.7 ± 0.3) with 5 N sodium
hydroxide and 5 N hydrochloric acid (Bischoff and Bold 1963;
Rowley, 2010). M4N medium and BG-11 Medium were prepared
according to reported media recipes (Mandalam and Palsson,
1998) and N-8 and M-8 Media were prepared according to reported
media recipes (Guillard and Ryther, 1962). Specific growth rate is a measure of number of generations (the
number of doublings) that occur per unit of time in an exponentially
growing culture. The exponential (straight line) phase of growth was
carefully determined and specific growth rate was obtained using
Equation (1) (Chisti, 2007). (1) 1716 Afr. J. Biotechnol. Figure 1. Schematic diagram of batch photobioreactor system. Figure 1. Schematic diagram of batch photobioreactor system. flocculant 1 to 5% (w/v) (with a unit difference of 1) were added
together at room temperature (25 to 28°C) to each 500 ml beaker
containing 400 ml of algal culture making both solutions finally
neutralized, then biomass removal was monitored at 2, 4 and 6 h
intervals. No is number of cells at the start of log phase, Nt is number of cells
at the end of log phase. To is Starting day of log phase, Tt is Final
day of log phase. CO2 fixation (g/L.d) In order to characterize harvesting of algal cells (as a result of
coagulation/flocculation) by a new flocculant prepared in our
laboratory as a solution form, the batch experiments were done
using a six Scott duran bottles 250 mL filled with 200 mL of S. dimorphus collected after stationary phase. The duran bottles allow
comparison of five different doses of flocculant to determine the
required dose for adequate removal of suspended microalgae and
the sixth bottle was used as a control. Doses used in these
experiments to achieve removal of freshwater algae were 1, 2, 3, 4
and 5% (v/v), and the last one (control) without any flocculant. Biomass and carbon production can be calculated based on fact
that the microalgae contain 53.55% carbon (Reddy, 2002), some
reports said micro algal biomass contains approximately 50%
carbon by dry weight which presents that 1 kg of dry algal biomass
utilize about 1.83 kg of CO2 (Dragone et al., 2010; Reddy, 2002). The percentage of carbon fixed based on input can be calculated
by using Equation (2). (2) In this process, it is essential that the flocculant was added by
slow mixing to allow good contact between the small flocs and to
agglomerate them into larger particles. The pH was measured
before and during the flocculation period, also cell concentration
and optical density at 730 nm was recorded. The biomass removal
percentage was calculated during the flocculation by using
Equation (3). (2) Biomass recovery using a newly designed flocculant So, Equation (3) could be compacted to this formula. RESULTS AND DISCUSSION The absorbance of the sample was measured using a
spectrophotometer and correlated to calculate the dry
weight or the number of cells per unit volume. The
empirical equation for the calibration curve showed a
linear relationship between optical densities and dry cell
weight (g/L), represented by Equation (6). Also, a linear
relationship between optical densities and biomass
concentrations (cell*E+6/L) is represented by Equation
(7). Biomass recovery using a newly designed flocculant A flocculant was prepared using palm oil industry waste (waste
activated bleaching earth). The flocculant was in possession of
aluminium
and
silica
contents
(unpublished
data
UKM,
Biotechnology Lab). The composition of flocculant was two
separate solutions of 350 ppm aluminum in 1N HCl and 400 ppm
silicon in 1N NaOH. Different concentration of both parts of (3)
So, Equation (3) could be compacted to this formula. (3) So, Equation (3) could be compacted to this formula. 1717 Al-Shatri et al. Figure 2. Growth of S. dimorphus in different media. Figure 2. Growth of S. dimorphus in different media. Figure 2. Growth of S. dimorphus in different media. (4) shown in Figure 3. Nature of metabolites secreted by the
algae was not examined but their relation with pH was
observed and evaluated. It was observed that M4N is the
only medium which did not allow any abrupt change in pH
and slowly increased the pH after 7th day of cultivation. In other words, algae in M4N medium were not secreting
alkaline or acidic metabolites to affect the pH like algae in
other media. S. dimorphis secreted alkaline metabolites
in all other media and increased the pH during first five
days and then got stable up to 15 days. This behaviour
shows that the same strain can be influenced differently
by different media constituent to produce metabolites of
different nature. (4) Cell removal efficiency percentage (%RE) was calculated using the
following Equation (5) (Fogg, 1966). (5) (5) Where; Final fluorescence is optical density after flocculation
process at limited time. Final fluorescence of control is optical
density for the control sample at same time. Effect of different media on growth rate Effect of different nutrient composition on growth pattern
forms the objective of present work that aimed at
selecting the best medium for S. dimorphus. Five runs
with various culture media were carried out to select a
suitable medium for cell growth of S. dimorphus. It can be
seen in Figure 2 that S. dimorphus was growing faster (in
15 days) in Bold’s Basal Medium (BBM) medium as
compared to the other media. M4N and BG-11 came
after BBM, while M-8 and N-8 came at the final as
arranged. The growth curve did not show lag phase and it
demonstrates that there was a quick adaptation of S. dimorphus to all media. pH was not controlled and the
changes were monitored during the growth period as y = 1.0885x (6)
y = 13.055x (7) (6) y = 1.0885x y = 1.0885x (7) y = 13.055x y = 13.055x The biomass productivity, specific growth rate, optical
density, cell concentration and CO2 fixation were Afr. J. Biotechnol. 1718 Figure 3. pH profile, while growing S. dimorphus in different media. Figure 3. pH profile, while growing S. dimorphus in different media. Figure 3. pH profile, while growing S. dimorphus in different media. Table 1. Results of growth rate in culturing media. Table 1. Results of growth rate in culturing media. Medium
BBM
M4N
BG11
N8
M8
Specific growth rate (µ)/day
0.10483
0.09781
0.09693
0.09492
0.09687
Final optical density (OD)
1.937
1.765
1.742
1.711
1.719
Final cell number (cell*E+6/L)
25.288
23.042
22.742
22.337
22.442
Biomass productivity ( g/L.d)
0.1406
0.12808
0.1264
0.1242
0.1247
CO2 fixation (g/L.d)
0.2573
0.2344
0.2313
0.2273
0.2282 and most of carbon lifted from the system quickly with low
dissolved amount, but it was best to keep culture on
agitation and prevent our strain from settling. Reddy
(2002) reported the carbon fixation based on the input
46.20 gC/day was approximately 3.65% for the flat-plate
photobioreactor (Reddy, 2002). Yun et al. (1997) found
that 0.624 g CO2/L.d fixation from flue gas in wastewater
medium at 15% (v/v) CO2 was supplied to the air-adapted
inoculum and at light intensity approximately 8 Klux, while
in this study it was found 0.258 g CO2/L.d in BBM with air
only (Osborne, 2009). and most of carbon lifted from the system quickly with low
dissolved amount, but it was best to keep culture on
agitation and prevent our strain from settling. Effect of different media on growth rate Reddy
(2002) reported the carbon fixation based on the input
46.20 gC/day was approximately 3.65% for the flat-plate
photobioreactor (Reddy, 2002). Yun et al. (1997) found
that 0.624 g CO2/L.d fixation from flue gas in wastewater
medium at 15% (v/v) CO2 was supplied to the air-adapted
inoculum and at light intensity approximately 8 Klux, while
in this study it was found 0.258 g CO2/L.d in BBM with air
only (Osborne, 2009). estimated for all media as shown in Table 1, based on
the equations (1), (6) and (7).The tabled values show the
good comparison between all checked media which
indicate that a bit better growth rate has been seen in the
BBM medium. The best result for growth of S. dimorphus was
obtained when BBM was used (Table 1). The biomass
productivity was 0.1406 g/L.d. Mass of C per day entering
the system was 3.4339 g/day. Microalgae contain 50%
carbon, so the amount of carbon production per day in
the system was 0.0703 g/day. Thus by using equation
(2), the percentage of carbon fixed based on input is: Biomass recovery using a novel flocculant The results for biomass recovery, removal percentage,
and removal efficiency percentage are tabulated in Table
2. The results were in the trends to increase the biomass
recovery by increasing the dose of flocculant from 1 to
5% (v/v). The maximal removal achieved at 5% (v/v)
during 2 h, while more effective removal can be obtained The theoretical carbon percentage found is less than 5 to
35% of the carbon fixed by marine phytoplankton and
immediately lost from the cells as excreted organic matter
(Yun et al., 1997). It may be due to high air flow rate used 1719 Al-Shatri et al. s recovery, biomass removal percentage and removal efficiency percentage at different doses during time Table 2. Biomass recovery, biomass removal percentage and removal efficiency percentage at differe Time (h)
2
4
6
Flocculant
(% v/v)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
0
9.60
46.27
0.00
15.50
74.70
0.00
18.48
89.04
0.00
1
17.80
85.78
54.38
19.66
94.75
67.55
20.17
97.18
15.84
2
19.30
93.01
84.38
20.10
96.88
87.09
15.84
20.61
99.33
3
19.50
93.98
89.06
20.48
98.69
88.08
20.66
99.57
68.32
4
19.75
95.18
89.84
20.59
99.20
90.40
20.67
99.63
75.25
5
20.10
96.87
92.19
20.66
99.55
92.38
99.82
78.22
18.48
Time (h)
Cell number E+6/ml
Figure 4. Biomass recovery during time with different doses of flocculant. Time (h)
2
4
6
Flocculant
(% v/v)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
Cell
(E+6/mL)
Removal
(%)
Efficiency
(%)
0
9.60
46.27
0.00
15.50
74.70
0.00
18.48
89.04
0.00
1
17.80
85.78
54.38
19.66
94.75
67.55
20.17
97.18
15.84
2
19.30
93.01
84.38
20.10
96.88
87.09
15.84
20.61
99.33
3
19.50
93.98
89.06
20.48
98.69
88.08
20.66
99.57
68.32
4
19.75
95.18
89.84
20.59
99.20
90.40
20.67
99.63
75.25
5
20.10
96.87
92.19
20.66
99.55
92.38
99.82
78.22
18.48 Time (h)
Cell number E+6/ml
Figure 4. Biomass recovery during time with different doses of flocculant. Figure 4. Biomass recovery during time with different doses of flocculant. is the optimal suitable medium for culturing S. dimorphus
due to the comparatively high productivity of biomass
(0.1406 g/L.d) at temperature 30°C, free air volumetric
flow rate 1.5 L/min, and under continuous light intensity
1.3 ± 0.05 KLux. Biomass recovery using a novel flocculant at 1% (v/v) by extending the period to 6 h as shown in
Figures 4 and 5, where the initial cell concentration was
20.75*106
cell/mL. The
removal
percentage
was
calculated by using Equation (4) and plotted per each
dosage of flocculant. The results showed the maximal
removal percentage that can be achieved at 5% (v/v) in 2
h (96.87%), but better removal can be achieved with 1%
(v/v) dose after 6 h (97.18%) as shown in the Figure 5. Regarding metabolite production, some
valuable
findings can be expressed here as, the metabolite
production from the same strain may vary to express
different pH in different media. It was observed that the
metabolites produced from S. dimorphus were mostly
neutral in the beginning while growing in M4N but the
other media presents some alkaline nature of metabolites
during all growth period. Also, the highest cell removal efficiency percentage (%
RE) was obtained at dose 5% (v/v) 92.19% after 2 h,
while the lowest (% RE) was obtained at dose 1% (v/v)
54.38% after 2 h. g
g
Variation in the pH of culture depends on the nature of
metabolites secreted in response to the nutrients in
different media but not due to the nature of algal strain. Conflict of interests The author(s) have not declared any conflict of interests. )
Harlioğlu AG (2012). Fat Soluble Vitamins and Cholesterol Content of
Farmed Rainbow Trout (Oncorhynchus mykiss). Pak. J. Zool
44(4):1013-1019. Conclusion The findings revealed that the Bold basal medium (BBM) Afr. J. Biotechnol. 1720 Time (h)
Cell removal percent
Figure 5. Removal percentage of biomass/algae using different doses of flocculant with time. Figure 5. Removal percentage of biomass/algae using different doses of flocculant with time. Hallenbeck PC, Benemann JR (2002). Biological hydrogen production;
fundamentals and limiting processes. Int. J. Hydrogen Energy 27(11-
12):1185-1193. ACKNOWLEDGEMENTS Huesemann M (2006). Can Advances in Science and Technology
Prevent Global Warming? Mitigation and Adaptation Strategies for
Global Change 11(3):539-577. The authors are thankful to Ministry of Higher Education
Malaysia for financial assistance through a research
grant ERGS/1/2011/STWN/UKM/02/5. Mandalam RK, Palsson B (1998). Elemental balancing of biomass and
medium composition enhances growth capacity in high-density
Chlorella vulgaris cultures. Biotechnol. Bioeng. 59(5):605-611. Narejo NT, Rahmatullah SM (2010). Studies on the Grazing Rate of
Culibaush, Labeo calbasu (Hamilton) on Periphyton. Pak. J. Zool. 42(1):53-56. REFERENCES Neenan B, Feinberg D, Hill A, McIntosh R, Terry K (1986). Fuels from
microalgae: Technology status, potential, and research requirements. Publ. No. SERI/SP-231-2550 Solar Energy Research Institute,
Golden, CO, 149 pp Amin S (2009). Review on biofuel oil and gas production processes
from microalgae. Energy Conversion and Management 50(7):1834-
1840. Osborne AL (2009). Harvesting Microalgae for Biofuel: Processes and
Mechanisms,
University
of
Texas,
Texas,
USA. http://hdl.handle.net/2152/ETD-UT-2009-12-711 Bischoff HW, Bold HC (1963). Some soil algae from Enchanted Rock
and related algal species. Phycological Studies IV. Univ. Texas Publ. 6318:1-95. Reddy MH (2002). Application of algal culture technology for carbon
dioxide and flue gas emission control, Arizona State University,
Arizona,
USA,
Arizona. http://www4.eas.asu.edu/pwest/Theses_Diss/Madhu_Thesis%20Alga
e%20Photosynthesis.pdf. Bruton T, Lyons, H, Lerat Y, Stanley M, BoRasmussen M (2009). A
review of the potential of marine algae as a source of biofuel in
Ireland. Sustainable Energy Ireland Publications, February 2009s. Chisti Y (2007). Biodiesel from microalgae. Biotechnol. Adv. 25(3):294-
306. Rowley WM (2010). Nitrogen and Phosphorus Biomass-Kinetic Model
for Chlorella vulgaris in a Biofuel Production Scheme. Ohaio: Air
Force Institute of Technology, Air University, NTIS Issue Number
1018 http://www.ntis.gov/search/product.aspx?abbr=ADA519649. Dragone G, Fernandes B, Teixeira JA (2010). Third generation biofuels
from microalgae. In. Mendez-Vilas A (eds.). Current research,
technology and education topics in applied microbiology and
microbial biotechnology. Vol. 1 ISBN (13) (pp. 1-788) g
Spolaore P, Joannis-Cassan, C, Duran E, Isambert A (2006). Commercial applications of microalgae. J. Biosci. Bioeng. 101(2):87-
96. Eliasson B, Riemer P, Wokaun A (1999). Greenhouse Gas Control
Technologies. Elsevier, 20-May-1999 - Science. 1205 p. Fogg GE (1966). The extracellular products of algae. Oceanography
and marine biology: an annual review 4:195-212. Yun YS, Lee SB, Park JM, Lee CI, Yang JW (1997). Carbon Dioxide
Fixation by Algal Cultivation Using Wastewater Nutrients. J. Chem. Technol. Biotechnol. 69(4):451-455. Guillard RRL, Ryther JH (1962). Studies of marine planktonic diatoms: i. Cyclotella nana Hustedt, and Detonula confervacea (cleve) gran. Can. J. Microbiol. 8(2):229-239.
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High Fragmentation Characterizes Tumour-Derived Circulating DNA
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PloS one
| 2,011
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cc-by
| 9,613
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Abstract Background: Circulating DNA (ctDNA) is acknowledged as a potential diagnostic tool for various cancers including
colorectal cancer, especially when considering the detection of mutations. Certainly due to lack of normalization of the
experimental conditions, previous reports present many discrepancies and contradictory data on the analysis of the
concentration of total ctDNA and on the proportion of tumour-derived ctDNA fragments. Methodology: In order to rigorously analyse ctDNA, we thoroughly investigated ctDNA size distribution. We used a highly
specific Q-PCR assay and athymic nude mice xenografted with SW620 or HT29 human colon cancer cells, and we correlated
our results by examining plasma from metastatic CRC patients. Conclusion/Significance: Fragmentation and concentration of tumour-derived ctDNA is positively correlated with tumour
weight. CtDNA quantification by Q-PCR depends on the amplified target length and is optimal for 60–100 bp fragments. Q-
PCR analysis of plasma samples from xenografted mice and cancer patients showed that tumour-derived ctDNA exhibits a
specific amount profile based on ctDNA size and significant higher ctDNA fragmentation. Metastatic colorectal patients
(n = 12) showed nearly 5-fold higher mean ctDNA fragmentation than healthy individuals (n = 16). bert B, Arnau Peyrotte E, Del Rio M, Ychou M, et al. (2011) High Fragmentation Characterizes Tumour-Derived Circulating DNA. PLoS
1371/journal.pone.0023418 ation: Mouliere F, Robert B, Arnau Peyrotte E, Del Rio M, Ychou M, et al. (2011) High Fragmentation Characterizes Tumour-Derived
E 6(9): e23418. doi:10.1371/journal.pone.0023418 Editor: Terence Lee, University of Hong Kong, Hong Kong Received May 5, 2011; Accepted July 16, 2011; Published September
, 2011
6 Copyright: 2011 Mouliere et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: A.R. Thierry is supported by INSERM (France). F. Mouliere is supported by a grant from the CNRS and the Region of Languedoc-Roussillon
(CNRS044406). The study was granted from the GEFLUC (France). The funders had no role in study design, data collection and analysis, decision to publish, or
preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Competing Interests: The authors have declared that no competing interests exist. * E-mail: alain.thierry@sysdiag.cnrs.fr biomarker
[9,10]. Particularly,
it
is
imperative
to
develop
technologies that can detect the number of mutant DNA molecules
and the specific mutation(s) in the same sample. Abstract For this it is
necessary to improve the sensitivity and specificity of ctDNA
analysis. In addition, in order to be able to compare results across
samples and studies, it is necessary to have detailed information
about the amount and the size distribution of ctDNA fragments and
to compare the data obtained for neoplastic and non-neoplastic cell
ctDNA. Necrosis, which is associated with tumour development, as
well as apoptosis and phagocytosis, which are associated with
defence mechanisms, lead to destruction of tumour cells and also of
the adjacent, non-tumour tissues [11,12,13,14], and to DNA
fragmentation as an hallmark of apoptosis. However, fragmentation
of cell-free DNA is higher following apoptosis than following
necrosis or phagocytosis [13]. Specifically, ctDNA fragments longer
than 10,000 bp are likely to originate from necrotic cells, whereas
DNA fragments shorter than 1000 bp, particularly of 180 bp or
multiples of this size, are reminiscent of the oligonucleosomal DNA
ladder observed in apoptotic cells [15]. Thus, a detailed analysis of
the size of the ctDNA fragments could allow discerning the source of
circulating nucleic acid in cancer patients. To this aim we developed
a highly specific Q-PCR assay that has already allowed us to
unequivocally identify tumour and non-tumour derived ctDNA Florent Mouliere1, Bruno Robert2, Erika Arnau Peyrotte1, Maguy Del Rio2, Marc Ychou3, Franck Molina1,
Celine Gongora2, Alain R. Thierry1* Florent Mouliere1, Bruno Robert2, Erika Arnau Peyrotte1, Maguy Del Rio2, Marc Ychou3, Franck Molina1,
Celine Gongora2, Alain R. Thierry1* 1 SysDiag UMR3145 – CNRS, National Centre of the Scientific Research/BIO-RAD, Montpellier, France, 2 U896 INSERM, National Institute of Health and Medical Research,
University Montpellier1, IRCM, Institute of Oncological Research of Montpellier, Montpellier, France, 3 CRLC, Regional Centre against Cancer, Val d’Aurelle-Paul Lamarque,
Montpellier, France 2. Xenograft model Female athymic nude mice (6–8 wk/old) were purchased from
Harlan (Gannat, France) and maintained in a specific pathogen-
free facility (study approval Nu B-34-172-27; Institut de Recherche
en
Cance´rologie
de
Montpellier-CRLC
Val
d’Aurelle-Paul
Lamarque, Montpellier, France). Mice were xenografted subcuta-
neously with 26106 cancer cells. Mice were euthanized with CO2
at different time points post-graft according to the tumour weight
desired. Peripheral blood was drawn into EDTA pre-coated tubes
and was used for plasma preparation within one hour. All
experiments complied with the current national and institutional
regulations and ethical guidelines and were performed by an
accredited person (Dr. B. ROBERT, Nu34-156). 6. Oligonucleotides The oligonucleotide primers used in this work are described in
Table S1. Primers were designed and selected according to the
previously described methods [16]. Fragmentation of Tumour-Derived Circulating DNA fluid protocol’’ in an elution volume of 60 ml. DNA samples were
kept at 220uC until use. No significant difference was found in the
results of the Q-PCR assays in which freshly extracted or stored
DNA was used. released during cancer progression in an animal model (athymic
nude mice xenografted with human colon cancer cells) [16]. Here,
we have used this assay to evaluate the amount and size distribution
of non-tumour and tumour-derived ctDNA fragments. We then
have applied the same system to quantify ctDNA fragments and
their size distribution in plasma samples from healthy individuals
and patients with metastatic colorectal cancer (CRC). Our data
revealed that analysis of ctDNA fragmentation provides valuable
information relative to cancer diagnosis. 1. Cell lines Human HCT116-s [17], SW620 (ATCC: CCL-227) and HT29
(ATCC: HTB-38) cells and mouse MC38 [18] colon carcinoma
cell lines were maintained in RPMI + 10% fetal bovine serum. Human HCT116-s [17], SW620 (ATCC: CCL-227) and HT29
(ATCC: HTB-38) cells and mouse MC38 [18] colon carcinoma
cell lines were maintained in RPMI + 10% fetal bovine serum. 7. CtDNA quantification by Q-PCR 7. CtDNA quantification by Q-PCR The Q-PCR method and ctDNA analysis were previously
described in detail [16]. Methodology and data description were
carried out according to the MIQE guidelines [19]. Q-PCR
amplifications were carried out in a reaction volume of 25 ml on a
Chromo4 instrument using the MJ Opticon Monitor 3 software
(Bio-Rad). Each PCR reaction mixture was composed of 12.5 ml
PCR mix (Bio-Rad Super mix SYBR Green); 2.5 ml of each
amplification primer (0.3 pmol/ml); 2.5 ml PCR-analyzed water
and 5 ml DNA extract. Thermal cycling was organised in 3
repeated steps: a first denaturation step of 3 min at 95uC, followed
by 40 repeated cycles of 95uC for 10 sec and 60uC for 30 sec. Melting curves were obtained by increasing the temperature from
55uC to 90uC with a plate reading every 0.2uC., Serial dilutions of
genomic DNA from human placenta cells were used as calibrators
for quantification, their concentration being assessed with a
Nanodrop spectrophotometer. Each sample was analyzed in
duplicate and each assay repeated at least once. The ctDNA
concentrations obtained with each primer set were normalized to
the precise concentration of a genomic DNA sample amplified
using the same primer set. The coefficient of variation due to
ctDNA extraction and of concentration value was calculated as
4% and 19% respectively (n = 12). The ctDNA amount was
arbitrarily estimated for the 60–100, 100–150, 150–400 and
.400 bp fragment size ranges. ctDNA amount was determined
for 1 ml and expressed as ng. The estimated amount of ctDNA in
each range of ctDNA size was calculated by subtracting the
amount obtained with the larger amplicon from the amount
obtained with the shorter amplicon (100 bp minus 60 bp, 150 bp
minus 100 bp, 400 bp minus 150 bp). The fraction corresponding
to ctDNA fragment higher than 400 bp corresponds to the
amount obtained when targeting sequence of 382 bp (mouse) or
409 bp (human). 8. Estimation of ctDNA fragmentation CtDNA fragmentation was assessed by calculating an index we
termed DNA Integrity Index (DII). DII was calculated as the ratio
of larger/shorter
fragment concentrations. DII is therefore
theoretically 1 if the template DNA is not truncated and ,1 if it
is truncated into fragments smaller than the template. Conven-
tionally, long and short fragments were longer or shorter than
180 bp (i.e., the mononucleosome average size). To note, DII did
not significantly vary when Q-PCR analysis was either carried out
2 hours following blood sampling or after plasma freezing,
provided that no blood degradation was observed after 4 hours
in
EDTA
(ethylenediaminetetraacetic
acid)
tubes
at
room
temperature (data not shown). In addition, samples were handled
in the same way in the same experiments. 3. Human blood samples Blood samples from patients with metastatic colorectal cancer
(CRC) (n= 12) and from selected human healthy individuals (human
healthy plasma, HHP, n = 16) were collected in EDTA tubes. The
overall process from blood collection to plasma preparation did not
exceed 3 hours and plasma was stored at 280Cu. Written, informed
consent was obtained from all participants prior to the onset of the
study. Protocols for the use of blood of healthy volunteers used in this
study were approved by the ‘‘Etablissement Franc¸ais du Sang’’ (EFS)
Ethics
Committee
(EFS-PM
agreement:
21/PVNT/MTP/
CNR14/2010-0029). According to the Code de Sante´ Publique
Article L1131-1 and next, no specific ethical approval is required for
this type of study. Each CRC and HHP sample was numbered
(CRCn and HHPn) and corresponded to the plasma of a single
subject with the exception of HHP3, which was a pool of 6 different
plasma samples. TNM classification of CRC 1–4 patients corre-
spond to T4N3M1, T3N2M1, T4N1M1 and TxN2M1, respective-
ly. All metastatic CRC patients did not received chemotherapy at
least one month before blood collection. 4. Plasma preparation Mouse and human blood was collected in 4 ml Venosafe K2E
tubes (Terumo) and centrifuged at 1200 g at 4uC in a Heraeus
Multifuge LR centrifuge with a CR 4j rotor for 10 minutes. Supernatants were isolated in sterile 1.5 ml Eppendorf tubes and
centrifuged at 16 000 g at 4uC for 10 min. Then, supernatants
were either immediately handled for DNA extraction or stored at
280uC. No significant difference was found in Q-PCR assays
when comparing freshly isolated or stored plasma. Introduction Recent advances in the understanding of the molecular
mechanisms of cancer enlarge the scope of diagnostic strategies in
oncology [1]. Many genetic alterations involved in tumourigenesis
are now positioned within a time sequence [2]. Moreover it is fully
acknowledged that tumours release cell-free nucleic acids in
biological fluids, especially blood [3]. Therefore, the detection of
the genetic alterations associated with tumour development in
circulating DNA (ctDNA) appears to be a particularly attractive
approach as a non-invasive, diagnostic or theranostic test for
cancers [4]. Previously, ctDNA was used as biomarker in two ways:
first by measuring its concentration and second by studying its
nucleotidic sequence [4,5]. Moreover, ctDNA levels in plasma have
been shown (i) to be significantly higher in CRC patients than in
healthy individuals [6]; (ii) to decrease progressively in the follow-up
period in tumour-free patients; and (iii) to increase in patients with
cancer recurrence or metastasis [5,7,8]. Many studies have been
focusing on the identification of abnormal forms of DNA in plasma
or serum; however, the reported results are so far contradictory,
although very high rates of cancer detection have been obtained in
such a way. These reports, although promising, have led to many
questions about the reliability of using abnormal ctDNA as cancer PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 1 PLoS ONE | www.plosone.org Fragmentation of Tumour-Derived Circulating DNA 1. ctDNA fragmentation increases with tumour size and
tumour ctDNA concentration The amount profile of ctDNA fragments of different size was
assessed using an integrated PCR system that target intronic
sequences of mouse or human origin within the same region. For
the human wild type specific primer set the designed sizes of
amplification were 60 bp, 73 bp, 101 bp, 145 bp, 185 bp, 249 bp,
300 bp, 357 bp and 409 bp (the reverse primer was the same for
each amplification system). These primers targeted intron 2 of
human KRAS (Fig. 1, C). For the mouse wild type specific primer
set the designed sizes of amplification were 63 bp, 95 bp, 150 bp,
196 bp, 256 bp, 284 bp, 354 bp and 382 bp (the reverse primer
was the same for each amplification system). These primers
targeted intron 7 of mouse KRAS (Fig. 1, D). The median of the
non-specific amplification of the mouse and human KRAS primer
sets, assessed by determining the concentration profile of aliquots
of human placental DNA and of MC38 DNA, was 7% and 9%,
respectively. Nude athymic mice xenografted with HT29 cells were sacrificed
at different time points, tumours were weighted and ctDNA
concentration in plasma samples determined using the HuACTB
133 primer pair that targets a 133 bp sequence in the human
ACTB gene (Fig. 2, A). The concentration of ctDNA increased
slightly (from 0.23 to 4.8 ng/ml) in samples from mice bearing
small tumours (,500 mg), but then rapidly augmented up to
19 ng/ml in samples from mice with bigger tumours (.500 mg). Then, the corresponding DII was calculated using the ctDNA
concentrations obtained with this primer pair and the values
obtained with the HuACTB 290 primer set that amplifies a longer
amplicon (290 bp). DII sharply decreased (from 0.44 to 0.1) in
samples from mice with tumours up to 400 mg and then remained
constant at approximately 0.05, notwithstanding the steadily
increase in tumour weight. 5. DNA extraction Targeted sequences are located in two genes: KRAS and ACTB
(Table S1). Evaluation of ctDNA fragmentation was performed
using an original double integrated PCR system that targets ctDNA was isolated from 200 ml plasma with the QIAmp DNA
Mini Blood kit (Qiagen, CA) according to the ‘‘Blood and body PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 2 Fragmentation of Tumour-Derived Circulating DNA Fragmentation of Tumour-Derived Circulating DNA human ACTB sequences [16] or intron 2 of the mouse or human
KRAS gene in which the reverse primer was the same for the short
and the long fragment (Fig. 1, A and B). 10. Statistical analysis Data were expressed as mean 6 SD. The Student’s t-test was
used for comparison of means. A probability of less than 0.05 was
considered to be statistically significant; *p # 0.05, **p # 0.01,
***p # 0.001. Since the previous results indicate that ctDNA fragmentation
increases with tumour progression, we then tested using our PCR
system whether the evaluation of ctDNA concentration could be Figure 1. Primer designs used in the study. Primer targeting an ACTB region for quantifying ctDNA and determining ctDNA fragmentation from
HT29 xenografted mice (A). Primer targeting a human KRAS region for studying the effect of the target sequence size on ctDNA concentration and
fragmentation in plasma from SW620 xenografted mice (B). Primer design for studying the size distribution profile of ctDNA fragments in plasma
from SW620 xenografted mice and human individuals by targeting human (C) and mouse (D) KRAS region. doi:10.1371/journal.pone.0023418.g001 Figure 1. Primer designs used in the study. Primer targeting an ACTB region for quantifying ctDNA and determining ctDNA fragmentation from
HT29 xenografted mice (A). Primer targeting a human KRAS region for studying the effect of the target sequence size on ctDNA concentration and
fragmentation in plasma from SW620 xenografted mice (B). Primer design for studying the size distribution profile of ctDNA fragments in plasma
from SW620 xenografted mice and human individuals by targeting human (C) and mouse (D) KRAS region. doi:10.1371/journal.pone.0023418.g001 September 2011 | Volume 6 | Issue 9 | e23418 PLoS ONE | www.plosone.org 3 Fragmentation of Tumour-Derived Circulating DNA Figure 2. Evolution of tumour-derived ctDNA fragmentation and concentration relative to tumour weight. (A): Athymic nude mice
were xenografted with HT29 cells and sacrificed at various time points and thus with tumours of increasing weight (137, 189, 310, 380, 400, 1019,
1080 mg from Mo1 to Mo7, respectively). CtDNA concentration (ng/ml plasma, left y axis) (dashed line) was assessed by Q-PCR using a primer set that
amplifies a 133 bp sequence from human ACTB (Fig. 1A). Tumour-derived ctDNA fragmentation (full line) was estimated by calculating the DII (right y
axis). Here, the DII corresponds to the ratio of the concentration found when targeting a 290 bp sequence of the human ACTB gene to the
concentration found when targeting a 133 bp sequence located within the former 290 bp region (Fig. 1A). 10. Statistical analysis (B): Tumour ctDNA was quantified by Q-
PCR with human KRAS 73 (empty bars), KRAS 145 (hatched bars) and KRAS 300 (full bars) primer sets (Fig. 1B) in plasma samples from SW620
xenografted mice. Mo8, Mo9 and Mo10 correspond to three non-xenografted nude mice. CtDNA fragmentation was estimated by calculating the
300/145 and 300/73 DII. DII was calculated as the ratio of the ctDNA concentrations obtained from the amplification of a short 73 bp, a medium
145 bp and a long 300 bp target sequence from the human KRAS gene (Fig. 1B). Bars represent ctDNA concentration expressed as ng/ml of plasma
(left y axis) and curve represents tumour weight (mg, right y axis). Data represent the mean values of the ctDNA concentrations obtained in
duplicates. NC, not calculated. No ctDNA was detected with the KRAS 300 bp primers in Mo11 and Mo12 plasmas. doi:10.1371/journal.pone.0023418.g002
Fragmentation of Tumour Derived Circulating DNA Figure 2. Evolution of tumour-derived ctDNA fragmentation and concentration relative to tumour weight. (A): Athymic nude mice
were xenografted with HT29 cells and sacrificed at various time points and thus with tumours of increasing weight (137, 189, 310, 380, 400, 1019,
1080 mg from Mo1 to Mo7, respectively). CtDNA concentration (ng/ml plasma, left y axis) (dashed line) was assessed by Q-PCR using a primer set that
amplifies a 133 bp sequence from human ACTB (Fig. 1A). Tumour-derived ctDNA fragmentation (full line) was estimated by calculating the DII (right y
axis). Here, the DII corresponds to the ratio of the concentration found when targeting a 290 bp sequence of the human ACTB gene to the
concentration found when targeting a 133 bp sequence located within the former 290 bp region (Fig. 1A). (B): Tumour ctDNA was quantified by Q-
PCR with human KRAS 73 (empty bars), KRAS 145 (hatched bars) and KRAS 300 (full bars) primer sets (Fig. 1B) in plasma samples from SW620
xenografted mice. Mo8, Mo9 and Mo10 correspond to three non-xenografted nude mice. CtDNA fragmentation was estimated by calculating the
300/145 and 300/73 DII. DII was calculated as the ratio of the ctDNA concentrations obtained from the amplification of a short 73 bp, a medium
145 bp and a long 300 bp target sequence from the human KRAS gene (Fig. 1B). Bars represent ctDNA concentration expressed as ng/ml of plasma
(left y axis) and curve represents tumour weight (mg, right y axis). September 2011 | Volume 6 | Issue 9 | e23418 3. Tumour-derived ctDNA fragments exhibit a specific
size distribution in xenografted mice The previous results indicate that quantification of tumour
ctDNA is influenced by the amplicon size. We thus asked whether
the size distribution profiles of neoplastic and non-neoplastic
ctDNA fragments were different. In order to reduce as much as Figure 3. CtDNA fragment size profiles determined by Q-PCR for amplicons of different length in xenografted mice. Evaluation of
tumour and non-tumour ctDNA concentration in plasma samples from mice xenografted with SW620 cells and of control ctDNA in non-xenografted
nude mice was determined using a multi-integrated Q-PCR system for detecting amplicons of increasing length in intron 2 of human wild type KRAS
(Fig. 1) or in intron 7 of mouse wild type KRAS (Fig. 1). Tumour ctDNA, non-tumour ctDNA and control ctDNA concentrations are expressed as ng/ml
of plasma (A) or as % of the highest value of each concentration set (B). Each point represents the mean of three pools of plasma from three mice
with 300–550 mg tumours. The relative percentage of tumour and non-tumour ctDNA from xenografted mice for each amplicon length is presented
in (C): Tumour (black bars) and non-tumour (hatched bars) ctDNA mean concentrations are expressed as ng/ml of plasma; the bar height is the sum
of tumour and non-tumour ctDNA concentrations (estimated as the total ctDNA concentration). For clarity human and mouse amplified targeted
sequences of close size were grouped together: 60 (60 and 63), 100 (101 and 95), 150 (145 and 150), 200 (185 and 196), 250 (249 and 256), 350 (354
and 357) and 400 (409 and 382) bp. Fractional fragment size distribution of ctDNA amount from tumour and non-tumour ctDNA in xenografted mice
and control ctDNA in non-xenografted mice (D). The ctDNA amount was arbitrarily estimated for the 60–100, 100–150, 150–400 and .400 bp
fragment size ranges. The estimated amount of ctDNA (ng) in one ml of plasma in each range of ctDNA size was calculated as described in Materials
and Methods section. doi:10.1371/journal.pone.0023418.g003 Figure 3. CtDNA fragment size profiles determined by Q-PCR for amplicons of different length in xenografted mice. Evaluation of
tumour and non-tumour ctDNA concentration in plasma samples from mice xenografted with SW620 cells and of control ctDNA in non-xenografted
nude mice was determined using a multi-integrated Q-PCR system for detecting amplicons of increasing length in intron 2 of human wild type KRAS
(Fig. 1) or in intron 7 of mouse wild type KRAS (Fig. 1). Fragmentation of Tumour-Derived Circulating DNA Fragmentation of Tumour-Derived Circulating DNA of ctDNA concentration is strongly influenced by the choice of the
amplicon length. For instance, the ctDNA concentration in mouse
#16 determined with the KRAS 73 primers was 5.3- and 40.5-fold
higher than the concentrations obtained with the KRAS 145 and
KRAS 300 primer pairs, respectively. The DII was then calculated
(Fig. 2, B). Although both KRAS 300/145 and KRAS 300/73 DII
decreased with tumour progression (13- and 30-fold decrease,
respectively), KRAS 300/145 DII remained more elevated than
300/73 DII (for instance the mouse #16 DII was 0.13 for the
KRAS 300/145 ratio and 0.02 for the KRAS 300/73 ratio). possible the effect of tumour size, ctDNA was quantified by
calculating the mean concentration values in three pools of plasma
from three mice with tumours weighing between 300 and 550 mg
(Fig. 3). The concentration profile of the different non-tumoral
ctDNA fragments in these pooled samples, obtained using mouse
KRAS primers that amplify fragments of increasing length up to
382 bp, was similar to that of control, non-xenografted mice
(Fig. 3A and B). With human KRAS primers that amplify tumour-
derived ctDNA fragments of increasing length up to 409 bp, the
highest ctDNA concentrations were obtained for amplicons of
length ,100 bp and were much higher than the values obtained
for mouse (non-tumoral and control) ctDNA. Conversely, when
detecting
amplicons
of
size
ranging
from
100
to
150 bp
comparable levels of non-tumour, tumour-derived and control
ctDNA were obtained, and for amplicons .185 bp lower
concentrations of tumoral than non-tumoral ctDNA were found
(p = 0.047 and 0.007 for the 185 bp and 250 bp amplicons, 10. Statistical analysis Data represent the mean values of the ctDNA concentrations obtained in
duplicates. NC, not calculated. No ctDNA was detected with the KRAS 300 bp primers in Mo11 and Mo12 plasmas. doi:10.1371/journal.pone.0023418.g002 influenced by the amplicon length. To this aim, the concentration
of tumour ctDNA in plasma samples of mice xenografted with
SW620 cells was quantified using three human KRAS primer pairs
that amplify sequences of 73 bp, 145 bp and 300 bp (Fig. 2, B)
and with KRAS primers targeting mouse and human KRAS (214
and 189 bp, Figure S1). With the KRAS 300 primers (300 bp
amplicon) ctDNA was not detected in plasma samples from the
mice with the smallest tumours (,174 mg), but then, in samples
from mice with tumours heavier than 174 mg, the ctDNA concentration decreased progressively with the increase of tumour
weight. On the other hand, the ctDNA concentrations obtained
using the KRAS 145 primer set seemed to slightly increase with
tumour size. Conversely, with the KRAS 73 primers the ctDNA
concentration highly increased in parallel with tumour weight. Data obtained by detecting 145 and 73 bp KRAS amplicon
confirmed the increase of ctDNA concentration with tumour
weight observed in HT29 cells by detecting 133 bp ACTB
amplicon (Fig. 2, A). These results indicate that the quantification September 2011 | Volume 6 | Issue 9 | e23418 PLoS ONE | www.plosone.org 4 4. The specific size distribution profile of tumour ctDNA
fragments appears to be relevant also in CRC patients HHP and CRC ctDNA concentration are
expressed as % of total ctDNA value of each profile (B). CtDNA
concentration was determined using a multi-integrated Q-PCR system
for detecting amplicons of increasing length in intron 2 of human wild
type KRAS (Fig. 1) as performed in Fig. 3. doi:10.1371/journal.pone.0023418.g004 Figure 4. ctDNA size profile upon amplicon length. Comparison
of ctDNA size profile upon amplicon length in healthy individuals (HHP)
and CRC patients (A). CtDNA amount was calculated as described in
Material and Methods section. The corresponding concentration values
for each CRC and HHP plasma sample are presented in Table 1. HHP is
the mean from plasmas of two healthy individuals (HHP1 and HHP2)
and of a pool of 7 healthy individuals (HHP3). The estimated amount of
ctDNA in each range of ctDNA size was calculated as described in
Materials and Methods section. HHP and CRC ctDNA concentration are
expressed as % of total ctDNA value of each profile (B). CtDNA
concentration was determined using a multi-integrated Q-PCR system
for detecting amplicons of increasing length in intron 2 of human wild
type KRAS (Fig. 1) as performed in Fig. 3. doi:10.1371/journal.pone.0023418.g004 5. Comparison of the fractional size distribution of ctDNA
in CRC patients and xenografted animals Figure 5 shows the relative ctDNA amount from plasmas of
human (Fig. 5A) and mouse (Fig. 5B) origin. The percentage of
60–150 bp ctDNA was rather similar in HHP and CRC samples
despite the higher proportion of tumour ctDNA observed for the
60–100 bp range. Decreased but significant ctDNA amounts for
the 150–400 range were detected in CRC samples, but not in
HHP samples. Conversely, the amount of .400 bp ctDNA in
HHP samples (approximately 35%) was 17-fold higher than in
CRC samples (2%). non-tumoral mouse ctDNA). Conversely, the percentage of tumour-
derived ctDNA in the 60–100 bp range was very high (71%) and
decreased sharply to 2% for fragments .400 bp. The fractional
fragment size distribution of tumour-derived ctDNA from xeno-
grafted mice and CRC patients was comparable (Fig. 5A and B). A similar method of evaluation of the ctDNA amount was
employed for the data (Fig. 3) obtained in xenografted mice (Fig. 5,
B). The amount of mouse ctDNA relative to fragment size range was
comparable in non-xenografted (control) and xenografted mice
(non-tumoral) and its distribution profile was similar to the one
observed in healthy controls (Fig. 5, A) with a noticeable very low
percentage in the 150–400 size range and a significant higher
proportion of .400 bp ctDNA (39% and 52% for control and 6. Comparison of the DII in healthy and CRC patients and
in xenografted animals 3. Tumour-derived ctDNA fragments exhibit a specific
size distribution in xenografted mice These results indicate that 1) ctDNA fragments
smaller than 150 bp are predominantly detected in tumour
ctDNA extract; 2) the proportion of ctDNA of tumour origin is
inversely correlated with the amplicon size (from less than 20% of
the total detectable ctDNA for the 400 bp amplicon to 85% for the
60 bp
amplicon)
(Fig. 3C). Finally,
ctDNA
amount
upon
arbitrarily selected size ranges (Fig. 3D) highlights that tumour-
derived ctDNA amount could be easily discriminated versus non
tumour and control ctDNA when targeting sequence ,100 bp (65
vs 2–5 ng) and when targeting sequence .400 bp (2 vs 9–10 ng). 3. Tumour-derived ctDNA fragments exhibit a specific
size distribution in xenografted mice Tumour ctDNA, non-tumour ctDNA and control ctDNA concentrations are expressed as ng/ml
of plasma (A) or as % of the highest value of each concentration set (B). Each point represents the mean of three pools of plasma from three mice
with 300–550 mg tumours. The relative percentage of tumour and non-tumour ctDNA from xenografted mice for each amplicon length is presented
in (C): Tumour (black bars) and non-tumour (hatched bars) ctDNA mean concentrations are expressed as ng/ml of plasma; the bar height is the sum
of tumour and non-tumour ctDNA concentrations (estimated as the total ctDNA concentration). For clarity human and mouse amplified targeted
sequences of close size were grouped together: 60 (60 and 63), 100 (101 and 95), 150 (145 and 150), 200 (185 and 196), 250 (249 and 256), 350 (354
and 357) and 400 (409 and 382) bp. Fractional fragment size distribution of ctDNA amount from tumour and non-tumour ctDNA in xenografted mice
and control ctDNA in non-xenografted mice (D). The ctDNA amount was arbitrarily estimated for the 60–100, 100–150, 150–400 and .400 bp
fragment size ranges. The estimated amount of ctDNA (ng) in one ml of plasma in each range of ctDNA size was calculated as described in Materials
and Methods section. doi:10.1371/journal.pone.0023418.g003 doi:10.1371/journal.pone.0023418.g003 September 2011 | Volume 6 | Issue 9 | e23418 PLoS ONE | www.plosone.org 5 Fragmentation of Tumour-Derived Circulating DNA Figure 4. ctDNA size profile upon amplicon length. Comparison
of ctDNA size profile upon amplicon length in healthy individuals (HHP)
and CRC patients (A). CtDNA amount was calculated as described in
Material and Methods section. The corresponding concentration values
for each CRC and HHP plasma sample are presented in Table 1. HHP is
the mean from plasmas of two healthy individuals (HHP1 and HHP2)
and of a pool of 7 healthy individuals (HHP3). The estimated amount of
ctDNA in each range of ctDNA size was calculated as described in
Materials and Methods section. HHP and CRC ctDNA concentration are
expressed as % of total ctDNA value of each profile (B). CtDNA
concentration was determined using a multi-integrated Q-PCR system
for detecting amplicons of increasing length in intron 2 of human wild respectively). 4. The specific size distribution profile of tumour ctDNA
fragments appears to be relevant also in CRC patients fragments appears to be relevant also in CRC patients
The same human KRAS primer sets that amplify DNA
fragments of different size (Fig. 1, C) was used to study the
ctDNA
size
distribution
in
plasma
samples
from
healthy
individuals (HPP) and four metastatic CRC patients who were
not under treatment (CRC1 to CRC4) (Fig. 4). Overall higher
ctDNA concentrations were detected in CRC plasma samples
than in HHP samples (Table 1). For instance, the plasma DNA
concentrations determined with the primers that amplify the 60 bp
amplicon were 8078, 2134, 618 and 402 ng/ml for CRC1, CRC2,
CRC3 and CRC4 and 24.8, 11.2 and 18.4 ng/ml in HHP1,
HHP2 and HHP3, respectively. Moreover, whereas among CRC
patients ctDNA levels were very different, in healthy individuals
they did not vary much as the mean +/2 SD was 18.1+/26.8. To
note, the value of HHP3, which is a pool of plasmas from 7
different healthy donors, was similar to that of the individual
HHP1 and HHP2 samples. The mean ctDNA amount in human
healthy individuals was found maximal at 60–100 bp (49%) range
while being very low at 150–400 bp range (1%). Except for the
.400 bp range HHP ctDNA amount was much lower (of several
order) than that of CRC ctDNA amount observed when fragments
are ,400 bp. In particular, CRC ctDNA amount observed within
150–400 bp rise to 178–1520 ng whereas HHP amount was very
poor (,1 ng). Conversely, the relative CRC ctDNA amount
detected for fragment .400 bp appears very low (1–5%),
suggesting that the relative determination of ctDNA amount
within a fragment size range might distinguish healthy vs CRC
plasmas. In this regard, the 300/60 DII calculated from the
concentration values obtained when detecting target sequence of
300 and 60 bp, was 0.45 for the HHP and 0.055 for the CRC
samples (Table 1). Figure 4. ctDNA size profile upon amplicon length. Comparison
of ctDNA size profile upon amplicon length in healthy individuals (HHP)
and CRC patients (A). CtDNA amount was calculated as described in
Material and Methods section. The corresponding concentration values
for each CRC and HHP plasma sample are presented in Table 1. HHP is
the mean from plasmas of two healthy individuals (HHP1 and HHP2)
and of a pool of 7 healthy individuals (HHP3). The estimated amount of
ctDNA in each range of ctDNA size was calculated as described in
Materials and Methods section. PLoS ONE | www.plosone.org 6. Comparison of the DII in healthy and CRC patients and
in xenografted animals DII was evaluated by calculating the ratio between the concentration of each amplicon and that of the 60 bp amplicon. HHP/CRC is the ratio between the mean HHP DII
and the mean CRC DII. doi:10.1371/journal.pone.0023418.t001 Since it was previously established that the shortest ctDNA should
theoretically be .180 bp (size of a nucleosome) and since Q-PCR
efficiency is conventionally (with genomic DNA) optimal when
amplifying sequence of 150–300 bp, most of the previous works
relied on the detection of amplicons of about 150 bp when
analyzing ctDNA. While Q-PCR assay was often used for ctDNA
study, only a couple of reports described the use of primers that
amplify segments ,100 bp and this choice was based solely on
technical aspects linked to primer design and/or to the targeted
gene region, but never on the assumption that a higher proportion
of ,100 bp ctDNA could be present in a specific physiological
condition or pathology. Here, we demonstrate, for the first time, the
presence of a higher proportion of ctDNA fragments ,100 bp that
is directly correlated with the increase of ctDNA concentration,
particularly in samples from cancer patients. Thus, contrary to the
analysis of genomic DNA in which the concentration of quantified
DNA is directly proportional to the number of amplified copies, this
proportionality varies upon target sequence length in the case of
ctDNA from CRC patients. Moreover, the targeting of sequences of
150 to 250 bp (the length commonly chosen for amplification)
generates a significant bias by not taking into account up to 80% of
the ctDNA amount. Thus, the size profile of ctDNA, as determined
by amplifying targets of increasing length, reveals that the optimal
detection is obtained with amplicons ,100 bp and that a much
higher proportion of ctDNA of size ranging from 150 to 400 bp is
present in non-tumour ctDNA than in tumour ctDNA. The choice
of the size of the amplified DNA region consequently appears
critical. Moreover 98% of human CRC tumour ctDNA fragments
were ,409 bp. amplicons (145–409 and .409 bp). DII from mouse plasmas
between the ctDNA concentration of each amplicon .60 bp and
that of the 60 bp amplicon was calculated (Table S2). The DII
ratios can clearly discriminate tumour ctDNA from non-tumour
ctDNA detected in xenografted or non-xenografted mice. 6. Comparison of the DII in healthy and CRC patients and
in xenografted animals The
profile of the DII ratios of non-xenografted and xenografted mice
remarkably appears similar to those calculated from human
healthy individuals and CRC patients. DII appears to similarly
discriminate tumour-derived ctDNA from non tumour-derived or
control ctDNA. Results strongly confirmed that 300/60 DII seems
optimal for discriminating healthy to CRC subjects (Fig. 6). DII
mean value of HHP (n = 16) is significatively different from DII
mean value of CRC (n = 12) (mean, 0.565 and 0.122, respectively;
p,0.001) (Fig. 7). A similar difference was observed in an animal
model where mean DII is 0.447 for healthy plasma ctDNA (n = 9),
0.645 for non tumour-derived ctDNA (n = 9) and 0.027 for
tumour derived ctDNA (n = 9). The DII values did not seem to be
influenced by the genome localization of the amplicons, since
comparable results were obtained with amplicons localized in
intron 2 and exon 2 of KRAS and in exon 15 of BRAF (0.036+/
20.006, data not shown). 6. Comparison of the DII in healthy and CRC patients and
in xenografted animals The ratio between the mean HHP DII and the CRC DII was
close to 1 for the 60/60 DII and 1.5 for 150/60 DII bp, but
sharply increased to 4–19 for the 300/60 DII (Fig. 6). This
indicates that the main differences in ctDNA fragmentation
between healthy and cancer patients are observed for long PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 6 Fragmentation of Tumour-Derived Circulating DNA Table 1. Total ctDNA concentration (ng/ml plasma) and DII in plasma samples from metastatic CRC patients (CRC1-4) and healthy
individuals (HHP1-3). Table 1. Total ctDNA concentration (ng/ml plasma) and DII in plasma samples from metastatic CRC patients (CRC1-4) and healthy
individuals (HHP1-3). Samples
Amplicons sizes (bp)
60
73
101
145
185
249
300
357
409
CRC 1
8077.8
6004.8
2395.5
1526.2
815.2
275.9
179.3
162.6
6.8
CRC 2
2133.9
2326.0
1016.5
596.2
409.3
267.2
175.1
105.1
123.9
CRC 3
618.1
591.5
289.9
187.7
93.2
46.1
37.5
20.3
9.9
CRC 4
402.2
286.3
179.0
77.4
81.3
135.3
62.9
28.1
34.1
HHP 1
24.8
16.5
9.8
5.9
7.0
6.0
3.6
2.4
3.0
HHP 2
11.2
12.8
7.6
6.1
6.4
6.8
8.0
5.2
7.4
HHP 3
18.4
17.3
11.6
6.4
5.2
9.8
9.0
4.4
4.7
Samples
Integrity index
60/60
73/60
101/60
145/60
185/60
249/60
300/60
357/60
409/60
CRC 1
1.000
0.743
0.297
0.189
0.101
0.034
0.022
0.020
0.001
CRC 2
1.000
1.090
0.476
0.279
0.192
0.125
0.082
0.049
0.058
CRC 3
1.000
0.957
0.469
0.304
0.151
0.075
0.061
0.033
0.016
CRC 4
1.000
0.712
0.445
0.192
0.202
0.336
0.156
0.070
0.085
mean CRC
1.000
0.876
0.422
0.241
0.161
0.143
0.080
0.043
0.040
HHP 1
1.000
0.664
0.394
0.239
0.284
0.244
0.147
0.098
0.123
HHP 2
1.000
1.145
0.680
0.547
0.571
0.607
0.712
0.463
0.659
HHP 3
1.000
0.941
0.628
0.348
0.282
0.531
0.490
0.237
0.257
mean HHP
1.000
0.916
0.567
0.378
0.379
0.461
0.450
0.266
0.346
HHP/CRC
1.000
1.047
1.345
1.568
2.348
3.232
5.597
6.180
8.673
DII was evaluated by calculating the ratio between the concentration of each amplicon and that of the 60 bp amplicon. HHP/CRC is the ratio between the mean HHP DII
and the mean CRC DII. doi:10.1371/journal.pone.0023418.t001 Table 1. Total ctDNA concentration (ng/ml plasma) and DII in plasma samples from metastatic CRC patients (CRC1-4) and healthy
individuals (HHP1-3). PLoS ONE | www.plosone.org Discussion CRC and healthy patients (A) and tumour and non-tumour
ctDNA in xenografted mice and control ctDNA in non-xenografted mice
(B). The estimated amount of each ctDNA fraction was expressed as the
percentage to the total ctDNA amount estimated as the sum of the
ctDNA amount of the four size fractions. Data were calculated from the
experiments presented in Table 1, Fig. 3 (mouse plasma samples) and
Fig. 4 (human plasma samples). doi:10.1371/journal.pone.0023418.g005 prostate [20] and testicular cancer patients [21] and suggested that
it witnesses the level of apoptosis among other mechanisms which
may release DNA from tumour cells to the circulation. In addition,
we think that DII could be very informative in regards to the
ctDNA fragment size profile and thus to the observation of the
tumour origin. Amplification of fragments of increasing size within the same
DNA region showed that non-tumoral ctDNA is less fragmented
than tumour ctDNA. In addition, the study revealed that tumour
ctDNA fragments have a distinct size profile in comparison to non-
tumoral ctDNA and ctDNA from non-xenografted mice. Frag-
ment size profile of non-tumoral, tumoral and control ctDNA are
similar in the animal model and in the clinical samples. Specifically, our results suggest that the concentration level and
the size distribution profile of ctDNA fragments can be used to
discriminate between healthy and cancer patients. Particularly,
three ctDNA size groups (,100, 150–400 and .400 bp) might be
relevant for such discrimination; exhibiting a higher, much higher
and much lower ctDNA amount in cancer patients as compared
with healthy individuals, respectively. Thus, the specific analysis of
tumour ctDNA can be performed by using a DII calculated from
the concentrations of ctDNA of size ,100 bp and between 145–
409 bp and appears to be optimal when using the 300/60 DII in
both xenografted mice and untreated CRC patients. The
determination of DII significatively discriminates between HHP
and CRC plasmas indicating a higher fragmentation of the
corresponding ctDNA. Higher number of subjects is needed to
confirm this observation. Ellinger et al determined an integrity
index by targeting 124 bp and 271 bp sequences in plasma of As a consequence the choice of target length is crucial when
determining the DII. Discussion Here we demonstrate that the length of the amplified fragment
dramatically influences the measurement of the ctDNA concen-
tration by Q-PCR. Specifically, a significant higher proportion of
tumour ctDNA is detected by primers that target amplicons
,100 bp, suggesting that the detection of amplicons ,100 bp is
more relevant for tumour ctDNA. PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 7 Fragmentation of Tumour-Derived Circulating DNA Figure 5. Comparison of the fractional fragment size distribu-
tion of ctDNA from clinical samples and from the animal
model. CRC and healthy patients (A) and tumour and non-tumour
ctDNA in xenografted mice and control ctDNA in non-xenografted mice
(B). The estimated amount of each ctDNA fraction was expressed as the
percentage to the total ctDNA amount estimated as the sum of the
ctDNA amount of the four size fractions. Data were calculated from the
experiments presented in Table 1, Fig. 3 (mouse plasma samples) and
Fig. 4 (human plasma samples). Figure 6. Discrimination between healthy and CRC subjects by
comparing DII values. DII ratio between HHP and CRC patients (A)
and between control non-xenografted and SW620 xenografted mouse
plasma (B) were determined from DII values represented in Table 1 and
S2, respectively. DII of plasma from each CRC patient was compared
with the mean of healthy individuals (HHP, n = 9). Mouse DII ratios are
determined from the mean of 3 pools from 3 mice (n = 9). The DII was
estimated by the ratio of the concentration of amplicons of increasing
size and the concentration to the 60 bp amplicons. doi:10.1371/journal.pone.0023418.g006 Figure 6. Discrimination between healthy and CRC subjects by
comparing DII values. DII ratio between HHP and CRC patients (A)
and between control non-xenografted and SW620 xenografted mouse
plasma (B) were determined from DII values represented in Table 1 and
S2, respectively. DII of plasma from each CRC patient was compared
with the mean of healthy individuals (HHP, n = 9). Mouse DII ratios are
determined from the mean of 3 pools from 3 mice (n = 9). The DII was
estimated by the ratio of the concentration of amplicons of increasing
size and the concentration to the 60 bp amplicons. doi:10.1371/journal.pone.0023418.g006 Figure 5. Comparison of the fractional fragment size distribu-
tion of ctDNA from clinical samples and from the animal
model. September 2011 | Volume 6 | Issue 9 | e23418 1. Sawyers CL (2008) The cancer biomarker problem. Nature 452: 548–552.
2. Smith G, Carey FA, Beattie J, Wilkie MJ, Lightfoot TJ, et al. (2002) Mutations in
APC, Kirsten-ras, and p53—alternative genetic pathways to colorectal cancer.
Proc Natl Acad Sci U S A 99: 9433–9438. Acknowledgments The authors wish to thank Dr. E. Crapez and P.J. Lamy (Institut de
Recherche en Cance´rologie de Montpellier-CRLC, Montpellier, France)
for useful discussions. The authors thank Dr. D. Pezet and B. Gillet (CHU
Estaing, Clermont-Ferrand, France) for their help with plasma collection. We thank K. Kaminski and J. Balicchi (SysDiag CNRS-BioRad) for
helping us on the statistical analysis. Vogelstein’s group reported that 1350 to 230 000 ctDNA
fragments/ml were found in CRC patients’ plasma samples in
comparison to the 1150 to 8280 fragments/ml in disease-free
patients [9], confirming the partial overlap between ctDNA level
in healthy and CRC individuals described also in other studies
[3,4,5,28]. In our study, the fragment number in healthy controls (DOC) Figure S1
CtDNA concentration determined by targeting a 214
bp sequence in intron 2 of mouse KRAS and a 189 bp sequence in
intron 2 of human KRAS in plasma samples from mouse 8 to 16
(previously tested in Fig. 1B). (DOC) Discussion This
might certainly be due to the Q-PCR analysis that targets a 60 bp
sequence and/or the clinical state of the CRC patients group at
blood sample time we examined (untreated mCRC). This
specificity level may be as well worth further study with a much
higher number of clinical samples. In conclusion, higher and more accurate ctDNA quantification
is now made possible enabling a novel examination of ctDNA as
cancer biomarker. Size profiling or DII determination could
appear as additional potential biomarker, especially during cancer
patient monitoring. However, more work has to be done
concerning the number of clinical samples and the robustness of
those observations to strengthen their relevance for a diagnostic
test. Studies are ongoing in our laboratory to follow these potential
biomarkers along with KRAS/BRAF SNP detection in regards to
the personalized anti-EGFR therapy for metastatic CRC [31]. This investigation might also be crucial to better understand
ctDNA release mechanisms. In addition, this study should be also
carried out in other cancer types or in various physiological or
pathological conditions that result or lead to ample release of cell-
free DNA. Figure 7. Comparison of the DII values. Comparison of the DII
values (A) from genomic DNA, and ctDNA from mice plasma (non-
xenografted and xenografted) and (B) from human plasmas (healthy
and CRC). The DII was estimated by the ratio of the concentration
obtained by targeting a 300 bp sequence and a 60 bp sequence in a
KRAS region. doi:10 1371/journal pone 0023418 g007 Discussion This could explain the contradictory results
reported by the few previous studies on the clinical use of the
integrity index to follow cancer progression that did not allow
drawing general conclusions (i.e., estimation of ctDNA fragmen-
tation was found lower [22,23] equivalent [24,25] or higher
[13,21,26,27] than in controls). As stated by Jung et al [28] those
discrepancies might be related to pre-analytical and analytical
factors as well as possible bias in the selection of the study groups. We suggest that they may be explained at least partially by the
possibility that a large proportion of tumour ctDNA is ,100 bp,
an option which has never been taken into account. In addition, it
is conceivable that our observation might be restricted to the
clinical status of the patients whose blood was here examined
(stage III mCRC patients not undergoing chemotherapy). Effect of
chemotherapy and influence of CRC stage on ctDNA analysis is
under investigation in our laboratory. Our results on the tumour ctDNA size profile disagree with
many earlier reports. Most of these works described that ctDNA PLoS ONE | w PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 8 Fragmentation of Tumour-Derived Circulating DNA Figure 7. Comparison of the DII values. Comparison of the DII
values (A) from genomic DNA, and ctDNA from mice plasma (non-
xenografted and xenografted) and (B) from human plasmas (healthy
and CRC). The DII was estimated by the ratio of the concentration
obtained by targeting a 300 bp sequence and a 60 bp sequence in a Figure 7. Comparison of the DII values. Comparison of the DII
values (A) from genomic DNA, and ctDNA from mice plasma (non-
xenografted and xenografted) and (B) from human plasmas (healthy
and CRC). The DII was estimated by the ratio of the concentration
obtained by targeting a 300 bp sequence and a 60 bp sequence in a
KRAS region. doi:10.1371/journal.pone.0023418.g007 was comparable (2539+/2952 ctDNA fragments/ml, n = 9). Conversely,
the
ctDNA
amount
in
CRC
plasma
samples
(1,130,920;
298,760;
86,520
and
56,280
fragments/ml
as
determined by detecting a 60 bp amplicon) was much higher
than in control samples, irrespective of the amplicon length. There
is a 25-fold disparity when comparing the highest HHP ctDNA
value to the lowest CRC ctDNA concentration in our study. Supporting Information Table S1
Characteristics of the selected primers and corre-
sponding amplicons. (DOC) g
doi:10.1371/journal.pone.0023418.g007 consisted of fragments of approximatively 100 to 500 bp [29]. To
our knowledge, tumour-derived ctDNA lower than 80 bp was
previously never examined and quantified on the basis of their
higher proportion in ctDNA total population. Precise ctDNA size
profiling was reported only in a few works. Liu et al [30], by using
microfluidic single molecule spectroscopy, found a size profile for
ctDNA fragments .100 bp from stage I and IV lung cancer
patient sera that is quite similar to our results. However, they
concluded that the greatest discriminatory power occurred at a
threshold of 800 bp; no comparison was made with non-neoplastic
ctDNA. The authors never focused on ctDNA of size lower than
320 bp, arguing that below 320bp the two curves are similar. D. Lo’s team [23] analyzed the size distributions of maternal and fetal
DNA in maternal plasma by using a Q-PCR assay with primer sets
that amplify sequences from 105 to 798 bp. They suggested that
‘‘most of the ctDNA molecules were in the range of 145–201 bp’’
(which is approximately the nucleosome size). None of the
previous reports did examine or discuss the possibility of ctDNA
of size lower than 105 bp [23,24,26,29,30]. consisted of fragments of approximatively 100 to 500 bp [29]. To
our knowledge, tumour-derived ctDNA lower than 80 bp was
previously never examined and quantified on the basis of their
higher proportion in ctDNA total population. Precise ctDNA size
profiling was reported only in a few works. Liu et al [30], by using
microfluidic single molecule spectroscopy, found a size profile for
ctDNA fragments .100 bp from stage I and IV lung cancer
patient sera that is quite similar to our results. However, they
concluded that the greatest discriminatory power occurred at a
threshold of 800 bp; no comparison was made with non-neoplastic
ctDNA. The authors never focused on ctDNA of size lower than
320 bp, arguing that below 320bp the two curves are similar. D. Lo’s team [23] analyzed the size distributions of maternal and fetal
DNA in maternal plasma by using a Q-PCR assay with primer sets
that amplify sequences from 105 to 798 bp. They suggested that
‘‘most of the ctDNA molecules were in the range of 145–201 bp’’
(which is approximately the nucleosome size). Author Contributions Conceived and designed the experiments: F. Mouliere ART. Performed
the experiments: F. Mouliere BR EAP CG. Analyzed the data: F. Mouliere
BR EAP MDR MY F. Molina CG ART. Contributed reagents/materials/
analysis tools: BR MY. Wrote the paper: F. Mouliere CG ART. Conceived and designed the experiments: F. Mouliere ART. Performed
the experiments: F. Mouliere BR EAP CG. Analyzed the data: F. Mouliere
BR EAP MDR MY F. Molina CG ART. Contributed reagents/materials/
analysis tools: BR MY. Wrote the paper: F. Mouliere CG ART. 3. Gormally E, Hainaut P, Caboux E, Airoldi L, Autrup H, et al. (2004) Amount of
DNA in plasma and cancer risk: a prospective study. Int J Cancer 111: 746–749.
4. Fleischhacker M, Schmidt B (2007) Circulating nucleic acids (CNAs) and
cancer—a survey. Biochim Biophys Acta 1775: 181–232. Supporting Information None of the
previous reports did examine or discuss the possibility of ctDNA
of size lower than 105 bp [23,24,26,29,30]. Table S2
Values of control DII, non-tumoral DII and tumoral
DII in mouse plasma samples determined using the ctDNA
concentrations presented in Fig. 3. (DOC) Table S2
Values of control DII, non-tumoral DII and tumoral
DII in mouse plasma samples determined using the ctDNA
concentrations presented in Fig. 3. (DOC) p
p
p
y
J
4. Fleischhacker M, Schmidt B (2007) Circulating nucleic acids (CNAs) and
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and qualitative characterization of plasma DNA identifies primary and recurrent
colorectal cancer. Cancer Lett 263: 170–181. 20. Ellinger J, Bastian PJ, Haan KI, Heukamp LC, Buettner R, et al. (2008)
Noncancerous PTGS2 DNA fragments of apoptotic origin in sera of prostate
cancer patients qualify as diagnostic and prognostic indicators. Int J Cancer 122:
138–143. 6. Schwarzenbach H, Stoehlmacher J, Pantel K, Goekkurt E (2008) Detection and
monitoring of cell-free DNA in blood of patients with colorectal cancer. Ann N Y Acad Sci 1137: 190–196. 21. Ellinger J, Wittkamp V, Albers P, Perabo FG, Mueller SC, et al. (2009) Cell-free
circulating DNA: diagnostic value in patients with testicular germ cell cancer. J Urol 181: 363–371. 7. Flamini E, Mercatali L, Nanni O, Calistri D, Nunziatini R, et al. (2006) Free
DNA and carcinoembryonic antigen serum levels: an important combination for
diagnosis of colorectal cancer. Clin Cancer Res 12: 6985–6988. 22. Ellinger J, Albers P, Muller SC, von Ruecker A, Bastian PJ (2009) Circulating
mitochondrial DNA in the serum of patients with testicular germ cell cancer as a
novel noninvasive diagnostic biomarker. BJU Int 104: 48–52. 8. Guadalajara H, Dominguez-Berzosa C, Garcia-Arranz M, Herreros MD,
Pascual I, et al. (2008) The concentration of deoxyribonucleic acid in plasma
from 73 patients with colorectal cancer and apparent clinical correlations. Cancer Detect Prev 32: 39–44. novel noninvasive diagnostic biomarker. BJU Int 104: 48–52. 23. Chan KC, Zhang J, Hui AB, Wong N, Lau TK, et al. (2004) Size distributi
maternal and fetal DNA in maternal plasma. Clin Chem 50: 88–92. 23. Chan KC, Zhang J, Hui AB, Wong N, Lau TK, et al. (2004) Siz maternal and fetal DNA in maternal plasma. Clin Chem 50: 8 24. Mamon H, Hader C, Li J, Wang L, Kulke M, et al. (2008) Preferential
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plasma DNA in patients with lung cancer and nonmalignant lung disease. Fragmentation of Tumour-Derived Circulating DNA Ann N Y Acad Sci 1137: 207–213. 10. Holdenrieder S, Holubec L, Jr., Topolcan O, Finek J, Stieber P (2005)
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Prediction of breast tumor progression by integrity of free circulating DNA in
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Increased plasma DNA integrity in cancer patients. Cancer Res 63: 3966–3968. 28. Jung K, Fleischhacker M, Rabien A (2010) Cell-free DNA in the blood as a solid
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17. References PLoS ONE | www.plosone.org September 2011 | Volume 6 | Issue 9 | e23418 9 Fragmentation of Tumour-Derived Circulating DNA Gongora C, Candeil L, Vezzio N, Copois V, Denis V, et al. (2008) Altered
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https://digital.csic.es/bitstream/10261/117315/1/assising-intraspecific-variation-effective-dispersal-along-altitudinal-gradient-Garcia-Fernandez-A2014.pdf
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English
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Assessing Intraspecific Variation in Effective Dispersal Along an Altitudinal Gradient: A Test in Two Mediterranean High-Mountain Plants
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PloS one
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cc-by
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Abstract Background: Plant recruitment depends among other factors on environmental conditions and their variation at different
spatial scales. Characterizing dispersal in contrasting environments may thus be necessary to understand natural
intraspecific variation in the processes underlying recruitment. Silene ciliata and Armeria caespitosa are two representative
species of cryophilic pastures above the tree line in Mediterranean high mountains. No explicit estimations of dispersal
kernels have been made so far for these or other high-mountain plants. Such data could help to predict their dispersal and
recruitment patterns in a context of changing environments under ongoing global warming. Methods: We used an inverse modelling approach to analyse effective seed dispersal patterns in five populations of both
Silene ciliata and Armeria caespitosa along an altitudinal gradient in Sierra de Guadarrama (Madrid, Spain). We considered
four commonly employed two-dimensional seedling dispersal kernels exponential-power, 2Dt, WALD and log-normal. Key Results: No single kernel function provided the best fit across all populations, although estimated mean dispersal
distances were short (,1 m) in all cases. S. ciliata did not exhibit significant among-population variation in mean dispersal
distance, whereas significant differences in mean dispersal distance were found in A. caespitosa. Both S. ciliata and A. caespitosa exhibited among-population variation in the fecundity parameter and lacked significant variation in kernel shape. Conclusions: This study illustrates the complexity of intraspecific variation in the processes underlying recruitment, showing
that effective dispersal kernels can remain relatively invariant across populations within particular species, even if there are
strong variations in demographic structure and/or physical environment among populations, while the invariant dispersal
assumption may not hold for other species in the same environment. Our results call for a case-by-case analysis in a wider
range of plant taxa and environments to assess the prevalence and magnitude of intraspecific dispersal variation. Editor: Giovanni G. Vendramin, CNR, Italy Editor: Giovanni G. Vendramin, CNR, Italy Received May 15, 2013; Accepted December 21, 2013; Published January 29, 2014 Copyright: 2014 Lara-Romero et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by the Spanish Ministry of Science and Innovation (http://micinn.es): ISLAS-CGL2009-13190-C03-01 and LIMITES-CGL2009-
07229; the Spanish Ministry of Economy and Competitiveness (http://www.idi.mineco.gob.es): AdAptA-CGL2012-33528; and the Government of Autonomous
Region of Madrid (http://madrid.org): REMEDINAL2. Abstract CLR and AG were supported by a F.P.I. fellowship (Spanish Ministry of Science). AG is also supported by a
postdoctoral contract (CGL2010-22234-C02/BOS). JJRA was supported by CGL2009-09428 project from the Spanish Ministry of Economy and Competitiveness. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests ex * E-mail: carlos.lara.romero@gmail.com Assessing Intraspecific Variation in Effective Dispersal
Along an Altitudinal Gradient: A Test in Two
Mediterranean High-Mountain Plants Carlos Lara-Romero1*, Juan J. Robledo-Arnuncio2, Alfredo Garcı´a-Ferna´ndez1,3, Jose M. Iriondo1
1 Departamento de Biologı´a y Geologı´a, Universidad Rey Juan Carlos, Madrid, Spain, 2 Department for Ecology and Forest Genetics, INIA-CIFOR, Madrid, Spain, 3 Institut
Botanic de Barcelona, IBB-CSIC-IQUB, Barcelona, Spain Citation: Lara-Romero C, Robledo-Arnuncio JJ, Garcı´a-Ferna´ndez A, Iriondo JM (2014) Assessing Intraspecific Variation in Effective D
Gradient: A Test in Two Mediterranean High-Mountain Plants. PLoS ONE 9(1): e87189. doi:10.1371/journal.pone.0087189 January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org Study Site and Species It is self-compatible,
although autogamy is restricted by pronounced protandry [39]. In
Sierra de Guadarrama, genetic diversity is quite homogeneous
across S. ciliata populations, which show significant levels of
inbreeding [39]. High-mountain habitats have been identified as one of the most
fragile environments in the world, and global warming is thought
to be especially critical for plant populations in mountain systems
[23,24]. The movement of plants to higher elevations tracking
their climatic niche seems to be one of the main responses to
ongoing global warming [25,26], although phenotypic plasticity
and adaptation may also play an important role in their response
[27–29]. Although all these processes are significantly conditioned
by dispersal and recruitment patterns [4–6,10,27], no explicit
measurements of dispersal kernels have been made for high-
mountain plants so far. A better understanding of dispersal and
recruitment patterns of high-mountain plants through the study of
effective dispersal kernels would, therefore, provide greater insight
into high-mountain plant species response to climate warming. Armeria caespitosa (Go´mez Ortega) Boiss. in DC. is a high-
mountain dwarf chamaephytic cushion plant, endemic to the
mountains of central Spain (Sierra de Guadarrama, Ayllo´n and
East Gredos), which occurs at altitudes from 1600 to 2430 m. It
grows in the same dry cryophilic pastures dominated by Festuca
curvifolia, although isolated individuals have also been found on the
ledges of granite and gneiss rocks. Its flowers are grouped in short-
scaped flowerheads with 1968 flowers each. Each flower has a
single ovule, which yields one seed that remains enclosed in the
papyraceous calyx. Average seed mass is 1.18 mg and its diameter
ranges between 2.0 and 4.7 mm [30]. The papyraceous calyx may
facilitate seed dispersal by wind [30]. A. caespitosa is self-
incompatible [31] and pollinated by bees, bumblebees and syrphid
flies [31]. Previous studies on the population genetics of the species
revealed relatively low genetic differentiation and a complex
genetic structure among populations [40,41]. We used an inverse modelling approach to measure the net
reproductive rate and effective dispersal kernel parameters along
an altitudinal gradient in a threatened Mediterranean high-
mountain pasture community in central Spain. Analyses were
performed on five populations of each of two representative
species of the community: Silene ciliata Poiret and Armeria caespitosa
(Go´mez Ortega) Boiss. in DC. Study Site and Species The study was carried out in the orophyllous cryophilic pastures
of Sierra de Guadarrama, a mountain range located in central
Spain. Mean annual precipitation, measured at the Navacerrada
Pass (40u 469N, 4u 199W; 1860 m a.s.l.), is 1330 mm with a
pronounced dry season (,10% of total annual rainfall) from May
to October. Mean annual temperature is 6.3uC, with mean
monthly temperatures ranging from 21uC in January to 16uC in
July (www.aemet.es). Dry cryophilic pastures occur in the higher
summits above the tree line between 1900 and 2430 m.a.s.l and
are dominated by Festuca curvifolia Lag. ex Lange and other
perennial plants interspersed in a shrub matrix characterized by
Cytisus oromediterraneus Rivas Mart. et al. and Juniperus communis
subsp. alpina (Suter) Cˇ elak. Most seed dispersal research, employing inverse modelling or
other methods, has been carried out in single populations,
disregarding potentially relevant environmental variation across
space that could affect the natural processes underlying the
recruitment of particular species [10,19]; but see [5,20]. Hetero-
geneity in environmental conditions and landscape properties (e.g. intra and interspecific plant density, fragmentation, soil moisture,
wind conditions) may indeed influence the dispersal kernel even
over local geographic scales [10,17,19,21]. However, this variation
is implicitly ignored in spatially unreplicated studies, which may
lead to local results that are unrepresentative of the average
dispersal pattern of the species. Hence, more comparative
empirical studies are needed to test whether effective dispersal
kernels exhibit intraspecific variation across contrasting environ-
ments and to find environmental correlates of potential variation
[10,19,22]. Such studies could assess the validity of kernel-based
approaches that assume a sole kernel for each species [22]. Silene ciliata Poiret (Caryophyllaceae) is a chamaephytic cushion
perennial plant that occurs in the Mediterranean mountain ranges
of southern Europe [35]. One of its southernmost distribution
limits is found in Sierra de Guadarrama at altitudes from 1900 to
2430 m, where it grows in dry cryophilic pastures dominated by
Festuca curvifolia. It blooms in late summer, with a peak in early
August [36]. Flowering stems reach 15 cm in height and have 1–5
flowers. Fruit capsules have up to 100 seeds which are wind
dispersed in August-September. Silene ciliata seeds have an average
mass of 0.59 mg and their diameter ranges between 1.1 and
1.5 mm [37]. The species is essentially barochorous (seeds lack any
specific structure to promote dispersal) and is pollinated by syrphid
flies, bumblebees and moth species [38]. Study Site and Species Previous studies carried out on
these species showed that the altitudinal gradient is associated with
an environmental stress gradient, with the lowest population
experiencing the most stressful conditions, constraining seedling
establishment and reproductive performance [30–34]. We expect-
ed that variation in population structure and physical environment
along the altitudinal gradient could modify the spatial recruitment
pattern and dispersal distance of the species. Specifically, we
addressed two main questions: (1) What is the spatial range of
effective seed dispersal in these two high-mountain species? (2) Do
effective seed dispersal and fecundity parameters vary among
populations at different altitudes? Field Data In August and September 2010, we established 10610 m plots
in five populations of both S. ciliata and A. caespitosa distributed
along an altitudinal gradient at the study site (Table 1). For each
plot, we estimated a set of climate variables for the growing season
(April-September). We calculated mean, minimum and maximum
rainfall and temperature for each population using the Digital
Climatic Atlas of Spain of the Spanish Ministry of Environment
[42] (Table 1), and mean wind speed at 80 m height using the
Spanish Wind Energy Atlas of the Spanish Ministry of Industry
(http://atlaseolico.idae.es, Table 1). Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants estimated without identifying the exact source of each seed or
seedling [12,13,16–18]; reviewed in [19]. Introduction comprises the environmental factors that are needed for seedling
recruitment [7,10,11]. Dispersal has important implications for plant species. It
determines gene flow rates and the spatial distribution of genetic
diversity [1–3], and it influences processes like colonization [4],
range expansion [5], local adaptation [6] and recruitment patterns
[7]. It also affects metapopulation dynamics and species coexis-
tence and diversity [8,9]. The fecundity of reproductive adults and the distribution of
dispersal distances are fundamental characteristics of the dispersal
process which have been extensively used as basic descriptors of
this process [7,12–14]. However, in many systems, measuring seed
production and dispersal distances poses a challenge, because
dispersing seeds cannot be tracked easily, and seed or seedling
shadows of neighboring plants typically overlap, making it difficult
to identify mother plants [15]. Inverse modelling is a useful
methodology that allows the fecundity of adult plants and the
shape of the dispersal kernel (the probability density function of
propagule dispersal distances from an individual plant) to be Dispersal studies distinguish between primary and effective
dispersal. Primary dispersal is the initial movement of seeds to the
location where they are first deposited, whereas effective dispersal
is the complex result of primary dispersal and the post-dispersal
processes that take place after the seed is deposited on the soil
surface until it is a successfully established seedling [7,10]. Effective
dispersal is more relevant to population dynamics, because it January 2014 | Volume 9 | Issue 1 | e87189 January 2014 | Volume 9 | Issue 1 | e87189 1 PLOS ONE | www.plosone.org January 2014 | Volume 9 | Issue 1 | e87189 Ethics Statement 1a the smaller the seedling plot areas and
the longer the distance between maternal plants and seedling plots. We assumed a linear relationship between seedling production
and plant diameter, Qi = bbi, because previous studies have shown
that plant size is linearly related to seed production in both species
[31,33]. Furthermore, as found in previous studies [14,48],
alternative allometric functions worsened inverse modelling fits,
including those using the number of inflorescences (results not
shown). The dataset used for this study is available from the institutional
repository of Rey Juan Carlos University (BURJC-Digital, http://
eciencia.urjc.es/handle/10115/11835). We assumed a linear relationship between seedling production
and plant diameter, Qi = bbi, because previous studies have shown
that plant size is linearly related to seed production in both species
[31,33]. Furthermore, as found in previous studies [14,48],
alternative allometric functions worsened inverse modelling fits,
including those using the number of inflorescences (results not
shown). Ethics Statement All necessary permits were obtained for the described field
studies. Mr. Juan Antonio Vielva from the Administration Bureau
of the Natural Park of Cumbres, Circo y Lagunas de Pen˜alara and
Mr. Antonio Sanz from the Administration Bureau of the
Regional Park of Cuenca Alta del Manzanares gave their
permission to work in the protected natural areas. Field studies
did not involve any endangered or protected species. We mapped every S. ciliata and A. caespitosa reproductive plant
and seedling in each study plot using two high-resolution
Differential Global Navigation Satellite System (DGNSS) receivers
(Viva GS15, Leica, Switzerland) with an accuracy of 5 cm for x
and y coordinates. We also measured plant diameter and the PLOS ONE | www.plosone.org PLOS ONE | www.plosone.org 2 January 2014 | Volume 9 | Issue 1 | e87189 Effective Dispersal Patterns in Mountain Plants Table 1. Description of A. caespitosa and S. ciliata sampling sites. Population
UTM Coordinates (x,y) m
Altitude m Pm (min, max) mm Tm (min, max) 6C
Ws m/s
Seedling
Reproductive
Silene ciliata
Pen˜alara
419427.30
4522814.15
2405
82.50 (32, 149)
8.77 (0.2, 14.1)
10.62
469
1068
Cabezas de Hierro
421169.50
4516859.61
2305
85.17 (34, 151)
8.37 (0.2, 13.7)
10.87
243
1085
Nevero
428863.74
4537167.57
2190
71.00 (30, 121)
9.67 (2.1, 14.6)
8.97
19
245
Najarra
430196.59
4518886.29
2080
72.17 (27, 112)
10.45 (2.8, 15.7)
9
89
523
Laguna
419931.16
4521082.09
1946
77.00 (31, 134)
10.27 (2.8, 15.2)
7.17
6
147
Armeria caespitosa
Cabezas de Hierro
420950.54
4516685.32
2336
85.2 (34, 151)
8.37 (0.2, 13.7)
10.87
145
318
Najarra
430196.79
4518886.33
2080
72.2 (27, 112)
10.45 (2.8, 15.7)
9
498
356
Loma de Cabezas
420013.12
4514794.49
1970
77.0 (30, 131)
9.78 (2.5, 14.8)
8.45
53
132
Collado de las Vacas
419168.95
4513371.38
1882
66.3 (25, 110)
11.80 (4.0, 17.2)
8.46
49
169
Sierra de los Porrones
420580.85
4512251.64
1647
64.5 (26, 102)
12.88 (4.4, 18.3)
7.4
32
52
Geographical coordinates, altitude, climatic variables for the growing season (April-September) and seedling and reproductive plant density (individuals per 10610 m
plot) in study populations. Pm: monthly precipitation, Tm: monthly temperature, Ws: wind speed at 80 m height. Minimum and maximum monthly precipitation and
temperatures are provided in brackets. Ethics Statement Population
UTM Coordinates (x,y) m
Altitude m Pm (min, max) mm Tm (min, max) 6C
Ws m/s
Seedling
Reproductive UTM Coordinates (x,y) m
Altitude m Pm (min, max) mm Tm (min, max) 6C
Ws m/s
Seedling
Repro Altitude m Pm (min, max) mm Tm (min, max) 6C
Ws m/s
Seedling
Reproductive Geographical coordinates, altitude, climatic variables for the growing season (April-September) and seedling and reproductive plant density (individuals per 10610 m
plot) in study populations. Pm: monthly precipitation, Tm: monthly temperature, Ws: wind speed at 80 m height. Minimum and maximum monthly precipitation and
temperatures are provided in brackets. rij~
ffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
(xi{xj)2z(yi{yj)2
q
with mean d and shape parameter u. The integral in eqn. 1a provides the exact probability of dispersal
from plant i into the whole surface Aj of plot j. All the studies we
are aware of have assumed that the probability of dispersal from
any given plant to any of the points lying within area Aj of each
plot is constant, yielding the usual simplified expression: number of inflorescences of each reproductive plant. To account
for potential seedling sources outside the study area, we also
recorded the location, diameter and number of inflorescences of
reproductive plants in a buffer zone of two meters around the
plots. The seedling stage in S. ciliata and A. caespitosa includes up to
one-year individuals (i.e. plants that had germinated in fall or
spring and had survived their first full summer), because the
seedlings that survive a second growth period grow into the
reproductive stage [43,44]. Summer is critical to the survival of
Mediterranean plants due to the incidence of severe droughts
[45,46]. This is also the case in Mediterranean high-mountain
plants [47] such as S. ciliata and A. caespitosa [30,35]. Within this
framework, we included all critical stages of effective dispersal:
seed dispersal, seed germination and seedling survival. ^sj(b,rj; b,d,u)~
X
m
i~1
Q(bi; b)f (rij; d,u)Aj,
ð1bÞ ð1bÞ where rij is the spatial distance between plant i and the centre of
plot j. The approximation in eqn 1b will be closer to the exact
values predicted by eqn. 1a the smaller the seedling plot areas and
the longer the distance between maternal plants and seedling plots. where rij is the spatial distance between plant i and the centre of
plot j. The approximation in eqn 1b will be closer to the exact
values predicted by eqn. Statistical Analysis Large deviances mean that stand-specific d improves the likelihood
of the data so substantially that the null hypothesis of invariant d
across populations can be rejected [5]. We first tested for
significant differences in d across all populations of each species. When the overall test was positive, we tested for specific
populations with d-values significantly different from the global
species-average by subsequently conducting q deviance tests, one
for each population i (x2 with 1 degree of freedom), using: Large deviances mean that stand-specific d improves the likelihood
of the data so substantially that the null hypothesis of invariant d
across populations can be rejected [5]. We first tested for
significant differences in d across all populations of each species. When the overall test was positive, we tested for specific
populations with d-values significantly different from the global
species-average by subsequently conducting q deviance tests, one
for each population i (x2 with 1 degree of freedom), using: the WALD kernel [14,53] fWALD(rijDa,u)~
u
8p3r5
ij
!1=2
exp { u(rij{a)2
2a2rij
"
#
,
ð5Þ and the exponential-power kernel [5] and the exponential-power kernel [5] Di~{2 ln
Ld(SqDbq,d,uq,hq)
L(SqDbq,di,d{i,uq,hq)
"
#
,
ð10Þ ð10Þ fEXPOW(rijDa,u)~
u
2pa2C(2=u) exp {
rij
a
u
:
ð6Þ ð6Þ where the numerator is the same likelihood as before, assuming
that the mean dispersal distance d is constant across all
populations, while the likelihood in the denominator assumes that
the i-th population has a mean dispersal distance di, while the
remaining q21 populations have a constant mean dispersal
distance d2i. We applied the sequential Bonferroni correction to
correct for multiple testing [56]. Analogous tests were conducted
to investigate among-population variation in the shape parameter
of the dispersal kernel (assuming constant u and variable d in eqn 7)
and in the fecundity parameter b (assuming constant b and
variable d in eqn 7). Although the field survey at the study sites was
exhaustive, some sample sizes were unbalanced and relatively
small due to natural density variation (Table 1), which may
compromise the large numbers approximation under which the x2
distribution is expected. Small samples can indeed make likelihood
ratio (LR) tests relatively lax in the case of unequal male
reproductive success analysis [57]. Smouse et al. Statistical Analysis We used inverse modelling to estimate seedling dispersal kernel
parameters for each population of each species and to test for
dispersal parameter variation among populations within species. Following Ribbens et al. [13], the expected number of seedling
recruits sˆj at a given plot or trap j of area Aj (sˆj) equals the sum of
the seedling shadows across this plot of all m maternal plants, in the
form: )
For each population of each species, we divided the 100-m2
study area into c = 100L22 equal adjacent cells of area L2, and for
each cell j, we computed observed (sj) and predicted (sˆj) seedling
densities. We used equations 1a and 1b to estimate sˆj, as cell areas
were relatively large compared to some of the mother-cell pairwise
distances (occasionally with maternal plants lying within the target
cell), and it was not obvious a priori that the approximation in
equation 1b would be sufficiently accurate. However, virtually
identical dispersal and fecundity estimates were obtained for all
populations and assumed kernels using each equation (results not
shown). Thus, we only present the results for the latter, whose
computational efficiency enabled us to calculate confidence
intervals and conduct hypothesis tests (see below) in a reasonable
number of CPU hours. We also tested the effect of three different
values for cell side length, L = 0.125, 0.25 and 0.50 m, on
parameter estimates, with lower and upper values approximating
DGNSS accuracy and dispersal range, respectively. Distribution of
seedling densities across cells exhibited overdispersion relative to a ^sj(b,rj; b,d,u)~
X
m
i~1
Q(bi; b)
ð
Aj
f
ffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
(xi{xj)2z(yi{yj)2
q
; d,u
dxjdyj
ð1aÞ ð1aÞ where b and rj are, respectively, m-length vectors of individual
plant size measurements bi and spatial distances rij between the
spatial coordinates (xj, yj) of plant i, and the coordinates of plot j (xi,
yi), Q is an allometric function with parameter b yielding the
number of seedlings produced by a single plant, and f is a two-
dimensional isotropic seedling dispersal probability density func-
tion
(dispersal
kernel)
as
a
function
of
dispersal
distance January 2014 | Volume 9 | Issue 1 | e87189 January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org 3 Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants Poisson distribution (p,0.05) in all populations except the Nevero
population of S. Statistical Analysis ciliata, and we thus modelled seedling counts using
a negative binomial distribution [5]: Ld(SqDbq,d,uq,hq)~ P
q
k~1 L(SkDbk,d,uk,hk),
ð7Þ ð7Þ where bq, uq, and hq are q-length vectors of population-specific
fecundity parameters bk, kernel shape parameters uk and negative
binomial parameters hk. This model has 3q degrees of freedom and
provides an average species-level estimate of d. Note that, unlike
Clark et al. [5], we allowed for different values of the negative
binomial parameter (i.e. different degrees of spatial seedling
clumping) across populations. We assumed the same kernel family
for all populations in conducting this test, choosing the one
yielding the best average fit over all populations (i.e. the one with
the smallest average AIC across the population-level fits obtained
using eqn. 2). We then obtained parameter estimates assuming that
all parameters (including d) vary among populations within species
(still assuming a single consensual kernel family), based on the
likelihood L(SDb,d,u,h)~ P
c
j~1
C(sjzh)
C(sjz1)C(h)
^s
sj
j (b,rj; b,d,u)hh
½^sj(b,rj; b,d,u)sjzh ,
ð2Þ ð2Þ where S is the set of c observed seedling counts sj, C is the gamma
function, and h is the negative binomial parameter, determining
the distribution variance (values of h ,1 indicate overdispersed
distributions, while h ..1 tend to the Poisson distribution). Maximum-likelihood estimates for b, d, u and h were obtained for
each population of each species by maximizing eqn 2. Confidence
intervals were computed using the profile-likelihood method [49]. As most studies show that no single dispersal function provides
consistently superior fits across species [14,18,19,50], the perfor-
mance of alternative dispersal kernels should always be compared
[14,17]. We considered four commonly employed two-dimension-
al seedling dispersal kernels, with scale parameter a and shape
parameter u: the 2Dt kernel [12]; see [51] for the parameterization
used here. L(SqDbq,dq,uq,hq)~ P
q
k~1 L(SkDbk,dk,uk,hk),
ð8Þ ð8Þ where dq is the q-length vector of population-specific mean
dispersal distances dk. The model in eqn 8 has 4q21 degrees of
freedom. The deviance f2Dt(rijDa,u)~ u{1
pa2
1z
r2
ij
a2
!{u
,
ð3Þ ð3Þ D~{2 ln Ld(SqDbq,d,uq,hq)
L(SqDbq,dq,uq,hq)
"
#
ð9Þ the log-normal kernel [52] ð9Þ fLN(rijDa,u)~
1
(2p)3=2ur2
ij
exp { ( ln (rij){a)2
2u2
"
#
,
ð4Þ ð4Þ is asymptotically distributed as x2 with q21 degrees of freedom. is asymptotically distributed as x2 with q21 degrees of freedom. January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org Results We mapped a total of 4095 reproductive individuals and 1603
seedlings across all populations of the two species (Table 1; Figure
S1). Reproductive plant and seedling densities varied between
populations in both species (Table 1). We focused on the results
obtained with a 0.125 m cell side length (Tables 2 and 3), because
some of the S. ciliata populations did not converge when 0.25 or
0.5 m cell side lengths were used (Table S1). In any case, similar
results were obtained for both species for all cell side lengths
(Tables 2, 3, S1 and S2). As none of the four models converged for
A. caespitosa in the Najarra population, this population was
excluded from further analyses. The fits of the most parsimonious models ranged from good in
A. caespitosa populations (r2 = 0.17–0.35) to fair or poor (r2 = 0.01–
0.12) in S. ciliata populations (Tables 2 and 3) and were highly
significant (p,0.0001) in all cases except in the population with the
lowest seedling number (p = 0.32 for the Laguna population of S. ciliata, which only had 6 seedlings). Population estimates of mean
dispersal distance (d) and fecundity (b) parameters were generally
consistent when assuming different kernel families (Table 2 and 3). No significant correlation was found between log b and log d
(r = 0.35, p = 0.65). Differences in AIC values indicated that no single kernel
function provided the best fit across all populations of either of the
two species (Table 2 and 3). For S. ciliata, the WALD kernel was
among the most parsimonious models (i.e. models whose AIC
values differ DAIC ,2 from the model with the lowest AIC [55])
in all five populations, while the log-normal model was among the
most parsimonious in four populations, the exponential-power
model in three populations and the 2Dt model in two populations
(Table 2). Results did not support a particular kernel shape in S. ciliata: the mesokurtic, thin-tailed exponential-power with u .1 Focusing on the cross-population comparisons of the WALD
dispersal model for S. ciliata, estimated mean dispersal distances
ranged
between
d = 0.23
and
0.71 m
(Table
2),
although
differences
were
only
marginally
significant
(x2
4d.f. = 8.846,
p = 0.065). Estimated shape parameters ranged between u = 0.55 Table 2. Estimated parameters for the models fitted to the seedling recruitment data of S. ciliata. Table 2. Statistical Analysis [58], proposed
the use of nonparametric (permutational) tests as an alternative to
LR tests, showing that the former can perform better than LR tests
when parent-offspring
genealogical information is available. Mean seedling dispersal distances for each kernel are given by
d2Dt = aC(3/2) C(u23/2)/C(u21), dLN = exp (a + 0.5u2), dWALD = a
and dEXPOW = aC(3/u)/C(2/u), respectively. Given that d provides
a more intuitive description of dispersal range than a, independent
of the assumed kernel, we parameterized the model (eqns 1 and 2)
in terms of d and u, rather than in terms of a and u, facilitating the
estimation of profile-likelihood confidence intervals for d [5,54]
and tests of its variation across populations (see below). We used
the Akaike Information criterion (AIC) [55] to assess method
performance and calculated the goodness of fit of the most
parsimonious model by measuring Pearson’s product-moment
correlation between observed vs. predicted seedling densities. We tested for potential differences in dispersal and fecundity
parameters across populations using likelihood ratio statistics as in
Clark et al. [5]. Assuming that mean dispersal distance d is
constant across populations within species, we obtained parameter
estimates based on the likelihood of the whole species data set Sq,
incorporating information from all q populations of the species: January 2014 | Volume 9 | Issue 1 | e87189 January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org PLOS ONE | www.plosone.org 4 Effective Dispersal Patterns in Mountain Plants Similar non-parametric tests have not been developed so far in the
different statistical framework of inverse modeling, for which LR
tests, such as those used here, are still widely employed as feasible
approximations for exploratory assessment of variation in dispersal
and fecundity parameters [5,59–61]. All statistical analyses were
performed in the open source software package R [62]. showed similarly good fits (DAIC ,2) as leptokurtic functions with
either exponential (WALD) or fat (2Dt and log-normal) tails within
the same populations. Using the WALD kernel across all S. ciliata
populations, species-level estimates of mean dispersal distance and
the shape parameters were d = 0.39 m (95% CI: 0.35–0.45 m) and
u = 0.80 (0.63–1.13). For A. caespitosa, the log-normal kernel ranked
among the best models in all four populations where estimation
converged, whereas the WALD, 2Dt and exponential-power
models showed a similarly good performance in three populations
(Table 3). Supported models in A. Statistical Analysis caespitosa always included
leptokurtic functions with exponential or fat tails (including the
exponential-power with u,1). Using the log-normal model across
all A. caespitosa populations, the species-level estimates of kernel
parameters were d = 0.43 m (0.32–0.60 m) and u = 0.98 (0.87–
1.14). January 2014 | Volume 9 | Issue 1 | e87189 Results Estimated parameters for the models fitted to the seedling recruitment data of S. ciliata. Population
Kernel
Mean dispersal
(d) (m)
Shape parameter (u)
Fecundity parameter
(b) (seedlings/cm)
Negative binomial
parameter (h)
2 logL
DAIC
r2
Pen˜alara
Log-norm
0.32 (0.27–0.39)
0.64
(0.52–0.77)
0.009
(0.008–0.010)
0.359
(0.260–0.523) 1624.80
–
0.12
WALD
0.33
(0.28–0.41)
0.64
(0.44–0.99)
0.009
(0.008–0.010)
0.360
(0.260–0.525)
1624.82
0.04
2Dt
0.32
(0.27–0.46)
3.80
(1.99–na)
0.009
(0.009–0.011)
0.351
(0.255–0.509)
1627.77
5.94
Exp-pow
0.31
(0.27–0.37)
1.51
(0.97–2.34)
0.009
(0.008–0.010)
0.349
(0.253–0.507)
1628.76
7.92
Cabezas de Hierro Exp-pow
0.37 (0.32–0.45)
9.86
(2.34–na)
0.003
(0.003–0.004)
0.449
(0.235–1.198) 1029.04
–
0.05
Log-norm
0.43
(0.35–0.60)
0.43
(0.23–0.70)
0.003
(0.003–0.004)
0.460
(0.240–1.246)
1029.04
0.01
WALD
0.43
(0.35–0.62)
2.26
(0.84–8.70)
0.003
(0.003–0.004)
0.459
(0.239–1.244)
1029.17
0.27
2Dt
0.43
(0.33–0.58)
172.54
(3.65–na)
0.003
(0.003–0.004)
0.450
(0.235–1.205)
1031.17
4.25
Nevero
Exp-pow
0.23 (0.17–0.50)
2.84
(0.74–21.91) 0.001
(0.001–0.002)
142.062
(0.045–na)
108.04
–
0.08
2Dt
0.23
(0.17–0.42)
171.06
(1.74–na)
0.001
(0.001–0.002)
142.162
(0.082–na)
108.12
0.17
WALD
0.30
(0.22–0.60)
1.23
(0.41–3.27)
0.001
(0.001–0.002)
141.865
(0.054–na)
109.04
1.99
Log-norm
0.30
(0.21–0.86)
0.47
(0.29–1.35)
0.001
(0.001–0.002)
50.393
(0.053–na)
109.14
2.19
Najarra
WALD
0.71 (0.37–na)
0.55
(0.27–1.24)
0.002
(0.003–0.003)
0.280
(0.092–8.456) 461.85
–
0.04
Log-norm
0.62
(0.36–3.01)
0.86
(0.58–1.46)
0.002
(0.002–0.003)
0.280
(0.092–8.401)
462.14
0.59
Exp-pow
0.61
(0.36–2.45)
0.83
(0.27–2.49)
0.002
(0.002–0.003)
0.275
(0.091–7.395)
463.15
2.59
2Dt
nc
nc
nc
nc
nc
Laguna
Exp-pow
0.29 (0.17–0.40)
1411.75
(0.03-na)
0.001
(0.0003–0.004) 0.003
(0.001–0.035) 38.74
–
0.01
WALD
0.24
(0.14-na)
1.00
(na-na)
0.001
(0.0003–0.029)
0.004
(0.001–0.049)
38.91
0.34
Log-norm
0.24
(0.16-na)
0.47
(0.23–na)
0.001
(0.0003-na)
0.004
(0.001–0.048)
38.95
0.42
2Dt
0.23
(0.15–0.86)
165.18
(1.00–na)
0.001
(0.0003 - 0.006)
0.003
(0.001–0.035)
39.12
0.76
For each parameter, 95% confidence intervals are presented along with the mean value. Log-norm, log-normal; Exp-pow, exponential-power. na denotes that the
confidence interval limit is not available because of flat likelihood function. nc denotes models that did not converge.–logL, log-likelihood; AIC, Akaike’s information
criterion; r, Pearson correlation coefficient between observed and predicted seedling density values. Models with the lowest AIC are marked in bold for each population. doi:10.1371/journal.pone.0087189.t002 January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org 5 Effective Dispersal Patterns in Mountain Plants Table 3. Estimated parameters for the models fitted to the seedling recruitment data of A. caespitosa. Results Population
Kernel
Mean dispersal
(d) (m)
Shape parameter (u)
Fecundity parameter
(b) (seedlings/cm)
Negative binomial
parameter (h)
2 logL
DAIC
r2
Cabezas de
Hierro
Log-norm
0.33
(0.24–0.56)
0.97
(0.78–1.29)
0.024
(0.019–0.029)
0.122
(0.076–0.208)
584.41
–
0.17
2Dt
0.80
(0.30 - na)
1.60
(1.24–2.48)
0.024
(0.020–0.030)
0.122
(0.076–0.207)
584.78
0.73
WALD
0.35
(0.24–1.44)
0.21
(0.13–0.35)
0.024
(0.020–0.030)
0.121
(0.076–0.206)
584.80
0.79
Exp-pow
0.29
(0.23–0.45)
0.60
(0.31–1.03)
0.024
(0.020–0.029)
0.120
(0.075–0.202)
585.01
1.19
Loma de
Cabezas
Exp-pow
0.66
(0.46–1.39)
0.69
(0.24–2.37)
0.014
(0.010–0.019)
0.061
(0.028–0.162)
247.54
–
0.12
Log-norm
0.78
(0.48–2.57)
1.02
(0.69–1.64)
0.014
(0.010–0.019)
0.063
(0.028–0.169)
247.82
0.57
WALD
0.77
(0.46 - na)
0.44
(0.19–1.01)
0.014
(0.010–0.020)
0.064
(0.028–0.171)
247.96
0.85
2Dt
0.73
(0.05 - na)
2.15
(0.12 - na)
0.014
(0.010–0.020)
0.058
(0.026–0.158)
248.64
2.20
Collado de
las Vacas
2Dt
0.30
(0.18–na)
1.84
(1.36–2.85)
0.011
(0.008–0.014)
0.286
(0.107–1.330)
206.84
–
0.22
WALD
0.28
(0.19–0.58)
0.25
(0.13–0.44)
0.011
(0.008–0.014)
0.298
(0.109–1.444)
207.11
0.56
Log-norm
0.25
(0.17–0.43)
0.84
(0.63–1.18)
0.011
(0.008–0.014)
0.298
(0.109–1.442)
207.47
1.26
Exp-pow
0.27
(0.18–0.45)
0.56
(0.29–0.96)
0.011
(0.008–0.015)
0.249
(0.096–1.057)
209.86
6.05
Sierra de los
Porrones
Log-norm
0.23
(0.13–0.50)
1.07
(0.84–1.56)
0.021
(0.013–0.033)
0.369
(0.118–2.966)
110.01
–
0.35
2Dt
0.28
(0.16 - na)
1.75
(1.27–3.12)
0.021
(0.013–0.033)
0.371
(0.117–3.291)
110.02
0.01
Exp-pow
0.21
(0.13–0.40)
0.48
(0.19–0.93)
0.020
(0.013–0.033)
0.370
(0.118–3.227)
110.15
0.27
WALD
0.20
(0.10 - na)
0.07
(0.03–0.12)
0.024
(0.015–0.043)
0.280
(0.098–1.231)
111.27
2.52
For each parameter, 95% confidence intervals are presented along with the mean value. Log-norm, log-normal; Exp-pow, exponential-power. na denotes that the
confidence interval limit is not available because of flat likelihood function. – logL, log-likelihood; AIC, Akaike’s information criterion; r, Pearson correlation coefficient
between observed and predicted seedling density values. Models with the lowest AIC are marked in bold for each population. doi:10.1371/journal.pone.0087189.t003 For each parameter, 95% confidence intervals are presented along with the mean value. Log-norm, log-normal; Exp-pow, exponential-power. na denotes that the
confidence interval limit is not available because of flat likelihood function. – logL, log-likelihood; AIC, Akaike’s information criterion; r, Pearson correlation coefficient
between observed and predicted seedling density values. Models with the lowest AIC are marked in bold for each population. doi:10.1371/journal.pone.0087189.t003 Fecundity parameter estimates exhibited significant differences in
A. caespiotsa (x2
3d.f. = 22.51, p,0.001), ranging from b = 0.011 to
0.024 seedlings/cm. Results Per-population tests indicated that maximum
estimates of b (at Cabezas de Hierro) and minimum estimates of b
(at Collado de las Vacas) were significantly higher and lower than
the species average, respectively (Table 6). and 2.26 and were not significantly different from each other
(x2
4d.f. = 7.022, p = 0.135). Fecundity parameters ranged between
b = 0.001 and 0.009 seedlings/cm across S. ciliata populations and
were significantly different (x2
4d.f. = 254.26, p,0.001). Per-popu-
lation tests indicated that all population estimates of b were
significantly different from the species average (Table 4). Assuming a log-normal dispersal model for A. caespitosa,
population estimates of d varied between 0.23 and 0.78 m
(Table
3)
and
were
significantly
different
(x2
3d.f. = 14.022,
p = 0.003). Per-population tests showed that only the population
with the largest d (Loma de Cabezas with 0.78 m) differed
significantly from the species average (Table 5). Estimated shape
parameters ranged between u = 0.84 and 1.07 and were not
significantly different from each other (x2
3d.f. = 1.426, p = 0.699). Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants Table 5. Comparison of estimated mean dispersal distances
(d) among A. caespitosa populations. Population
di (m)
d-i (m)
-logL
x2
1d.f. (p-value)
Cabezas de
Hierro
0.33
0.49
1155.80
1.84 (0.1744)
Loma de
Cabezas
0.78
0.29
1150.46
12.53 (0.0003)*
Collado de las
Vacas
0.25
0.50
1154.47
4.50 (0.0338)
Sierra de los
Porrones
0.23
0.47
1155.20
3.04 (0.0809)
Average
0.43
–
1156.72
–
di is the mean dispersal distance for the corresponding population and d-i is the
average d across the three remaining populations. –logL is the log-likelihood of
the model considering di and separately d-i. The last column shows the
deviance (x2
1d.f. -distributed) of the model considering di and separately d-i
relative to the model (shown in the last row) assuming constant d across all four
populations. The asterisk denotes a significant test after sequential Bonferroni
correction. doi:10.1371/journal.pone.0087189.t005 Most effectively dispersed seeds were established near conspe-
cific adult plants (Figure S1). Consequently, estimated mean
dispersal distances were low, suggesting that both A. caespitosa and
S. ciliata have low effective seed dispersal ranges. Effective dispersal
scale is influenced by factors operating at the level of both primary
seed dispersal and germination and establishment. According to
mechanistic wind dispersal models based on the WALD kernel,
the seeds of six high-mountain species are transported no farther
than a few meters by primary wind dispersal, probably due to low
seed release height [65]. Release height, determined by inflores-
cence length, is indeed low (about 5–25 cm) in both A. caespitosa
and S. ciliata (see Castroviejo et al. [66] for morphological
characteristics). However, according to the mechanistic simula-
tions in Dullinger et al. [65], the primary seed dispersal range of
these two species should be longer than that estimated by our
effective dispersal models. This suggests that effective seed
dispersal is further limited by post-dispersal processes. Environ-
mental harshness in summer might be the main factor in reducing
primary seed dispersal distance. In the two species studied here,
recruitment is mainly limited at the seedling emergence and
survival stages by summer drought [30,34]. Furthermore, Garcı´a-
Camacho et al. [30] found a positive effect of A. caespitosa adult
cover on seedling emergence and survival in 0.560.5 m samplings
subplots. Effective Dispersal Patterns in Mountain Plants This suggests that adults act as nurse plants, facilitating
seedling emergence and survival along the altitudinal gradient
[30], as observed in other Mediterranean mountain regions [67]. The effect of adult cover on post-dispersal survival might lead to
unrealistic conclusions of short effective dispersal distance, because
our modelling approach assumes a distance-invariant survival
function. This assumption may be violated in the case of distance-
dependent survival because the primary and effective seed
dispersal kernel would attain different shapes [68]. Hence, the
modelling approach will implicitly assume that seedlings close to a
conspecific adult originate from this adult, even though facilitation
can also favour seeds from other adults after more substantial
dispersal [68]. Given the mean distance between the adult plants
recorded in our study plots (mean 6 SD: 5.3560.38 m for S. ciliata; 5.2160.58 m for A. caespitosa) and the essentially barochor-
ous seed dispersal mechanism, the ratio of seeds from other adults
to seeds from the closest adult is likely to be very small, and,
therefore, the effect of this bias negligible. Moreover, all the
studied S. ciliata populations exhibited significant positive fine-scale
spatial genetic structure (FSGS) at the 0–1 m distance class (Lara-
Romero et al. unpublished data), indicating that the spatial
aggregation of genotypes is consistent with the low effective
dispersal distance observed in our study. Mean dispersal distance provides a limited characterization of Table 5. Comparison of estimated mean dispersal distances
(d) among A. caespitosa populations. the source. No single kernel function provided the best fit across all
populations. Furthermore, fecundity parameters were significantly
different among populations in both species. Mean dispersal
distance was significantly different among A. caespitosa populations,
while differences in mean dispersal distance among S. ciliata
populations were only marginally significant. The kernel shape
parameter did not vary significantly in either of the two species. The inverse modelling approach proved to be useful in
characterizing effective seed dispersal patterns in both species. Parameter estimates were consistent among populations and
species independent of the assumed kernel. In addition, estimates
conformed to independent fecundity measures for the same species
[30,34], and to primary seed dispersal estimates obtained with
direct and indirect methods in congeneric species (Silene latifolia:
0.17–0.85 m, [63]; Armeria maritima: 0.6 m, [64]). Although r2
values were slightly lower than those obtained in analogous studies
[13,18,20], they were highly significant (except in the population
with the lowest seedling number). Discussion We used an inverse modelling approach to analyse the variation
in effective seed dispersal patterns of two coexisting species along
an altitudinal gradient in a high mountain pasture community of
central Spain. Estimated mean dispersal distances were short in
both species, as most seedlings established less than a meter from Table 4. Comparison of estimated fecundity parameters (b) among S. ciliata populations. Population
bi (seedlings/cm)
b-i (seedlings/cm)
2logL
x2
1d.f. (p-value)
Pen˜alara
0.009
0.003
23276.28
229.28 (,0.0001)*
Cabezas de Hierro
0.003
0.007
23365.25
51.35 (,0.0001)*
Nevero
0.001
0.006
23361.72
58.41 (,0.0001)*
Najarra
0.002
0.006
23366.10
49.64 (,0.0001)*
Laguna
0.001
0.005
23380.92
19.99 (,0.0001)*
Average
0.005
–
23390.92
–
bi is the fecundity parameter for the corresponding population and b-i is the average b across the remaining four populations. –logL is the log-likelihood of the model
considering bi and separately b-i. Last column shows the deviance (x2
1d.f. -distributed) of the model considering bi and separately b-i relative to the model (shown in the
last row) assuming constant b across all five populations. Asterisks denote a significant test after sequential Bonferroni correction. doi:10.1371/journal.pone.0087189.t004 Table 4. Comparison of estimated fecundity parameters (b) among S. ciliata populations. bi is the fecundity parameter for the corresponding population and b-i is the average b across the remaining four populations. –logL is the log-likelihood of the model
considering bi and separately b-i. Last column shows the deviance (x2
1d.f. -distributed) of the model considering bi and separately b-i relative to the model (shown in the
last row) assuming constant b across all five populations. Asterisks denote a significant test after sequential Bonferroni correction. doi:10.1371/journal.pone.0087189.t004 January 2014 | Volume 9 | Issue 1 | e87189 6 Effective Dispersal Patterns in Mountain Plants [20], variation in dispersal patterns within
species was found across tree stands of temperate forests that
spanned gradients in moisture and canopy openness. Similarly, we
expected higher effective dispersal ranges in high-altitude popu-
lations because less stressful conditions could reduce the intensity
of the adult nurse effect on the survival of emerged seedlings. Furthermore, the lower encroachment by subalpine shrubs at high
elevation sites [71] would provide a more open habitat with
greater exposure to wind that might favour longer-distance seed
dispersal. However, contrary to our expectations, differences in the
shape parameter value were not significant among populations in
either of the two species. Only the A. caespitosa population with the
largest mean dispersal distance differed significantly from the
species average value for this parameter. Despite the strong
variation in demographic structure and/or physical environment
along the altitudinal gradient, dispersal parameter estimates
tended to be consistent among populations within the species. This implies that the inherent seed dispersal traits of the species
(e.g., inflorescence length, fruit morphology, seed size and shape),
which are essentially the same in all populations, were the most
important factors in determining the effective dispersal kernel in
our study sites during the study period. Post-dispersal processes
controlling seedling emergence and establishment seem to operate
similarly across populations, regardless of evident variation in local
demographic and environmental conditions (intra and interspecific
plant density, local topography, soil moisture, temperature regime,
etc.). This provides new insight into the prevalence and magnitude
of intraspecific dispersal variation. Although our results are
consistent with previous studies [5,20], showing that effective
mean dispersal distances vary among populations of particular
species, they also indicate that effective dispersal kernels can
remain invariant across populations of other co-occurring species,
even if there is significant variation in demographic structure and
the environment. The estimates provided by empirical dispersal studies such as
this one could help to explicitly incorporate real migration
constraints in predictive species distribution models. This infor-
mation is all the more relevant because accurate predictions about
dispersal and migration capacities are considered to be among the
most significant uncertainties in projecting climate impacts on
plant species ranges [72]. Our results, together with previous seed
dispersal mechanistic simulations of alpine plant species [65] show
that the majority of dispersal events occur within a few meters
from the source. Effective Dispersal Patterns in Mountain Plants kernel such as the kurtosis and fatness of the distribution tail may
have important demographic, ecological and genetic consequences
[12,19,69,70]. Our results did not support a particular kernel
shape in the case of S. ciliata. Although the behavior near the origin
and the tail is not independent in the assumed phenomenological
kernels (see [16] and references therein), the virtual absence of
seedlings beyond the close proximity of S. ciliata adults (Figure S1)
may result in similarly good fits for kernels with very different tails
but with sufficiently fast probability decay near the origin. That is,
fat-tailed kernels could fit well in S. ciliata, because the actual
dispersal probability decreased fast near the origin and not
necessarily because the actual process had a fat tail. In A. caespitosa,
results supported leptokurtic functions with exponential or fat tails
with consistently more abundant isolated seedlings than for S. ciliata (Figure S1). This would suggest a larger proportion of long-
distance effective dispersal events in A. caespitosa than in S. ciliata. small number of individuals of each population; data not shown). Furthermore, seedling density was positively related to altitude
(t = 0.8, p,0.05 for both species). These results suggest different
seedling mortality rates at different altitudes, which have probably
influenced the observed differences in the fecundity parameter
among populations. These results are also congruent with previous
studies on the demography of the two species [30,33], indicating
that more benign conditions at the populations located at the
highest altitudes allow the emergence and establishment of larger
numbers of seedlings. Hence, the altitudinal gradient appears to
influence effective seed dispersal patterns in S. ciliata and A. caespitosa through variation in effective seedling establishment
probabilities, but not as much through variation in the effective
seed dispersal range. The explicit incorporation of finer-scale
environmental factors and landscape features (e.g. shrub cover) in
our models (for instance through the application of the movement
space concept; Schurr et al. [14]) might help to estimate and model
the potential effects of environmental variables on fine-scale spatial
patterns of effective dispersal more accurately, and thus increase
the amount of explained variance in within-population recruit-
ment patterns. p
p
The few studies that have previously used inverse modelling
with a multi-population approach found local variation in
dispersal parameter estimates within species [5,20]. In Clark et al. [5] and LePage et al. Effective Dispersal Patterns in Mountain Plants This is consistent with [73], who found
widespread post-glacial dispersal constraints on the current
distribution of plants in the European Alps. These findings raise
doubts about the capacity of high mountain plants to track their
climatic niche under the rapid climate warming predicted for
mountain systems during the 21st century [23]. However, spatial
spread and colonization rates are not necessarily governed by
mean seed dispersal distance but by the frequency of rare long-
distance dispersal (LDD) events [11,74,75]. LDD is usually caused
by extreme events in terms of horizontal wind speed or turbulence
[21,74]. High-mountain habitats are exposed to frequent and
strong updrafts [76,77]. Consequently, LDD may occur frequently
in high-mountain environments [78]. These events could greatly
increase the chance of threatened populations to track the
altitudinal-zone displacement induced by warming. Therefore,
further research should accurately estimate the impact of LDD on
seed dispersal patterns in mountain ecosystems using adequate
methods
(for
instance
through
the
application
of
recently
developed genetic methods [79–81]). Effective Dispersal Patterns in Mountain Plants This suggests that inverse
modelling was able to provide information on the scale of dispersal
in the studied high mountain plants, even though there seemed to
be substantial distance-independent fluctuations in dispersal and
establishment probabilities that cannot be characterized by simple
kernel fits as frequently found in heterogeneous environments
[19]. Mean dispersal distance provides a limited characterization of
dispersal range [19,69]. Other characteristics of the dispersal Table 6. Comparison of estimated fecundity parameters (b) among A. caespitosa populations. Table 6. Comparison of estimated fecundity parameters (b) among A. caespitosa populations. Population
bi (seedlings/cm)
b-i (seedlings/cm)
-logL
x2
1d.f. (p-value)
Cabezas de Hierro
0.024
0.013
1152.64
16.65 (,0.0001)*
Loma de Cabezas
0.014
0.020
1159.56
2.82 (0.0932)
Collado de las Vacas
0.011
0.021
1153.55
14.84 (0.0001)*
Sierra de los Porrones
0.021
0.017
1160.66
0.61 (0.4337)
Average
0.017
–
1160.97
–
bi is the fecundity parameter for the corresponding population and b-i is the average b across the three remaining populations. –logL is the log-likelihood of the model
considering bi and separately b-i. The last column shows the deviance (x2
1d.f. -distributed) of the model considering bi and separately b-i relative to the model (shown in
the last row) assuming constant b across all four populations. Asterisks denote significant tests after sequential Bonferroni correction. doi:10.1371/journal.pone.0087189.t006 n of estimated fecundity parameters (b) among A. caespitosa populations. bi is the fecundity parameter for the corresponding population and b-i is the average b across the three remaining populations. –logL is the log-likelihood of the model
considering bi and separately b-i. The last column shows the deviance (x2
1d.f. -distributed) of the model considering bi and separately b-i relative to the model (shown in
the last row) assuming constant b across all four populations. Asterisks denote significant tests after sequential Bonferroni correction. doi:10 1371/journal pone 0087189 t006 January 2014 | Volume 9 | Issue 1 | e87189 7 Effective Dispersal Patterns in Mountain Plants PLOS ONE | www.plosone.org References Levine JM, Murrell DJ (2003) The community-level consequences of seed
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mountain island. Plant Biol 11: 515–524. 9. Supporting Information eter; – logL, log-likelihood; nc denotes models that did not
converge. eter; – logL, log-likelihood; nc denotes models that did not
converge. Figure S1
Spatial distribution of adults (circles) and seedlings
(crosses) in each study plot of A. caespitosa and S. ciliata. (PDF) Author Contributions Table S2
Estimated parameters for the four models fitted to the
seedling recruitment data of Armeria caespitosa. (cell size length 0.25
and 0.50 m). d, mean dispersal distance (m); u shape parameter; b,
fecundity parameter (seedlings/cm); h, negative binomial param- Conceived and designed the experiments: CLR JJRA JMI. Performed the
experiments: CLR AGF. Analyzed the data: JJRA CLR. Wrote the paper:
CLR JJRA JMI AGF. Conceived and designed the experiments: CLR JJRA JMI. Performed the
experiments: CLR AGF. Analyzed the data: JJRA CLR. Wrote the paper:
CLR JJRA JMI AGF. Conceived and designed the experiments: CLR JJRA JMI. Performed the
experiments: CLR AGF. Analyzed the data: JJRA CLR. Wrote the paper:
CLR JJRA JMI AGF. Acknowledgments Table S1
Estimated parameters for the models fitted to the
seedling recruitment data of Silene ciliata (cell size length 0.25 and
0.50 m). d, mean dispersal distance (m); u shape parameter; b,
fecundity parameter (seedlings/cm); h, negative binomial param-
eter; – logL, log-likelihood; nc denotes models that did not
converge. The authors especially thank Dr. Marı´a Jose Albert for her help on the
design of the study and field work. They also thank two anonymous
reviewers for their valuable comments on the draft version of the
manuscript, Gema Escribano-Avila who helped in the fieldwork and made
comments on the manuscript and Lori De Hond for linguistic assistance. Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants 44. Gimenez-Benavides L, Albert MJ, Iriondo JM, Escudero A (2011) Demographic
processes of upward range contraction in a long-lived Mediterranean high
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Environ Chang 17: 420–428. January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org January 2014 | Volume 9 | Issue 1 | e87189 9 Effective Dispersal Patterns in Mountain Plants Effective Dispersal Patterns in Mountain Plants Can J Forest Res 35: 978–989. 77. Whiteman DC (2000) Terrain-Forces Flow. Mountain Meteorology: Funda-
mentals and Applications. New York: Oxford University Press. 141–170. 78. Tackenberg O, Stocklin J (2008) Wind dispersal of alpine plant species: A
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62. R Core Team (2013) R: A language and environment for s Vienna, Austria: R Foundation for Statistical Computing. 81. Robledo-Arnuncio JJ, Garcı´a C (2007) Estimation of the seed dispersal kernel
from exact identification of source plants. Mol Ecol 16: 5098–5109. 63. Jongejans E, Schippers P (1999) Modeling seed dispersal by wind in herbaceous
species. Oikos 87: 362–372. January 2014 | Volume 9 | Issue 1 | e87189 PLOS ONE | www.plosone.org 10
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German
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Replicatio rei secundum actorem iudicatae
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Zeitschrift der Savigny-Stiftung für Rechtsgeschichte. Romanistische Abteilung
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327 327 Miszellen. [Replicati«) rei secundum actorem indicatae.J In zwei Digesten-
fragmenten wird mittelst einer replicatio nicht das Dasein, sondern
der I n h a l t eines Urteils geltend gemacht. Diese Stellen sind 20,11.16 § 5 : Creditor hypothecam sibi per sententiam adiudica-
tam quemadmodam habituras sit, quaeritur: nam dominium eius
vindicare non potest, sed hypothecaria agere potest, et si exceptio
obicietur a possessore rei indicatae, repli cet: si secundum me iudi-
catum non est, 20,11.16 § 5 : Creditor hypothecam sibi per sententiam adiudica-
tam quemadmodam habituras sit, quaeritur: nam dominium eius
vindicare non potest, sed hypothecaria agere potest, et si exceptio
obicietur a possessore rei indicatae, repli cet: si secundum me iudi-
catum non est, und 44, 2 1. 9 § 1: Si quis fundum, quem putabat se possidere, defen-
derit, mox emerit, re secundum petitorem iudicata an reetituere
cogatur? et ait Neratius, si actori iterum petenti obiciatur exceptio
rei iudicatae, replicare eum oportere de re secundum se iudicata. Es ist nicht zu bestreiten, daß hier vom Kläger der Inhalt des
Urteils geltend gemacht wird, und ich habe dies auch anerkannt (diese
Ztschr. 21 S. 51), zugleich aber geltend gemacht, daß man hiervon auf
eine exceptio rei iudicatae zur Geltendmachung des Urteilsinhalts
nicht schließen könne. Es wäre auch handgreiflich falsch, die in den
beiden allegierten Stellen angeführte replicatio als replicatio rei iudi-
catae zu bezeichnen; denn diese müßte ja lauten: aut si ea res iudi-
cata est, was sich im Anschluß an eine exceptio: si ea res iudicata
non est merkwürdig genug ausnehmen würde! Die beiden Fragmente stellen uns aber vor folgende Frage : warum
berücksichtigt hier der iudex den Inhalt des zwischen den Parteien
früher ergangenen Urteils nicht schon kraft seines ihn auf das ius civile
verweisenden officium, ohne Anweisung in der formula? g
Die Antwort lautet: er tut dies deshalb nicht, weil er es nicht
tun kann, und er kann es nicht aus demselben Grunde, ans dem er
bei der Formel der actio Publiciana auf die Behauptung des Beklagten,
er sei ziviler Eigentümer, keine Rücksicht nehmen kann, hierzu viel-
mehr einer besonderen Ermächtigung durch inserierte exceptio iusti
dominii bedarf (vgl. materielle Grundlage der exceptio S. 103f.). Der
iudex ist in erster Linie an die in der Formel enthaltene Weisung des
Prätors gebunden. In den beiden uns beschäftigenden Stellen ist durch
die Formel gegeben, daß der iudex absolvieren muß, si ea res iudicata
est. 327 Das ist aber ebenso der Fall, wenn secundum actorem, als wenn
contra actorem erkannt ist. Es wird also durch die vom iudex in
erster Linie zu beachtende exc. rei iudicatae demselben die Möglichkeit,
den Inhalt des Urteils als ius inter partes zu berücksichtigen und zur
Geltung zu bringen, genommen, und erst durch die replicatio wird ihm
diese Möglichkeit eröffnet. g
Mit dem Gesagten haben wir aber nicht nur die replicatio rei
iudicatae erklärt und dargelegt, daß sie mit unserer Annahme betr. die ipso-iure-Wirkung der materiellen Rechtskraft nicht im Widerspruch
steht, sondern wir können nun gerade auf Grund der beiden Stellen,
die als die festesten Stützen der Theorie von der positiven Funktion
der exc. rei iudicatae gegolten haben, die Frage stellen: gibt es einen 328 Miszellen. stärkeren Beweis gegen die positive Funktion dieser exceptio, als
unsere beiden Stellen, die dartnn, daß die exc. rei iudicatae es dem
iudex geradezu unmöglich machen kann, den Inhalt des Urteils zur
Geltung zu bringen? Auch nach einer anderen Richtung hin läßt sich diese replicatio
rei secundum actorem iudicatae verwerten. Sie beweist, daß die Be-
achtung eines früheren inter partes ergangenen Urteils seitens des
iudex nicht etwa in dem Sinn im Belieben desselben steht, wie ein
Unterrichter eine Entscheidung eines höchsten Gerichts als „Präjudiz"
in einem ähnlichen Fall für sich maßgebend sein lassen kann, oder
nicht. Denn es ist keine Frage, daß der iudex der replicatio rei
secundum actorem iudicatae parieren muß, auch wenn er das dem
Kläger günstige frühere Urteil für unrichtig hält. Wie sollte denn
der Prätor dazu kommen, den iudex in dieser Weise zu fesseln, wenn
der Regel nach res iudicata inter partes für den iudex ohne verbind-
liche Kraft wäre? Fr. Ei s el e. Freiburg i. Br. [Zum Eigentunisttbergang beim Kauf.] Die Redaktion der Sav.-Z. gestattet mir, einen kurzen Vorbericht über einen Teil meiner demnächst
erscheinenden Abhandlung: Der Kauf mit fremdem Geld (Studien zum
griechischen und römischen Recht) zu geben. — Die Abhängigkeit des
Eigentumsüberganges von der Preiszahlung oder deren Surrogaten ist
auf Grund griechischer Rechtsanschauung von den Byzantinern her-
gestellt. Das klassische römische Recht zeigt Gaius Inst. 2, 18—20,
das justinianische Just. Inst. 2,1,41. In einigen diokletianischen Konsti-
tutionen wird der Kampf der griechischen und der römischen Auf-
fassung sichtbar (C. 3,32,12 ; 4, 38,8 ; 4,38, 9 ; 4, 38,12 ; 4, 49, 6 ; 3, 32, 27 ;
vgl. auch C. 4, 49, 1 ; 4, 54, 3). 327 Die Digestenfragmente, die den Satz der
Institutionen Justinians zu bestätigen scheinen, sind teils interpoliert
(D. 7, 1, 12, 5; 7, 1, 25, 1; 14, 4, 5, 18; 18, 1, 19; 18, 1, 53; 40, 12, 38, 2),
teils ohne Beweiskraft (D.19,1,11, 2; 47, 2,14, 1; 49, 14, 5,1). Jherings
Annahme, daß der Satz, den Just. Institutionen wiedergeben, für die
Manzipation durch einen Zusatz zu deren Formular überwunden worden,
fur die Tradition aber in Geltung geblieben sei (Geist des röm. Rechts
5. Aufl., 2, 541 ff.), ist in ihrem zweiten Teile unglaubhaft. Keine klassische
Stelle, die von Tradition handelt, kennt das Erfordernis (Gai Inst. 2, 19-20; D. 6,1, 41,1; 6,1, 50; 41,1,13; 41, 1,31 pr.; Just. Inst. 2,1,43;
2, 23, 3»), Von den erwähnten Konstitutionen, die das Erfordernis ab-
lehnen, handelt keine mit Sicherheit von Manzipation, C. 4, 38, 8 und
4, 38, 9 sicher, C. 4, 38, 12; 3, 32, 12 und 3, 32, 27 vielleicht, C. 4, 49,1
und 4,54, 3 wahrscheinlich von Tradition, D. 19,1,11,2 (si modo — satis-
factum T.); 14, 4, 5, 18; 18,1,19; 18,1, 53; 40,12, 38, 2 zeigen, daß die
Byzantiner nicht nur dort die Preiszahlung einfügten, wo sie die
Manzipation durch die Tradition ersetzten (so schon Mitteis, Röm.
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English
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Development of a point-of-care-device for fast detection of periodontal pathogens
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BMC oral health
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cc-by
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© 2015 Gaertig et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Abstract Background: A number of pathogens can cause severe destruction of the periodontal apparatus during the course
of periodontitis. The aim of this work was the development of a diagnostic device for the use at the point-of-need
for the detection of periodontal pathogens to enable a personalized therapy for treatment of periodontitis. Methods: This test system is based on the polymerase chain reaction of DNA isolated from periodontal pathogens
and was examined to precisely detect species-specific sequences on a rotating chip with lyophilized reagents for
polymerase chain reaction. The preservation of the reagents was optimized to ensure their stability during the
storage. Results: In the current work, we have developed a model point-of-care device and showed a proof of concept. It
requires low sample volume, is timesaving and can therefore facilitate early diagnosis and treatment of periodontal
diseases. Conclusions: The developed device can provide fast diagnosis of the composition and amount of patients’ oral
flora and might help to assess the stage of periodontitis infection. This can facilitate an optimization of therapeutic
approaches in order to prevent some of the more serious consequences of the disease. Keywords: Bacterial quantification, Periodontitis, Real-time PCR, Microbial diagnostics, Lyophilization For the diagnosis of periodontitis, several practices
have been established. Direct periodontal examinations
include patients’ anamnesis, measurements of probing
pocket depth and clinical attachment level, and taking
dental radiographs. Additionally, genetic susceptibility
tests or examination of the subgingival microflora and
gingival crevicular fluid are optional methods to evaluate
periodontal disease [5]. Depending on the results of the
medical diagnosis and stage of the disease, an appropri-
ate therapy follows. Approaches such as the optimization
of oral hygiene, regular professional cleaning, scaling/de-
bridement and root planing, the use of antibiotics for
controlling bacterial plaque formation and inflammation,
and even in extreme cases surgery can be necessary to
prevent tooth loss [6]. * Correspondence: cornelia.gaertig@izi.fraunhofer.de
1Nanotechnology Unit, Department of Diagnostics, Fraunhofer Institute for
Cell Therapy and Immunology, Perlickstraße 1, 04103 Leipzig, Germany
2Friedrich Schiller University, Jena, Germany
Full list of author information is available at the end of the article Development of a point-of-care-device for
fast detection of periodontal pathogens Cornelia Gaertig1,2*, Katja Niemann1, Jana Berthold1, Lisa Giel1, Nadine Leitschuh1, Christoph Boehm1,
Liudmila Roussak1, Katja Vetter3 and Dirk Kuhlmeier1 Gaertig et al. BMC Oral Health (2015) 15:165
DOI 10.1186/s12903-015-0155-y Gaertig et al. BMC Oral Health (2015) 15:165
DOI 10.1186/s12903-015-0155-y Background Approximately 90 % of the world’s population experi-
ence dental or oral problems during their lifetimes
[1]. 11.2 % of adults are affected by severe periodon-
titis, thus making it a pertinent topic in the dental
health care sector [2]. Periodontitis is a complex inflammatory response of
the human body against microorganisms, which can lead
to severe tissue destruction in the oral cavity [3]. Risk factors leading to infections have been identified
as smoking, diabetes, occurrence of special microorgan-
isms, obesity, psychological and hygienic aspects. Fur-
thermore, genetic factors, age, gender and ethnicity
might also play a role in the development of periodontal
disease [4]. Of the approximately 600–700 bacterial species that
are living in subgingival areas, only a few of them are as-
sociated with periodontitis and can lead to significant
changes in oral microbial community, quantity and
composition [7, 8]. In order to assess the stage of Gaertig et al. BMC Oral Health (2015) 15:165 Page 2 of 8 Page 2 of 8 Page 2 of 8 rising amplicon amount can be detected via fluorescence
measurements. periodontitis infections and initiate effective treatments,
it is helpful to determine both the identities of the inter-
acting species as well as the overall number of patho-
gens
living
in
periodontal
pockets. The
work
presented here focuses on bacteria species shown to
be associated with periodontitis, which can be delin-
eated into three classes based on pathogenicity: E. corrodens, F. nucleatum, P. micra are slightly patho-
genic, P. intermedia, C. rectus, E. nodatum are mod-
erately pathogenic and A. actinomycetemcommitans,
P. gingivalis, T. denticola strongly pathogenic organ-
isms [9]. Despite the fact that further species, the ex-
pression
of
typical
biomarkers
and
the
individual
lifestyle of people can influence the development of
disease as well, the detection of these marker species
can be a first step in the process of identifying a dis-
ease or the medical condition in more detail. Conceptually similar rotation devices exist, such as
the Focus 3M™Integrated Cycler, which uses the
combination of centrifugal force and infrared energy
as heating source for the PCR. Sample analysis is re-
alized either in a disposable universal disc for user as-
says or for particular diseases in a chip with specific
kits requiring many pipetting steps or cooling of re-
agents [13]. Also Sun et al. Background [14] showed 2007 a circu-
lar microchip for amplification applying ferrofluids
and magnetic forces, revealing the problem of low
sample throughput. The lab-on-a-chip based amplification and measure-
ment system presented in this work has the potential to
optimize conventional periodontitis diagnostics in the
future by minimizing laboratory handling steps and en-
able fast realization of PCR, avoiding a long shipping
time of patient samples. Furthermore, the chip-based
setup can be integrated with modules for DNA isolation
in later versions. In this way, we aim to eliminate time-
consuming and labor-intensive steps during sample
preparation. If the types of bacterial species living in periodontal
pockets can be identified, this information can be used
to better classify the nature and severity of infection and
tailor a personalized therapy best suited to reversing its
course [10]. Only a few point-of care devices for nucleic acid based
detection have been developed and established on the
market so far. Cepheid’s GeneXpert, for example, is an
automated device for polymerase chain reaction (PCR)
with special sample preparation cartridges for tubercu-
losis pathogens, MRSA or group B streptococci. How-
ever, this high-cost system does not enable the option to
implement several multiplexed reactions in one test run. IQuum recently presented the PCR-driven analysis of
HIV in 1.5 h using the LIAT analyzer and Lutz et al. demonstrated a lab-on-a-foil system realizing an amplifi-
cation reaction for MRSA in less than 20 minutes [11]. Furthermore, Van Oordt et al. [12] reported in 2012
about an integrated system for nucleic acid detection or
immunoassays with the help of a disposable Labdisk
platform. Methods Bacterial strains/ cultivation conditions/ DNA isolation
Periodontitis-causing bacteria were obtained from the
German collection of Microorganisms and Cell Cultures
(DSMZ, Braunschweig, Germany) and grown in appro-
priate medium (see Additional file 1: Table S1). Colonies
or bacteria pelleted from liquid media by centrifugation
could be picked for DNA isolation. DNA purification
was accomplished with the help of the QIAamp DNA
Mini Kit® (Qiagen, Hilden, Germany) according to the
manufacturer’s instructions. DNA yield and purity were
measured with the NanoDrop instrument 1000 (Thermo
Scientific, Wilmington, USA). In the present study, the development of a simple to
use, low-cost lab-on-chip system for the detection, iden-
tification and quantification of periodontal bacteria is
the primary consideration in the design of the molecular
biological assay and diagnostic device. The diagnostic
system consists of two major elements: a microfluidic
cartridge, which contains all necessary PCR reagents in a
dehydrated format, and a hardware device responsible
for driving and controlling the work-flow. The latter is a
radially-divided heating block assembly containing mul-
tiple stable temperature zones for the different steps of
the PCR, combined with a centrally located rotary motor
containing a mounting for a polycarbonate (PC) PCR
chip. This chip harbors the patient sample in reaction
chambers and rotates through the temperature zones on
the heating block in order to carry out the PCR. The PCR The detection and quantification of periodontal bacteria
amplification were carried out using a Lightcycler 480 II. Primer sequences are given in Additional file 2: Table
S2. The primers for amplification of the species-specific
DNA fragment were designed during this project with
the help of the Primer3 software. Universal 16S primers
were adopted from Kommedal et al. [15]. Cycling conditions for the amplification were as fol-
lows: 5 minutes of initial incubation at 95 °C, followed
by 40 cycles of amplification (95 °C for 10 s, 51 °C for 10
s and 72 °C for 15 s). After each PCR run, an additional
melting curve analysis was carried out to characterize
the produced DNA molecules. Gaertig et al. BMC Oral Health (2015) 15:165 Page 3 of 8 Southern blot To determine the 16S copy number of E. corrodens via
Southern Blot, approximately 10 μg DNA were digested
with restriction enzymes EcoRI HF, HindIII (New
England Biolabs, Ipswich, USA), NcoI, PstI, XbaI (Jena
Bioscience
GmbH,
Jena,
Germany)
and
XhoI
FD
(Thermo Fisher Scientific, Waltham, USA) at 37 °C
overnight. The samples, along with the 16S PCR
amplicon as positive control, were separated on a 1 %
agarose gel at 4 °C. The gel was prepared for capillary
blot with depurination (0.25 M HCl), denaturation
(1.5 M NaCl, 0.5 M NaOH) and neutralization (3 M
NaCl, 0.5 M Tris-HCl), followed by overnight blotting
onto a nylon membrane. The reaction mix itself was optimized with several ad-
ditives as described in the following ‘lyophilization sec-
tion’ in order to increase the PCR-compatibility of PC
and to enable lyophilization of the mix. PCR-on-a-chip The PCR was implemented on 1 mm thick PC chips
with cavities that hold a volume of 10 μl of reaction mix. The chips were sealed with sealing foils (nerbe plus
GmbH, Winsen/Luhe, Germany) from both sides with
the PCR mix embedded. For amplification, the PCR chip rotates on the thermo-
cycling device, which is shown in Fig. 1. Briefly, it is
composed of six circularly arranged pie-slice-shaped
heating blocks, three of them for accomplishing denatur-
ation, annealing and elongation steps and three add-
itional blocks for achieving rapid temperature changes
inside the cavities. For
proof
of
concept
of
the
apparatus,
a
well-
established PCR of S. aureus (ATCC 2592), with an
amplicon size of 279 bp was used. Cycling conditions
were an initial 5 min denaturation, followed by 30 s at
each of the sequential 95, 61 and 72 °C steps for amplifi-
cation for a total of 40 cycles, with 5 s spent on the
plates located between each of the three. Standard curves amplicon to the control DNA in an agarose gel was car-
ried out. For establishing standard curves, dilution series of bac-
terial DNA ranging from 2 to 2 million genome equiva-
lents were used in a four-fold attempt for quantitative
real-time PCR. Standard curves were created by default
with
the
fit
points
method
of
the
Lightcycler
programme. For the overall bacterial count, Porphyromo-
nas gingivalis served as a representative for 16S PCR
with an average copy number of four 16S gene copies. The standard was set up to achieve an estimated patho-
gen number for quantification. It is known that several
copies of the 16S gene can occur within one prokaryotic
genome, with a maximum of 15 copies for Bacillus thur-
ingiensis. This fact makes it difficult to estimate a realis-
tic overall bacterial count in a diverse population of
bacteria with strongly varying copy numbers between
the individual species. The periodontitis-causing organ-
isms used in our investigation are believed to be in the
lower copy number range according to information
given in the Ribosomal RNA Operon Copy Number
Database [16]. To prove this, the unknown copy number
for E. corrodens was determined during the course of
this work. For unsequenced organisms, copy number de-
termination can be achieved with the help of Southern
Blot analysis. A 16S probe can detect the number of
equivalent regions in the restricted genome, as previ-
ously shown by Lee et al. [17]. After pre-hybridization overnight at room temperature
followed by a 2-hour incubation at 42 °C, hybridization
with
200
ng
probe/ml
buffer
was
carried
out. Hybridization and detection procedures were performed
as described in the Biotin Chromogenic Detection Kit
(Thermo Scientific, Schwerte, Germany), resulting in a
coloured precipitate. Southern Blot The restriction endonucleases EcoRI, HindIII, NcoI, PstI,
XbaI and XhoI were used to estimate the number of 16S
operons present in the genome of E. corrodens. These
enzymes were additionally tested to assure that there
was no cutting site within the 16S region, where the de-
tection probe hybridizes during the Southern Blot, in
order to prevent occurrence of too many bands resulting
from binding of the probe to cut target sequences. As
shown in Fig. 3, a maximum number of four bands
could be detected after cutting with EcoRI, HindIII and
PstI. The presence of fewer visible bands for the restric-
tion with NcoI, XbaI and XhoI could be the result of the
emergence of long DNA strands containing two or more
16S regions after enzymatic incubation. According to
our investigations, a 16S operon number of four is very
likely for E. corrodens, therefore using P. gingivalis,
which contains the same 16S operon number, as the
standard for estimating the overall bacterial count,
seems to be very appropriate. Lyophilization
A
d A test to determine the effect of lyophilization of PCR
reagents upon the amplification reaction was conducted
using the E. corrodens specific PCR as well-characterized
test system. The PCR mix contained a final concentra-
tion of 0.25 μM for each primer, 50 % 2x SBYR Green
Mastermix (Roche, Mannheim, Germany), 5 % trehalose,
0.25 μg/μl BSA, 0.75 % PEG 8000, 10 ng DNA and the
remainder ddH2O. The lyophilization mix was frozen at
-80 °C for 15 min and finally freeze-dried for three hours
in a precooled lyophilizer. Long-time storage up to four
months at 4 °C and room temperature were compared,
with mixes protected from light, both with and without
the addition of polymerase stabilizing trehalose, using
the Lightcycler system for real-time PCR. A negative
control was run during lyophilization to exclude any
contamination during the process. For hybridization, the Biotin PCR Labeling Master
(Jena Bioscience, Jena, Germany) was used to generate a
probe. 16S primers and template DNA from E. corrodens
were applied to produce a biotinylated amplicon by in-
corporation of biotinylated dUTPs during PCR. The
probe was purified with the NucleoSpin® Gel and PCR
Clean-up Kit (Macherey-Nagel, Düren, Germany). To
ensure a complete target sequence for hybridization, an
examination of possible cutting sites within the sequence
was conducted: an enzymatic incubation of the 16S
amplicon with the aforementioned enzymes and subse-
quent check by comparing the band size of digested Gaertig et al. BMC Oral Health (2015) 15:165 Page 4 of 8 Fig. 1 Thermocycling device with different temperature zones and a rotating PCR chip. Left: A schematic drawing illustrates the arrangement of
the heating elements (Ta = annealing temperature) and the fluorescence detector. A rotating chip passes each zone and at 72 °C the
fluorescence signal is measured. The right image shows the lab set-up, including the driving motor and temperature control Fig. 1 Thermocycling device with different temperature zones and a rotating PCR chip. Left: A schematic drawing illustrates the arrangement of
the heating elements (Ta = annealing temperature) and the fluorescence detector. A rotating chip passes each zone and at 72 °C the
fluorescence signal is measured. The right image shows the lab set-up, including the driving motor and temperature control Standard curves For
the
detection
and
quantification
of
the
ten
periodontitis-causing bacteria used in this study, and to
determine the detection limits, quantitative PCR assays
were established. By utilizing dilution series of bacterial
genome copy numbers, standard curves were created ac-
cording to the resulting amplification curves. As an ex-
ample, the standard curves of C. gingivalis and E. corrodens are given in Fig. 2. To obtain reliable and re-
producible results, the efficiency of the PCR should be
greater than 1.8. Standard curves were set up for the
remaining periodontal bacteria in the same way. Detection limits determined by the Lightcycler 480 II
were as follows: P. intermedia 2000 copies/reaction, C. gingivalis 20 copies/reaction, C. rectus 2 copies/reaction,
A. actinomycetemcommitans 10 copies/reaction, P. gingi-
valis 10 copies/reaction, E. corrodens 200 copies/reac-
tion, P. micra 2000 bacteria, T. denticola 20000 copies/
reaction, overall count 200 copies/reaction. -
y= 3.9071x + 25.768
0
10
20
30
40
50
-5
-4
-3
-2
-1
0
1
2
CP
log c
C. gingivalis
y = - 3.3468x + 13.977
0
5
10
15
20
25
30
-4
-3
-2
-1
0
1
CP
log c
E. corrodens
A
B
Fig. 2 Standard curves: crossing point (CP) versus sample concentration. a C. gingivalis with an efficiency of 1.80 ranging from 20 to 2 million
bacteria, and (b) E. corrodens with an efficiency of 1.99 ranging from 200 to 2 million bacteria. All data are presented as means ± SD -
y= 3.9071x + 25.768
0
10
20
30
40
50
-5
-4
-3
-2
-1
0
1
2
CP
log c
C. gingivalis
A y = - 3.3468x + 13.977
0
5
10
15
20
25
30
-4
-3
-2
-1
0
1
CP
log c
E. corrodens
B B CP Fig. 2 Standard curves: crossing point (CP) versus sample concentration. a C. gingivalis with an efficiency of 1.80 ranging from 20 to 2 million
bacteria, and (b) E. corrodens with an efficiency of 1.99 ranging from 200 to 2 million bacteria. All data are presented as means ± SD Page 5 of 8 Gaertig et al. BMC Oral Health (2015) 15:165 Page 5 of 8 amplification as exemplarily shown in cycle 20 and 39 of
sample 2 (see Fig. 4a). Nevertheless, the expected in-
crease of fluorescence and a positive PCR result of the
targeted 279 bp region of S. Lyophilization Preservation of the biological activity of the polymerase
after freeze-drying was demonstrated for the examined
period of four months. E. corrodens-specific PCR evalu-
ation shows that this long-term storage at 4 °C after
lyophilization did not lead to any changes of the amplifi-
cation curve, whereas storage at room temperature
flattened the typical course of the curve and its charac-
teristic exponential increase. In this case, the real-time
PCR crossing points occurred in a delayed manner (see
Fig. 5 and Additional file 3: Table S3). The addition of trehalose to the PCR mix proved to be
essential for
long-term
storage,
especially
at
room
temperature. Amplification with the reaction mix with-
out trehalose led to a poor signal, lower overall fluores-
cence levels and delayed CPs after storage at room
temperature for four months. Standard curves aureus were demonstrated
in the initial tests (see Fig. 4b). Fig. 3 16S copy number of E. corrodens determined by Southern
hybridization. Genomic DNA was cut with 1) EcoRI, 2) HindIII, 3) NcoI,
4) PstI, 5) XbaI, 6) XhoI. 7) positive control. EcoRI, HindIII and PstI show
the maximum number of 4 bands Discussion and conclusions Periodontitis is a ubiquitous disease worldwide with the
need for effective diagnostics, fast treatment and inhib-
ition of its progression in order to prevent tooth loss. Furthermore, oral infections can potentially lead to se-
vere secondary diseases such as sepsis, systemic or car-
diovascular disorder and pneumonia [18]. Fig. 3 16S copy number of E. corrodens determined by Southern
hybridization. Genomic DNA was cut with 1) EcoRI, 2) HindIII, 3) NcoI,
4) PstI, 5) XbaI, 6) XhoI. 7) positive control. EcoRI, HindIII and PstI show
the maximum number of 4 bands In order to facilitate the detection and quantification
of bacteria species indicative of the disease, we developed
a
PCR-driven
point-of-care
device
for
periodontitis-
associated pathogen identification and showed basic
functionality. PCR-on-a-chip The quantitative PCR assay for bacterial strains was
developed successfully and standard curves were set up
with the Lightcycler system. For absolute quantification,
unknown concentrations of samples can be compared
with already established standard curves, where at least
one single standard sample of known concentration fall-
ing within the range of the imported curve has to be in-
cluded in each PCR run to serve as a reference. Ximénez-Fyvie et al. already showed in 2000 that sub-
gingival plaque samples of healthy patients often contain
significantly lower bacterial numbers, so the achieved
detection limits here of as few as two bacteria in some
cases are suitable to analyze the often very high bacterial
loads in cases of existing periodontitis [19]. Developing
the device further, the described quantification proced-
ure will be transferred onto the point-of-care device to
enable testing of patient-derived samples in the future. First fluorescence measurements were performed with
the Qiagen ESElog fluorescence measurement system
placed directly above the 72 °C temperature zone of the
thermocycling device, which is responsible for the elong-
ation step of the PCR. The ESElog USB E470/D520 is
suitable for detection of the DNA-intercalating SYBR
Green dye contained in the PCR mastermix. For every cycle during which the sample-containing
chip passed through each of the different temperature
zones on the thermocycling device, the continuous rise
in fluorescence corresponding to the amount of S. aur-
eus-specific amplicon present at that time was measured. The resulting rise in fluorescence as measured over 40
cycles by the detector can be seen in Fig. 4. During the heating steps, bubble formation in the
PCR chambers led to aberrations in the course of Gaertig et al. BMC Oral Health (2015) 15:165 Page 6 of 8 0
20
40
60
80
100
120
0
2
4
6
8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40
Fluorescence 470 - 520 [mV]
Cycles
sample 2
sample 1
300 bp
sample: 1 2
A
B
Fig. 4 Real-time fluorescence detection. Singleplex PCR shown on the lab-on-chip system. a Fluorescence measurement exemplarily shown for
two samples. b Resulting amplicon on a 2 % agarose gel B Southern Blot analysis could identify four copies of the
16S region in the genome of E. corrodens. This method
circumvents sequencing of the complete E. corrodens
genome, whose full sequence is not available yet. PCR-on-a-chip How-
ever, this number might be higher in the case of incom-
plete cutting by the restriction enzymes or in the case of
two or more 16S areas being located on one DNA frag-
ment arising from restriction digest. Nevertheless, these
results show that using P. gingivalis as a representative
for periodontopathogenic bacteria is appropriate for gen-
erating the standard curve for overall bacterial counts
with a typical 16S copy number of four. The use of this
bacterium for the generation of a universal standard curve for the assessment of total microbial counts was
already described by Kirakodu et al. [20]. As a proof of concept, the PCR assay could be shown
to work in PCR chips rotating over several heating zones
on a specially designed thermocycling device. Due to the
small reaction volume required and the lack of necessity
for heating and cooling steps by the heating blocks
themselves, the reaction time was one-third faster than
in conventional thermocyclers, even without any extra
care taken to further optimize the individual PCR steps. In the future, this reaction will be optimized with
shortened
denaturation/annealing
steps
and
PCR-
additives or fast polymerases, for example a KAPA2G 0
2
4
6
8
10
12
14
16
18
20
0
10
20
30
40
Fluorescence 483 - 533
Cycles
4 Months
0
2
4
6
8
10
12
14
16
18
20
0
10
20
30
40
Fluorescence 483 - 533
Cycles
1 Month
1-3
4
5-7
8
9
11
10
1-3
4
5-7
8
11
10
9
A
B
Fig. 5 Lyophilization test. Comparison of the polymerase activity after 1 (a) and 4 (b) months of storage of freeze-dried PCR mixes.1-3) storage at
4 °C, 4) storage at 4 °C without addition of trehalose, 5-7) storage at room temperature, 8) storage at room temperature without addition of trehalose,
9) PCR negative control, 10) negative control stored at 4 °C, 11) positive control after lyophilization 0
2
4
6
8
10
12
14
16
18
20
0
10
20
30
40
Fluorescence 483 - 533
Cycles
1 Month
1-3
4
5-7
8
9
11
10
A 0
2
4
6
8
10
12
14
16
18
20
0
10
20
30
40
Fluorescence 483 - 533
Cycles
4 Months
1-3
4
5-7
8
11
10
9
B A B Fig. 5 Lyophilization test. PCR-on-a-chip Comparison of the polymerase activity after 1 (a) and 4 (b) months of storage of freeze-dried PCR mixes.1-3) storage at
4 °C, 4) storage at 4 °C without addition of trehalose, 5-7) storage at room temperature, 8) storage at room temperature without addition of trehalose,
9) PCR negative control, 10) negative control stored at 4 °C, 11) positive control after lyophilization Gaertig et al. BMC Oral Health (2015) 15:165 Page 7 of 8 Page 7 of 8 Fast DNA Polymerase (Kapabiosystems, Wilmington,
USA) with an elongation time of approximately 1 s/kb,
thus leading to a significantly shortened overall reaction
time. Moreover, a further reduction of the reaction vol-
ume can accelerate the heat transfer, consequently low-
ering the required amount of reagents and costs [21]. showed 1998 in their complex theory that there are typ-
ical early and late colonizers in the periodontal disease
progression, indicating different manifestations of peri-
odontitis requiring respective individualized treatment. To find out about the microbial composition of
subgingival patient samples, dentists usually have to
send plaque samples to a clinical laboratory. Cultiva-
tion, immunological, or DNA probe analysis and re-
cently more sensitive nucleic acid based test systems
such as the microIDent® (Hain Lifescience, Nehren,
Germany) can deliver information for optimal therapy,
however still with the necessity and cost of shipment
to laboratories [24]. In our point-of-care system, pipetting steps for individ-
ual reagents are minimized leading to fewer sources of
error and increasing the convenience for the end user. Therefore, the use of lyophilized reaction mixes was
tested in order to enable long-term storage. It was dem-
onstrated that Taq polymerase
could
be
protected
against degradation and loss of inactivity when stored at
4 °C, while some reduced activity was evident when
stored at room temperature for up to four months. Based on these results the reaction mix is suitable for
storage in conventional refrigerators. The simple, model point-of-care system presented
here, which can be easily applied in every dental office,
reduces both time and costs for diagnostics and elimi-
nates the necessity for transport to external laboratories
and therefore facilitates crucial fast treatments in the fu-
ture. Additionally, the chip can be adjusted and used for
further applications in medical areas with a need for fast
detection and analysis of pathogens, such as those caus-
ing sepsis or regional/global pandemics. Authors’ contributions CG drafted the manuscript, carried out lyophilization experiments and PCR-
on-a-chip together with KN. LG contributed to Southern Blot analysis and JB,
NL, LR have been involved in real-time PCR experiments. CB helped with the
device’s assembly and developed the software for PCR analysis. KV and DK
conceived the project, participated in the design and coordination. All au-
thors read and approved the final manuscript. Availability of supporting data The supplemental data of Additional file 1: Table S1,
Additional file 2: Table S2 and Additional file 3: Table S3
supporting the methods and results of this article are in-
cluded within the additional Excel files. Competing interests
The authors declare that they have no competing interests. p
g
The authors declare that they have no competing interests. Abbreviations Abbreviations
BSA: Bovine serum albumin; CP: Crossing point; PCR: Polymerase chain
reaction; PC: Polycarbonate; PEG: Polyethylene glycol. In general, a significant problem for treatment and
prevention of periodontitis is that, up to now, standard
home dental care products such as toothbrushes and
floss cannot completely eliminate bacterial plaque for-
mation and prevent occurrence of the disease [9]. Thera-
peutic
measures
e.g. surgery,
tooth
extraction
and
eventually dentures are a massive financial burden for
patients or dental insurance, if purchased. Taken to-
gether, this shows the importance of developing a quick
to apply, reliable point-of-care system for the diagnosis
and quantitative classification of periodontal infections
in order to facilitate the immediate implementation of
the optimal healthcare plan that is available and for the
diagnosed stage of the disease. Socransky et al. [10] PCR-on-a-chip However, long-term studies to examine the shelf life of
the reaction mixes are currently being carried out in
order to see whether Taq is damaged after a longer
period of time following the dehydration process. The optical system for fluorescence detection was inte-
grated onto the device, revealing the problem of bubble
formation within the sample during PCR. PCR mix de-
gassing, higher glycerol content and changing hydropho-
bicity as suggested by Trung et al. and Jankowski et al. [22, 23] did not completely eliminate this problem and
occasional aberrations in the course of fluorescence
measurements were observed. Retrofitting of the system
with a heating block or a hot air fan at the top of the
chip with taller cavities might help to prevent evapor-
ation and subsequent bubble formation. Additional files Additional file 1: Table S1. Organisms and cultivation conditions. (XLSX 11 kb)
Additional file 2: Table S2. Primer sequences and product size. (XLSX 12 kb)
Additional file 3: Table S3. Crossing points (CP) corresponding to
Fig. 5 (XLSX 10 kb) Additional file 1: Table S1. Organisms and cultivation conditions. (XLSX 11 kb)
Additional file 2: Table S2. Primer sequences and product size. (XLSX 12 kb)
Additional file 3: Table S3. Crossing points (CP) corresponding to
Fig. 5 (XLSX 10 kb) Additional file 1: Table S1. Organisms and cultivation conditions. (XLSX 11 kb)
Additional file 2: Table S2. Primer sequences and product size. (XLSX 12 kb)
Additional file 3: Table S3. Crossing points (CP) corresponding to
Fig. 5 (XLSX 10 kb) In the further development of this device, an on-chip
DNA purification module will be integrated into the
chip and multiplex reactions with sequence-specific
fluorescent probes should be established in order to
avoid the use of unspecific intercalating dyes. Further-
more, biomarker tests could be implemented into the
chip construction, which helps to characterize and
optimize patients’ state of health as well. Additional file 3: Table S3. Crossing points (CP) corresponding to
Fig. 5 (XLSX 10 kb) Author details
1N
h
l 1Nanotechnology Unit, Department of Diagnostics, Fraunhofer Institute for
Cell Therapy and Immunology, Perlickstraße 1, 04103 Leipzig, Germany. 2Friedrich Schiller University, Jena, Germany. 3Scanbec GmbH,
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Thi
j This project was supported by the Federal Ministry for Economic Affairs and
Energy (AIF-project KF2302705AK1). Furthermore we thank Dr. David M. Smith for constructive criticism and reviewing the manuscript. Page 8 of 8 Page 8 of 8 References 18. Li X, Kolltveit KM, Tronstad L, Olsen I. Systemic Diseases Caused by Oral
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• Our selector tool helps you to find the most relevant journal
• We provide round the clock customer support
• Convenient online submission
• Thorough peer review
• Inclusion in PubMed and all major indexing services
• Maximum visibility for your research
Submit your manuscript at
www.biomedcentral.com/submit
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English
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Recognition for a black couple in a mock silver alert: Comparing couples presented together or separately with or without glasses
|
Current psychology
| 2,022
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public-domain
| 13,051
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Abstract Demographic trends indicate an increasing elderly population accompanied by an increase in the prevalence of individuals
with Alzheimer’s disease (AD). These trends are likely to result in increasing numbers of elderly individuals who wander
away from home or care facilities. There is limited research on the efficacy of systems for alerting the public about missing
elderly individuals, such as Silver Alerts (SA). Previous research on SAs was limited to alerts featuring White senior citizens. The present study is the first to extend SA research to Black senior citizens. A sample of college students (N = 210) viewed
a mock SA along with a short video of a “missing” couple and later attempted to recognize the two individuals from a series
of photos. The male and female targets were shown in the SA either together or separately and with or without glasses, and
participants were shown photos with and without glasses. The results indicated no effect of whether the couple was shown
together or separately, but participants were more likely to recognize the missing male without glasses when he had been
shown without glasses in the SA. The female target was recognized more often when wearing glasses than when not wearing
glasses, irrespective of how she had been shown in the SA. The results suggest that the appearance of the target at encoding
and at recognition may affect ability to identify the target, but that such effects may depend on individual characteristics. Keywords African American · Silver alerts · Face recognition · Attitudes toward elderly · Conscientiousness words African American · Silver alerts · Face recognition · Attitudes toward elderly · Conscientious Humans are considered face recognition experts for people
with whom we are familiar; however, studies have shown
that recognition of faces is error-prone (Young & Burton,
2017a, 2017b). Unfamiliar faces of older adults who are
seen in missing person alerts, such as Silver Alerts (SAs)
are therefore hard for people to recognize. a mock SA. Additionally, we tested whether wearing glasses
has an effect on face recognition. Furthermore, we measured
how recognition may vary with individual personality dif-
ferences including Attitudes Towards Older Adults (ATOA),
conscientiousness, experience with older adults, and con-
fidence ratings in responses to target photos. SAs are not
as well-known as AMBER alerts (AA) outside of the U.S.;
therefore, we will describe SAs and the importance of hav-
ing SA type programs worldwide. Current Psychology (2023) 42:12424–12438
https://doi.org/10.1007/s12144-021-02531-8 Current Psychology (2023) 42:12424–12438
https://doi.org/10.1007/s12144-021-02531-8 Abstract In the present study, we investigate face recognition in the
context of public alert systems intended to help locate indi-
viduals who have gone missing. Past studies on face recogni-
tion in relation to SAs have only used White targets (Gier,
2019; Gier & Kreiner, 2019; Gier & Kreiner, 2020; Gier, V. S., Kreiner, D. S. Recognition of a missing elderly couple
in a Silver Alert (under review)); therefore, in the present
study we investigated face recognition for a Black couple in Recognition for a black couple in a mock silver alert: Comparing
couples presented together or separately with or without glasses Vicki S. Gier1 · David S. Kreiner2 Accepted: 14 November 2021 / Published online: 4 January 2022
© The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature 2022 Accepted: 14 November 2021 / Published online: 4 January 2022
© The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature 2022 2
Departments of Psychology, Nutrition, and Kinesiology,
University of Central Missouri, Warrensburg, MO, USA 1
Department of Arts & Sciences, Mississippi State
University-Meridian Campus, 1000 Hwy 19 North,
Meridian, MS 39307, USA Importance of Silver Alerts Research on face recognition relating to a SA is particularly
important as our population is aging and life expectancy is
increasing (Petonito et al., 2013, p. 17). According to the
U.S. Census Bureau (2019), the number of older Ameri-
cans is expected to increase from 40.3 million (2010) to a
predicted 85.7 million by 2050. According to a 2019 report
by the United Nations, “Older people account for more than
one fifth of the population in 17 countries today, and the
United Nations Department of Economic and Social Affairs 2
Departments of Psychology, Nutrition, and Kinesiology,
University of Central Missouri, Warrensburg, MO, USA .(123456789
3 Current Psychology (2023) 42:12424–12438 12425 12425 Population Division’s projections to the end of the century
indicate that this will be the case in 2100 for 155 countries,
covering a majority (61 per cent) of the world’s population”
(United Nations, 2019, p. 5). These statistics are alarming
when factoring in the number of older adults who will suffer
from neurodegenerative diseases such as Alzheimer’s dis-
ease (AD), Parkinson’s disease, and Amyotrophic Lateral
Sclerosis (ALS), and Huntington’s disease (HD) (Lu et al.,
2013). of finding a missing older adult who is believed to be in
immediate danger. g
The aging of America's increasingly diverse population
requires a revised narrative about the value of immigrants,
refugees, and minorities to elders' well-being. The preva-
lence of AD appears significantly higher in Black people
than in White people. Based on a meta-analysis, Steenland
et al. (2016) concluded that the prevalence in the U.S of
AD for individuals between the ages of 65 to 90 is 5.5% for
White people, compared to 8.6% for Black people. Steen-
land et al. noted that the reasons for the difference in AD
prevalence are not clear, but could be related “biological,
psychological, and socioeconomic factors” (p.71). Reese
(2015, np.) found that, “African-Americans are 64 percent
more likely to develop AD than CA after adjusting for age,
gender and education.” The Alzheimer's Association (2019,
np.) recently reported that, “for all adults 65 and older, Afri-
can- Americans are about twice as likely to have Alzheimer's
or other dementias as older Whites and Hispanics.” Interest-
ingly, “a higher estimated prevalence of AD in AAs versus
CCs is consistent not only with higher incidence rates but
also higher survival after AD diagnosis for AAs” (Steenland
et al, p. 5). Importance of Silver Alerts Four studies have found consistent evidence of an
approximately 20% longer survival after AD diagnosis for
Black individuals versus White individuals. These studies
support the urgency to learn more about the ability of the
public to recognize missing elderly Black individuals. Bar-
riers for recognition of an older Black person who is missing
in a SA include biases people have for recognizing faces of
different groups of people. Alzheimer’s disease is a type of dementia that slowly
deteriorates the person’s brain, affecting memory as well
as all bodily functions (Alzheimer’s Organization, 2020). Over time, both the person with AD and their families suf-
fer physical and emotional consequences of serious memory
loss. It is important to note that not all older adults have
dementia or AD. Many older adults living into their 90s
are still cognitively alert; however, the National Institute on
Aging (NIA) stated that “as many as 5.5 million Americans
age 65 and older may have Alzheimer’s” (National Institute
on Aging/National Institute on Health, Feb 17, 2021). “An
estimated 5.8 million Americans age 65 and older are living
with Alzheimer's dementia in 2020 and 80% are age 75 or
older” (Alzheimer’s disease Organization, 2020). The Alz-
heimer’s Association (2020) predicted that by “2050, one
new case of AD is expected to develop every 33 seconds,
resulting in nearly 1 million new cases per year” (np). Vespa
et al. (2020) noted that by “2030, all Baby Boomers will be
older than 65 and after 2050, all Baby Boomers will be older
than age 85” (p.23). One common symptom of AD and other forms of demen-
tia is wandering and elopement. . One particular behavior
associated with AD in the elderly is pacing. Pacing can then
evolve into wandering which is a potentially life-threatening
event. Thomas (1995) defined wandering as “a purposeful
behavior that attempts to fulfill a particular need (from
the context of the wanderer), is initiated by a cognitively
impaired and disoriented individual, and is characterized
by excessive ambulation that often leads to safety and/or
nuisance-related problems” (p. 35). The act of wandering
can especially lead to dangerous outcomes when the AD
patient goes from simple wandering to elopement. Elope-
ment, as stated by Aud (2004), “the act of wandering away
safe residence, is very dangerous for the older adult with
dementia” (p. 362). Previous Research on Silver Alerts Africa and England, where both White and Black confeder-
ates approached both Black and White people in shopping
centers who were later “asked to identify the confederate
from both a sequential lineup, in which photographs are seen
one at a time, and a forced-choice recognition test, where
photographs are seen simultaneously. In both countries and
with both identification tasks the confederates were better
recognized by people of their own race” (Wright, et al, p. 119). Scores on the attitude scale were not related to recognition
of the target; however, experience with elderly individuals
was related to target recognition. Gier and Kreiner (2019)
suggested that “Silver Alerts can be effective under cer-
tain circumstances” (p. 410) and more research is needed
to investigate whether increasing experience with older
adults increases the efficacy of Silver Alerts. fi
Gier and Kreiner (2020) had participants view an edu-
cational video on the importance of SA before viewing a
SA showing a White female. This study also produced a
high overall recognition rate (70%), with even higher rec-
ognition when participants had watched the educational
video (79%). Furthermore, females correctly identified the
missing woman more than males, and White participants
correctly identified the missing woman more than Black
participants. Experience with older adults was also related
to recognition of the woman in the SA.f Researchers such as Bernstein et al. (2007), Fisk and Neu-
ger (1990), and Rodin (1987) conducted studies focused on
how faces are categorized as either being a member of an
out-group versus in-group and how categorization can result
in either deep processing (processing individuating features)
of in-group faces compared to less effective processing of
individuating features of out-group faces (Bernstein et al.,
2007; Fisk & Neuberg, 1990; Rodin, 1987). “Thinking cat-
egorically about out-group members leads individuals to
search for category-specifying features (e.g., skin tone) in
cross-race faces, instead of the individuating features that
distinguish one face from another (see also MacLin & Mal-
pass, 2001, 2003)” (in Bernstein et al., 2007, p.706). In the
present study, we investigate recognition for an adult Black
couple in a mock SA. Although testing for ORB was not
the purpose of the present study, it is important to consider
that Black participants may show better overall performance
that participants of other ethnicities in recognizing Black
individuals shown in a SA. Previous Research on Silver Alerts Gier (2019) conducted two studies on individual difference
factors and the effect of an educational video on recognizing a
man in a mock SA. In Study 1, college students viewed both
a video of the missing man and an educational video on Sil-
ver Alerts before attempting to recognize the man. In Study 2,
Gier experimentally manipulated whether participants viewed
the educational video. Both studies resulted in high recognition
rates (83.90% and 75.1%, respectively). Those who recognized
the target had significantly higher empathy scores and metacog-
nitive ratings (i.e., confidence) in both studies, with inconsist-
ent differences on other individual difference variables. White
participants were more likely than Black participants to recog-
nize the White male target, consistent with previous research on
Own-Race Bias in face recognition. Although there are a few published studies on missing
older adults in mock SA, the target seniors in all of the previ-
ous studies were White (Gier, 2019; Gier et al., 2017; Gier &
Kreiner, 2019; Gier & Kreiner, 2020). The present study is
the first to our knowledge to investigate ability to recognize
Black individuals in the context of a SA. Gier and Kreiner (unpublished) used a SA featuring a White
couple (in their 80s) who were videotaped together and viewed
in a mock SA. They investigated recognition of the targets in
relation to individual difference variables and whether the cou-
ple appeared together or separately in the SA. In study 1, partici-
pants correctly identified the missing female target 5.3% of the
time and the missing male target 84.7% of the time. In study 2,
the researchers replicated study 1 by adding a condition where
participants viewed the targets either together or separately. Gier
and Kreiner (unpublished) and found (56.7%) correctly recog-
nized the female target while 75% correctly identified the male
target. In both studies, participants’ attitudes towards older adults
were related to recognition of the targets. In order to understand
why recognition for all face recognition studies, we will now
discuss biases towards older adults. Own‑Age Bias (OAB) An OAB has been demonstrated with studies finding signifi-
cant differences in accuracy depending on the relationship
between the age of the participants and the age of the targets. McKelvie (1987), for instance, has indicated superior rec-
ognition of faces of one’s own gender relative to the other
gender. Although the OAB has been recognized within face
recognition research literature, the sample in the present
study was comprised of college students; therefore, it was
not practical to investigate OAB in our current study. An OAB has been demonstrated with studies finding signifi-
cant differences in accuracy depending on the relationship
between the age of the participants and the age of the targets. McKelvie (1987), for instance, has indicated superior rec-
ognition of faces of one’s own gender relative to the other
gender. Although the OAB has been recognized within face
recognition research literature, the sample in the present
study was comprised of college students; therefore, it was
not practical to investigate OAB in our current study. Previous Research on Silver Alerts Gier et al. (2017) conducted a ‘mock’ Silver Alert showing
an older woman in her 80s and found that recognition rates
were 6.8% when she was shown in typical (casual) clothing
and 14.3% when she was shown in atypical clothing (wear-
ing a nightgown), although the difference between condi-
tions was not significant. However, recognition was signifi-
cantly greater in comparison to a control condition in which
the target did not appear in the SA. Gier et al. reported no
significant differences in recognition related to participant
gender or ethnicity. Gier and Kreiner (2019) reported a much higher recog-
nition rate (84.6%) in another SA study. In this study, the
target was a White male in his 60’s. Females had better
recognition of the target than males, and those participants
reporting higher experience with older adults were more
likely to identify the target. Recognition of the target was
not related to participant age, race, or measures of empa-
thy or conscientiousness. The researchers replicated the
study and added a scale measuring attitudes about older
adults. Again, the recognition rate was high (83.90%). Many people are familiar with programs for missing and
abducted persons (e.g., AMBER alerts); however, SAs are
relatively new in the U.S. compared to AMBER alerts. Other names used for similar alert systems in the U.S. include Safe Return, Project Lifesaver, Missing Silver Alert,
Mattie’s Call (named after a 67 year old AD patient who
wandered away from her home and was found deceased eight
months later just feet from her home), and Golden Alert. Regardless of what the alert is called, all have the same goal 1 3 Current Psychology (2023) 42:12424–12438 12426 Africa and England, where both White and Black confeder-
ates approached both Black and White people in shopping
centers who were later “asked to identify the confederate
from both a sequential lineup, in which photographs are seen
one at a time, and a forced-choice recognition test, where
photographs are seen simultaneously. In both countries and
with both identification tasks the confederates were better
recognized by people of their own race” (Wright, et al, p. 119). Experience with Older Adults In addition to investigating appearance factors (wearing
glasses and appearing together vs. separately), we were
also interested in differences among participants that might
predict ability to recognize the missing individuals shown
in a SA. Attitudes towards outgroups (e.g., ethnicity, age,
gender), the amount of contact/experience one has with
the group, and whether the contact is positive or negative
can affect face recognition performance. Söderhamna et al. (2002) administered an instrument measuring attitudes
toward elderly individuals to nursing students in Sweden
at the beginning of their nursing program. The results sup-
ported earlier findings that limited positive experience
with older adults predicted less favorable attitudes and
feelings towards older adults. Gorelik et al. (2000) found
that students who had contact with older adults and those
who chose to take classes on aging reported more positive
attitudes towards older adults. Other studies (Brown et al.,
1999) found that overall positive experience and contact
with older adults tended to predict more positive attitudes
towards older adults compared to individuals who had nega-
tive experiences. Own‑Race Bias The OGB occurs when one has better recognition and accu-
racy for one’s own gender. Herlitz and Lovén (2013) found
the bias is highly asymmetrical meaning the bias occurs The ORB has been found between multiple races. For exam-
ple, a study by Wright et al. (2001) was conducted in South 1 3 Current Psychology (2023) 42:12424–12438 12427 more commonly with females. Only a handful of studies
have found the full crossover bias in both men and women
(Ellis et al., 1973; Man & Hills, 2016; Wright & Sladden,
2003). There is strong evidence in the literature showing
that females display an OGB more than males do. In stud-
ies focusing on OGB the participants view both males and
females sequentially followed by a recognition phase where
are presented with target face which was previously seen,
or distractor faces (foils). The majority of face recognition
studies following this type of research paradigm tend to
show an OGB for females, but not for males (e.g. Herlitz
et al., 1997; Herlitz et al., 2013; Shapiro & Penrod, 1986). Females tend to recognize faces of females more accurately
and quickly than males recognize female faces. Interestingly,
males and females performed similarly in recognizing male
faces (Lewin & Herlitz, 2002), showing a lack of OGB for
males. more commonly with females. Only a handful of studies
have found the full crossover bias in both men and women
(Ellis et al., 1973; Man & Hills, 2016; Wright & Sladden,
2003). There is strong evidence in the literature showing
that females display an OGB more than males do. In stud-
ies focusing on OGB the participants view both males and
females sequentially followed by a recognition phase where
are presented with target face which was previously seen,
or distractor faces (foils). The majority of face recognition
studies following this type of research paradigm tend to
show an OGB for females, but not for males (e.g. Herlitz
et al., 1997; Herlitz et al., 2013; Shapiro & Penrod, 1986). Females tend to recognize faces of females more accurately
and quickly than males recognize female faces. Interestingly,
males and females performed similarly in recognizing male
faces (Lewin & Herlitz, 2002), showing a lack of OGB for
males. attractiveness. Most importantly, they found that ability
to match faces was significantly worse when one face was
shown with glasses and one without. Couples Presented Together vs. Separately When a SAs of a missing couple is shown on the news as a
special alert, the couples are typically first shown together,
then sometimes, viewed separately. Although the SA context
is not the same as conducting an eyewitness lineup, the num-
ber of individuals shown to participants may be an impor-
tant factor in both contexts. Studies of eyewitnesses asked to
pick a perpetrator of a lineup together versus separately have
shown that sequential presentation appears to be the supe-
rior method, possibly because simultaneous presentation of
the possible perpetrators tends to lead victims to compare
the faces presented, often ending in false alarms when non-
perpetrators are identified from the lineup (Patterson & Bad-
deley, 1977). In the present study, we manipulated whether
the couple was shown together or separately in the SAs to
determine whether that factor would affect ability to recog-
nize the two people in the recognition phase. We should keep in mind that many gender studies show
more differences within genders than between genders (Bur-
ton et al., 2010; Darling et al., 2009; Herlitz & Lovén, 2013). Phenomena such as OGB present interesting challenges as
people may identify with different genders apart from their
biological sex. It may be important to distinguish between
differences in genders as compared to differences between
sexes. Impact of Wearing Eyeglasses In the interest of exploring factors that may impact ability to
recognize missing individuals shown in SAs, we investigate
the potential impact of showing the missing individuals with
or without eyeglasses, both in the SAs and later in photos
shown during a recognition test. Research has shown that
eyeglasses can act as an occlusion, preventing face recogni-
tion (Graham & Ritchie, 2019). Graham and Ritchie (2019)
found that wearing glasses and sunglasses can interfere with
face recognition. Kramer and Ritchie (2016) found that
wearing glasses can fool people who are not familiar with
the person. Patterson and Baddeley (1977) conducted a face
recognition study in which the targets were either wearing
or not wearing glasses, as one of several stimulus changes
in the study. They found that participants were more accu-
rate if the encoding and recognition phase were the same,
meaning that the individual wore glasses or did not wear
glasses both at encoding and recognition. However, McKel-
vie (1993) found no difference in face recognition depending
on whether the individuals were shown with glasses. McKel-
vie presented the faces for 3s each (20 faces, 10 with glasses
& 10 faces without glasses) and had the participants return
seven days later for the recognition phase of the study. Leder
et al. (2011) found that wearing rimless glasses resulted in
faces appearing less distinctive; however, rimless glassed
increased perceived trustworthiness, but did not decrease In a recent SA study (Gier & Kreiner, unpublished)
the researchers administered the Attitudes Toward Older
Adults scale (ATOA; Kogan, 1961). They hypothesized that
more positive attitudes towards older adults would relate
to increased face recognition of the elderly target individu-
als. Their hypothesis was supported for a female target, but
not for a male target. That pattern of results may suggest 1 Current Psychology (2023) 42:12424–12438 12428 target male in his late-fifties, found high confidence for those
accurately recognizing the target. that recognition of missing females is subject to differ-
ences in observers’ attitudes toward the target population
to a greater extent than for men. However, we should be
cautious about generalizing such findings when based on
only one target of each gender, as any differences could be
related to other characteristics of the targets. These results
do imply that relationships between individual differences in
observers and recognition of missing persons could depend
on the characteristics of the missing person. Impact of Wearing Eyeglasses We included
the ATOA measure in the present study to explore whether
attitudes toward the elderly would predict face recognition
when the targets were Black individuals rather than White,
as in the Gier and Kreiner (unpublished) study. Conscientiousness Another relevant individual difference variable is conscien-
tiousness. Komarraju et al. (2011) stated that conscientious-
ness is related to fact retention, elaborative processing, and
motivation. Fact retention, elaborative processing, and moti-
vation are important factors in face recognition studies, espe-
cially if the faces are not the same age, gender, or ethnicity
as the participants. Other researchers have described con-
scientious people as reliable, responsible, and persevering
(McCrae and Costa, 1997), and to have a tendency to follow
and adhere to ethical principles and moral activity (Blasi,
1980). Therefore, we hypothesized that individuals scor-
ing higher in conscientiousness would be more motivated
to pay close attention to details from the SAs and would
tend to follow directions more than those scoring lower on
conscientiousness. Participants high in conscientiousness
may correctly identify the male and female in the SAs. In
the recognition phase of the study, however, being conscien-
tious, they may also question their confidence unless they
were very sure their response was correct. A second purpose of the study was to investigate char-
acteristics of the SAs that may affect ability to later recog-
nize the individuals shown in the SAs. Older adults often
wear glasses; however, if older adults with AD wander or
elope, they may not be wearing their eyeglasses. Although
this may not seem important, if someone in the public sees
a missing older adult wearing glasses, but the SAs showed
the older adult without glasses, the mismatch based on the
presence of glasses may result in not recognize the missing
older adult. In the present study, we manipulated whether
the targets appeared in the SAs and video with or without
glasses as well as in the recognition phase of the study. We
hypothesized that recognition would be greater for the target
wearing glasses when the person had been shown in the SA
wearing glasses. In the present study, the SAs and an accompanying video
presented the Black couple either together or separately. Gier
and Kreiner (2020, unpublished) found no difference in rec-
ognition of the older adults whether presented together or
separately using White targets, thus we did not expect to find
a difference in recognition depending on whether the older
adults were presented together or separately.f Purpose of the Present Study The present study is the first research study, to our knowl-
edge, to include a Black couple as the targets in a mock SA
study. We located five previous studies on SAs; all five stud-
ies had White target individuals. Although we did not, in the
present study, directly compare Black older adult face rec-
ognition to recognition of White older adults, it is of interest
to determine whether recognition rates are similar to those
found in previous studies. As the number of older adults are
increasing rapidly, both in the U.S. as well as internation-
ally, more SAs will be posted for those older adults with
Alzheimer’s who wander away from their residence, or even
worse, who are elopers. As mentioned earlier, Black indi-
viduals have a relatively high prevalence of AD; therefore, it
is important to add to literature on face recognition relating
to SAs showing Black individuals. Metacognition and Face Recognition Participants’ confidence in their decisions about whether
they have recognized a missing elderly individual are impor-
tant in addition to whether they recognize the missing person
or not. Fleming and Lau (2014) stated that, “metacognitive
sensitivity refers to the ability of a person to discriminate
between different levels of performance, such as correct or
incorrect trials, also known as type 2 bias, over-or-under
confidence or calibration” (Fleming & Lau, 2014, np). The complexity of the task can have an effect on whether
a participant has a tendency to report either a high or low
confidence level, regardless of performance (Mazancieux
et al., 2018). An important metacognitive measurement in
face recognition studies is Retrospective Confidence Judg-
ments (RCJ). Gier and Kreiner (2020), in a SA study with a In terms of individual differences across participants,
we hypothesized that individuals who report greater con-
tact with the elderly and who have more positive attitudes
toward the elderly would be more likely to recognize the
targets. Gier and Kreiner (unpublished) found that scores on
the ATOA were related to recognition of a missing person
shown in a SA. They presented both a White male and White
female in their middle 80s and found that more favorable
attitudes toward older adults were associated with better
recognition of females, but not males. We hypothesized for 1 3 3 Current Psychology (2023) 42:12424–12438 12429 12429 dichotomous variable of response to the recognition photos
(yes/no) and the continuous variable of confidence ratings
in responses to the recognition photos. The correlational
aspect of the design explored relationships between target
recognition and the individual difference variables of par-
ticipant scores on measures of contact with the elderly, atti-
tudes toward the elderly, and conscientiousness scores (See
Figure 1 for flowchart of the study). the present study that there would be a significant difference
in ATOA scores related to target recognition, but possibly
more likely for the female than the male in the study based
on Gier and Kreiner’s (unpublished) findings. the present study that there would be a significant difference
in ATOA scores related to target recognition, but possibly
more likely for the female than the male in the study based
on Gier and Kreiner’s (unpublished) findings. i
Finally, we hypothesized that those scoring higher on
conscientiousness would more likely correctly recognize
both the male and female in the Silver Alert. Metacognition and Face Recognition If a person
scores high on the conscientious trait, we predicted they
would more likely focus more closely on the faces and
details in the study. Materials The study was conducted using the online Qualtrics plat-
form (https://www.qualtrics.com/). Mock SAs were created
in a PowerPoint presentation that was programed to auto-
matically progress during the slides. The SAs consisted of
a photo of the target couples viewed separately (either with
or without glasses) that included the physical descriptions of
the male and female targets’ age, height, weight, eye color,
and hair color, as well as a home-video where the couples
were videotaped separately in the SA (see Appendix A). A
phone number was also provided to make the poster appear
as realistic to a SA as possible. Another SA was created
showing a photo of the couples together both in the SA as
well as the home-video. The control condition consisted of
a different Black elderly coupled viewed both together and
separately in the SA and only together in the home-video
wearing glasses (see Appendix B for couples together seen
in the SA). Appendix C shows targets with and without
glasses. Participants The participants were recruited from a southeastern uni-
versity using SONA, a research management system that
allows students to sign up for research studies online and to
obtain credit for participation. Of the 222 individuals who
responded, 12 reported having technical difficulties (video
and/or audio). Data from those individuals were excluded
from the analyses, yielding a Sample size of 210. The sam-
ple size of 210 yielded a power greater than .90 for detecting
medium size effects in a chi-square contingency test or two-
way between subjects ANOVA. The majority of the sample
identified as female (71.4%, N = 150), with 59 identifying
as male (28.1%) and one identifying as transgender male
(0.5%). The sample included 147 participants identifying
as White (70.0%), 50 as Black participants (23.8%), 7 as
Hispanic participants (3.3%), 4 as Asian participants (1.9%),
and 2 as other ethnicities (1.0%). The mean age of partici-
pants was 20.07 years (SD =3.67, range of 18 to 45 years). We created the SAs so that they would be similar to alerts
in current use. We created a 42 second video of the SAs cou-
ple in the presence of the couple’s son and daughter-in-law
to make the couple feel comfortable conversing while being
videotaped on a cell phone. This created a more relaxed,
natural video of the couple than being videotaped alone by a
stranger. In the condition where the targets were videotaped
separately, the length of the videos was 21 seconds each for
the target male and female. The videos were counterbal-
anced when the couple was shown separately so that half
of the participants in that condition saw the SA male target
first then the female target and then reversed with the female
target first followed by the male target. Design The design of the study included both experimental and cor-
relational components. We manipulated whether the SAs
showed the couple together or separately (between-subjects). We also included a control condition which consisted of two
different elderly Black individuals. This constituted a control
condition in the sense that it provided a baseline recognition
rate when the target individuals were not included in the
SA; if participants in the four experimental conditions do
not recognize the targets at a rate above that of the control
condition, it would suggest that participants were performing
at chance. The couple in the control condition SA did not
appear in the recognition phase of the study. We manipu-
lated whether the SAs showed the target individuals wearing
glasses or not (between-subjects), and included photos in
the recognition phase that showed the individuals wearing
glasses or not (within-subjects, as all participants viewed
all the recognition photos). Dependent variables were the i
Before the recognition phase of the study we asked par-
ticipants to complete surveys that included Kogan’s (1961)
Attitudes Towards Older Adults (ATOA) scale to measure
participants’ attitudes towards older adults. This instrument
is intended to measure stereotypes of the aged and people’s
image of older adults. The scale consists of three domains:
personal appearance, resemblance, and the nature of inter-
personal relations across age generations. The scale consists
of 34 questions in total with 17 positively worded and 17
negatively wording, using a 6-point response scale rang-
ing from strongly agree (1) strongly agree to (6) strongly 1 3 Current Psychology (2023) 42:12424–12438 12430 Fig. 1 Flowchart and hypotheses Fig. 1 Flowchart and hypotheses Fig. 1 Flowchart and hypotheses Past research,
such as Wixted and Wells (2017), suggests that foil’s faces
should not look exactly like the description from the eye-
witness (such as wearing the exact same clothing) as this
can contribute to false recognitions. Foils should include
important features of the perpetrator such as color of eyes or
specific facial features. Therefore, in our study we included
people looking somewhat similar to the missing target male
and female, and some foils who were dissimilar in their
appearance (see Appendix E). Similarity was determined by
a panel of ten undergraduate students who viewed an array
of 60 photos of AA elderly males and females. The photos
were ranked by the judges, then the photos were arranged by
ranking the most similar to photos not similar to the target
couple. This process continued until we had 20 similar and
10 not similar photos. which stated: “You are going to be watching a short video
that lasts approximately 42 seconds. Please make sure the
volume is turned down so as not to distract others (i.e. if you
are in a computer lab or library participating in the study).”
The video showed the participants sitting in a living room
talking to their son and daughter-in-law (only the targets
were in the video). Equal time was given to the couple to
talk about their lives and about activities they had planned
for their week. After the participants viewed the video, they were then
asked to respond to demographic questions and a short sur-
vey on the amount of contact they had with older adults’
group in ages of 50-59, 60-69, 70-79, and 80+. Participants
responded with the choices of the following: I have no con-
tact with adults in this age range; I have very little contact
with adults in this age range; I have a moderate amount of
contact with this age range; and I have a great deal of contact
with this age. The participants were then asked to respond
to the IPIP (Goldberg et al., 2006) Conscientiousness items
and the ATOA questionnaire (Kogan, 1961). The photos in the recognition phase displayed the tar-
gets separately, dressed differently, with one photo of each
target wearing glasses and another photo without glasses
(Appendix E). . Fig. 1 Flowchart and hypotheses We also measured how much experience each partici-
pant reported with older adults. This measure was a 4-point
response scale. The age ranges consisted of four age ranges
(adults 50-59, 50-69, 70-79, and 80 and older). The prompt
indicated, “Below is a list of different age ranges. We would
like to know about your experience/contact with members
of those groups. For each group, indicate how much experi-
ence you have interacting with those age ranges in the past
five years. I have _______ experience interacting with adults
this age” with the four options being no/very little/moder-
ate amount of/great deal of experience. At the end of the
study we asked the participants demographic questions that
included the participant’s age, race, gender, and whether
they received AMBER Alerts or Silver Alerts on their cell
phones. The surveys and demographic questions served
as the retention interval of our study and took participants
approximately 20 minutes to complete. disagree. Negatively worded items were reverse scored; thus,
higher total scores on the ATOA indicated more negative
attitudes towards older adults. Kogan (1961) reported the
overall Cronbach’s alpha reliability for the ATOA (Kogan,
1961) to be .81. Content and construct validity for the ATOA
have been supported in more recent studies, such as a Chi-
nese study with a reported construct validity of .92 (Yen
et al., 2009) and a Turkish version of the scale at .94 (Küçük-
güçlü et al., 2011). We measured conscientiousness by using the Interna-
tional Personality Item Pool (IPIP, Goldberg et al., 2006). The participants were asked to rate 20 items on a 0 to 100
very inaccurate to very accurate scale. Scores for positively
and negatively worded items were each averaged, then the
overall conscientiousness score was calculated as the differ-
ence between the positive and negative scores. Cronbach’s
alpha for this scale ranges from .79 to .87 (Goldberg et al.,
2006). We used a total of 34 photos in the recognition phase of
the study. Thirty were foils (meaning the photograph was 1 3 Current Psychology (2023) 42:12424–12438 12431 not the target male or female in the study). We included two
different photos of each target, one wearing glasses and the
other without glasses. The control couple wore glasses (See
Appendix D). The target photos in the recognition phase
were not the exact photos shown in the SAs. Procedure Participants were randomly assigned to one of five condi-
tions. Four of the conditions contained the target couple. These conditions consisted of the target couple shown
together with glasses, together without glasses, separately
with glasses, or separately without glasses. The fifth con-
dition was a control condition in which a different elderly
Black male and female appeared; these individuals did not
appear in the recognition photos. Fig. 1 Flowchart and hypotheses Four photos of the target male and female
seniors were presented (one photo with and without glasses),
two photos of each of the targets wearing different clothes
and in different background settings. The majority of the
foils looked similar to the older AA couple; however, we
also added foils who appeared physically different than the
targets in the study, for example, differing in hair length,
skin tone, and facial features such as having either a round
or more square shaped jaw. For the females some facial fea-
tures such as the cheekbones, eyebrows, and lip size (also
including the color of lipstick) were different than the target
female’s facial features. The recognition phase of the study was presented at the
end of the study. We did not mention to participants that the
targets may not be in the recognition phase. We presented
photos of 34 elderly adults in their 80s, where 30 of the pho-
tos were foils. All 34 photos were presented in random order
(in other words, the male and female photos were in random
order such that each photo had an equal chance to be seen in
each position from 1-34). Each photo appeared separately
on the screen. The participants were asked to respond Yes or
No to the question: “Was this male (or female) in the Silver
Alert?” Next, the participants were asked to rate how confi-
dent they were in their decision of whether the person was
in the Silver Alert or not on a visual analogue scale of 0-100,
with 0 indicating they were not confident in their decision of
whether or not the person was in the SA, and 100 meaning
they were 100% confident the person in the photograph was
one of the targets in the SA. Overall Recognition Rates Each participant responded to four target photos: two for
the female and two for the male, with one photo of each
shown wearing glasses and one without. Overall recogni-
tion rates (percentage of participants responding Yes to the
target) were as follows: female with glasses (57.6%); female
without glasses (15.2%); male with glasses (33.3%); male
without glasses (31.0%). Mean false alarm rates were 11.9%
for female foils and 12.3% for male foils. Once the participants agreed to participate in the study,
they saw the following message: “You are now going to
see a Silver Alert Poster. You may take your time to study
the poster.” The next screen showed a photo of the couple. Immediately following the photo of the missing senior citi-
zens, the participants saw a slide of the description of the
couple. Next, a message was presented to the participants In order to determine whether participants could identify
the targets at a rate above chance, we conducted a chi-square 1 3 Current Psychology (2023) 42:12424–12438 12432 Table 1 Target recognition rates (%) by glasses condition and target
gender
Percentages shown are percentage of participants responding “Yes” to
the target photo. Glasses in Silver Alert
No
Yes
χ2
p
Female Target
No Glasses
21.1
12.0
2.54
.111
Glasses
61.8
63.5
0.06
.813
Male Target
No Glasses
50.0
20.2
16.63
<.001
Glasses
36.4
39.3
0.16
.693 Table 1 Target recognition rates (%) by glasses condition and target
gender
Percentages shown are percentage of participants responding “Yes” to
the target photo. Glasses in Silver Alert
No
Yes
χ2
p
Female Target
No Glasses
21.1
12.0
2.54
.111
Glasses
61.8
63.5
0.06
.813
Male Target
No Glasses
50.0
20.2
16.63
<.001
Glasses
36.4
39.3
0.16
.693
Table 2 Target recognition rates (%) by whether the couple appeared
together or separately in the silver alert
Percentages shown are percentage of participants responding “Yes” to
the target photo. Overall Recognition Rates Couple Shown Together
No
Yes
χ2
p
Female Target
No Glasses
14.3
17.9
0.36
.546
Glasses
60.3
63.8
0.20
.658
Male Target
No Glasses
32.1
37.1
0.40
.527
Glasses
31.5
40.7
1.33
.248 Table 1 Target recognition rates (%) by glasses condition and target
gender test of independence for each of the four targets (female and
male with and without glasses) to determine if proportions
of Yes and No responses to the targets differed between the
control condition and the four conditions (combined) in
which the target couple appeared in the SAs. For the female target wearing glasses, 41.4% of partici-
pants in the control condition responded Yes, compared to
62.6% in the other conditions, χ 2 (1, N=203) = 4.67, p =
.031. For the female target without glasses, there was not a
significant difference in identification, with 9.7% of those
in the control condition responding Yes compared to 16.8%
in the other conditions, χ 2 (1, N=204) = 1.00, p = .318. For the male target wearing glasses, 16.7% of those in the
control condition responded Yes, compared to 37.8% in the
other conditions, χ 2 (1, N=202) = 5.03, p = .025. For the
male target without glasses, 12.9% of participants in the
control condition responded Yes, compared to 35.5% in the
other conditions, χ 2 (1, N=203) = 6.14, p = .013. In sum-
mary, participants who had seen the target in the Silver Alert
identified the target at a significantly higher rate than those
who had not, except for responses to the female target who
was not wearing glasses. Percentages shown are percentage of participants responding “Yes” to
the target photo. Table 2 Target recognition rates (%) by whether the couple appeared
together or separately in the silver alert
Percentages shown are percentage of participants responding “Yes” to
the target photo. Couple Shown Together
No
Yes
χ2
p
Female Target
No Glasses
14.3
17.9
0.36
.546
Glasses
60.3
63.8
0.20
.658
Male Target
No Glasses
32.1
37.1
0.40
.527
Glasses
31.5
40.7
1.33
.248 Table 2 Target recognition rates (%) by whether the couple appeared
together or separately in the silver alert Percentages shown are percentage of participants responding “Yes” to
the target photo. Effect of Wearing Glasses We compared recognition rates across the four target pho-
tos, using z-tests for dependent proportions, and excluding
data from the control condition, in which the target male
and female did not appear in the SA. Because the overall
recognition rates indicated differences between the male and
female targets, we did not collapse data for the male and
female photos but report the results separately.i Silver Alert. In other words, when the male senior citizen
appeared without glasses in the Silver Alert, participants
were more likely to correctly identify his photo when he
was again shown without glasses. For the other three target
photos, there was not a significant difference in recognition
depending on whether the target was shown in the Silver
Alert with glasses or not. For male targets, the recognition rate was not significantly
different when the target wore glasses in the photo (37.8%)
than when the target did not wear glasses (35.5%), z = 0.35,
p = .363. However, the female target was recognized sig-
nificantly more often (62.6%) when the target wore glasses
than without glasses (16.8%), z = 7.90, p < .001. When not
wearing glasses, the male target was recognized more often
than the female target, z = 3.51, p < .001, but when wearing
glasses, the female target was recognized more often than
the male target, z = 3.35, p < .001. Effect of Couple Shown Together or Separately Table 2 shows parallel analyses comparing recognition rate
between conditions with the target appearing together or
separately in the Silver Alert. There was not a significant
relationship between the Together-Separate condition and
recognition of any of the four target photos. Confidence Ratings To test the relationship between whether the targets were
shown in the SAs with or without glasses and whether they
appeared in the recognition phase with or without glasses,
we conducted a chi-square contingency test for each of the
target photos (female and male with and without glasses)
to compare frequencies of participants responding Yes and
No in relation to whether the Silver Alert showed the target
wearing glasses or not. As shown in Table 1, participants
correctly responded Yes to the male target without glasses
more often when the target appeared in the Silver Alert with-
out glasses than when the target appeared with glasses in the For each of the four targets, we compared participants’ con-
fidence ratings (on a scale of 0-100%) for their responses
across the four conditions using 2 x 2 between subjects
factorial ANOVAs, with the two factors being whether the
Silver Alert showed the couple together or separately and
whether it showed them with or without glasses. Table 3
shows mean confidence ratings across the four conditions. For the female target without glasses, there was a significant
main effect of the Glasses condition, F(1,168) = 7.59, p = 3 3 Current Psychology (2023) 42:12424–12438 12433
Current Psychology (2023) 42:12424–12438
Table 3 Mean target confidence
ratings by glasses condition
and whether targets appeared
together or separately in the
silver alert
Couple Shown Together
Couple Shown Separately
No Glasses
Glasses
No Glasses
Glasses
M
SD
M
SD
M
SD
M
SD
Female Target
No Glasses
67.93
24.23
82.32
19.44
73.15
25.98
80.82
25.50
Glasses
67.37
25.52
73.58
24.32
71.92
23.86
74.67
23.69
Male Target
No Glasses
67.18
25.64
75.93
21.44
75.81
21.60
79.02
24.67
Glasses
71.11
26.33
69.52
27.46
72.69
25.18
76.86
22.90 12433 Table 3 Mean target confidence
ratings by glasses condition
and whether targets appeared
together or separately in the
silver alert .007, ηp
2= .04, indicating significantly higher confidence
ratings when the target had been shown with glasses than
without in the Silver Alert. No other main effects or interac-
tions were significant. For predicting recognition of the male wearing glasses,
the final logistic regression model included participant age
(odds ratio = 1.10), with greater ages predicting higher like-
lihood of recognizing the target individual; and confidence
rating for the target photo (odds ratio = 0.98), with higher
confidence ratings associated with lower likelihood of recog-
nition. Discussion i
We conducted a logistic regression analysis, using the
Backward Wald method, to predict recognition of each of
the four target photos from confidence rating to the target
photo, confidence to same-gender distractor photos, partici-
pant age, IPIP conscientious score, ATOA score, and a total
experience with the elderly score, which was calculated by
summing responses for reported experience with each of the
four age ranges (ages 50-59, 60-69, 70-79, and 80 or older),
with higher totals representing greater amounts of reported
experience with elderly individuals.i We explored the recognition of an elderly Black couple
shown in a mock Silver Alert in relation to whether the cou-
ple appeared together or separate in the Silver Alert and
with or without glasses in the SAs, as well as the relation-
ship between recognition and several individual difference
variables. Notably, this is the first face recognition study
using a mock SAs showing Black people. Previous studies
on Silver Alerts (Gier et al., 2017; Gier & Kreiner, 2019a;
Gier & Kreiner, 2019b; Gier, 2019) used White people as
their target seniors. Black older adults are a minoritized and
marginalized group in the U.S. which needs more represen-
tation in studies relating to AD. Black older adults are at
the highest risk of developing AD (Steenland et al., 2016). Several studies have revealed that in the U.S, Blacks are the
majority, rather than the minority, in many health-related
illnesses such as diabetes (Blacks and Hispanics are over
50% more likely to have diabetes than non-Hispanic Whites,
American Diabetes Association, 2020) and chronic kidney
disease (Laster et al., 2018, p.3). Breathett et al. (2018)
found not only that Blacks have the highest risk of heart fail-
ure, but “among patients admitted to an ICU for HF, African
Americans were less likely than Caucasians to receive pri-
mary care by a cardiologist. Primary care by a cardiologist
was associated with higher survival for both Caucasians and
African Americans” (2018, p. 415). Healthcare and biases
often prevent Black individuals from equal access. Health
conditions, and age in general, can have negative effects on
an older adult’s vision, resulting in many older adults need-
ing to wear eyeglasses. However, given that a missing older The logistic regression model was not significant for
predicting recognition of the female wearing glasses in the
recognition photo, as all predictors were eliminated from
the model. Confidence Ratings The final model containing those two predictors was
significant, χ2 (2) = 12.96, p = .002, Nagelkerke R2 = .086. See Figure 2 for classification plots for the two significant
logistic regression final models. Individual Differences and Target Recognition We compared recognition of the target across ethnicity
group (Black, White, Other) to determine whether there was
an advantage for Black participants in recognizing the Black
targets. For all four targets, the chi-square did not approach
significance, all p’s > .705. Discussion The initial model including all predictors was not
significant, χ2 (6) = 3.17, p = .717, Nagelkerke R2 = .021. Similarly, there were no significant predictors for recogni-
tion of the male not wearing glasses, χ2 (6) = 4.03, p = .673,
Nagelkerke R2 = .028. For predicting recognition of the female not wearing
glasses in the recognition photo, the final logistic regres-
sion model included participant age (odds ratio = 1.12),
with greater ages predicting higher likelihood of recogniz-
ing the target individual; ATOA (odds ratio = 0.96), with
higher scores (indicating more negative attitudes toward the
elderly) associated with lower likelihood of recognition; and
confidence rating for the target photo (odds ratio = 0.98),
with higher confidence ratings associated with lower likeli-
hood of recognition. The final model containing those three
predictors was significant, χ 2 (3) = 20.63, p < .001, Nagel-
kerke R2 = .166. 1 3 12434 Current Psychology (2023) 42:12424–12438 Current Psychology (2023) 42:12424–12438 Fig. 2 Classification plots from
logistic regressions predicting
recognition from individual dif-
ference variables (final models). (Symbols: N – No Y – Yes each
symbol represents 20 Cases) Fig. 2 Classification plots from
logistic regressions predicting
recognition from individual dif-
ference variables (final models). (Symbols: N – No Y – Yes each
symbol represents 20 Cases) adult may or may not be wearing glasses, more research is
needed on whether that aspect of appearance has an effect
on face recognition of Black individuals in a SA. especially an unfamiliar individual, is harder to recognize if
the target in the study phase of the study wore glasses, but
in the recognition phase the target did not wear glasses (and
vice-versa), than when the person wore glasses or did not
wear glasses at both exposure and recognition. For the two
individuals who appeared as targets in the present study,
the male target was more likely to be recognized when not
wearing glasses in the recognition photo when he had pre-
viously appeared in the SAs without glasses, but there was
no such matching effect when he was wearing glasses, and
this effect did not occur for the female target. However, the
female target was more likely to be recognized when wear-
ing glasses than when not wearing glasses, irrespective of
how she appeared in the SAs. Fig. 2 Classification plots from
logistic regressions predicting
recognition from individual dif-
ference variables (final models).
(Symbols: N – No Y – Yes each
symbol represents 20 Cases) Discussion (2020)
found that ratings of intelligence were higher for people
seen wearing glasses, even when the participants were
instructed to ignore the glasses and when viewing time was
very short. Gerger et al. (2017), Leder et al. (2011) found
that individuals wearing glasses were rated higher on suc-
cessfulness and intelligence than those not wearing glasses. These results suggest that people may make rapid inferences
about an individual based on whether the person is wearing
glasses. Fleischmann et al. (2019) conducted a study on the
effect glasses have perception of intelligence for politicians
running in an electoral race, both in the United States as well
as in India. Politicians who wore glasses in the U.S. were
viewed as being intelligent; however, the effect of glasses
making the politician appear intelligent was not significant
in India. Thus, the inferences that are drawn may vary across
cultures. It is possible that any of these inferences about
individual characteristics could serve as recall cues.ifi had the lowest recognition rate. It is possible that differences
in attitudes toward the elderly become more important in
relation to recognizing the missing person when particular
characteristics of the person make it more challenging to
make an identification. These results were partially consist-
ent with those of Gier and Kreiner (2020, unpublished), who
found that participants who recognized a White female or
male scored significantly lower on the ATOA than those who
did not (indicating lower levels of negative attitudes). We
did not find significant differences based on target recogni-
tion in relation to conscientiousness scores or self-reported
experience with the elderly. SAs are relatively new compared to missing children
alerts, such as AMBER alerts, but the two programs serve
the same purpose: to alert the public of a missing person
considered in possible grave danger and vulnerable due
to age, and physical as well as psychological disabilities. Research on missing children in AMBER alerts has shown
that people claim to look at missing people posters when in
a supermarket, yet when this question was experimentally
tested (Lampinen et al., 2009) the results showed partici-
pants had become complacent. Although customers reported
that they believe AMBER alerts are very important, nearly
none of the customers reported looking at the posters. Participants seemed to find it especially difficult to rec-
ognize the female target when she was not wearing glasses. Discussion Overall, she was not recognized significantly more often
when she appeared in the SAs than when she did not appear,
suggesting that participants were not better than chance at
identifying her photo during the recognition phase. We also
found that participants who did identify the female target
without glasses had a higher false alarm rate to foils than
individuals who did not identify this particular target, imply-
ing that those who identified the target were just more likely
to respond Yes in general, consistent with guessing. Researchers studying the fate of our older adults with
dementia fear similar complacency. The U.S. Census Bureau
(2019) reported that, “Older people are projected to outnum-
ber children for first time in U.S. history.” We encourage
researchers to focus on face recognition older of adults with
the same degree of importance of face recognition of miss-
ing children, runaway teens, and people with physical and/
or mental disabilities. Gier and Kreiner’s study (unpublished) with a White
older adult couple in their mid-80s presented the couples
either together or separately. They found no significant dif-
ferences between presenting the missing couple separately
in the SA compared to presenting them together. Similarly,
we found no evidence in the present study that whether the
couple appeared separately or together in the SA affected
later recognition. Those elderly adults who do wander from
home are sometimes alone, but often are with their spouse
or significant other who may or may not have AD (Examples
of actual alerts can be seen in the Appendices A-E). The
present results may be beneficial for those presenting SAs
on TV, allowing the alert to be presented with the couple
together, or separately as it does not appear to make a dif-
ference in recognition of the missing senior. However, we
should keep in mind that the present results do not conclu-
sively indicate the absence of a together-separate effect, just
a lack of evidence to support such a difference.f We recommend that every state should have a campaign
on educating the state residents on the different alert pro-
grams for people who are missing. Education on AMBER
alerts and Silver Alerts could be incorporated in school
assemblies, having someone from law enforcement, or
someone working with relatives of missing people, speak to
the students on the importance of all missing people alerts,
including SAs. Discussion In fact, recognition of the
target woman when she was shown without glasses was no
better when she was shown in the SA than when she was not
shown in the SA (control condition). These results suggest
that the importance of whether the person appears with or
without glasses at encoding and recognition may depend on
characteristics of the particular person. The results of the present study indicated that wearing
glasses is an important consideration in the public’s ability
to recognize missing elderly individuals. However, there was
only partial support for the matching hypothesis that recog-
nition would be greatest when the target’s appearance in the
SAs matched the recognition photo (e.g., wearing glasses
at both encoding and recognition). Matching appearance,
in terms of wearing glasses, may only affect recognition in
some situations, possibly dependent on unique factors asso-
ciated with each individual face. It is also interesting to note
that the results were not consistent with the idea that partici-
pants could rely entirely on the presence of the glasses as a
visual cue to identify the target. The results suggest that we
should not assume that individuals are equal or even similar
in the likelihood that they will be recognized by the public,
as ability to recognize them may depend on specific facial
characteristics as well as how they appear in the SAs and
when people are attempting to recognize them. One possible explanation for the importance of the
glasses is that observers may draw inferences about people
based on whether they wear glasses or not. It is possible that
when these inferences are made, they could affect memory
for those individuals. For example, glasses can give a per-
ception of intelligence (Fleischmann et al., 2019). An early
study by Harris (1991) found that people wearing glasses
were perceived as less attractive, more intelligent, and more Previous research has indicated a variety of factors
involving wearing glasses that could impact face recogni-
tion. Wearing glasses can reduce the accuracy of face rec-
ognition due to the emphasis on the eye region of the face
(Graham & Ritchie, 2019). Other research has indicated that
glasses can serve as an occlusion and produce the ‘superman
effect’ (Ritchie & Kramer, 2016) meaning that a person, 1 3 1 3 Current Psychology (2023) 42:12424–12438 12435 intense than those without glasses. Eggleston et al. Discussion For example,
older adults could be invited to visit classrooms to read to
young children, or volunteer to tutor students if they have the
academic background (e.g., a retired math teacher may enjoy
tutoring struggling math students a few days per week).f As the world is suffering from COVID-19, older adults
are especially isolated and becoming increasing over-
whelmed as they cannot be in touch with their family;
therefore, this would be an ideal time to create a virtual
Adopt a Grandparent program. Once it is safe again to
interact with older adults, then the meeting between the
student and older adult could continue both in person as
well as online. Perhaps a coordinator could match students
and older adults’ interests in order for their connection to
have more meaning (e.g., a retired police officer, nurse,
computer programmer; or interests such as playing chess,
painting, video-gaming). We encourage researchers from multiple disciplines to
consider collaborating on face-recognition studies relating
to SAs. For example, researchers from disciplines such as
psychology, criminology, computer science, and graphic
design could combine their skills to create SAs. Because
there are so many venues for SAs (TV, online newspapers,
social media, cellphone alerts, etc.), it could be beneficial to
develop collaborative multidisciplinary programs to create
SAs for different media outlets. One limitation to the present study is that we only
included one Black couple in their mid-80s. We picked that
particular age range as most SA for both all races tend to be
in their 80s; however, having multiple target ages could help
us better understand what older adult ages are easier to rec-
ognize in a SA. Including individuals in the SAs of only one
ethnicity is also a limitation. It is also important to consider
that the majority of the participants in the study identified
as White. This could have affected recognition of the Black
couple as the ORE would predict relatively low recognition
of Black individuals by White participants. However, the
results did not indicate a significant difference in recognition
according to participant ethnicity. The increasingly diverse
population in the U.S. has brought attention to the value of
all elders’ well-being and safety. All couples of any race or
ethnicity may go missing, including multiracial couples, and
couples who have migrated from other countries. Discussion Although our results did not provide support for the
hypothesis that higher levels of contact with elderly peo-
ple were associated with increased recognition, we did find
some support for attitudes toward the elderly as a predictive
variable for recognition. Burnes et al. (2019) conducted a
meta-analysis on different interventions with older and
younger people in order to reduce ageism. The research-
ers concluded “Interventions are associated with substantial
reduction in ageism and should be part of an international
strategy to improve perceptions of older people and the
aging process” (American Journal of Public Health, np). We
recommend that school systems consider incorporating pro-
grams designed to reduce ageism. Such programs could be f
In terms of individual’s difference variables, we had
hypothesized that attitudes toward the elderly, measured by
ATOA scores, would predict greater likelihood of recog-
nizing a missing senior citizen. Our results indicated that
ATOA scores were predictive of recognition but only for
the female target without glasses. Note that this target photo 1 12436 Current Psychology (2023) 42:12424–12438 Hockley et al. (1999) found that, during the initial encod-
ing of a face and the target is wearing glasses, less informa-
tion may be available later during the recognition phase. However, during encoding, or the learning phase, if facial
features are emphasized, such as any unique facial features
other than the eyes, then participants may adapt to focusing
on the central part of the face, not just the eyes. Based on
Hockley et al. (1999) study, future mock SAs studies could
include directing half of the participants to focus on specific
areas of a targets face and no directions to the other half
of participants. For example, participants could focus on
whether the target has any unusual markings such as moles
or freckles, or a noticeable facial feature as full lips, dimpled
chin, or high forehead. integrated into clubs, honor societies as well as incorporated
into class service and volunteering projects. Discussion f
SA research has just begun to appear in journals, and with
the population of the elderly increasing exponentially, even
more research on how to help the public better recognize
older adult faces from Silver Alerts is necessary. With the
number of older adults over the age of 65 and older increas-
ing at a high rate, further research on SAs is imperative. It is
also important to note that Black people are at particularly
high risk of Alzheimer’s disease. Barnes et al. (2005) esti-
mated that Black people are about twice as likely as White
people to develop AD, although, once diagnosed, they may
experience slower cognitive decline (Alzheimer's disease
facts and figures, 2020). This pattern highlights the need
for research on recognition of elderly Black individuals. In
searching for examples of SA for a Black couple we were
surprised to find how few alerts there were for missing Black
couples. Research should be conducted to explore whether
alerts are less likely to be issued for Black people than for
other groups, especially in light of the risk of AD among
Black people. The present study provides an initial step in
extending the limited literature to recognition of an elderly
Black couple. Further, all of our participants were all from general
psychology classes or careers in psychology classes. Thus,
our sample was limited to college students taking particular
classes, which also resulted in limited age variability of par-
ticipants. The present study was also administered online. Although all of the research protocols were followed, and
participants were randomly assigned to conditions, the
results may differ if the study had been in-person, in the
physical presence of a researcher. In future research, it may be helpful to include an experi-
ence/contact scale specifically for Black seniors (or whatever
race is used in the SA study) in addition in addition to the
general contact/experience measures as we used in our study. Also, it would be important to explore the effects of different
delay periods between viewing the SA and encountering the
recognition photos, as there was a relatively short delay in
the present study. Supplementary Information The online version contains supplemen-
tary material available at https://doi.org/10.1007/s12144-021-02531-8. Data Availability Data is Available at: https://osf.io/e3hwd/?view_
only=a5184772978b4f1bb37cbc82715372bf Declarations Conflict of Interest We have no conflict of Interest to disclose. 3 3 References Gier, V. S., & Kreiner, D. S. (2020). Recognizing a missing senior
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Forgiveness or Striking Back? The Influence of Social Interaction on Adolescents' Conflict Coping Strategy
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International journal of educational psychology
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The Link Between Social Interaction with Adults and Adolescent
Conflict Coping Strategy in School Context Zhuojun Yao1 & Robert Enright1 Zhuojun Yao1 & Robert Enright1 Date of publication: February 24th, 2018
Edition period: February 2018 – June 2018 To cite this article: Yao, Z., & Enright, R. (2018). The link between social
interaction with adults and adolescent conflict coping strategy in school
context. International Journal of Educational Psychology, 7(1), 1-20. doi:
10.17583/ijep.2018.2872 j p
To link this article: http://dx.doi.org/10.17583/ijep.2018.2872 Instructions for authors, subscriptions and further details:
http://ijep.hipatiapress.com
The Link Between Social Interaction with Adults and Adolescent
Conflict Coping Strategy in School Context
Zhuojun Yao1 & Robert Enright1
1) University of Wisconsin-Madison, United States of America
Date of publication: February 24th, 2018
Edition period: February 2018 – June 2018
To cite this article: Yao, Z., & Enright, R. (2018). The link between social
interaction with adults and adolescent conflict coping strategy in school
context. International Journal of Educational Psychology, 7(1), 1-20. doi:
10.17583/ijep.2018.2872
To link this article: http://dx.doi.org/10.17583/ijep.2018.2872
PLEASE SCROLL DOWN FOR ARTICLE Instructions for authors, subscriptions and further details:
http://ijep.hipatiapress.com Instructions for authors, subscriptions and further details: PLEASE SCROLL DOWN FOR ARTICLE The terms and conditions of use are related to the Open Journal System and
to Creative Commons Attribution License (CC-BY). IJEP – International Journal of Educational Psychology, Vol. 7 No. 1
February 2018 pp. 1-20 IJEP – International Journal of Educational Psychology, Vol. 7 No. 1
February 2018 pp. 1-20 The Link between Social
Interaction with Adults and
Adolescent Conflict Coping
Strategy in School Context Zhuojun Yao, Robert Enright
University of Wisconsin-Madison Keywords: conflict coping strategy; school context; adolescence; social interaction. Abstract Based on social learning theory, this study aimed at providing a better understanding
of the influence of social interaction on adolescents' conflict coping strategy. This
study used the data from the Taiwan Educational Panel Survey (N=8717) to test the
unique contribution of religious involvement, parent-child interaction, teacher-
student interaction on adolescents' conflict coping strategy when they encountered
an interpersonal offense in school. Findings showed that religious involvement,
being physically hurt by father, being understood by father, and positive teacher-
student interaction could increase the possibility of positive conflict coping strategy. And being verbally hurt by mother and negative teacher-student interaction would
decrease the possibility of positive conflict coping strategy. Based on these results,
implications for research and practice were discussed. 2018 Hipatia Press
ISSN: 2014-3591
DOI: 10.17583/ijep.2018.2872 IJEP – International Journal of Educational Psychology, Vol. 6 No. 2
J
2017
97 119 IJEP – International Journal of Educational Psychology, Vol. 6 No. 2
June 2017 pp. 97-119 La Relación entre la
Interacción Social con Adultos
y la Estrategia de
Afrontamiento de Conflicto en
el Contexto Escolar Zhuojun Yao, Robert Enright
University of Wisconsin-Madison IJEP – International Journal of Educational Psychology, 7(1) IJEP – International Journal of Educational Psychology, 7(1) 3 3 h
(
o
M h
(
o
M any problems in living, both clinically severe and normal ones,
have their roots in or are exacerbated by interpersonal offenses
(Wade & Worthington, 2003). If the response to interpersonal
offense is expressed in forms of maladaptive coping, it can damage
relationships and lead to avoidance and revenge (Worthington & Scherer,
2004). Especially for now, youth violence is an increasing concern in our
school and society. A significant factor in the involvement of violence is the
inability to resolve conflict. Many adolescents who have shown violent
behavior say that they are motivated by anger and revenge (Pfefferbaum &
Wood, 1994). These situations might be improved if positive conflict coping
strategy is involved. Although there are many positive coping strategies to
deal with interpersonal offense, forgiving is an effective coping response
which can diminish negative emotions and repair relationships, it is
positively associated with conflict resolution, advice and support seeking
strategies, and negatively associated with revenge seeking (Ahmed &
Braithwaite, 2006; Worthington, Berry, & Parrott, 2001; Worthington &
Wade, 1999). M Resumen Sobre la base del aprendizaje social, este estudio se dirigió a conseguir una mejor
comprensión sobre la influencia de la interacción social sobre la estrategia de
adolescentes relativa al afrontamiento del conflicto. Este estudio usó los datos de la
Taiwan Educational Panel Survey (N=8717) para medir la contribución de la
implicación religiosa, la interacción padre-hijo, la interacción profesor/a-estudiante
sobre la estrategia de adolescentes relativa al afrontamiento del conflicto cuando se
encontraban con una ofensa interpersonal en la escuela. Los resultados muestran que
la implicación religiosa, ser dañado por el padre, ser comprendido por el padre, y
una interacción positiva profesor/a-estudiante podría aumentar la posibilidad de una
estrategia de afrontamiento del conflicto positiva. Y ser dañado verbalmente por la
madre y una interacción negativa profesor/a-estudiante disminuiría la posibilidad de
una estrategia de afrontamiento positiva. Basado en estos resultados, se discuten
implicaciones para la investigación y la práctica. Palabras clave: estrategia de afrontamiento del conflicto; contexto escolar;
adolescencia; interacción social. 2017 Hipatia Press
ISSN: 2014-3591
DOI: 10.17583/ijep.2017.2584 Forgiving as a positive conflict coping strategy In literature, Worthington et al (2007) suggest that there are two types of
forgiveness: decisional forgiveness and emotional forgiveness. Emotional
forgiveness will lead to decisional forgiveness, and decisional forgiveness
may influence emotional forgiveness (DiBlasio & Benda, 2008). Emotional
forgiveness is the emotional replacement of negative unforgiving emotions
(like bitterness, resentment, and anger) by positive other-oriented emotions
such as empathy, sympathy, compassion, or love. Decisional forgiveness is a
behavioral intention to eliminate negative behavior and increase positive
behavior toward the transgressor. People who grant decisional forgiveness
are inclined neither to seek revenge nor avoiding the transgressor but treat
the person well even though they might not have completely forgiven the
person emotionally (Worthington, Jennings, David, & Diblasio, 2010). Decisional and emotional forgiveness are different processes. Emotional
forgiveness is more conducive to mental health because negative affect and
stress reactions can be overcome by positive affect. Decisional forgiveness is Yao & Enright – Interaction and Forgiveness 4 a motivation statement about controlling one’s future behavior which might
improve reconciliatory processes and relationships (Worthington, Witvliet,
Pietrini, & Miller, 2007). Therefore, decisional forgiveness is more suitable
to be used as coping response when people encounter interpersonal offense,
it is a motivational transformation that inclines people to inhibit relationship-
destructive responses and behave constructively toward someone who has
behaved destructively toward them (McCullough, Worthington, & Rachal,
1997; Rusbult et al, 1991). ;
,
)
Peer world is important for adolescents and the subtlety of social
interaction grows exponentially in this period (Denham et al., 2002), such
complex peer interactions are complemented by increased social cognitive
ability and forgiveness reasoning (Crick & Dodge, 1994; Enright, 1994). However, age may be related to increasingly abstract reasoning about
conflict resolution, but it is not a strong predictor of it: many adolescents at
this period still act in ways that lead to social rejection and relational
aggression (Crick & Grotpeter, 1995; Park & Enright, 1997; Worthington,
2007). The Influence of Social Interaction on forgiving strategy Noddings (2002) emphasized the need to develop and maintain an
environment in which moral life could flourish, “How I treat you may bring
out the best or worst in you.” Exposure to more mature others will stimulate
maturity in adolescents’ own value processes, and these virtues will
eventually be internalized and become a part in adolescents (Windmiller,
Lambert, & Turiel, 1980). On the other hand, positive social interaction is
associated with adolescents’ positive personality and emotion. The nurturing
adult caregiver provides a model of concern for others, in the interaction
with them, children will develop the capacity for empathy (Windmiller,
Lambert, & Turiel, 1980). Youth who have positive personality traits such as
agreeableness and emotional stability may treat the transgression less severe
or less intentional, as well as interpret apologies as more sincere, thus they
can forgive the transgressor more easily (Boon & Sulsky, 1997;
McCullough, 2001). learning theory, forgiveness as a moral virtue can be initiated through
observation and imitation, children observe the behavior of parents over time
and imitate what they see. Bandura and Walters (1963) stated that children
from homes where heavy punishment predominated tended to believe the
effectiveness of punishment and retributive justice. In addition to the
observation of behavior, learning also occurs through the observation of
rewards and punishments, a process known as vicarious reinforcement. Children can be induced to say that he or she grants forgiveness at a very
early age, because they tend to think that forgiveness will help them avoid
punishment and get rewards (Park & Enright, 1997; Enright & Fitzgibbons,
2000; Worthington, 2006). Adolescents become more social and capable of
realizing social disapproval and approval for their responses to
transgressions, forgiveness may happen under the conditions of social
pressure, such as demands or suggestions from peers, family, or certain
institutions that encouraging a forgiving response (Park & Enright, 1997). Noddings (2002) emphasized the need to develop and maintain an
environment in which moral life could flourish, “How I treat you may bring
out the best or worst in you.” Exposure to more mature others will stimulate
maturity in adolescents’ own value processes, and these virtues will
eventually be internalized and become a part in adolescents (Windmiller,
Lambert, & Turiel, 1980). On the other hand, positive social interaction is
associated with adolescents’ positive personality and emotion. The Influence of Social Interaction on forgiving strategy Forgiveness is a concept with deep religious roots. Religious traditions,
beliefs, and rituals can influence people’s interpersonal thoughts, feelings,
and behaviors. Forgiveness (decisional or emotional) in response to a
transgression, is valued by every major religion such as Christianity,
Judaism, and Islam, and they firmly advocate forgiveness as a way of
controlling one’s negative emotion and behavior (Worthington, Jennings, &
DiBlasio, 2010). If religions emphasize the value of forgiveness, it would be
unsurprising to find that people higher in religious involvement tend to be
more forgiving than people lower in religious involvement. However,
empirical research suggests that religious involvement related to forgiveness
at a general, abstract level, but is not as strongly related to forgiveness in
specific, real-life circumstances (McCullough & Worthington, 1999). In this
study, we measured adolescents’ response to a specific circumstance, but we
are not sure whether there is a strong relationship between religious
involvement and forgiving response to peer transgression. Besides religious interaction, significant others also remain mostly
responsible for the moral development of adolescents. According to social IJEP – International Journal of Educational Psychology, 7(1) 5 learning theory, forgiveness as a moral virtue can be initiated through
observation and imitation, children observe the behavior of parents over time
and imitate what they see. Bandura and Walters (1963) stated that children
from homes where heavy punishment predominated tended to believe the
effectiveness of punishment and retributive justice. In addition to the
observation of behavior, learning also occurs through the observation of
rewards and punishments, a process known as vicarious reinforcement. Children can be induced to say that he or she grants forgiveness at a very
early age, because they tend to think that forgiveness will help them avoid
punishment and get rewards (Park & Enright, 1997; Enright & Fitzgibbons,
2000; Worthington, 2006). Adolescents become more social and capable of
realizing social disapproval and approval for their responses to
transgressions, forgiveness may happen under the conditions of social
pressure, such as demands or suggestions from peers, family, or certain
institutions that encouraging a forgiving response (Park & Enright, 1997). The Influence of Social Interaction on forgiving strategy The nurturing
adult caregiver provides a model of concern for others, in the interaction
with them, children will develop the capacity for empathy (Windmiller,
Lambert, & Turiel, 1980). Youth who have positive personality traits such as
agreeableness and emotional stability may treat the transgression less severe
or less intentional, as well as interpret apologies as more sincere, thus they
can forgive the transgressor more easily (Boon & Sulsky, 1997;
McCullough, 2001). For the role of parent-child interaction on forgiveness, Worthington
(2007) demonstrated that parental socialization and emotional climate within
the parent-child relationship could affect adolescents’ temperament and
emotion-regulation capability (e.g. empathy, lessened anger and shame,
appropriate guilt), which might influence forgiveness reasoning, motivation,
and behavior. Mincic et al. (2004) also claimed that children’s perceptions of
positive childrearing practices were related to children’s forgiveness. However, parents are not the sole models for the child. Durkheim (2012) Yao & Enright – Interaction and Forgiveness 6 suggested that social unit of family was not constituted to be the agent to
impart what was important to the culture, family was too small and too
personal to reflect the whole of the social system, and school had the
function of linking the child to this society, and teachers’ role is extreme
important in creating a social and moral being. In Taiwan and also many
Asian societies, when children enter into senior high school, school life
becomes more important. Most students immerse themselves in school at
least eight hours a day, so teachers spend the longest time with students even
compared with their parents. A teacher's classroom behavior is constantly
under scrutiny by students. As a result, students learn a lot from a teacher's
nonverbal behavior as well as their verbal behavior (Galloway, 1976). Children who perceive greater levels of support from teachers have fewer
behavioural problems, higher levels of social competency, and better school
adjustment (Birch & Ladd, 1997; Pianta &Steinberg, 1992; Khamis, 2009). Besides, teachers as adult model in the school context can directly and
indirectly tell and show students what emotions and behaviors are acceptable
to express. To date there is no research investigating the relationship
between teacher-student interaction and adolescents' forgiving coping
strategy but based on the social support theory and related literature about
parent-child interaction, it is reasonable to assume that teacher-student
interaction is one influencing factor. The Influence of Social Interaction on forgiving strategy We should keep in mind that school is a
complex system different from family context, where students encounter
more than one teacher, so rather than measuring the degree or frequency of a
specific teacher’s behaviour, the present study measured the number of
teachers which students perceive with positive or negative interaction. IJEP – International Journal of Educational Psychology, 7(1) JEP – International Journal of Educational Psychology, 7(1) 7 This research added to the literature on adolescent conflict coping strategy
by assessing the influence of social interaction, especially teacher-student
interaction, which has not been widely examined in previous research. This research added to the literature on adolescent conflict coping strategy
by assessing the influence of social interaction, especially teacher-student
interaction, which has not been widely examined in previous research. Participants
hi
d p
This study made use of data from Taiwan Educational Panel Survey (Chang,
2008) for Wave 4 (2007) Senior High School. Taiwan Education Panel
Survey is a national longitudinal project initiated by Academia Sinica and
jointly funded by the Ministry of Education, the National Science Council,
and Academia Sinica. A multistage stratified sampling method was used,
and three classes from each school were sampled, with 15 students selected
at random in each class. The dataset contained a sample of 12th graders in
senior high school. The sample size in this study was 8717; the number of
male was 4468 (51.3%), and the number of female was 4249 (48.7%). The
full data set of students eligible for inclusion in the study contained 4.6
percent missing data, finally 8320 participants were included in this study. The Current Study Building on previous research, we examined the extent to which religious
involvement, parent-child interaction and teacher-student interaction
influenced high-school students’ conflict coping strategy when individual
factors (Gender and Angry) were controlled. We expected to find that
religious involvement and positive interaction with parents and teachers
were positively associated with adolescents’ positive conflict coping
strategy, whereas, negative interaction with parents and teachers are
negatively associated with adolescents’ positive conflict coping strategy. Measures Gender
Gender was coded 1 for males, 0 for females. Religious Involvement Religious Involvement Respondents’ religious involvement was measured by two items, they were
asked: “whether you have religious affiliation” and “whether you attended
religious service activities last summer vacation”. Each question allowed a
(0, 1) response.0 means No, and 1 means Yes. Yao & Enright – Interaction and Forgiveness Yao & Enright – Interaction and Forgiveness 8 Parent-child interaction Participants were asked to indicate whether their father or mother did any of
the following things: (1) listen to my idea and communicate with me
patiently; (2) give me help when I encounter some big problems or
difficulties; (3) criticize me heavily when I make a small mistake; (4) beat
me when I make a small mistake. Each question allowed a (0, 1) response. The first two questions measured positive interaction with parents, and the
last two questions measured negative interaction with parents. Teacher-student interaction Angry g y
Angry as the control variable was measured by 2 items. Participants were
asked: “In this semester, did these following things happen to you?” Items
included, “want to scream, fight, and quarrel”, “feel irritated”. These 2 items
assessed the self- reported frequency of angry experienced over the semester
on a four-point Likert scale: never (assigned 1), sometimes (assigned 2),
frequently (assigned 3), very frequently (assigned 4). The scores for each
item were averaged to form a new variable called angry. The higher the
scores were, the more severe the angry was considered to be. Cronbach's
alpha was .63. Teacher-student interaction There were four items self-assessed by asking how many teachers behaved
in a certain manner since the student started senior high school. Two of these
items were averaged to measure positive interaction with teachers
(Cronbach's alpha was .73): “When I express my ideas, the teacher will
listen and try to understand.” and “The teacher will praise me when I work
hard.” Another two items were averaged to measure negative interaction
with teachers (Cronbach's alpha was .61): “The teacher may hurt my self-
esteem when he or she tutors me.”, and “The teacher sometime may punish
us physically when we make a mistake.” Each item was measured on a five-
point Likert scale: none (assigned 1), 1~2 (assigned 2), 3~4 (assigned 3),
5~6 (assigned 4), more than 6 (assigned 5). Higher scores on the measure
were indicative of more positive or negative interaction with teachers. Positive conflict coping strategy as the dependent variable was measured by
1 item. Participants were asked: “If one of your classmates treats you badly,
what will you do?” Options of this item included, “A: Treat him or her
better”, “B: Treat him or her the same as before”, “C: Ignore and avoid him
or her,” “D: Treat him or her badly”. Participants who chose A and B
showing forgiveness toward transgressor were considered as positive IJEP – International Journal of Educational Psychology, 7(1 9 conflict coping strategy and scored as 1. In contrast, participants who choose
C and D deemed to be negative conflict coping strategy and were coded as 0. Results Descriptive statistics showed that the number of adolescents who involved in
religion activity were less than the number of those who didn’t, χ²
(1,8605)=5706.72,
p<.001. For the
parent-child
interaction,
more
adolescents experienced verbal hurt rather than physical hurt from parents, χ²
(1,8594)=831.85, p<.001, and more adolescents experienced positive
interactions with mothers than fathers, χ² (1, 8695) =467.45, p<.001. In
terms of conflict coping strategy, there were fewer students who had the
intention to behave in a forgiving way when they are offended by a
classmate, χ² (1, 8716) =338.13, p<.001. Descriptive statistics of the
continuous variable showed that adolescents perceived more positive
(M=2.28, SD=0.9) rather negative (M=1.42, SD=0.58) interaction with
teachers, t0.05(8544) =75.9, p<.001. Table 1 display the regression coefficients, standard errors, odds ratio, and
model statistics for three models. The baseline model (model 1) consisted of
the control variables: gender and angry. And according to the previous
research, religious and parental variables have an influence on adolescents'
conflict coping strategy, but the influence of teacher-student interaction has
not been tested. In addition, the influence of religious and parental variables
happened before the influence of teacher variables on students, so we put
them in model 2 simultaneously，and teacher-student interaction was added
in model 3. Results for Model 1 revealed that female (B=-0.23, SE=0.05,
OR=0.8, p<0.01) and adolescents with lower level of angry (B=-0.5,
SE=0.03, OR=0.61, p<0.01) tended to show forgiveness to the offender. In
Model 2, Likelihood ratio was decreased, Nagelkerke R2 and Correct
predicted percentage were all increased, which mean the regression equation
in Model 2 was better than Model 1. The statistically significant and positive
coefficients for Religion (B=0.23, SE=0.05, OR=1.25, p<0.01) and
Religious service activity (B=0.62, SE=0.08, OR=1.86, p<0.01) supported
the notion that religious interaction had a positive effect on adolescents’
forgiving response to peer transgression. For the role of parent-child
interaction, there are three noteworthy results. First, contrary to our
hypothesis that negative parent-child interaction was related to negative Yao & Enright – Interaction and Forgiveness 10 coping strategy, we found that adolescents who experienced physically hurt
by fathers were more inclined to be forgivable (B=0.57, SE=0.12, OR=1.77,
p<0.01). Second, verbal hurt from mother (B=-0.13, SE=0.06, OR=0.88,
p<0.05) but not from father (B=-0.1, SE=0.07, OR=0.9, p>0.05) decreased
the probability of forgiveness. Results Last, only positive interaction with father
(Understood: B=0.15 SE=0.06, OR=1.16, p<0.01; Helped: B=0.12, SE=0.06,
OR=1.13, p<0.05) not mother (Understood: B=0.08 SE=0.06, OR=1.08,
p>0.05; Helped: B=0.08, SE=0.06, OR=1.09, p>0.05) increased the
probability of positive conflict coping strategy toward the offender. However, when parent-child interactions and teacher-student interactions
were jointly examined in Model 3, the effect of “Helped by father” was not
significant (B=0.08, SE=0.06, OR=1.08, p>0.05), and both positive and
negative interaction with teachers emerged statistically significant (Positive:
B=0.38 SE=0.03, OR=1.46, p<0.01; Negative: B=-0.14, SE=0.04, OR=0.87,
p<0.01). According to Model statistics, the regression equation of Model 3
(Likelihood ratio=10888.80, Nagelkerke R2=0.098, Correct predicted
percentage=61) could better predict the intention of forgiveness than Model
1 and Model 2, and Hosmer-Lemeshow test (Chi-square=6.40, p=0.60)
showed Model 3 had goodness of fit. Good model fit would be evidence in
support of our hypothesis that teacher-student interaction played an
important role in adolescents' positive conflict coping strategy. IJEP – International Journal of Educational Psychology, 7(1) Yao & Enright – Interaction and Forgiveness
12 12 IJEP – International Journal of Educational Psychology, 7(1) 11 Table 1. Summary of Logistic Regression Analysis for Variables Predicting Positive Conflict Coping Strategy. (N=8320)
Note *p < 05 **p < 01 is for Variables Predicting Positive Conflict Coping Strategy. (N=8320) Yao & Enright – Interaction and Forgiveness
12 Conclusions and Discussion The present work provided a preliminary theoretical and empirical basis for
the influencing factors of conflict coping strategy in adolescents. In this
paper, forgiving is viewed as a positive conflict coping strategy: When
people forgive, they inhibit their normal destructive responses that would
increase the probability of further relational disintegration and instead
become more likely to enact constructive responses that would help to
restore the damaged relationship to health (McCullough, Worthington, &
Rachal, 1997; Rusbult et al, 1991). Forgiveness is a deeply religious concept
for people from many faiths and cultures, issues of guilt, reconciliation,
salvation, and redemption to many religions directly or indirectly question
about forgiveness and its place in the life of individuals and communities
(Paloutzian & Park, 2014). The result of this study showed that religious
involvement was positively related to adolescents’ forgiving response to
peer transgression. This supported a large body of work showing that
religion was positively related to forgivingness (for a meta-analytic review,
see Davis et al, 2013). As Toussaint and Jorgensen (2008) pointed out,
whether in formal religious services, prayer groups, or other venues,
adolescents have many opportunities to study and learn about the reason and
benefits of forgiveness, in the long run this can help shaping the forgiving
personality. In terms of relationship between parent-student interaction and forgiving
response toward transgressors, our results revealed that adolescents who
experienced physically hurt from father would be more forgiving, and this is
inconsistent with our hypothesis that negative parent-student interaction is
negatively associated with positive coping strategy. One possible
explanation for this is that the negative experience such as being threatened,
criticized, and punished excessively by adults could lead to a strict superego,
which will make adolescents feel guilty. Guilt-prone individuals adopt more
proactive and constructive strategies for managing anger, they are more
likely to engage in constructive behaviors, such as non-hostile discussion
with the target of their anger (Baumeister, Stillwell, & Heatherton, 1994). The second explanation is adolescents who often experienced physically hurt
from fathers may tend to be afraid of the conflict with others, they are
inclined to maintain a relationship rather than destroy it. IJEP – International Journal of Educational Psychology, 7(1) IJEP – International Journal of Educational Psychology, 7(1) 13 Another question raised by negative parent-child interaction is why only
physically hurt by father (but not mother) and verbally hurt by mother (but
not father) have an influence on adolescents' conflict coping strategy. Even
for now there is no empirical research investigating the difference between
the pattern of father and mother in punishment. A possible explanation
based on our experience is that the degree of physically hurt from father is
much more serious than mother, and the frequency of verbally hurt from
mother is higher than father. The future research can measure specific
components of maternal and paternal punishment and exam their influence
on adolescents’ outcome. Besides, we also found that only positive
interaction with father not mother can increase the probability of positive
conflict coping strategy. Consistent with our finding, previous research
found that mothers’ and fathers’ contributions are different on the function
of outcome (Williams & Kelly, 2005; Hastings, McShane, Parker, & Ladha,
2007). For the positive parent-child interaction, Day and Padilla-Walker
(2009) asserted that connectedness and involvement from fathers (but not
mothers) was negatively related to adolescents’ internalizing and
externalizing behaviors, whereas connectedness and involvement from
mothers (but not fathers) was positively related to adolescents’ prosocial
behaviors and hope. Therefore, we assumed that the real mechanism
underlying the association between positive conflict coping strategy and
positive father-child interaction was that perceived care from father could
significantly decrease the probability of negative conflict coping strategy
(e.g. revenge or striking back) rather than increase the probability of positive
conflict coping strategy. This assumption is still an open question and
definitely an area for future research. The specific hypothesis tested in the current research focused on the role
of teacher-student interaction. While less commonly addressed, the way that
teachers and students interact is a critical factor in determining the student’s
conflict resolution. Based on the result of binary logistic regression, teacher-
student interaction played a more significant role in adolescent's conflict
coping strategy than parent-child interaction. There are two possible
explanation for this result. On the first hand, Pianta (1999) concluded that
emotionally warm relationship between teachers and students (characterized
by open communication, support, and involvement) provided students with a
sense of security within school settings, which in turn promoted exploration
and comfort, as well as social and emotional competence. IJEP – International Journal of Educational Psychology, 7(1) We look forward to the possibility that the ideas IJEP – International Journal of Educational Psychology, 7(1 15 we have presented might help to build more efficient ways to increase
adolescents’ positive conflict coping strategy, and this will play a protective
role for interpersonal relationship in school and might well be extended to
community and society. IJEP – International Journal of Educational Psychology, 7(1) Adolescents with Yao & Enright – Interaction and Forgiveness 14 higher social and emotional competence had a greater willingness to forgive,
because they could replace negative emotions with positive emotions such as
emotional empathy (Fincham, Paleari, & Regalia, 2002; McCullough et al.,
1998; Fincham et al., 2002) and had more rumination about the transgression
(McCullough et al, 1998). On the other hand, a teacher who acts as a
desirable social model can facilitate student social development and
students’ acquisition of appropriate social skills (Kahn & Cangemi, 1979). Teachers can make a sizeable difference in the social lives of students by
serving as either positive or negative behavioral models (Englehart, 2009). Children learn basic conversational rules and conflict resolution in school
context from seeing teachers as the adult models. Teachers who fail to
manage their emotions and externalize anger and frustration in public
display are not only missing an opportunity to model a socially desirable
response but are also modeling an undesirable response in its place. When
teachers respond to conflict in a calm, rational manner, respectfully
acknowledge disagreement, and welcome multiple points of view, these
behaviors can “rub off” on students, making them more able to confront the
complexities of human relationships (Englehart, 2009). A clear limitation of the current work was it measured conflict coping
strategy on imagined scenario, so the dependent variable in present paper is
intention rather than actual behavior. An extension of this work would
involve collecting data from adolescents who have experienced a
classmate’s offense, and how they deal with this situation. Maybe the actual
offense is harder to forgive than imaginary one. Another limitation was that
parent-child interaction and teacher-student interaction are culturally
embedded. A useful next step for research would be determine whether
adolescents in other cultures share the same patterns in Taiwanese samples. Contrasting more family oriented or collectivistic cultures would be
informative. The last limitation was that, in order to match up the items in
TEPS, we only exam the influence of social interaction from adults on
adolescents' conflict coping strategy, future research should examine
whether social interaction from peers or siblings may influence adolescents’
conflict coping strategy In conclusion, positive conflict coping strategy not only diminishes
victim’s motivation to seek revenge and maintain estrangement from an
offending relationship partner, but also increase their motivation to pursue
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psychology, 35(1), 69-79. doi: 10.1037//0012-1649.35.1.69. Rusbult, C. E., Verette, J., Whitney, G. A., Slovik, L. F., & Lipkus, I. (1991). Accommodation processes in close relationships: Theory and
preliminary empirical evidence. Journal of Personality and Social
Psychology, 60(1), 53-78. doi: 10.1037//0022-3514.60.1.53. Toussaint, L., & Jorgensen, K. M. (2008). Inter-Parental Conflict, Parent-
Child Relationship Quality, and Adjustment in Christian Adolescents:
Forgiveness as a Mediating Variable. Yao & Enright – Interaction and Forgiveness Journal of Psychology &
Christianity, 27(4), 337-346. Wade, N. G., & Worthington Jr, E. L. (2003). Overcoming interpersonal
offenses: Is forgiveness the only way to deal with unforgiveness? Journal of Counseling and Development: JCD, 81(3), 343-353. doi:
10.1002/j.1556-6678.2003.tb00261.x. Williams, S. K., & Kelly, F. D. (2005). Relationships among involvement,
attachment, and behavioral problems in adolescence: Examining
father’s influence. Journal of Early Adolescence, 25, 168–196. doi:10.1177/0272431604274178. Windmiller, M., Lambert, N. M., & Turiel, E. (1980). Moral development
and socialization. Allyn & Bacon. Worthington Jr, E. L. (2006). Forgiveness and reconciliation: Theory and
application. Routledge. Worthington Jr, E. L. (2007). Handbook of forgiveness. Routledge. Worthington Jr, E. L., Berry, J. W., & Parrott III, L. (2001). Unforgiveness,
forgiveness, religion, and health. In T. G. Plante & A. C. Sherman IJEP – International Journal of Educational Psychology, 7(1) IJEP – International Journal of Educational Psychology, 7(1) 19 (Eds.), Faith and health: Psychological perspectives, 107-138. New
York: Guilford Press. Worthington Jr, E. L., Jennings, I. I., David, J., & DiBlasio, F. A. (2010). Interventions to promote forgiveness in couple and family context:
conceptualization, review, and analysis. Journal of Psychology &
Theology, 38(4), 231-245. Worthington, E. L., & Scherer, M. (2004). Forgiveness is an emotion-
focused coping strategy that can reduce health risks and promote health
resilience: Theory, review, and hypotheses. Psychology &
Health, 19(3), 385-405. doi: 10.1080/0887044042000196674. Worthington Jr, E. L., & Wade, N. G. (1999). The psychology of
unforgiveness and forgiveness and implications for clinical
practice. Journal of Social and clinical psychology, 18(4), 385-418. doi:
10.1521/jscp.1999.18.4.385. Worthington Jr, E. L., Witvliet, C. V. O., Pietrini, P., & Miller, A. J. (2007). Forgiveness, health, and well-being: A review of evidence for
emotional versus decisional forgiveness, dispositional forgivingness,
and reduced unforgiveness. Journal of behavioral medicine, 30(4), 291-
302. doi: 10.1007/s10865-007-9105-8. Yao & Enright – Interaction and Forgiveness 20 Zhuojun YAO is a PhD student in the Department of Educational
Psychology, University of Wisconsin-Madison, USA. Robert ENRIGHT is Professor in the Department of Educational
Psychology, University of Wisconsin-Madison, USA. Contact Address: Department of Educational Psychology, University of
Wisconsin-Madison, 1025 W Johnson St, Madison, WI 53706, EE. UU. Email: zyao37@wisc.edu Robert ENRIGHT is Professor in the Department of Educational
Psychology, University of Wisconsin-Madison, USA.
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European Medicines Agency Policy 0070: an exploratory review of data utility in clinical study reports for academic research
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Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204
https://doi.org/10.1186/s12874-019-0836-3 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204
https://doi.org/10.1186/s12874-019-0836-3 Open Access European Medicines Agency Policy 0070:
an exploratory review of data utility in
clinical study reports for academic research Jean-Marc Ferran1† and Sarah J. Nevitt2*† Abstract Background: Clinical study reports (CSRs) have been increasingly utilised within academic research in recent years. European Medicines Agency (EMA) Policy 0070 ‘Phase 1,’ which came into effect in January 2015, requires the
publication of regulatory documents such as CSRs from central applications in an anonymised format. EMA Policy
0070 requires sponsors to demonstrate careful consideration of data utility within anonymised CSRs published
within the scope of the policy, yet the concept of data utility is not clearly defined in the associated anonymisation
guidance. Objective: To review the use of data from CSRs in published academic research and to hypothesise the potential
data utility of CSRs anonymised under the objectives of EMA Policy 0070 for future academic research. Methods: Review of the objectives, research methodologies and findings of academic research reports using
unpublished data from CSRs (prior to EMA Policy 0070). Semi-structured interviews with authors of academic
research reports, including questions related to data utility of anonymised CSRs published under EMA Policy 0070. Results: Thirteen academic research reports were identified and reviewed. The research purposes ranged from
assessment of reporting bias, comparison of methods and results with published data sources, detailed evaluation
of harms and adverse events, re-analysis and novel analyses including systematic reviews and meta-analysis. All of
the examples identified required access to the methods and results sections of CSRs (including aggregated
summary tables) and research purposes relating to evaluation of adverse events also required access to participant
narratives. Retaining anonymised participant narratives relating to interventions, findings and events, while
maintaining an acceptably low risk of participant re-identification, may provide an important gain in data utility and
further understanding of drug safety profiles. Conclusions: This work provides an initial insight into the previous use of CSR data and current practices for
including regulatory data in academic research. This work also provides early guidance to qualitatively assess and
document data utility within anonymised CSRs published under EMA Policy 0070. Keywords: Policy 0070, Regulatory, Anonymised, Clinical study reports, Data utility, Review * Correspondence: sjn16@liverpool.ac.uk Correspondence: sjn16@liverpool.ac.uk
†Jean-Marc Ferran and Sarah J. Nevitt contributed equally to this work. 2Department of Biostatistics, University of Liverpool, Block F, Waterhouse
Building, 1-5 Brownlow Street, Liverpool L69 3GL, UK
Full list of author information is available at the end of the article Correspondence: sjn16@liverpool.ac.uk
†Jean-Marc Ferran and Sarah J. Nevitt contributed equally to this work.
2Department of Biostatistics, University of Liverpool, Block F, Waterhouse
Building, 1-5 Brownlow Street, Liverpool L69 3GL, UK
Full list of author information is available at the end of the article Background The Organization for Economic Co-operation and Devel-
opment (OECD) propose the following definition for ‘Data
Utility’: “A summary term describing the value of a given
data release as an analytical resource. This comprises the
data’s analytical completeness and its analytical validity. Disclosure control methods usually have an adverse effect
on data utility. Ideally, the goal of any disclosure control re-
gime should be to maximise data utility whilst minimising
disclosure risk. In practice disclosure control decisions are a
trade-off between utility and disclosure risk [18].” Clinical Study Reports (CSRs) represent a wealth of
information related to design, conduct and analysis of clin-
ical trials, in addition to more comprehensive trial results
compared to publicly available sources such as journal
manuscripts and clinical trial registries. Doshi et al. 2013
[1] refer to CSRs as a “hitherto mostly hidden and untapped
source of detailed and exhaustive data on each trial” and
make reference to the concept of a “compression factor,”
defined as the ratio of CSR page length compared to the
page length of the journal publication of the same trial, ran-
ging from 1 to 8805 based on the review of 78 CSRs [1]. Clearly, the ‘analytical completeness and analytic valid-
ity’ of anonymised CSR data, and therefore the relative
‘data utility’ of anonymised CSRs made public under EMA
Policy 0070 are directly dependent on the consumers of
the data and their reasons or purposes for using this data. g g
Previous work has highlighted the impact of selective
outcome reporting [2, 3], in that the data and results
published within a journal manuscript may be incom-
plete or misleading, and the biases that originate from
this selective reporting. Increasingly, researchers under-
taking novel secondary analyses of clinical trial data such
as systematic reviews and meta-analyses are seeking ac-
cess to previously confidential regulatory documents,
such as CSRs. These documents are used as a means of
appraising, revaluating and reducing the impact of any
misreporting or selective reporting bias, generating more
complete and reliable information and investigating clin-
ical questions which could not have previously been
considered using published data sources alone [4–16]. Therefore the objective of this work is to review the
previous use of data from CSRs in academic research
and with these research purposes in mind, to hypothe-
sise the potential data utility of CSRs anonymised under
the objectives of EMA Policy 0070. © The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Page 2 of 10 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Methods This paper begins with a short reflection of requests
made under EMA Policy 0043, a policy which enables
requests for access to regulatory documents (related to
medicinal products for human and veterinary use),
followed by a summary and discussion of previous re-
search conducted using CSR data [4–16]. The European Medicines Agency (EMA) policy on the
publication of clinical data for medicinal products for hu-
man use (EMA Policy 0070 ‘Phase 1’, herein referred to as
EMA Policy 0070), allowing global access to regulatory
documents for non-commercial purposes, came into effect
in January 2015. The policy requires ‘clinical reports’ (de-
fined as clinical overviews, clinical summaries, and CSRs
together with the following appendices to the CSRs: proto-
col and protocol amendments, sample case report form
and documentation of statistical methods (clinical trial
statistical analysis plan (SAP)) from central regulatory ap-
plications to be published in an anonymised portable
document format, such that the risk of re-identification of
trial participants from the information available contained
within these documents, including any retained narrative
information relating to individual participants, is deemed
to be acceptably low for public disclosure of data [17]. Insights from EMA policy 0043 An EMA Policy on access to documents (related to medi-
cinal products for human and veterinary use), known as
‘EMA Policy 0043’, came into effect in 2010. Requesting
documents through EMA Policy 0043 is a controlled
process and requesters must identify themselves and their
affiliation but it are not required to document the purpose
of their request as part of the requesting process. Figure 1
summarises the number of requests and number of pages
released per affiliation and was made public in 2016 [19]. Source: Annexes to the annual report of the European
Medicines Agency 2016, Annex 19 [19]. 1. ‘Legal’ and ‘Consultant’ requesters refers to mostly
professionals from or contracted by the
pharmaceutical industry. The category
“Pharmaceutical industry” both includes companies
from the innovative and generic industry; no
distinction was made. The concept of ‘Data Utility’ appears to be an important
criterion for anonymised CSRs to meet the objectives of
EMA Policy 0070. Reference is made within several sections
of EMA Policy 0070 anonymisation external guidance and
within the ‘Anonymisation Report’ template that the appli-
cant must demonstrate careful consideration of “the impact
of the anonymisation methodology used on data utility
[17].” However, despite the apparent importance of this
criterion, the EMA Policy 0070 external guidance [17] does
not define or quantify the utility of anonymised CSRs (and
“Data Utility” is absent within section “3. Definitions”) [17]. The concept of ‘Data Utility’ appears to be an important
criterion for anonymised CSRs to meet the objectives of
EMA Policy 0070. Reference is made within several sections
of EMA Policy 0070 anonymisation external guidance and
within the ‘Anonymisation Report’ template that the appli-
cant must demonstrate careful consideration of “the impact
of the anonymisation methodology used on data utility
[17].” However, despite the apparent importance of this
criterion, the EMA Policy 0070 external guidance [17] does
not define or quantify the utility of anonymised CSRs (and
“Data Utility” is absent within section “3. Definitions”) [17]. 2. The request process of EMA Policy 0043 includes
an appeal process in the case of an initial rejection It should be noted that the scope of EMA Policy 0043 is
not limited to requests for clinical reports (i.e. CSRs and
other documents related to clinical trial conduct and results Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Page 3 of 10 Fig. Insights from EMA policy 0043 1 Policy 0043 documents requests per affiliation each consumer group via EMA Policy 0043 is merely an
assumption. Nonetheless,
the
affiliations
described
within Fig. 1 provide an indication of who the main data
consumers of EMA Policy 0070 may be and further
research on EMA Policy 0043 requests, particularly re-
lating to the types of documents requested by each data
consumer group, could help in understanding the likely
consumers of EMA Policy 0070 and their purposes. (such as clinical trial protocols and SAPs) and a range of
other documents such as commercial, technical or legal
documents as well as meeting minutes could also be re-
quested. A breakdown of the information within Fig. 1 re-
lating to the type of documents requested was not available. Grouping together the first seven rows to represent
‘Non-commercial’ requests from “Not-for-profit organ-
isation” to “Academia/Research institute”, the number of
requests received represent 18.6% of total requests and
‘Commercial’ requests from ‘Legal’, ‘Pharmaceutical In-
dustry’ and ‘Consultant’ represents 76% of total requests. However, when comparing the number of pages re-
leased, the ‘Non-Commercial’ and ‘Commercial’ groups
are approximately equivalent with 52.3 and 45.6% re-
spectively. This shift is mainly explained by requests for
documents with a large number of pages (likely to be
clinical reports such as CSRs, trial protocols, trial SAPs)
by Academia/Research institutes (31.59%) while other
subgroups may be more likely to request other docu-
ments (such as technical documents or meeting mi-
nutes) that represent fewer pages. It is also of note that
requests from “Patients or Consumers” represent 6.68%
of all requests and 9.55% of pages released. Use of clinical study reports in academic research We note that direct extrapolation of these figures from
EMA Policy 0043 (a controlled process covering all types
of documents) to EMA Policy 0070 (public access to
clinical reports as part of a central application) is not
recommended and the type of documents requested by Page 4 of 10 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Table 1 Examples of the objectives of research using CSR data
Objective
CSR section(s) that may be required
Examples
Assessment of reporting and evaluation
of bias
Methods, results (aggregate summary
tables and text), narratives, participant
listings
Eyding et al [4], Schroll et al [9], Hodkinson et al [10],
Jefferson et al [11], Vedula et al [15]
Comparison of methods and/or results
(including adverse events) with data
registries or manuscripts
Methods, results (aggregate summary
tables and text)
Eyding et al [4], Le Noury et al [8], Schroll et al [9], Hodkinson
et al [10], Jefferson et al [11], Maund et al [12–14], Vedula
et al [15], Nevitt et al [16]
Detailed evaluation of harms and
adverse events
Results (aggregate summary tables
and text), narratives, participant listings
Eyding et al [4], Maund et al [12–14]
Systematic review and meta-analysis
(evidence synthesis)
Methods, results (aggregate summary
tables and text), narratives
Eyding et al [4], Jefferson et al [11],
Maund et al [14], Nevitt et al [16]
Re-analysis (repeating original analysis)
Methods, results (aggregate summary
tables and text), narratives, participant
listings
Le Noury et al [8] (also using individual participant data)a
Re-analysis (different method or
objective to the original analysis)
Methods, results (aggregate summary
tables and text), narratives, participant
listings
Maund et al [14], Nevitt et al [16]
aIn this example, Le Noury et al [8], re-analysis was conducted using both individual participant data (IPD), requested via data sharing platform
clinicalstudydatarequest.com [22] in addition to supporting information from CSRs and case report forms Eyding et al [4], Schroll et al [9], Hodkinson et al [10],
Jefferson et al [11], Vedula et al [15] Eyding et al [4], Le Noury et al [8], Schroll et al [9], Hodkinson
et al [10], Jefferson et al [11], Maund et al [12–14], Vedula
et al [15], Nevitt et al [16] Eyding et al [4], Le Noury et al [8], Schroll et al [9], Hodkinson
et al [10], Jefferson et al [11], Maund et al [12–14], Vedula
et al [15], Nevitt et al [16]
Eyding et al [4], Maund et al [12–14]
Eyding et al [4], Jefferson et al [11],
Maund et al [14], Nevitt et al [16]
Le Noury et al [8] (also using individual participant data)a Le Noury et al [8] (also using individual participant data)a Maund et al [14], Nevitt et al [16] Methods, results (aggregate summary
tables and text), narratives, participant
listings aIn this example, Le Noury et al [8], re-analysis was conducted using both individual participant data (IPD), requested via data sharing platform
clinicalstudydatarequest.com [22] in addition to supporting information from CSRs and case report forms aIn this example, Le Noury et al [8], re-analysis was conducted using both individual participant data (IPD), requested via data sharing platform
clinicalstudydatarequest.com [22] in addition to supporting information from CSRs and case report forms For example, by comparing the information in CSRs
and other regulatory documents (such as protocols) to
journal publications for 20 trials of Gabapentin, Vedula
et al [15] identified selective outcome reporting for trials
of off-label use of gabapentin which ‘threatens the valid-
ity of evidence for the effectiveness of off-label interven-
tions’. Use of clinical study reports in academic research Use of clinical study reports in academic research
A number of examples of journal publications using CSRs
in academic research [4, 8–11, 14, 15] were cited within a
conference presentation delivered by Tom Jefferson in 2017
[20] and within a related published report in 2018 [21]. This section summarises and discusses the research pur-
poses and methodology applied within these examples, in
addition to a recently published Cochrane Review involving
SJN which has made use of CSR data in analysis [16]. Some of the research objectives from these examples,
outlined in general terms including the sections of the
CSRs that may be required for the research, are sum-
marised in Table 1. We further discuss some of the re-
search objectives outlined within the examples within this
section. Further specific details of the all regulatory data
sources provided for the research, the methods employed
within each example and the main findings of the research
are summarised in Additional file 1: Table S1. Use of clinical study reports in academic research Eyding et al [4] also discovered that published
data overestimated the benefit of reboxetine versus
placebo by up to 115% and reboxetine versus SSRIs by
up to 23%, and also underestimated harm compared to
the information presented in CSRs and other regulatory
documents. Similarly, by comparing publications of Orli-
stat trials to their corresponding CSRs, both Schroll et al
[9] and Hodkinson et al [10] identified that journal pub-
lications provided ‘insufficient’ information on harms
outcomes of clinical trials and in some cases the authors
noted ‘important disparities’ in the definitions and num-
bers of adverse events were reported across different
documents relating to the same trial. We note that many of the research purposes outlined in
Table 1 overlap and are not intended to be an exhaustive
list; rather a general summary of a sample of published re-
search using CSR data at the time of writing. It must be
noted that the examples discussed within this section are
a ‘selective sample’ of academic work which has mostly
shown changes in conclusions, particularly regarding
harms of drugs, when analysing or re-analysing clinical
trial data using CSRs and the methodological approaches
taken by some of the academic research groups have been
challenged by the pharmaceutical companies in question
(see Additional file 1: Table S1). This sample should not
be considered completely reflective of academic objectives
for accessing regulatory documents or a comprehensive
list of all research using CSRs (which is likely much wider
as indicated by the number of requests from Fig. 1) The
selective nature of the sample summarised here must be
taken into account when interpreting the findings within
the context of all published research making use of
unpublished regulatory documents. Use of previously unpublished summary data for
systematic reviews and meta-analyses Previous work has shown that publicly available informa-
tion (such as from journal publications and trial registries)
of primary and secondary outcomes, as well as patient-
relevant outcomes and harms, may not be sufficient [5–7,
11, 16], particularly for the objectives of a novel systematic
review or meta-analysis [11, 16]. Additionally, the format
that the summary results are provided in may not readily
allow the inclusion of the information within meta-
analysis; for example, where a measure of precision of the
treatment effect is not published or if numerical results
are not reported for all measurement times [16]. A related publication from Maund et al [14], presents a
meta-analysis relating to the benefit and harms of duloxe-
tine for the treatment of stress urinary incontinence, with a
specific interest in harms related to suicide and violence, in
response to FDA concerns about an association between
duloxetine and these serious adverse events. The authors
use data from CSRs (summary tables and narratives) to per-
form meta-analysis of four trials and compare their analysis
to the results of a Cochrane review of the same topic con-
ducted only with data available in the public domain (i.e. from trial publications). The two pooled analyses come to
the overall same conclusion that clinical benefits of duloxe-
tine for stress urinary incontinence do not outweigh the po-
tential harms, but the two analyses consider different
outcomes sets and show slightly different results for com-
mon outcomes. Notably, the authors note that the analysis
of CSR data has allowed more detailed considerations of
specific AEs which would not have been possible without
access to CSRs and narratives. Therefore, in order to provide the most clinically inform-
ative and high quality systematic review and meta-analysis
results as possible, researchers may wish to use unpublished
summary (aggregated) data from one or more CSRs within
systematic reviews and meta-analyses. In 2014, Jefferson
et al [11, 27, 28] reported on the first Cochrane review to be
based on all relevant full CSRs of a drug (oseltamivir), aug-
mented by ‘regulatory comments’ (i.e. any other relevant in-
formation
submitted
to
regulatory
authorities
for
oseltamivir). For example, “one patient had a “nervous breakdown,”
which was coded as mental disorder, and another patient
reported “feeling drugged,” which was coded as somnolence. Assessment of reporting and evaluation of bias nine trials of duloxetine for the treatment of major depres-
sive order shows that coded events and narratives within
CSRs suggest different numbers of events related to sui-
cide and the authors conclude that in this case the narra-
tives are more informative and coded events in summary
tables may be misleading and may not fully capture the
true nature of the event. Specifically: “…narratives of adverse events can provide additional
information, including original investigator reported ad-
verse event terms, which can enable a more accurate esti-
mate of harms [13].” “Using the patient’s trial identification number we were
able to reconcile data reported in the patient listings with
those in the narrative. Secondly, using data (treatment as-
signment, coded term, and timing of event) from the patient
listings and narratives, we were able to reconcile data from
these two formats with the data in summary tables [13].” Use of previously unpublished summary data for
systematic reviews and meta-analyses In addition, 5 patients, all receiving duloxetine, experienced
a total of 8 events that were mentioned only in the narrative
text [14].” A CSR would typically contain more details about the
choice of statistical method, interpretation of results and
the full set of endpoints, results and statistics at all time
points measured and therefore may provide a useful sup-
plementary source of data for systematic reviews and meta-
analysis. For example, a review of 101 CSRs conducted by
Wieseler et al [5] shows that CSRs provided complete infor-
mation on 78 to 100% of benefit outcomes (compared to
20 to 53% from publicly available sources), CSRs also pro-
vided considerably more information on harms and on
patient-relevant outcomes such as outcomes describing
morbidity, mortality, and health-related quality of life. These publications highlight the potential utility of the
participant narratives and participant listings for novel ana-
lyses relating to AEs and further understanding of drug
safety profiles beyond summary statistics of dictionary-
coded events. It must be noted that participant listings are
out-of-scope for ‘Phase 1’ of EMA Policy 0070 and work
conducted in November 2017 showed that the majority of
CSRs published under EMA Policy 0070 have ‘fully redacted’
participant narratives [25], likely due to concerns regarding
the risk of re-identification from narrative information relat-
ing to a specific participant and technical challenges to
retrospectively anonymise unstructured data (i.e. narratives Assessment of reporting and evaluation of bias It is mandatory that a summary of adverse events (AEs)
occurring during a trial is made public via registries such
as ClinicalTrials.gov or EudraCT [24]. However, a num-
ber of examples have made use of the more detailed
harms and AE information available within CSRs, both
in the context of appraising ‘selective outcome reporting’
as outlined in the section above [4, 9–11, 15] as well as
for novel analyses of previously unpublished harms and
AE information [11–14]. p
g
A common reason for using unpublished information,
such as the trial protocols and CSRs of specific trials to
access to detailed trial methodology and comprehensive
results, is to perform an appraisal, assessment and evalu-
ation of any bias in the trial design, misreporting or any
selective outcome reporting bias in journal manuscripts
[2, 3]. It is recommended within the Cochrane Handbook
for Systematic Review of Interventions that selective out-
come reporting should be appraised within Cochrane
Reviews by comparing trial protocols with published re-
ports [23] and examples identified within this review
have also shown the use of CSRs for identifying misre-
porting or selective outcome reporting. For example, Maund et al [12, 13] present methodo-
logical investigations relating to the conclusions that can
be drawn from reading summary tables of AEs which are
usually dictionary coded, compared to reading verbatim
descriptions in the narratives. Their illustrative example of Page 5 of 10 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Page 5 of 10 and body text), despite EMA Policy 0070 external guidance
specifying that case narratives should be anonymised rather
than removed [17]. With the valid re-identification concerns
in mind, primarily, the ability to follow the ‘journey’ of a par-
ticipant through a trial, particularly preserving sequences
and distances between interventions, findings and events
(and access to different definitions of events, including inves-
tigator reported terms) has clear advantages for the ‘utility’
of CSR data and is essential for detailed evaluations of harms
outcomes, such as in the examples of Maund et al [12–14]. Secondarily, availability of demographics and medical history
among others could also support more detailed analyses, in-
cluding analyses relating to harms and AEs. Use of regulatory data in Cochrane reviews: current
practice and attitudes A survey was conducted among Cochrane authors between
June and September 2016 regarding any experiences and Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Page 6 of 10 Page 6 of 10 case report forms etc.) that were gathered before the im-
plementation of EMA Policy 0070. opinions they had of using CSRs and ‘other regulatory doc-
uments’ (defined as any document produced by or held by
a regulatory agency) in Cochrane Reviews [20, 21, 29]. In order to further explore the use of CSRs in the pro-
jects and the potential impact that EMA Policy 0070
may have on the data utility of anonymised CSRs in aca-
demic research, in August to September 2017, we
attempted to make contact with the authors of the jour-
nal publications identified and discussed within this re-
view (Table 1 and Additional file 1: Table S1). Full
details
of
the
correspondence
with
authors
who
responded to us is provided in Additional file 2, shared
with the permission of the authors. There were 156 respondents out of 7181 invited
Cochrane authors who had worked on a Cochrane review
in the previous 2 years (around 2% response rate). Results
of the survey showed only 10% of the 156 respondents
used or requested regulatory data (mostly CSRs to supple-
ment meta-analyses) from pharmaceutical companies dir-
ectly or from regulatory agencies such as the EMA and
the FDA. A further 5% had considered using regulatory
data and the remaining 85% had not considered using
regulatory data. Of those who had experience of using or
requesting regulatory data, 80% believed that regulatory
data should be used on Cochrane reviews, yet two thirds
of these respondents who had accessed and included CSR
data in Cochrane reviews mentioned barriers to using this
data such as limited or restricted access to the data (such
as not being able to print documents and download data)
and lack of experience or skills required to interpret these
detailed documents [29]. In summary, all of the authors stated that their ana-
lyses would not have been possible without access to
CSRs. None of the authors raised any specific concerns
about anonymised or redacted CSRs (in line with EMA
Policy 0070). In fact, one research team had used CSRs
publicly available from a sponsor website which were
redacted and this redaction did not impact upon the
analysis from the author’s recollection [8]. Use of regulatory data in Cochrane reviews: current
practice and attitudes Furthermore,
none of the authors stated that their team had any diffi-
culties in interpreting the information from the CSRs (a
potential barrier raised within the Cochrane Survey) [20,
29]; the only problems related to ‘illegible’ text or the
portable document format of the documents which pre-
vented electronic searching. The proportion of individuals who believed that regula-
tory data should be used in Cochrane Reviews falls to 38
and 32% respectively for those who had considered or had
not considered using regulatory data and results of the
survey also showed that out of all respondents, only 12%
claimed to know where to access regulatory information
from clinical trials and 32% of respondents admitted to
having no understanding of regulatory submissions and
the documents produced for this process [29]. All of the authors stated that some or all of their ana-
lyses or research would not have been possible if narra-
tives and/or appendices (with participant listings) were
removed from anonymised CSRs under EMA Policy 0070. One author stated that: “Anonymised CSRs are ok, but the
current EMA policy redacts important information about
when the adverse events appeared as well as what they
were. Newer CSRs do not have individual adverse event
listings and the EMA are not even in possession of them.” These results demonstrate that using regulatory data,
such as the data from CSRs is rather new for the academic
Cochrane community as a whole but researchers who are
requesting or using regulatory documents to complete
Cochrane Reviews consider access to these documents im-
portant and valuable for their analyses. Results of the sur-
vey also suggested that the availability of further guidance
on how to interpret and use regulatory data in secondary
analyses would help to promote the use of CSRs in
Cochrane Reviews [29]. Conference proceedings have
been presented regarding advantages and disadvantages in
using CSRs versus published articles for Cochrane reviews,
highlighting the extra information and opportunities of
such data sources as well as challenges working with CSRs
for Cochrane authors [30, 31]. Guidance of when to use
Clinical Study Reports and other regulatory documents in
systematic reviews has been developed as part of a
‘Cochrane Methods Innovation Fund’ [21]. Another author stated that: “I have actually looked at
data that are released under the EMAs new EMA Policy
0070, and they do provide fully redacted CSRs. Use of regulatory data in Cochrane reviews: current
practice and attitudes So yes, I
would say you could use these provided the drug is centrally
licenced. But redactions may permit what data can be used,
and they may not be of use for creating IPD datasets with-
out the subject IDs and other patient-level information.” It should be emphasised that these observations are
anecdotal and rhetorical as these projects were based on
CSRs that were obtained before the implementation of
EMA Policy 0070. We must also note that the quotes
above reflect the opinions of the authors and we have
not verified the accuracy of any statements made by the
authors. However, our observations as well as the obser-
vations and views of the academic authors, and the ratio-
nales for the type of analyses being conducted using
CSRs do raise some potential issues relating to ‘data util-
ity’ of documents anonymised under EMA Policy 0070
(Phase 1). The full extent and any impact of such issues Summary of findings and implications CSRs have previously been considered as an ‘untapped’
source of detailed information relating to design, con-
duct and analysis of clinical trials [1]. The value of the
information within CSRs is becoming increasingly recog-
nised within the academic research community, particu-
larly within the Cochrane Collaboration [20, 21, 29]. Publicly available summary data of clinical trial results
from journal publications and trial registries may be
suitable and sufficient to support some types of second-
ary analyses. However, an anonymised CSR provides
complete information and data on study design and stat-
istical methods, interpretation of results and the full set
of endpoints’ results and statistics. Such a resource
would certainly allow a more in depth appraisal includ-
ing verification of numerical results, detailed assessment
relating to bias such as selective outcome reporting and
the conduct of novel analyses, such as systematic reviews
and meta-analyses using data from all endpoints. EMA Policy 0070 ‘Phase 1’, where anonymised CSRs
are made public, is likely to further facilitate the second-
ary use of the information within CSRs for appraisal and
secondary analysis. However, little consideration has
been given to the data utility of the anonymised infor-
mation within CSRs under EMA Policy 0070. The ob-
jective of this review was to investigate the type of
research purposes and research methodologies employed
in previous work using data from CSRs in academic re-
search, and to hypothesize the impact of EMA Policy
0070 on the ‘data utility’ of future research of this kind. Two main general types of analysis emerged from this
research: Appraisal and Secondary Analysis. The different
objectives across these two general analysis types should
help prioritise anonymisation methods from a data utility
perspective in addition to data privacy considerations. The
classification of research objectives also provides more
guidance for developing a qualitative approach to assess
and document data utility of anonymised CSRs in-line
with EMA Policy 0070 [17], Health Canada Public Release
of Clinical Information Policy [33] or other contexts. Further understanding of the safety profile of the drugs
and verification of how conclusions of clinical studies are
derived would certainly provide added value for many
stakeholders and data consumers, including patients them-
selves. However, several journal publications that were
reviewed within this review and described in Additional file
1: Table S1 have had their findings challenged by the spon-
soring pharmaceutical companies through comments on
journal websites. Potential impact of EMA policy 0070 on data utility of
CSRs: academic author interviews The examples considered within this review are based on
CSRs and other regulatory documents (such as protocols, Page 7 of 10 Page 7 of 10 Page 7 of 10 Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 will not become apparent until sufficient research pro-
jects are conducted and published using anonymised
CSRs prepared in line with EMA Policy 0070. Table S1). Handling of narratives, together with the hand-
ling of in-text listings, seems to be the most challenging
aspect of EMA Policy 0070 from a technical standpoint
and various levels of or approaches to anonymisation
would further define different levels of data utility. Discussion y
Certain free-text fields such as e.g. Adverse Events Re-
ported Terms has been shown to be instrumental for
certain secondary analysis to e.g. verify dictionary coding
and conduct re-analysis [12–14]. Further, preserving
Subject IDs and Dates in an anonymised format in order
to follow the events of a participant, using sequences
and distances to further understand the drug safety pro-
file. An ongoing review of CSRs published under EMA
Policy 0070 indicates that free-text variables (such as
narratives) tend to be fully redacted when a dictionary-
coded variable is available and deemed to be better
suited for analysis [25]. The PhUSE De-Identification
standard [32] recommends as a primary rule in the case
of pro-active release of data to follow such rational and
a secondary rule to “Review and redact PII” in such free-
text variables. It is therefore advised to researchers to
make it clear in their requests in the context of their re-
search objective whether certain free-text variables (with
all PII redacted) are required, even if a dictionary-coded
variable is available in the given data domain. Summary of findings and implications Discussion of academic findings and in-
terpretations should always be encouraged but there is a
risk that ‘rapid-response’ additional analyses as a challenge
to published research may confuse readers and secondary
data consumers such as clinical practitioners, patients and
patient associations who cannot interpret which of the
many published results are the ‘correct’ ones. Bonini et al. 2014 [34] also note that “access to clinical data imposes a
high ethical standard on anyone using those data, lest in-
appropriate reanalyses breed unjustified concern about the
efficacy or safety of marketed drugs.” We (SJN and JMF) are Based on the number of requests made under EMA Pol-
icy 0043, we anticipate that academic researchers or re-
search groups and the Pharmaceutical Industry are likely
to be the primary recipients of anonymised CSRs under
EMA Policy 0070. The research examples we discuss
within this review indicate that the objectives and scopes
of secondary analyses and novel research that have been
conducted using CSR data are vast (Table 1, Additional
file 1: Table S1). Authors of such research have communi-
cated with us their concerns over the type of research that
could be conducted in the future if information such as
participant listings or narratives are redacted or removed
completely under EMA Policy 0070 (Additional file 2). Keeping the same conclusions and comparable numer-
ical results of all primary, secondary and safety endpoints
in the Anonymised CSRs to those of the original CSR
(prior to any anonymisation) is of utmost importance. There are examples in the literature on how narratives are
used to verify safety conclusions (see Additional file 1: Ferran and Nevitt BMC Medical Research Methodology (2019) 19:204 Page 8 of 10 Page 8 of 10 of the opinion that communication (and potentially collab-
oration) between academic research groups and pharma-
ceutical companies regarding interpretations of regulatory
documents such as protocols, statistical analysis plans and
CSRs, and the interpretation of results of secondary ana-
lyses from their different perspectives during the research
projects should be encouraged. Such communication and
discussions occurring before any results of secondary ana-
lyses are published within journals are likely to provide the
most informative novel results and in turn, provide the
most benefit to readers and data consumers. method and full listings of participant narratives are out-
of-scope [37]. Supplementary information
S
l
i f
i pp
y
Supplementary information accompanies this paper at https://doi.org/10. 1186/s12874-019-0836-3. Additional file 1: Table S1. Examples of academic work using CSRs in
secondary research. Additional file 1: Table S1. Examples of academic work using CSRs in
secondary research. Additional file 2. Correspondence with authors of journal publications
using CSRs for academic research. A list of semi-structured interview
questions posed to corresponding authors of journal publications using
CSRs for academic research and corresponding author responses to
questions. Abbreviations AE: Adverse Event; CCI: Company Confidential Information;
ClinicalTrials.gov: US Clinical Trials Database; CSR: Clinical Study Report;
DIA: Drug Information Association; EFPIA: European Federation of
Pharmaceutical Industries and Associations; EMA Policy 0043: European
Medicines Agency Policy on access to documents (related to medicinal
products for human and veterinary use); EMA Policy 0070: European
Medicines Agency policy on the publication of clinical data for medicinal
products for human use (Phase I for Clinical Study Reports); EMA: European
Medical Agency; EU: European Union; EudraCT: European Clinical Trials
Database; FDA: Food and Drug Administration; HTA: Health Technology
Assessment; IPD: Individual Patient Data; LPLV: Last Patient Last Visit;
OECD: Organization for Economic Co-operation and Development;
PII: Personal Identifying Information; SAE: Serious Adverse Event; SAP: Clinical
Trial Statistical Analysis Plan In the context of EMA Policy 0070, where anonymised
CSRs are made public, a myriad of data recipient groups
could be considered together with various objectives for
reviewing and using the information within these anon-
ymised CSRs. Their needs could differ from the academic
research community and could be worth investigating at a
later point. In addition, at the time of writing, other regulatory
agencies are defining, finalising and publishing their data
transparency initiatives. FDA made an announcement in
January 2018 [36] and started pilots with pharmaceutical
companies where redaction is the only anonymization Summary of findings and implications Health Canada started developing regula-
tions around public access to clinical documents in 2017
and released a draft guidance for review in the second
quarter of 2018 [33]. Difference of requirements between
EMA Policy 0070 guidance, FDA and Health Canada ap-
proaches (under development) [38] may also lead to dif-
ferent anonymized versions of the same document in the
public domain. Only when all policies are finalised will it
become clear which versions under which jurisdiction
serve best the needs of the academic community and
others. Conclusions It must be emphasised that the examples of academic re-
search using CSRs summarised within this review (Table
1 and Additional file 1: Table S1) are a selective sample
and do not necessarily represent all research objectives
which would make use of anonymised CSRs under EMA
Policy 0070. Further, most observations provided to us
by the authors of this research and our interpretations
are rhetorical and speculative, rather than based on dir-
ect experience of anonymised CSRs published under
EMA Policy 0070 and the validity of these observations
may not become clear for some time. In conclusion, EMA Policy 0070 guidance [17] refers to
various levels of anonymisation but based only on level
of risk of re-identification of participants without refer-
ence to different levels of data utility. The extent of the
data utility of CSRs published under EMA Policy 0070
for academic research may not be fully known for some
time, therefore in this interim time period, this review
provides an initial insight into the previous use of CSR
data, current practices for including regulatory data in
academic research such as Cochrane systematic reviews
and some early indications for the potential use of the
data, and therefore the utility of anonymised data, from
CSRs published under EMA Policy 0070. y
The planned ‘Phase 2’ of EMA Policy 0070 which ex-
tends to sharing of individual participant data (IPD)
should provide the next level of data utility that is required
to conduct robust secondary analyses. A number of spon-
sors already provide access to anonymised IPD via data
sharing platforms [22, 26] based on research requests for
studies under the European Federation of Pharmaceutical
Industries and Associations (EFPIA) principle of respon-
sible clinical trial data sharing [35]. ‘Phase 2’ of EMA Pol-
icy 0070, when in effect, should in principle standardise
the access to anonymised IPD for studies part of a central
application in European Union regardless of the outcome
of the application. The current needs of the research com-
munity which may include access to individual participant
information such as full patient listings, which is out of
scope of EMA Policy 0070 ‘Phase 1’, will be better ad-
dressed in ‘Phase 2’ of the policy where Individual Patient
Datasets are in scope. Consent for publication
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JMF: I have worked for various Pharmaceutical companies as a consultant
since January 2010 on compound specific clinical projects or cross-
compound system-related projects. I have also entered a strategic alliance in
January 2017 with d-Wise Inc. (North Carolina) where I contribute as an SME
to the design and development of their data de-identification products. I am
an appointed member of the European Medicines Agency Technical Anon-
ymisation Group and Health Canada Stakeholder Reference Group on Public
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h We would like to thank Dr. Tom Jefferson and Dr. Alex Hodkinson for
providing initial results of the Cochrane Survey and we would like to thank
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Khaled El-Emam, Professor Catrin Tudur Smith, Dr. Jesse Berlin and Stephen
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published maps and institutional affiliations.
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Introduksi Produk Olahan Ubi Jalar Pada Anggota Ika Tanjung Sakti, Bandar Lampung
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Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat Vol. 4, No. 2 Juni 2020, Hal. 263-267 Abstrak Ubi jalar merupakan pangan karbohidrat nonberas yang mempunyai manfaat kesehatan dan gizi,
sehingga perlu disebar-luaskan di masyarakat. Program pengabdian kepada masyarakat ini bertujuan
meningkatkan pemahaman masyarakat sasaran terkait diversifikasi pengolahan ubi jalar menjadi berbagai
produk pensubstitusi nasi, serta memberikan pengetahuan manfaat ubi jalar dari aspek gizi, dan
kesehatan.Khalayak sasaran dalam kegiatan ini adalah ibu-ibu kelompok IKA Tanjung Sakti, Sukarame,
Bandar Lampung. Metode yang digunakan adalah ceramah, diskusi dan evaluasi. Hasil kegiatan
menunjukkan persentase perubahan pengetahuan peserta tentang manfaat ubi Jalar meningkat sebesar
85%, dan terhadap produk olahan ubi jalar sebesar 67-100 % bergantung pada jenis produk. Secara umum,
peserta menganggap penyuluhan atau introduksi tentang ubi jalar ini bermanfaat. Kata kunci: Tanjung Sakti IKA, manfaat kesehatan, produk ubi jalar Neti Yuliana*1, Dewi Sartika2, Samsu Udayana Nurdin3, Novita Herdiana4, Pramita Sari
Anungputri5
1,2,3,4,5 Jl Program Studi Teknologi Industri Pertanian, Fakultas Pertanian, Universitas Lampung
*e-mail: neti.yuliana@fp.unila.ac.id Neti Yuliana*1, Dewi Sartika2, Samsu Udayana Nurdin3, Novita Herdiana4, Pramita Sari
Anungputri5
1,2,3,4,5 Jl Program Studi Teknologi Industri Pertanian, Fakultas Pertanian, Universitas Lampung
*e-mail: neti.yuliana@fp.unila.ac.id Keywords: Tanjung Sakti IKA, health and nutritional benefits, sweet potatoes product Keywords: Tanjung Sakti IKA, health and nutritional benefits, sweet potatoes product Abstract Sweet potato is a non-rice carbohydrate food that has health and nutritional benefits, so it needs to be
disseminated in the community. This community extention program aimed to increase the understanding of
the target community regarding the diversification of sweet potatao processing into various rice substitution
products, as well as providing knowledge of the benefits of sweet potato from the nutritional and health
aspects. The participant audiences was the Tanjung Sakti IKA group members, Sukarame, Bandar Lampung. The method used was lecture, discussion, and evaluation. The results of the activity showed that the change
percentage in participants' knowledge about the benefits of sweet potatoes increased by 85%, and that about
the processed sweet potato products by 67-100% depending on the type of product. In general, participants
consider counseling or introduction of sweet potatoes was useful. 1. PENDAHULUAN Varietas-varietas lokal seperti singkong, garut, gembili, ubi jalar dan lain-lain berpotensi
menjadi bahan pangan alternatif ataupun substitusi beras, dan terigu. Keadaan ini mendorong
masyarakat, peneliti dan pemerintah untuk kembali mengenalkan, membudidayakan, dan
memanfaatkan secara maksimal bahan-bahan karbohidrat lokal untuk memperkuat ketahanan
pangan lokal menuju kemandirian pangan. Bahan pangan lokal sumber karbohidrat yang semula
terpinggirkan perlu diperkenalkan kembali, dan dikembangkan dalam berbagai diverisifikasi
produk. Beberapa produk non beras yang telah dikembangkan antara lain dalam bentuk mie,
roti, makanan jajan pasar, serta beras tiruan (beras analog)(Kumalasari et al., 2018; Muthoharoh
dan Sutrismo, 2017; Pasparingi dan Hudi, 2014; Noviasari et al., 2017). Kekhawatiran sebagai pengimpor terigu terbesar (Nurhalimah, 2019) dan impor beras
yang meningkat (Kusumah, 2019), menjadi tantangan bagi para akademisi perguruan tinggi
untuk mensosialisasi dan menyebarluaskan pentingnya produk pangan nonberas bagi
masyarakat yang diikuti dengan gerakan massal yang terencana dan berkelanjutan. Pemikirian
masyarakat perlu diarahkan bahwa banyak alternatif produk pangan nonberas yang bisa
menggantikan fungsi nasi sebagian atau seluruhnya. Hal ini dapat dilakukan dengan introduksi,
sosialisasi dan pengembangan pengolahan pascapanen bahan pangan berkarbohidrat nonberas
yang melibatkan masyarakat. P-ISSN 2614-7424 | E-ISSN 2614-8927 263 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat Vol. 4, No. 2 Juni 2020, Hal. 263-267 Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 Tim peneliti dari Jurusan Teknolgi Hasil Pertanian, Fakultas Pertanian, Universitas
Lampung, telah berhasil mengembangkan tanaman lokal ubi jalar menjadi tepung melalui
berbagai proses modifikasi sifat tepung sehingga dapat meningkatkan fungsinya sebagai
pensubstitusi terigu (Yuliana dan Nudjanah, 2013). Ubi jalar merupakan produk pertanian ke
dua yang banyak diproduksi di provinsi Lampung, sehingga dengan mudah banyak di temui di
pasar-pasar. Selain berdaging umbi putih, varian ubi jalar ada yang berwarna kuning dan ungu,
yang mempunyai khasiat kesehatan seperti anti cancer dan anti oksidant (Ayeloso et al., 2016). Pengembangan kudapan (snack) berbasis ubi jalar mempunyai kelebihan karena dapat dibuat
menggantikan porsi terigu sampai separuh atau seluruhnya sebagai substitusi, merupakan
sumber karbohidrat yang mengenyangkan seperti beras dan mempunyai indek glikemik yang
rendah dibandingkan ubi lainnya dan beras, sehingga juga sesuai untuk penderita diabetes. Dalam bentuk snack rerotian, berbagai varian produk yang bergizi juga dapat diturunkan
dengan tambahan mentega dan sayur, daging dan ikan. Umumnya, pangan yang disubtitusi
dengan tepung ubi jalar belum tersebar luas di masyarakat. Berdasar kenyataan di atas, upaya
introduksi dan sosialisasi perlu dilakukan terkait pentingnya diversifikasi produk pangan
olahan non beras dari ubi jalar sebagai upaya mendukung ketahanan pangan nasional. 2. METODE Kegiatan introduksi dan pelatihan dilakukan di pusat kegiatan kelompok yang
beralamatkan di Sukarame-Bandar Lampung. Metode kegiatan terdiri dari ceramah, diskusi, dan
evaluasi. Penyebaran informasi melalui metode ceramah dilengkapi dengan contoh-contoh
terkait materi pelatihan untuk memotivasi peserta untuk bertanya. Setelah penyampaian materi
ceramah, kemudian dilakukan diskusi (tanya jawab) yang memberikan kesempatan kepada
peserta untuk melakukan proses pemahaman materi atau menyampaikan gagasan dan
permasalahan yang terkait dengan pentingnya diversifikasi produk, manfaat ubi jalar dan
produk ubi jalar bagi kesehatan dan gizi. 1. PENDAHULUAN Secara umum program pengabdian kepada masyarakat ini bertujuan memberikan
pelatihan tentang pengolahan ubi jalar menjadi berbagai produk yang dapat dikonsumsi sebagai
pengganti nasi, serta memberikan pengetahuan terkait manfaat ubi jalar dari aspek gizi,
kesehatan dan pengembangan produknya. Khalayak sasaran dalam kegiatan ini adalah ibu-ibu
kelompok IKA Tanjung Sakti, Sukarame, Bandar Lampung. Khalayak sasaran ini dipilih dengan
pertimbangan komunitasnya melakukan kegiatan rutin selama ini secara aktif, mempunyai
anggota yang beragam latar belakang pendidikan dan ragam sosial ekonomi. Khalayak sasaran
belum pernah mendapatkan penyuluhan dan pelatihan pengolahan ubi jalar dan manfaatnya. 3. HASIL DAN PEMBAHASAN Secara keseluruhan, kegiatan pelatihan ini telah dapat meningkatkan pengetahuan
peserta terkait manfaat kesehatan dan gizi ubi jalar, dan produk olahan ubi jalar (tepung ubi
jalar, snack kue-rerotian dan stik ubi jalar). Hasil evaluasi yang dilakukan untuk mengukur
keberhasilan peserta pada awal proses dan akhir tersaji pada Tabel 1,2 dan Grafik 1. Hasil
kegiatan
menunjukkan
bahwa
60%
peserta
dapat
ditingkatkan
pengetahuan
dan
ketrampilannya, yang mengindikasikan proses penyuluhan berhasil diterapkan. Pada
penyuluhan terdapat suatu proses pendidikan, metode penyebaran informasi untuk membawa
perubahan pengetahuan, sikap, ketrampilan dan motivasi, seperti yang telah sukses dilakukan
pada program pengabdian gizi (Nindrea, 2017), penyuluhan pembangunan desa (Prahati et al.,
2017), dan penerapan cara produksi pangan (Esthy, 2019). Table 1. Persentase Perubahan Pengetahuan Peserta tentang Ubi Jalar
Kondisi
Pengetahuan
jenis ubi jalar
nilai gizi
manfaat kesehatan
sebelum
sesudah
sebelum
sesudah
sebelum
sesudah P-ISSN 2614-7424 | E-ISSN 2614-8927 264 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat Vol. 4, No. 2 Juni 2020, Hal. 263-267 Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 Ya tahu
90
100
10
90
0
85
Sedikit
10
0
45
10
40
5
Tidak
0
0
45
0
60
0 Pengetahuan peserta terkait nilai gizi dan kesehatan meningkat terlihat dari banyaknya
prosenstase peserta sebelum penyuluhan yang tahu hanya sedikit atau bahkan tidak tahu,
meningkat menjadi tahu. Ketertarikan peserta terutama pada informasi bahwa warna kuning
ubi jalar tidak kalah khasiatnya pada carotene wortel. Ubi jalar diketahui mengandung
betacarotene yang bermanfaat sebagai provitamin A dan antioksidan (Christides et al., 2015;
Ayeloso et al., 2016). Demikian pula pengetahuan tekait olahan pembuatan tepung ubi jalar yang
kemudian dapat dibuat produk turunannya seperti stik, bolu, tepung, brownis dan kue Bugis. Hal ini dimungkinkan karena ubi jalar merupakan sumber karbohidrat yang dengan mudah
dapat diproduksi menjadi tepung yang mempunyai daya simpan tinggi serta dapat dimanfaatkan
pada produk olahan agroindustri lainnya (Tamam et al., 2014; Yuliana et al., 2014; Yuliana et al.,
2018]. Tepung ubi jalar kemudian dapat mensubstitusi terigu sebagai bahan dasar berbagai
snack. Table 2. 4. KESIMPULAN Penyuluhan meningkatkan pengetahuan peserta mengenai manfaat ubi jalar dari aspek
gizi, kesehatan dan pengolahan ubi jalar menjadi berbagai produk yang dapat dikonsumsi. Peserta juga berminat untuk mempraktekkannya di rumah. Hasil pengabdian ini disarankan
untuk ditindak lanjuti dengan cara mengakomodasi peserta yang aktif melakukan praktek agar
dapat diberdayakan sehingga dapat menunjang ekonomi keluarga, melalui skim program
pengabdian yang sesuai. UCAPAN TERIMA KASIH Penulis mengucapkan terima kasih kepada DIPA Fakultas Pertanian Unila, yang telah
memberi dukungan financial terhadap pengabdian ini. 3. HASIL DAN PEMBAHASAN Persentase Perubahan Pengetahuan Peserta tentang Olahan Ubi Jalar
Tingkat
mengetahui
stik ubi jalar
tepung ubi jalar
brownis
sebelum
sesudah
sebelum
sesudah
sebelum
sesudah
ya
0
100
0
100
33
100
sedikit
90
0
40
0
67
0
tidak
10
0
60
0
0
0
bolu
bola-bola
bugis
sebelum
sesudah
sebelum
sesudah
sebelum
sesudah
ya
42
100
30
100
33
100
sedikit
42
0
46
0
42
0
tidak
16
0
24
0
25
0 Table 2. Persentase Perubahan Pengetahuan Peserta tentang Olahan Ubi Jalar
stik ubi jalar Seluruh peserta menganggap penyuluhan atau introduksi tentang ubi jalar ini
bermanfaat, dan sebagain peserta mempunyai minat untuk memprakekkannya di rumah
(Gambar 1). Berdasarkan hal ini, tindak lanjut kegiatan perlu diusulkan pada kegiatan
pengabdian berikutnya yaitu praktek pengolahan ubi jalar menjadi berbagai snack
mengenyangkan sebagai substitusi terhadap terigu dan beras. Kajian produk-produk ini telah
banyak dilaporkan antara lain oleh Noer et al. (2017), Aulia et al. (2017), Izza et al. (2019),
sehingga introduksi dan praktek dapat dengan mudah diterapkan oleh masyarakat. Gambar 1. Pendapat peserta tentang penyuluhan dan minat peserta Gambar 1. Pendapat peserta tentang penyuluhan dan minat peserta P-ISSN 2614-7424 | E-ISSN 2614-8927 265 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat Vol. 4, No. 2 Juni 2020, Hal. 263-267 Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 Gambar 2. Aktivitas Penyuluhan Gambar 2. Aktivitas Penyuluhan DAFTAR PUSTAKA Aulia, S.S., Rustanti, N., & Fitranti, D.Y. (2017). Fotifikasi NaFeEDTA pada cookies ubi Jalar kuning
sebagai poduk alternatif untuk menanggulangi anemia defesiensi besi. Jurnal Gizi Pangan,
12(3),161-168. Ayeleso, T.B., Ramachela, K., & Mukwevho, E. (2016). A review of therapeutic potentials of
sweet potato: Pharmacological activities and influence of the cultivar. Tropical Journal of
Pharmaceutical Research, 15(12), 2751-2761, (
)
Christides, T., Amagloh, F.K., & Jane, C. (2015). Iron bioavailability and provitamn A from potato
and ceraal-based complementary foods. Foods, 4, 463-476 Esthy, E. R. A. (2019). Penerapan cara produksi pangan yang baik untuk meningkatkan kualitas
bawang goreng pada industri rumah tangga di Kelurahan Tangkerang Tengah Kecamatan
Marpoyan Damai Pekanbaru. Dinamisia : Jurnal Pengabdian Kepada Masyarakat, 3(2), 221-
227. https://doi.org/10.31849/dinamisia.v3i2.3229 Izza, N.K., Hamidah, N., & Ira, Y. (2019). Kadar lemak dan air pada cookies dengan substitusi
tepung ubi ungu dan kacang tanah. Jurnal Gizi, 8(2),106-114. Kusumah, F.P. (2019). Ekonomi politik dan kebijakan impor beras: Membaca arah kebijakan
pemerinth 2014-2019. Politika: Jurnal Ilmu Politik, 10(2), 137-162, Muthoharoh, D.F. & Sutrisno,A, (2017). Pembuatan roti tawar bebas gluten berbahan baku
tepung garut, tepung beras, dan maizena (Konsentrasi Glukomanan dan waktu proofing). Jurnal Pangan dan Agrindustri vol 5(2), 34-44. P-ISSN 2614-7424 | E-ISSN 2614-8927 266 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat
Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 DINAMISIA: Jurnal Pengabdian Kepada Masyarakat Vol. 4, No. 2 Juni 2020, Hal. 263-267 Vol. 4, No. 2 Juni 2020, Hal. 263-267
DOI: https://doi.org/10.31849/dinamisia.v4i2.2718 Nindrea, R.D. (2017). Pengaruh penyuluhan gizi dengan perubahan pilaki sarapan agi siswa
sekolah dasar. Jurnal Endurance 2(3),239-244. Nindrea, R.D. (2017). Pengaruh penyuluhan gizi dengan perubahan pilaki sarapan agi siswa
sekolah dasar. Jurnal Endurance 2(3),239-244. Nindrea, R.D. (2017). Pengaruh penyuluhan gizi dengan perubahan pilaki sarapan agi siswa
sekolah dasar. Jurnal Endurance 2(3),239-244. Noer, S.W.M., Mohammad Wijaya, & Kadirman. (2017). Pemanfaatan tepung ubi jalar (Ipomea
batatas L) berbagai varietas sebagai bahan baku pembuatan kue bolu kukus. Jurnal
Pendidikan Teknologi Pertanian, 3,S60-S71. g
Noviasari, S., Kusnandar, F., Setiyono, A., & Budijanto, S. (2017). Karakteristik fisik, kimia, dan
sensori beras anaog berbasis bahan pangan non beras. Pangan, 26 (1),1-11. Nurhalimah, F. 2019. Kepentingan Indonesia melakukan impor gandum dai Australia tahun
2012-2016. JOM FISIP 6, 1-8. Prahati, P., Zuhdi, S., & Aguswan, A. (2017). Penyuluhan penyusunan rencana pembangunan
desa di Desa Kuapan Kecamatan Tambang Kabupaten Kampar . Dinamisia : Jurnal
Pengabdian Kepada Masyarakat, 1(1), 94-99. https://doi.org/10.31849/dinamisia.v1i1.410 g
p
y
( )
p //
g/
/
Pasparingi, M. DAFTAR PUSTAKA & Hudi, L. (2014). Karakteristik kue ranginyang tebuat dai berbagai
tepung. Nabanatia Vol11 (1), 11-20. Tamam, B., Agustini, Ni.P., & Antarini, A.A.N. (2014). Karakterisrik gizi dan fisik tepung ubi jalar
dan talas temodisfikasi dengan fermentasi enzim amylase. Jurnal Sekala Husada, 11(1), 24-
28. Yuliana, N., & Nurdjanah, S. (2013). Pengembangan pikel fermentasi ubi jalar sebagai
bahan baku tepung termodifikasi: upaya mengimbangi ketergantungan terigu. Laporan
Penelitian Hibah Strategis Nasional. Lembaga Penelitian, Unila. j
(
)
g
g
p
j
g
bahan baku tepung termodifikasi: upaya mengimbangi ketergantungan terigu. Laporan
Penelitian Hibah Strategis Nasional. Lembaga Penelitian, Unila. g
g
Yuliana, N., Nudjanah, S., Sugiharto, R., & Amethy, D. (2014). Effect of Spontaneous Lactic Acid
Fermentation on Physico-Chemical Properties of Sweet Potato. Microbiology Indonesia 8
(1):1-8 Yuliana, N., Nudjanah, S., Sugiharto, R., & Amethy, D. (2014). Effect of Spontaneous Lactic Acid
Fermentation on Physico-Chemical Properties of Sweet Potato. Microbiology Indonesia 8
(1):1-8 Yuliana, N., Nurdjanah, S. & Dewi, Y.R. (2018). Physicochemical properties of fermented sweet
potato flour in wheat composite flour and its use in white bread. International Food
Research Journal, 25 (3),1051-1059. Yuliana, N., Nurdjanah, S. & Dewi, Y.R. (2018). Physicochemical properties of fermented sweet
potato flour in wheat composite flour and its use in white bread. International Food
Research Journal, 25 (3),1051-1059. P-ISSN 2614-7424 | E-ISSN 2614-8927 267
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English
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Three-dimensional Graphene with MoS 2 Nanohybrid as Potential Energy Storage/Transfer Device
|
Scientific reports
| 2,017
|
cc-by
| 9,523
|
Three-dimensional Graphene with
MoS2 Nanohybrid as Potential
Energy Storage/Transfer Device Received: 16 March 2017
Accepted: 17 July 2017
Published: xx xx xxxx Received: 16 March 2017
Accepted: 17 July 2017
Published: xx xx xxxx Kulvinder Singh, Sushil Kumar, Kushagra Agarwal , Khushboo Soni, Venkata Ramana
Gedela & Kaushik Ghosh Portable and matured energy storage devices are in high demand for future flexible electronics. Flowery shaped MoS2 nanostructures with porous and flake like morphology was used to study the
supercapacitive nature with specific capacitance (Csp) of 169.37F/g, the energy density of 28.43 Wh/
Kg and power density of 10.18 W/Kg. This nanoflower like architecture was decorated on 3D-graphene
on Graphite electrode to design the solid-state-supercapacitor prototype device of dimensions of
23.6 × 22.4 × 0.6 mm3 having considerable high Csp of 58.0F/g and energy density of 24.59 Wh/Kg,
and power density of 8.8 W/Kg. Four fabricated supercapacitors were connected in series for real state
practical demonstration using the light emitting diode that remains enlightened for 40 s by charging
it only for 25 s. This study demonstrates the 3D-graphene/MoS2 nanohybrid has a quite high overall
potential window nearly about 2.7 V (−1.5 to +1.2 V) in KOH-PVA medium which can be used for the
development of solid-state supercapacitors thereby completely eliminating the need for any expensive
ionic liquid mediums thus building an exciting potential for high-performance energy storage/transfer
devices. Current economy and population growth rates project a demand for energy of 40 terawatts (TW) with a total
population of 10 billion by 20501. Due to the increase in demand for energy, the non-renewable sources like fossil
fuels are continuously decreasing and will automatically come to an end in the near future. To meet this demand,
alternate renewable energy sources has been projected like solar, wind and hydropower. However, a suitable stor-
age device like Li-ion batteries and Na-air batteries and so on are currently lagging behind with a storage capacity
of only 1% of the total renewable energy with longer charging time. Presently, batteries are used in wide variety
not only for energy storage but also for the energy supply devices2. The use of a battery in large scale sophisti-
cated electronic devices is limited due to its heavy weight, portability, flexibility and other issues3, 4. Currently,
researchers are looking for alternative energy storage devices that effectively store and provide energy as per
requirement. www.nature.com/scientificreports www.nature.com/scientificreports www.nature.com/scientificreports Institute of Nano Science and Technology, Sec. 64, Mohali, Punjab, India. Correspondence and requests for materials
should be addressed to K.G. (email: kaushik@inst.ac.in) Received: 16 March 2017
Accepted: 17 July 2017
Published: xx xx xxxx Three-dimensional Graphene with
MoS2 Nanohybrid as Potential
Energy Storage/Transfer Device Supercapacitors are portable, flexible5 and transparent unlike conventional batteries that draw the
attention of the researchers to extract its interesting properties. Along with these physical advantages, superca-
pacitor has high charge storage capacity, high power density, low energy density, longlife time and short charging
time as compared to traditional batteries6–8. A notable improvement in performance has been achieved through
recent advances in understanding charge storage mechanisms and the development of advanced nanostructured
materials in supercapacitor devices6, 9. Supercapacitors are classified mainly in two categories such as, pseudoca-
pacitor and electrical double layer capacitance (EDLC)8. In case of pseudo capacitor, the electrical energy is stored
via faradaic reaction in electrolyte and electro-active species on the electrode surface. There are several reports
in the literature based on pseudocapacitors made of various nanomaterials as well as their composite, such as
SnO2-V2O5, Cobalt pyrophosphate, Metal oxide-CNT, RGO-metal sulfide, MnO2-veritcally aligned graphene, etc. having a specific capacitance in the range of 190–3480F/g with energy density ranging from 14–89 Wh/kg10–12. It
has been reported that the capacitance due to faradaic peaks increases the specific capacitance of supercapacitor
10–100 times higher than the electrostatic capacitance of an EDLC. Recently, Ko et al.13. has demonstrated porous
graphitic carbon with Ni2P2O7, heterostructure has shown high specific capacitance of 1893 F/g and Liao et al.14. has reported ultrahigh supercapacitance of 3480F/g using Co3O4 nanoparticles on vertically aligned graphene
sheets supported in carbon fibers. Although pseudocapacitors are shown to have high initial capacitance, but due
to limited durability, lack of stability during cycling and poor power density, its practical use in solid-state energy Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 1 www.nature.com/scientificreports/ storage devices is limited8, 15, 16. In addition to this, crystallinity and morphology also play an important role in
specific capacitance of pseudo capacitors, lower the crystallinity higher is the specific capacitance, which is due
to the availability of chemically active dangling units which can easily take part in oxidation-reduction cycle8. To
overcome these limitations, there is an extensive drive to develop EDLC type supercapacitor that carries various
advantages over pseudo capacitor. In case of EDLC, electrical energy is stored over the layer of current collector
without any redox couple. Due to the absence of faradaic current, EDLC type supercapacitors are more reliable
and have shown long term durability with minimum loss of capacitance under multiple charging-discharging
cycles. y
Xu et al. Three-dimensional Graphene with
MoS2 Nanohybrid as Potential
Energy Storage/Transfer Device have fabricated graphene hydrogel for the EDLC type supercapacitor where storage performance is
related to the high surface area17. Sundaram et al. have fabricated MnO2 nanostructures for EDLC type super-
capacitance in a lower applied potential window having Csp of 50F/g18. Various carbonaceous materials till now
have been developed for the testing of EDLC type supercapacitors due to its cost efficiency, high surface area,
porous nature and versatile existing forms19, 20. Significant enhancement in capacitive performance is not only
governed by the properties of electroactive materials but also by the separators and electrolytes. Lin et al. have
fabricated graphene based supercapacitor (Csp of 130F/g) in ionic liquid electrolyte21 where ionic liquids shows
high chemical stability as compared to conventional electrolytes at a wide potential range22. Further development
of the storage device lies on the various tricky parameters such as, proper choice of electrolyte, electrochemically
stable current collector and thin porous spacer matrix between the electrode materials etc3, 4, 23, 24. The current
developmental trend of miniaturized autonomous electronic equipment such as implantable medical devices
and active radio frequency identification (RFID) tags have raised the demand for solid-state-supercapacitors25. Significant efforts have been devoted to improve flexibility and energy storage/transfer capacity of solid-state
supercapacitors based on various carbonaceous materials and their composites26, out of which graphene shows
a promising choice of material due to high conductivity, flexibility, transparency, large surface area and layer like
structure that drags attention to various scientific communities for its large scale application in energy storage
devices17, 23, 24. In most of the cases, graphene based solid-state-supercapacitors demonstrate very low specific
capacitance which is mainly due to the parallel restacking of graphene sheets resulting in decrease of the specific
capacitance17, 27, 28. Recently, 3D graphene proves to be more prominent material for solid state supercapacitor
due to the partial stacking of graphene sheets in random orientation in three dimensions resulting in formation
of micro porous structures17. This unique orientation of graphene not only helps to improve the charge storage
capacity of graphene but helps the electrolytic ions to move freely via porous geometry. To further improve the
response of the 3D graphene as solid-state-supercapacitor various layer/porous nanomaterial hybrids have been
reported which develops the charge storage capacity of the 3D graphene via increase in surface area and porous
nature of the hybrid materials29–33. Three-dimensional Graphene with
MoS2 Nanohybrid as Potential
Energy Storage/Transfer Device It is already reported that graphene based hybrid material possess superior
charge storage capacity by providing enhanced surface area, electrical conductivity, thermal stability and mechan-
ical strength to graphene34. Out of the various nanomaterials, 2D transition-metal dichalcogenides have been a
perfect candidate for charge storage applications due to their layer like structures that shows weak Van der Waal
force of attraction with graphite or graphene, especially 1T-MoS2 nanostructures37–41. Due to their band gap of
~1.9 eV, which reveals the semi-insulating nature, they are thus not immediately thought to be an electrode mate-
rial for energy storage application42. But still MoS2 nanosheets display an excellent performance in wide, negative
potential in neutral electrolytes, such as high specific capacitance and rate performance, which promotes the
MoS2 nanosheets to be a promising electrode material for supercapacitor applications43. Till date various MoS2
based supercapacitor have been developed but only handful reports have been presented which shows the prac-
tical applicability of solid state supercapacitor42–46. The main research challenges associated with electrochemical
supercapacitors are to develop new electrode materials, to enhance specific capacitance by modification of the
electro- active material9, 37. In this work, we report the simpler and cost effective process via chemical route of synthesis of hybrid
MoS2 nanoflowers with 3D graphene heterostructure as an active material for energy storage device. The real
demonstration of the solid state supercapacitor has been performed here using the 3D graphene-MoS2 hybrid
over graphite current collector. Herein, the specific capacitance of bare MoS2 nanostructures comes out to be
169.37F/g with an energy density of 28.43 Wh/kg and power density of 10.17 W/kg. The capacitive performance
of 3D graphene-graphite electrode has been improved significantly when the current collector is incorporated
with MoS2 nanoflowers. This remarkable supercapacitor performance of the hybrid could be attributed to the
combined effect between layered MoS2 and 3D graphene which could have some real phase application in mod-
ern energy storage devices. Results and Discussion
h
i
i
f Characterization of 3D graphene and MoS2 nanoflowers. As-prepared MoS2 nanoflowers were char-
acterized by XRD, SEM, and Raman spectroscopy. Figure 1(a) displays the XRD pattern of the MoS2 nanoflowers. It shows the diffraction patterns at 2θ = 13.55, 33.06,35.22, 42.5, 49.82, 58.01, and 69.77 which arises from the (0 0
2, (1 0 0), (1 0 2), (006), (105), (110) and (201) planes of MoS2
47. The observed diffraction patterns are broadened
in nature with a slightly lower diffraction angle shift as compared to the bulk 2H-MoS2, suggesting the presence
of randomly stacked layers of MoS2 which may be due to the intercalation of NH3 in between the layers during
the synthesis48. The XRD pattern well matches with the already common works on randomly stacked MoS2 nano-
structures49, 50. No peak other than MoS2 has been observed in the XRD pattern revealing that the synthesized
MoS2 nanoflowers are highly pure in nature. Figure 1(b) shows the SEM image of MoS2 nanoflowers at a low
magnification, it is observed that the particles are spherical in nature with porous nanostructure in the order of
nanometers. It is well established in the literature that porous nanomaterials possess a superior charge storage
capacity as compared to that of their bulk counterpart51. The formation of the porous MoS2 nanoflowers is due to
the reaction conditions as well as the precursors used in the reactions. Coagulation into a sphere is most likely due Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 2 www.nature.com/scientificreports/ www.nature.com/scientificreports/ Figure 1. (a) Diffraction pattern of as-prepared MoS2 nanoflowers with peaks at 13.55, 33.06, 35.22, 42.5, 49.82,
58.01, and 69.77. (b) SEM image of MoS2 nanoflowers (Inset Raman Spectrum of MoS2 nanomaterial with the
band at 296.86, 346.87 and 390 cm−1). (c) Raman band at 1357 and 1596 cm−1 of 3D graphene. (d) SEM images
of 3D graphene. (e) EDX spectrum of MoS2-3D graphene hybrid. The flower like MoS2@3D-graphene hybrid
architecture has also been observed in SEM and high resolution TEM images shown in (f) and (g), respectively,
which reveals the petals of MoS2 nano-flowers along with the 3D-graphene network effectively enhance the
overall surface area that impacts on high storage capacity, (h) The hybrid interface is further confirmed by the
HRTEM analysis where the lattice spacing are calculated to be 0.65 nm for MoS2 and 0.34 nm for graphene,
respectively. Figure 1. (a) Diffraction pattern of as-prepared MoS2 nanoflowers with peaks at 13.55, 33.06, 35.22, 42.5, 49.82,
58.01, and 69.77. (b) SEM image of MoS2 nanoflowers (Inset Raman Spectrum of MoS2 nanomaterial with the
band at 296.86, 346.87 and 390 cm−1). (c) Raman band at 1357 and 1596 cm−1 of 3D graphene. (d) SEM images
of 3D graphene. (e) EDX spectrum of MoS2-3D graphene hybrid. The flower like MoS2@3D-graphene hybrid
architecture has also been observed in SEM and high resolution TEM images shown in (f) and (g), respectively,
which reveals the petals of MoS2 nano-flowers along with the 3D-graphene network effectively enhance the
overall surface area that impacts on high storage capacity, (h) The hybrid interface is further confirmed by the
HRTEM analysis where the lattice spacing are calculated to be 0.65 nm for MoS2 and 0.34 nm for graphene,
respectively. to the hydrothermal reaction conditions and the precursors used. During the reaction, ammonium molybdate
releases MoO4−. These ions, having a layered structure, react with sulfide ions and intercalation by the residual
ammonia prevents the stacking of MoS2 layers which leads to the self-assembly of a sphere-like morphology48. Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 3 www.nature.com/scientificreports/ The Raman spectrum of MoS2 nanostructure was carried out at room temperature (Inset Fig. 1(b)). Three distinct
characteristic band positions were observed in the raman spectrum of MoS2 nanostructures i.e. E1g, E2g and A1g
at 296.86, 346.87 and 390 cm−1 respectively. www.nature.com/scientificreports/ The raman band appearing at 296.86 cm−1 with strong intensity (E1g
symmetry), arises due to S atom in the basal plane. Band appearing at 347.86 cm−1 (E2g symmetry) arises due to
intralayer vibrational mode of Mo and S atoms in the basal plane. The A1g mode which appears at 390 cm−1 is
due to the intralayer mode involving the motion of S atoms52, 53. Figure 1(c) depicts the Raman spectrum of 3D
graphene in which two prominent raman bands were clearly visible that corresponds to D and G band at 1357
and 1596 cm−1, respectively54–56. The appearance of D band arises due to the activation of the first order scattering
process of sp3 hybrid carbons, which is attributed to dislocation defects in graphene sheets57. Similarly, the SEM
studies of as-synthesized 3D graphene were carried out, see (Fig. 1(d)). Graphene hydrogels are formed by the
partial overlapping of graphene sheets via hydrophobic and π-π interaction in the 3D space helps it to possess
an interconnected porous network. The elemental analysis of 3D graphene-MoS2 hybrid was carried out by EDX
studies (Fig. 1(e)). Elemental analysis shows that the hybrid contains C, Mo and S elements. In addition, Si ele-
ment was also detected in the EDX studies which appear due to the usage of Si wafer for the elemental detection. Inset of Fig. 1(e) reveals the complete percentage composition of the hybrid materials, where the wt% of C, O, Mo,
S, and Si are 32.32, 23.54, 19.63, 22.51 and 2.06 respectively. For morphological characterization of 3D graphene-
MoS2 hybrid, SEM studies were carried out. 2 y
Figure 1(f) shows that the MoS2 nanospheres, as well as 3D graphene, were successfully deposited over graph-
ite. Inset of Fig. 1(f) shows the flaky nature of MoS2. SEM images clearly indicate that the MoS2 has completely
formed a hybrid network with 3D graphene. Figure 1(g,f) displays the TEM images of 3D graphene-MoS2 hybrid
nanomaterial revealing the flower like morphology of MoS2 nanostructure decorated on 3D graphene hybrid and
is fully consistent with SEM results.h y
The MoS2 nanosheets along with 3D-Graphene will noticeably improve the number of the exposed electro-
chemically active sites, which will considerably enhance the ion diffusion efficacy during the reversible electro-
chemical reactions58. www.nature.com/scientificreports/ There is a possibility to enhance the energy density of supercapacitor by tuning the porosity
of the active material which should be lower than the hydrodynamic size of the active ion or equivalent or higher
than the desolvated ions6. This porous nature of the active material will help to reduce the rate of discharg-
ing hereby enhancing the energy density of supercapacitor. Further deep insight of the nanomaterial (Fig. 1(h))
demonstrate the 3D graphene-MoS2 interface and the characteristic lattice fringes, shown in HRTEM image,
corresponds to 0.65 nm of MoS2 and 0.34 nm of graphene respectively, that are attributed to the (002) planes of
MoS2 and multilayer graphene59. Cyclic Voltammetry studies of MoS2, 3D graphene and hybrid in solution phase. Figure 2(a)
displays the cyclic voltammetry response of MoS2 modified glassy carbon electrode in 0.1 M KOH solution from
low (10 mV/s) to higher (300 mV/S) scan rate. From the CV it is clear that no significant redox peak was observed
which reveals that MoS2 nanostructure behaves as typical electrical double layer capacitance which is similar to
already published in the literature41, 48, 60. Also the working potential of MoS2 nanoflowers were quite high i.e. −0.9 to 0.2 i.e. 1.1 V of working potential was observed. The specific capacitance of MoS2 was calculated using
given equation: IdV
V
m
(1)
∫
υ × ∆
× (1) where I is the current, ʋ the scan rate ΔV is the working potential window and m is the active mass of the material
present on the surface of the electrode61. By this equation, the specific capacitance of MoS2 nanostructure comes
out to be 145.73F/g (100 mV/s).h where I is the current, ʋ the scan rate ΔV is the working potential window and m is the active mass of the material
present on the surface of the electrode61. By this equation, the specific capacitance of MoS2 nanostructure comes
out to be 145.73F/g (100 mV/s).h g
The energy density of the present system was calculated by using equation62: g
The energy density of the present system was calculated by using equation62: =
∆
E
C
V /2
(2)
2 (2) where E is the energy density, C is specific capacitance, ΔV is the working potential. To calculate the power den-
sity equation 3 was used ref. www.nature.com/scientificreports/ (a) Cyclic voltammogram of MoS2 modified electrode in the presence of 0.1 M KOH solution in
low scan rate (10–100 mV/s), (b) Cyclic Voltammograms of MoS2 modified electrode at higher scan rates
(100–300 mV/s). (c) Galvanostatic charge/discharge curves of MoS2 modified electrode in 0.1 M KOH solution. (d) Comparison of 3D graphene, MoS2 and 3D graphene-MoS2 hybrid response over 1 × 1 cm2 graphite piece in
0.1 M KOH. good charging and discharging behavior. For further calculation of specific capacitance by charge-discharge scan
Fig. 2(c), following relation was used: C
I
t m
V
/
(4)
sp = ∆
∆ (4) where I is the discharge current (mA), Δt is the discharge time (s), m is the mass of the electro-active materia
mg) and ΔV is the potential window.hi g
p
The specific capacitance obtained by from CV curves is about 169.37, 95.66, 62.52 and 46.17F/g for 1, 2, 3 and
4 A/g respectively. This observed specific capacitance shows a significant improvement over previously reported
values65 and is in good agreement with the specific capacitance using equation (1) 40, 48. For comparison, the
supercapacitor performance of 3D graphene, graphite and MoS2 have been explored over 1 × 1 cm2 graphite sheet
(Fig. 2(d)). Green, red and black line denotes the response of graphite, 3D graphene and hybrid material (3D
graphene and MoS2) in 0.1 M KOH. By comparing their respective cyclic voltammograms, it is observed that the
hybrid material not only increases the working potential window but also increases the current response, Csp and
energy density for graphite, 3D graphene over graphite and of 3D graphene-MoS2 hybrid comes out to be 7.83,
46.82, 58.0 F/g and 1.09, 13.0, 24.59 Wh/Kg, respectively which shows that the hyrid material has more superca-
pacitive performance as compared to both 3D graphene and graphite material. The complete values of specific
capacitance, energy density, and power density was given in Tables 1 and 2 of Supplementary Information. Solid state supercapacitor studies. Figure 3 displays the complete schematic of fabrication of solid state
device. For synthesis of solid state device first the graphite sheet of dimension 2.2 × 2.2 cm2 was scotch taped and
then put in teflon line stainless steel vessel, 5 mg/mL solution of graphene oxide was added along with 2 M ascor-
bic acid solution and afterwards hydrothermal was sealed. www.nature.com/scientificreports/ 19: (3) P
E t/
(3)
= P
E t/
= where P is the power density and t is the discharge time. where P is the power density and t is the discharge time. b d
l
h
l
l where P is the power density and t is the discharge time. p
y
g
Figure 2(b) displays the cyclic voltammetry response of MoS2 nanospheres in 0.1 M KOH solution at various
scan rates. As seen from the figure the approximately rectangular shape with high symmetry is an indicator to
double layer capacitance with fast charge-discharge rates. These results i.e. electrical double layer formation, are
in good agreement with the previously published literature47, 48, 63. For various scan rates i.e. 100, 150, 200, 250,
300 mV/s, the specific capacitance using equation (1) comes out to be 145.73, 124.31, 116.58, 110.39, 104.79 F/g
respectively. With the increase in scan rate, the current density increases revealing the existence of ideal super-
capacitor behaviour and specific capacitance decreases thus indicating that at lower scan rates the electrolyte
ions diffuse into the inner layers of the active material quite easily, leading to higher interactions with the active
sites of the active material for the charge transfer (Fig. S1 in Supplementary)64. Effect of scan rate on the energy
density was displayed in Fig. S2 of Supplementary. These results indicate that MoS2 is a very important candidate
for supercapacitor applications. Further for specific capacitance calculation charge discharge was carried out in
between the potentials of −0.8 to 0.2 V (Fig. 2(c)). From charge discharge studies, it is observed that the charging
time for MoS2 nanoflowers is ~200 s and for discharging it is ~190 s which reveals the MoS2 nanoflowers show Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 4 www.nature.com/scientificreports/ Figure 2. (a) Cyclic voltammogram of MoS2 modified electrode in the presence of 0.1 M KOH solution in
low scan rate (10–100 mV/s), (b) Cyclic Voltammograms of MoS2 modified electrode at higher scan rates
(100–300 mV/s). (c) Galvanostatic charge/discharge curves of MoS2 modified electrode in 0.1 M KOH solution. (d) Comparison of 3D graphene, MoS2 and 3D graphene-MoS2 hybrid response over 1 × 1 cm2 graphite piece in
0.1 M KOH. Figure 2. www.nature.com/scientificreports/ www.nature.com/scientificreports/ Nanomaterial
Energy density
Power density
Capacitance (F/g)
Reference
Co3O4
80 Wh/Kg
20 W/Kg
3560
14
RuO2.nH2O
10.62 WhJ/g
4.456 kW/Kg
570
71
BCN
244
72
Carbon with PHEMA-co-TMPA
—
—
130
73
CNT and PEDOT
8.85 mWh/cm3
9.4 W/cm3
0.354
74
Carbon cloth with phosphoric acid doped
polybenzimidazole
10 Wh/kg
300 W/kg
290
75
High specific surface Area (SSA) carbon
and poly [2,5-benzimidazole]
—
—
248
76
polyaniline and sulfonated polymers
—
—
98
77
Graphene
82 kW/kg
32.3 W/Kg
98
78
rGO
34 µWh/cm2
56.11
79
3D graphene-MoS2 hybrid
24.59 Wh/Kg
8.8 W/kg
58 F/g
Present work
Table 1. Comparative studies of recently reported solid state supercapacitor utilizing various electroactive
materials. Table 1. Comparative studies of recently reported solid state supercapacitor utilizing various electroactive
materials. Successful implementation of the supercapacitor in solid-state without a loss in performance can be inferred. Figure 4(a) depicts comparative cyclic voltammetry curves of the fabricated solid-state supercapacitor as well
as the hybrid electroactive material in solution. This reveals that area under the curve almost doubles due to
the contribution of two parallel electrode plates in the device with large potential window ranging from -1.5 V
to 1.2 V. It can be clearly understood, that the solid-state supercapacitor is performing quite well as compared
to the hybrid material in solution. The potential window as well as the area under the curve has increased sig-
nificantly in solid-state devices, due to the combination of two electrode parallel plates, thus indicating that the
hybrid materials shows potential development of supercapacitive performance over 3D-graphene and graphite
based electroactive materials. Specific capacitance for 3D graphene-MoS2 comes out to be 74.94F/g and the spe-
cific capacitance of 3D graphene-MoS2 hybrid in solid state device comes out to be 58.0F/g (using equation 1). Similarly using equation (2) energy density comes out to be 15.77 and 24.59 Wh/kg for solution and solid state
device, respectively. The performance of the hybrid material over graphite plate at various scan rates was also
studied, shown in Fig. 4(b). Similar to MoS2, no oxidation reduction peak appeared in the device indicating the
existence of pure EDLC type behavior. The current density of the hybrid material increases with the increase in
scan rate, confirming its ideal nature. A large operating current indicates that the existence of low internal resist-
ance between 3D graphene to graphite electrode. www.nature.com/scientificreports/ This is due to the excellent electrical conductivity of graphene
sheets and the absence of electrical barrier at the 3D graphene-graphite sheet interface via π-π interactions. For
the solid-state supercapacitor, two sheets of 2.2 × 2.2 cm2 graphite, coated with 3D graphene-MoS2 hybrid were
pasted using a filter paper soaked in PVA-KOH gel and kept for drying in a vacuum desiccator. These results
indicate that the performance of the solid-state supercapacitor is quite high as compared to that in solution and
that it can work as a standalone solid-state device. It is observed from the galvanostatic charge-discharge curves
in Fig. 4(c) that the charging and discharging time of the solid-state device is found to be significantly shorter
compared to that of MoS2 nanoflowers (see Fig. 2c). This is suggesting a marked improvement in supercapacitor
performance due to the presence of 3D graphene. In Fig. 4(c), it is also noted that the charging time (~50 s) is
lower than the discharging times (~68 s), emphasizing upon the ideal supercapacitor behavior of the device. Further, at high current density the galvanostatic charge discharge (GCD) curves show typically triangular shape
which is the characteristic of nearly ideal capacitive performance. However, at low (<3 A/g) current density the
deviation of the GCD pattern is attributed possible due to the redox reaction occurs at the dangling bond of the
3D graphene and defect site of the MoS2, which is quite similar to the recent report9, 66, 67. The formation of the
plateau at low current density at high applied voltage (2.7 V) fully opens the stacked 3 D graphene as well as MoS2
nano flaks which allow ion intercalation and creates abundant ion-accessible sites for adsorption/intercalation,
and dramatically increases the cell’s capacitance68.h y
p
The weight calculation of the active material was done by TGA analysis. Figure 4(d) displays the TGA study
of MoS2 3D graphene hybrid over graphite. TGA spectrum shows two weight losses one at ~350 °C and other
at around 500 °C. Weight loss i.e. 5% at 350 °C corresponds to 3D graphene69. While the weight loss i.e. 2.5% at
around 500 °C corresponds to MoS2 nanoflowers59. From the TGA calculation, the weight of the active material
comes out to be 1.1 mg, which has been used to calculate the specific capacitance (using eq. 1) of 58.0F/g. www.nature.com/scientificreports/ The hyrothermal bomb was then kept at 120 °C for 4 h
resulting in the formation of hydrogel of 3D graphene over graphite piece. This modified graphite was kept in
5 mg/mL solution of MoS2. Similiary other plate of supercapacitor was fabricated using same protocol. After this
the two plate was assembled by using filter paper soaked with PVA/KOH solution. Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 5 www.nature.com/scientificreports/ i
To further understand the superior performance of the MoS2-3D graphene hybrid electrode, electrochemical
impedance spectroscopy (EIS) is performed by electroactive materials to analyze the kinetic feature of the ion
transportation and ion diffusion in the porous hybrid electrode in aqueous electrolyte system. Figure 4(e) shows
the Nyquist plot obtained at the frequency range from 100 kHz to 0.1 Hz with different amplitude of single sin
wave (0.005, 0.01, 0.015, and 0.02 V RMS value). The calculated internal resistance (Rs) for MoS2-3D graphene
hybrid is 18.66 ohm. The corresponding equivalent circuit has been shown in Figure S5 in Supplementary
Information. A sharp increase of the impedance plot at low frequencies indicates the capacitive behavior of the
electrode. It is observed that like an ideal double-layer capacitor, the plot comes out to be a vertical line, parallel
to the imaginary axis70. The enchantment in energy density is found to be attributed to the enlarge operating
potential voltage of 2.7 V in the gel electrolyte (PVA with KOH) and the relatively high specific capacitance of
both electrodes, especially the negative electrode, was found which is very important for the solid state devices. Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 6 www.nature.com/scientificreports/ Figure 3. Schematic process flow for the fabrication of device using graphite as current collector. On top of
graphite 3D graphene was grown further which was kept in 5 mg/mL solution of MoS2 of isopropyl alcohol
and finally by using PVA-KOH solution two plates of graphite were assembled to prepare real solid state
supercapacitor. Figure 3. Schematic process flow for the fabrication of device using graphite as current collector. On top of
graphite 3D graphene was grown further which was kept in 5 mg/mL solution of MoS2 of isopropyl alcohol
and finally by using PVA-KOH solution two plates of graphite were assembled to prepare real solid state
supercapacitor. The stability curve strongly indicates that about 90% of the initial capacitance can still be retained after 1400 cycle
at the scan rate of 100 mV/s (Fig. 4f), refers to the superior cycling performance. g
p
y
g p
Figure 5(a) displays the digital image of fabricated solid state device. Finally, the solid state supercapacitor
response has been monitored by using red LED. Four devices were connected in series to monitor the response of
red LED. www.nature.com/scientificreports/ www.nature.com/scientificreports/ Figure 4. Comparison of MoS2-3D graphene hybrid in solution and solid state supercapacitor (b) scan rate
studies of device (c) charge discharge studies. (d) TGA analysis of the hybrid material over graphite. (e) Nyquist
plot for MoS2-3D graphene hybrid, (f) Cycle performance measured at 100 mV/s, and measured loss of stability
is found to be 10% after 1400 cycles. Figure 4. Comparison of MoS2-3D graphene hybrid in solution and solid state supercapacitor (b) scan rate
studies of device (c) charge discharge studies. (d) TGA analysis of the hybrid material over graphite. (e) Nyquist
plot for MoS2-3D graphene hybrid, (f) Cycle performance measured at 100 mV/s, and measured loss of stability
is found to be 10% after 1400 cycles. dying out. The longer discharge time compared to the charging time makes the device suitable for use in practical
energy storage and transfer application.h The Table 1 depicts the comparative statements of recently reported solid state supercapacitors utilizing var-
ious electroactive materials along with our present findings. Although the specific capacitances of the devices
made of active materials like RuO2, high-SSA Carbon and BCN, are quite high but the corresponding energy
density is low that may be due to the shorter range of working potential. However, it is quite challenging to expand
the working potential of the electrode for the practical use of device in real field of application. Common tech-
nique is to utilize the ionic liquid based electrolyte like BMIM, which screens the ionic interaction of ions with
C ≡ N moieties and make them free to move. This may assist the fast ion transport. Thus, expanding the working
potential without introducing ionic liquid can lead to a successful attempt for the development of solid state
supercapacitor. The carbon cloth with phosphoric acid doped polybenzimidazole, Co3O4 and polyaniline with
sulfonated polymers show significant enhancement of Csp due to pseudocapacitance, which is better to avoid for
long term durability of the practical devices. The present report focus on the 3D graphene-MoS2 hybrid for the
development of real solid state supercapacitor which not only works well in negative potential but also expands
significantly in positive working potential. The overall potential window for solid state supercapacitor is quite
high which is about to be 2.7 V (i.e. −1.5 V to +1.2 V) in KOH-PVA medium without utilizing any expensive
ionic liquid mediums. Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 www.nature.com/scientificreports/ The devices were charged with 9 V battery for 25 s.t h
g
y
After that the LED was connected to the devices, the response was monitored through LED glow. With 25 s
of charging the LED glows for 40 s. The video recording is added to the Supplementary data. This reveals that the
present fabrication of the device completely work as solid state supercapacitor. Figure 5(b) displays the charging
of four devices in series connection and the inset represents the response of the LED after charging of the solid
state supercapacitor. The practical applicability of the device is demonstrated by the lighting of a red LED bulb. Four devices of the exact same dimensions and similar mass loading were connected in series and charged by a
9 V battery for 25 s. On connecting the device array to the LED, it remained lit up for an impressive 40 s before Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 7 Conclusions MoS2 was successfully paired with 3D graphene to fabricate a flexible, solid-state supercapacitor with outstanding
performance. The facile method of fabrication can easily be scaled up at minimal cost to create a large array of
supercapacitors which is lightweight and reliable. An immediate application can be found in solar energy storage
due to its large energy storage capacity in a short span of time in purely EDLC type nature. The specific capac-
itance of the solid state supercapacitor comes out to be 58 F/g with energy density of 24.59 Wh/Kg and power
density of 8.8 W/kg. It can be a partial replacement of conventional batteries and can be easily incorporated into
flexible and thin electronics due to its high stability, thinness and flexibility. The operating potential window can
be further enhanced by using an ionic-liquid gel-based electrolyte. Other transition metal dichalcogenides can
also be investigated along with MoS2 for their supercapacitor performance on top of 3D graphene matrix. www.nature.com/scientificreports/ Recently a solid state architecture has been designed using a similar heterostructure66,
however a costlier growth technique has been adopted where for the graphene growth was performed in 1000 °C
in CVD setup. In contrast our study shows a very simpler and cost effective process via chemical route to design
heterostructure. In comparison to the device performance, they report the specific capacitance of 19.44 F/cm3 in
contrast to our device as 58.0 F/g. Here, it is noticed that the straight forward comparison is not feasible because
the analytical approaches are entirely different. 8 Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 www.nature.com/scientificreports/ www.nature.com/scientificreports/ Figure 5. (a) Digital image of the single device. (b) Four devices fabricated and connected in series. (c) Digital
image of the four devices connected in series (Charging for 25 s), Inset figure digital image of LED glow
(Discharging for 40 s). Figure 5. (a) Digital image of the single device. (b) Four devices fabricated and connected in series. (c) Digital
image of the four devices connected in series (Charging for 25 s), Inset figure digital image of LED glow
(Discharging for 40 s). Experimental Section
i l Materials. Graphite, ammonium molybdate, ascorbic acid, isopropyl alcohol, thiourea were purchased from
sigma aldrich. All the solutions were prepared in deionized water. KOH was purchased from TCI chemicals. Synthetic and fabrication procedures. Synthesis and Purification of MoS2. The MoS2 nanoflowers were
synthesized by a hydrothermal method using ammonium molybdate and thiourea as a starting precursor. Briefly,
1 M solution of ammonium molybdate and 5 M solution of thiourea were prepared separately and transferred into
a teflon-lined stainless steel autoclave (100 mL capacity) and hydrothermal reaction was carried out at 180 °C for
24 h. Black precipitate was obtained followed by washing with deionized water and absolute ethanol. The washing
procedure is repeated several times until the final product is free of any trace amount of residual ions. The final
product was dried at 80 °C for 12 h. Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 9 www.nature.com/scientificreports/ Modification of the electrode. The as-synthesized MoS2 nanospheres were coated on a polished GC electrode
surface (GC, diameter = 2 mm) to modify the Glassy Carbon (GC) electrode. For the coating, a solution of 1 mg/
mL of MoS2 was prepared in isopropyl alcohol from which 5 µL was drop casted over the GC electrode surface. The prepared electrode was kept in a vacuum desiccator for 24 h to form a uniform layer over the surface. Cyclic
voltammetry was carried out using a CHI electrochemical workstation with a three electrode system. While the
MoS2-modified electrode was the working electrode, Ag/AgCl in sat. KCl was used as the reference electrode and
platinum electrode was the counter electrode. These were dipped in 0.1 M KOH electrolyte. Preparation of square shaped graphite electrodes. We followed the modified hydrothermal reduction method
for the deposition of 3D graphene onto the graphite electrodes. Briefly, two rectangular pieces were taken from a
graphite sheet and they were dipped into 10 mL of 5 mg/mL aqueous dispersion of graphene oxide (GO). This was
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poly [2,5-benzimidazole] for high temperature application. Electrochim. Acta 59, 296–303 (2012). Scientific REPorTS | 7: 9458 | DOI:10.1038/s41598-017-09266-2 Additional Information Supplementary information accompanies this paper at doi:10.1038/s41598-017-09266-2h Supplementary information accompanies this paper at doi:10.1038/s41598-017-09266-2h Competing Interests: The authors declare that they have no competing interests. Competing Interests: The authors declare that they have no competing interests. Competing Interests: The authors declare that they have no competing interests. Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and
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Determination of recent tectonic deformations in the vicinity of Adana–Osmaniye–Hatay–Gaziantep triple junction region by half-space modeling
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Comptes rendus. Géoscience/Comptes rendus. Géoscience
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cc-by
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Géoscience
Sciences de la Planète Selim Serhan Yıldız, Ali Özkan, Hasan Hakan Yava¸so˘glu, Frédéric
Masson, ˙Ibrahim Tiryakio˘glu, Mehmet Nurullah Alkan and Serdar
Bilgi ∗Corresponding author. Determination of recent tectonic deformations in the vicinity of
Adana–Osmaniye–Hatay–Gaziantep triple junction region by half-space
modeling
Volume 352, issue 3 (2020), p. 223-232
<https://doi.org/10.5802/crgeos.39> Volume 352, issue 3 (2020), p. 223-232 Les Comptes Rendus. Géoscience — Sciences de la Planète sont membres du
Centre Mersenne pour l’édition scientifique ouverte
www.centre-mersenne.org Les Comptes Rendus. Géoscience — Sciences de la Planète sont membres du
Centre Mersenne pour l’édition scientifique ouverte
www.centre-mersenne.org Comptes Rendus
Géoscience — Sciences de la Planète
2020, 352, no 3, p. 223-232
https://doi.org/10.5802/crgeos.39 Comptes Rendus
Géoscience — Sciences de la Planète
2020, 352, no 3, p. 223-232
https://doi.org/10.5802/crgeos.39 https://comptes-rendus.academie-sciences.fr/geoscience/ 1. Introduction erating earthquakes around the southern end of the
EAFZ. Accordingly, we intend to estimate the recent
slip rates along these faults and fault locking param-
eters with high accuracy and spatial resolution by
modeling the observed GPS data. For this purpose,
campaign observation sites were used along profiles
perpendicular to the related faults. A wide geodetic
network consisting of campaign observation sites to-
gether with permanent GPS stations was observed
during the execution of the above proposed study. Raw data obtained from previous studies related to
geodesy, paleoseismology, and active tectonics were
analyzed together with the latest GPS observations to
estimate current strain accumulations for the East-
ern Mediterranean region. By analyzing all the ob-
tained data, slip rates along the faults and fault lock-
ing depths were studied spatially and temporally in
detail. The East Anatolian Fault Zone (EAFZ), one of the
most important neotectonic and seismogenic fault
systems in Turkey, is a sinistral strike-slip active fault
zone extending from Karliova in the east of Erzincan
to the Hatay region, where it reaches the Dead Sea
Fault Zone further south [Arpat and Saroglu, 1972,
Westaway and Arger, 1996]. The EAFZ has intense
seismic activity depending on the northward rela-
tive motions of Arabian and African plates, the west-
ward motion of the Anatolian plate, and continental
collision occurring along the Bitlis–Zagros Fold and
Thrust Belt [Saroglu and Yilmaz, 1990]. In terms of seismicity, the Karasu Fault (KF;
Amanous
Fault)
segment
of
the
EAFZ
and
Toprakkale, Karatas, and Yumurtalık Faults are con-
siderable fault segments within the complex struc-
ture of the northeast Mediterranean region. These
faults were evaluated as active faults on the Active
Fault Map of Turkey updated recently [Emre et al.,
2018]. On the other hand, the fact that few geodetic
studies conducted to investigate the kinematics of
these active faults makes it difficult to understand
the structure of these faults and analyze earthquake
hazards [Aktu˘g et al., 2016, Mahmoud et al., 2012,
Nocquet, 2012, Meghraoui et al., 2011, Reilinger
et al., 2006]. To estimate the potential hazard of a
probable earthquake, there is a need to study the slip
rates along these active faults and also fault lock-
ing parameters using geodetic methods. Determination of recent tectonic deformations in
the vicinity of Adana–Osmaniye–Hatay–Gaziantep
triple junction region by half-space modeling The Karatas–Osmaniye Fault has a 3.4 mm/yr dextral strike-slip rate together with a reverse-slip rate
of 3.1 mm/yr, which corresponds to a possible magnitude of 6.8–7.1 for an earthquake on the south-
west end of the East Anatolian Fault. The Karasu Fault extends from the Turkoglu Triple Junction on
the EAFZ in the north to the Hatay Triple Junction in the south. This critical fault segment has a sinis-
tral 4.4–5.4 mm/yr slip rate with a normal component slip rate of 3.0 mm/yr, supporting earthquake
prediction with a possible magnitude of 6.8–7.2. As a result, the determination of such kinematic indi-
cations in active tectonic zones by GPS observations plays a key role in seismic hazard analysis for the
Eastern Mediterranean region. Keywords. Active tectonics, GPS, East Anatolian Fault Zone, Slip rate, Fault locking depth. Manuscript received 28th May 2020, revised 6th October 2020, accepted 13th October 2020. Determination of recent tectonic deformations in
the vicinity of Adana–Osmaniye–Hatay–Gaziantep
triple junction region by half-space modeling Selim Serhan Yıldıza, Ali Özkan∗, b, Hasan Hakan Yava¸so˘gluc, Frédéric Massond,
˙Ibrahim Tiryakio˘glue, Mehmet Nurullah Alkanf and Serdar Bilgic a Department of Geomatics Engineering, Osmaniye Korkut Ata University, Osmaniye,
Turkey b Osmaniye Vocational School, Osmaniye Korkut Ata University, Osmaniye, Turkey c Department of Geomatics Engineering, Istanbul Technical University, Istanbul,
Turkey d Institut de Physique du Globe, UMR 7516, Université de Strasbourg/EOST CNRS,
Strasbourg, France e Department of Geomatics Engineering, Afyon Kocatepe University, Afyonkarahisar,
Turkey f Osmancık Vocational School, Hitit University, Corum, Turkey E-mails: serhan@osmaniye.edu.tr (S. Serhan Yıldız), aliozkan@osmaniye.edu.tr
(A. Özkan), yavasoglu@itu.edu.tr (H. Hakan Yava¸so˘glu), frederic.masson@unistra.fr (F. Masson), itiryakioglu@gmail.com (˙I. Tiryakio˘glu), nurullahalkan@hitit.edu.tr (M. Nurullah Alkan), bilgi@itu.edu.tr (S. Bilgi) Abstract. Recent earthquakes on the East Anatolian Fault Zone (EAFZ) and its surroundings in the
Eastern Mediterranean region reveal a potential seismic hazard. It is thus needed to constrain defor-
mations within tectonically active zones and clarify kinematics of the triple junction near Adana, Os-
maniye, Hatay, and Gaziantep provinces. To understand strain accumulations and interseismic pat-
terns of these active zones, one of the most popular geodetic methods was applied. Slip rates derived
from Global Positioning System (GPS) observations by half-space modeling lead us to predict possible
magnitudes and even probable locations of future destructive earthquakes. In this context, we form
a region-specific GPS network consisting of both campaign observation sites and permanent stations
to monitor strain accumulations across major active faults and investigate fault kinematics in detail. The campaign GPS data sets acquired in 2009, 2010, 2011, and 2019 were merged with data from dif-
ferent local permanent GPS networks for an integrated analysis of the determination of recent tec-
tonic deformations. The TDEFNODE software was used to construct a kinematic model in an elastic
and homogeneous half-space. The modeling approach needs inputs such as block and fault geome-
tries, GPS site velocities, and locking depth assumptions. Our kinematic model indicates that an al-
most purely sinistral strike-slip rate of 7.5 mm/yr without any significant normal or reverse compo-
nent is dominant on the main branch of the EAFZ. This estimated slip rate supports a prediction for a ISSN (electronic) : 1778-7025 https://comptes-rendus.academie-sciences.fr/geoscience/ 224 Selim Serhan Yıldız et al. magnitude margin of 7.2–7.6 for the next probable large earthquake on the Turkoglu–Golbasi segment. C. R. Géoscience — 2020, 352, no 3, 223-232 2. Kinematic setting Since the motions of tectonic plates in the Earth’s
lithosphere are defined in accordance with plate tec-
tonics theory, it is believed that the Arabian, Anato-
lian, and Sinai plates interact around the region of
the Eastern Mediterranean. According to this hypoth-
esis, the Arabian plate moves to the north at an ap-
proximate rate of 20 mm/yr and the Sinai plate con-
verges to a subduction zone along the Cyprus Arc
at approximately 10 mm/yr. Furthermore, the west-
ward motion of the Anatolian plate is accommodated
by a sinistral strike slip along the EAFZ and a dex-
tral strike slip along the North Anatolian Fault Zone
at rates of 10 mm/yr and 20–25 mm/yr, respectively
[Reilinger et al., 1997, McClusky et al., 2000, Reilinger
et al., 2006, Mahmoud et al., 2012, Aktu˘g et al., 2016]. Considering the role of geodetic techniques in
active tectonics, it is now possible to estimate the
slip rates more precisely along active faults. The slip
rates for EAFZ segments are estimated in different
geodetic studies as 15 ± 3 mm/yr [Reilinger et al.,
1997] for the whole EAFZ and 10.0 ± 0.3 mm/yr for
the Palu–Hazar Lake segment and 9.9 ± 0.2 mm/yr
for the Hazar Lake–Turkoglu segment along the EAFZ
[Reilinger et al., 2006]. The slip rates are calcu-
lated to be 9.7 ± 0.9 mm/yr for EAFZ and 5.5 ±
1.5 mm/yr for the Karatas–Osmaniye Fault (KOF) seg-
ment [Bertrand et al., 2006]. Since the motions of tectonic plates in the Earth’s
lithosphere are defined in accordance with plate tec-
tonics theory, it is believed that the Arabian, Anato-
lian, and Sinai plates interact around the region of
the Eastern Mediterranean. According to this hypoth-
esis, the Arabian plate moves to the north at an ap-
proximate rate of 20 mm/yr and the Sinai plate con-
verges to a subduction zone along the Cyprus Arc
at approximately 10 mm/yr. Furthermore, the west-
ward motion of the Anatolian plate is accommodated
by a sinistral strike slip along the EAFZ and a dex-
tral strike slip along the North Anatolian Fault Zone
at rates of 10 mm/yr and 20–25 mm/yr, respectively
[Reilinger et al., 1997, McClusky et al., 2000, Reilinger
et al., 2006, Mahmoud et al., 2012, Aktu˘g et al., 2016]. Figure 1. GPS network in the study area. 1. Introduction However,
the use of permanent Global Positioning System
(GPS) stations together with campaign GPS obser-
vations is the most efficient method for this pur-
pose, but the geometry and distinctive features of
relevant faults are important criteria for geodetic
networks. During the past decades, strain accumulations on
the EAFZ have sometimes caused life-threatening
earthquakes in eastern Turkey such as Elazig–
Karakocan (Mw 6.1, 2010) and Bingol (Mw 6.4, 2003)
earthquakes. The last destructive earthquake on the
EAFZ occurred with an epicenter close to the town of
Sivrice in Elazig province on the Puturge segment of
the EAFZ in January 2020. The earthquake solution
published by the United States Geological Survey
(USGS) is a magnitude of Mw 6.7 and a depth of
10.0 km. However, seismic hazard analysis may pre-
vent substantial loss of life and property associated
with such relatively major earthquakes. In this con-
text, the potential moment magnitude for a possible
earthquake within the modeled segment of an active
fault can be predicted using empirical relationships. These involve either total displacement considering
historical and instrumental seismic data [Wells and
Coppersmith, 1994] or fault slip rate together with
the length of fault rupture [Anderson et al., 1996]. This study was conceived with the aim of mon-
itoring active faults with a high probability of gen- C. R. Géoscience — 2020, 352, no 3, 223-232 Selim Serhan Yıldız et al. 225 Figure 1. GPS network in the study area. Ab-
breviations for faults are EAF (East Anatolian
Fault), DSF (Dead Sea Fault), KF (Karasu Fault),
KOF (Karatas–Osmaniye Fault), and CA (Cyprus
Arc). Fault mapping from Meghraoui et al. [2011]. Orange squares show provinces in the
region. Since the predicted moment magnitude is converted
into average displacement with the help of the ap-
proach proposed by Wells and Coppersmith [1994],
the recurrence interval of an earthquake can be cal-
culated. From this perspective, the determination of
tectonic deformations in active fault zones within the
Adana, Osmaniye, Hatay, and Gaziantep provinces is
very crucial in terms of drawing attention to seismic
hazards in the Eastern Mediterranean region. 3. GPS data set and analysis Using the GPS network presented in Section 2,
campaign GPS observations were carried out at
24 campaign sites for four campaign sets in 2009,
2010, 2011, and 2019. Since 4 of the 24 campaign
sites were destroyed, the last campaign in 2019 was
achieved with the remaining 20 sites. The strategy
adopted during campaign GPS observations is based
on data logging in two sessions on consecutive days. In addition, each session covers an observation span
of at least 8 hours. All campaign observations were
performed within the same time periods of years to
mitigate seasonal effects on GPS observations. gf
The GPS data analysis was performed in the
GAMIT/GLOBK software [King et al., 2009] using
validated strategies [Reilinger et al., 2006] for both
campaign GPS observations and for also data ob-
tained from municipalities, Turkish National Per-
manent GNSS Network-Active, and IGS permanent
GNSS stations. First, in the GAMIT stage, station co-
ordinates, zenith delay of the atmosphere at each
station, and orbital and Earth orientation parame-
ters (EOPs) were estimated from daily GPS phase ob-
servations using a weighted least squares algorithm. Second, in the GLOBK stage, Kalman filtering was ap-
plied to loosely constrained estimates of station co-
ordinates, orbits, and EOPs and their covariances for
a reference frame definition with a consistent set of
coordinates and velocities. Eighteen stations selected
from the IGS global networks were incorporated both
to tie our local network with a global network and
to estimate the orbital and earth rotation param-
eters with high accuracy. Apart from IGS stations,
a total of 30 permanent GPS stations from local Following the procedures for the definition of
the reference frame for both coordinates and ve-
locities [Aktu˘g et al., 2009], transformation parame-
ters to ITRF2008 coordinates of selected IGS stations
were estimated for the stabilization frame. The sta-
bilization was realized for the Eurasia-fixed reference
frame by minimizing the adjustments to the horizon-
tal velocities of the 17 stations where these velocities
were obtained with a post root mean square (rms)
value of 0.31 mm/yr. The GPS velocity field obtained from this study
was combined individually with published velocities
while introducing the velocities by Reilinger et al. [2006] as the reference solution. The transformation
accuracy of the velocity field solutions was expressed
by the wrms parameter depending on the common
observation sites (Table 1). 2. Kinematic setting Ab-
breviations for faults are EAF (East Anatolian
Fault), DSF (Dead Sea Fault), KF (Karasu Fault),
KOF (Karatas–Osmaniye Fault), and CA (Cyprus
Arc). Fault mapping from Meghraoui et al. [2011]. Orange squares show provinces in the
region. Considering the role of geodetic techniques in
active tectonics, it is now possible to estimate the
slip rates more precisely along active faults. The slip
rates for EAFZ segments are estimated in different
geodetic studies as 15 ± 3 mm/yr [Reilinger et al.,
1997] for the whole EAFZ and 10.0 ± 0.3 mm/yr for
the Palu–Hazar Lake segment and 9.9 ± 0.2 mm/yr
for the Hazar Lake–Turkoglu segment along the EAFZ
[Reilinger et al., 2006]. The slip rates are calcu-
lated to be 9.7 ± 0.9 mm/yr for EAFZ and 5.5 ±
1.5 mm/yr for the Karatas–Osmaniye Fault (KOF) seg-
ment [Bertrand et al., 2006]. Tectonic processes in the Eastern Mediterranean
region trigger seismic activities around the area. This
is why active fault zones generating seismic haz-
ards need to be monitored continuously using geo-
detic techniques (Figure 1). Some well-documented
destructive historical seismic activities indicate po-
tential risks in seismic zones, namely between the
Turkoglu–Golbasi segment of the EAFZ and the KOF
(1114 AD earthquake, Ms > 7.8), northern segment
of the Dead Sea Fault (DSF) (1408 AD earthquake,
M 7.4), KOF (1513 AD earthquake, M ∼7.4), and KF
(1822 AD earthquake, M 7.0 and 1872 AD earthquake,
M 7.2) [Ambraseys and Jackson, 1998, Sbeinati et al.,
2005, Ambraseys, 2004]. More recently, one of the GPS block modeling
studies about slip rate estimations has reported the
results as 8.8 ± 0.3 mm/yr for the EAFZ and 3.6 ±
0.6 mm/yr for the KOF segment [Meghraoui et al.,
2011]. However, Aktu˘g et al. [2016] obtained slip rates
ranging from 10.3±0.7 mm/yr to 13.5±1.3 mm/yr for
the segments between Turkoglu and Karliova. Addi-
tionally, the rates were indicated to be 4.5±1.1 mm/yr
for the KF and 2.7 ± 1.4 mm/yr along the KOF in the
same study. C. R. Géoscience — 2020, 352, no 3, 223-232 226 Selim Serhan Yıldız et al. Table 1. Transformation results for GPS velocity field solutions
Solution
wrms (mm/yr)
No. of common observation sites
This study
0.76
97
Aktu˘g et al. [2013]
0.67
5
Mahmoud et al. [2012]
0.57
6
Al-Tarazi et al. [2011]
0.72
20
Alchalbi et al. [2010]
0.74
10
Le Béon et al. 2. Kinematic setting [2008]
0.71
32
Gomez et al. [2007]
0.77
5
Reference solution: Reilinger et al. [2006] Table 1. Transformation results for GPS velocity field solutions municipalities and TUSAGA-Aktif networks were in-
troduced into the daily GAMIT processing. Loosely
constrained daily solutions from GAMIT processing
were investigated on expected normalized root mean
square (nrms) values between 0.15 and 0.25. After
discarding outliers, coordinate repeatabilities were
generated by controlling weighted root mean square
(wrms) parameters approximately 2–4 mm for hori-
zontal components and 10–15 mm for vertical com-
ponents. Since the monumentation of GPS stations
fluctuates [Langbein and Johnson, 1997], a random
walk of 2 mm/pyr in horizontal positions of the sta-
tions was applied to obtain more realistic uncertain-
ties for GPS velocities [McClusky et al., 2000]. C. R. Géoscience — 2020, 352, no 3, 223-232 3. GPS data set and analysis The combined velocity
field obtained from the transformation model was
referenced to the ITRF2000 Eurasia-fixed reference
frame (Figure 2). 227 Selim Serhan Yıldız et al. tors. The block modeling approach on the basis of
inversion of GPS velocities provides the estimation
of block rotation rates, fault locking parameters, and
internal strain rates in an elastic and homogeneous
half-space [Okada, 1985]. The components of the
strain-rate tensor aim to reveal internal deformations
within the blocks that are caused by unsettled faults. Apart from GPS site velocities, block and fault geome-
tries formed in a three-dimensional (3D) elastic half-
space, euler poles for blocks and locking depths are
considered as model inputs for parameter estimation
in the inversion. However, defects in block modeling
emerge from poor density of observation sites. In this
case, slip rate estimations become more susceptible
to introduced block geometries and a priori locking
depths. The TDEFNODE software [McCaffrey, 2009]
based on the simulated annealing method [Press
et al., 1986] was used for block modeling by minimiz-
ing the residuals between observed and model veloc-
ities. According to the approach in McCaffrey [2002],
minimizing the misfit of the velocities allows the si-
multaneous estimation of block rotations using Euler
poles and coupling fractions along block boundaries. The data misfit minimization is achieved by reduced
chi-square statistic tests. Figure 2. Eastern Mediterranean GPS velocity
field in the ITRF2000 Eurasia-fixed reference
frame with 95% confidence ellipses (fault map-
ping from GEM GAF-DB). The locking depth in a seismogenic zone roughly
reflects the characteristic mechanism of the fault. The state of being locked or creeping for a fault seg-
ment is controlled by coupling fraction. This parame-
ter denoted by phi (ϕ) depends on the short-term slip
rate (Vc) and the plate velocity (V ), where ϕ = 0 cor-
responds to creeping and ϕ = 1 to a fully locked fault
while partially locked faults have values between 0
and 1 as per (1). Figure 2. Eastern Mediterranean GPS velocity
field in the ITRF2000 Eurasia-fixed reference
frame with 95% confidence ellipses (fault map-
ping from GEM GAF-DB). ϕ = 1−Vc
V . (1) (1) C. R. Géoscience — 2020, 352, no 3, 223-232 4. Kinematic modeling In a deeper layer below
the locked zone, the fault is assumed to be locked
partially down to the depth below where full creep-
ing begins. In our model, the faults were introduced
as not creeping with a constraint of ϕ = 1 from 0
to 7 km in the zone close to the surface. Following
this, ϕ parameters were estimated between 0 and 1
in the inversion for depths ranging from 7 km to the
fully creeping limit. Below the partially locked zone,
the faults creep fully with ϕ = 0. Between locked and
fully creeping zones, which is called the “effective
transition zone” by Wang et al. [2003], the down dip
transition was estimated using a factor of λ = 0.2
based on an exponential function, which indicates
the shape of the slip distribution. During the inver-
sion process, along-strike smoothing was introduced
using a penalty function with a smoothing factor re-
ferred to above. No special data weighting strategy
was followed since all the velocity solutions in model-
ing were equally weighted. However, the velocity field
published by Mahmoud et al. [2012] was not incorpo-
rated into the kinematic model since the site veloc-
ity uncertainties on average are greater than those in
each independent solution. Figure 3. Fault geometry for Eastern Mediter-
ranean region (red and black vectors repre-
sent the model and observed velocities with
95% confidence ellipses, respectively). Pur-
ple lines are fault segments and blue circles
are nodes connecting fault segments to each
other. Orange boxes denote block names de-
fined in the model. EAF (East Anatolian Fault),
KOF (Karatas–Osmaniye Fault), DSF (Dead
Sea Fault), CA (Cyprus Arc), N-AMNS (North
Boundary of Amanous Block), E-AMNS (East
Boundary of Amanous Block), and S-AMNS
(South Boundary of Amanous Block). kinematic models from Mahmoud [2012] were con-
sidered. However, evaluating the statistical results
of the inversion, the geometry of the proposed
kinematic model fits better with observations. The
simplest model with the blocks of arab, sina, and
anat comprises EAF, KF, CA, and DSF. The reduced
chi-square parameter of this simplest model was
estimated as 4.816 with a wrms of 1.476 mm and an
nrms of 2.124. The more complex model consists of
one additional block and one fault segment com-
pared to the simplest model. 4. Kinematic modeling Before inversion, fault geometries are considered
along the strike and dip directions as all the faults
introduced in the kinematic model reach a depth
of 30 km downward in almost vertical planes. The
nodes along strike and down dip surrounding the
blocks in the model define the fault surfaces in 3D
with horizontal positions in degrees and depth in
kilometers. Since the fault surface is composed of
patches (quadrilaterals) between nodes, the kine-
matic model herein assembles patches bounded by
2 × 2 km along the strike and dip directions on
the faults to estimate strain accumulations in the
region. Kinematic modeling relying on GPS velocity fields
instead of interpreting site velocities individually
strengthens tectonic analysis in the region. Blocks
bounded by faults have different characteristics in-
side the mass and along their boundaries. Although
the block motions are systematic, the resistance due
to the friction of rocks prevents free movement at
their boundaries. This velocity difference, known as
“slip deficit”, causes strain accumulations generally
ending in an earthquake. Slip deficit describes the
divergence of surface velocities from expected ve-
locities of the blocks depending on geological fac- 228 Selim Serhan Yıldız et al. Figure 3. Fault geometry for Eastern Mediter-
ranean region (red and black vectors repre-
sent the model and observed velocities with
95% confidence ellipses, respectively). Pur-
ple lines are fault segments and blue circles
are nodes connecting fault segments to each
other. Orange boxes denote block names de-
fined in the model. EAF (East Anatolian Fault),
KOF (Karatas–Osmaniye Fault), DSF (Dead
Sea Fault), CA (Cyprus Arc), N-AMNS (North
Boundary of Amanous Block), E-AMNS (East
Boundary of Amanous Block), and S-AMNS
(South Boundary of Amanous Block). The kinematic model consists of creeping, fully
or partially locked segments [Wang et al., 2003],
while the segments in the tectonic structure ac-
commodate purely strike-slip or along with dip-slip
components in the Eastern Mediterranean region
[Yava¸so˘glu, 2009, Tiryakio˘glu, 2012]. The kinematic
model in our study is assumed to be locked from
the surface down to a locking depth fixed at 7 km
in the crust. This approach constrains the fault from
creeping since a thicker zone greater than 7 km in
depth is recommended when exploring continental
transform faults [Genrich et al., 2000, Le Béon et al.,
2008, Walters et al., 2011]. C. R. Géoscience — 2020, 352, no 3, 223-232 4. Kinematic modeling Figure 5. Green boxes indicate the strike-slip
rates in which negative values correspond to
left-lateral motion. White boxes indicate dip-
slip rates corresponding to extension, while
yellow boxes denote contraction. Red vectors
demonstrate GPS-derived block velocities and
black rotated arrows denote block rotation
rates with estimated values on the right side
when the Arabian block is assumed as the ref-
erence block. Blue double-headed arrows in-
dicate the estimated internal block strain-rate
axes. Orange boxes denote block names de-
fined in the model. of 1.122 and an nrms of 1.943. On the other hand,
the Amanous block (amns) was introduced addition-
ally into the model in the last stage. Eventually, the
GPS observations fit well with the kinematic model
formed with five blocks and five fault segments (Fig-
ure 3). The significant nonsystematic residuals are
noted with 1.35 mm overall data rms between the
model and observations (Figure 4). Depending on
the block and fault geometries introduced, the re-
duced chi-square parameter of the model converges
to 1.814. In light of the findings from this research, a left-
lateral strike slip at a rate of 7.5 mm/yr on the
EAF allows significant stress transfer to the KF at
4.4–5.4 mm/yr sinistral strike-slip rates (Figure 5). Moreover, it contributes to a sinistral strike slip at
a rate of 3.4 mm/yr and a reverse slip at a rate of
3.1 mm/yr along the KOF. Reverse faulting on the
KOF indicates contraction in agreement with the re-
sult by Aktu˘g et al. [2016]. Normal faulting on the
KF has a slip at a rate approximately 3 mm/yr indi-
cating extension, which is consistent with the result
by Reilinger et al. [2006]. The internal block strain-
rate tensors, which are shown by blue double-headed arrows in the same figure, clearly point out a com-
pression of approximately 30 nanostrain/year within
the Amanous Block. However, the greatest strain ac-
cumulation was loaded on the Iskenderun Block at
approximately 40 nanostrain/year, inducing exten-
sion parallel to the main faults and compression per-
pendicular to them. It is also possible to see a simi-
lar pattern on the Sina Block with a strain rate almost
half of the Iskenderun Block. The slowest straining re-
gion seems to be the Anatolian Block with an internal
deformation of approximately 10 nanostrain/year. Furthermore, a locking depth of less than 10 km was
estimated for the EAF. C. R. Géoscience — 2020, 352, no 3, 223-232 4. Kinematic modeling The addition of the
Iskenderun Block (iskd) and the KOF improves the
reduced chi-square parameter to 3.869 with a wrms This study was initiated from a point of view
comprising several blocks and main faults, namely
Arabia (arab-reference block), Anatolia (anat), and
Sinai (sina) blocks and East Anatolian Fault (EAF), KF,
Cyprus Arc (CA), and DSF. The east of the Amanous
(E-AMNS) block boundary equates to the KF, while
the western boundary of the Amanous block was
assumed to be free slipping (nonlocked) without any
elastic strain accumulated. As discussed in Mah-
moud [2012] and Mahmoud et al. [2012], block
and fault geometries in the kinematic model vary
from the simplest to the most complex related to
the tectonic structures in the region. The different Selim Serhan Yıldız et al. 229
Figure 4. Residuals between the model and ob-
served velocities with 95% confidence ellipses. of 1.122 and an nrms of 1.943. On the other hand,
the Amanous block (amns) was introduced addition-
ally into the model in the last stage. Eventually, the
GPS observations fit well with the kinematic model
formed with five blocks and five fault segments (Fig-
ure 3). The significant nonsystematic residuals are
noted with 1.35 mm overall data rms between the
model and observations (Figure 4). Depending on
the block and fault geometries introduced, the re-
duced chi-square parameter of the model converges
to 1.814. In light of the findings from this research, a left-
lateral strike slip at a rate of 7.5 mm/yr on the
Figure 5. Green boxes indicate the strike-slip
rates in which negative values correspond to
left-lateral motion. White boxes indicate dip-
slip rates corresponding to extension, while
yellow boxes denote contraction. Red vectors
demonstrate GPS-derived block velocities and
black rotated arrows denote block rotation
rates with estimated values on the right side
when the Arabian block is assumed as the ref-
erence block. Blue double-headed arrows in-
dicate the estimated internal block strain-rate
axes. Orange boxes denote block names de-
fined in the model. arrows in the same figure, clearly point out a com-
pression of approximately 30 nanostrain/year within Selim Serhan Yıldız et al. 229
Figure 4. Residuals between the model and ob-
served velocities with 95% confidence ellipses. Figure 5. Green boxes indicate the strike-slip
rates in which negative values correspond to
left lateral motion White boxes indicate dip Selim Serhan Yıldız et al. 4. Kinematic modeling Selim Serhan Yıldız et al. 229 Figure 4. Residuals between the model and ob-
served velocities with 95% confidence ellipses. of 1.122 and an nrms of 1.943. On the other hand,
the Amanous block (amns) was introduced addition-
ally into the model in the last stage. Eventually, the
GPS observations fit well with the kinematic model
formed with five blocks and five fault segments (Fig-
ure 3). The significant nonsystematic residuals are
noted with 1.35 mm overall data rms between the
model and observations (Figure 4). Depending on
the block and fault geometries introduced, the re-
duced chi-square parameter of the model converges
to 1.814. In light of the findings from this research, a left-
lateral strike slip at a rate of 7.5 mm/yr on the
EAF allows significant stress transfer to the KF at
4.4–5.4 mm/yr sinistral strike-slip rates (Figure 5). Moreover, it contributes to a sinistral strike slip at
a rate of 3.4 mm/yr and a reverse slip at a rate of
3.1 mm/yr along the KOF. Reverse faulting on the
KOF indicates contraction in agreement with the re-
sult by Aktu˘g et al. [2016]. Normal faulting on the
KF has a slip at a rate approximately 3 mm/yr indi-
cating extension, which is consistent with the result
Figure 5. Green boxes indicate the strike-slip
rates in which negative values correspond to
left-lateral motion. White boxes indicate dip-
slip rates corresponding to extension, while
yellow boxes denote contraction. Red vectors
demonstrate GPS-derived block velocities and
black rotated arrows denote block rotation
rates with estimated values on the right side
when the Arabian block is assumed as the ref-
erence block. Blue double-headed arrows in-
dicate the estimated internal block strain-rate
axes. Orange boxes denote block names de-
fined in the model. arrows in the same figure, clearly point out a com-
pression of approximately 30 nanostrain/year within
the Amanous Block. However, the greatest strain ac-
cumulation was loaded on the Iskenderun Block at
approximately 40 nanostrain/year, inducing exten-
sion parallel to the main faults and compression per-
pendicular to them. It is also possible to see a simi-
lar pattern on the Sina Block with a strain rate almost
half of the Iskenderun Block. The slowest straining re-
gion seems to be the Anatolian Block with an internal
deformation of approximately 10 nanostrain/year. Figure 4. Residuals between the model and ob-
served velocities with 95% confidence ellipses. 4. Kinematic modeling However, it is partially locked 230 Selim Serhan Yıldız et al. Figure 6. Distribution of fault locking depths. Phi represents fully locked fault zones in red,
while purple stands for free slipping. Color
tones between red and purple demonstrate
partially locked zones. have great importance for understanding the present
kinematics of the Eastern Mediterranean region. The
campaign GPS observations in 2019 helped improve
the data quality in estimating velocity uncertainties. In addition to the new campaign data set acquired in
2019, permanent GPS stations from local networks
operated by municipalities were merged into our re-
gional network to densify observation sites spatially
and temporally. Both the new campaign data set and
permanent GPS stations, regarded as the true value
of this study, constitute unique data sets that were
not available in the past. Accordingly, slip rates for the faults mentioned
above were published in several studies recently. Earlier, Aktu˘g et al. [2016] and Mahmoud et al. [2012] estimated 10.5 mm/yr and 9.0 mm/yr strike-
slip rates, respectively, while Reilinger et al. [2006]
reported 10.0 mm/yr slip rates for the left-lateral
EAFZ. However, there seems to be no consensus on
normal/reverse fault components of the EAF since
Reilinger et al. [2006] argued for extension at a
5.1 mm/yr dip-slip rate, while Aktu˘g et al. [2016]
proposed contraction at 2.4–6.3 mm/yr reverse-slip
rates. On the other hand, Mahmoud et al. [2012]
and our study assert no significant normal/reverse
fault components for the EAF. Similarly, the KF was
also studied many times recently so that the esti-
mations for strike-slip rates are 4.5 mm/yr by Ak-
tu˘g et al. [2016], 4.0 mm/yr by Mahmoud et al. [2012], 6.8 mm/yr by Reilinger et al. [2006], and 4.4–
5.4 mm/yr from this study. There are also some de-
bates about normal/reverse faulting on the KF. In
contrast, the KOF needs to be studied further in
detail. Figure 6. Distribution of fault locking depths. Phi represents fully locked fault zones in red,
while purple stands for free slipping. Color
tones between red and purple demonstrate
partially locked zones. down to a 25 km depth. In contrast, just like the KOF,
the KF has a locking depth of 20 km (Figure 6). C. R. Géoscience — 2020, 352, no 3, 223-232 5. Discussion and conclusion In this study, we explored the strain accumula-
tions on major tectonic features around the Eastern
Mediterranean region by the inversion of GPS veloc-
ity estimations to obtain block rotation rates, fault
locking parameters, and internal strain rates in a 3D
elastic dislocation model. For this purpose, a region-
specific GPS network, composed of campaign obser-
vation sites and permanent stations, was formed to
clarify kinematic characteristics of the tectonically
active zones around the Hatay Triple Junction in
the region. Initially, former campaign data sets from
2009, 2010, and 2011 acquired in joint campaign ob-
servation sites from previous studies were collected. The campaign data sets from previous years provide
a historical kinematic background for the region. Moreover, it was required to determine recent tec-
tonic deformations. Ongoing strain accumulations
lead this research to review the estimations for slip
distributions and locking depths using new GPS ob-
servations across major faults. Therefore, GPS ob-
servations performed in 2019 on the same network,
which was the fourth time after 2009, 2010, and 2011, In conclusion, a sinistral slip rate of 7.5 mm/yr
estimated from our model was initialized to accu-
mulate a strain of approximately 6.8 m in total af-
ter the 1114 AD earthquake (Ms > 7.8) between the
Turkoglu–Golbasi segment of the EAFZ and the KOF. The total strain accumulation within the Golbasi–
Turkoglu segment supports the prediction of the next
probable major earthquake with a magnitude of 7.2–
7.6 if it ruptures entirely over its full 90 km length. Similarly, a 3.4 mm/yr dextral slip rate for the KOF
corresponds to a 1.7 m total strain since the 1513 AD
earthquake (M ∼7.4) and makes M 7.1 possible for
the next probable major earthquake. However, Mw
6.8–7.0 is predicted in the case of two segments with a
rupture length of approximately 40 km in each. Con- C. R. Géoscience — 2020, 352, no 3, 223-232 Selim Serhan Yıldız et al. 231 GPS measurements in Syria: slow slip rate along the
northern Dead Sea Fault. Geophys. J. Intl, 180:125–
135. sidering the 1822 AD (M 7.0) and 1872 AD (M 7.2)
earthquakes on the KF, a total strain accumulation
of 0.65–0.80 m at a 4.4–5.4 mm/yr sinistral slip rate
confirms the prediction of a possible M 6.8–6.9 earth-
quake along 150 km based on Wells and Coppersmith
[1994]. As per the approach by Anderson et al. Acknowledgments Anderson, J. G., Wesnousky, S. G., and Stirling, M. W. (1996). Earthquake size as a function of fault slip
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this project who helped during the field work. This
article was supported financially by the Coordinator
of Scientific Research Projects (BAP) of Hitit Univer-
sity (project no. ODMYO19001.19.002) and Istanbul
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ware developed by Massachusetts Institute of Tech-
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Reproducibility of CT radiomic features in lung neuroendocrine tumours (NETs) patients: analysis in a heterogeneous population
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Abstract Background The aim is to find a correlation between texture features extracted from neuroendocrine (NET) lung cancer
subtypes, both Ki-67 index and the presence of lymph-nodal mediastinal metastases detected while using different computer
tomography (CT) scanners. Methods Sixty patients with a confirmed pulmonary NET histological diagnosis, a known Ki-67 status and metastases, were
included. After subdivision of primary lesions in baseline acquisition and venous phase, 107 radiomic features of first and
higher orders were extracted. Spearman’s correlation matrix with Ward’s hierarchical clustering was applied to confirm the
absence of bias due to the database heterogeneity. Nonparametric tests were conducted to identify statistically significant
features in the distinction between patient groups (Ki-67 < 3—Group 1; 3 ≤ Ki-67 ≤ 20—Group 2; and Ki-67 > 20—Group
3, and presence of metastases). Results No bias arising from sample heterogeneity was found. Regarding Ki-67 groups statistical tests, seven statistically
significant features (p value < 0.05) were found in post-contrast enhanced CT; three in baseline acquisitions. In metastasis
classes distinction, three features (first-order class) were statistically significant in post-contrast acquisitions and 15 features
(second-order class) in baseline acquisitions, including the three features distinguishing between Ki-67 groups in baseline
images (MCC, ClusterProminence and Strength). g
g
Conclusions Some radiomic features can be used as a valid and reproducible tool for predicting Ki-67 class and hence the
subtype of lung NET in baseline and post-contrast enhanced CT images. In particular, in baseline examination three features
can establish both tumour class and aggressiveness. La radiologia medica (2023) 128:203–211
https://doi.org/10.1007/s11547-023-01592-y La radiologia medica (2023) 128:203–211
https://doi.org/10.1007/s11547-023-01592-y CHEST RADIOLOGY Reproducibility of CT radiomic features in lung neuroendocrine
tumours (NETs) patients: analysis in a heterogeneous population Eleonora Bicci1 · Diletta Cozzi1,2 · Edoardo Cavigli1 · Ron Ruzga1 · Elena Bertelli1 · Ginevra Danti1 · Silvia Bettarini3 ·
Paolo Tortoli3 · Lorenzo Nicola Mazzoni4 · Simone Busoni3 · Vittorio Miele1 Eleonora Bicci1 · Diletta Cozzi1,2 · Edoardo Cavigli1 · Ron Ruzga1 · Elena Bertelli1 · Ginevra Danti1 · Silvia Bettarini3 ·
Paolo Tortoli3 · Lorenzo Nicola Mazzoni4 · Simone Busoni3 · Vittorio Miele1 Received: 30 September 2022 / Accepted: 4 January 2023 / Published online: 13 January 2023
© The Author(s) 2023 * Diletta Cozzi
dilettacozzi@gmail.com
Eleonora Bicci
eleonora.bicci92@gmail.com
Edoardo Cavigli
edoardocavigli@yahoo.it
Ginevra Danti
ginevra.danti@gmail.com
Silvia Bettarini
silvia.bettarini@unifi.it
Paolo Tortoli
paolo.tortoli@unifi.it
Lorenzo Nicola Mazzoni
mazzoniln@gmail.com Abstract Keywords Lung carcinoids · Lung cancer · Computed tomography · Radiomics · Ki-67 Vol.:(0123456789)
1 3
* Diletta Cozzi
dilettacozzi@gmail.com
Eleonora Bicci
eleonora.bicci92@gmail.com
Edoardo Cavigli
edoardocavigli@yahoo.it
Ginevra Danti
ginevra.danti@gmail.com
Silvia Bettarini
silvia.bettarini@unifi.it
Paolo Tortoli
paolo.tortoli@unifi.it
Lorenzo Nicola Mazzoni
mazzoniln@gmail.com
Simone Busoni
busonis@aou-careggi.toscana.it
Vittorio Miele
vmiele@sirm.org
1
Department of Emergency Radiology, Careggi University
Hospital, Largo Brambilla 3, 50134 Florence, Italy
2
Italian Society of Medical and Interventional Radiology
(SIRM), SIRM Foundation, 20122 Milan, Italy
3
Department of Health Physics, L.Go Brambilla, Careggi
University Hospital, 50134 Florence, Italy
4
Department of Health Physics, AUSL Toscana Centro, Via
Ciliegiole 97, 51100 Pistoia, Italy Simone Busoni
busonis@aou-careggi.toscana.it Vittorio Miele
vmiele@sirm.org
1
Department of Emergency Radiology, Careggi University
Hospital, Largo Brambilla 3, 50134 Florence, Italy
2
Italian Society of Medical and Interventional Radiology
(SIRM), SIRM Foundation, 20122 Milan, Italy
3
Department of Health Physics, L.Go Brambilla, Careggi
University Hospital, 50134 Florence, Italy
4
Department of Health Physics, AUSL Toscana Centro, Via
Ciliegiole 97, 51100 Pistoia, Italy (0121 3456789)
3 La radiologia medica (2023) 128:203–211 204 Abbreviations
NET
Neuroendocrine tumour
CT
Computed tomography
TC
Typical carcinoid
AC
Atypical carcinoid
LCNEC
Large-cell neuroendocrine carcinoma
SCLS
Small-cell lung carcinoma
GEP-NEN
Gastrointestinal neuroendocrine tumour
ROI
Regions of interest
MRI
Magnetic resonance imaging
PET-CT
Positron emission tomography computed
tomography
PACS
Picture archiving and communication system
GLCM
Grey level co-occurrence matrix
GLSZM
Grey level size zone matrix
GLRLM
Grey level run length matrix
NGTDM
Neighbouring grey tone difference matrix
GLDM
Grey level dependence matrix
MCC
Maximal correlation coefficient of the grading system of lung NETs, it is currently used
in gastrointestinal neuroendocrine tumours (GEP-NENs)
according to the 2019 WHO classification. Well-differen-
tiated NENs are further divided into grades based solely on
Ki-67 proliferation index and mitotic index: into grade 1
(G1, mitotic rate < 2, Ki-67 index < 3), grade 2 (G2, mitotic
rate 2–20, Ki-67 index between 3 and 20) and grade 3 (G3,
mitotic rate > 20, Ki-67 index > 20) [12, 13]. It has been
studied how these grading values can play a fundamental
role in prognostic evaluation and differentiation between
various tumour histotypes and in particular in discriminat-
ing between TCs and ACs tumours or high-grade SCLS and
LCNEC from carcinoid tumours [12, 13]. Background Pulmonary neuroendocrine tumours (NETs) are a group
of neoplasms that account for about 25% of all NETs and
2% of lung cancers, they are divided into different groups
according to their aggressiveness [1, 2]. In particular, NETs
are classified as low-grade or typical carcinoids (TCs),
intermediate grade or atypical carcinoids (ACs) and high
grade, divided into large-cell neuroendocrine carcinomas
(LCNECs) and small-cell lung carcinomas (SCLC) [3]. This
division into successively more aggressive forms is based on
the progressive increase in the number of mitoses and the
presence of necrosis at the histological evaluation: usually
typical carcinoid does not show necrosis unlike the atypical
ones, whereas the highest percentage of mitoses are found in
SCLC, the most aggressive form of NETs [4–7]. Abstract In recent years, radiomics, with the use of texture analy-
sis, is becoming an increasingly used tool, capable of giving
more precise structural information, not always visible by
the human eye and not subject to interindividual variabil-
ity [14–21]. Radiomics is therefore an innovative technique
used to characterize the inhomogeneity of a given tissue,
and more specifically, as in our case, the lung NET lesions,
through the extraction and analysis of features obtained
by investigating regions of interest (ROI) from different
imaging modalities such as CT, magnetic resonance imag-
ing (MRI) or positron emission tomography CT (PET-CT)
[22–26]. The application of texture analysis in NET of the
lung could therefore be useful in both the diagnosis and
early differentiation of distinctive NET tumour histotypes
[27, 28]. In this paper, we faced the reproducibility of CT
radiomics features in lung NETs using different CT scanners,
trying to integrate different features or to find new metrics
in assessing tumour aggressiveness and histotype, that could
be reproducible in daily practice. Patients and ethics issues Textural features extraction was car-
ried out by means of SlicerRadiomics tool. A total of 107
features of the PyRadiomics lists were selected, belonging to
first-order, 3D shape-based, grey level co-occurrence matrix
(GLCM), grey level size zone matrix (GLSZM), grey level
run length matrix (GLRLM), neighbouring grey tone differ-
ence matrix (NGTDM) and grey level dependence matrix
(GLDM) classes. 237–1226 mGy*cm; for contrast enhanced scans acquisi-
tion parameters were: matrix size 512 × 512 pixels with
slice thickness between 1 and 5 mm, 120 kVp, 156 ± 101
mAs, CTDIvol of 11.4 ± 6.6 mGy and DLP in the range
253.5–1228 mGy*cm. All studies were reviewed by two
radiologists, with 5 and 15 years’ experience in thoracic
imaging. The entire volume of the primary tumour was visu-
ally segmented in both unenhanced and enhanced acquisi-
tions employing a volumetric ROI (region of interest) using
3DSlicer software version 4.10.2 (open source software;
https://www.slicer.org/). The ROI was delineated slice by
slice for each patient. Textural features extraction was car-
ried out by means of SlicerRadiomics tool. A total of 107
features of the PyRadiomics lists were selected, belonging to
first-order, 3D shape-based, grey level co-occurrence matrix
(GLCM), grey level size zone matrix (GLSZM), grey level
run length matrix (GLRLM), neighbouring grey tone differ-
ence matrix (NGTDM) and grey level dependence matrix
(GLDM) classes. Fig. 1 Workflow of patients’ selection Fig. 1 Workflow of patients’ selection patients). Our study population resulted in 60 patients, 29
males and 31 females. As there is still no standardization
in the use and class division of lung NETs using the Ki-67
index, we relied in our study on the most recent WHO 2019
GEP-NENs grading. Selected patients were then divided in
three groups: Ki-67 < 3 (Group 1), 3 ≤ Ki-67 ≤ 20 (Group 2)
and Ki-67 > 20 (Group 3). This retrospective observational
study was approved by the Ethics Committee of our Institu-
tion (study protocol n:14776_oss). patients). Our study population resulted in 60 patients, 29
males and 31 females. As there is still no standardization
in the use and class division of lung NETs using the Ki-67
index, we relied in our study on the most recent WHO 2019
GEP-NENs grading. Selected patients were then divided in
three groups: Ki-67 < 3 (Group 1), 3 ≤ Ki-67 ≤ 20 (Group 2)
and Ki-67 > 20 (Group 3). Patients and ethics issues This retrospective observational
study was approved by the Ethics Committee of our Institu-
tion (study protocol n:14776_oss). Statistical analysis The intrinsic heterogeneity of the database due to the use of
different contrast agents, scanners and reconstruction kernels
could lead to the presence of bias in the statistical differ-
entiation of the patient classification groups; to verify the
absence of this bias a Spearman’s correlation matrix with
Ward’s hierarchical clustering was created using R software
(https://www.R-project.org/). Nonparametric tests were per-
formed to identify features that showed significant differ-
ences between the three classes of Ki67 (Ki-67 < 3 (Group
1), 3 ≤ Ki-67 ≤ 20 (Group 2) and Ki-67 > 20 (Group 3)) or
between the presence or absence of mediastinal lymph-node
metastases. This statistical analysis was performed sepa-
rately on unenhanced and on contrast enhanced CT scans
databases using SPSS (IBM SPSS Statistics for Windows,
version 27.0. Armonk, NY: IBM Corp). For the Ki-67 class
distinction, the Kruskal–Wallis test was used and the post
hoc analysis was performed with the Dunn’s test, consider-
ing the Bonferroni correction. For the metastases grouping
distinction, the Mann–Whitney test was employed. Signifi-
cance threshold was set at p = 0.05. CT scanners
Bright Speed, Optima CT 660, LightSpeed VCT, Revolution HD—General Electric Healthcare
SOMATOM Emotion 16, Definition Flash, Definition AS + , Sensation 16, Sensation 64, Sensation
Open—Siemens Healthineers
iCT SP—Philips Healthcare Patients and ethics issues This is a single-centre, observational, retrospective study. Between September 2008 and October 2021, all patients
with a histological diagnosis of pulmonary NET who
underwent pre-treatment CT exam were selected by search-
ing our Picture Archiving and Communication System
(PACS). Inclusion criteria were: patients aged between 18
and 99 years; histological diagnosis of pulmonary NET
confirmed by biopsy or by surgical specimen; Ki-67 value;
CT examination performed in our department with non-
enhanced acquisition and venous phase; at least one CT or
PET-TC in the follow-up. The workflow of patient’s selec-
tion is shown in Fig. 1. The initial population included 91
patients; of these 15 had no pre-treatment CT performed in
our hospital. To make the sample more homogeneous, we
excluded those who did not have a baseline-CT (six patients)
and those who did not have venous phase acquisition (10 Computed tomography (CT) is the imaging of choice
in diagnosing this pathology, being able not only to detect
changes related to the presence lesion, but also necessary
for loco-regional staging of disease [8, 9]. The most fre-
quent findings on CT are the presence of a solid consolida-
tive lesion within the lung parenchyma, frequently polylo-
bate, especially in the case of low-grade forms, with vivid
enhancement after administration of contrast medium. The
lesion may also present as endo-bronchial or with a mixed
parenchymal and bronchial component. Parenchymal atelec-
tasis may also be present in the case of bronchial obstruction
[10, 11]. Increasingly important in assessing tumour aggressive-
ness and thus meaning in the prognosis of these patients by
the correlation of nuclear antigen expressed by proliferat-
ing cells (Ki-67). Although this classification is not yet part 1 3 1 3 La radiologia medica (2023) 128:203–211 205 237–1226 mGy*cm; for contrast enhanced scans acquisi-
tion parameters were: matrix size 512 × 512 pixels with
slice thickness between 1 and 5 mm, 120 kVp, 156 ± 101
mAs, CTDIvol of 11.4 ± 6.6 mGy and DLP in the range
253.5–1228 mGy*cm. All studies were reviewed by two
radiologists, with 5 and 15 years’ experience in thoracic
imaging. The entire volume of the primary tumour was visu-
ally segmented in both unenhanced and enhanced acquisi-
tions employing a volumetric ROI (region of interest) using
3DSlicer software version 4.10.2 (open source software;
https://www.slicer.org/). The ROI was delineated slice by
slice for each patient. Bright Speed, Optima CT 660, LightSpeed VCT, Revolution HD—General Electric Healthcare
SOMATOM Emotion 16, Definition Flash, Definition AS + , Sensation 16, Sensation 64, Sensation
Open—Siemens Healthineers
iCT SP—Philips Healthcare Images acquisitions and analysis CT images were acquired using different CT scanners, as
shown in Table 1. The study protocol consisted of a baseline
acquisition followed by a venous phase acquisition with a
70s’ delay after administration of an intravenous contrast
medium (flow 3 mL/s, followed by bolus of saline with a
dose of 40 ml). Two types of contrast medium were injected,
Ultravist®370 (Bayer Healthcare) and Iomeron®400
(Bracco Imaging Italia). The acquisition parameters for
the basal scan were: matrix size 512 × 512 pixels with
slice thickness between 1 and 5 mm, 120 kVp, 145 ± 97
mAs, CTDIvol of 10.3 ± 6.7 mGy and DLP in the range Table 1 List of computed
tomography (CT) scanners used
in our study 1 3 La radiologia medica (2023) 128:203–211 206 from the Mann–Whitney test, in the distinction between
the absence (0) and presence (1) of metastases, is also
reported. Results This retrospective observational study was approved by
the Ethics Committee of our Institution (study protocol
n:14776_oss). The 60 patients selected were aged between
35 and 90 years old (mean age 71 years; 35 women, 25
men). Histopathological analysis revealed 45 cases of
TCs (75.0%), 12 SCLCs (20.0%) and 3 LCNECs (5.0%). According to Ki-67 expression, patients in Group 1 were
18/60 (30.0%), Group 2 24/60 (40.0%) and Group 3 18/60
(30.0%). Typical carcinoids showed mean Ki-67 values of
7.02, while SCLCs of 74.58 and LCNECs of 80,0. Medi-
astinal lymph-nodal metastases were present in 30/60
(50.0%) at the first follow-up made by CT or PET-CT. The
absence of bias in the database due to possible confound-
ing factors was verified through Spearman’s correlation
matrix with Ward’s hierarchical clustering, as shown in
Fig. 2. No evidence of a bias associated to these factors
was found. In Tables 2 and 3, the features that showed
significant differences among the Ki-67 classes, after
Kruskal–Wallis with Dunn’s post hoc test, are listed for
unenhanced and contrast enhanced CT scans, respectively
(Tables 2, 3). The list of significant features, resulting In the correlation analysis between the different types
of tumour and Ki-67 classes, three features were statis-
tically significant in non-contrast enhanced scans: MCC
(p = 0.048), ClusterProminence (p = 0.046) and Strength
(p = 0.040). While in the contrast enhanced scans, seven
features resulted statistically significant: Correlation
(p = 0.046), Median (p = 0.004), Maximum (p = 0.005),
RootMeanSquared (p = 0.002), 90th Percentile (p = 0.001),
10th Percentile (p = 0.020), Mean (p = 0.006). All of them,
except for Correlation, are first-order class features. i
When assessing the correlation between tumour his-
totype and the presence of metastases, three characters
(Median (p = 0.049), RootMeanSquared (p = 0.045) and
90th Percentile (p = 0.030) were found in the feature’s
statistical analysis of the venous phase contrast medium
images. In the analysis performed on the unenhanced
images, 15 features were statistically significant, includ-
ing those that distinguished the presence or absence of
metastases also in post-contrast medium acquisitions,
namely MCC (p = 0.016), ClusterProminence (p = 0.033)
and Strength (p = 0.003). Fig. 2 Heatmaps. Heatmap representing Spearman’s correlation
matrix with Ward’s hierarchical clustering for unenhanced (a) and
contrast enhanced (b) CT acquisitions. The colour of the heatmap
goes from red (high correlation) to blue (low correlation). Results The top of
the matrix shows the dendrogram of the clustering and three coloured
bars which represent exam characteristics that could lead to a bias
in the statistical correlation between groups (CT model, convolution
kernel and contrast medium). It can be seen that all of those charac-
teristics are randomly distributed over the database and none of them
can be associated to a specific cluster Fig. 2 Heatmaps. Heatmap representing Spearman’s correlation
matrix with Ward’s hierarchical clustering for unenhanced (a) and
contrast enhanced (b) CT acquisitions. The colour of the heatmap
goes from red (high correlation) to blue (low correlation). The top of
the matrix shows the dendrogram of the clustering and three coloured bars which represent exam characteristics that could lead to a bias
in the statistical correlation between groups (CT model, convolution
kernel and contrast medium). It can be seen that all of those charac-
teristics are randomly distributed over the database and none of them
can be associated to a specific cluster Fig. 2 Heatmaps. Heatmap representing Spearman’s correlation
matrix with Ward’s hierarchical clustering for unenhanced (a) and
contrast enhanced (b) CT acquisitions. The colour of the heatmap
goes from red (high correlation) to blue (low correlation). The top of
the matrix shows the dendrogram of the clustering and three coloured bars which represent exam characteristics that could lead to a bias
in the statistical correlation between groups (CT model, convolution
kernel and contrast medium). It can be seen that all of those charac-
teristics are randomly distributed over the database and none of them
can be associated to a specific cluster 1 3 3 La radiologia medica (2023) 128:203–211 207 Discussion
To the best of our knowledge this is the first study evaluating
and tumour histotype of pulmonary NETs by analysing the
possibility of results’ standardization and reproducibility on
different CT scanners, therefore avoiding an important bias
Table 2 List of features that showed significant differences among the Ki-67 classes in Kruskal–Wallis test for unenhanced CT scans
For each feature are reported median and Tukey’s 1° and 3° quartile for each class, p value and classes pairs that passed Dunn’s test, if present. List of significant features, resulting from Mann–Whitney test, in the distinction between the absence (0) and presence (1) of metastases. Results For
each feature are reported median and Tukey’s 1° and 3° quartile, separately for the two groups, and the p value
Feature
Ki67 class
Median
1° quartile
3° Quartile
p value
Dunn’s test
Baseline-CT
1
0.525
0.33
0.6225
MCC
2
0.44
0.34
0.5825
0.048
/
3
0.34
0.265
0.43
1
182.46
9.1325
1968.1775
Cluster prominence
2
17.01
3.4025
194.9325
0.046
1–3
3
3.66
9.01
18.8
1
0.67
0.2075
2.24
Strength
2
0.385
0.1275
0.9875
0.040
1–3
3
0.16
0.045
0.215
Feature
Metastasis
Median
1° Quartile
3° Quartile
p value
Grey level variance (gldm)
0
0.83
0.5725
2.04
0.041
1
0.58
0.415
0.885
Small dependence emphasis
0
0.05
0.04
0.0675
0.025
1
0.04
0.03
0.05
MCC
0
0.505
0.345
0.6525
0.016
1
0.36
0.285
0.47
Sum squares
0
0.715
0.5
2.1
0.048
1
0.53
0.395
0.845
Cluster prominence
0
39.575
5.3075
1152.3575
0.033
1
9.01
3.69
21.935
Imc1
0
-0.1
-0.16
-0.07
0.048
1
-0.07
-0.12
-0.045
Cluster tendency
0
1.895
1.1625
6.6975
0.041
1
1.16
0.905
2.225
Variance
0
468.395
310.345
1196.237
0.039
1
306.71
195.815
500.74
Grey level variance (glrlm)
0
0.995
0.7175
3.0175
0.045
1
0.71
0.525
1.005
Grey level non uniformity normalized (glszm)
0
0.205
0.1325
0.27
0.017
1
0.26
0.215
0.32
Grey level non uniformity (glszm)
0
7.8
4.64
21.1475
0.017
1
23.82
8.765
40.975
Large area emphasis
0
7606.69
1944.933
34,346.72
0.047
1
40,481.62
5396.74
115,192.56
Zone percentage
0
0.05
0.03
0.07
0.035
1
0.03
0.02
0.05
Large area low grey level emphasis
0
189.485
92.2725
1300.74
0.035
1
1181.95
244.225
2984.82
Strength
0
0.67
0.225
2.12
0.003
1
0.15
0.05
0.415 For each feature are reported median and Tukey’s 1° and 3° quartile for each class, p value and classes pairs that passed Dunn’s test, if present. List of significant features, resulting from Mann–Whitney test, in the distinction between the absence (0) and presence (1) of metastases. For
each feature are reported median and Tukey’s 1° and 3° quartile, separately for the two groups, and the p value and tumour histotype of pulmonary NETs by analysing the
possibility of results’ standardization and reproducibility on
different CT scanners, therefore avoiding an important bias
due to the heterogeneity of the machine used for imaging. Discussion For each feature are reported median and Tukey’s 1° and 3° quartile separately for the two groups and the p value are not affected by variations secondary to different techni-
cal acquisition parameters is therefore a fundamental factor
[30]. One of the main results of our study is that none of the
features extracted and selected is affected by bias arising
from sample heterogeneity. In our previous study, we have selected significant features
in Ki-67 classes and aggressiveness stratification, using a
small number of patients trying to have the most homogene-
ous group of subjects, using only one CT scanner [27]. We
therefore attempted with this additional study to evaluate a
larger number of patients, assessing whether with hierarchi-
cal clustering analysis there was any bias resulting from the
use of different CTs. Different studies have been conducted
to assess how variation in the acquisition parameters and
reconstruction techniques of different CTs could affect radi-
omics features [29]. The study by Meyer et al. assessed how
most features can be altered secondary to variations in acqui-
sition parameters: in particular slice thickness showed the
greatest impact on the reproducibility of these features. This
demonstrates how the selection of reproducible features that In our previous study, we have selected significant features
in Ki-67 classes and aggressiveness stratification, using a
small number of patients trying to have the most homogene-
ous group of subjects, using only one CT scanner [27]. We
therefore attempted with this additional study to evaluate a
larger number of patients, assessing whether with hierarchi-
cal clustering analysis there was any bias resulting from the
use of different CTs. Different studies have been conducted
to assess how variation in the acquisition parameters and
reconstruction techniques of different CTs could affect radi-
omics features [29]. The study by Meyer et al. assessed how
most features can be altered secondary to variations in acqui-
sition parameters: in particular slice thickness showed the
greatest impact on the reproducibility of these features. This
demonstrates how the selection of reproducible features that An important goal of our study was the identification of
features capable of differentiating tumour histotypes accord-
ing to the Ki-67 value. In particular, three features (maxi-
mal correlation coefficient—MCC, ClusterProminence and
Strength) were significant in non-contrast acquisitions, as
explained in the Results section. Cluster Prominence and
MCC are second-order features belonging to the class of
the grey level co-occurrence matrix (GLCM). Discussion To the best of our knowledge, this is the first study evaluating
the efficacy of radiomics in considering the aggressiveness 1 3 La radiologia medica (2023) 128:203–211 208 Table 3 List of features that showed significant differences among the Ki-67 classes in the Kruskal–Wallis test between the Ki-67 classes for
contrast enhanced CT scans. For each feature are reported median and Tukey’s 1° and 3° quartile for each class, p value and classes pairs that had passed Dunn’s test, if pre-
sent. List of significant features, resulting from the Mann–Whitney test, in the distinction between the absence (0) and presence (1) of metasta-
ses. For each feature are reported median and Tukey’s 1° and 3° quartile separately for the two groups and the p value
Feature
Ki67 class
Median
1° Quartile
3° Quartile
p value
Dunn’s test
BASELINE-CT
1
0.4115
0.3325
0.58875
Correlation
2
0.4015
0.294
0.52
0.046
1–3
3
0.293
0.229
0.411
1
93
79
116.5
Median
2
78
59.75
95
0.004
1–3
3
60
52.5
70
1
233
147.5
378.5
Maximum
2
196
139
267.25
0.005
1–3
3
134
122
153
1
98.965
83.5675
128.56
Root mean squared
2
81.3765
67.108
102.3775
0.002
1–3
3
64.473
58.3855
72.0255
1
120.5
102
158.75
90th Percentile
2
104
84.75
124.2
0.001
1–3
3
86
79.5
93.7
1
63.9
47.5
77.75
10th Percentile
2
53
35
58.35
0.020
1–3
3
32
29.5
46
1
91.549
74.37
116.8468
Mean
2
76.712
59.16125
89.25025
0.006
1–3
3
58.515
52.965
69.818
Feature
Metastasis
Median
1° Quartile
3° Quartile
p value
Median
0
86.5
66.25
104.75
0.049
1
68
55
84.5
Root mean squared
0
95.6215
72.4565
118.7953
0.045
1
71.166
61.829
91.037
90th Percentile
0
115
94.5
137.4
0.030
1
93
83.2
111.5 Table 3 List of features that showed significant differences among the Ki-67 classes in the Kruskal–Wallis test between the Ki-67 classes for
contrast enhanced CT scans. Feature
Ki67 class
Median
1° Quartile
3° Quartile
p value
Dunn’s test For each feature are reported median and Tukey’s 1° and 3° quartile for each class, p value and classes pairs that had passed Dunn’s test, if pre-
sent. List of significant features, resulting from the Mann–Whitney test, in the distinction between the absence (0) and presence (1) of metasta-
ses. Discussion The former
is a measure of the skewness and asymmetry of the GLCM. The latter represents a quantification of the complexity of 1 3 3 La radiologia medica (2023) 128:203–211 209 the texture. Strength belongs to neighbouring grey tone dif-
ference matrix (NGTDM), representing a measure of the
primitives in an image. Equally, these features were also
significant in acquisitions without contrast medium in cor-
relating with the presence of metastases. features showed higher values in tumours with metastasis
in relation to the tissue heterogeneity of poorly differenti-
ated NETs. As evidence of this, Grey Level non-uniformity
(GLSMZ) also showed higher values in tumours that had
metastases and were therefore more aggressive. This study still has some limits: first of all, we acknowl-
edge the relatively small number of patients and we are
collecting further exams for future radiomics applications. Moreover, we lack validation of our results with a control
group. Another limit is the segmentation performed manu-
ally by one radiologist: even if an expert one, it could be
interesting to repeat in the next future these evaluations
applying an automatic approach based on deep learning. Finally, enhanced exams were performed using two differ-
ent types of contrast media that may have affected the CT
texture of NETs imaging. However, a previous study by
Botta et al. demonstrates that radiomic features were not
influenced by different contrast media, in fact this was not
investigated in our study [36–38]. Considering that the majority of patients have lesions
with high contrast enhancement, which is a typical char-
acteristic of all neuroendocrine tumours, an evaluation
after the administration of contrast medium is manda-
tory [31–33]. Concerning the correlation between Ki-67
classes and tumour histotypes in enhanced CT images,
six first-order features and one second-order feature were
significant. Median is the median of grey level intensity
within the region of interest and was higher in the class
with lower Ki-67 values in relation to the greater pres-
ence of solid tissue without necrotic or necrotic areas [34]. Maximum represents the highest grey level intensity within
the region of interest and even this was higher in typi-
cal carcinoids due to the absence of necrosis and greater
uptake of contrast medium in a more homogeneous tissue
structure. Discussion 10th and 90th Percentiles are mirror of the 10th
and 90th Percentiles of the grey level intensity within the
region of interest, while Mean represents the average grey
level intensity within the region of interest. As mentioned
above, also these features showed higher values in class
1, therefore representing low-grade tumours. Root mean
squared (RMS) is the square root of the mean of all the
squared intensity values. Median, RMS and 90th Percen-
tile were also significant when analysing the correlation
with the presence or absence of metastases in enhanced
images, showing higher values in tumours with no medi-
astinal lymph-node metastases, underlining their ability to
detect low-grade tumours. This represents another impor-
tant point in our study as it identifies these three features
as highly useful in differentiating tumours with a high
risk of metastasis and high Ki-67 values: these tumours
are therefore more inhomogeneous due to the presence of
necrosis or colliquation from those with a low tendency
to metastasise and with low grading, thus allowing the
early identification of those tumours at high risk [35]. Cor-
relation is a second-order feature representing the linear
dependency of grey level values to their respective voxels
in the grey level co-occurrence matrix (GLCM). Higher
values of this feature were found to be present in Group
1: this result needs further investigation, maybe not be
related to a real tissue inhomogeneity, not common in typi-
cal carcinoids. Other features correlate with the presence
of metastases, including Grey Level Variance belonging
to grey level dependence matrix (GLDM), representing the
variance in grey level in the image and Grey Level Vari-
ance of grey level size zone matrix (GLSZM) that is the
variance in grey level intensities for the zone Both these In conclusion, texture analysis can be a useful tool in
the stratification of lung NET tumour histotypes in corre-
lation with Ki-67 values and the presence of metastases. Limitations resulting from sample inhomogeneities can be
overcome by selecting features unaffected by acquisition
parameters, making the results reproducible and stand-
ardized. Although radiomics is not yet used in clinical
practice, it may become in the future a valuable aid in
the evaluation of both tumour class and aggressiveness of
NETs tumour and, therefore, in decision-making process. Discussion Authors’ contribution DC, EB and EC were involved in conceptualiza-
tion; DC, LNM and SB helped in methodology; LNM, SB, PT and SB
contributed to formal analysis and investigation; EB, LNM, SB,RR,
PT and SB were involved in data curation; DC, EB, LNM, SB, RR, PT
and SB helped in writing—original draft preparation; DC, EC, RR,
LNM and SB helped in writing—review and editing; VM contributed
to visualization; VM helped in supervision and project administra-
tion. All authors have read and agreed to the published version of the
manuscript. Funding Open access funding provided by Università degli Studi di
Firenze within the CRUI-CARE Agreement. This research received
no external funding. Availability of data and materials The datasets used and/or analysed
during the current study are available from the corresponding author
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copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. 16. Palatresi D, Fedeli F, Danti G et al (2022) Correlation of CT
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212(3):497–504 Publisher's Note Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations. 36. Botta F, Raimondi S, Rinaldi L et al (2020) Association of a
CT-based clinical and radiomics score of non-small cell lung
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ing decision support (QDIS™) tool consistency evaluation and 1 3 1 3 3
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Table S4 from Simultaneous Measurement of 92 Serum Protein Biomarkers for the Development of a Multiprotein Classifier for Ovarian Cancer Detection
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Supplemental Table 3. Log2 values for all 92 proteins comparing healthy versus ovarian cancer
serum samples Supplemental Table 3. aMean log2 value with standard deviation in parenthesis. Supplemental Table 3. Log2 values for all 92 proteins comparing healthy versus ovarian cancer
serum samples Log2 values for all 92 proteins comparing healthy versus ovarian cancer
serum samples Healthy Patients
Ovarian Cancer Patients
FGF.BP1
5.79 (0.54)
4.17 (0.44)
< 0.001
MUC.16
2.4 (0.84)
6.48 (1.33)
< 0.001
S100A4
3.58 (0.52)
2.45 (0.6)
< 0.001
WFDC2
6.54 (0.6)
7.82 (0.7)
< 0.001
ITGAV
2.59 (0.41)
1.83 (0.47)
< 0.001
ICOSLG
5.86 (0.79)
4.11 (1.18)
< 0.001
SCF
8.58 (0.86)
7.52 (0.86)
< 0.001
MSLN
3.43 (0.96)
2.16 (1.08)
< 0.001
FADD
1.81 (0.76)
1 (0.64)
< 0.001
EGF
10.02 (1.47)
7.5 (2.41)
< 0.001
CD207
2.19 (0.6)
1.51 (0.59)
< 0.001
MK
5.38 (1.16)
6.67 (1.12)
< 0.001
ITGB5
6.66 (0.63)
5.93 (0.67)
< 0.001
FASLG
8.44 (0.78)
7.55 (0.88)
< 0.001
XPNPEP2
6.29 (0.74)
5.46 (0.88)
< 0.001
KLK13
2.61 (0.66)
3.33 (0.78)
< 0.001
IFN.gamma.R1
3.33 (0.56)
2.79 (0.58)
< 0.001
GPNMB
5.53 (0.74)
4.99 (0.42)
< 0.001
CEACAM5
2.15 (0.85)
1.47 (0.64)
< 0.001
CD48
5.82 (0.67)
5.16 (0.73)
< 0.001
TGFR.2
6.48 (0.71)
5.8 (0.76)
< 0.001
LY9
4.39 (0.66)
3.79 (0.68)
< 0.001
ADAM.TS.15
0.33 (0.59)
1.06 (0.92)
< 0.001
IL6
1.78 (1.01)
3.01 (1.6)
< 0.001
hK11
4.62 (0.65)
5.35 (0.95)
< 0.001
GPC1
3.82 (0.71)
3.21 (0.73)
< 0.001
LYPD3
3.27 (0.7)
2.57 (0.89)
< 0.001
CRNN
4.91 (1.02)
4.11 (0.93)
< 0.001
TLR3
4.75 (0.83)
4.05 (0.87)
< 0.001
WIF.1
4.85 (0.61)
4.28 (0.75)
< 0.001
RSPO3
1.17 (0.71)
1.9 (1.02)
< 0.001
MIA
9.15 (0.65)
8.62 (0.69)
< 0.001
ERBB2
6.28 (0.66)
5.75 (0.71)
< 0.001
FR.alpha
6.44 (0.79)
7.24 (1.16)
< 0.001
CD70
3.89 (0.91)
3.29 (0.7)
< 0.001
GZMB
2.09 (0.87)
1.42 (0.91)
< 0.001
RET
3.7 (0.68)
3.18 (0.77)
< 0.001
TNFRSF19
3.74 (0.91)
3.11 (0.89)
< 0.001
VEGFR.2
6.29 (0.54)
5.91 (0.56)
< 0.001
log2 value on Proseek® Oncology II Platea
Proseek® Protein
Abbreviation
p-values CEACAM1
6.67 (0.47)
6.33 (0.54)
< 0.001
ERBB3
7.12 (0.56)
6.74 (0.62)
< 0.001
TNFRSF6B
3.02 (1.05)
3.73 (1.22)
< 0.001
PPY
3.82 (1.53)
2.96 (1.32)
< 0.001
DLL1
8.65 (0.8)
8.18 (0.78)
< 0.001
CYR61
4.72 (1.09)
4.09 (1.08)
< 0.001
HGF
6.36 (0.82)
5.87 (0.93)
0.001
DKN1A
1.01 (0.78)
0.64 (0.63)
0.001
FCRLB
0.05 (0.52)
0.39 (0.7)
0.002
CD27
7.48 (0.63)
7.18 (0.63)
0.004
SPARC
5.94 (0.23)
5.81 (0.31)
0.005
TGF.alpha
2.41 (0.97)
1.96 (0.93)
0.005
PVRL4
5.09 (1.07)
5.61 (1.12)
0.005
ESM.1
7.62 (0.7)
7.95 (0.75)
0.006
VEGFR.3
5.69 (0.49)
5.43 (0.64)
0.008
EPHA2
1.23 (0.56)
0.98 (0.58)
0.008
AREG
1.92 (0.84)
1.54 (0.95)
0.014
TCL1A
3.95 (1.35)
3.4 (1.34)
0.016
IGF1R
2.9 (0.54)
2.66 (0.66)
0.021
ANXA1
2.31 (0.9)
1.97 (0.96)
0.032
MetAP.2
2.06 (0.9)
2.36 (0.9)
0.048
MAD.homolog.5
1.22 (0.87)
1.48 (0.85)
0.073
ABL1
1.27 (0.74)
1.53 (0.99)
0.087
ADAM.8
3.97 (0.66)
3.79 (0.68)
0.116
TXLNA
1.63 (0.97)
1.53 (0.86)
0.507
VEGFA
9.68 (0.91)
9.69 (1.06)
0.95
CPE
1.83 (0.79)
1.7 (0.89)
0.381
TNFSF13
7.55 (0.63)
7.53 (0.72)
0.89
SYND1
5.7 (0.97)
5.86 (1.17)
0.397
SCAMP3
1.71 (1)
1.68 (1.19)
0.876
TRAIL
6.26 (0.78)
6.39 (0.7)
0.293
TFPI.2
6.76 (0.81)
6.98 (1.04)
0.177
hK8
5.43 (0.69)
5.46 (0.97)
0.798
PODXL
3.34 (0.4)
3.43 (0.61)
0.307
GZMH
2.73 (1.17)
2.58 (1.27)
0.447
FURIN
3.31 (0.63)
3.2 (0.79)
0.351
hK14
4.73 (0.95)
4.8 (0.96)
0.677
Gal.1
5.53 (0.49)
5.65 (0.56)
0.208
SEZ6L
3.9 (0.92)
3.71 (0.77)
0.175
CAIX
2.5 (1.4)
2.24 (1.2)
0.23
CTSV
1.86 (1.06)
1.9 (1.02)
0.805
ERBB4
4.34 (0.57)
4.19 (0.74)
0.185
X5..NT
8.88 (0.95)
8.89 (0.86)
0.905
LYN
-0.07 (0.39)
-0.11 (0.39)
0.592
VIM
3.78 (0.97)
3.64 (1.18)
0.441
CD160
3.71 (0.74)
3.54 (0.77)
0.183
TNFRSF4
2.69 (0.66)
2.57 (0.65)
0.252 MIC.A.B
3.24 (1.3)
3.14 (1.44)
0.672
WISP.1
4.17 (0.76)
4.18 (0.84)
0.945
CXL17
2.12 (1.12)
2.35 (0.92)
0.163
S100A11
2.97 (0.71)
3 (0.71)
0.799
CXCL13
7.94 (0.73)
7.91 (0.85)
0.82
FR.gamma
7.3 (2.24)
6.94 (2.03)
0.318 aMean log2 value with standard deviation in parenthesis.
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Turbulent rectangular compound open channel flow study using multi-zonal approach
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1
School of Engineering, Faculty of Engineering and Informatics, University of Bradford,
Bradford DB7 1DP, UK Turbulent rectangular compound open channel flow study
using multi‑zonal approach Jaan H. Pu1 Received: 10 September 2018 / Accepted: 12 December 2018 / Published online: 29 December 2018
© The Author(s) 2018 * Jaan H. Pu
j.h.pu1@bradford.ac.uk Environmental Fluid Mechanics (2019) 19:785–800
https://doi.org/10.1007/s10652-018-09655-9 Environmental Fluid Mechanics (2019) 19:785–800
https://doi.org/10.1007/s10652-018-09655-9 ORIGINAL ARTICLE Abstract In this paper, an improved Shiono–Knight model (SKM) has been proposed to calculate the
rectangular compound open channel flows by considering a multi-zonal (MZ) approach in
modelling turbulence and secondary flows across lateral flow direction. This is an effort to
represent natural flows with compound shape more closely. The proposed model improves
the estimation of secondary flow by original SKM model to increase the accuracy of depth-
averaged velocity profile solution formed within the transitional region between different
sections (i.e. between main-channel and floodplain) of compound channel. This proposed
MZ model works by sectioning intermediate zones between floodplain and main-channel
for running computation in order to improve the modelling accuracy. The modelling results
have been validated using the experimental data by national UK Flood Channel Facility. It
has been proven to work reasonably well to model secondary flows within the investigated
compound channel flow cases and hence produce better representation to their flow lateral
velocity profile. Keywords Shiono–Knight method · Secondary flow · Turbulence · Natural flow · Lateral
velocity distribution · Depth-averaged model List of symbols List of symbols
b
Half of the in-bank channel width (m)
B
Half of the whole channel surface width (m)
Dr
Relative depth (−)
f
Friction factor (−)
g
Gravitational acceleration (m/s2)
h
Height of in-bank channel (m)
H
Flow depth (m)
So
Bed slope (−)
t
Time (s)
U*
Shear velocity (m/s)
U
Velocity in x direction (m/s) 123456789)
1 3 456789)
1 3 56789)
3 Environmental Fluid Mechanics (2019) 19:785–800 786 Ud
Depth-averaged velocity (m/s)
Uw
Width-averaged velocity (m/s)
V
Velocity in y direction (m/s)
W
Velocity in z direction (m/s)
x
Streamwise coordinate (m)
X
Water body forces (Pa)
y
Lateral coordinate (m)
z
Coordinate normal to bed (m)
휌
Density of water (kg/m3)
휏
Shear stress (Pa)
휇
Kinematic viscosity (m2/s)
휎
Normal stresses (Pa)
ε
Eddy viscosity (kg/ms)
훤
Secondary flow parameter (kg/(ms2))
휆
Dimensionless eddy viscosity (−)
̄𝜀yx
Depth-averaged eddy viscosity (m2/s)
휏b
Local boundary shear stress (Pa) 1 Introduction In recent years, studies into compound channels have been undertaken due to its impor-
tance in representing the seasonal-impact rivers and floodplains. The naturally formed
compound channel usually consisted of a deeper main-channel known as the in-bank and
shallower floodplains known as the over-bank. This un-even channel could affect the flow
velocity distribution, which could alter its sediment transport pattern [8]. In order to rep-
resent these flows, Shiono and Knight [11] introduced an analytical Shiono–Knight model
(SKM) based on the simplified two-dimensional (2D) Reynolds Averaged Navier–Stokes
(RANS) equations. The model ignored the effect of turbulence created from secondary
currents. The SKM was improved when Shonio and Knight [12] included the effects of
secondary flows within the SKM model, which they also found that the turbulence and
secondary flow occurrences are complicated in compound channel flows due to the lateral
exchange of flow momentum between the main-channel and floodplains. ll
Some compound channel studies further considered the channel-shape-generated turbu-
lence as well as secondary flows to improve flow velocity profile representation (i.e. Ervine
et al. [2] and Tang and Knight [13]). Studies by Yang et al. [17] showed that large-scale
eddies are formed by instability in regions with high velocity variation, such as at the inter-
face between main-channel and its floodplains of a compound channel flow. All these stud-
ies showed that the accurate calculation of secondary currents and hence turbulence are
crucial for estimating the flow velocity.l l
To ease the consideration of secondary flows in SKM model, Shonio and Knight [12]
assigned a secondary flow parameter, 훤 , into their model to calculate flow velocity, where
this approach was commonly adopted in later studies [13–15, 18]. The parameter was cho-
sen to represent each section in those studies to ensure the appropriate flow representa-
tion at main-channel and floodplain. Liao and Knight [3] and [4] further investigated 훤
together with local friction factor (f) and dimensionless eddy viscosity ( 휆 ) for trapezoidal
and rectangular compound channels respectively. They proved the importance of all these
parameters in SKM model, in particular 훤 . From their studies, it has been proposed that 1 3 Environmental Fluid Mechanics (2019) 19:785–800 787 velocity profile mis-representation could happen due to secondary flow assumption used by
the SKM model. This mis-representation error usually occurs due to the lumping of single
parameter within a section (i.e. 1 Introduction within main-channel or floodplain) as a simplifying step at
SKM modelling. Tang and Knight [13] considered the SKM’s boundary conditions associ-
ated with complex compound geometries and proved that by modifying SKM its velocity
profile calculation can be improved for flows with different aspect ratios. Consequently, in
Tang and Knight [14], it has been acknowledged that there is a lack of experimental and
theoretical understanding to 훤 which can lead to the inaccuracy of velocity computation. In Tang and Knight [15], several analytical models have been reviewed and further
tested, including by Ervine et al. [2], Castanedo et al. [1] and Van Prooijen et al. [16]. Ervine et al. [2] based their model by assuming the temporal-mean transverse veloc-
ity component in the flow momentum equation to be fraction of the streamwise depth-
averaged velocity Ud, hence replacing 훤 by KUd
2, where K is a weighted coefficient. In
their model, K was used to account for complex 3D interfacing of the main-channel and
floodplain, which includes the effect of horizontal shear layer, mass exchange in and out
of each section and any expansion and contraction losses from non-prismatic channels. In
Castanedo et al. [1], three different forms of the turbulent shear stress were investigated,
however their model did not take into account the secondary flow effect. Van Prooijen et al. [16] proposed an analytical eddy viscosity model that included the effects of the horizontal
coherent structures as well as the effects of 3D wall-induced turbulence. Tang and Knight
[15] suggested that the large horizontal eddies induced by strong shearing in the sections’
mixing region play a significant role in the flow transverse mixing process. As a result,
various eddy viscosity models that take into account the horizontal coherent structures and
turbulence (i.e. Van Prooijen et al. [16]) have shown promising modelling outcomes.li From studies above, the turbulence and secondary flow are identified to be crucial for
analytical model to represent velocity profile reasonably. The key forming of secondary
flow usually takes place at the interval between sections (i.e. between main-channel and
floodplain) of compound channel where its effect towards velocity is hard to be modelled
accurately. Shonio and Knight [12] and Tang and Knight [13] have suggested that more
complex sub-divided section members within a compound channel may be able to improve
velocity modelling accuracy, but there is complexity to represent additional section’s tur-
bulence and secondary flow. 1 Introduction Due to this, a novel multi-zonal (MZ) model is proposed and
investigated in this study for its suitability to improve common SKM model to represent
velocity profile across rectangular compound channel. The proposed MZ model utilises
extra section to improve the modelling of turbulence and secondary flow within the investi-
gated compound channel flows and hence enhance the accuracy of velocity profile calcula-
tion. The proposed model has also been validated against the UK Flood Channel Facility
(FCF) experimental data. 2.1 Shiono–Knight (SKM) and turbulence modelling Originated from the SKM model by Shiono and Knight [11, 12], Castanedo et al. [1] and Van
Prooijen et al. [16] has proposed methods to improve the SKM’s turbulence modelling. How-
ever, their respective comparisons with experimental data demonstrated that the flow model-
ling improvement mainly restricted to flow regions away from compound sections’ interval. 3 Environmental Fluid Mechanics (2019) 19:785–800 788 Yang et al. [18] considered a model attempting to represent flow velocity profile in rectangular
compound channel. Their proposal was different to the SKM at designing secondary flows and
eddy viscosity by considering momentum transfer.l Yang et al. [18] considered a model attempting to represent flow velocity profile in rectangular
compound channel. Their proposal was different to the SKM at designing secondary flows and
eddy viscosity by considering momentum transfer.l Consequently, it appears that several studies for the compound channel flows were based
around modifying the SKM model due to its modelling capability. Their proposed modifi-
cations to the SKM presented improved results from the original SKM model suggesting its
versatility to adapt to different improved functions. Ergo, in this study, the SKM model will be
adapted and improved. The proposed governing model from Navier–Stokes equations suggested by Shiono and
Knight [11] SKM model can be described as follows (1)
휌X + 휕휎xx
휕x +
휕휏yx
휕y + 휕휏zx
휕z = 휌
(
U 휕U
휕x V 휕U
휕y + W 휕U
휕z + 휕U
휕t
) (1) where 휌 is water density; X is water body force; 휏 is shear stress; 휎 is normal stress; x, y
and z denote streamwise, lateral and vertical directions; t denotes time; and U, V and W are
velocity at x, y and z-direction. The subscripts for 휏 and 휎 indicate their acting direction. In order to form the governing equation for lateral variation of depth-averaged velocity, Ud,
the depth-averaged momentum equation has to be solved for steady uniform turbulent flow in
the streamwise direction. And by considering the dominant horizontal eddies across the chan-
nel, one can get that 휕U∕휕x = 휕휎xx∕휕x = 0 . Furthermore, it is assumed that the velocity vari-
ation is steady i.e. 휕U∕휕t = 0 . As a result, the governing equations becomes (when combined
with the continuity equation) (2)
휌H 휕
휕y(UV) = 휌gSoH +
휕휏yx
휕y H + 휕휏zx
휕z H (2) where H is water flow depth. 2.1 Shiono–Knight (SKM) and turbulence modelling where H is water flow depth. l
Combining the bed shear stress theory with the Darcy-Weisbach equation, the following
uation can be deduced l
Combining the bed shear stress theory with the Darcy-Weisbach equation, the following
equation can be deduced (3)
휕휏zx
휕z =
fU2
d휌
8H (3) where f is friction factor. where f is friction factor. Shiono and Knight [12] stated that the depth-averaged transverse shear stress 휏yx can be
expressed in terms of lateral gradient of Ud as (4)
𝜏yx = 𝜌̄𝜀yx
𝜕Ud
𝜕y (4) where (5)
̄𝜀yx = 𝜆U∗H (5) and ̄𝜀yx is depth-averaged eddy viscosity; 휆 is dimensionless eddy viscosity; and U∗ is shear
velocity. and ̄𝜀yx is depth-averaged eddy viscosity; 휆 is dimensionless eddy viscosity; and U∗ is shear
velocity. Substituting the above into the Eq. (2) together with using the expression of shear velocity
U∗= (휏b∕휌) 1
2 , we could deduce that (6)
휕
휕y
[
H(휌UV)d
]
= 휌gSoH −
f휌U2
d
8
+ 휕
휕y
{
휌휆H2
( f
8
) 1
2
Ud
휕Ud
휕y
} (6) 1 3 1 3 Environmental Fluid Mechanics (2019) 19:785–800 789 where 휏b is the local boundary shear stress; and subscript d denotes depth-averaged charac-
teristic of a parameter.l where 휏b is the local boundary shear stress; and subscript d denotes depth-averaged charac-
teristic of a parameter.l The secondary flows in the depth-averaged Eq. (6) can be assumed as follows [12, 13] (7)
휕
휕y
[H(휌UV)d
] = 훤 (7) In uniform flow, the depth-averaged velocity equation can be expressed as follows [from
Eqs. (6)–(7)] In uniform flow, the depth-averaged velocity equation can be expressed as follows [from
Eqs. (6)–(7)] (8)
Ud =
[
A1e훾y + A2e−훾y + k
] 1
2 where
in which
(8)
Ud =
[
A1e훾y + A2e−훾y + k
] 1
2
(9)
k = 8gSoH
f
(1 −훽) (8) where (9)
k = 8gSoH
f
(1 −훽) (9) in which in which (10)
훽=
훤
휌gSoH and 훾=
√
2
휆
( f
8
) 1
4 1
H (10) In Eqs. (8)–(10), 훤 is secondary flow parameter; So is bed slope; g is gravitational accelera-
tion; and A1, A2, k, 훽 and 훾 are all coefficients used for depth-averaged velocity formulation. In Eqs. 2.1 Shiono–Knight (SKM) and turbulence modelling (8)–(10), 훤 is secondary flow parameter; So is bed slope; g is gravitational accelera-
tion; and A1, A2, k, 훽 and 훾 are all coefficients used for depth-averaged velocity formulation. 2.2 Boundary conditions The schematic diagram for the rectangular compound channel flow is showing at Fig. 1, where
it consists of two sections within the channel: deeper main-channel and shallower floodplain. The schematic diagram for the rectangular compound channel flow is showing at Fig. 1, where
it consists of two sections within the channel: deeper main-channel and shallower floodplain. Liao and Knight [3, 4] discussed the boundary conditions produced by compound channel
in which they suggested each section should possess separate constants 훤 , f and 휆 . This is
because the velocity at the main-channel and floodplain is not continuous but affected from
the region where the sections meet. They further proposed that (11)
휕U(1)
d
휕y
||||||y=0
= 0 (11) 1 3
Fig. 1 Rectangular compound
channel and dimensions Fig. 1 Rectangular compound
channel and dimensions Fig. 1 Rectangular compound
channel and dimensions Fig. 1 Rectangular compound
channel and dimensions 3 3 Environmental Fluid Mechanics (2019) 19:785–800 790 (12)
U(2)
d
|||y=B = 0 (12) The superscript in each boundary refers to the section number (i.e. 1 for main channel
and 2 for floodplain). By approximating this relationship using proportional approach, Liao
and Knight [3] produced (13)
Ud(2) ≈훼uUd(1) (13) where the following relationships can describe 훼u at Eq. (13) above (14)
훼ub
B
= KuDr + Bu (14) in which (15)
Dr = (H −h)∕H (15) (16)
Ku = −0.123601
(B
b
)2
+ 0.518198
(B
b
)
+ 0.019269 (16) (17)
Bu = 0.0179595
(B
b
)2
−0.166066
(B
b
)
+ 0.600033 (17) 훼u , Ku , Bu are coefficient constants; Dr is relative depth [as showing at Eq. (14)]; and B, b
and h are half of the whole channel surface width, half of the in-bank channel width and
flow depth at in-bank channel respectively (as can be referred to Fig. 1). 훼u , Ku , Bu are coefficient constants; Dr is relative depth [as showing at Eq. (14)]; and B, b
and h are half of the whole channel surface width, half of the in-bank channel width and
flow depth at in-bank channel respectively (as can be referred to Fig. 1). l
p
p
y
g
Therefore, from Eq. where (25)
C1 = 8gSoH
f1
(1 −훽1), C2 = 8gSo(H −h)
f2
(1 −훽2), 훽1 =
훤1
휌gSoH , and 훽2 =
훤2
휌gSo(H −h) 2.2 Boundary conditions (13), the boundary conditions at y = b can be written as (18)
훼uU(1)
d
|||y=b = U(2)
d
|||y=b (18) (19)
휕U(1)
d
휕y
||||||y=b
=
휕U(2)
d
휕y
||||||y=b (19) After all four boundary conditions at Eqs. (11), (12), (18) and (19) are applied into
Eq. (8), A coefficients become (20)
A1 = A2 = A3훾2e훾2b −A4훾2e−훾2b
2훾1훼u sinh(훾1b) (20) where (21)
A3 = −C2e−훾2B −A4e−2훾2B (21) (22
C2 −훼2
uC1 +
[
훼u훾2C2e훾2(b−B)]
∕
[
훾1 tanh(훾1b)
]
−C2e
훾2(b−B)
−e−훾2b −
[
훼u훾2e훾2(b−2B)]
∕
[
훾1 tanh(훾1b)
]
−(훼u훾2e−훾2b)∕
[
훾1 tanh(훾1b)
] 1 3
(22)
A4 =
C2 −훼2
uC1 +
[
훼u훾2C2e훾2(b−B)]
∕
[
훾1 tanh(훾1b)
]
−C2e
훾2(b−B)
e
훾2(2b−B) −e−훾2b −
[
훼u훾2e훾2(b−2B)]
∕
[
훾1 tanh(훾1b)
]
−(훼u훾2e−훾2b)∕
[
훾1 tanh(훾1b)
] (22) 1 3 3 Environmental Fluid Mechanics (2019) 19:785–800 791 in which, A3, A4, C1, C2, 훾1 and 훾2 are all coefficients for SKM boundary conditions. which, A3, A4, C1, C2, 훾1 and 훾2 are all coefficients for SKM boundary conditions. fi
In Eqs. (20)–(22), A1 and A2 are related to the symmetrical and wall-slip conditions of
the channel, whereas their associated A3 and A4 are related to flow continuity between sec-
tions. Substituting A coefficients into the depth-averaged velocity equation gives (23)
Ud(1) =
√
2A1 cosh(훾1y) + C1 (23)
Ud(1) =
√
2A1 cosh(훾1y) + C1
(24)
Ud(2) =
√
A3e훾2y + A4e−훾2y + C2 (23) (24)
Ud(2) =
√
A3e훾2y + A4e−훾2y + C2 (24) where 3.1 Modelling parameters In analytical modelling, the parameters for secondary flow and eddy viscosity are crucial
to ensure a precise representation to the compound channel flows. As proven in the cali-
bration processes by Shiono and Knight [12], Tang and Knight [13, 14, 15], and Liao and
Knight [3, 4], those parameters significantly influenced the performance of modelling cal-
culation, in particular at near-wall and section transitional regions. Through the calibra-
tion using the UK-FCF experimental data, this study obtains a set of calibrated secondary
flow and dimensionless eddy viscosity parameters. As for the friction factor, it has been
obtained using the measured shear stress in the experiment, hence same coefficients used
by the SKM model will be employed in this study [3, 14].l In Figs. 2, 3, 4, 5, 6 and 7, the results of the proposed secondary flow and dimensionless
eddy viscosity parameters are compared to the UK-FCF experimental data for rectangular Fig. 2 FCF0804 depth-averaged velocity comparison between the proposed model and literatures Fig. 2 FCF0804 depth-averaged velocity comparison between the proposed model and literatures Environmental Fluid Mechanics (2019) 19:785–800 792 compound channel flows. The experimental conditions and width-averaged velocity (Uw)
for tests in Figs. 2, 3, 4, 5, 6 and 7 are described in Table 1, where the utilised B/H aspect
ratio range from 1.98 to 15.99. For comparison purpose, the models by Tang and Knight
Fig. 3 FCF0807 depth-averaged velocity comparison between the proposed model and literatures
Fig. 4 ROS224 depth-averaged velocity comparison between the proposed model and literatures Fig. 3 FCF0807 depth-averaged velocity comparison between the proposed model and literatures Fig. 3 FCF0807 depth-averaged velocity comparison between the proposed model and literatures Fig. 3 FCF0807 depth-averaged velocity comparison between the proposed model and literatures Fig. 4 ROS224 depth-averaged velocity comparison between the proposed model and literatures Fig. 4 ROS224 depth-averaged velocity comparison between the proposed model and literatures compound channel flows. The experimental conditions and width-averaged velocity (Uw)
for tests in Figs. 2, 3, 4, 5, 6 and 7 are described in Table 1, where the utilised B/H aspect
ratio range from 1.98 to 15.99. For comparison purpose, the models by Tang and Knight
[14], and Liao and Knight [3] are also plotted in same Figs. 1 3 3.1 Modelling parameters B/H (−)
So (−)
b (m)
h (m)
B (m)
H (m)
Uw (m/s)
FCF0804
15.99
0.001027
0.750
0.150
3.00
0.18760
0.469
FCF0807
11.99
0.001027
0.750
0.150
3.00
0.25020
0.783
ROS224
8.37
0.002024
0.400
0.050
0.60
0.07165
0.464
ROS245
6.93
0.002024
0.400
0.050
0.60
0.08660
0.692
DWK0404
2.67
0.000966
0.076
0.076
0.30
0.11420
0.379
DWK0406
1.98
0.000966
0.076
0.076
0.30
0.15380
0.497
Table 2 Coefficients summary for compound channel flow tests
Tang and Knight [14]
Liao and Knight [3]
Present study
Experiment no. B/H
λ (1&2)
Γ (1&2)
λ (1&2)
Γ (1&2)
λ (1&2)
Γ (1&2)
FCF0804
15.99
0.07, 0.07
0.25, − 0.10
0.01, 0.10
0.25, −0.15
0.10, 0.09
0.30, − 0.095
FCF0807
11.99
0.08, 0.07
0.60, − 0.25
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.65, − 0.25
ROS224
8.37
0.07, 0.07
0.25, − 0.10
0.01, 0.10
0.25, − 0.15
0.035, 0.055
0.24, − 0.10
ROS245
6.93
0.07, 0.07
0.35, − 0.25
0.01, 0.10
0.25, − 0.15
0.035, 0.055
0.40, − 0.30
DWK0404
2.67
0.07, 0.07
0.34, − 0.092
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.39, − 0.091
DWK0406
1.98
0.03, 0.07
0.55, − 0.187
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.60, − 0.18 Table 1 Experimental conditions for compound channel flow tests Table 1 Experimental conditions for compound channel flow tests
Experiment no. B/H (−)
So (−)
b (m)
h (m)
B (m)
H (m)
Uw (m/s)
FCF0804
15.99
0.001027
0.750
0.150
3.00
0.18760
0.469
FCF0807
11.99
0.001027
0.750
0.150
3.00
0.25020
0.783
ROS224
8.37
0.002024
0.400
0.050
0.60
0.07165
0.464
ROS245
6.93
0.002024
0.400
0.050
0.60
0.08660
0.692
DWK0404
2.67
0.000966
0.076
0.076
0.30
0.11420
0.379
DWK0406
1.98
0.000966
0.076
0.076
0.30
0.15380
0.497 Table 2 Coefficients summary for compound channel flow tests
Tang and Knight [14]
Liao and Knight [3]
Present study
Experiment no. 3.1 Modelling parameters 2, 3, 4, 5, 6 and 7.i It can be observed from the figures that the proposed parameter set gives reasonable
representation to the experimental data, especially at the main-channel flow region. How-
ever, it has not presented adequate representation to the measurements at interface region
between main-channel and floodplain, except for a few flow cases. The models by Tang and
Knight [3, 14] have also shown non-precise representation to the measured velocity profile 1 3 Environmental Fluid Mechanics (2019) 19:785–800 793 793
Environmental Fluid Mechanics (2019) 19:785–800
at the main-channel and floodplain interface. To further compare the proposed parameter
sets with suggestions from Tang and Knight [14] and Liao and Knight [3], Table 2 has
Fig. 5 ROS245 depth-averaged velocity comparison between the proposed model and literatures
Fig. 6 DWK0404 depth-averaged velocity comparison between the proposed model and literatures Fig. 5 ROS245 depth-averaged velocity comparison between the proposed model and literatures Fig. 5 ROS245 depth-averaged velocity comparison between the proposed model and literatures Fig. 6 DWK0404 depth-averaged velocity comparison between the proposed model and literatures Fig. 6 DWK0404 depth-averaged velocity comparison between the proposed model and literatures at the main-channel and floodplain interface. To further compare the proposed parameter
sets with suggestions from Tang and Knight [14] and Liao and Knight [3], Table 2 has
been produced. From the table, it can be observed that the proposed calibrated parameters
fall within a reasonable range to those suggested in literatures.l From the model validation process, it has been found that the flow tests with high
aspect ratio of B/H (i.e. FCF0804, FCF0807 and ROS224) can be represented quite
accurately by SKM models (by Liao and Knight [3] and Tang and Knight [14]), and by
the present calibrated model. For other flow tests with smaller B/H ratios, all models
show incapability to represent the compound channel flows with high accuracy. The key 1 3 Environmental Fluid Mechanics (2019) 19:785–800 794 Fig. 7 DWK0406 depth-averaged velocity comparison between the proposed model and literatures Fig. 7 DWK0406 depth-averaged velocity comparison between the proposed model and literatures Table 1 Experimental conditions for compound channel flow tests
Experiment no. 3.1 Modelling parameters B/H
λ (1&2)
Γ (1&2)
λ (1&2)
Γ (1&2)
λ (1&2)
Γ (1&2)
FCF0804
15.99
0.07, 0.07
0.25, − 0.10
0.01, 0.10
0.25, −0.15
0.10, 0.09
0.30, − 0.095
FCF0807
11.99
0.08, 0.07
0.60, − 0.25
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.65, − 0.25
ROS224
8.37
0.07, 0.07
0.25, − 0.10
0.01, 0.10
0.25, − 0.15
0.035, 0.055
0.24, − 0.10
ROS245
6.93
0.07, 0.07
0.35, − 0.25
0.01, 0.10
0.25, − 0.15
0.035, 0.055
0.40, − 0.30
DWK0404
2.67
0.07, 0.07
0.34, − 0.092
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.39, − 0.091
DWK0406
1.98
0.03, 0.07
0.55, − 0.187
0.01, 0.10
0.25, − 0.15
0.10, 0.09
0.60, − 0.18 Table 2 Coefficients summary for compound channel flow tests inaccuracy happens at the flow interface between main-channel and floodplain prompt-
ing the use of different approach to model this region of flow is needed. The following
section will describe a new multi-zonal (MZ) approach to improve compound channel
study. Environmental Fluid Mechanics (2019) 19:785–800 795 Table 3 Experimental conditions for flows with small aspect ratios for multi-zonal (MZ) model tests
Experiment no. B/H (−)
So (−)
b (m)
h (m)
B (m)
H (m)
Uw (m/s)
DWK0406
1.98
0.000966
0.076
0.076
0.30
0.1538
0.497
ROS255
6.36
0.002024
0.400
0.050
0.60
0.0950
0.783
ROS250
6.55
0.002024
0.400
0.050
0.60
0.0920
0.738
ROS245
6.93
0.002024
0.400
0.050
0.60
0.0866
0.692
ROS240
7.10
0.002024
0.400
0.050
0.60
0.0846
0.674
Fig. 8 Secondary flow compo-
nents at interface zone between
main-channel and floodplain. (1)–(4) represent zones within
the compound channel flow 3.2 Proposed multi‑zonal (MZ) approach
Table 3 Experimental conditions for flows with small aspect ratios for multi-zonal (MZ) model tests
Experiment no. B/H (−)
So (−)
b (m)
h (m)
B (m)
H (m)
Uw (m/s)
DWK0406
1.98
0.000966
0.076
0.076
0.30
0.1538
0.497
ROS255
6.36
0.002024
0.400
0.050
0.60
0.0950
0.783
ROS250
6.55
0.002024
0.400
0.050
0.60
0.0920
0.738
ROS245
6.93
0.002024
0.400
0.050
0.60
0.0866
0.692
ROS240
7.10
0.002024
0.400
0.050
0.60
0.0846
0.674
Fig. 8 Secondary flow compo-
nents at interface zone between
main-channel and floodplain. (1)–(4) represent zones within
the compound channel flow xperimental conditions for flows with small aspect ratios for multi-zonal (MZ) model tests 3.2 Proposed multi‑zonal (MZ) approach As proven in the previous section, some compound channel flows have not been represented
precisely by SKM approach in particular flows with small aspect ratio. Motivated by this fact,
a multi-zonal (MZ) approach has been proposed in this study to improve the flow modelling
at the interface flow region between floodplain and main-channel. The UK-FCF experiments
with small aspect ratio of B/H and flow depth H are benchmarked to find out the accuracy of
the proposed MZ model. These small aspect ratio tests include various ROS and DWK flow
cases, where their flow conditions and Uw are outlined at Table 3. All tests in this section have
aspect ratio from 1.98 to 7.10.l The MZ model works by sectioning out an additional zone in the interface between flood-
plain and main-channel. To model this zone, the MZ model will consider stronger secondary
flow created at the step of compound channel and implement this additional impact into 훽 for
SKM modelling, which is presented at the following Eqs. (26)–(27). Fundamentally, the second-
ary flow at compound step consists of two components acting in opposed manner to each other
(refer to schematic diagram at Fig. 8), as suggested in Nezu and Nakagawa [5]. By considering
these components of secondary flow, the SKM calculation could be improved and hence the
consideration of Ud. 1 3
(26)
훽
1 =
훤
1
휌gSoH
(27)
훽
2 =
훤
2
휌gSo(H −h) (26)
훽
1 =
훤
1
휌gSoH (26) (27)
훽
2 =
훤
2
휌gSo(H −h) (27) 1 3 Environmental Fluid Mechanics (2019) 19:785–800 796 where, 훽
′
1 and 훽
′
2 are coefficients at zones 2 and 3 within the flow interface area (showing
at Fig. 8 below); and 훤
1 = 훤1 + 휑1 and 훤
2 = 훤2 + 휑2 , in which 휑1 and 휑2 represent the 훤
components due to the escalated secondary flow at zones 2 and 3 within the flow interface
area. where, 훽
′
1 and 훽
′
2 are coefficients at zones 2 and 3 within the flow interface area (showing
at Fig. 8 below); and 훤
1 = 훤1 + 휑1 and 훤
2 = 훤2 + 휑2 , in which 휑1 and 휑2 represent the 훤
components due to the escalated secondary flow at zones 2 and 3 within the flow interface
area. 3.2 Proposed multi‑zonal (MZ) approach It has been found in this study that the interface zones 2 and 3 in Fig. 8 are accounted
for about 10-25% of the distance for main-channel and floodplain respectively (agreed with
findings by experimental studies on various channel flows in [6, 10]. In comparison, the
shallower zone 3 is lengthier than zone 2 as the secondary flow has less depth of water to
evolve vertically. Since both secondary flow 휑1 and 휑2 components act at the counter-rota-
tional manner, they are estimated to have positive and negative impact to 훤
′
1 and 훤
′
2 respec-
tively. In the exact middle of the meeting point between main-channel and floodplain, 휑1
and 휑2 are set at zero as both secondary flow components neutralised. 1 3
2
yl
p
Fig. 9 DWK0406 depth-averaged velocity comparison between models with and without MZ model
Fig. 10 ROS255 depth-averaged velocity comparison between models with and without MZ model Fig. 9 DWK0406 depth-averaged velocity comparison between models with and without MZ model Fig. 9 DWK0406 depth-averaged velocity comparison between models with and without MZ model 1 3
Fig. 10 ROS255 depth-averaged velocity comparison between models with and without MZ model Fig. 10 ROS255 depth-averaged velocity comparison between models with and without MZ model Fig. 10 ROS255 depth-averaged velocity comparison between models with and without MZ model 1 3
Fig. 10 ROS255 depth-averaged velocity comparison between models with and without MZ model 1 3 Environmental Fluid Mechanics (2019) 19:785–800 797 Fig. 11 ROS250 depth-averaged velocity comparison between models with and without MZ model Fig. 11 ROS250 depth-averaged velocity comparison between models with and without MZ model Fig. 11 ROS250 depth-averaged velocity comparison between models with and without MZ model Fig. 12 ROS245 depth-averaged velocity comparison between models with and without MZ model Fig. 12 ROS245 depth-averaged velocity comparison between models with and without MZ model For validation, it can be observed from Figs. 9, 10, 11, 12 and 13 that for all compound
channel flow cases tested, the conventional model without considering MZ approach shows
inaccuracy in calculating the flow velocity at the main-channel and floodplain interface
region due to the mis-interpretation of secondary current intensity. This interface zone
velocity profile modelling has been improved by the present MZ model as benchmarked by
the UK-FCF experimental data. 3.2 Proposed multi‑zonal (MZ) approach Table 4 shows the root-mean square error (RMSE) com-
parison for interface zone’s velocity calculations with and without MZ approach bench-
marked by the measured data. The compared error fraction in the table reveals that the MZ
model works better in flow cases with higher velocity, i.e. ROS245, ROS250 and ROS255, 1 1 3 3 Environmental Fluid Mechanics (2019) 19:785–800 798 Fig. 13 ROS240 depth-averaged velocity comparison between models with and without MZ model Fig. 13 ROS240 depth-averaged velocity comparison between models with and without MZ model Table 4 Root-mean square error (RMSE) analysis benchmarked by measured data at the main-channel and
floodplain interface region
Test
Width-average of
measured velocity
(m/s)
Averaged RMSE for
calculation Without
MZ (m/s)
Averaged RMSE for
calculation with MZ
(m/s)
Comparative error
fraction (non-MZ to
MZ model) (−)
DWK0406
0.4968
0.036894
0.013479
2.737
ROS240
0.6737
0.097826
0.033446
2.925
ROS245
0.6922
0.066959
0.012258
5.462
ROS250
0.7381
0.125526
0.028812
4.357
ROS255
0.7834
0.108704
0.031258
3.478 le 4 Root-mean square error (RMSE) analysis benchmarked by measured data at the main-channel and
dplain interface region when compared to flows with low velocity. This is due to the fact that those flow cases with
higher flow velocity were subjected to more intense secondary current at the interface area,
which in terms causing larger differences between MZ and non-MZ models.ll when compared to flows with low velocity. This is due to the fact that those flow cases with
higher flow velocity were subjected to more intense secondary current at the interface area,
which in terms causing larger differences between MZ and non-MZ models.ll f
The investigated narrow channel flow cases have been influenced by secondary cur-
rents which impact the flows towards left and right of the compound step. Compared
to wide channel flows, the side-wall and compound step effects are crucial for nar-
row flow cases and it has been proven that the 2D depth-averaged model could not
effectively represent them without combining with innovative approach (i.e. by Nezu
and Nakagawa [5], Pu et al. [9], Pu [7]). In the tests of this study, the proposed MZ
model has proven to reasonably improve the conventional SKM model deduced from
2D depth-averaged Navier–Stokes flow assumption. It has been shown that by addi-
tional sectioning of the flow model laterally, the compound channel 2D modelling
accuracy could be improved. 3.2 Proposed multi‑zonal (MZ) approach Theoretically, the MZ approach works by considering
smaller width of the channel sections and hence able to reduce the inaccuracy of SKM 1 3 1 Environmental Fluid Mechanics (2019) 19:785–800 799 calculation, by reducing error caused by representing whole section with a single sim-
plified 훤. calculation, by reducing error caused by representing whole section with a single sim-
plified 훤. 4 Conclusions The Shiono–Knight model (SKM) has been investigated to estimate the flow velocity pro-
file within rectangular compound channel which has been subjected to turbulence and sec-
ondary flows. Due to its 2D Navier–Stokes assumption, SKM has found to represent the
narrow channel flow with less accuracy compared to wide channel flows. A multi-zonal
(MZ) approach has been proposed to improve this modelling weakness. In the proposed
MZ model, the investigated compound channel flows have been sectioned with extra zones
in the interface region between main-channel and floodplain where the escalated secondary
flow effect has been considered. To validate the proposed MZ model, it was used to calcu-
late and compare with the experimental data by national UK-FCF facility. The comparison
with measured data showed that the proposed MZ model represents the narrow flows with
better accuracy than the SKM approach for the rectangular compound channel flow cases. This study has also proven that the modelling of narrow 3D-characterised secondary flow
could be improved by considering extra zones’ turbulence and secondary current model-
ling in the analytical model. Acknowledgement The help of Ms. Faye Lynch (the author’s former undergraduate student) is greatly
appreciated in the model testing and formulations. OpenAccess This article is distributed under the terms of the Creative Commons Attribution 4.0 Interna-
tional License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution,
and reproduction in any medium, provided you give appropriate credit to the original author(s) and the
source, provide a link to the Creative Commons license, and indicate if changes were made. OpenAccess This article is distributed under the terms of the Creative Commons Attribution 4.0 Interna-
tional License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution,
and reproduction in any medium, provided you give appropriate credit to the original author(s) and the
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A technique to train new oculomotor behavior in patients with central macular scotomas during reading related tasks using scanning laser ophthalmoscopy: immediate functional benefits and gains retention
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BMC Ophthalmology
BioMed Central
Open Access
Research article
A technique to train new oculomotor behavior in patients with
central macular scotomas during reading related tasks using
scanning laser ophthalmoscopy: immediate functional benefits and
gains retention
Anouk Déruaz*1,2, Mira Goldschmidt1, Andrew R Whatham1,
Christophe Mermoud1, Erika N Lorincz1, Armin Schnider3 and
Avinoam B Safran1
Address: 1Ophthalmology Clinic, Department of Clinical Neurosciences and Dermatology, Geneva University Hospitals, Switzerland, 2Perception
and Eye Movement Laboratory, Department of Neurology and Department of Clinical Research, University of Bern, Inselspital, Switzerland and
3Division of Rehabilitation, Department of Clinical Neurosciences and Dermatology, Geneva University Hospital, Switzerland
Email: Anouk Déruaz* - aderuaz@yahoo.fr; Mira Goldschmidt - mira@urbanet.ch; Andrew R Whatham - a_whatham@hotmail.com;
Christophe Mermoud - cmermoud@bluewin.ch; Erika N Lorincz - erika.lorincz@hcuge.ch; Armin Schnider - armin.schnider@hcuge.chj;
Avinoam B Safran - a.b.safran@hcuge.ch
* Corresponding author
Published: 23 November 2006
BMC Ophthalmology 2006, 6:35
doi:10.1186/1471-2415-6-35
Received: 10 August 2006
Accepted: 23 November 2006
This article is available from: http://www.biomedcentral.com/1471-2415/6/35
© 2006 Déruaz et al; licensee BioMed Central Ltd.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Background: Reading with a central scotoma involves the use of preferred retinal loci (PRLs) that enable both letter resolution
and global viewing of word. Spontaneously developed PRLs however often privilege spatial resolution and, as a result, visual span
is commonly limited by the position of the scotoma. In this study we designed and performed the pilot trial of a training
procedure aimed at modifying oculomotor behavior in subjects with central field loss. We use an additional fixation point which,
when combined with the initial PRL, allows the fulfillment of both letter resolution and global viewing of words.
Methods: The training procedure comprises ten training sessions conducted with the scanning laser ophthalmoscope (SLO).
Subjects have to read single letters and isolated words varying in length, by combining the use of their initial PRL with the one
of an examiner's selected trained retinal locus (TRL). We enrolled five subjects to test for the feasibility of the training technique.
They showed stable maculopathy and persisting major reading difficulties despite previous orthoptic rehabilitation. We evaluated
ETDRS visual acuity, threshold character size for single letters and isolated words, accuracy for paragraphed text reading and
reading strategies before, immediately after SLO training, and three months later.
Results: Training the use of multiple PRLs in patients with central field loss is feasible and contributes to adapt oculomotor
strategies during reading related tasks. Immediately after SLO training subjects used in combination with their initial PRL the
examiner's selected TRL and other newly self-selected PRLs. Training gains were also reflected in ETDRS acuity, threshold
character size for words of different lengths and in paragraphed text reading. Interestingly, subjects benefited variously from the
training procedure and gains were retained differently as a function of word length.
Conclusion: We designed a new procedure for training patients with central field loss using scanning laser ophthalmoscopy.
Our initial results on the acquisition of newly self-selected PRLs and the development of new oculomotor behaviors suggest that
the procedure aiming primarily at developing an examiner's selected TRL might have initiated a more global functional adaptation
process.
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Background
Loss of reading ability is a leading complaint of subjects
with central scotomas due to macular disorders, such as
age-related macular degeneration (AMD) [e.g. [1,2]]. As
fixation frequently shifts to a non-foveal location, an
adaptation of the oculomotor system is necessary to
search for, position and stabilize visual targets [3]. During
such an adaptation, individuals develop one or several
eccentric locations in the visual field that enable them to
achieve fixation and reading [1,3-11]. These eccentric
locations are referred to as preferred retinal loci (PRLs)
[9]. Eccentric PRLs develop in approximately eighty-four
percent of affected eyes [12]. The factors determining the
development of a PRL at a precise location relative to the
scotoma are still not well known. Previous studies
reported that the PRL was located more frequently to the
left of the scotoma (34% to 63% of the cases) [12,13].
Other studies showed that different PRLs could be used
predictably according to various parameters, such as target
size or background illumination [14-16]. In subjects with
macular disorders, two factors strongly affect reading ability, namely global viewing of words related to the reading
visual span, and character discrimination related to spatial
resolution [17-19]. Global viewing of words refers to the
ability to perceive the word in its entirety, to estimate its
number of letters and to perform saccades to completely
uncover it. Character discrimination refers to the ability to
correctly read individual letters. Spontaneously selected
PRLs do not consistently meet both requirements. In our
previous investigations we found that multiple PRLs with
complementary functional advantages could be used in a
coordinated reading strategy [5,6,20] and we emphasized
that the two essential capacities for reading could be allocated to different PRLs used in a combined fashion [5].
Whether and how individuals with central visual field loss
could be trained in eccentric fixation is an important issue
for the management of people suffering from age-related
macular degeneration. A limited number of training procedures using the Scanning Laser Ophthalmoscope (SLO)
to readapt fixation behavior during reading have been
reported so far. Nilsson, Frennesson and Nilsson [21]
investigated the influence of the SLO training structured
in sessions scheduled to be one hour a week over two to
seven weeks. In this study, subjects with age-related macular degeneration were trained to use a proposed TRL
located above or below the lesion and to renounce to their
spontaneously developed PRL. To our knowledge, the feasibility of a training process aiming at developing multiple PRLs has not yet been demonstrated.
Therefore the purpose of this investigation was to evaluate, in subjects with age-related macular degeneration, the
use of the SLO to train eye movement strategies for reading, based on a combined use of two preferred retinal loci.
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The rationale for our selection of SLO training procedures
was based on the observation that the use of more than
one PRL in a structured reading strategy can meet the
requirements for both global viewing and detailed discrimination [5-7,20]. Moreover, considering that intensive training over short periods of time is beneficial in
neuro-rehabilitation [22-26], and that reorganization of
the oculomotor system for eccentric reading can be
achieved using an intensive training over a two-weeks
period in experimental conditions [27], we designed our
training protocol accordingly. We tested the feasibility of
our procedure on subjects who reported an altered perception of words, despite conventional reading rehabilitation. Their main complaint was that they missed parts of
words, thus expressing their lack of adapted global viewing abilities. We therefore attempted to train them to use
in combination with their spontaneously developed PRL,
an additional retinal locus, the trained retinal locus or
TRL, fulfilling the needs for global viewing, i.e. allowing
the perception of the entire word.
Methods
The training procedure comprises three steps. Firstly, a
baseline assessment of visual function and reading ability
on isolated words is carried out. Secondly, the SLO training is conducted over ten sessions of one hour each.
Finally, reading performance and reading strategies are
evaluated both immediately after the completion of the
SLO training process and three months later.
General procedure
Subjects' evaluation includes visual acuity measurement,
scotoma delineation, identification of spontaneously
developed PRLs and characterization of oculomotor
behavior during reading. Visual acuity is first determined
for both eyes with ETDRS charts (Precision Vision Ltd,
USA) (table 1). Based on the values obtained, the remaining evaluation and the training procedure are conducted
in the eye with the best visual acuity. For the SLO experiments, subjects are comfortably seated forehead rested at
46 cm in front of the SLO screen, which provides images
of 32 × 22 degrees of visual angle [7]. Stimuli are presented on the SLO screen at a luminance of 30 000 trolands (obtained using the conversion factor reported in
Nygaard & Schuchard (2001) [28]). Experiments are conducted in a darkened room.
Step I: initial evaluation
The scotoma is delineated using the microperimetry software of the scanning laser ophthalmoscope (SLO,
Scotometry 3.0, Rodenstock, Germany). The testing
method is based on the one described by Sunness et al.
(1995) [29]. Briefly, the patient is instructed to fixate a
cross. The investigator firstly chooses a retinal landmark
on the SLO real-time image that serves as a reference
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Table 1: Subject's clinical characteristics
Subjects
MT
SL
LG
AA
GM
Age
(years)
81
76
80
78
79
Visual acuity (logMAR)
Right eye
Left eye
1.1
1.46
1.24
1.12
1.5
1.43
2.2
1.5
1.02
1.36
Selected eye
Contrast sensitivity
in the selected eye
(log unit)
RE
RE
RE
LE
LE
1.6
1.3
1.2
1.6
1.55
Visual acuity was measured with ETDRS charts. Selected eye: eye with the better visual acuity selected for training.
point. Subsequently stimulus points are presented for 500
msec on sites of interest. The order of presentation is randomized. The patient has to press a button each time he
sees a stimulus and the response is recorded on the computer. After the completion of the testing a final fundus
image is grabbed, where the investigator specifies the
position of the landmark. The data points are displayed
on this image with different symbols for seen and unseen
stimuli. Subsequently we present through the SLO to each
subject a series of 32 words varying in length to enable the
subjects to become familiar with the testing procedure.
Baseline parameters of the training procedure are evaluated for the threshold character size for single letters, isolated words and paragraphed text, for the accuracy of
words recognition during paragraphed text reading and
for the oculomotor strategies while reading isolated words
and paragraphed text.
Evaluation of threshold character size for single letters and isolated
words
Threshold character size for single letters and isolated
words is assessed using three different lists of 32 items of
8 different character sizes (1.5 to 0.8 logMAR) that we
project with the SLO in a random order onto the retina.
Threshold character size is defined as the smallest size at
which a correct verbal report of the isolated letter or word
can be made at the first attempt. The results obtain for the
three different lists are averaged. Isolated words are of
three types: 3 and 4-letter words, 6 and 7-letter words and
9 and 10-letter words (table 2). Words are matched for frequencies (> 1/1000) and grammatical category between
word-length using the Brulex database [30]. Matching
words according to these two parameters makes impossible to use words of a single length in a group. Presenting
words of different lengths in a random order is intended
to restrict subjects' prediction of the word based on its
number of letters. Each word is presented only once to
each subject during the whole experiment.
Evaluation of paragraphed text reading
Paragraphed text reading is evaluated using texts from elementary school handbooks (children between 8 and 12
years old) to reduce possible difficulties linked with lexicon and grammar contents. Threshold character size is
evaluated using the same eight sizes previously used for
isolated words and is defined as the smallest size at which
the subject is able to read the text without complaining
about character size. For each text size read, we evaluate
reading accuracy by determining the percentage of words
read correctly at first attempt for each character size. Presentation time of visual stimuli is not restricted.
Analyses of reading strategies
Analyses of reading strategies are conducted according to
the method we previously described [5]. To summarize,
the SLO is used in conjunction with a program developed
in our laboratory for projecting letters, words or paragraphed texts directly onto the ocular fundus. Images of
the ocular fundus and the superimposed words obtained
during the reading task are recorded onto digital videotape, at a frequency of 25 frames/second. Another program developed in our laboratory allows extracting eye
movements automatically by comparing the frame-byframe shift of the image position [31]. Scotoma borders
and some major retinal landmarks, such as the optic disc
and blood vessels are outlined from the microperimetry
results to produce a schematic image. Finally, the program
reconstructs a "cartoon" of the sequence of the eye movements during reading (e.g. figures 3, 4, 5, 6, 7). From these
cartoons, we analyse the oculomotor behavior and identify the PRLs employed. We defined a PRL as an area
repeatedly and consistently used during reading isolated
words and paragraphed text. This area is used by the
patient to perform saccades and fixations while deciphering the presented word or segment of text.
Step II: training procedure
As previously mentioned, reading with a central scotoma
involves the use of PRLs that enable both letter resolution
and global viewing of words. The spontaneously devel-
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Patients
Before
training
Immediately
after training
Threshold character size for words (logMAR)
3 months
later
Before training
Threshold character size for paragraphed text (logMAR)
Immediately after SLO training
3 months later
1
3 and 4
6 and 7
9 and 10
1
3 and 4
6 and 7
9 and 10
1
3 and 4
6 and 7
9 and 10
Before
training
Immediately
after training
3 months later
MT
1.1
0.96
0.9
1.0
1.1
1.4
1.4
0.9
1.3
1.2
1.3
0.9
1.2
1.4
1.5
1.4
1.3
1.5
SL
1.46
1.1
1.14
1.0
1.2
1.4
1.4
0.9
1.1
1.2
1.2
0.9
1.2
1.2
1.2
1.4
1.2
1.4
LG
1.24
1.2
1.12
1.1
1.2
1.5
1.4
0.8
1.0
1.2
1.0
0.8
0.8
1.1
1.2
1.4
1.2
1.2
AA
1.02
0.84
0.86
1.0
1.1
1.1
1.1
0.8
1.0
1.1
1.0
0.8
1.0
1.0
1.1
1.2
1.1
1.2
GM
1.36
1.2
1.1
1.15
1.5
> 1.5
> 1.5
0.8
1.1
1.2
1.1
0.9
1.2
1.2
1.3
1.5
1.4
1.5
1: 1 letter; 3 and 4: words of 3 and 4 letters; 6 and 7: words of 6 and 7 letters; 9 and 10: words of 9 and 10 letters. The mean value obtained for each patient
and for each stimulus type is reported.
Table 3: Retinal areas used for reading and eye movements to completely uncover presented words
BMC Ophthalmology 2006, 6:35
Subjects
Areas used to decipher single letters and isolated words
Areas used to read paragraphed text
Ocular movements to entirely
uncover presented words
Before SLO
training
Immediately after SLO
training
Three months after SLO
training
Before SLO
training
Immediately after SLO
training
Three months after SLO
training
Before SLO
training
Immediately
after SLO
training
Three
months after
SLO training
MT
PRL: 1
PRL: 1
PRL: 1
PRL: 1
PRL: 1 nPRLs: 1
PRL: 1 nPRLs: 2
No
Yes
No
SL
PRL: 1
PRL: 1 TRL: 1
PRL: 1 TRL: 1 nPRLs: 2
PRL: 1
PRL: 1 nPRLs: 2
PRL: 1 TRL: 1 nPRLs: 2
No
Yes
Yes
LG
PRL: 1
PRL: 1 TRL: 1 nPRL: 1
PRL: 1 TRL: 1 nPRL: 1
PRL: 1
PRL: 1 TRL: 1 nPRLs: 1
PRL: 1 TRL: 1
No
Yes
Yes
AA
PRL: 1
PRL: 1 TRL: 1 nPRL: 1
PRL: 1 TRL: 1 nPRL: 1
PRL: 1
PRL: 1 TRL: 1
PRL: 1 TRL: 1
Yes
Yes
Yes
GM
PRL: 0
TRL: 1
TRL: 1 nPRLs: 2
PRL: 0
TRL: 1
TRL: 1
Yes
Yes
Yes
PRL: initial PRL; TRL: examiner's selected TRL; nPRL: additional newly self-selected PRLs. Following the training four subjects (SL, LG, AA, GM) used either the examiner's selected TRL and/or newly
self-selected PRLs both for isolated words' deciphering and for paragraphed text reading. Note that, before training, only two subjects were able to perform eye movements to entirely uncover
isolated words, either sequentially using the initial PRL, or at once using the examiner's selected TRL. Immediately following training, all subjects were able to achieve the task. In four out of five
subjects this capacity was retained three months following SLO training. Note also that the subject that did not retain the ability to uncover words also did not retain any functional training gains.
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ETDRS acuity (logMAR)
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Table 2: ETDRS acuity and threshold character sizes for single letters, isolated words and paragraphed text
BMC Ophthalmology 2006, 6:35
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Figure 1topography
Scotoma
Scotoma topography. Scotoma borders were plotted using scotometry, with the brightest (Goldman III) stimulus. In subjects MT, SL and GM, parts of scotoma borders could not be delineated, as the scotoma was not always entirely visible within
the SLO analysis area. Locations of the initial preferred retinal locus (PRL
locus (TRL
) and the examiner's selected trained retinal
) are indicated, as well as the visual acuity measured at these locations with visumetry.
oped PRLs often privilege spatial resolution. We thus
designed a procedure that intended to train the use of an
additional area, the examiner's selected trained retinal
locus (TRL), where the presented word could be perceived
in its entirety. This area is generally situated either above
or below the lesion on the SLO image. However, it is
worth mentioning that our SLO approach is also valuable
to train areas of high spatial resolution to optimize reading.
Among potentially valuable retinal locations, the area
closest to the initial PRL is selected. Visual acuity is evaluated at the initial PRL and at the examiner's selected TRL
(figure 1). This involves the presentation of a tumbling
"E" in one of four orientations with the SLO using the visumetry program 3.0. Stimuli are presented in 0.1 logMAR
steps. With this technique, visual acuity is defined as the
smallest stimulus size at which a correct answer is
obtained for three out of four presented stimuli.
The training process involves the projection of single letters and isolated words onto the retina using the SLO. To
facilitate the control of eye position at the beginning of
the procedure, fixation is anchored using a cross while
words are projected onto the examiner's selected TRL (figure 2). Subjects are instructed to fixate the words by alternating between the initial PRL and the examiner's selected
TRL. The PRL intends to fulfill one of both functions, i.e.
letter discrimination or global viewing, and the TRL aims
at fulfilling the remaining function. When applicable,
subjects are trained to use the initial PRL for fine spatial
discrimination and the examiner's selected TRL for global
viewing, i.e. to estimate word length and to reach the end
of the word. The anchored cross is eventually removed
and the subject is asked to repeatedly attempt to scrutinize
the word alternating by himself between these fixation
locations. When the subject cannot voluntarily shift fixation onto the examiner's selected TRL, the training procedure is repeated from the previous stage using the fixation
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BMC Ophthalmology 2006, 6:35
Figure 2 for SLO training
Procedure
Procedure for SLO training. First, fixation was anchored
using a cross at the initial preferred retinal locus (PRL) and
words were projected onto the location of the examiner's
selected trained retinal locus (TRL). Projected words and
crosses were randomly presented at two different locations
on the SLO screen to induce changes in eye position. The
anchored cross was eventually removed and the subject was
asked to repeatedly attempt to scrutinize the word alternating by himself between fixation locations.
cross. Verbal feedback by the examiner assists in learning
the use of the TRL.
In each training session the subject is asked to read a 32
single letters and isolated words' list, displaying a wordlength distribution similar to that used in the pre-training
evaluation, but including only four different character
sizes. Character sizes are selected according to the threshold determined during the baseline testing, starting with
two initially readable and two initially unreadable character sizes. Once the subject can read all words, the set of
sizes is readjusted by removing the largest character size
and including a smaller one. Our training process
involves presenting the fixation cross and the associated
word randomly at two different positions on the SLO
screen to avoid subject's anticipation of the spatial location of the stimulus. Ten training sessions of one hour
each are conducted at the same time of the day, with three
to four sessions a week. We conduct the sessions at the
same time each day to avoid variations of performances
due to the circadian cycle. At the end of the SLO training,
subjects are encouraged to apply the trained strategy at
home. For this purpose, they are advised to use the reading device, which was prescribed by the low vision specialist during their previous rehabilitation training.
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Step III: post-training evaluations
To assess for possible functional changes, an evaluation is
conducted both immediately after SLO training and three
months later, and findings are compared to those collected before starting the training procedure. Our rationale for performing these two post-training evaluations is
based on results obtained in normal subjects while learning to read using their eccentric visual field [27]. Those
subjects were able to establish and develop an oculomotor strategy within two weeks but needed afterwards a
period of at least one month to reach a plateau in performances [27] Assessment included ETDRS visual acuity
and threshold character size for single letters and isolated
words, reading accuracy for paragraphed text, as well as
analysis of the oculomotor behavior and reading strategies. We finally ask the subjects to give his subjective
impressions on the possible improvement linked to the
training procedure.
Results
To test for the feasibility of our training procedure, we
conducted a pilot study involving 5 subjects.
Subjects
Criteria used for inclusion of subject were the following:
(1) presence of a bilateral AMD condition stable over the
last year (table 1); (2) visual acuity ranging between 1.5
logMAR and 0.9 logMAR in the best eye; and (3) complaints of reading disturbances consisting essentially of
gross difficulty to decipher long words. Six subjects were
initially enrolled in the study (MT, SL, LG, AA, GM, ML).
Subject ML, however, was subsequently discarded, as he
showed severe signs of memory impairment three months
following the training. His statement that he did not
remember to have been trained in our laboratory emphatically demonstrated his memory loss. Ages ranged from
76 to 81 years (mean: 78.8 years ± 1.92 SD). In each subject a stable bilateral macular condition had been present
in average for 2.3 years. Contrast sensitivity of subjects'
best eye, as measured with the Pelli-Robson contrast sensitivity chart (Clement Clarke Intl, Haag-Streit, UK) was
lower than the one measured in subjects without macular
disorder of similar age [32]. These clinical features are
shown in table 1. A regular low-vision rehabilitation procedure, including scanning and reading exercises similar
to those described by Beaunoir et al. (2000) [33], had
been conducted by a low vision specialist, for four to ten
training sessions, extending over a period of more than
three months. Despite rehabilitation, subjects presented
reading difficulties during the SLO evaluation as for example reading only the first five letters of a ten letter-word, or
guessing, often incorrectly the end of presented words. In
general they were unable to uncover the end of words
(table 3). Stability of the clinical condition and previous
rehabilitation training allowed the use of each subject as
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The
lateddevelopment
Figure
words
3 – before
of the
andcombined
immediately
useafter
of initial
training
PRL, examiner's selected TRL, and two new self-selected PRLs for reading isoThe development of the combined use of initial PRL, examiner's selected TRL, and two new self-selected PRLs
for reading isolated words – before and immediately after training. The figure shows, as seen on SLO images, two
uninterrupted sequences of eye movements performed while attempting to decipher an isolated word presented in 1.3 logMAR. Recordings were obtained in subject LG (A) before SLO training and (B) immediately after SLO training. In each reading
sequence images are numbered in chronological order. Previous to SLO training, the inability to perform eye movements to
reach the end of the word "conscience" prevented reading. The subject therefore only read "con". Immediately following SLO
training, LG adapted his reading strategies and was able to read entirely the word "révolte". The arrows indicate the position of
the retinal areas being presumably employed: initial PRL (
(
), examiner's selected TRL (
) and newly self-selected PRLs
). The examiner's selected TRL was consistently used combined with the initial PRL and/or with newly self-selected PRLs.
Please note that, on the SLO image, an upside down inversion of the image allows the word and the presented reading strategies to be viewed normally.
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The
lateddevelopment
Figure
words
4 – three
of months
the combined
after training
use of initial PRL, examiner's selected TRL, and two new self-selected PRLs for reading isoThe development of the combined use of initial PRL, examiner's selected TRL, and two new self-selected PRLs
for reading isolated words – three months after training. This figure shows an uninterrupted sequence of eye movements performed while attempting to decipher an isolated word presented in 1.3 logMAR, like in figure 3. Recordings were
obtained in subject LG three months following the training procedure. Adaptation due to the training procedure was maintained allowing LG to read entirely the word "compagnie". The arrows indicate the position of the retinal areas being presumably employed: initial PRL (
) and newly self-selected PRLs (
). The initial PRL was consistently used with newly self-
selected PRLs. Please note that, on the SLO image, an upside down inversion of the image allows the word and the presented
reading strategies to be viewed normally.
his own control, as no spontaneous recovery was
expected.
All subjects gave their written informed consent for the
procedure and testing methods, all of which were in
accordance with the Declaration of Helsinki and the study
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The
Figure
development,
5
with SLO training, of a progressive uncovering strategy for reading isolated words
The development, with SLO training, of a progressive uncovering strategy for reading isolated words. The figure
shows three uninterrupted sequences of eye movements while reading an isolated word presented in 1.5 logMAR as seen on
SLO images. Recordings were obtained in subject MT (A) before SLO training, (B) immediately after SLO training and (C) three
months later. In each reading sequence images are numbered in chronological order. Before training, the subject was unable to
read entirely the word "maison". He read only "mais". Immediately following SLO training, he became able to read entirely the
word "horloge", performing successive eye movements uncovering segment by segment the entire word with the same retinal
area (
). Three months following completion of the training process, however, the subject could not anymore read
entirely the word "liaison". The vertical arrow indicates the retinal area presumably being employed, here exclusively the initial
PRL (
). MT was, in the study, the only subject of the experiment to lose both SLO training gains, and modification of the
eye movement pattern. Note that, on the SLO image, an upside down inversion of positions allows the word and the presented reading strategies to be viewed normally.
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Figure
The
text
development,
– before
6
and immediately
with SLO training,
after training
of combined use of the initial PRL and examiner's selected TRL for reading paragraphed
The development, with SLO training, of combined use of the initial PRL and examiner's selected TRL for reading paragraphed text – before and immediately after training. This figure shows, as seen on SLO images, two uninterrupted sequences of eye movements while reading lines of paragraphed text presented in 1.4 logMAR. Recordings were
obtained for subject LG (A) before SLO training and (B) immediately after SLO training. Vertical arrows indicate the retinal
area presumably being employed for fixation. The line being read is shown darker than the other lines. Note that previous to
SLO training, the subject exhibited restricted eye movements. Immediately following SLO training, the subject used consistently the examiner's selected TRL (
) in combination with the initial PRL (
) and/or with a newly self-selected PRL (
).
Note that on the SLO image, the text begins at the lower left side of the figure and ends in the upper right corner. An upside
down inversion of positions allows the text and the reading strategies to be viewed normally.
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Figure
The
text
development,
– three
7 months
with
after
SLO
training
training, of combined use of the initial PRL and examiner's selected TRL for reading paragraphed
The development, with SLO training, of combined use of the initial PRL and examiner's selected TRL for reading paragraphed text – three months after training. This figure shows, an uninterrupted sequence of eye movements
while reading lines of paragraphed text presented in 1.4 logMAR, like figure 6. Recordings were obtained for subject LG three
months after training. Vertical arrows indicate the retinal area presumably being employed for fixation. Three months after
training, the subject used consistently the examiner's selected TRL (
a newly self-selected PRL (
) in combination with the initial PRL (
) and/or with
).
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had been accepted by the ethical committee for human
experimentation of Geneva University Hospitals.
Reading strategies for isolated words and paragraphed
text
Initial evaluation
The results of the initial evaluation step showed that for
reading isolated words and paragraphed text four (MT, SL,
LG, AA) out of five subjects had spontaneously developed
an initial PRL that fulfilled the needs for detailed discrimination capacity although it prevented global viewing.
Indeed, the PRL showed better visual acuity than the
examiner's selected TRL, but was not optimally located to
allow the viewing of the whole word, thus altering reading
ability (figure 3A, 4, 5A).
Three subjects (MT, SL, LG) had an initial PRL to the left
of the scotoma and consequently ignored the end of the
words (figure 3A, 5A and 6A). Two of these subjects (SL,
LG) were additionally unable to perform search movements to detect single letters and isolated words initially
projected within the scotoma, and to relocate the projected image onto a healthy peripheral area. The third subject (MT) showed an additional small scotoma in the
vicinity of the left border of the central scotoma on the
SLO picture that repeatedly superimposed on parts of the
scrutinized characters (figure 1, 5). We trained these three
subjects (MT, SL, LG) to use an examiner's selected TRL
above the scotoma on the SLO image (i.e. below the
scotoma in the visual field).
The fourth subject (AA) had an initial PRL located above
the scotoma on the SLO image, but too close to the
scotoma's border, the upper part of the letters scrutinized
being often projected into the scotoma. Consequently he
often missed the center of presented words. We trained
him to use an examiner's selected TRL located on the same
side of the scotoma as the initial PRL, i.e. above the lesion
on the SLO image, but farther from the scotoma's border
(figure 1).
The remaining fifth subject (GM) had not developed a
PRL despite previous rehabilitation. He was found to use
an ill-defined area with preserved global viewing but nonoptimal spatial resolution. In addition, he presented a
highly instable fixation.
Based on the results of visumetry, he was trained to use an
area below the scotoma on the SLO picture, which presented a better acuity than the initial PRL while still preserving global viewing capacity (figure 1).
Post-training evaluations
Immediately after SLO training, analyses of reading strategies revealed that four subjects (SL, LG, AA, GM) used the
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examiner's selected TRL, or additional newly self-selected
PRLs (figure 3B and 6B) alone or in combination with
their initial PRL for reading isolated words and paragraphed text. One of these subjects (GM) (no defined PRL
before SLO training) used the examiner's selected TRL
alone. The remaining fifth subject (MT) showed an
improved ability to scan the word but only used the initial
PRL (figure 5B).
Three months following SLO training, four subjects (SL,
LG, AA, GM) consistently used the examiner's selected
TRL either in isolation (GM), or in combination with the
initial PRL (LG, AA, SL) for reading both isolated words
and paragraphed texts (table 3, figures 4 and 7). The use
of newly developed self-selected PRLs was observed in
three subjects (SL, AA, GM) to read isolated words and in
two subjects to read paragraphed text (MT, SL). Interestingly, one subject (MT) lost the ability to perform eye
movements to scan the entire words, but kept some adaptation, i.e. used newly self-selected PRLs, during text reading (table 2).
ETDRS visual acuity
ETDRS values are reported in table 2 and figure 8. Visual
acuity evolution across the three evaluation periods was
analyzed using a one-way repeated measure ANOVA. This
analysis yielded a significant difference (F(2,3) = 17.85; p
= 0.022). Values measured before and three months following the SLO training significantly improved compared
to those measured before the training process (Bonferroni
pairwise comparison p = 0.012), while the difference was
marginally significant between acuities measured before
and immediately after SLO training (Bonferroni pairwise
comparison p = 0.085). However when considering individual data, the majority of improvements were no greater
than the test repeatability of 0.2 [34,35].
Threshold character size for single letters and isolated
words
Threshold character sizes for single letters and isolated
words are shown in table 2 and figure 9. A three-way
repeated measure ANOVA with subjects (MT, SL, LG, AA,
GM), stimulus types (single letters, 3 and 4 letter-words, 6
and 7 letter-words and 9 and 10 letter-words) and evaluation periods (before, at the end of training and three
months later) was conducted to explore the impact of the
training procedure as measured by the threshold character
size.
There was a highly significant main effect of the evaluation periods (F(2, 19) = 46.04; p < 0.0005). The post-hoc
analysis using Bonferoni correction showed that all subjects benefited from the training procedure both, immediately after SLO training (mean ± standard error: before =
1.255 ± 0.017, immediately after SLO training = 1.055 ±
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0.016; p < 0.0005) and three months later (1.084 ± 0.014;
p < 0.0005). There was, however, no significant evolution
after completion of the training procedure (p = 0.563).
There was also a main effect of stimulus type confirming
that threshold character size varied as a function of word
length (F(3, 20) = 73.921; p < 0.0005). Threshold character size for single letters (0.9 ± 0.019) was lower than for
any other word lengths and threshold for 3 and 4 letterwords (1.13 ± 0.019) was lower than those for 6 and 7
(1.23 ± 0.019) and 9 and 10 letter-words (1.26 ± 0.019;
Newman-Keuls: p < 0.05). Additionally, the retention of
the training gains varied with stimulus types (interaction
evaluation periods and stimulus type: F(6, 38) = 3.135; p
= 0.014). To further investigate the retention of the training gains on the different stimulus types we conducted a
post-hoc analysis using Bonferroni adjustment for multiple comparisons with α = 0.00125. Subjects read significantly smaller character sizes immediately after the end of
the SLO training procedure at all word lengths but 3/4 letter-words, despite the fact that a slight improvement was
noticeable for the latter (1 letter: before = 1.02 ± 0.029,
immediately after SLO training = 0.82 ± 0.022; p = 0.006;
6 and 7-letter words: before = 1.41 ± 0.062, immediately
after SLO training = 1.15 ± 0.027; p = 0.008; 9 and 10-letter words: before = 1.38 ± 0.057, immediately after SLO
training = 1.13 ± 0.045; p = 0.006; 3 and 4-letter words:
before = 1.21 ± 0.062, immediately after SLO training =
1.10 ± 0.045; p = 0.513). Three months following the
training procedure, gains were retained for single letters
Figure 8 of ETDRS visual acuity
Evolution
Evolution of ETDRS visual acuity. Measures are
reported for each subject. before: before SLO training; after:
immediately after SLO training; three months later: three
months following SLO training. Changes in ETDRS values
could be considered significant when greater than 0.2 logMAR [38, 39]. Note that ETDRS visual acuity consistently
increased with the SLO training and, globally, gains were
retained three months later.
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and 6 and 7-letter words, as shown by the non-significant
difference between results obtained at the end of training
and three months later (1 letter: three months later = 0.86
± 0.016; p = 1.0; 6 and 7 letter-words: three months later
= 1.13 ± 0.056; p = 1.0). In contrast, performances were
not retained for 9 and 10-letter words (three months later
= 1.28 ± 0.057; p = 0.02) and returned to the initial score
(p = 0.32). For 3 and 4-letter words the statistical test did
not reveal any additional change over the whole procedure either (immediately after SLO training – three
months later: p = 1.0: before – three months later: p =
0.456).
Another main result was that subjects benefited differently from the training procedure, as shown by the interaction between evaluation periods and subjects (F(8, 38)
= 5.371; p < 0.0005). Immediately after the SLO training
procedure, all subjects had improved their performances
regardless of the type of stimulus. Three months later subjects SL, LG and AA totally retained and even slightly
improved, training gains. Subject GM did not fully preserve the improvements, even if his performances were far
better than before the SLO training. Subject MT did not
retain training gains at all (figure 9). Despite the fact that
the interaction subjects, stimulus type and evaluation
periods was not significant (F(24; 38) = 0.1014; p =
0.497), it is interesting to describe individual behavior.
When looking at individual data (figure 9), it appears that
all subjects had better performances three months following the training procedure than before for 3 and 4-letter
words. For 6 and 7-letter words, three subjects (SL, LG,
AA) progressed in deciphering capacities, one subject
(GM) retained training benefits, and the remaining subject (MT) partially lost training gains. For the 9 and 10-letter words, one subject (SL) improved, three subjects (LG,
AA, GM) partially lost the gains, and the remaining subject (MT) lost all benefits of the SLO training (see arrows
in figure 9). Finally there was a main effect of subjects, as
expected regarding the individual variations of threshold
character sizes (F(4, 20) = 16.443; p < 0.0005). The interaction subject and stimulus type was not significant (F(12;
20) = 1.100; p = 0.087).
Paragraphed text reading
Threshold character size for paragraphed text and percentage of words read are shown in table 2 and figure 10.
Immediately following SLO training, three subjects (SL,
AA, GM) became able to read paragraphed text one character size smaller (0.1 logMAR) and one subject (LG)
could read two character sizes smaller (0.2 logMAR) than
evaluated before training (table 2). In all subjects, the proportion of words correctly read at the first attempt, i.e.
reading accuracy, increased with training (figure 9). Three
months following the completion of the SLO training
procedure, the smallest readable character size for para-
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Figure 9 of threshold character size for single letters and isolated words
Evolution
Evolution of threshold character size for single letters and isolated words. Threshold character sizes' values in logMAR, are provided for each subject. before: before SLO training; after: immediately after SLO training; three months later: three
months following SLO training. Isolated letters and word length categories are considered independently. With subject GM,
threshold character size for 6 and 7 and 9 and 10 letter words could not be determined before training, because he was unable
to decipher the largest presented stimuli (1.5 logMAR). Therefore, in the graph, he was assigned the minimum 1.6 logMAR
level. Note the invariance of the results for isolated letters, contrasting with the variability of the changes obtained for isolated
words. Loss of training gains mainly occurred when deciphering longer words. In this regard, measures in subject LG are particularly illustrative, showing continuous improvement in reading single letters and 3/4-letter words, stabilized performance for
6 and 7-letters words, results which contrast with the gain loss observed for 9 and 10-letter words.
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graphed text was similar to that of the initial evaluation in
four subjects (MT, SL, AA, GM), but the percentage of
words correctly read had increased (figure 9) allowing
subjects to have an adequate comprehension on smaller
character sizes considering that an accuracy of 85% allows
a complete understanding of the text (27).
Patients' comments on the modification of their visual
perception before and after the training
We asked the patients to give a subjective evaluation of the
influence of the SLO training on their everyday life. All
patients reported less visual fatigue and improved awareness of the location of the scotoma. MT reported that two
weeks after the completion of the SLO training, she had
the impression of an improvement in reading. SL noticed
that SLO training enabled a more efficient reading of the
panels in the street as well as bus numbers. LG could not
report specific subjective improvements but said that, in
general, she better coped with her visual field defect. AA
applied the taught strategy to other visual tasks. She said
she recovered the ability to hand-knit. After the SLO training process, GM started to use a closed circuit television
(CCTV) and was able to read with it. He stated that the
SLO training allowed him to locate his scotoma in his visual field and, as a result, several visual tasks were easier to
accomplish.
Discussion
SLO training in subjects with macular conditions had
been emphasized previously [21]. This technique allows
local visual acuity measurements, estimation of potential
global viewing capacity in different areas of the retina and
direct understanding of the nature of reading problems
[21,36]. It also enables the guidance of subject's fixation
onto the presented words with a continuous interaction
between the subject and the examiner during the training
session. The particularity of our training procedure consisted of an attempt to structure new oculomotor behavior
involving the use of additional fixation points, without
preventing the patients to use his initially spontaneously
developed PRL. We also established the schedule of our
training process based on recent findings in neurorehabilitation showing that intensive and short-term training was
beneficial [reviewed in 25], and applied, for example, in
individuals with chronic aphasia [24]. Thus, as retaining
new oculomotor behaviors require brain plasticity and as
the occurrence of such phenomenon is comparable with
those occurring in patients suffering from other neural
damages, we schedule the SLO training accordingly.
Based on our first results, we can already discuss possible
events underlying the adaptation to a second fixation
area.
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Immediate functional benefits and gains retention
Our analyses of reading strategies demonstrated essentially that our SLO training allowed subjects with limited
global viewing to acquire the capacity to perform eye
movements, which allow the perception of the presented
words in their whole. The newly adapted oculomotor
behavior included the use of the examiner's selected TRL,
either in isolation or in combination with the initial spontaneously developed PRL and the associated use of newly
self-selected PRLs. Three subjects used the examiner's
selected TRL in combination with other areas, whereas the
remaining subject, who had no defined initial PRL, used
only the examiner's selected. Interestingly, the development of the newly self-selected PRLs and new oculomotor
patterns suggest that the structured training procedure,
aiming primarily at developing an examiner's selected
TRL, in fact is likely to initiate a more global functional
adaptation process. As already suggested in neuro-rehabilitation studies, training might trigger general changes in
adaptation strategies and thus may "prime" the brain for
future learning [26].
In our subjects, the adaptation of reading strategies was
associated with improvements measured in different reading related tasks. The only subject (MT) who showed no
adaptation of oculomotor behavior also did not improve
performances in reading related tasks. ETDRS acuity and
threshold character size for single letter measurements
evolved, as expected, similarly throughout the training.
Increased visual acuity was likely related to an increased
ability to detect and then relocate stimuli that fell into
scotoma's area. One might also consider whether gains in
ETDRS acuity and threshold character size for single letters and isolated words observed following SLO training
were non-specific results of the training process, and unrelated to the use of the SLO. This hypothesis however, is
unlikely, as all subjects had previously undergone a conventional low vision training procedure, which proved to
be unsuccessful in allowing isolated words to be deciphered.
Assessment conducted three months following the end of
the SLO training showed that gains persisted and that
improvements remained for reading single letters but
were not retained for 9 and 10-letter words. Deciphering
9 and 10-letter words requires both accurate fixation and
adapted saccadic movements, probably more so than
those required for isolated letters and short words, which
necessitate essentially accurate eccentric fixation. Differences between various reading tasks in terms of gain retention presumably reflect the variation in complexity of
mechanisms involved in specific adaptation processes.
Indeed, it is conceivable that complex tasks more
demanding in terms of plasticity may be more difficult to
retain and/or need more extensive training. A similar var-
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Figure 10of paragraphed text reading performance over the training process
Evolution
Evolution of paragraphed text reading performance over the training process. This figure shows the percentage of
words correctly read at the first attempt in paragraphed text before, immediately after, and three months following SLO training. Percentage is plotted for each character size – from the largest size presented (1.5 logMAR, upper graph) to the smallest
readable character size (1.1 logMAR, lower graph). The percentage was assigned to zero when, after attempting to read, the
subject felt unable to decipher a single word. Dashed are used when a value was assigned to zero at one of the training phases.
It appears that, at the end of the training procedure, all subjects were able to read correctly about 82% or more words at first
attempt at the largest character size presented. Note that, previous to SLO training, two of these subjects had demonstrated a
very low percentage at that character size. The three subjects (SL, LG, AA) who, already before training, exhibited a high percentage of words correctly read, improved their ability to read smaller character sizes immediately following SLO training.
Finally, it should be emphasized that gains observed at the end of training period were often not retained three months later.
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iability in the retention of performances as a function of
the complexity of the task was observed in 400 patients
who had followed short-term, intensive procedures for
rehabilitation on motor tasks [37].
Eye movements' adaptation
The adaptation of the oculomotor system studied here
involved at least two different mechanisms: the first one
to achieve eccentric fixation, and the second to reorganize
the control of saccades around the coordinates of the initial PRL and the examiner's selected TRL. The distinction
between these mechanisms and their respective complexity has been demonstrated experimentally. Heinen and
Skavenski (1992) [38] found in adult monkeys that one
day after bilateral foveal lesions, fixation was performed
with a new retinal locus, although saccades at this stage of
adaptation still maladaptively brought visual targets onto
the lesioned fovea. Subsequently, over a period of several
weeks, saccades trajectories progressively changed and targets projected directly onto the intact retina [38]. In our
study, individual practice of oculomotor strategies during
the period following the SLO training might also have
influenced the degree of retention of reading ability. Particularly, in one subject (MT) who could not pursue any
reading practice at home, the gains observed immediately
after SLO training were eventually reduced, and even
return to baseline performances. We therefore propose
that to retain improvements on complex tasks "brush up"
periods of training might be advised following structured
procedure.
It is of interest to investigate to what extent the improvements obtained following training for isolated words
might reflect subjects' increased ability to read paragraphed texts in everyday' life conditions. We previously
reported in subjects with macular scotoma, a strong correlation between the threshold character size for reading
words longer than five letters and for reading paragraphed
text in the SLO [39]. Consequently, following reading
training, improvements on longer words are likely to be
associated with an improvement on paragraphed text,
although the latter requires a more complex adaptation of
eye movements – to perform accurate return sweeps, for
example. In this pilot training study we showed that
patients did not acquired the capacity to read paragraphed
text of smaller character sizes but improved the number of
words correctly read at first attempt and thus reached levels of complete text comprehension. We limited our analysis of paragraphed text reading to an evaluation of
reading threshold character size and the determination of
the percentage of words correctly read at first attempt
because the SLO is not adapted to determine reading
speed as a important number of images are lost while the
subject performs eye movements.
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Conclusion
In subjects with macular disorders, we previously suggested that learning to control eye movements improved
the use of the PRL and facilitates the development of additional PRLs [20]. Here we proposed a procedure allowing
the training of the use of multiple PRLs in reading related
tasks, which seemed to favor the development of additional individual reading strategies involving the examiner's selected TRLs and newly self-selected PRLs to bypass
problems linked to the presence of the central macular
lesion. SLO training aiming at combining the use of multiple PRLs is feasible and likely to improve reading ability.
The SLO has unfortunately been out of production for
many years and for this reason is presently not a widespread instrument. However, our study suggested that
similar and simpler devices allowing real time view of the
retina and projection of chosen stimuli might be introduced into the market for the purpose of reading training,
particularly as the incidence of AMD is expected to
improve in the following years. The first results with such
a training that we obtained on five patients are encouraging but to reach definite results the training of a larger
group of subjects and a comparison of data with those of
a control group would be more compelling.
Competing interests
The author(s) declare that they have no competing interests.
Authors' contributions
AD participated in the design of the study, the acquisition,
analysis and interpretation of the data, and the drafting of
the final manuscript. MG participated in the design of the
study, the interpretation of the data and the drafting if the
final manuscript. ARW participated in the design of the
study and the drafting of the manuscript. CM designed the
computer software necessary to all analyses of the data.
ENL performed the statistical analysis and revised the
manuscript. AS participated in the design of the study and
in the drafting of the final manuscript. ABS participated in
the design of the study, in the interpretation of data and
in drafting the final manuscript. All authors read and
approved the final manuscript.
Acknowledgements
The authors thank the therapists from the Association pour le Bien des
Aveugles and Mrs Claire Ligny for their help in subjects' recruitment. This
investigation was supported by the Pro Visu Foundation and the Fondation
en Faveur des Aveugles, Geneva, Switzerland.
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Autodisplay for the co-expression of lipase and foldase on the surface of E. coli: washing with designer bugs
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Abstract Background: Lipases including the lipase from Burkholderia cepacia are in a main focus in biotechnology research
since many years because of their manifold possibilities for application in industrial processes. The application of
Burkholderia cepacia lipase for these processes appears complicated because of the need for support by a
chaperone, the lipase specific foldase. Purification and reconstitution protocols therefore interfere with an economic
implementation of such enzymes in industry. Autodisplay is a convenient method to express a variety of passenger
proteins on the surface of E. coli. This method makes subsequent purification steps to obtain the protein of interest
unnecessary. If enzymes are used as passengers, the corresponding cells can simply be applied as whole cell
biocatalysts. Furthermore, enzymes surface displayed in this manner often acquire stabilization by anchoring within
the outer membrane of E. coli. Results: The lipase and its chaperone foldase from B. cepacia were co-expressed on the surface of E. coli via autodisplay. The whole cell biocatalyst obtained thereby exhibited an enzymatic activity of 2.73 mU mL-1 towards the substrate
p-nitrophenyl palmitate when applied in an OD578 =1. Outer membrane fractions prepared from the same culture
volume showed a lipase activity of 4.01 mU mL-1. The lipase-whole cell biocatalyst as well as outer membrane
preparations thereof were used in a standardized laundry test, usually adopted to determine the power of washing
agents. In this test, the lipase whole cell biocatalyst and the membrane preparation derived thereof exhibited the same
lipolytic activity as the purified lipase from B. cepacia and a lipase preparation which is already applied in commercial
washing agents. Conclusions: Co-expression of both the lipase and its chaperone foldase on the surface of E. coli yields a lipid
degrading whole cell biocatalyst. Therefore the chaperone supported folding process, absolutely required for the
lipolytic activity appears not to be hindered by surface display. Furthermore, the cells and the membrane preparations
appeared to be stable enough to endure a European standard laundry test and show efficient fat removal properties
herein. high enantio- and/or regio-selectivity as well as a high
catalytic activity and thermostability in organic solvents
[2,3]. Contrary to esterases, which preferentially break
ester bonds of short chain fatty acids, lipases are able to
catalyze the hydrolysis of water-insoluble long-chain acyl-
glycerols [1]. Interestingly, activation of lipases often de-
pends on the presence of a lipid-water interface, which
can be explained by their three-dimensional structure. © 2014 Kranen et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative
Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication
waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise
stated. * Correspondence: joachim.jose@uni-muenster.de
1Institute of Pharmaceutical and Medicinal Chemistry, PharmaCampus,
Westfalian Wilhelms-University Münster, Corrensstr. 48, 48149 Münster,
Germany
Full list of author information is available at the end of the article Abstract In
an enzymatically inactive state, a surface loop, the so-
called lid, covers the active site of the lipase. Upon con-
tacting the lipid-water interface the lid switches open, and
the active site becomes accessible for the substrate [4]. Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19 Autodisplay for the co-expression of lipase and
foldase on the surface of E. coli: washing with
designer bugs Eva Kranen2, Christian Detzel2, Thomas Weber3 and Joachim Jose1* Background Lipolytic enzymes are attractive biotechnological tools
[1]. Among them lipases (triacylglycerol acylhydrolases
EC 3.1.1.3), which catalyze the hydrolysis of triglycerides
in aqueous media, liberating free fatty acids and glycerol,
or the reverse reaction in organic solvents as well, have
gained particular interest, since they simultaneously show Page 2 of 12 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 So far, lipases have been established in numerous in-
dustries, such as the food industry, paper manufacturing,
pharmaceutical processing [5], and detergents industry,
reflecting their great importance [4]. Despite this enor-
mous industrial interest, not more than around 20 lipases
have been established for industrial applications yet [6]. The sometimes troublesome and time-consuming purifi-
cation procedures to obtain pure enzyme preparations for
particular applications seem to be one possible obstacle in
broadening the use of lipases in industrial processes [7]. Moreover, to express lipases from Burkholderia and
Pseudomonas species in an active form, lipases which have
advantageous features regarding thermal stability, alkaline
pH tolerance and high substrate selectivity, and there-
fore making them promising industrial biocatalysts
[8-10], bears an additional problem. These enzymes
are dependent on the presence of a personal chaperon,
the so-called lipase-specific foldase (Lif), responsible
for correct folding of the lipase [1,11]. As a consequence,
former heterologous expression of the Burkholderia cepacia
lipase in E. coli resulted in a very low yield of active soluble
lipase, whereas the majority of the enzyme was expressed as
insoluble inclusion bodies. Significant amounts of active lip-
ase were only achieved by applying an additional in-vitro
refolding protocol [12]. encodes a fusion protein comprised of an N-terminal
signal peptide derived from cholera toxin β-subunit
(CtxB), a variable passenger domain and the C-terminal
AIDA-I autotransporter including a linker to enable full
surface access of the passenger domain (Figure 1B). Most
probably, the linker and the β-barrel are responsible
for the translocation of the passenger protein across
the E. coli outer membrane (Figure 1A). One of the
most striking features of the Autodisplay system is the mo-
bility of the β-barrel serving as an anchor within the outer
membrane. This enables the self-driven dimerization or
multimerization of subunits to active or functional en-
zymes on the surface of E. coli, even in case they were
expressed as monomers. Examples for this self-driven
dimerization or multimerization of passsenger proteins on
the cell surface of E. Background coli are the active display of dimeric
adrenodoxin [23], dimeric sorbit dehydrogenase [24], mul-
timeric nitrilase [25] and dimeric prenyl transferase [26]. Moreover, Autodisplay has proven to be a robust expres-
sion platform for the surface display of enzymes in general Figure 1 Passenger transport across two membranes by
Autodisplay. A: The N-terminal signalpeptide facilitates the transport
across the inner membrane by the so called Sec-pathway [32] and is
then cut off by periplasmic signal peptidases. The C-terminal part forms
a porin-like β-barrel structure inside the outer membrane through
which the passenger is translocated to the surface by the linker. B and
C: The structure of the precursor proteins for LipBC-FP (B) and
FoldBc-FP (C) is shown schematically. The mature fusion proteins
anchored inside the outer membrane only consist of the passenger
(which would be here lipase or foldase, depicted in lightgrey) and the
autotransporter structure (linker (white) and β-barrel (black)). (The
genes for lipase and foldase were amplified from plasmid pHES8,
containing the complete sequence of the B. cepacia lipase [GenBank:
FJ638612] and cloned into appropriate Autodisplay-vectors via
restriction sites XhoI (5′-end) and KpnI (3′-end). An innovative way to gain access to the synthetic po-
tential of lipases is their display on the surface of a living
cell, in particular an E. coli cell [13]. Since the enzyme is
directly accessible for its substrate, costly purifications as
mentioned above are not necessary. So far, various anchoring motifs like OmpC [14], ice
nucleation protein [15], OprF [16] and FadL [17] have
been used to display Pseudomonas and Bacillus lipases
on the surface of E. coli. Wilhelm et al. [18] were able to
display the LipH chaperone of P. aeruginosa in an active
state on the surface of E. coli by using the P. aeruginosa
autotransporter protein EstA. With these cells displaying
the lipase specific foldase, reconstitution of a purified
but denatured lipase into an active form was facilitated. In another report, Yang et al. described the display of ac-
tive P. aeruginosa and B. cepacia lipases on the surface
of E. coli via co-expression of lipase and the Lif protein
within a single fusion protein [19]. Autodisplay, a bacter-
ial surface display system, appeared to be a convenient
tool for the expression of B. cepacia lipase, since it has
been proven to be well adapted for the surface display of
challenging enzymes. Background coli BL21
(DE3)pAT-LipBc without addition of IPTG were culti-
vated and outer membranes were prepared and analyzed
identically (Figure 2A, lanes 1 and 2). Inducing the pro-
tein expression of E. coli BL21(DE3) pAT-LipBc resulted
in expression of the lipase fusion protein with a size
of ~82 kDa (Figure 2A, lane 3). A lipase specific anti-
body was available, so the correct surface exposure of
lipase could be evaluated by fluorescence-activated
cell sorting (FACS). Since antibodies are too large to
cross the outer membrane, they can only bind on sur-
face exposed structures [36]. Therefore, cells express-
ing a passenger protein on their surface which is then
marked by fluorescently labeled antibodies can easily
be detected by FACS and will thereby cause an increase in
fluorescence values compared to cells without such sur-
face displayed protein. To identify effects caused by un-
specific binding, the native host strain E. coli BL21(DE3)
and another autodisplay strain displaying a different en-
zyme (NADH oxidase) on its surface (E. coli BL21(DE3)
pAT-NOx) were used as controls. It turned out that the
sample containing the lipase expressing cells showed a
tenfold increase in mean fluorescence intensity values
(Figure 3C) compared to the samples used as controls
which showed no increased fluorescence signal (Figure 3A
and B). The lipase antibody thus effectively bound the
enzyme but did not show unspecific binding effects. Therefore the lipase expressed via autodisplay can be
regarded as surface exposed. Interestingly, like Yang
et al. [19] were already able to demonstrate, antibody la-
beling of the surface exposed lipase does not require the
involvement of its chaperone foldase. including cytochrome P450 enzymes of bacterial and hu-
man origin [27-29]. More recently, it was shown that
Autodisplay, which is defined as the surface display of a
recombinant protein by the autotransporter secretion
pathway [30], relies on a set of periplasmic chaperones in-
cluding a complex of proteins which corresponds to the
so-called Bam machinery in E. coli [31]. This makes the
prefix “auto” somewhat obsolete, but for clarity reasons it
appears to be favorable not to change the term Autodis-
play on these findings. Background As an example it was possible to
express enzymatically active human hyaluronidases in
E. coli, a group of enzymes which are known to form
inclusion bodies, when expressed by other means [20]. Autodisplay is based on AIDA-I, the adhesin involved
in diffuse adherence in enteropathogenic E. coli (EPEC)
[21,22], a naturally occurring autotransporter protein in
E. coli. The gene construct applied in Autodisplay Figure 1 Passenger transport across two membranes by Figure 1 Passenger transport across two membranes by Figure 1 Passenger transport across two membranes by
Autodisplay. A: The N-terminal signalpeptide facilitates the tra g
g
p
y
Autodisplay. A: The N-terminal signalpeptide facilitates the transport
across the inner membrane by the so called Sec-pathway [32] and is
then cut off by periplasmic signal peptidases. The C-terminal part forms
a porin-like β-barrel structure inside the outer membrane through
which the passenger is translocated to the surface by the linker. B and
C: The structure of the precursor proteins for LipBC-FP (B) and
FoldBc-FP (C) is shown schematically. The mature fusion proteins
anchored inside the outer membrane only consist of the passenger
(which would be here lipase or foldase, depicted in lightgrey) and the
autotransporter structure (linker (white) and β-barrel (black)). (The
genes for lipase and foldase were amplified from plasmid pHES8,
containing the complete sequence of the B. cepacia lipase [GenBank:
FJ638612] and cloned into appropriate Autodisplay-vectors via
restriction sites XhoI (5′-end) and KpnI (3′-end). Autodisplay. A: The N-terminal signalpeptide facilitates the transport
across the inner membrane by the so called Sec-pathway [32] and is
then cut off by periplasmic signal peptidases. The C-terminal part forms
a porin-like β-barrel structure inside the outer membrane through
which the passenger is translocated to the surface by the linker B and Page 3 of 12 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 protein was then induced by addition of isopropyl-β-
thiogalactosid (IPTG) to a final concentration of 1 mM
and incubation for one hour. Adjacently cells were har-
vested and the outer membrane proteins were isolated
according to the protocol of Hantke [34], modified by
Schultheiss et al. [35]. The obtained outer membrane
preparations were then subjected to SDS-PAGE to
analyze the expression of the lipase fusion protein. As a
control host cells E. coli BL21(DE3) and E. Background In order to elucidate, whether
Autodisplay is not only capable of permitting subunits of
enzymes to aggregate on the cell surface, but can also be
used for the expression of two different enzymes on a sin-
gle cell, we chose Burkholderia cepacia lipase and its spe-
cific foldase as candidates. Lipolytic activity was tested in
common lab scale assays as well as in a standardized laun-
dry test which is typically used to evaluate the quality of
washing agents. Since the presence of recombinant bac-
teria in clothes after washing could cause some resistance
in application, also membrane preparations of the cells
co-expressing lipase and foldase were applied in the iden-
tical test as well. Results: Construction of the plasmid for autodisplay of lipase
By analyzing the amino acid sequence of B. cepacia
ATCC 21808 lipase using the SignalP computer program
[33], a classical signal peptide was identified at its N
terminus. Since this lipase inherent signal peptide is pro-
posed to interfere with the signal peptide used in auto-
display and thus constrain a proper transport across
the inner membrane, the lipase signal peptide encod-
ing 120 bp sequence was deleted by PCR. PCR-primers
were designed according to the deposited sequence of
the B. cepacia lipase [GenBank: FJ638612] and added
an XhoI (5′end) and a KpnI restriction site (3′end) to
the PCR fragment in order to enable an in frame fu-
sion with the plasmid DNA encoding the autodisplay
domains. For PCR plasmid pHES8 was used, which re-
sembles pHES12 described by Quyen et al. [12] and
encodes the complete B. cepacia lipase operon (i.e. lip-
ase and its corresponding foldase) for intracellular ex-
pression in E. coli. After insertion into plasmid pCD003
[25] cleaved with XhoI and KpnI as well, plasmid pAT-
LipBc was obtained encoding a fusion protein comprising
the signal peptide of CtxB at the N terminus followed
by the lipase as a passenger, the linker region and the
β-barrel from the AIDA-I autotransporter needed for
outer membrane translocation and full surface accessi-
bility (Figure 1B). Construction of the plasmid for autodisplay of foldase
According to Quyen et al. [12] the gene for foldase con-
tains a possible N-terminal 70 aa membrane anchor. This
structure is not required for the chaperone function of fol-
dase, but may interfere with correct surface expression via
autodisplay. Therefore foldase also was amplified from
plasmid pHES8, which encodes the whole lipase operon
[12], deleting the first 210 bp encoding this particular an-
chor structure. PCR primers, designed using the deposited
sequence of the whole B. cepacia lipase [GenBank:
FJ638612] added an XhoI site at the 5′-end and a KpnI
site at the 3′-end of the foldase gene, analogously as
described for the construction of plasmid pAT-LipBc. Surface display of lipase M: protein marker; IPTG: protein expression was induced by
adding IPTG (final concentration: 1 mM); Proteinase K: whole cells were treated with Proteinase K; concentrations are given in mg mL-1. The
lipase and foldase fusion proteins are indicated by using black arrows. OmpA/OmpF: native E. coli outer membrane proteins are also indicated
by a black arrow Surface display of lipase E. coli BL21(DE3) pAT-LipBc were grown until an
OD578 of 0.5 was reached. Expression of the lipase fusion Page 4 of 12 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Figure 2 Expression of lipase fusion protein, expression and surface display of foldase fusion protein. A: SDS-PAGE of the outer membrane
protein preparation of E. coli BL21(DE3)pAT-LipBc. Lane 1 shows an outer membrane preparation of E. coli BL21(DE3), used as a control. Lanes 2
and 3 show outer membrane preparations of E. coli BL21(DE3)pAT-LipBc. B: SDS-PAGE of the outer membrane protein preparation of E. coli BL21
(DE3)pAT-FoldBc. Molecular weight markers are indicated on the left hand side. M: protein marker; IPTG: protein expression was induced by
adding IPTG (final concentration: 1 mM); Proteinase K: whole cells were treated with Proteinase K; concentrations are given in mg mL-1. The
lipase and foldase fusion proteins are indicated by using black arrows. OmpA/OmpF: native E. coli outer membrane proteins are also indicated
by a black arrow. Figure 2 Expression of lipase fusion protein, expression and surface display of foldase fusion protein. A: SDS-PAG Figure 2 Expression of lipase fusion protein, expression and surface display of foldase fusion protein. A: SDS-PAGE of the outer membrane
protein preparation of E. coli BL21(DE3)pAT-LipBc. Lane 1 shows an outer membrane preparation of E. coli BL21(DE3), used as a control. Lanes 2
and 3 show outer membrane preparations of E. coli BL21(DE3)pAT-LipBc. B: SDS-PAGE of the outer membrane protein preparation of E. coli BL21
(DE3)pAT-FoldBc. Molecular weight markers are indicated on the left hand side. M: protein marker; IPTG: protein expression was induced by
adding IPTG (final concentration: 1 mM); Proteinase K: whole cells were treated with Proteinase K; concentrations are given in mg mL-1. The
lipase and foldase fusion proteins are indicated by using black arrows. OmpA/OmpF: native E. coli outer membrane proteins are also indicated
by a black arrow. Figure 2 Expression of lipase fusion protein, expression and surface display of foldase fusion protein. A: SDS-PAGE of the outer membrane
protein preparation of E. coli BL21(DE3)pAT-LipBc. Lane 1 shows an outer membrane preparation of E. coli BL21(DE3), used as a control. Lanes 2
and 3 show outer membrane preparations of E. coli BL21(DE3)pAT-LipBc. B: SDS-PAGE of the outer membrane protein preparation of E. coli BL21
(DE3)pAT-FoldBc. Molecular weight markers are indicated on the left hand side. Surface display of foldase The derived fragment was ligated into autodisplay vector
pBL001, digested with XhoI and KpnI before. Vector
pBL001 is a pCOLA DuetTM derivative, encoding the do-
mains needed for autodisplay. Vector pBL001 furthermore
provides a kanamycin resistance. Insertion of the foldase
gene into pBL001 resulted in plasmid pAT-FoldBc encod-
ing an in frame fusion of the autodisplay domains with fol-
dase as a passenger (Figure 1C). E. coli BL21 (DE3) pAT-FoldBc cells were grown to mid-
log phase and autotransporter fusion protein expression
(FoldBc-FP) was induced by adding 1 mM IPTG to the
fermentation broth and incubating the culture for an-
other hour. After preparation of the outer membrane
fraction, obtained protein samples were subjected to
SDS-PAGE. As can be seen in Figure 2B, induction of Figure 3 FACS analysis of lipase surface display. Whole cells were treated with rabbit-anti-lipase-antibody and anti-rabbit Dylight coupled
secondary antibody. A: host cells E. coli BL21(DE3) used as a control, B: Cells displaying another enzyme on their surface (E. coli BL21(DE3)
pAT-NOx). These were used as control cells to test whether the anti-lipase-antibody binds parts of the autotransporter. C: BL21(DE3)pAT-LipBc
expressing the lipase via Autodisplay. Only the reaction with BL21(DE3)pAT-LipBc and the lipase-antibody caused a tenfold increase in mean
fluorescence values, which means that the lipase can be regarded as surface exposed. Figure 3 FACS analysis of lipase surface display. Whole cells were treated with rabbit-anti-lipase-antibody and anti-rabbit Dylight coupled
secondary antibody. A: host cells E. coli BL21(DE3) used as a control, B: Cells displaying another enzyme on their surface (E. coli BL21(DE3)
pAT-NOx). These were used as control cells to test whether the anti-lipase-antibody binds parts of the autotransporter. C: BL21(DE3)pAT-LipBc
expressing the lipase via Autodisplay. Only the reaction with BL21(DE3)pAT-LipBc and the lipase-antibody caused a tenfold increase in mean
fluorescence values, which means that the lipase can be regarded as surface exposed. Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 5 of 12 protein expression resulted in the appearance of a pro-
tein band with an apparent molecular mass of around
80 kDa (Figure 2B, lane 2), which is in good accordance
with the calculated molecular mass of 78.5 kDa for
FoldBc-FP. The SDS-analysis revealed the location of the
autotransporter fusion protein in the outer membrane
protein fraction. The investigation of surface exposure
via FACS was not possible for foldase, since there was
no specific antibody against foldase available. Surface display of foldase Lanes 2-5 show outer membrane
preparations of E. coli BL21(DE3)pAT-LiFoBc. M: protein marker;
IPTG: protein expression was induced by adding IPTG (final concentration:
1 mM); Proteinase K: whole cells were treated with Proteinase K,
concentrations are given in mg mL-1; OmpA/OmpF: native E. coli
outer membrane proteins. The foldase and lipase fusion proteins
are indicated by black arrows. and 5). The decrease in intensity of the fusion protein bands
in comparison to the non-treated sample (Figure 4, lane 3)
indicated their surface exposure. Additionally, the constant
intensity of OmpA protein band indicates, that the cell in-
tegrity was sustained throughout this experiment. Surface display of foldase Therefore,
to elucidate if the passenger domain of FoldBc-FP is
truly surface exposed and not directed to the periplasm,
the accessibility of the fusion protein for proteases was
tested. Since proteases are too large to pass the outer
membrane, only surface exposed proteins will be de-
graded. In order to perform this degradation test whole
cells of E. coli BL21(DE3) pAT-FoldBc were incubated
with different concentrations of proteinase K. This treat-
ment resulted in degradation of FoldBc-FP (Figure 2B,
lanes 3 and 4). To demonstrate the integrity of the
outer membrane during protease treatment, outer mem-
brane protein A (OmpA) can be used as a reporter. The
C-terminal part of OmpA directs into the periplasmic
space while the N-terminal part builds a compact β-barrel
structure inside the outer membrane [37]. A digestion of
OmpA therefore can only occur from the periplasmic side,
indicating that the outer membrane lost its integrity to en-
able the access for proteases into the periplasm. Thus, the
fact, that the performed protease accessibility test led to a
strong decrease of FoldBc-FP intensity (Figure 2B, lanes 3
and 4), without affecting OmpA intensity, provides strong
evidence for the surface exposure of FoldBc-FP. Figure 4 Coexpression and surface display of both lipase and
foldase fusion protein. SDS-PAGE of membrane preparations of
E. coli BL21(DE3)pAT-LiFoBc, coexpressing the lipase and foldase
fusion protein. Molecular weight markers are indicated on the
left hand side. Lane 1 shows a membrane preparation of E. coli
BL21(DE3), used as a control. Lanes 2-5 show outer membrane
preparations of E. coli BL21(DE3)pAT-LiFoBc. M: protein marker;
IPTG: protein expression was induced by adding IPTG (final concentration:
1 mM); Proteinase K: whole cells were treated with Proteinase K,
concentrations are given in mg mL-1; OmpA/OmpF: native E. coli
outer membrane proteins. The foldase and lipase fusion proteins
are indicated by black arrows. Figure 4 Coexpression and surface display of both lipase and
foldase fusion protein. SDS-PAGE of membrane preparations of
E coli BL21(DE3)pAT-LiFoBc coexpressing the lipase and foldase Figure 4 Coexpression and surface display of both lipase and
foldase fusion protein. SDS-PAGE of membrane preparations of foldase fusion protein. SDS PAGE of membrane preparations of
E. coli BL21(DE3)pAT-LiFoBc, coexpressing the lipase and foldase
fusion protein. Molecular weight markers are indicated on the
left hand side. Lane 1 shows a membrane preparation of E. coli
BL21(DE3), used as a control. Lipase activity of outer membrane preparations from E.
Coli BL21(DE3) pAT-LiFoBc In order to apply not only whole cells but membrane
preparations for further washing experiments, the de-
scribed enzyme assay was carried out with samples of
membrane preparations as well. Membrane preparations
were derived from E. coli BL21(DE3) pAT-LiFoBc and
from previously combined E. coli BL21(DE3) pAT-LipBc
and E. coli BL21(DE3) pAT-FoldBc. To obtain the outer
membrane proteins, the preparation was performed ac-
cording to a protocol described by Schultheiss et al [35]
(see materials and methods). After the washing steps,
outer membrane proteins were suspended in 1 mL of
25 mM phosphate buffer (pH 7.4). 20 μL of a 200 μL
assay sample volume was composed of the membrane
protein suspension which was corresponding to an
amount of cells with a final OD578 of 2. As we antici-
pated that outer membrane preparation could lead to a
loss in proteins and/or enzymatic activity, the amount of
outer membrane proteins were taken from double the
amount of cells assayed in the whole cell activity deter-
mination. The photometrical assays were then carried
out at 25°C according to the same protocol as was used In addition, we investigated whether mixing the cells
displaying only the lipase with cells displaying only the
foldase could lead to whole cell lipase activity. This ap-
proach was somehow similar to that of Wilhelm et al. [18], who mixed cells displaying foldase with a dena-
tured lipase and ended up with lipase activity. In our in-
vestigation, for the combination of both types of cells, E. coli BL21(DE3) pAT-LipBc and E. coli BL21(DE3) pAT-
FoldBc were cultivated separately and protein expression
was induced as described above. Each type of cells was
washed and suspended to an OD578 of 10 as described
before. Subsequently E. coli BL21(DE3) pAT-LipBc and
E. coli BL21(DE3) pAT-FoldBc were mixed in a ratio of
1:1. Half of the sample was incubated for one hour, the
other half was incubated for 24 hours at 20°C with vigor-
ous shaking (200 rpm) to avoid sedimentation. After the
incubation enzymatic activity was determined as de-
scribed for the cells co-expressing lipase and foldase. However, mixing the cells displaying the foldase with Figure 5 Enzyme activity of whole cells displaying lipase and foldase. p-nitrophenyl palmitate was used as substrate and the increase of
absorption at 405 nm was observed photometrically. The assay was performed in potassium phosphate buffer pH 7.4 at a constant temperature
of 25°C. Lipase Activity of whole cells co-expressing LipBc-FP and
FoldBc-FP Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 6 of 12 Page 6 of 12 cells displaying the lipase did not yield any activity at all,
neither after 1 h nor after 24 h. This is to indicate that
the surface displayed lipase needs to be co-expressed
with its chaperone foldase on the surface of a single cell
to gain its enzymatic activity. temperature of 25°C. An increase of absorption values
could only be measured in the samples containing E. coli
BL21(DE3) pAT-LiFoBc (Figure 5). The host strain E. coli
BL21(DE3) showed no significant increase in absorption
at all. By using the initial enzyme reaction at min 1-4, the
extinction coefficient of p-NPP and a pathway of 0,52 cm
for a 200 μl reaction volume in the microplate reader, an
activity of 2.73 mU/ml could be calculated for E. coli BL21
(DE3) pAT-LiFoBc cells co-expressing lipase and foldase,
applied at an OD578 of 1. Lipase Activity of whole cells co-expressing LipBc-FP and
FoldBc-FP Coexpression of both LipBc-FP and FoldBc-FP
Activity of the lipase from Burkholderia cepacia is
dependent on the presence of foldase, a specific chaperone,
enabling the correct folding of the lipase [1,12]. Since E. coli
BL21(DE3) pAT-LipBc cells showed no lipase activity at all
(data not shown), co-expression of pAT-LipBc together with
pAT-FoldBc in one host was conducted. To bring both plas-
mids into one E. coli expression strain, plasmid pAT-FoldBc
was transformed into electrocompetent cells of E. coli BL21
(DE3)pAT-LipBc. Since both plasmids encode for different
antibiotic resistances, transformants harboring pAT-LipBc
and pAT-FoldBc could be identified by using selection
media containing carbenicillin as well as kanamycin. The
obtained strain was named E. coli BL21(DE3)pAT-LiFoBc. Cells co-expressing both LipBc-FP and FoldBc-FP were also
investigated for correct surface display of both autotranspor-
ter fusion proteins. Therefore co-expression of both proteins
was induced and cells were treated with proteinase K as de-
scribed above in order to determine the accessibility of lipase
and foldase fusion protein on the surface of one E. coli strain
for externally added proteases. Proteinase K treatment re-
sulted in digestion of both fusion proteins (Figure 4, lanes 4 Lipases are known to split ester bonds and an established
and easily performable assay to determine lipase activity is
the lipolytic degradation of p-nitrophenyl palmitate
(p-NPP) into p-nitrophenolate and palmitate. The
nitrophenolate anion is colored yellow and its forma-
tion can be followed spectrophotometrically at 405 nm
(εnitrophenol = 17,000 L mol-1 cm-1). To determine the lipase
activity of whole cells, E. coli BL21(DE3)pAT-LiFoBc was
cultivated and protein expression was induced as de-
scribed above. As a control the host strain E. coli BL21
(DE3) without a plasmid was cultivated analogously. Cells
were then washed twice and resuspended to an OD578 of
10 in potassium phosphate buffer (25 mM, pH 7.4). For
enzymatic conversion 20 μl of these cells were added to
180 μl of a 0.29 mM p-NPP solution in phosphate buffer
(25 mM, pH 7.4) resulting in a final substrate concentra-
tion of 0.26 mM and a final OD578 =1. The assay was per-
formed in in a 96-well plate and the kinetics of lipase
reaction was measured as the increase in absorption at
405 nm for 25 min in a microplate reader at a constant Kranen et al. Lipase activity of outer membrane preparations from E.
Coli BL21(DE3) pAT-LiFoBc The increase in absorption is caused by the nitrophenylate anion after lipolytic cleavage of the ester bond conducted by the surface
displayed lipase. ◯= E. coli BL21(DE3)pAT-LiFoBc coexpressing both lipase and foldase ▲= E. coli BL21(DE3) pAT-LipBc and E. coli UT5600(DE3)
pAT-FoldBc mixed and preincubated for one hour ■= E. coli BL21(DE3), host strain used as a control, ☐= substrate solution in buffer. Mean
values with standard deviations are shown, n = 3. Figure 5 Enzyme activity of whole cells displaying lipase and foldase. p-nitrophenyl palmitate was used as substrate and the increase of
absorption at 405 nm was observed photometrically. The assay was performed in potassium phosphate buffer pH 7.4 at a constant temperature
of 25°C. The increase in absorption is caused by the nitrophenylate anion after lipolytic cleavage of the ester bond conducted by the surface
displayed lipase. ◯= E. coli BL21(DE3)pAT-LiFoBc coexpressing both lipase and foldase ▲= E. coli BL21(DE3) pAT-LipBc and E. coli UT5600(DE3)
pAT-FoldBc mixed and preincubated for one hour ■= E. coli BL21(DE3), host strain used as a control, ☐= substrate solution in buffer. Mean
values with standard deviations are shown, n = 3. Page 7 of 12 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 was determined in a standardized test imitating a con-
ventional machine washing process. During this test,
cells and membrane fractions were compared to soluble,
reconstituted lipase from B. cepacia and Lipex® a lipase
preparation, which is already applied in washing agents. It turned out, that there was no significant difference in
lipase activity between the soluble enzyme from B. cepa-
cia, the lipase-whole cell biocatalyst and membrane
preparations thereof. These results indicate that the
lipase-whole cell biocatalyst and its membrane prepar-
ation endured the mechanically demanding procedure
(test cloth and steel balls within the washing vessel,
40°C, 45 rpm) yielding up to 100% of the lipolytic per-
formance given as relative brightening effect of Lipex®
against
Butaris®
(Figure
7). Lipolytic
performance
against the other tested fat and grease spots moved in
the range of 90-95% relative activity compared to
Lipex®. The membrane stabilization of lipase by auto-
display therefore obviously revealed no significant im-
provement in efficiency compared to soluble lipase
within this test. Lipase activity of outer membrane preparations from E.
Coli BL21(DE3) pAT-LiFoBc Nevertheless, the low differentiation
values between the tested enzyme preparations and
the relatively high standard deviations are presumably
due to the small scale testing which was applied here. Since this might be a statistical problem, a more exact
determination of differences between the several prep-
arations of lipase may be overcome by an enlargement
of the test set up and the application of a larger num-
ber of samples. Furthermore a better differentiation for whole cells. Only membrane protein preparations of
the strain co-expressing enzyme and chaperone (E. coli
BL21(DE3) pAT-LiFoBc) showed lipase activity (Figure 6). From the linear part of the curve in Figure 6 the enzym-
atic activity was determined to be 4.01 mU/ml, whereas
membrane preparations of native E. coli BL21(DE3) cells
as well as those of the pre-incubated cell mixture of E. coli
BL21(DE3) pAT-LipBc and E. coli BL21(DE3) pAT-Fold-
Bc showed no lipase activity at all (Figure 6). The deter-
mined activity for the membrane preparation from the
cells coexpressing lipase and foldase on the surface was
only by a factor of 1.5 higher than the activity of whole
cells when applied in the same assay. But as described
above the outer membrane proteins from double the
amount of cells were applied, referring to the correspond-
ing OD578.This indicates a loss of function or even a loss
of the lipase and/or foldase during the preparation proto-
col, but could also been due to a general loss in cellular
material during the centrifugation step. Nevertheless the
enzyme, co-expressed with its chaperone, showed activity
not only on the surface of E. coli cells but also in prepara-
tions of outer membrane proteins derived thereof. Application of cells and membrane preparations in a
standardized laundry test The brightening effect caused by soluble
lipase, the lipase-whole cell biocatalyst and the membrane preparation, respectively are shown in per cent relatively to the brightening effect
caused by the detergent lipase. Average values determined from three measurement points and standard deviations are depicted. White bars:
detergent lipase, light grey bars: soluble lipase from B. cepacia, dark grey bars: the herein described lipase-whole cell biocatalyst, shaded
bars: membrane preparations thereof. may be obtained by a more precise determination of
the exact number of enzymes on a single whole-cell-
biocatalyst and thus the number of enzymes applied in
one sample, which is possible by flow cytometry, for
example. Nonetheless it needs to be considered, that
this was the first time, whole cells with a surface dis-
played lipase and membrane preparations thereof were
subjected to a process like this. et al., [18], which displayed a foldase on the surface of
E. coli and added the corresponding lipase as a purified
protein subsequently and it goes an important step fur-
ther than the work of Yang et al. [19] who obtained the
surface display of an active lipase after co-expression
with foldase in a single fusion protein. Our report is
the first time description of the separate expression
and surface display of two enzymes that finally inter-
acted with each other in order to obtain an enzymatic
activity. It paves the way for the surface display of
other multiprotein or multienzyme complexes by a
similar strategy, which was to the best of our know-
ledge up to now not taken into consideration. Our data
show, that this interaction and the anchorage within
the E. coli outer membrane deliver a biocatalyst stable
enough to endure even a stressing and mechanically
demanding procedure like the standardized laundry
tests which had been conducted here. The whole cell
biocatalyst and the membrane preparations yielded an
activity in the same order of magnitude to the purified
enzyme and a standard lipase formulation already used
in detergents (Lipex®). Taken the activity 0f 4.01 mU/ml at
an OD578 = 1 as an example, the whole cell lipase/foldase
biocatalyst described here would reduce the costs in a 30
qm fermenter to 35% of those required for the purified en-
zyme to get the same amount of product, taken into con-
sideration fermentation, purification and stabilization of the
catalysts, as well as the necessary raw materials (G. Festel,
unpublished). Application of cells and membrane preparations in a
standardized laundry test One major aim of this study was the application of an
autodisplay whole-cell biocatalyst in a real-life laundry
process. Therefore the lipolytic capability of E. coli BL21
(DE3) pAT-LiFoBc and membrane preparations thereof Figure 6 Enzyme activity of outer membrane preparations obtained from cells displaying lipase and foldase. Outer membranes were
prepared as described in materials and methods and then applied to an assay with p-nitrophenyl palmitate as substrate and in which the
increase of absorption at 405 nm was observed photometrically. The assay was performed in potassium phosphate buffer pH 7.4 at a constant
temperature of 25°C. The increase in absorption is caused by the nitrophenylate anion after lipolytic cleavage of the ester bond conducted by
the surface displayed lipase. ◯= outer membrane preparations of E. coli BL21(DE3)pAT-LiFoBc coexpressing both lipase and foldase, ■= outer
membrane preparations of E. coli BL21(DE3) used as control, ☐= substrate solution in buffer. Mean values with standard deviations are shown, n = 3. Figure 6 Enzyme activity of outer membrane preparations obtained from cells displaying lipase and foldase. Outer membranes were
prepared as described in materials and methods and then applied to an assay with p-nitrophenyl palmitate as substrate and in which the
increase of absorption at 405 nm was observed photometrically. The assay was performed in potassium phosphate buffer pH 7.4 at a constant
temperature of 25°C. The increase in absorption is caused by the nitrophenylate anion after lipolytic cleavage of the ester bond conducted by
the surface displayed lipase. ◯= outer membrane preparations of E. coli BL21(DE3)pAT-LiFoBc coexpressing both lipase and foldase, ■= outer
membrane preparations of E. coli BL21(DE3) used as control, ☐= substrate solution in buffer. Mean values with standard deviations are shown, n = 3. Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 8 of 12 Figure 7 Laundry cleaning with different lipase samples in the Linitest plus testing system. The brightening effect caused by soluble
lipase, the lipase-whole cell biocatalyst and the membrane preparation, respectively are shown in per cent relatively to the brightening effect
caused by the detergent lipase. Average values determined from three measurement points and standard deviations are depicted. White bars:
detergent lipase, light grey bars: soluble lipase from B. cepacia, dark grey bars: the herein described lipase-whole cell biocatalyst, shaded
bars: membrane preparations thereof. Figure 7 Laundry cleaning with different lipase samples in the Linitest plus testing system. Application of cells and membrane preparations in a
standardized laundry test But it would be also possible to gain an even
higher enzymatic activity by E. coli BL21(DE3) pAT-LiFoBc
which exceeds the activity of purified and reconstituted
B. cepacia lipase and the detergent lipase by further
optimization of the culturing conditions and culture
medium for instance. Moreover directed evolution ap-
proaches or site-directed mutagenesis could be applied
in order to gain higher lipase activities finally. Discussion
Si
l Since ecologically friendly housekeeping processes be-
come more and more important for a broad public and
within a steadily growing biotechnological industry the
need for cost efficient and easy accessible lipase prepara-
tions increases. By means of Autodisplay a new method
to make the challenging lipase from B. cepacia easily
available was developed: Within this study we were for
the first time able to use Autodisplay for the co-
expression of two different proteins, which need to
interact with each other, a lipase and its implicitly re-
quired chaperone, foldase. By co-expression of both
these proteins on the surface of one single E. coli cell we
obtained a functional lipase-whole cell biocatalyst. Sim-
ply combining two cell types, each displaying one of the
proteins, either lipase or foldase was not sufficient to
create a functional whole cell biocatalyst. This indicates
that the interaction between lipase and foldase can only
take place if they are expressed on the surface of a single
cell. Therefore, it can be assumed that a certain vicinity
of lipase and foldase is needed for the process of folding
supported by the chaperone. The novelty of the present
investigation is, that the lipase and its specific foldase
were expressed separately and both proteins interacted
spontaneously and self driven, finally yielding an enzy-
matically active lipase at the cell surface of E. coli. In this
respect the study goes beyond the aims of Wilhelm Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 9 of 12 Page 9 of 12 Conclusion Recombinant DNA techniques Recombinant DNA techniques
For construction of plasmid pAT-LipBc, which contains
the gene encoding LipBc-FP, the lipase gene was ampli-
fied by PCR. Plasmid pHES8 served as a template for
primers EK009 (CGCTCGAGGCGAGCGCGCCCGCC-
GAC) and EK010 (GGTACCCACGCCCGCGAGCTT-
CAGCCG). To facilitate cloning of the lipase-PCR fragment
into the autotransporter cassette, a XhoI restriction site was
added to the 5′-end and a KpnI restriction site was added to
the 3′-end via PCR. For construction of plasmid pAT-
FoldBc, containing the gene which encodes for FoldBc-FP,
the foldase gene was amplified by PCR, again using pHES8
as a template for primers CD004 (CTCGAGCCGTCGTC
GCTGGCCGGCTCC) and CD005 (GGTACCCTGCGCG
CTGCCCGCGCCGCG). 5′-XhoI and 3′-KpnI-restriciton
sites were attached to the PCR fragment analogously. Both PCR products were each inserted into vector
pCR®4-TOPO® and first brought to site directed muta-
genesis according to the protocols delivered by Strata-
gene to remove unwanted restriction sites within the
genes of interest. Mutated plasmids were then restricted
with XhoI and KpnI. The restriction fragment containing
the lipase gene was ligated into pET-derivative pCD003
[25] restricted with the same enzymes. The restriction
fragment containing the foldase gene was ligated into
pCOLA-DuetTM-1–derivative pBL001 restricted with the
same enzymes before. Both ligation steps yielded an in
frame fusion of lipase or foldase respectively, with the
autotransporter domains under the control of a T7/lac
promoter. Plasmid DNA preparation, restriction digestion,
ligation, DNA electrophoresis and transformation were
performed according to standard protocols [38]. Gel ex-
traction of digested fragments was performed using a gel
extraction kit from Qiagen (Hilden, Germany). Autodisplay offers once more a convenient alternative
to obtain a functional biocatalyst without precedent
laborious purifying steps and in the special case of
B. cepacia lipase and its chaperone foldase without a
strongly required reconstitution protocol. The suc-
cessful removal of fat or grease spots respectively dur-
ing standard washing procedures was possible by
simply applying surface engineered cells and E. coli
outer membrane preparations containing active sur-
face displayed lipase. Working with a cell-free prepar-
ation which achieves the same activities like the
whole cell biocatalyst is therefore also feasible. These
results give an outlook of possible applications for en-
zymes utilized by Autodisplay beyond laboratory scale
testing. Methods Bacterial strains, plasmids and culture conditions
Escherichia coli strains UT5600(DE3) [Fˉ, ara-14, leuB6,
secA6, lacY1, proC14, tsx-67, Δ(ompT-fepC)266, entA403,
trpE38, rfbD1, rpsL109(Strr), xyl-5, mtl-1, thi-1, λ(DE3)]
and E. coli BL21(DE3) [B, Fˉ, dcm, ompT, lon, hsdS(rBˉ
mBˉ), gal, λ(DE3)] were used for the expression of auto-
transporter fusion proteins. E. coli TOP10 (F- mcrA
Δ(mrrhsdRMS-mcrBC) ϕ80lacZDM15 ΔlacX74 deoR
recA1 araD139 Δ(ara-leu) 7697 galU galK rpsL (StrR)
endA1 nupG) and the vector pCR®4-TOPO® were used
for subcloning of polymerase chain reaction (PCR)
products, using the TOPO-TA cloning kit (Invitrogen,
Carlsbad, CA, USA). Site directed mutagenesis of the
restriction sites for XhoI and KpnI inside the genes of
interest was performed using the QuikChange Site Di-
rected Mutagenesis Kit (Stratagene, Santa Clara, CA,
USA) and appropriate mutagenesis primers. Construc-
tion of plasmid pCD003 which encodes the AIDA-I
autotransporter has been described elsewhere [25]. Plas-
mid pBL001 is a pCOLA-DuetTM-1–derivative. The sec-
ond MCS had been removed and the autotransporter
cassette was inserted using NcoI and BlpI restriction sites. Plasmid pHES8, encoding the lipase and foldase from
Burkholderia cepacia, is a derivative of pHES12, which
has been described by Quyen et al. [12]. Bacteria were
routinely grown at 37°C in Lysogeny broth (LB) contain-
ing carbenicillin (100 mg L-1) or kanamycin (30 mg L-1) or
both antibiotics, respectively. For co-expression of both,
lipase and foldase, a culture from strain E. coli BL21(DE3)
pAT-LipBc, already containing the plasmid encoding for
lipase-autotransporter fusion protein, was prepared to ob-
tain electrocompetent cells according to a modified proto-
col from Sambrook et al. [38]. Plasmid pAT-FoldBc was
then transformed into an aliquot of these cells by electro-
poration resulting in strain BL21(DE3)pAT-LiFoBc which
contains both plasmids Outer membrane protein preparation p
p
p
E. coli cells were grown overnight and 1 ml of the cul-
ture was used to inoculate LB medium (40 ml). Cells
were cultured at 37°C with vigorous shaking (200 rpm)
for about 2 hours until an OD578 of 0.5 was reached. The culture was separated into two aliquots and protein
expression was induced by adding IPTG at a final con-
centration of 1 mM to one of the aliquots. Cultures then
were incubated at 30°C and shaking (200 rpm) for one
hour. Induction was stopped by incubating the cells on
ice for 15 min. After harvesting and washing of the cells
with Tris-HCl (0.2 M, pH 8), differential cell fraction-
ation was performed according to the method of Hantke
[34] as modified by Schultheiss et al. [35] In detail, cell
lysis was obtained by adding lysozyme (0.04 mg/mL
end concentration) in the presence of 10 mM sacchar-
ose and 1 μM EDTA in a final volume of 1.5 mL of
Tris-HCl (0.2 M, pH 8) and incubation for 10 min at
room temperature. Subsequently aprotinin (10 μg/mL), Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 10 of 12 phenylmethylsulfonyl fluoride (PMSF) (0.5 mM), as
well as 5 mL of extraction buffer (50 mM Tris-HCl
pH8.0, 10 mM MgCl2, 2% Triton × 100) and DNAseI
(10 μg/mL) were added. After incubation on ice for
30 min the samples were centrifuged (2,460 g, 5 min,
4.0°C) to remove intact bacteria and large cell debris. The supernatants representing the clarified bacterial
lysate were retained and centrifuged at higher speed
(38,700 × g, 30 min, 4.0°C) in order to obtain the
membrane protein fraction. The resulting supernatant,
containing soluble cytoplasmic and periplasmic pro-
teins, was completely aspirated. The pellet was sus-
pended in 10 ml phosphate-buffered saline (PBS) plus
1% Sarcosyl (N-lauryl sarcosinate, sodium salt) and
centrifuged again (38,700 × g, 60 min, 4°C). The super-
natant after this step contained the sarcosyl-soluble
cytoplasmic membrane proteins and was completely
aspirated. The sediment representing the outer mem-
brane protein fraction was washed twice with 10 ml of
water and dissolved in 30 μl water for SDS-PAGE or
an adequate volume of 25 mM potassium phosphate
buffer pH 7,4 for activity determination. For whole cell
protease treatment, E. coli cells were harvested, washed and
resuspended in 1 ml Tris-HCl (0.2 M, pH 8). Lipase activity assay
h
l A y
y
Photometrical Assays to determine lipolytic activity of
the lipase-whole cell biocatalyst were performed accord-
ing to a modified protocol by Winkler and Stuckmann
[39] with p-nitrophenylpalmitate (p-NPP) as substrate. For this purpose cells were routinely cultivated in LB
medium until an optical density at 578 nm (OD578) of
1.0 was reached. Induction of protein expression was
started by adding IPTG at a final concentration of 1 mM
and incubating the cells another hour at 30°C and
200 rpm. Cells were then harvested by centrifugation
and
washed
twice
in
potassium
phosphate
buffer,
25 mM, pH 7.4, and stored in the same buffer at 4°C in
an OD578 = 10 until used for assays. In case of mixing
different types of cells, they were used in a 1:1 ratio at
OD578 =10 and incubated at 20°C on a rocking platform
to avoid sedimentation For activity assays a stock solu-
tion of the substrate p-NPP was prepared in ethanol to a
final concentration of 7.9 mM) and finally diluted in po-
tassium phosphate buffer, 25 mM, pH 7.4 under con-
stant stirring to a working concentration of 0.29 mM. This working solution was prepared freshly, kept at 25°C
for one hour before its application and was not used
when a visible turbidity or a yellow coloring occurred. Activity measurement was started by adding 180 μl of this
working solution to 20 μl of cells with an OD578 = 10. This
yielded a final substrate concentration of 0.26 mM and a
final OD578 = 1 of the cells in the assay. The lipolytic pro-
duction of yellow colored nitrophenylate at 25°C was mea-
sured at 405 nm in a 96 well plate using a microplate
reader (Mithras LB940, Berthold, Bad Wildbach, Germany). The linear increase in absorption was used to calculate the
enzymatic activity according to the law of Lambert and For outer membrane proteins that were applied for ac-
tivity assays, cells were not treated with Proteinase K. SDS-PAGE Outer membrane isolates were diluted (1:1.5) with sam-
ple buffer (100 mM Tris/HCl (pH 6.8) containing 4%
SDS, 0.2% bromophenol blue, 200 mM dithiothreitol
and 20% glycerol), boiled for 10 minutes and analyzed
on 10% polyacrylamid gels. Proteins were stained with
Coomassie brilliant blue (R250). To correlate molecu-
lar masses of protein bands of interest, a molecular
weight standard was used (PageRuler unstained, Fermentas,
Burlington, Canada). Outer membrane protein preparation Proteinase K
was added to final concentrations between 0.2 mg mL-1 and
0.5 mg mL-1 and cells were incubated for 1 hour at 37°C. Digestion was stopped by washing the cells twice with
Tris-HCl (0.2 M, pH 8) containing 10% fetal calf serum
(FCS) and outer membrane proteins were prepared as
described above. to a final OD578 of 0.25/mL for further experiments. 100 μl of these cells were again centrifuged and resus-
pended in 500 μL PBS (pH 7.4) containing 3% bovine
serum albumin (BSA, filter sterilized) and incubated for
10 min at room temperature. After centrifuging the cells
for 60 sec with 17,000 g (Mikro200R, Hettich, Tuttlingen,
Germany), the obtained cell pellet was suspended with
100 μL of rabbit anti lipase antibody (diluted 1:50 in PBS
(pH 7.4) + 3% BSA, filter sterilized) and incubated for an-
other 30 min at room temperature. Subsequently cells
were washed twice with 500 μL of PBS (pH 7.4) + 3% BSA. Cell pellets were resuspended in 100 μL of secondary anti-
body solution (goat-anti-rabbit, DylightTM 633, Thermo
Scientific, diluted 1:25 in PBS (pH 7.4) +3% BSA) and in-
cubated for 30 min in the dark at room temperature. After
washing twice in 500 μL of PBS (pH 7.4) the cell pellet
was finally suspended in 1.5 mL of PBS (pH 7.4, filter ster-
ilized to avoid external particles). The samples were ana-
lyzed using a flow cytometer (Cyflow Space, Partec,
Münster, Germany) at an excitation wavelength of 647 nm. References 1. Arpigny JL, Jaeger KE: Bacterial lipolytic enzymes: classification and
properties. Biochem J 1999, 343:177–183. 2. Reetz MT: Lipases as practical biocatalysts. Curr Opin Chem Biol 2002,
6:145–150. 3. Tadashi E: Mechanism of enantioselectivity of lipases and other
synthetically useful hydrolases. Curr Org Chem 2004, 8:1009–1025. 3. Tadashi E: Mechanism of enantioselectivity of lipases and other
synthetically useful hydrolases. Curr Org Chem 2004, 8:1009–1025. 4. Jaeger KE, Reetz MT: Microbial lipases form versatile tools fo
biotechnology. Trends Biotechnol 1998, 16:396–403. 5. Sharma R, Chisti Y, Banerjee UC: Production, purification, characterization,
and applications of lipases. Biotechnol Adv 2001, 19:627–662. 6. Shu ZY, Jiang H, Lin RF, Jiang YM, Lin L, Huang JZ: Technical methods to
improve yield, activity and stability in the development of microbial
lipases. J Mol Catal B-Enzym 2010, 62:1–8. 5. Sharma R, Chisti Y, Banerjee UC: Production, purification, characterization,
and applications of lipases. Biotechnol Adv 2001, 19:627–662. 6. Shu ZY, Jiang H, Lin RF, Jiang YM, Lin L, Huang JZ: Technical methods to
improve yield, activity and stability in the development of microbial
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7. Gupta R, Gupta N, Rathi P: Bacterial lipases: an overview of production,
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Characterization of an extracellular lipase in Burkholderia sp. HY-10
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chaperone-dependent lipase in Escherichia coli using the dual
expression cassette plasmid. Microb Cell Fact 2012, 11:29. 12. Acknowledgements
W
h
k T V
lk
f We thank T. Voelker for the preparation of cells and membranes used in the
laundry tests. In addition, we acknowledge support by Deutsche
Forschungsgemeinschaft and Open Access Publication Fund of University of
Muenster. We thank T. Voelker for the preparation of cells and membranes used in the
laundry tests. In addition, we acknowledge support by Deutsche Forschungsgemeinschaft and Open Access Publication Fund of University of
Muenster. Flow cytometer analysis E. coli BL21(DE3) pAT-LipBc cells were grown and ex-
pression of lipase fusion protein was induced as de-
scribed above by adding IPTG to a final concentration
of 1 mM and incubating the cells for another hour at
30°C under shaking (200 rpm). Cells were harvested by
centrifugation (2400 g, 2 min, 4°C, Mikro200R, Hettich,
Tuttlingen, Germany) and washed twice with filter steril-
ized (0.2 μm pore size, ethylethersulfone membrane)
phosphate buffered saline (PBS, pH 7.4) before suspending Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Kranen et al. Microbial Cell Factories 2014, 13:19
http://www.microbialcellfactories.com/content/13/1/19 Page 11 of 12 Beer (εnitrophenol = 17,000 L mol-1 cm-1, d = 0,52 cm path
length correction for the layer thickness of 200 μL assay
volume in one well). One unit was defined as the amount
of enzyme which caused the release of 1 μmol of p-NPP
per minute [12]. For measuring the lipolytic activity of
outer membrane protein preparation we proceeded simi-
larly, with the exception, that the 20 μl which were added
to the 180 μl assay buffer with the substrate were derived
from an amount of cells corresponding to an OD578 = 2. For these activity measurements, absorption values at
405 nm obtained with outer membrane preparations in po-
tassium phosphate buffer without the addition of p-NPP
were used for blank correction. application on parts of the study (autodisplay of an active lipase). EK and CD
are working now for a company called “Autodisplay Biotech”. application on parts of the study (autodisplay of an active lipase). EK and CD
are working now for a company called “Autodisplay Biotech”. Authors’ contributions EK carried out cloning of the lipase fusion protein, performed the
coexpression of lipase and foldase, arranged and conducted all lipase activity
tests in laboratory scale and drafted the main part of the manuscript. CD
carried out cloning of the foldase fusion protein, participated in the
coexpression studies and wrote minor parts of the manuscript. TW carried
out the standardized laundry test for fatty stain removal. JJ conceived of the
study, guided its design and coordination and wrote parts of the manuscript. All authors read and approved the final manuscript. Author details
1 1Institute of Pharmaceutical and Medicinal Chemistry, PharmaCampus,
Westfalian Wilhelms-University Münster, Corrensstr. 48, 48149 Münster,
Germany. 2Autodisplay Biotech GmbH, Merowingerplatz 1a, 40225
Düsseldorf, Germany. 3International R&D/Technology Laundry & Homecare,
Henkel AG & Co. KGaA, 40191 Düsseldorf, Germany. The capability of lipase was tested on five different, stan-
dardized, lipase sensitive staining. The staining con-
tained either Biskin® (Peter Kölln KGaA, Elmshorn,
Germany), Butaris® (DFF Dairy Fine Food GmbH,
Ratzeburg, Germany) or butter oil or a mixture of
soot and mineral oil (C01, Center for Test Materials,
Vlaardingen, The Netherlands) and a mixture of cutaneous
sebum and pigment (20D, wfk Testgewebe GmbH, Krefeld,
Germany) respectively. Tested lipases were a) a standard
lipase preparation which is already used for washing pur-
poses, b) soluble lipase from B. cepacia, c) the herein de-
scribed lipase-whole cell biocatalyst and d) a membrane
preparation thereof. To allow comparability, all lipases
were applied in the same amounts, related to enzymatic ac-
tivity. The washing process was carried out in a Linitest
Plus (Atlas, Rock Hill, SC, USA), which represents the
minituarized form of a standard machine washing process. The washing solution was prepared with 3.53 g of an en-
zyme free liquid detergent similar to a european premium
detergent in water (16 °dH) buffered with 50 mM sodium
phosphate pH 7.0. The washing process took place in a
total volume of 170 mL at 40°C and 45 rpm for 60 mi-
nutes. To simulate the mechanism of a standard washing
process, 10 steel balls were added and filled up with test
cloth to a total amount of 14.3 g textile weight. Subse-
quently the test cloth was rinsed three times with deion-
ized water and dried at room temperature in the dark. Color measurement of the staining was then carried out
with a Minolta colorimeter (Konica-Minolta, München-
Neuperlach, Germany), calibrated against
producer’s
standards, applying CIE L*a*b*, D65/10°/SCI settings. Each staining was measured three times and the average
L* value was determined. Received: 8 January 2014 Accepted: 16 January 2014
Published: 29 January 2014 References Quyen DT, Schmidt-Dannert C, Schmid RD: High-level formation of active
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The presented study arose from a larger project within the scope of a patent
value fund. In this context we acknowledge the financial support provided
by the Zyrus Beteiligungsgesellschaft mbH & Co. Patente I KG (Germany).
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Th
d
d The presented study arose from a larger project within the scope of a patent
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d Submit your next manuscript to BioMed Central
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and take full advantage of: 38. Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning. A Laboratory Manual. Cold Spring Harbour, NY: Cold Spring Harbor Press; 1989. 38. Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning. A Laboratory Manual. Cold Spring Harbour, NY: Cold Spring Harbor Press; 1989. • Convenient online submission 39. Winkler UK, Stuckmann M: Glycogen, hyaluronate, and some other
polysaccharides greatly enhance the formation of exolipase by Serratia
marcescens. J Bacteriol 1979, 138:663–670. • Thorough peer review doi:10.1186/1475-2859-13-19
Cite this article as: Kranen et al.: Autodisplay for the co-expression of
lipase and foldase on the surface of E. coli: washing with designer
bugs. Microbial Cell Factories 2014 13:19. doi:10.1186/1475-2859-13-19
Cite this article as: Kranen et al.: Autodisplay for the co-expression of
lipase and foldase on the surface of E. coli: washing with designer
bugs. Microbial Cell Factories 2014 13:19.
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https://openalex.org/W3131294833
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Evaluation of Tomato-Based Packing Material for Retention of Ammonia, Nitrous Oxide, Carbon Dioxide and Methane in Gas Phase Biofilters: A Laboratory Study
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Agronomy
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| 9,210
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Citation: Pereira, J.L.S.; Perdigão, A.;
Marques, F.; Coelho, C.; Mota, M.;
Fangueiro, D. Evaluation of
Tomato-Based Packing Material for
Retention of Ammonia, Nitrous Oxide,
Carbon Dioxide and Methane in Gas
Phase Biofilters: A Laboratory Study. Agronomy 2021, 11, 360. https://
doi.org/10.3390/agronomy11020360 Keywords: ammonia; biofilter; GHG emissions; mitigation measure; tomato waste Academic Editor: Katja Klumpp
Received: 21 December 2020
Accepted: 11 February 2021
Published: 17 February 2021 agronomy agronomy agronomy agronomy Citation: Pereira, J.L.S.; Perdigão, A.;
Marques, F.; Coelho, C.; Mota, M.;
Fangueiro, D. Evaluation of
Tomato-Based Packing Material for
Retention of Ammonia, Nitrous Oxide,
Carbon Dioxide and Methane in Gas
Phase Biofilters: A Laboratory Study.
Agronomy 2021, 11, 360. https://
doi.org/10.3390/agronomy11020360 Evaluation of Tomato-Based Packing Material for Retention of
Ammonia, Nitrous Oxide, Carbon Dioxide and Methane in Gas
Phase Biofilters: A Laboratory Study a 1,2,*
, Adelaide Perdigão 1,2
, Francisco Marques 1,2, Catarina Coelho 1,2, Mariana Mota 3
ueiro 3 José L. S. Pereira 1,2,*
, Adelaide Perdigão 1,2
, Francisco Marques 1,2, Catarina Coelho 1,2, Mariana Mota 3
and David Fangueiro 3 1
Agrarian School of Viseu, Polytechnic Institute of Viseu, Quinta da Alagoa, 3500-606 Viseu, Portuga
aperdigao@esav.ipv.pt (A.P.); fm90@esav.ipv.pt (F.M.); ccoelho@esav.ipv.pt (C.C.) 1
Agrarian School of Viseu, Polytechnic Institute of Viseu, Quinta da Alagoa, 3500-606 Viseu, Portugal;
aperdigao@esav.ipv.pt (A.P.); fm90@esav.ipv.pt (F.M.); ccoelho@esav.ipv.pt (C.C.)
2
Centre for the Research and Technology of Agro-Environmental and Biological Sciences (CITAB),
University of Trás-os-Montes and Alto Douro, Quinta de Prados, 5000-801 Vila Real, Portugal
3
LEAF, Instituto Superior de Agronomia, Universidade de Lisboa, Tapada da Ajuda, 1349-017 Lisboa, Portugal;
mariana@isa.ulisboa.pt (M.M.); dfangueiro@isa.ulisboa.pt (D.F.)
*
Correspondence: jlpereira@esavipvpt *
Correspondence: jlpereira@esav.ipv.pt Abstract: Biofilters are an effective air pollution control technology to break down gaseous contam-
inants and produce innocuous end products. This laboratory study aimed to evaluate a biofilter
media, mainly composed by tomato waste, as packing material to reduce NH3, N2O, CO2 and CH4
losses from stored pig slurry. Three mixtures of packing materials, with and without oxalic acid,
were arranged in treatments, namely: mixture of tomato waste, pine bark and agricultural compost;
mixture of tomato waste and rice husk; tomato waste only. A control treatment (no biofilter) was also
included. The experiments were conducted using a system of laboratory scale biofilters connected to
jars filled with pig slurry and under a constant airflow rate. The gas concentrations were measured
for 14 days and the physicochemical of the packing materials were assessed. Results showed that
biofilter media mixtures had a potential for NH3 retention ranging from 51 to 77% and the addition
of oxalic acid to these biofilters increased NH3 retention to 72–79%. Additionally, the biofilter media
mixtures with and without oxalic acid showed a potential retention for CH4 (29–69%) but not for
N2O, yet with no impact on the global warming potential. It can be concluded that tomato based
biofilters had the potential to reduce gaseous emissions from slurry. 1. Introduction Intensive animal production is a significant source of ammonia (NH3), nitrous oxide
(N2O), carbon dioxide (CO2) and methane (CH4) emissions, all with important impacts
on climate change, acid rain and ozone formation in the troposphere [1,2]. The Integrated
Pollution Prevention and Control (IPPC) Directive (96/61/EC) requires the implementation
of best available techniques at large livestock production facilities tin order to comply with
environmental protection and the National Emissions Ceiling (NEC) Directive (2016/2284),
which sets emission reduction commitments for five main air pollutants by 2030, includ-
ing NH3. Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations. g
Ammonia comes principally from the urine, which contains most of the volatile
N excreted. Ammonium (NH4+) is in equilibrium with the NH3 that will be lost by
volatilisation, leading also to CO2 emissions. Nitrous oxide is emitted by the nitrification
and denitrification processes [3]. The anaerobic decomposition of organic matter by bacteria
and methanogenic Archaea leads to CH4 and CO2 emissions [1]. Carbon dioxide is also
produced via aerobic processes. The production and emissions of these gases is influenced
by the manure characteristics such as concentration of total ammoniacal N and dry matter
content [4]. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article
distributed
under
the
terms
and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/). https://www.mdpi.com/journal/agronomy Agronomy 2021, 11, 360. https://doi.org/10.3390/agronomy11020360 2 of 12 2 of 12 Agronomy 2021, 11, 360 Biofiltration is recommended as best available technique for the treatment of exhaust
air from mechanically ventilated animal houses, with a removal efficiency of about 50%
for NH3 [5]. Biofiltration is a biological air pollution control in which gases are treated
by passage through a biologically active porous medium [6]. In a biofilter, a contami-
nated/odorous gas stream passes through a biologically enriched layer of a filter material,
followed by biodegradation of the absorbed/adsorbed pollutant. However, N2O might be
produced in the biofilter as a by-product from both nitrification and denitrification [7]. The
by-products of microbial oxidation are primarily CO2, water, mineral salts, some volatile
organic compounds and microbial biomass [8]. Several important variables affect the
performance of a biofilter such as optimal microbial environment (nutrients, pH, humidity
and temperature), large specific surface area, structural integrity, high humidity retention,
high porosity and low bulk density [6,9]. 1. Introduction Biofiltration has been regarded as a promising technology for the treatment of exhaust
air of livestock production facilities, but information on its effectiveness is scarce [10,11]. Many different media materials have been tested as suitable biofilters. In order to be easily
implemented at farm scale, any biofilter must be simple, cheap, robust, as compact as
possible and efficient, with quick-response and low-cost maintenance [11]. The media
materials for biofilters should be locally available because the construction costs are a major
consideration when adapting biofilters to animal housing. The most widely considered
media in agriculture are organic materials such as soil, compost, wood chips, peat, bark
mulch and mixtures of these [12], but more studies are needed to assess the proper biofilters
base mixtures for each region. The aim of this laboratory study was to evaluate biofilter media, mainly composed
by tomato waste as packing material, to reduce NH3 and greenhouse gas (N2O, CO2 and
CH4) losses. 2. Materials and Methods 2.1. Biofilter Packing Materials 2.1. Biofilter Packing Materials The tomato waste and rice husk used in the present study were collected from agricul-
tural crops located in central Portugal and the pine bark and agricultural compost were
purchased from GEOLIA (Leroy Merlin, Viseu, Portugal). The particles of the wastes
were cut by hand to a size <5 mm. The agricultural wastes were sampled and analyzed
by standard laboratory methods [13–16] to assess the following parameters: pH value
by EN 13037; dry matter content by gravimetric method (24 h at 105 ◦C); total carbon
by the Dumas method; total nitrogen by the Kjeldahl method by EN 13654-1; NH4+ and
NO3−by absorption spectrophotometry; bulk density and porosity by the volumetric
method. Briefly, the bulk density was determined from the mass and volume of a circular
container, that was filled with a known volume of a waste sample, whereas the porosity
was calculated from Equation (1): POR = Ms −Md
VOL
× 100
(1) (1) where, POR was the porosity (%), Ms was the mass (kg) of the sample 24 h after being
saturated with deionised water, Md was the mass (kg) of the sample 24 h after being
oven-dried at 105 ◦C, VOL was the volume of the circular container (m−3). The physical
and chemical characteristics of the wastes used as packing materials were summarized in
Table 1. Agronomy 2021, 11, 360 3 of 12 Table 1. Physicochemical characteristics of the agricultural wastes and pig slurry used in the experiment (n = 15). Wastes
pH
DM
TC
TN
NH4+
NO3−
C/N
MD
PO
Tomato waste
6.1 b
853.3 b
848.7 b
31.0 b
31.3 a
5.7 a
28 b
90.6 e
87 a
Pine bark
4.6 d
750.0 c
855.9 b
2.2 c
0.3 b
0.1 b
411 a
276.0 c
20 d
Rice husk
6.4 b
895.6 a
843.4 b
3.3 c
0.2 b
4.3 a
257 a
149.3 d
44 c
Agricultural compost
5.1 c
363.3 d
987.6 a
12.7 b,c
0.3 b
1.9 a,b
78 b
434.3 b
60 b
Pig slurry
7.3 a
28.1 e
685.2 c
73.8 a
57.3 a
3.3 a,b
10 b
1000.0 a
Note: pH: pH (H2O), DM: dry matter (g kg−1), TC: total C (g C kg−1 DM), TN: total N (g N kg−1 DM), NH4+: NH4+-N (mg N kg−1 DM),
NO3−: NO3−-N (mg N kg−1 DM), C/N: C:N ratio, MD: mass density (kg m−3), PO: porosity (%). 2.1. Biofilter Packing Materials Data expressed on a dry weight basis. Values presented with different superscripts within columns, are significantly different (p < 0.05) by Tukey test. ochemical characteristics of the agricultural wastes and pig slurry used in the experiment (n = 15). Table 1. Physicochemical characteristics of the agricultural wastes and pig slurry used in the experim Note: pH: pH (H2O), DM: dry matter (g kg−1), TC: total C (g C kg−1 DM), TN: total N (g N kg−1 DM), NH4+: NH4+-N (mg N kg−1 DM),
NO3−: NO3−-N (mg N kg−1 DM), C/N: C:N ratio, MD: mass density (kg m−3), PO: porosity (%). Data expressed on a dry weight basis. Values presented with different superscripts within columns, are significantly different (p < 0.05) by Tukey test. The packing materials tested to reduce NH3 gas losses from pig slurry, were mainly
composed by tomato waste at a rate that ranged from 75 to 100%, compared in lab-scale
biofilters as treatments. The following three mixtures of packing materials, combined
with and without oxalic acid at a rate of 4.5%, were arranged in treatments with three
replicates, namely: tomato + pine, mixture of tomato waste, pine bark and agricultural
compost; tomato + rice, mixture of tomato waste and rice husk; tomato, tomato waste
only. For the addition of the acid to the assigned treatments, oxalic acid (2H2O, pure solid
formulation, Panreac, Odivelas, Portugal) was dissolved with 25 mL of ethanol 96% and
then was carefully sprayed by hand on the biofilter media mixtures. A control treatment
(without biofilter media and oxalic acid) was also included, which gives a total of seven
treatments considered in this study (Table 2). Table 2. Amounts of agricultural wastes and additives added to each biofilter considered as a treatment in the experiment. Treatments
Agricultural Wastes Added (g)
Additive Added (%
w/w)
Tomato Waste
Pine bark
Agricultural Compost
Rice Husk
Oxalic Acid
Control
0
0
0
0
0
Tomato + Pine
75
15
10
0
0
Tomato + Pine + Acid
75
15
10
0
4.5
Tomato + Rice
75
0
0
25
0
Tomato + Rice + Acid
75
0
0
25
4.5
Tomato
100
0
0
0
0
Tomato + Acid
100
0
0
0
4.5 wastes and additives added to each biofilter considered as a treatment in the experiment. nts of agricultural wastes and additives added to each biofilter considered as a treatment in the experimen Table 2. 2.1. Biofilter Packing Materials Amounts of agricultural wastes and additives added to each biofilter considered as a treatment in t 2.2. Lab-Scale Biofilter Setup 2.2. Lab-Scale Biofilter Setup The experiments were carried out using a system of 12 laboratory scale biofilters
(H = 135 mm, Ø = 95 mm, volume = 1.0 L) connected to Kilner jars (H = 230 mm, Ø = 105 mm,
volume = 2.0 L) filled with 1.0 L (H = 115 mm) of pig slurry each and under a constant
airflow rate (2.5 L min−1) (Figure 1). Ammonia, N2O, CO2 and CH4 concentrations were
measured during 14 days after application of constant amounts (100 g) of biofilter media
mixtures (Table 2). The temperature was recorded every 600 s by sensors (CS107, Campbell
Scientific, Leicestershire, UK) connected to a micrologger (CR3000, Campbell Scientific)
and the composition of the pig slurry used in the experiments is described in Table 1. gg
p
and the composition of the pig slurry used in the experiments is described in Table 1. Each Kilner jar had a headspace of 1.0 L between the surface of the slurry and the
lid. One air inlet and one air outlet were positioned symmetrically in the jar lid, being
inserted a Teflon tube (3 mm internal diameter) through one of the septa and the end
kept 20 mm above the slurry surface. The airflow through the headspace of each jar was
achieved by a pump (KNF, model N010.KN.18, Neuberger GmbH, Freiburg, Germany),
with a 2.5 L min−1 flowrate regulated by a needle valve coupled to a flow meter (AalborgTM
FT10201SAVN, Aalborg, Denmark) (Figure 1). The outlet air from the jars passed inside
the biofilter media mixtures and finally has been exhausted out of the climatic room by a
fume hood (Figure 1). 4 of 12 Agronomy 2021, 11, 360 The concentrations of NH3, N2O, CO2 and CH4 were measured in the exhaust air
of the biofilter media mixtures by a photoacoustic multigas monitor (INNOVA 1412i-
5, Lumasense Technologies, Ballerup, Denmark) and air samples collected, in sequence
(120 s intervals), through one sampling point (Teflon tube with 3 mm internal diame-
ter) per biofilter, by a multipoint sampler (INNOVA 1409-12, Lumasense Technologies,
Ballerup, Denmark) provided with PTFE-filters (0.001 mm pore size, Whatman, Tokyo,
Japan) (Figure 1). 2 3 Data Analysis
2.3. Data Analysis 2.3. Data Analysis
The gas concentrations of NH3, N2O, CO2 and CH4 from the exhaust air of the biofilter
media mixtures were used to calculate means per hour and day, after discounting the
following background coefficients: 0.00266 mg m−3 for NH3, 0.58942 mg m−3 for N2O,
628.71429 mg m−3 for CO2 and 1.07411 mg m−3 for CH4. The gas (NH3, N2O, CO2 or CH4)
elimination efficiency (EE, %) of each biofilter media mixture was calculated by compar-
ing the daily mean gas concentrations of the Control (exhaust sample without biofilter)
and biofilter treatments (exhaust sample of each biofilter evaluated) [17]. The overall EE
(%)
f NH
N O CO
d CH
d
i
d f
E
i
(2)
The gas concentrations of NH3, N2O, CO2 and CH4 from the exhaust air of the
biofilter media mixtures were used to calculate means per hour and day, after discounting
the following background coefficients: 0.00266 mg m−3 for NH3, 0.58942 mg m−3 for N2O,
628.71429 mg m−3 for CO2 and 1.07411 mg m−3 for CH4. The gas (NH3, N2O, CO2 or CH4)
elimination efficiency (EE, %) of each biofilter media mixture was calculated by comparing
the daily mean gas concentrations of the Control (exhaust sample without biofilter) and
biofilter treatments (exhaust sample of each biofilter evaluated) [17]. The overall EE (%) of
NH3, N2O, CO2 and CH4 was determined from Equation (2): was determined from Equation (2):
EE = ∑Cl,i − CE,i
C
n
(2)
EE =
n
∑
i = 1
Cl,i −CE,i
Cl,i
(2) (2) Cl,i
i = 1
where, Cl,i was the i-th daily mean gas concentration (NH3, N2O, CO2 or CH4) from the
exhaust air of the Control treatment (mg m−3), CE,i was the i-th daily mean gas concentra-
tion (NH3 N2O CO2 or CH4) from the exhaust air of the biofilter treatments (mg m−3) and
where, Cl,i was the i-th daily mean gas concentration (NH3, N2O, CO2 or CH4) from
the exhaust air of the Control treatment (mg m−3), CE,i was the i-th daily mean gas
concentration (NH3, N2O, CO2 or CH4) from the exhaust air of the biofilter treatments
(mg m−3), and n was the total data days. tion (NH3, N2O, CO2 or CH4) from the exhaust air of the biofilter treatments (mg m 3), and
n was the total data days. 2.2. Lab-Scale Biofilter Setup The photoacoustic multigas monitor was equipped with an optical filter
for water vapour (filter type SB0527) and the detection limits for NH3 (filter type UA0973),
N2O (filter type UA0985), CO2 (filter type UA0982) and CH4 (filter type UA0969) were,
respectively, 0.1521, 0.0589, 2.9471 and 0.2864 mg m−3. The photoacoustic multigas monitor
was calibrated by the manufacturer before the beginning of the experiment, being operated
in a mode that compensates for water interference and cross-interference. Lumasense Technologies, Ballerup, Denmark) and air samples collected, in sequence (120
s intervals), through one sampling point (Teflon tube with 3 mm internal diameter) per
biofilter, by a multipoint sampler (INNOVA 1409-12, Lumasense Technologies, Ballerup,
Denmark) provided with PTFE-filters (0.001 mm pore size, Whatman, Tokyo, Japan) (Fig-
ure 1). The photoacoustic multigas monitor was equipped with an optical filter for water
vapour (filter type SB0527) and the detection limits for NH3 (filter type UA0973), N2O
(filter type UA0985), CO2 (filter type UA0982) and CH4 (filter type UA0969) were, respec-
tively, 0.1521, 0.0589, 2.9471 and 0.2864 mg m−3. The photoacoustic multigas monitor was
calibrated by the manufacturer before the beginning of the experiment, being operated in
a mode that compensates for water interference and cross-interference. Figure 1. Layout of the laboratory system used for measuring gas concentrations in the outlet air of each biofilter. BIOFILTER
(INNOVA 1412i-5)
(INNOVA 1409-12)
MULTIGAS ANALYSER
OUTLET AIR
INLET AIR
MULTIPOINT SAMPLER
AIR PUMP
FLOW METER
KILNER JAR
Figure 1. Layout of the laboratory system used for measuring gas concentrations in the outlet air of each biofilter. INLET AIR gure 1 Layout of the laboratory system used for measuring gas concentrations in the outlet air of each biof
igure 1. Layout of the laboratory system used for measuring gas concentrations in the outlet air of each biofi 3.1. Composition of the Biofilters The initial (0 days) and final (14 days) composition of the biofilter media mixtures are
shown in Table 3. The initial and final values of pH did not differ significantly (p > 0.05)
among treatments tomato + pine and tomato + rice (average pH = 6.2 at 0 days and average
pH = 6.6 at 14 days for treatments tomato + pine and tomato + rice) but were significantly
higher (p < 0.05) in these treatments when compared with treatment tomato (pH = 5.7)
(Table 3). The addition of oxalic acid had no significant effect (p > 0.05) on the initial
and final values of pH of amended treatments, except in treatment tomato + pine + acid
(pH = 4.7 at 0 days and pH = 7.5 at 14 days), that was significant (p < 0.05) lower at 0 days
and higher at 14 days relative to the same treatment without acid (Table 3). The final
values of pH were increased significantly (p < 0.05) when compared with the initial values
(Table 3). The initial content of dry matter was significantly lower (p < 0.05) in treatment
tomato + pine (DM = 780 g kg−1) relative to treatments tomato+ rice and tomato (DM > 840
g kg−1) (Table 3). The final content of dry matter was not significantly different (p > 0.05)
in almost all treatments (Table 3). Table 3. Physico-chemical characteristics of the biofilters at the beginning and at end of the experiment (n = 3). 3.1. Composition of the Biofilters Treatments
pH
DM
TC
TN
NH4+
NO3−
C/N
MD
PO
0 days of experiment
Tomato + Pine
6.1 d,e
776.9 c,d
844 b,c
26.5 d
28 bcd
4 a
32 a,b
237 a,b
86 a
Tomato + Pine + Acid
4.7 g
606.7 e
860 a,b
28.3 b,c,d
17 d
8 a
31 a,b,c
153 b,c
86 a
Tomato + Rice
6.3 c,d
893.3 a
823 c
25.0 d
24 d
6 a
33 a
326 a
81 a
Tomato + Rice + Acid
6.4 c,d
804.4 b,c
780 d
30.7 a,b,c,d
26 c,d
7 a
26 a,b,c,d
105 c
81 a
Tomato
5.7 f
842.2 a,b,c
780 d
39.6 a,b
15 d
7 a
20 b,c,d
91 c
87 a
Tomato + Acid
5.8 e,f
862.2 a,b
748 e
38.1 a,b,c
40 a,b,c,d
8 a
20 c,d
91 c
87 a
14 days of experiment
Tomato + Pine
6.6 b,c
853.3 a,b
882 a
24.7 d
41 a,b,c,d
5 a
37 a
238 a
nd
Tomato + Pine + Acid
7.5 a
711.1 d
835 b,c
30.3 a,b,c,d
57 a,b,c
7 a
28 a,b,c,d
153 a,b
nd
Tomato + Rice
6.5 b,c,d
802.2 b,c
828 c
32.7 a,b,c,d
69 a
5 a
25 a,b,c,d
105 b,c
nd
Tomato + Rice + Acid
6.7 b,c
824.4 a,b,c
775 d
32.0 a,b,c,d
70 a
5 a
24 a,b,c,d
105 c
nd
Tomato
6.8 b
800.0 b,c
776 d
42.7 a
62 a
6 a
18 d
91 c
nd
Tomato + Acid
6.7 b,c
802.2 b,c
756 de
41.7 a
58 a,b
5 a
18 d
91 c
nd
p treatment (A)
***
***
***
***
ns
*
ns
***
***
p day (B)
***
ns
ns
ns
***
*
***
***
A × B
***
***
***
ns
*
ns
ns
***
Note: pH: pH (H2O), DM: dry matter (g kg−1), TC: total C (g C kg−1 DM), TN: total N (g N kg−1 DM), NH4+: NH4+-N (mg N kg−1 DM),
NO3−: NO3−-N (mg N kg−1 DM), C/N: C:N ratio, MD: bulk density (kg m−3), PO: porosity (%), nd: not determined. Data expressed on a
dry weight basis. Values from the interaction treatment × date are presented with different superscripts within columns (0 and 14 days
together), are significantly different (p < 0.05) by Tukey test. 2 3 Data Analysis
2.3. Data Analysis The pollutant elimination capacity (EC, mg m−3 s−1) was the mass (mg m−3) of pollu-
tant (NH3 N2O CO2 or CH4) reduced by the biofilter media mixtures expressed per unit
g
y
The pollutant elimination capacity (EC, mg m−3 s−1) was the mass (mg m−3) of
pollutant (NH3, N2O, CO2 or CH4) reduced by the biofilter media mixtures, expressed per
unit volume of the media per unit time [17]. The EC was calculated by Equation (3): time [17]. The EC was calculated by Equation (3):
EC
∑Qi(Cl i − CE i)
n
( )
EC =
n
∑
i = 1
Qi(Cl,i −CE,i)
VB
(3) (3) EC
∑
VB
i = 1
(3)
where, Qi was the i-th daily mean treatment airflow rate (m3 s−1), Cl,i was the i-th daily
mean gas concentration (NH3, N2O, CO2 or CH4) from the exhaust air of the Control treat-
ment (mg m−3), CE,i was the i-th daily mean gas concentration (NH3, N2O, CO2 or CH4)
where, Qi was the i-th daily mean treatment airflow rate (m3 s−1), Cl,i was the i-th daily
mean gas concentration (NH3, N2O, CO2 or CH4) from the exhaust air of the Control
treatment (mg m−3), CE,i was the i-th daily mean gas concentration (NH3, N2O, CO2 or
CH4) from the exhaust air of the biofilter treatments (mg m−3), and VB was the filter media
volume (m3). 5 of 12 Agronomy 2021, 11, 360 The global warming potential (GWP) for each biofilter media mixture was determined
for a time horizon of 100 years, using the GWP coefficients for direct greenhouse gas
emissions (265 for N2O, 1 for CO2 and 28 for CH4) and indirect N2O emissions (1% of
NH3-N volatilised for N2O-N) [1]. All data obtained were analysed by two-way analysis of
variance (ANOVA) to test the effects of each treatment and time on biofilter characteristics
and gas concentrations and emissions. The statistical significance (p < 0.05) of the means
difference between treatments was determined by the Tukey Honestly Significant Differ-
ence test and using the statistical software package STATISTIX 10 (Analytical Software,
Tallahassee, FL, USA). 3.1. Composition of the Biofilters ns, *, ** and *** mean that the factor or interaction effects were, respectively, not
significant or significant at the 0.05, 0.01 and 0.001 probability level. Table 3. Physico-chemical characteristics of the biofilters at the beginning and at end of the expe Note: pH: pH (H2O), DM: dry matter (g kg
1), TC: total C (g C kg
1 DM), TN: total N (g N kg
1 DM), NH4+: NH4+-N (mg N kg
1 DM),
NO3−: NO3−-N (mg N kg−1 DM), C/N: C:N ratio, MD: bulk density (kg m−3), PO: porosity (%), nd: not determined. Data expressed on a
dry weight basis. Values from the interaction treatment × date are presented with different superscripts within columns (0 and 14 days
together), are significantly different (p < 0.05) by Tukey test. ns, *, ** and *** mean that the factor or interaction effects were, respectively, not
significant or significant at the 0.05, 0.01 and 0.001 probability level. Comparatively to all other treatments, the initial values of total C in treatments
tomato + pine and tomato + pine + acid were higher and rather not statistically significant
(p > 0.05) whereas the final values of total C were significantly higher (p < 0.05) in these
same two treatments (Table 3). The addition of oxalic acid to treatment tomato + pine +
acid treatments seems reduced significantly (p < 0.05) the initial and final values of total
C relative to the same treatments without acid (Table 3). The initial and final values of
total N were higher (but not always statistically significant) in treatment tomato relative Agronomy 2021, 11, 360 6 of 12 to treatments tomato + pine and tomato + rice (Table 3). The addition of oxalic acid to
treatments had no significant effect (p > 0.05) on the initial and final values of total N of the
same treatments without acid (Table 3). The initial and final values of NH4+ did not differ
significantly (p > 0.05) among all treatments but increased significantly (p < 0.05) in the end
of the experiment (15 to 40 mg kg−1 DM at day 0 against 41 to 70 mg kg−1 DM at day 14)
(Table 3). The initial and final values of NO3−did not differ significantly (p > 0.05) among
all treatments (Table 3). 3.2.1. Ammonia Emissions The daily concentrations of NH3 from treatments along the experiment are shown in
Table 4. Comparatively to biofilter treatments without acid, the daily NH3 concentrations of
the control treatment were significantly higher (p < 0.05) during the 14 days of experiment,
with a peak in the first 6 days (14 to 31 mg NH3 m−3) followed by a progressive decrease (31
to 21 mg NH3 m−3) until the end of the experiment (Table 4). The daily NH3 concentrations
did not differ significantly (p > 0.05) among all biofilter treatments without acid, except in
the first 2 days for treatment tomato (Table 4). The mean NH3 concentrations (0–14 days)
decreased but not always significantly by the following order: treatments tomato and
tomato + rice < treatment tomato + pine < treatment control (Table 4). The addition of
oxalic acid had no significant effect (p > 0.05) on mean NH3 concentrations (0–14 days)
of biofilter treatments without acid, except in treatment tomato + pine + acid that was
significant lower (p < 0.05) relative to the same treatment without acid (4.9 mg NH3 m−3
for treatment tomato + pine + acid against 14.9 mg NH3 m−3 treatment tomato + pine)
(Table 4). Table 4. Average ammonia and nitrous oxide concentrations (mg m−3) in the outlet air of each biofilter (n = 3). 3.1. Composition of the Biofilters Comparatively to 0 days of experiment, the NO3−values did not
increase significantly (p > 0.05) in the 14 days of experiment (Table 3). The initial and final
values of C/N ratio and bulk density ratio were numerically lower in treatments tomato
and tomato + acid when compared with all other treatments (Table 3). The initial values of
porosity did not increase significantly (p > 0.05) as increased the amount of tomato-waste
in biofilter treatments (Table 3). 3.2.1. Ammonia Emissions As can be observed in Table 5, the NH3 elimination efficiency did not vary significantly
(p > 0.05) among biofilter treatments without acid (tomato + pine, tomato + rice and tomato)
and these treatments had a significant reduction (p < 0.05) from 51 to 77% relative to Agronomy 2021, 11, 360 7 of 12 treatment control. Comparatively to treatments without acid, the addition of oxalic acid
decreased significantly (p < 0.05) the NH3 elimination efficiency in treatment tomato + pine
+ acid (79% reduction with acid against 51% reduction without acid) but not in the other
treatments that received acid (Table 5). The NH3 elimination capacity of biofilter treatments
without acid (tomato + pine, tomato + rice and tomato) decreased significantly (p < 0.05)
when compared with treatment control, but no were found significant differences (p > 0.05)
among these biofilter treatments (reduction of 644 to 908 µg NH3 m3 s−1) (Table 5). No was
found significant differences (p > 0.05) among treatments with and without oxalic acid on
the NH3 elimination capacity (Table 5). Table 5. Gas elimination efficiency and pollutant elimination capacity from each biofilter (n = 3). Treatments
Gas Elimination Efficiency (EE) (%)
Pollutant Elimination Capacity (EC) (µg m−3 s−1)
NH3
N2O
CO2
CH4
NH3
N2O
CO2
CH4
GWP
Control
0 c
0 a
0 a
0 a
0 b
0 a
0 a
0 b
0 a,b,c
Tomato + Pine
↓51.4 b
↑23.4
a,b,c
↑8.8 a
↓59.2 a
↓644 a
↑5 a,b,c
↑2814 a
↓216 a,b
↓1807
a,b
Tomato + Pine + Acid
↓79.2 a
↑21.9
a,b,c
↑65.1 b
↓72.2 a
↓854 a
↑5 a,b,c
↑21425
b
↓243 ab
↑15978 c
Tomato + Rice
↓77.1 a,b
↑10.2 a,b
↓8.4 a
↓68.8 a
↓908 a
↑2 a,b
↓5542 a
↓245 a
↓11795 a
Tomato + Rice + Acid
↓72.4 a,b
↑59.4 b,c
↑1.0 a
↑139.5 b
↓809 a
↑14 b,c
↓1263 a
↑320 c
↑11347
b,c
Tomato
↓66.8 a,b
↑60.5 c
↑18.1 a
↓28.9 a
↓831 a
↑15 c
↑7169
a,b
↓99 a,b
↑8084
b,c
Tomato + Acid
↓72.9 a,b
↑63.7 c
↑19.1 a
↓49.0 a
↓908 a
↑15 c
↑7639
a,b
↓232 a,b
↑5015
a,b,c
(↓): Reduction of gas emission in comparison to control, (↑): increase of gas emission in comparison to control. Values presented with
different superscripts within columns, are significantly different (p < 0.05) by Tukey test. GWP: global warming potential as CO2-eq. 3.2.1. Ammonia Emissions and
expressed in µg (CO2 −eq.) m−3 s−1 = 0.01 × [NH3] + 265×[N2O] + 1 × [CO2] + 28 × [CH4]. Table 5. Gas elimination efficiency and pollutant elimination capacity from each biofilter (n = 3). (↓): Reduction of gas emission in comparison to control, (↑): increase of gas emission in comparison to control. Values presented with
different superscripts within columns, are significantly different (p < 0.05) by Tukey test. GWP: global warming potential as CO2-eq. and
expressed in µg (CO2 −eq.) m−3 s−1 = 0.01 × [NH3] + 265×[N2O] + 1 × [CO2] + 28 × [CH4]. The mechanism of NH3 removal occur by nitrifying bacteria that grow on the moist
packing material of biofilter [7]. The NH3 dissolves in the water phase and is converted to
nitrite and nitrate by nitrification. These compounds could be removed with the percolate
water as dissolved NH4NO2 and NH4NO3 and to some extent also are accumulated in the
organic packing material [7]. In the present study, the three biofilters evaluated presented
similar NH3 elimination efficiency (51 to 77% of reduction) and elimination capacity (644
to 908 µg NH3 m−3 s−1 of reduction) (Table 5). Although not statistically different, the
differences among biofilters evaluated in this study (Table 3) could be related with the
moisture content and characteristics of the media [11]. Chen et al. [12], studying three
moisture levels (20, 40 and 60%) of biofilters filled with wood chips, reported significant
decreases in the NH3 removal efficiency according to the type of material. y
g
yp
As can be observed in Table 6, Liu et al. [18] reported that a biofilter using woodchips
as packing material could reduce NH3 emissions from 64 to 78%, being comparable with
the value observed in the present study for the three tomato based biofilters (51 to 77% of
reduction). Also, excluding differences among packing materials referred in Table 6, the
reduction of NH3 emissions by tomato based biofilters was comparable with conventional
packing materials like woodchip, pine bark and compost, being a promising solution as
packing materials in regions where are available. 3.2.1. Ammonia Emissions Treatments
Days of Experiment
1
2
3–4
5–6
7–8
9–10
11–12
13–14
0–14
NH3 concentrations
Control
14.2 a
22.8 a,b
28.4 a
31.2 a
28.2 a
21.9 a
22.9 a
20.7 a
24.6 a
Tomato + Pine
1.8 b
2.8 b
4.2 c
7.2 c
11.0 b
14.8 a,b
17.8 a,b
17.3 a,b
14.9 b
Tomato + Pine + Acid
0.3 b
0.7 b
3.0 c
6.7 c,d
8.7 b,c
6.8 b
9.3 a,b
8.1 a,b,c
4.9 c
Tomato + Rice
4.3 b
2.7 b
3.5 c
3.0 c,d
2.9 c
4.4 b
8.0 a,b
9.8 a,b,c
5.0 c
Tomato + Rice + Acid
1.3 b
8.5 a,b
12.8 b
11.7 b
9.6 b,c
3.2 b
3.4 b
4.2 b,c
7.1 b,c
Tomato
19.1 a
30.8 a
4.2 c
3.4 c,d
3.3 c
3.4 b
4.0 b
3.3 c
6.7 b,c
Tomato + Acid
21.3 a
8.6 a,b
2.2 c
2.6 d
3.2 c
4.0 b
4.6 b
3.5 c
5.0 c
N2O concentrations
Control
0.5 cd
0.5 b
0.5 a
0.5 c
0.5 c
0.5 d
0.5 d
0.5 d
0.5 a
Tomato + Pine
0.6 b,c
0.6 b
0.6 a
0.6 b,c
0.6 b
0.6 b
0.6 b
0.6 c
0.6 a
Tomato + Pine + Acid
0.7 b,c
0.7 a,b
0.6 a
0.6 b,c
0.6 b
0.6 b
0.6 b
0.6 c
0.6 a
Tomato + Rice
0.6 b,c
0.5 b
0.6 a
0.5 b,c
0.5 c
0.6 c
0.5 c
0.5 d
0.6 a
Tomato + Rice + Acid
0.3 d
0.7 a,b
0.6 a
0.6 a,b
0.7 a
0.8 a
0.7 a
0.6 c
0.8 a
Tomato
0.8 ab
1.2 a
0.7 a
0.8 a
0.7 a
0.7 a
0.9 a
0.8 b
0.8 a
Tomato + Acid
0.9 a
1.0 ab
0.7 a
0.8 a
0.7 a
0.8 a
1.0 a
0.9 a
0.8 a
For each gas, values presented with different superscripts within columns, are significantly different (p < 0.05) by Tukey test. le 4. Average ammonia and nitrous oxide concentrations (mg m−3) in the outlet air of each biofilter (n = 3). 3.2.1. Ammonia Emissions When the biofilters were coated with
oxalic acid crystals to absorb NH3 from airflow that passed by the packing material [19,20],
a significant NH3 elimination efficiency was observed only in treatment tomato + pine +
acid, what could be related with the lower pH value and higher moisture content relative to
the other two biofilters that were coated with oxalic acid [20]. In addition, minor advantages
were gained from the addition of oxalic acid to the three tomato-based biofilters, with 72
to 79% reduction (average of 75%) with acid against 51 to 77% reduction without acid
(average of 66%). Agronomy 2021, 11, 360 8 of 12 Table 6. Pollutant removal efficiencies of biofilters for animal housing facilities. Type of Biofilter
NH3 (%)
N2O (%)
CO2 (%)
CH4 (%)
Country
Reference
Biofilter
9 to 99
Netherlands
[21]
Biofilter
−60 to 93
−167 to 37
5 to 21
Netherlands
[22]
Woodchips
64 to 78
−19 to −13
China
[18]
Woodchips
−10 to 90
France
[11]
Woodchips
7 to 83
−400 to 13
−43 to 41
Netherlands
[23]
Pine bark and lava rock
56
1
25
[24]
Coconut and peat
−9 to 81
Netherlands
[25]
Coconut and peat
11 to 26
−29 to −26
−1 to 0
−2 to 10
USA
[26]
Perlite and compost
90 to 100
85
Netherlands
[24]
Tomato based
51 to 77
−61 to −10
−18 to −9
29 to 69
Portugal
This study
3.2.2. Nitrous Oxide Emissions Table 6. Pollutant removal efficiencies of biofilters for animal housing facilities. 3.2.2. Nitrous Oxide Emissions 3.2.2. Nitrous Oxide Emissions The daily concentrations of N2O from all biofilter treatments and control did not varied
along the experiment (Table 4). The daily N2O concentrations, including the average values
(0–14 days), from biofilter treatments and control did not differed significantly (p > 0.05) in
most measurement days (0.5 to 1.2 mg N2O m−3), being numerically higher in treatment
tomato (Table 4). Comparatively to treatment control, the mean N2O concentrations
(0–14 days) were numerically higher in biofilter treatments without acid by the following
order: treatment tomato > treatments tomato + rice and tomato + pine > control (Table 4). The addition of oxalic acid to assigned treatments did not reduce significantly (p > 0.05) the
mean N2O concentrations (0–14 days) relative to biofilter treatments without acid (Table 4). 3.3. Carbon Emissions from Biofilters
3.3.1. Carbon Dioxide Emissions 3.3. Carbon Emissions from Biofilters 3.2.1. Ammonia Emissions The N2O elimination efficiency was not significantly decreased (p > 0.05) in biofilter
treatments without acid relative to treatment control, being observed a significant increase
(p < 0.05) in treatment tomato when compared with treatments tomato + pine and tomato +
rice (61% increase for treatment tomato against 17% increase for treatments tomato + pine
and tomato + rice) (Table 5). The addition of oxalic acid did not affect significantly (p > 0.05)
the N2O elimination efficiency of biofilter treatments (tomato + pine, tomato + rice and
tomato) relative to these same biofilter treatments without acid (Table 5). The N2O elimi-
nation capacity was not significantly reduced (p > 0.05) in treatment tomato (15 µg N2O
m3 s−1 of increase) relative to all other biofilter treatments (2 µg N2O m3 s−1 of increase)
(Table 5). In addition, no significant differences (p > 0.05) were found among treatments
with and without oxalic acid on the N2O elimination capacity (Table 5). p
y
Ammonia gas from contaminated air streams recovered through absorption is then
oxidized into nitrite and nitrate by microorganisms that grow on the surface of packing
media [7]. In anaerobic zones of the biofilter, denitrification can also take place, meaning
that part of the nitrite and/or nitrate is converted to N2 [7]. In the present study, the three
tomato-based biofilters without acid increased significantly the N2O emissions (Table 5),
particularly for biofilter with 100% tomato, which could be related with the high amount
of N in tomato waste (Table 3). Thus, N2O is influenced by moisture content and might be
produced in the biofilter as a by-product from both nitrification and denitrification [7,11]. p
y p
The results of this study followed the same trend than previous studies [18,23,26]
reported in Table 6, where biofilters using different packing materials increased the N2O
emissions. In the present study, the addition of oxalic acid crystals to the biofilters had
no effect on N2O losses, which could be related with the absence of effect on the reduc-
tion of pH of packing materials (Table 3) and then did not affected the nitrification and
denitrification processes. 3.3.1. Carbon Dioxide Emissions In all biofilter treatments and control, the daily concentrations of CO2 were quite
similar during the 14 days of experiment (Table 7). For almost all measurement days of
experiment, the daily CO2 fluxes, including the average values (0–14 days), from biofilter Agronomy 2021, 11, 360 9 of 12 treatments without acid and control did not varied significantly (p > 0.05) (870 to 1660 mg
CO2 m−3), being observed numerically lower fluxes in treatment tomato + rice and higher
fluxes in treatment tomato (Table 7). The addition of oxalic acid to assigned biofilter treat-
ments increased, but not significantly (p > 0.05), the mean CO2 concentrations (0–14 days)
comparatively to biofilter treatments without acid and control (Table 7). Table 7. Average carbon dioxide and methane concentrations (mg m−3) in the outlet air of each biofilter (n = 3). 3.3.1. Carbon Dioxide Emissions Treatments
Days of Experiment
1
2
3–4
5–6
7–8
9–10
11–12
13–14
0–14
CO2 concentrations
Control
1114 c,d
1023 b,c
1014 b
1046 b
1104 a,b
990 b,c
934 b
1402 a
1080 a
Tomato + Pine
1182 c,d
1132 b,c
1057 b
1068 b
1033 a,b
1165 a,b
1184 a,b
1220 a
1141 a,b
Tomato + Pine + Acid
1365
a,b,c
1599 a
2206 a
1555 a
1278 a
1334 a
1460 a
1475 a
1716 a,b
Tomato + Rice
956 d
968 c
982 b
948 b
868 c
903 c
996 b
1064 a
961 b
Tomato + Rice + Acid
1314 b,c
1112 b,c
893 b
1011 b
1004 b,c
1077 b,c
1141 a,b
1030 a
1053 b
Tomato
1657 a
1109 b,c
1171 b
1181 a,b
1220 a
1207 a,b
1288 a,b
1191 a
1234 a,b
Tomato + Acid
1555 a,b
1234 b
1194 b
1215 a,b
1227 a
1201 a,b
1289 a,b
1192 a
1244 a,b
CH4 concentrations
Control
5.4 a
7.8 a
8.1 a
4.8 a
5.6 a
8.5 a
10.7 a
9.8 a
7.9 a
Tomato + Pine
2.5 a
3.0 a
3.2 a
1.7 a
2.2 a
3.7 a,b
5.0 a,b,c
4.3 a,b
3.3 a
Tomato + Pine + Acid
5.4 a
1.0 a
1.1 a
1.1 a
1.4 a
2.6 b
6.1 a,b
3.4 b
2.1 a
Tomato + Rice
2.6 a
1.6 a
1.7 a
2.3 a
1.5 a
1.8 b
4.3 b,c,d
4.8 b
2.6 a
Tomato + Rice + Acid
68.1 a
49.4 a
23.6 a
9.3 a
6.6 a
2.6 b
2.0 c,d
0.8 b
14.8 a
Tomato
27.2 a
19.0 a
4.1 a
1.7 a
1.0 a
0.4 b
0.2 c,d
0.2 b
4.4 a
Tomato + Acid
28.7 a
4.9 a
1.8 a
0.9 a
0.6 a
0.1 b
0.1 d
0.1 b
2.9 a
For each gas, values presented with different superscripts within columns, are significantly different (p < 0.05) by Tukey test. Average carbon dioxide and methane concentrations (mg m−3) in the outlet air of each biofilter (n = 3). The CO2 elimination efficiency was not significantly reduced (p > 0.05) in biofilter
treatments without acid relative to treatment control (Table 5). 3.3.1. Carbon Dioxide Emissions The addition of oxalic acid
increased significantly (p < 0.05) the CO2 elimination efficiency in treatment tomato + pine
+ acid relative to treatment tomato + pine, but not in all other treatments (Table 5). The CO2
elimination capacity was not significantly reduced (p > 0.05) in biofilter treatments with
and without acid relative to treatment control, except in treatment tomato + pine + acid
(Table 5). Carbon dioxide is originated by decomposition of urea and by anaerobic decomposi-
tion of the organic matter present in animal manure, as well as by microbial oxidation of
gaseous contaminants in the packing material of biofilter [8,27]. Although the decomposi-
tion of packing material of biofilters evaluated in the present study, the three tomato- based
biofilters without acid evaluated had similar CO2 elimination efficiency and elimination
capacity than values reported by Martinec et al. [26] for biofilters equipped with coconut
and peat as pack materials. The addition of oxalic acid to assigned biofilter treatments
increased significantly (p < 0.05) the CO2 elimination capacity in treatment tomato + pine +
acid relative to treatment tomato + pine, but not in all other biofilter treatments (Table 5). 3.3.2. Methane Emissions In each biofilter treatment and control, the daily concentrations of CH4 did not
varied greatly during the 14 days of experiment (Table 7). On most all days of the ex-
periments, the daily CH4 concentrations and average values (0–14 days) did not differ
significantly (p > 0.05) among all biofilter treatments with and without acid and treatment
Control (0 to 70 mg CH4 m−3) (Table 7). The mean CH4 concentrations (0–14 days) were
higher, although not statistically different, in treatment control (7.9 mg CH4 m−3) when
compared with all other biofilter treatments without acid (treatments tomato + pine, tomato
+ rice and tomato) (2.6 to 4.4 mg CH4 m−3). Comparatively to biofilter treatments without Agronomy 2021, 11, 360 10 of 12 10 of 12 acid, the addition of oxalic acid did not reduce significantly (p > 0.05) the mean CH4
concentrations of these same biofilter treatments (Table 7). acid, the addition of oxalic acid did not reduce significantly (p > 0.05) the mean CH4
concentrations of these same biofilter treatments (Table 7). The CH4 elimination efficiency did not decrease significantly (p > 0.05) in biofilter
treatments without acid relative to treatment control (Table 5). The addition of oxalic acid to
assigned biofilter treatments reduced significantly (p < 0.05) the CH4 elimination efficiency
in treatment tomato + pine + acid relative to treatment tomato + pine, but not in all other
biofilter treatments (Table 5). The CH4 elimination capacity was not significantly reduced
(p > 0.05) in biofilter treatments without acid when compared with treatment Control
(Table 5). The addition of oxalic acid to assigned treatments reduced significantly (p < 0.05)
the CH4 elimination capacity in treatment tomato + rice + acid relative to treatment tomato
+ rice, but not in all other treatments (Table 5). On other hand, the GWP elimination
efficiency was not reduced significantly (p > 0.05) in biofilter treatments without acid
relative to treatment control (Table 5). The addition of oxalic acid to assigned biofilter
treatments reduced significantly (p < 0.05) the GWP elimination capacity in treatments
tomato + pine + acid and tomato + rice + acid relative to the same biofilter treatments
without acid (Table 5). In the present study, the three tomato-based biofilters had similar CH4 elimination
efficiency (29 to 69% of reduction) and elimination capacity (99 to 245 µg CH4 m3 s−1
of reduction) (Table 5). 3.3.2. Methane Emissions The elimination of CH4 in a biofilter is mainly through aerobic
conversion of CH4 to CO2 and water by the so-called methanotrophic bacteria [28]. Also,
the adsorption of CH4 in the packing materials, low water solubility and biodegradation
rate may also contribute to the CH4 removal [7,29]. The achieved CH4 removal efficiencies (29 to 69% of reduction) in the three tomato
based biofilters are in line with previous biofilter studies [22–24,26,30] reported in Table 6,
with average values that varied from 0 to 85% for biofilters equipped with different type of
pack materials. At the present study, the addition of oxalic acid crystals to the biofilters did
not reduced the CH4 losses in treatment tomato + rice + acid (140% of increase) relative to
the other two acidified treatments (49 to 72%), which may be due to the higher pH value in
this treatment (pH = 6.4 for treatment tomato + rice + acid against pH < 5.8 for treatments
tomato + pine + acid and tomato + acid) (Table 3). Conflicts of Interest: The authors declare no conflict of interest. Conflicts of Interest: The authors declare no conflict of interest. 4. Conclusions The present study suggests that biofilter media mixtures, mainly composed by tomato
waste, had a potential for NH3 retention between 51 and 77% and the addition of oxalic
acid to these biofilters increased NH3 retention to 72–79%. Additionally, the biofilter media
mixtures with and without oxalic acid showed a potential retention for CH4 (29–69%)
but not for N2O, yet with no impact on the global warming potential. The tomato based
biofilters had the potential to reduce gaseous emissions from livestock production facilities,
but further full-scale studies are needed to confirm these inferences. Author Contributions: Conceptualization, J.L.S.P.; methodology, J.L.S.P., A.P. and D.F.; software,
J.L.S.P., A.P. and D.F.; validation, J.L.S.P., A.P., F.M., C.C., M.M. and D.F.; formal analysis, J.L.S.P.,
A.P. and D.F.; investigation, J.L.S.P., A.P., F.M., C.C., M.M. and D.F.; resources, J.L.S.P., A.P., F.M.,
C.C., M.M. and D.F.; data curation, J.L.S.P., A.P., F.M., C.C., M.M. and D.F.; writing—original draft
preparation, J.L.S.P. and D.F.; writing—review and editing, J.L.S.P. and D.F.; visualization, J.L.S.P. and
D.F.; supervision, D.F.; project administration, J.L.S.P. and D.F.; funding acquisition, J.L.S.P. and D.F. All authors have read and agreed to the published version of the manuscript. Funding: This research was funded by National Funds by FCT—Portuguese Foundation for Science
and Technology, under the projects UIDB/04033/2020 and UID/AGR/04129/2020, and project
Cleanslurry PTDC/ASP-SOL/28769/2017. Institutional Review Board Statement: Not applicable. Informed Consent Statement: Not applicable. Data Availability Statement: Not applicable. 11 of 12 Agronomy 2021, 11, 360 References 1. IPCC. 2019 Refinement to the 2006 IPCC Guidelines for National Greenhouse Gas Inventories; Calvo Buendia, E., Tanabe, K., Kranjc, A.,
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2019; Available online: http://www.ipcc-nggip.iges.or.jp (accessed on 17 December 2020). p
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27. Pereira, J.L.S.; Figueiredo, V.; Pinto, A.F.M.A.; Silva, M.E.F.; Brás, I.; Perdigão, A.; Wessel, D.F. Effects of Biochar and Clinoptilolite
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27. Pereira, J.L.S.; Figueiredo, V.; Pinto, A.F.M.A.; Silva, M.E.F.; Brás, I.; Perdigão, A.; Wessel, D.F. Effects of Biochar and Clinoptilolite
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https://openalex.org/W3130577588
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https://pure.uva.nl/ws/files/59310422/s41467_021_21349_3.pdf
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English
| null |
Ice-nucleating proteins are activated by low temperatures to control the structure of interfacial water
|
Nature communications
| 2,021
|
cc-by
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UvA-DARE (Digital Academic Repository) Ice-nucleating proteins are activated by low temperatures to control the structure
of interfacial water Roeters, S.J.; Golbek, T.W.; Breg
G.; Šantl-Temkiv, T.; Finster, K.; P
DOI
10.1038/s41467-021-21349-3
Publication date
2021
Document Version
Final published version
Published in
Nature Communications
License
CC BY
Link to publication Citation for published version (APA):
Roeters, S. J., Golbek, T. W., Bregnhøj, M., Drace, T., Alamdari, S., Roseboom, W., Kramer,
G., Šantl-Temkiv, T., Finster, K., Pfaendtner, J., Woutersen, S., Boesen, T., & Weidner, T. (2021). Ice-nucleating proteins are activated by low temperatures to control the structure of
interfacial water. Nature Communications, 12, Article 1183. https://doi.org/10.1038/s41467-
021-21349-3 Citation for published version (APA):
Roeters, S. J., Golbek, T. W., Bregnhøj, M., Drace, T., Alamdari, S., Roseboom, W., Kramer,
G., Šantl-Temkiv, T., Finster, K., Pfaendtner, J., Woutersen, S., Boesen, T., & Weidner, T. (2021). Ice-nucleating proteins are activated by low temperatures to control the structure of
interfacial water. Nature Communications, 12, Article 1183. https://doi.org/10.1038/s41467-
021-21349-3 Citation for published version (APA): General rights General rights
It is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s)
and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open
content license (like Creative Commons). UvA-DARE is a service provided by the library of the University of Amsterdam (https://dare uva nl)
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If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please
let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material
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to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You
will be contacted as soon as possible. 1 Department of Chemistry, Aarhus University, Aarhus C, Denmark. 2 Van ‘t Hoff Institute for Molecular Sciences, University of Amsterdam, Amsterdam, The
Netherlands. 3 Department of Molecular Biology and Genetics, Aarhus University, Aarhus C, Denmark. 4 Department of Chemical Engineering, University of
Washington, Seattle, WA, USA. 5 Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, The Netherlands. 6 Department of Biology,
Aarhus University, Aarhus C, Denmark. 7 The Stellar Astrophysics Centre – SAC, Department of Physics and Astronomy, Aarhus University, Aarhus
C, Denmark. 8 Interdisciplinary Nanoscience Center – iNano, Aarhus University, Aarhus C, Denmark. 9These authors contributed equally: Steven J. Roeters,
Thaddeus W. Golbek, Mikkel Bregnhøj. ✉email: weidner@chem.au.dk Disclaimer/Complaints regulations
f UvA-DARE is a service provided by the library of the University of Amsterdam (https://dare.uva.nl)
will be contacted as soon as possible. Download date:24 Oct 2024 ARTICLE ARTICLE ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 I
ce-nucleation active (INA) bacteria are the most effective ice
nucleators known1–3. Using ice-nucleating proteins (INPs)
attached to their outer membrane, they promote ice growth at
high sub-zero temperatures to gain access to nutrients from
plants4. INA bacteria can cause severe frost damage to crops and
other types of vegetation. Water and ice are fundamental for life
and INA bacteria play important roles in this context as they
affect interfacial water, the hydrological cycle, local and global
climate, and vegetation5–7. Recent field studies have revealed large
amounts of INA bacteria at high altitudes8, which have been
identified as an important cause of cloud glaciation, snow and
hail precipitation, and cloud formation5,6,9. Pure water droplets
in the atmosphere can remain supercooled and liquid down to
~−40 °C. It has been shown that, in the presence of the ice-active
bacterium Pseudomonas syringae, water freezes at temperatures as
high as −2 °C1,10. INA bacteria may therefore play an important
role especially in mixed-phase, low-altitude clouds, where abiotic
particles are far less effective ice-nucleating particles than
bacteria1,11. With their record-holding ice activity, P. syringae has
also caught attention for technological applications such as arti-
ficial snow formation, food preservation, cryo-medicine, and
freezing technologies2,12–14. I visible and infrared laser pulses when the infrared pulse is reso-
nant with a surface vibration30. The selection rules of SFG dictate
that only ordered species at an interface are detected. Therefore,
in the amide-I region, SFG can probe the structure of interfacial
proteins and in the water stretching region one can use the
technique to observe water molecules interacting with the protein
interface. Previous SFG studies have shown that P. syringae and INPs
anchored to the surface of hollow Escherichia coli cell envelopes
can order water at the bacterial surface31–33, and that water
interaction is enhanced close to the water melting point31,34. However, the molecular basis for water ordering and the
mechanism behind the activation at low temperatures remain
unclear. Bacterial cell surfaces contain a variety of different pro-
teins and other biomolecules and therefore these experiments,
performed with entire P. syringae cells, could not identify the
molecular origin of water ordering. Here, we report on experiments with isolated model INPs. We
use a truncated version of InaZ that contains 9 out of 67 repeat
units (repeats 1–4 and 63–67) of the native INA protein (see
Fig. 1) of P. Results and discussion
W
h Results and discussion
Water structure at the InaZ interface. We investigate the
interaction of water in direct contact with InaZ9R with SFG
spectroscopy in the water region. Heavy water was used in the
experiments for two reasons. First, the vibrational spectrum of
D2O is narrower than that of H2O and can therefore be probed
more conveniently using broadband SFG. Second, using D2O
avoids any overlap of water-bending modes with the amide-I
region. Ice nucleation takes place at the very interface between single
layers of protein interacting with only a few layers of water
molecules. The current lack of experimental data about INP
activity can be traced back to the general challenges involved in
studying the conformation of proteins and water at such thin
interfaces. Vibrational sum-frequency generation (SFG) spectro-
scopy allows one to probe the structure of InaZ at the air–water
interface as a model system for the cell–water interface. SFG relies
on the resonant enhancement of frequency mixing between g
Figure 2 displays temperature-dependent SFG spectra collected
with the ssp (s-polarized SFG, s-polarized visible, and p-polarized
infrared) polarization combination for InaZ9R in phosphate-
buffered saline (PBS) buffer at the air–water interface (panel A)
and for neat PBS buffer (panel B). For both the InaZ and pure
buffer spectra, resonances corresponding to O–D stretching and Fig. 1 The InaZ repeat sequence and the model InaZ9R construct. Based on current models, the blue sequences are ice-nucleation active sites. A Repeat
structure of the ice-nucleation active protein InaZ without the N- and C-terminal domains. B Sequence of the full InaZ9R construct. It contains an N-
terminal tag, the membrane-binding N-terminal domain, the ice-active repeat units, and a C-terminal domain of unknown function15,17,61,62. Fig. 1 The InaZ repeat sequence and the model InaZ9R construct. Based on current models, the blue sequences are ice-nucleation active sites. A Repeat
structure of the ice-nucleation active protein InaZ without the N- and C-terminal domains. B Sequence of the full InaZ9R construct. It contains an N-
terminal tag, the membrane-binding N-terminal domain, the ice-active repeat units, and a C-terminal domain of unknown function15,17,61,62. Fig. 1 The InaZ repeat sequence and the model InaZ9R construct. Based on current models, the blue sequences are ice-nucleation active sites. A Repeat
structure of the ice-nucleation active protein InaZ without the N- and C-terminal domains. B Sequence of the full InaZ9R construct. ARTICLE syringae embedded in the native N- and C-terminal
domains. Although the truncation may lead to a reduced ice-
nucleation activity, the strategy behind the design has been to
provide a native environment for the INA repeat units. For this
construct (termed InaZ9R), we show that the central repetitive
region of InaZ adopts a β-helical structure and provide conclusive
evidence for the ordering of water molecules by INPs. In addition,
our data indicate that InaZ reorients at low temperatures and
thereby increases contact to water molecules, which increases ice-
nucleation activity. g
g
Despite the numerous environmental and economic aspects
where bacterial ice nucleation is of critical importance4,7, the
fundamental mechanisms of protein-driven ice nucleation are still
not clear. The most studied model system is InaZ, the INP of P. syringae. Although the InaZ sequence (containing ~1200 resi-
dues) has been known for over 30 years15,16, and there have been
several theoretical studies of INP folding17–20, only limited
experimental
information
on
the
structure
of
INPs
is
available16,21–26. The current picture of the structure of INPs and
their interaction with water is still based on theoretical models
and molecular-dynamics (MD) simulations17,27,28, with contra-
dicting conclusions. The first model was put forward by Kajava
and Lindow and is based on parallel β-sheets19. Later, based on
homology modeling, Graether and coworkers proposed that InaZ
folds into a β-helical structure (Fig. 1), similar to insect antifreeze
proteins17,27,29. Ice-nucleating proteins are activated by
low temperatures to control the structure
of interfacial water Steven J. Roeters1,2,9, Thaddeus W. Golbek1,9, Mikkel Bregnhøj
1,9, Taner Drace3, Sarah Alamdari4,
Winfried Roseboom5, Gertjan Kramer5, Tina Šantl-Temkiv
6,7, Kai Finster
6,7, Jim Pfaendtner
4,
Sander Woutersen2, Thomas Boesen
3,8 & Tobias Weidner
1,4,8✉ Steven J. Roeters1,2,9, Thaddeus W. Golbek1,9, Mikkel Bregnhøj
1,9, Taner Drace3, Sarah Alamdari4,
Winfried Roseboom5, Gertjan Kramer5, Tina Šantl-Temkiv
6,7, Kai Finster
6,7, Jim Pfaendtner
4,
Sander Woutersen2, Thomas Boesen
3,8 & Tobias Weidner
1,4,8✉ Ice-nucleation active (INA) bacteria can promote the growth of ice more effectively than any
other known material. Using specialized ice-nucleating proteins (INPs), they obtain nutrients
from plants by inducing frost damage and, when airborne in the atmosphere, they drive ice
nucleation within clouds, which may affect global precipitation patterns. Despite their evident
environmental importance, the molecular mechanisms behind INP-induced freezing have
remained largely elusive. We investigate the structural basis for the interactions between
water and the ice-nucleating protein InaZ from the INA bacterium Pseudomonas syringae. Using vibrational sum-frequency generation (SFG) and two-dimensional infrared spectro-
scopy, we demonstrate that the ice-active repeats of InaZ adopt a β-helical structure in
solution and at water surfaces. In this configuration, interaction between INPs and water
molecules imposes structural ordering on the adjacent water network. The observed order of
water increases as the interface is cooled to temperatures close to the melting point of water. Experimental SFG data combined with molecular-dynamics simulations and spectral calcu-
lations show that InaZ reorients at lower temperatures. This reorientation can enhance water
interactions, and thereby the effectiveness of ice nucleation. 1 NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications TURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/ NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 Although the
N–D-modes may potentially affect a ~100 cm−1 region, the whole
O–D region (~500 cm−1 broad) exhibits an increasing SFG signal
for decreasing temperatures, thus making it probable that the
influence of the N-D modes on the spectra is rather limited. Temperature changes can potentially affect the density of
interfacial protein layers and thereby influence the water signal. We therefore determined the protein coverage of the air–water
interface at 5, 10, and 20 °C with X-ray photoelectron spectro-
scopy. This data, recorded for InaZ9R lifted off the air–water
interface, showed that the surface coverage was largely unaffected
by the temperature change (see Supplementary Note 1). InaZ secondary structure in solution. This raises the question as
to what is the structural basis for the enhanced water order at
lower temperatures. Previous studies with INA bacterial cells have
not been able to answer this question because of the structural
heterogeneity of bacterial cell surfaces. With the InaZ9R con-
struct, we now attempt to address this question—by firstly
determining the solution-state structure of InaZ9R and thereafter
resolving the binding geometry of InaZ9R at the air–water
interface. To determine the secondary structure in solution, we use
Fourier-transform infrared (FT-IR) and two-dimensional infrared
(2D-IR) spectroscopy. Figure 3A displays the experimental FT-IR
and 2D-IR amide-I (1600–1700 cm−1) spectra of InaZ9R in PBS
buffer. Two-dimensional IR spectra can be recorded by exciting
the sample with a narrowband pump pulse with a given frequency
ωpump, after which the IR absorbance is recorded across the whole
spectral range with a broadband probe pulse. By plotting the
differential absorbance with versus without the pump excitation
as a function of ωprobe and the scanned ωpump, 2D-IR spectra can
be obtained, which can be regarded as the vibrational analog of
2D-NMR spectra41. If two modes, A and B, are coupled, the
absorption of one is affected when the other is excited. So, if
mode A is excited by the pump beam (ωpump = ωA), then an
absorption change will be recorded with the probe beam at
ωprobe = ωB, and vice versa, which gives rise to a distinct cross-
peak pattern at (ωA, ωB) and (ωB, ωA) in the 2D-IR spectrum. In
the case of the amide-I mode of proteins, the individual cross-
peaks can generally not be resolved, but the shape of the amide-I
2D-IR spectrum is still very sensitive to the secondary structure42. NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 As the buffer temperature
is decreased to 10 °C, and subsequently to 5 °C, the water signal is
strongly increased. Figure 2C summarizes the intensity changes of
the different water modes (see Table S3 for fitting parameters). Potential N–D contributions from the backbone, the so-called
amide-A modes, or from side chains may also contribute to this
spectral region as a narrow peak at ~2550 cm−1 39. Although the
N–D-modes may potentially affect a ~100 cm−1 region, the whole
O–D region (~500 cm−1 broad) exhibits an increasing SFG signal
for decreasing temperatures, thus making it probable that the
influence of the N-D modes on the spectra is rather limited. Temperature changes can potentially affect the density of
interfacial protein layers and thereby influence the water signal. We therefore determined the protein coverage of the air–water
interface at 5, 10, and 20 °C with X-ray photoelectron spectro-
scopy. This data, recorded for InaZ9R lifted off the air–water
interface, showed that the surface coverage was largely unaffected
by the temperature change (see Supplementary Note 1). free O–D are observed; however, the presence of the InaZ at the
interface results in an extra resonance near 2408 cm−1, probably
related to water molecules interacting with InaZ9R35. N–H
resonances originating from protein amine groups can in
principle also contribute to the spectra. However, as hydrophilic
side chains tend to remain disordered at hydrophobic surfaces
such as the air–water interface36, at most minor contributions are
expected. For PBS, there is also a weak, but discernible signature
of the free O–D resonance near 2655 cm−1. Already at 20 °C, the
intensities of the observed resonances are significantly stronger
when InaZ9R is present, compared to the neat buffer spectrum,
indicating an increased water order. This is often observed for
biological interfaces, such as protein films and lipid monolayers,
and can be explained by the interaction and alignment of water
molecules at the InaZ9R interface37,38. As the buffer temperature
is decreased to 10 °C, and subsequently to 5 °C, the water signal is
strongly increased. Figure 2C summarizes the intensity changes of
the different water modes (see Table S3 for fitting parameters). Potential N–D contributions from the backbone, the so-called
amide-A modes, or from side chains may also contribute to this
spectral region as a narrow peak at ~2550 cm−1 39. Results and discussion
W
h It contains an N-
terminal tag, the membrane-binding N-terminal domain, the ice-active repeat units, and a C-terminal domain of unknown function15,17,61,62. NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunicatio 2 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 Fig. 2 Temperature-dependent OD-stretch SFG spectra collected in the ssp polarization combination, along with fits. For clarity, the 10 °C and 5 °C
spectra are offset from the corresponding 20 °C spectrum by 0.01 and 0.02 arbitrary units, respectively. A Spectra of water molecules interacting with the
ice-nucleating active protein InaZ9R in D2O-based PBS buffer. The intensity of the water signal is increased significantly for lower temperatures in the
presence of inaZ9R. B Spectra of the neat buffer surfaces remain essentially unchanged. C Plot of the SFG amplitudes for the different water modes fitted to
the InaZ9R spectra. Fig. 2 Temperature-dependent OD-stretch SFG spectra collected in the ssp polarization combination, along with fits. For clarity, the 10 °C and 5 °C
spectra are offset from the corresponding 20 °C spectrum by 0.01 and 0.02 arbitrary units, respectively. A Spectra of water molecules interacting with the
ice-nucleating active protein InaZ9R in D2O-based PBS buffer. The intensity of the water signal is increased significantly for lower temperatures in the
presence of inaZ9R. B Spectra of the neat buffer surfaces remain essentially unchanged. C Plot of the SFG amplitudes for the different water modes fitted to
the InaZ9R spectra. temperature”—close to the water melting point at which INPs are
typically active. The similarity of this behavior to what has been
observed for ice bacteria provides strong experimental evidence
that InaZ is indeed the water-ordering agent at the surface of P. syringae cells. free O–D are observed; however, the presence of the InaZ at the
interface results in an extra resonance near 2408 cm−1, probably
related to water molecules interacting with InaZ9R35. N–H
resonances originating from protein amine groups can in
principle also contribute to the spectra. However, as hydrophilic
side chains tend to remain disordered at hydrophobic surfaces
such as the air–water interface36, at most minor contributions are
expected. For PBS, there is also a weak, but discernible signature
of the free O–D resonance near 2655 cm−1. Already at 20 °C, the
intensities of the observed resonances are significantly stronger
when InaZ9R is present, compared to the neat buffer spectrum,
indicating an increased water order. This is often observed for
biological interfaces, such as protein films and lipid monolayers,
and can be explained by the interaction and alignment of water
molecules at the InaZ9R interface37,38. NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 While these sites are typically deprotonated at neutral pH, the pH
and protonation state within the interior of proteins/protein
layers can differ significantly from the pH of the surrounding
solution44–46. Based on literature about infrared absorption
resonances and SFG studies, the feature at ~1630 cm−1 can be
assigned to β-sheet type protein structures47–49 (see Supple-
mentary Note 5 for details about the spectral fits). Since only
ordered structures are visible in SFG spectra, the presence of the
amide-I mode shows that InaZ9R forms a well-aligned layer at
the air–water interface. This is also supported by SFG spectra in
the C–H stretching region, where aliphatic and aromatic modes
are visible, which demonstrates a high degree of alignment within
the hydrophobic side chains (see Supplementary Figure 8/Sup-
plementary Note 5). y
The models are briefly equilibrated using a 100 ns molecular-
dynamics (MD) simulation in GROMACS. The 2D-IR spectra are
calculated with an amide-I Hamiltonian model (see Supplemen-
tary Notes 2 and 6 for details of the simulations and spectral
calculations). To calculate the spectra depicted in Fig. 3B and C,
an additional inhomogeneous broadening was applied to the
local-mode frequencies in order to match the broad experimental
peak shape. This is probably due to the C- and N-termini of the
full InaZ9R sequence used in the experiments (Fig. 1) that are
likely to adopt a less-well defined structure in solution, as is also
indicated by UV-circular dichroism (CD) spectroscopy (Supple-
mentary Figure 5 and Note 3). It is clear that the β-helical model
results in a closer spectral match than the β-sheet model. The
calculated β-sheet model spectra are significantly shifted with
respect to the experimental resonance positions, and exhibit high-
frequency peaks that are absent in the experimental data. The 2D-
IR spectra calculated for the β-helix model, on the other hand,
capture the experimental spectra very well. Minor deviations of
experiment and calculation observed for the β-helix model might
be explained by spectral contributions of coiled structures in the
N- and C-terminal domains that are different from the purely
random-coil structure (here modeled by the included inhomo-
geneous broadening). In summary, we conclude that the β-helix
model agrees with the IR data and describes the solution structure
of InaZ9R well. At 5 °C, the resonance positions remain largely unchanged
while the peak ratio of the 1630 cm−1 protein resonance between
the ssp and ppp spectra is dramatically increased. NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 Fig. 3 Comparison between experimental and calculated FT-IR and 2D-IR spectra of InaZ in solution. A Experimental, and (B, C) calculated spectra, with
the FT-IR spectra shown on top and the 2D-IR spectra below. The IR spectra are sensitive to the secondary structure of proteins41 and can be regarded as a
fingerprint of their conformation. The comparison between the experimental and calculated spectra shows that the β-helix model captures the
experimental data very well, whereas the β-sheet model shows significant deviations. The 2D-IR spectra are recorded with a pump polarization that is
parallel with respect to the probe polarization. Color bar indicates differential absorption (in ΔOD) for all 2D-IR spectra. Fig. 3 Comparison between experimental and calculated FT-IR and 2D-IR spectra of InaZ in solution. A Experimental, and (B, C) calculated spectra, with
the FT-IR spectra shown on top and the 2D-IR spectra below. The IR spectra are sensitive to the secondary structure of proteins41 and can be regarded as a
fingerprint of their conformation. The comparison between the experimental and calculated spectra shows that the β-helix model captures the
experimental data very well, whereas the β-sheet model shows significant deviations. The 2D-IR spectra are recorded with a pump polarization that is
parallel with respect to the probe polarization. Color bar indicates differential absorption (in ΔOD) for all 2D-IR spectra. calculation on the published 48-mer repeat unit composed of
stacked antiparallel β-sheets19. For the β-helix model, we use a
structural model that was proposed in a previous INP simulation
study17. can promote water ordering at low temperatures, we record
amide-I SFG spectra of InaZ9R at the air–water interface. Amide-
I SFG spectra are sensitive to secondary structure, and can also
provide information about the orientation of interfacial pro-
teins30. Fig. 4 displays amide-I SFG spectra collected at 20 °C and
5 °C. The spectra recorded at 20 °C show modes centered near
1630 cm−1 and 1720 cm−1. The 1720 cm−1 mode can be assigned
to lipid molecules, which could indicate the presence of protein-
associated lipids. Both sodium dodecyl-sulfate polyacrylamide gel
electrophoresis and intact-protein mass spectrometry point to an
increased molecular weight of the protein compared with the
expected mass based on its assumed amino-acid sequence (see
Supplementary Note 3). It is also possible that the mode is due
to protonated aspartic acid and glutamic acid side chain modes. NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 The FT-IR and 2D-IR spectra of InaZ9R are composed of a single
broadband that peaks at 1632 cm−1, and the 2D-IR spectrum
does not contain any strong cross-peaks. The spectra show a close
resemblance to previously measured IR spectra of proteins
composed mainly of β-helices, with segments that contain other
secondary structures43. To test which of the theoretical models
agrees best with the experimental data, we calculate 2D-IR spectra
based on both the β-sheet and the β-helix model to compare with
the experimental data (Fig. 3). For the β-sheet model, we base the SFG spectra of the neat PBS buffer surface do not change
appreciably with temperature. This is in agreement with previous
studies
of
non-ice-nucleating
interfaces31,40. Temperature-
dependent SFG experiments with model lipid monolayers, non-
INPs such as lysozyme and lysed ice bacteria have been used to
test whether the increased water order is indeed related to ice
activity. In those reports, an increase of the water signal has only
been observed for ice-active proteins, which showed that the
increased water order at lower temperatures is related to INPs
and not a generic property of aqueous biological interfaces31. For
the INP InaZ9R, we clearly observe effective water ordering that
becomes more efficient at temperatures near its “operating 3 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 In addition, InaZ9R
rotates along the long axis of the helix, such that the ice-active
strands are oriented more parallel to the water surface. As
summarized in Fig. 6, the low-temperature geometry increases the
exposure of INA sites to the interfacial water network when the
protein is part of a densely packed layer. Lateral assembly of InaZ
in the room temperature state (Fig. 6A) will lead to burial of INA
sites within the protein film, while at low temperatures (Fig. 6B)
the protein film exposes a large area of INA sites to the water. Evidently, the reorientation of InaZ at lower temperatures
enhances water ordering and is thereby the probable cause of
the observed increase of SFG water signal at low temperatures51. At cell surfaces, based on current models, InaZ is anchored
through its N-terminal domain17 and would presumably be
flexible enough to reorient from perpendicular to parallel to the
cell surface with decreasing temperatures. Similar to the air–water
interface studied here, InaZ assembles into large patches at the
cell surface. Such a surface assembly maximizes the size of the ice-
nucleation site and thereby increases the ice-nucleation activity. The current picture of the assembly of InaZ at the bacterial
surface with the long protein axis parallel to the surface (Fig. 6C)
is in excellent agreement with the low-temperature pose we
identified with SFG, where the long helix axis is oriented parallel
to the air–water interface. InaZ9R is a model INP with a
significantly shortened ice-active domain. Therefore, the extent of
water interactions may differ from the native INP. Yet, based on
the present data, we can conclude that the reason why water close
to ice-active bacterial cells becomes more ordered when the water
is cooled to lower temperatures is likely not to be found in
changes of INA sites as such, but is driven by a reorientation of
InaZ into a “flat”, carpet-like geometry at a lower temperature,
which maximizes the availability of ice-nucleation sites to water. p
y
y
In summary, the data indicate a reorientation to a larger tilt
angle θ and a Ψ rotation around the helical axis with decreasing
temperature, which changes the orientation of the INA sites
considerably (see Fig. 5C). NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 Fig. 4 Temperature-dependent amide-I SFG spectra of InaZ9R at the air–water interface in ssp and ppp polarization. A Spectra recorded at 20 °C show
a resonance near 1630 cm−1 related to β-sheet type structure and a resonance near 1720 cm−1 related to lipids. B Spectra collected for 5 °C show a strong
increase in amide-I ssp intensity, which indicates the protein has changed orientation at the interface. Fig. 4 Temperature-dependent amide-I SFG spectra of InaZ9R at the air–water interface in ssp and ppp polarization. A Spectra recorded at 20 °C show
a resonance near 1630 cm−1 related to β-sheet type structure and a resonance near 1720 cm−1 related to lipids. B Spectra collected for 5 °C show a strong
increase in amide-I ssp intensity, which indicates the protein has changed orientation at the interface. reproduced well by the calculated spectral response of the central
repeat domain (see Fig. 5). Based on the selection rule of SFG that
only ordered species will generate a signal, this might indicate
that the N- and C-terminal domains adopt a coiled or less defined
structure at the interface and that their contribution to the
measured signal is small. Figure 5a presents an overview of the
comparison between calculated and experimental SFG spectra for
a range of combinations of tilt and twist angles (θ, Ψ) with a step
size of 2.5°, indicated by the residual sum-of-squares (RSS). There
is a narrow range of angles for which theory and experiment
closely match. The closest spectral match is found for small tilt
angles for 20 °C, whereas the minimal RSS values for 5 °C are
found at larger tilt angles. Going from 20 °C to 5 °C, fits that start
at the minimal RSS values result in a transition from (θ, Ψ) =
(21°, 45°) to (59°, 290°) (see Fig. 4B). The calculated spectra for
these orientations capture both the resonance positions and the
relative intensities of the experimental data well. Deviations could
be explained by weak contributions by the terminal domains. provides an overview of the temperature-induced transition of
InaZ9R. Although at room temperature the long axis of the
protein is oriented perpendicular to the water surface, at lower
temperatures the protein reorients and adopts an orientation that
is aligned more parallel with the interface. NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 The fact that
the resonance positions are similar at both temperatures indicates
that the folding of InaZ9R does not change considerably. IR spectra recorded at different temperatures show that the
secondary structure of InaZ9R does not change as a function of
temperature (see Supplementary Figure 8 and Note 4), which
indicates that the observed temperature-dependent water order-
ing (Fig. 2) is caused by another effect. Often, a change in amide-I peak ratios indicates a reorientation
within protein monolayers. We have calculated SFG spectra based
on the IR-derived β-helix model, which encompasses the central
repeat domain of InaZ9R17. The experimental SFG spectra can be InaZ reorientation at low temperatures. To determine the
structure of InaZ9R at the water interface and how the protein NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications 4 ARTICLE NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 A normal-mode analysis of the two
strongest types of modes in the spectrum, the one around 1625
cm−1 and the one at ~1670 cm−1, reveals that these can be
assigned to the B2 mode and to β-turns (see Supplementary
Figure 3 and Supplementary Note 2), in line with previous
literature49. InaZ structure and activation. The IR data provides experi-
mental evidence that the central repeat region of InaZ has a β-
helical fold. In addition, the SFG data support that this con-
formation is also stable in protein films interacting with inter-
facial water. Simulations have predicted a β-helical folding motif
based on sequence homology with insect AFPs17,29. This study
now provides evidence that indeed bacterial INPs and insect
AFPs have a similar β-helical structure, despite having contrary
effects on water molecules. The difference in ice activity is
probably caused by the difference in size of the ice-interacting
surface—insect AFPs have sizes around 10 kilo-Dalton, whereas
INPs have sizes of several mega-Dalton. The reorientation of InaZ9R could be caused by a subtle shift
in the sensitive balance of forces between protein side chains,
water molecules and lateral protein interactions. In principle such
reorientations are independent of ice activity and could occur also
in other protein films. For ice-active proteins these effects could
be more pronounced because studies of INPs and related AFP β-
helices have shown that for this class of proteins all of these
factors are affected by temperature. For example, it has been
shown that their hydration52 and lateral protein interactions53
change significantly when decreasing the temperature close to the g
Earlier studies have reported that the ice activity of INA
bacteria
is
increased
when
the
bacteria
are
exposed
to
temperatures ~4 °C50. The SFG analysis of the interfacial protein
structure shows that this effect can be explained by INP
reorientation when cooled to lower temperatures. Figure 5c 5 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 5 Theoretical and experimental SFG amide-I spectra of InaZ9R at the air–water interface. A RSS plots illustrating the match between theory an
iment for different tilt (θ) and twist (Ψ) angles. B Calculated spectra resulting from (θ, Ψ) fits starting at the minimum RSS position, resulting in bes
hing tilt and twist angles of θ = 21° +31°−40° and Ψ = 41° +72°−110° for 20 °C, and θ = 59° +18° −8° and Ψ = 303° +42°−36° for 5 °C (positive and
ive uncertainties are given as super- and subscripts, see SI). C Schematic representation of the best matching protein orientations. InaZ9R reorients a
temperatures, with a more inclined orientation at 5 °C. In a tightly packed protein layer, the low-temperature pose increases the interaction of th
sites (marked blue) with water. TICLE
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349- Fig. 5 Theoretical and experimental SFG amide-I spectra of InaZ9R at the air–water interface. A RSS plots illustrating the match between theory and
experiment for different tilt (θ) and twist (Ψ) angles. B Calculated spectra resulting from (θ, Ψ) fits starting at the minimum RSS position, resulting in best
matching tilt and twist angles of θ = 21° +31°−40° and Ψ = 41° +72°−110° for 20 °C, and θ = 59° +18° −8° and Ψ = 303° +42°−36° for 5 °C (positive and
negative uncertainties are given as super- and subscripts, see SI). C Schematic representation of the best matching protein orientations. InaZ9R reorients at
lower temperatures, with a more inclined orientation at 5 °C. In a tightly packed protein layer, the low-temperature pose increases the interaction of the
INA sites (marked blue) with water. leads to an increased water order and, consequently, surface
freezing. water melting point. The protein–water interface of INPs has also
been observed to become less dynamic at lower temperatures28. Together, these factors can alter protein–protein as well as
protein–water contacts and impact the lateral organization of
InaZ at interfaces. Vibrational spectroscopy 4. Lindow, S. E., Arny, D. C. & Upper, C. D. Bacterial ice nucleation: a factor in
frost injury to plants. Plant Physiol. 70, 1084–1089 (1982). Sample preparation. The sample solutions for SFG and IR spectroscopy were
prepared using D2O-based buffers. Using D2O is an established strategy in
vibrational protein spectroscopy to prevent overlap of the amide-I modes of the
protein backbone with the OH-bending mode of water. In addition, using D2O is
advantageous in broadband SFG spectroscopy because the water spectrum is
narrower, which facilitates the acquisition of spectra representing the entire water
region in one experiment. D2O forms slightly stronger hydrogen bonds and H2O/
D2O replacement may cause small changes, so the temperature dependencies
observed in our experiments might be slightly shifted in D2O as compared with
H2O55. At the same time, replacing H2O by D2O does not result in any qualitative
differences, and our conclusions will also apply to the case of H2O. 5. Huffman, J. A. et al. High concentrations of biological aerosol particles
and ice nuclei during and after rain. Atmos. Chem. Phys. 13, 6151–6164
(2013). 6. Pöschl, U. et al. Rainforest aerosols as biogenic nuclei of clouds and
precipitation in the Amazon. Science 329, 1513–1516 (2010). 7. Christner, B. C., Morris, C. E., Foreman, C. M., Cai, R. & Sands, D. C. Ubiquity of biological ice nucleators in snowfall. Science 319, 1214 (2008). 8. DeLeon-Rodriguez, N. et al. Microbiome of the upper troposphere: species
composition and prevalence, effects of tropical storms, and atmospheric
implications. Proc. Natl Acad. Sci. USA 110, 2575–2580 (2013). 8. DeLeon-Rodriguez, N. et al. Microbiome of the upper troposphere: species
composition and prevalence, effects of tropical storms, and atmospheric
implications. Proc. Natl Acad. Sci. USA 110, 2575–2580 (2013). 9. Fröhlich-Nowoisky, J. et al. Bioaerosols in the Earth system: climate, health,
and ecosystem interactions. Atmos. Res. 182, 346–376 (2016). p
9. Fröhlich-Nowoisky, J. et al. Bioaerosols in the Earth system: climate, health,
and ecosystem interactions. Atmos. Res. 182, 346–376 (2016). 2D-IR. Transmission 2D-IR spectra were recorded by overlapping a chopped
narrowband pump IR laser pulse and a broadband probe IR laser pulse in the
sample with a delay of 1.5 ps56. An additional reference beam, identical to the
probe, was passed through the sample at a few mm displacement. Code availability Code used to calculate SFG and 2D-IR spectra is available from the authors upon
reasonable request. Received: 22 June 2020; Accepted: 22 January 2021; Spectral calculations 2D-IR. A two-exciton amide-I Hamiltonian was constructed based on the solution-
state MD trajectory at room temperature. The couplings in the Hamiltonian were
based on the transition-dipole coupling model57 for non-nearest neighbors, whose
interaction is dominated by through-space effects, while the nearest neighbor
couplings, dominated by through-bond effects, are modeled with a parameterized
map of an ab initio calculation with the 6-31 G + (d) basis set and B3LYP-
functional58. The local-mode frequencies on the diagonal of the Hamiltonian are
estimated using the same hydrogen bond-shift model as published previously59,
which is based on the effect that various types of hydrogen bonds to the amide
group have on the amide-I frequency60. By diagonalizing the Hamiltonian, we
obtain the amide-I eigenvectors and eigenvalues from which the 2D-IR spectra are
calculated for each of the frames, after which the response is averaged over all
frames. SFG. A one-exciton Hamiltonian was constructed based on the 100 ns-relaxed β-
helix model, using the same coupling models as for the 2D-IR calculations. To
determine the orientation of the protein from the amide-I ssp and ppp lineshapes
and intensities for 5 and 20 °C, we calculated 10,000 spectra as a function of θ and
Ψ (with a grid size of 2.5°) and determined the deviation between calculation and
experiment for each (θ, Ψ) point. Subsequently, Levenberg-Marquardt least-square
fits were performed with the best (θ, Ψ) values as the initial guess. ARTICLE ARTICLE Fig. 6 Illustration of the interaction of InaZ9R with water molecules. A At
room temperature, the ice-nucleating sites (blue arrows, N- and C- terminal
domains marked yellow and pink, respectively) are buried in the protein film
resulting in relatively low water order. B At 5 °C, InaZ9R reorients with the
long axis of the helix more parallel to the surface. In this orientation the ice-
nucleating sites are exposed to the interfacial water layer. Although the
specific water orientation cannot be inferred from homodyned SFG data,
the results clearly show increased water order, which promotes ice
nucleation. C The low-temperature protein pose, with the β-helix (green)
axis of InaZ parallel to the surface, and the N- and C termini again marked
yellow and pink, supports recent models for INP assembly at bacterial
surfaces. SFG. SFG spectra were recorded in standard reflection geometry, by overlapping a
tunable broadband IR laser pulse with a narrowband visible (~800 nm) laser pulse
in space and time. The generated SFG light was routed to a spectrograph and
recorded with an EMCCD camera. All spectra were background subtracted and
subsequently normalized by a spectrum from a neat gold surface. Protein samples
were prepared at 0.44 mg/mL, 10 μM, in phosphate-buffered D2O at pD = 7.4, to
avoid spectral interference from water bending modes. SFG. SFG spectra were recorded in standard reflection geometry, by overlapping a
tunable broadband IR laser pulse with a narrowband visible (~800 nm) laser pulse
in space and time. The generated SFG light was routed to a spectrograph and
recorded with an EMCCD camera. All spectra were background subtracted and
subsequently normalized by a spectrum from a neat gold surface. Protein samples
were prepared at 0.44 mg/mL, 10 μM, in phosphate-buffered D2O at pD = 7.4, to
avoid spectral interference from water bending modes. Data availability All spectroscopic data are available from the authors upon request and on https://
figshare.com under https://doi.org/10.6084/m9.figshare.13372793. Further details about
experiments and data analysis can be found in the Supplementary Information document
(Supplementary Notes 1–6). Fig. 6 Illustration of the interaction of InaZ9R with water molecules. A At
room temperature, the ice-nucleating sites (blue arrows, N- and C- terminal
domains marked yellow and pink, respectively) are buried in the protein film
resulting in relatively low water order. B At 5 °C, InaZ9R reorients with the
long axis of the helix more parallel to the surface. In this orientation the ice-
nucleating sites are exposed to the interfacial water layer. Although the
specific water orientation cannot be inferred from homodyned SFG data,
the results clearly show increased water order, which promotes ice
nucleation. C The low-temperature protein pose, with the β-helix (green)
axis of InaZ parallel to the surface, and the N- and C termini again marked
yellow and pink, supports recent models for INP assembly at bacterial
surfaces. References 1. Murray, B., O’sullivan, D., Atkinson, J. & Webb, M. Ice nucleation by particles
immersed in supercooled cloud droplets. Chem. Soc. Rev. 41, 6519–6554
(2012). fractions were either flash-frozen in liquid nitrogen and stored for later use in the
size exclusion buffer (20 mM Tris-HCl pH 7.5; 150 mM NaCl), or buffer- fractions were either flash-frozen in liquid nitrogen and stored for later use in the
size exclusion buffer (20 mM Tris-HCl pH 7.5; 150 mM NaCl), or buffer-
exchanged into (D2O) PBS buffer for further characterization. Further details on
materials and primers used are provided in Supplementary Note 3. Biological
materials are available from the authors upon reasonable request. fractions were either flash-frozen in liquid nitrogen and stored for later use in the
size exclusion buffer (20 mM Tris-HCl pH 7.5; 150 mM NaCl), or buffer-
exchanged into (D2O) PBS buffer for further characterization. Further details on
materials and primers used are provided in Supplementary Note 3. Biological
materials are available from the authors upon reasonable request. exchanged into (D2O) PBS buffer for further characterization. Further details on
materials and primers used are provided in Supplementary Note 3. Biological
materials are available from the authors upon reasonable request. 2. Białkowska, A., Majewska, E., Olczak, A. & Twarda-Clapa, A. Ice binding
proteins: diverse biological roles and applications in different types of
industry. Biomolecules 10, 274 (2020). 3. Lorv, J. S., Rose, D. R. & Glick, B. R. Bacterial ice crystal controlling proteins. Scientifica 2014, 976895 (2014). NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunicatio Methods Protein purification. The truncated InaZ protein construct (termed InaZ9R),
containing nine repeat units, was prepared following protocols developed for a
related 16 repeat InaZ construct54. The InaZ9R sequence was cloned into the
pET30 Ek/LIC vector (Novagen, Merck Biosciences) according to manufacturer’s
instructions, and used for transformation of E. coli Rosetta (DE3) competent cells. A large-scale culture was grown in lysogeny broth (LB) medium and induced with
1 mM isopropyl β-d-1-thiogalactopyranoside overnight at 20 °C. The protein was
purified by immobilized metal affinity chromatography. Fractions containing
InaZ9R were subsequently loaded on an anion exchange chromatography column
attached to a fast protein liquid chromatography (FPLC) system and finally on a
size exclusion chromatography column attached to an FPLC system. The individual In summary, the presented data provides direct experimental
evidence that InaZ adopts a β-helical fold in solution and in
protein layers in contact with water. Furthermore, the data
supports that InaZ is indeed the driving force for the water
ordering at lower temperatures observed for P. syringae. InaZ
reorients at the interface when cooled from room temperature to
the “operating temperature” of INPs, leading to an increased
exposure of the INA sites. This temperature-induced activation NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications 6 NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 ARTICLE ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-21349-3 13. Voets, I. From ice-binding proteins to bio-inspired antifreeze materials. Soft
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aqueous biological interfaces. Biointerphases 7, 20 (2012). This article is part of a project that has received funding from the European Research
Council (ERC) under the European Union’s Horizon 2020 research and innovation
program (Grant agreement no. 819039 F-BioIce). T.W., S.J.R. and T.Š.T. thank the Villum
Foundation for financial support (Experiment Grants 22956 and 23175). K.F. and T.Š.T. also acknowledge the support by The Danish National Research Foundation (Grant
agreement no.: DNRF106, to the Stellar Astrophysics Centre, Aarhus University). We
thank Sean A. Fischer for support with the spectral calculations, and Rolf Mertig for help
with the Mathematica code used to perform the spectral calculations. We thank Daniel
Otzen for support with CD spectroscopy. S.A. would like to acknowledge funding from
the National Science Foundation Graduate Research Fellowship Program under Grant no. DGE-1762114 to support this research. We thank Søren Jensen and Meilee Ling for help
with the InaZ9R construct. The simulations were done on the Hyak supercomputer
system funded in part by the STF at the University of Washington. We thank Andrey
Kajava and Peter Davies for providing us with structure files for the InaZ models. q
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adhesion at aqueous interfaces. Acc. Chem. Vibrational spectroscopy The difference-
absorption spectrum between the unchopped and chopped pulses was recorded on
a spectrograph with an IR-sensitive MCT (HgCdTe) array, and corrected for peak-
to-peak stability with the reference beam. The 7 μL IR samples (prepared at 0.8 mg/
mL, 18 μM, in D2O PBS buffer at pD = 7.4) were pipetted in between two CaF2
windows that were spaced 50 μm apart and sealed by a greased Teflon spacer. y
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growth, and recrystallization. Acc. Chem. Res. 51, 1082–1091 (2018). 7 TURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications Competing interests Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative
Commons license, and indicate if changes were made. The images or other third party
material in this article are included in the article’s Creative Commons license, unless
indicated otherwise in a credit line to the material. If material is not included in the
article’s Creative Commons license and your intended use is not permitted by statutory
regulation or exceeds the permitted use, you will need to obtain permission directly from
the copyright holder. To view a copy of this license, visit http://creativecommons.org/
licenses/by/4.0/. p
g
The authors declare no competing interests. Author contributions 41. Hamm P., Zanni M. Concepts and Methods of 2D Infrared Spectroscopy. Cambridge University Press (2011). S.J.R. and T.W. have designed the research. S.J.R., T.W.G., and M.B. performed and
analyzed the SFG, IR, and UV-CD experiments. T.D. expressed the protein construct. SA
and JP performed and analyzed the MD simulations. W.R. and G.K. performed and
analyzed the MS experiments. T.S.-T., K.F., and T.B. designed the protein construct. S.J.R. and S.W. performed and designed the spectral calculations. S.J.W. and T.W. wrote
the communication with input from all co-authors. 42. Ganim, Z. et al. Two-dimensional infrared spectroscopy of proteins. Acc. Chem. Res 41, 432–441 (2008). 43. Khurana, R. & Fink, A. L. Do parallel β-helix proteins have a unique Fourier
transform infrared spectrum? Biophys. J. 78, 994–1000 (2000). 8 NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunications Additional information Supplementary information The online version contains supplementary material
available at https://doi.org/10.1038/s41467-021-21349-3. Correspondence and requests for materials should be addressed to T.W. Peer review information Nature Communications thanks Chengcheng Zhang and the
other, anonymous, reviewer(s) for their contribution to the peer review of this work. Reprints and permission information is available at http://www.nature.com/reprints © The Author(s) 2021 Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations. NATURE COMMUNICATIONS | (2021) 12:1183 | https://doi.org/10.1038/s41467-021-21349-3 | www.nature.com/naturecommunication
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Larissa Vital Britto Vinhas¹, Luis Felipe Paixao Batalha Jardim¹, Maísa Raquel
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AO
DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR
EM ÁREA ENDÊMICA DO ESTADO DO MARANHÃO
DURANTE O PERÍODO DE 2011 A 2021
Larissa Vital Britto Vinhas¹, Luis Felipe Paixao Batalha Jardim¹, Maísa Raquel
Guimaraes De Araujo¹ Marcone Douglas Braga Filho¹, Nayana Pers
Marques¹. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS
FATORES
DE
RISCO
RELACIONADOS
AO
DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR
EM ÁREA ENDÊMICA DO ESTADO DO MARANHÃO
DURANTE O PERÍODO DE 2011 A 2021
Larissa Vital Britto Vinhas¹, Luis Felipe Paixao Batalha Jardim¹, Maísa Raquel Larissa Vital Britto Vinhas¹, Luis Felipe Paixao Batalha Jardim¹, Maísa Raquel
Guimaraes De Araujo¹ Marcone Douglas Braga Filho¹, Nayana Pers
Marques¹. Larissa Vital Britto Vinhas¹, Luis Felipe Paixao Batalha Jardim¹, Maísa Raquel
Guimaraes De Araujo¹ Marcone Douglas Braga Filho¹, Nayana Pers
Marques¹. RESUMO A Leishmaniose Tegumentar Americana (LTA) é uma doença causada por protozoários
do gênero Leishmania, transmitida ao homem pela picada das fêmeas do inseto
flebotomíneos. No Brasil existem atualmente 6 espécies de Leishmania responsáveis
pela doença humana, e mais de 200 espécies de flebotomíneos implicados em sua
transmissão. Devido a fatores morfoclimáticos e socioeconômicos, o Brasil apresenta-se
como uma das áreas de maior incidência desta patologia, sobretudo na região
amazônica, sendo responsável por mais de um terço de todas as ocorrências dessa
doença a nível global. Este artigo tem como objetivo analisar o perfil epidemiológico da
Leishmaniose Tegumentar Americana na região Pré-Amazônica Maranhense,
determinando o padrão dessa endemia e avaliando os fatores que que favorecem a
proliferação da LTA. Trata-se de estudo retrospectivo, descritivo e observacional de tipo
analítico ecológico a partir de dados disponíveis na plataforma DataSUS, através da
utilização e tratamento dos dados obtidos pelo filtro Microrregião por município, etnia,
faixa etária, nível de escolaridade e forma clínica de apresentação da afecção. A LTA
possui grande prevalência na região pré-amazônica maranhense com. 9.112 casos
notificados ao longo do decênio analisado. Espera-se contribuir para uma melhor
compreensão do perfil epidemiológico dessa parasitose e assim propor medidas de
profilaxia para evitar o estabelecimento e surgimento de novos casos. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. ABSTRACT American Tegumentary Leishmaniasis (ATL) is a disease caused by protozoa of the genus
Leishmania, transmitted to humans through the bite of female sand flies. In Brazil, there
are currently 6 species of Leishmania responsible for the human disease, and more than
200 species of sandflies implicated in its transmission. Due to morphoclimatic and
socioeconomic factors, Brazil is one of the areas with the highest incidence of this
pathology, especially in the Amazon region, being responsible for more than a third of
all occurrences of this disease globally. To analyze the epidemiological profile of
American Cutaneous Leishmaniasis in the Pre-Amazon region of Maranhão, determining
the pattern of this endemic and evaluating the factors that favor the proliferation of ATL. This is a retrospective, descriptive and observational study of na ecological analytical
type based on data available on the DataSUS platform, through the use and treatment
of data obtained by the Microregion filter by municipality, ethnicity, age group, level of
education and clinical form presentation of the condition. Keywords: Cutaneous Leishmaniasis. Epidemiology. Pre-Amazônia. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135.
Keywords: Cutaneous Leishmaniasis. Epidemiology. Pre Amazônia.
This work is licensed under a Creative Commons Attribution 4.0
International License.
Instituição afiliada – UNIVERSIDADE CEUMA
Dados da publicação: Artigo recebido em 02 de Janeiro e publicado em 12 de Fevereiro de 2024.
DOI: https://doi.org/10.36557/2674-8169.2024v6n2p1122-1135
Autor correspondente: Larissa Vital Britto Vinhas larissa024615@ceuma.com.br Instituição afiliada – UNIVERSIDADE CEUMA
Dados da publicação: Artigo recebido em 02 de Janeiro e publicado em 12 de Fevereiro de 2024. DOI: https://doi.org/10.36557/2674-8169.2024v6n2p1122-1135
Autor correspondente: Larissa Vital Britto Vinhas larissa024615@ceuma.com.br Instituição afiliada – UNIVERSIDADE CEUMA
Dados da publicação: Artigo recebido em 02 de Janeiro e publicado em 12 de Fevereiro de 202
DOI https //doi org/10 36557/2674 8169 2024v6n2p1122 1135 Instituição afiliada – UNIVERSIDADE CEUMA
Dados da publicação: Artigo recebido em 02 de Janeiro e publicado em 12 de Fevereiro de 2024. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. METODOLOGIA Trata-se de um estudo retrospectivo, descritivo e observacional de tipo analítico
ecológico a partir de dados disponíveis na plataforma DataSUS. Para esta pesquisa
foram utilizados dados disponíveis no Sistema Nacional de Notificações e Agravos
(SINAM), vinculado ao Ministério da Saúde disponíveis para consulta pública através do
sítio eletrônico datasus.saude.gov.br referentes ao estado do Maranhão, em especial a
região pré-amazônica que compreende 181 dos 217 municípios do estado com áreas de
confluência entre biomas do Cerrado, Amazônia e Caatinga resultando em vários
ecossistemas diferentes. Foram utilizados para a pesquisa o banco de dados do DATASUS, com os
descritores: municípios/microrregiões, raça, faixa etária, nível de escolaridade, incidência por sexo,
forma clínica de principal apresentação na faixa temporal de 2011 a 2021 para entender
como essa afecção se desdobra e seus principais achados sociodemográficos. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. desta patologia com a Síndrome da Imunodeficiência Adquirida (SIDA). Ademais,
propõe-se identificar quais são os principais fatores de risco para o desenvolvimento da
LTA. desta patologia com a Síndrome da Imunodeficiência Adquirida (SIDA). Ademais,
propõe-se identificar quais são os principais fatores de risco para o desenvolvimento da
LTA. INTRODUÇÃO A Leishmaniose Tegumentar Americana (LTA) é uma doença que possui como
agente etiológico protozoários do gênero Leishmania, subgêneros Leishmania e Viannia,
que tem como espécie e subespécie predominante, no Brasil, a Leishmania braziliensis. Este protozoário pode ser transmitido ao homem, a animais silvestres e domésticos, por
meio da picada do mosquito flebotomíneo fêmea do gênero Lutzomyia que esteja
infectado, durante o repasto sanguíneo [1]. A afecção por LTA é uma enfermidade polimórfica que afeta pele e mucosas,
provocando lesões ulcerosas e indolores, as quais podem ser únicas ou múltiplas [2]. Destacam- se, assim, dois tipos de manifestações: a cutânea e a mucosa ou
mucocutânea. São presentes as seguintes apresentações clínicas: cutânea localizada,
cutânea disseminada, recidiva cútis e cutânea difusa, mucosa tardia, mucosa de origem
indeterminada, mucosa concomitante, mucosa contígua, mucosa primária. Pode, ainda,
ser uma infecção inaparente ou de acometimento linfonodal [3]. Essa enfermidade tem proliferação a nível global, sendo a maioria dos casos
encontrados na Ásia, África e Américas (regiões tropicais e subtropicais), e está presente
em mais de 80 países. Contudo, segundo a Organização Pan-Americana de Saúde [4],
em 2017, só o Brasil foi responsável por 35% das ocorrências dessa doença, o que
demonstra a necessidade de enfoque sobre tal parasitose [5]. A proliferação acentuada no Brasil, sobretudo, na região Amazônica, se dá em
razão de fatores climáticos, históricos e socioeconômicos [6]. Dessa forma, a doença,
que é de notificação compulsória, representa um grave problema de saúde de cunho
nacional, uma vez que foi intensamente negligenciada em razão da vulnerabilidade
social dos infectados, sobretudo no período pandêmico, em razão do enfoque público e
governamental dado à Covid- 19 e o status de esquecimento a outras doenças [7]. Nesse contexto de grande necessidade de implementação de políticas públicas
eficazes ao combate da enfermidade no Brasil, o presente estudo analisa aspectos
clínico-epidemiológicos da LTA na região pré-amazônica do Maranhão, uma vez ser este
o segundo estado com maior número de casos da federação [1]. Para tanto, serão
avaliados fatores ambientais, étnicos, etários e socioambientais, bem como a correlação Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. RESULTADOS A presente pesquisa tem como expectativa somar com a comunidade
acadêmica e científica, com o fito de conscientizar a população acerca do
reconhecimento da LTA como agravo de saúde pública e vistas a redução dos casos de
infecção, uma vez que, por meio deste trabalho será possível identificar, aspectos
epidemiológicos, fatores de risco associados a esta doença bem como diretriz de
tratamento, permitindo assim, a criação de políticas públicas realmente eficazes
direcionadas a região Pré-Amazônica e medidas de orientação que reduzam a
prevalência dos altos índices vistos por essa antropozoonose. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Figura 1: Número de casos Região Pré-Amazônica Maranhense de 2011 a 2021 no estado
no Maranhão. Figura 1: Número de casos Região Pré-Amazônica Maranhense de 2011 a 2021 no estado
no Maranhão. O Gráfico 1 relaciona o número de casos confirmados de Leishmaniose
Tegumentar Americana nas Microrregiões maranhenses, revelando um número total de
casos em 18.959 casos no decênio de 2011 a 2021. Conforme os dados analisados e
registrados na Tabela 1 pode-se concluir que o maior número de casos confirmados se
deu na Microrregião de Pindaré, região pré-amazônica que registrou 4539 casos ao
longo do decênio analisado, correspondendo a 23,94% do total. A Microrregião de Pindaré corresponde aos municípios de Santa Inês, Santa
Luzia, Buriticupu, Zé Doca, Lago da Pedra, Bom Jardim, Bom Jesus das Selvas, Pindaré-
Mirim, Alto Alegre do Pindaré, Vitorino Freire, Santa Luzia do Paruá, Paulo Ramos, Nova
Olinda do Maranhao, São João do Caru, Araguanã, Lagoa Grande do Maranhão,
Governador Newton Bello, Brejo de Areia, Altamira do Maranhão, Marajá do Sena,
Presidente Médici, Tufilândia. RESULTADOS Com resultados expressivos, verifica-se também um alta taxa de notificação por
Leishmaniose Tegumentar Americana – LTA na Microrregião pré-amazônica do Gurupi
com 2.271 casos, correspondendo a 11,9% do total; Imperatriz com 2423 casos
correspondendo a 12,78% do total, Chapadinha com 1.649 casos notificados
correspondendo a 8,69% do total, Alto do Mearim e Grajaú com 1.206 casos notificados
correspondendo a 6,36% do total. Desse modo, salienta-se também a necessidade de
compreensão desses dados em virtude da LTA ser uma afecção que predispõe o
surgimento de outros agravos de saúde pública. Da análise da literatura disponível sobre o tema é possível observar que as Da análise da literatura disponível sobre o tema é possível observar que as Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. regiões maranhenses, em especial as pré-amazônicas possuem um elevado número de
casos confirmados de Leishmaniose Tegumentar Americana, em virtude do seu clima
favorável e da variedade de flebotomíneos presentes na região amazonense que se
mistura ao território maranhense, cujos períodos de chuvas maranhenses propiciam ao
surgimento de várias doenças tropicais, dentre elas a Leishmaniose Tegumentar
Americana [7]. Desse modo, em virtude de o Maranhão possuir altos índices de notificação por
Leishmaniose Tegumentar Americana, e ter apresentado uma breve diminuição ao
longo dos anos, isso não representa, necessariamente, um fator de melhora nos índices
de infecção por Leishmaniose, haja vista os casos conhecidos de subnotificação e, ainda,
da pandemia de covid-19 que alterou as dinâmicas em saúde no mundo todo. Figura 2: Incidência da Leishmaniose Tegumentar Americana segundo etnia de 2011 a 2021
no estado do Maranhão. Figura 2: Incidência da Leishmaniose Tegumentar Americana segundo etnia de 2011 a 2021
no estado do Maranhão. O gráfico 2, demonstra uma maior incidência de Leishmaniose Tegumentar
Americana na população parda, em sequência em indivíduos de etnia branca, seguido
pela raça de pessoas negras, tendo em vista os dados de enfermos de 1 a 4 anos de
idade. De 5 a 9 anos de idade, prepondera maior incidência em pardos, além de haver
uma paridade entre pacientes das raças negra e branca quanto ao número de casos
notificados. De 10 a 14 anos de idade, permanece a prevalência da afecção em pessoas
pardas, contudo a população negra começa a apresentar um significante aumento de
casos em relação a população branca, o que ocorre novamente em indivíduos de 15 a Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Vinhas et. al. 19 anos de idade. Ademais, de 20 a 59 anos de idade, percebe-se um aumento considerável na
quantidade de casos da afecção, de modo que a raça parda permanece de forma
determinante. Todavia, dos 60 anos em diante, existe uma diminuição do número de
afetados pela LTA. Nesse sentido, pode-se perceber também uma maior incidência na
população parda, seguida pela branca e, em sequência, pela raça negra. Faz-se válido frisar que a população indígena e os asiáticos também foram
considerados, estando em 4º e 5º lugar no ranking de incidência desta doença em toda
as idades avaliadas. Em um estudo sobre epidemiologia da LTA no Brasil, concluiu-se que: a
ocorrência da enfermidade acometeu mais indivíduos de cor parda. Contudo, outros
autores afirmam que não se pode admitir que a LTA acomete mais indivíduos de uma
determinada etnia, pois isto pode ser influenciado pela raça predominante na região em
análise, a depender da herança genética dos grupos de indivíduos e da história do
povoamento que existe em cada região. Ademais, não há estudos que indiquem a
existência da resistência de alguma raça ao agente etiológico da LTA, desse modo,
necessita-se de mais estudos quanto às predisposições genéticas para que se possa
afirmar de modo contundente esse padrão da infecção por LTA em determinada etnia
[8,9]. Figura 3: Incidência de infecção por Leishmaniose Tegumentar Americana segundo o nível Figura 3: Incidência de infecção por Leishmaniose Tegumentar Americana segundo o ní Figura 3: Incidência de infecção por Leishmaniose Tegumentar Americana segundo o nível Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. de escolaridade no estado do Maranhão durante o decênio de 2011 a 2021. O Gráfico 3 demonstra que a escolaridade é um fator preponderante na
avaliação do perfil epidemiológico da LTA, haja vista que 64,4% dos indivíduos com casos
notificados de 2011 a 2021 apresentam baixa escolaridade: eram analfabetos ou que
não tinham Ensino Fundamental completo. A categoria mais acometida foi a de
pacientes que não concluíram seus estudos da 1ª a 4ª série Ensino Fundamental (4073
casos), representando 21,4% dos casos totais entre os anos de 2011 e 2021. A partir da análise de outros estudos sobre a epidemiologia da doença, é
possível perceber que cerca de 58,3% de indivíduos acometidos pela LTA no Maranhão
de 2015 a 2017 apresentaram baixa escolaridade, sendo Analfabetos e Ensino
Fundamental Incompleto as categorias mais acometidas pela doença [1,10]. Além disso, um estudo sobre perfil epidemiológico da LTA realizado em Minas
Gerais entre 2007 e 2018 evidenciou padrões semelhantes quanto a incidência da
doença e escolaridade: 51,73% das fichas de notificação eram referentes a indivíduos
com no máximo 4ª série do ensino fundamental completa e a tendência de queda no
número de casos com o aumento da escolaridade. Portanto, o nível de escolaridade é
um fator crucial na avaliação do risco da afecção de LTA. Figura 4: Incidência da Leishmaniose Tegumentar Americana por sexo segundo ano de
notificação de 2011 a 2021 na Região Pré-Amazônica no Estado do Maranhão. Figura 4: Incidência da Leishmaniose Tegumentar Americana por sexo segundo ano de
notificação de 2011 a 2021 na Região Pré-Amazônica no Estado do Maranhão. Figura 4: Incidência da Leishmaniose Tegumentar Americana por sexo segundo ano de
notificação de 2011 a 2021 na Região Pré-Amazônica no Estado do Maranhão. O Gráfico 4 relaciona o número de casos de Leishmaniose Tegumentar
Americana na àrea Pré Amazônica no Estado do Maranhão com a variável sexo, segundo
o ano de notificação, no decênio de 2011 a 2021. Por meio de sua análise, evidenciou- Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Vinhas et. al. se que 13.599 indivíduos do sexo masculino foram acometidos pela LTA neste período,
o que representa 71,7% do número total de casos, enquanto 5.358 foram os casos
notificados em indivíduos do sexo feminino, representando 28,2% do total de casos
(18.959 em números absolutos). Deste modo, observa-se uma incidência de casos de Leishmaniose
Tegumentar Americana bem superior em homens, quando se compara àquela em
mulheres. Em números absolutos, 8.241 homens a mais que mulheres foram
acometidos pela LTA segundo o gráfico, o que acende um importante alerta sobre os
motivos dessa maior vulnerabilidade dos indivíduos do sexo masculino à afecção pela
doença. Embora seja irrefutável que o maior número de casos de LTA se dê em homens,
a justificativa para tanto não o é, segundo os estudos e pesquisas acerca do tema. Dessa
forma, não há qualquer comprovação de que indivíduos do sexo masculino sejam
geneticamente mais suscetíveis ou vulneráveis a esta enfermidade do que mulheres, em
razão do seu sexo. O que se observa, contudo, é que são os homens, jovens e adultos,
em fase reprodutiva, os mais expostos aos fatores de risco para o surgimento da doença. Isto se relaciona, sobretudo, com o tipo de atividade ocupacional realizada por estes
indivíduos, tais como agricultura, pecuária, garimpo, extração de madeira, construção
de estradas, usinas hidrelétricas, instalação de povoados, treinamentos militares, etc. Estas acabam por inseri-los de forma mais intensa, quando se compara às mulheres, em
locais endêmicos, como florestas, por exemplo, em que há maior proliferação dos
flebotomíneos e, por essa razão os indivíduos do sexo masculino tornam-se mais
suscetíveis e são mais acometidos pela doença [3]. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Figura 5: forma clínica por ano de notificação da LTA na região pré-amazônica do estado do
Maranhão de 2011 a 2021. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Figura 5: forma clínica por ano de notificação da LTA na região pré-amazônica do estado do
Maranhão de 2011 a 2021. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. 2011 2012 2013 2014 2015 2016 2017 2018 2019 2020 2021 Total
Mudança de Diagnóstico
6
4
1
1
2
2
1
2
1
4
6
30
Tranferência
8
2
7
7
10
1
0
5
5
4
2
51
Óbito por outra causa
0
0
2
2
1
4
5
3
3
2
2
24
Óbito por LTA
0
0
0
0
0
0
3
0
0
0
0
3
Abandono
9
8
7
8
9
3
9
6
9
3
20
91
Cura
1229 929
570
635
382
217
380
558
478
410
521 6309
Ign/Branco
320
376
399
495
361
117
166
240
99
142
10
2725
0
1000
2000
3000
4000
5000
6000
7000
8000
9000
10000
Evolução dos casos por ano de notificação
Ign/Branco
Cura
Abandono
Óbito por LTA
Óbito por outra causa
Tranferência
Mudança de Diagnóstico AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. 2011 2012 2013 2014 2015 2016 2017 2018 2019 2020 2021 Total
Mudança de Diagnóstico
6
4
1
1
2
2
1
2
1
4
6
30
Tranferência
8
2
7
7
10
1
0
5
5
4
2
51
Óbito por outra causa
0
0
2
2
1
4
5
3
3
2
2
24
Óbito por LTA
0
0
0
0
0
0
3
0
0
0
0
3
Abandono
9
8
7
8
9
3
9
6
9
3
20
91
Cura
1229 929
570
635
382
217
380
558
478
410
521 6309
Ign/Branco
320
376
399
495
361
117
166
240
99
142
10
2725
0
1000
2000
3000
4000
5000
6000
7000
8000
9000
10000
Evolução dos casos por ano de notificação
Ign/Branco
Cura
Abandono
Óbito por LTA
Óbito por outra causa
Tranferência
Mudança de Diagnóstico O gráfico 4, por sua vez, apresenta a relação da evolução dos casos por ano de
notificação, permitindo avaliar os índices de cura e a letalidade da LTA. Nesse sentido, a
avaliação da quantidade de óbitos por LTA no período de 2011 a 2021 com a quantidade
de casos registrados nesse mesmo período – estes apresentados no gráfico 1 – obtém-
se uma letalidade de aproximadamente 0,015%, enquanto cerca de 33,30% dos
pacientes apresentaram cura da doença e 0,13% óbito por outra causa. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Portanto, é
possível percebe que a LTA possui baixa letalidade na região pré-amazônica do
Maranhão e seus índices de cura são favoráveis. Contudo, é válido ressaltar que não há
informações a cerca da evolução de aproximadamente 15% dos casos – ign/branco. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. No Gráfico 5 em que é feita a avaliação da forma clínica por ano de notificação
durante o decênio de 2011 a 2021, é possível observar que a LTA se manifesta
principalmente em sua forma clínica cutânea, tendo o seu maior número de casos
registrados no ano de 2011, 98,28% (1545) dos casos notificados nesse mesmo ano. A
forma mucosa, menos prevalente, apresentou um maior número de casos no ano de
2018, compondo 9,21% (75) dos casos confirmados no referente ano. No total, 96,02%
das notificações realizadas no decênio de 2011 a 2021 apresentaram lesões cutâneas,
enquanto 3,98% manifestaram a forma clínica mucosa. Além disso, é pertinente
ressaltar que após um grande declínio no número de casos de 2014 a 2016, houve uma
estabilização de 2016 a 2017. Dados apresentados pelo Ministério da Saúde inferem que somente cerca de
3% a 5% dos casos de leishmaniose tegumentar americana tendem apresentar a sua
forma clínica mucosa. Ademais, outra análise do perfil epidemiológico da doença na
região pré-amazônica Maranhão apresentou somente 4,4% do total em sua
manifestação cutânea. Desse modo, nota-se uma maior predominância da forma clínica
cutânea em relação à mucosa, diante da congruência de dados apresentados [1, 11]. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. CONSIDERAÇÕES FINAIS A Leishmaniose Tegumentar Americana (LTA) representa um problema de saúde
pública crescente e de considerável repercussão no território maranhense,
apresentando 18.959casos registrados no decênio de 2011 a 2021. Por ser uma doença
com influência de condições ambientais, tal afecção encontra um habitat favorável na
região Pré-Amazônica Maranhense para se disseminar através da variedade de
flebotomíneos e, ainda, pelo clima propício. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. Vinhas et. al. Da análise das microrregiões pré-amazônicas obtém-se um total de 9112 casos
notificados de LTA no decênio estudado. Uma prevalência de mais de 50% dos casos
está relacionada à região pré-amazônica maranhense o que corrobora com os fatores
climáticos e ambientais da área e propicia um melhor habitat para o vetor transmitir a
doença. Além dos fatores biológicos, a questão socioeconômica também merece
destaque. Em virtude do aumento das atividades humanas ao longo da região pré-
amazônica no território maranhense, o padrão de casos notificados permanece
crescente. Tendo em vista as circunstâncias epidemiológicas em relação as pressões
antrópicas produzidas que, aliadas ao baixo nível educacional dos acometidos e às
precárias condições de saúde, favorecem o cenário exposto. A distribuição de casos de LTA apresentou uma ocorrência maior no ano de 2011
com 2919 casos, considerando a avaliação do perfil epidemiológico do ano de 2011 a
2021. Destes, os maiores números de casos registrados foram na microrregião de
Pindaré, que também apresenta o maior número de casos totais durante todo o período. Nesse sentido, também se evidenciou que a forma clínica predominante no Maranhão,
durante esse período, é cutânea. Além disso, o perfil predominante de indivíduos
acometidos pela LTA no Maranhão é: sexo masculino, raça parda, faixa etária entre 20
e 39 anos e com baixa escolaridade (principalmente, analfabetos e pessoa com ensino
fundamental incompleto). AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. https://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegume
ntar_americana_2edicao.pdf. https://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegum
ntar_americana_2edicao.pdf. [4] Leishmanioses: informe epidemiológico das américas [Internet]. México; 2020
[acesso
em:23
out
2022]. Disponível
em:
https://panaftosa.org/leish/inf2020_es/info_mex_2020_esp.pdf. [4] Leishmanioses: informe epidemiológico das américas [Internet]. México; 2020
[acesso
em:23
out
2022]. Disponível
em:
https://panaftosa.org/leish/inf2020_es/info_mex_2020_esp.pdf. [4] Leishmanioses: informe epidemiológico das américas [Internet]. México; 2020
[acesso
em:23
out
2022]. Disponível
em:
https://panaftosa.org/leish/inf2020_es/info_mex_2020_esp.pdf. [5] Abraão LS, José BM, Gomes CB, Nunes PC, Santos DR, Varela AP, et al. Perfil
epidemiológico dos casos de Leishmaniose Tegumentar Americana no estado do Pará,
Brasil, entre 2008 e 2017. Rev Pan-Amaz Saúde [Internet]. 2020 [acesso em: 23 out
2022];
11:
e202000612. Disponível
em:
http://scielo.iec.gov.br/scielo.php?script=sci_arttext&pid=S217662232020000100022
&lng=pt. Epub 18-Nov-2020. http://dx.doi.org/10.5123/s21766223202000612. [5] Abraão LS, José BM, Gomes CB, Nunes PC, Santos DR, Varela AP, et al. Perfil
epidemiológico dos casos de Leishmaniose Tegumentar Americana no estado do Pará,
Brasil, entre 2008 e 2017. Rev Pan-Amaz Saúde [Internet]. 2020 [acesso em: 23 out
2022];
11:
e202000612. Disponível
em:
http://scielo.iec.gov.br/scielo.php?script=sci_arttext&pid=S217662232020000100022
&lng=pt. Epub 18-Nov-2020. http://dx.doi.org/10.5123/s21766223202000612. [6] Sabroza PC, Toledo LM, Osanai CH. A organização do espaço e os processos
endêmico-epidêmicos. 3. ed. São Paulo: Hucitec; Abrasco; 1992. 55-77 p. [7] Lustosa FD, Minuzzo EA, Nunes AL, Sá LC, Filho HF, Pinto AR, et al. Perfil clínico-
epidemiológico da Leishmaniose Tegumentar em Redenção, Pará [Internet]. Pará:
TheBrazilian Journal Of Infectious Diseases; 22 jan 2022 [acesso em: 23 out 2022]. Disponível em: http://dx.doi.org/10.1016/j.bjid.2021.101971. [7] Lustosa FD, Minuzzo EA, Nunes AL, Sá LC, Filho HF, Pinto AR, et al. Perfil clínico-
epidemiológico da Leishmaniose Tegumentar em Redenção, Pará [Internet]. Pará:
TheBrazilian Journal Of Infectious Diseases; 22 jan 2022 [acesso em: 23 out 2022]. Disponível em: http://dx.doi.org/10.1016/j.bjid.2021.101971. [8] Vasconcelos JM, Gomes CG, Sousa A, Teixeira AB, Lima JM. American integumentary
leishmaniasis: epidemiological profile, diagnosis and treatment. Revista Brasileira de
Análises Clínicas [Internet]. 2018;50(3). Available from: http://www.rbac.org.br/wp-
content/uploads/2019/01/RBAC-vol-50-3-2018-ref-722-final.pdf REFERÊNCIAS [1] Perfil epidemiológico dos casos de Leishmaniose Tegumentar Americana no estado
do Maranhão no período de 2015 a 2017 [Internet]. Maranhão; 2018 [acesso em: 23 out
2022]. Disponível em: http://dx.doi.org/10.24863/rib.v10i3.340. [2] Neves DP, Melo AL, Lenardi PM, Vitor RW. Parasitologia humana. 13. ed. São Paulo:
Atheneu; 2016. [3] Ministério da Saúde. Manual de Vigilância da Leishmaniose Tegumentar Americana
[Internet]. Brasília: Editora MS; 2007 [acesso em: 23 out 2022]. Disponível em: Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135. AVALIAÇÃO DO PERFIL CLÍNICO-EPIDEMIOLÓGICO E DOS FATORES DE RISCO
RELACIONADOS AO DESENVOLVIMENTO DA LEISHMANIOSE TEGUMENTAR EM ÁREA
ENDÊMICA DO ESTADO DO MARANHÃO DURANTE O PERÍODO DE 2011 A 2021
Vinhas et. al. https://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegume
ntar_americana_2edicao.pdf. [4] Leishmanioses: informe epidemiológico das américas [Internet]. México; 2020
[acesso
em:23
out
2022]. Disponível
em:
https://panaftosa.org/leish/inf2020_es/info_mex_2020_esp.pdf. [5] Abraão LS, José BM, Gomes CB, Nunes PC, Santos DR, Varela AP, et al. Perfil
epidemiológico dos casos de Leishmaniose Tegumentar Americana no estado do Pará,
Brasil, entre 2008 e 2017. Rev Pan-Amaz Saúde [Internet]. 2020 [acesso em: 23 out
2022];
11:
e202000612. Disponível
em:
http://scielo.iec.gov.br/scielo.php?script=sci_arttext&pid=S217662232020000100022
&lng=pt. Epub 18-Nov-2020. http://dx.doi.org/10.5123/s21766223202000612. [6] Sabroza PC, Toledo LM, Osanai CH. A organização do espaço e os processos
endêmico-epidêmicos. 3. ed. São Paulo: Hucitec; Abrasco; 1992. 55-77 p. [7] Lustosa FD, Minuzzo EA, Nunes AL, Sá LC, Filho HF, Pinto AR, et al. Perfil clínico-
epidemiológico da Leishmaniose Tegumentar em Redenção, Pará [Internet]. Pará:
TheBrazilian Journal Of Infectious Diseases; 22 jan 2022 [acesso em: 23 out 2022]. Disponível em: http://dx.doi.org/10.1016/j.bjid.2021.101971. [8] Vasconcelos JM, Gomes CG, Sousa A, Teixeira AB, Lima JM. American integumentary
leishmaniasis: epidemiological profile, diagnosis and treatment. Revista Brasileira de
Análises Clínicas [Internet]. 2018;50(3). Available from: http://www.rbac.org.br/wp-
content/uploads/2019/01/RBAC-vol-50-3-2018-ref-722-final.pdf
[9] Neves T. EPIDEMIOLOGIA DA LEISHMANIOSE TEGUMENTAR AMERICANA NO BRASIL. [UniCEUB]; 2017. [10] Vi
AG S
FV d
P
l AMB d
Sil
i
MF B
lh AC d C A
lí i [9] Neves T. EPIDEMIOLOGIA DA LEISHMANIOSE TEGUMENTAR AMERICANA NO BRASIL.
[UniCEUB]; 2017. [10] Viana AG, Souza FV de, Paula AMB de, Silveira MF, Botelho AC de C. Aspectos clínico-
epidemiológicos da leishmaniose tegumentar americana em Montes Claros, Minas
Gerais. wwwrmmgorg [Internet]. 2012 [cited 2023 May 30];22(1):1–128.Disponível em:
https://www.rmmg.org/artigo/detalhes/125 [11] Rocha Thiago José Matos, Barbosa Ana Clara André, Santana Elizabete Priscila Costa,
Calheiros Cláudia Maria Lins. Aspectos epidemiológicos dos casos humanos confirmados
de leishmaniose tegumentar americana no Estado de Alagoas, Brasil. Rev Pan-Amaz
Saude [Internet]. 2015 Dez [citado 2023 Maio 26];6(4):49-54. Disponível
em:http://scielo.iec.gov.br/scielo.php?script=sci_arttext&pid=S2176622320150004000
07&lng=pt. Brazilian Journal of Implantology and Health Sciences
Volume 6, Issue 2 (2024), Page 1122-1135.
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https://www.degruyter.com/document/doi/10.1515/ohe-2022-0031/pdf
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English
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World Heart Day: Clinical case to raise awareness on cardiovascular disease in women
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Open Health
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cc-by
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2 Objective In the context of World Heart Day, which is marked on 29
September of each year, and in line with the WHF aware-
ness campaign with the theme USE HEART FOR EVERY
HEART [5], we hereby use heart for every heart and
spread awareness on CVD in women. The sharing of
this clinical case about a merry episode of drinking and
singing during a social gathering has the objective to
sensitize health professionals and the public in general
about CVD in women. Keywords: cardiovascular disease, women, vulnerability,
awareness, transient ischemic attack, World Heart Day Short Communication https://doi.org/10.1515/ohe-2022-0031
received September 28, 2022; accepted January 22, 2023 contraceptive use, and postmenopausal hormone therapy
[3]. Moreover, women who develop stroke more commonly
have atrial fibrillation, hypertension, and migraine, while
men with stroke are more likely to have coronary artery
and peripheral vascular disease [3]. In light of the female-
specific risk factors and/or predisposing conditions, there
is a need to change the paradigm of underdiagnosing and
undertreating CVD in women [4]. contraceptive use, and postmenopausal hormone therapy
[3]. Moreover, women who develop stroke more commonly
have atrial fibrillation, hypertension, and migraine, while
men with stroke are more likely to have coronary artery
and peripheral vascular disease [3]. In light of the female-
specific risk factors and/or predisposing conditions, there
is a need to change the paradigm of underdiagnosing and
undertreating CVD in women [4]. Abstract: The World Heart Federation highlights that car-
diovascular disease (CVD) is the leading cause of death in
women worldwide. Indeed, women have unique female-
specific risk factors for stroke, which include pregnancy,
history of preeclampsia/eclampsia or gestational dia-
betes, duration of reproductive life, oral contraceptive
use, and postmenopausal hormone therapy. In light of
the female-specific risk factors and/or predisposing con-
ditions, there is need to sensitize health professionals
and the public in general on CVD in women. This clinical
case is being shared with the written consent of the
patient so as to avoid misdiagnosis or underdiagnosis
of CVD among women. Open Health 2023; 4: 20220031 This work is licensed under the Creative Commons Attribution 4.0 International Open Access. © 2023 the author(s), published by De Gruyter.
This work is licensed under the Creative Commons Attribution 4.0 International
License. ), published by De Gruyter.
This work is licensed under the Creative Commons Attribution 4.0 International 5 Discussion TIA diagnosis constitutes a challenge for physicians [16]. In this particular case, its somehow non-conventional
clinical presentation was masked by the “chameleon”
context of wedding celebration with binge drinking. The
context lent support to attribute the symptoms to the con-
sumption of alcoholic drinks. However, there is a need to
consider the fact that binge drinking is associated with a
higher risk of CVD [17,18]. Findings by Piano et al. support
the notion that binge drinking is associated with a higher
risk of stroke, myocardial infarction, and hypertension
among middle-aged and older adults [17]. The possible
mechanism of action, though not completely understood,
is that binge drinking induces oxidative stress and vas-
cular injury while being atherogenic [17]. Public health
awareness campaigns regarding binge drinking need to
include the effects of binge drinking on the cardiovascular
system [17]. A systematic review by Tasnim et al. showed
that within 6 h of consumption, high-dose alcohol (>30 g)
decreased systolic blood pressure by 3.5 mmHg, decreased
diastolic blood pressure by 1.9 mmHg, and increased heart
rate by 5.8 beats per minute. With reference to Tasnim
et al., the highly probable drop in blood pressure that
affected cerebral circulation was the TIA mechanism [19]. In addition, the INTERSTROKE Study highlighted alcohol
intake as a risk factor for acute stroke: a high level of
alcohol intake was consistently associated with all stroke
types, ischemic stroke, and intracerebral hemorrhage [20]. The diagnosis of a Transient Ischemic Attack (TIA),
which constitutes sudden neurologic dysfunctions caused
by transient brain ischemia, was clinically put forward. Nonetheless, the investigations undertaken were revealed
to be negative. This finding is in line with Madsen et al.,
who put forward that women admitted for suspected TIA
are less likely to have imaging evidence of infarction than
men [9]. Indeed, TIA represents a considerable risk for a
subsequent ischemic stroke [10] to occur within the first
weeks after a TIA [11]. An estimated 7.5–17.4% of patients
with TIA will have a stroke in the next 3 months [12]. The risk of ischemic stroke following a TIA is directly
related to its etiopathogenesis, which is commonly cate-
gorized within one of three domains: cardiac dysfunction,
cerebrovascular pathology, or hematologic disorder [10]. Patients with TIA, therefore, require urgent evaluation and
treatment that has the potential to significantly reduce the
risk of recurrent stroke [10,11]. 4 Decision-making sequential cardiac monitoring [15]. Atrial fibrillation was
not detected in more than three-quarters of patients with
stroke or TIA [15]. As a precautionary measure, the family decided to go
to the nearest clinic to monitor the situation overnight. The patient was known to have episodic migraine and
had hypertensive problems during her second pregnancy. Upon admission, the specialist doctor performed a general
clinical examination, including a neurological assessment,
which showed the impossibility to raise the left arm. The patient was clinically alert with spontaneous opening
of the eyes, normal temporospatial orientation, and an
appropriate response to instructions by the examiner. The Barré and Mingazzini tests were used to look for any
subtle hemiparesis of the arm and leg [6]. While the Glasgow
Coma Scale is commonly used to objectively describe the
extent of impaired consciousness in all types of acute medical
and trauma patients [7], the National Institutes of Health
Stroke Scale is recommended as the gold standard [8]. 3 Clinical case The World Heart Federation (WHF) highlights that cardi-
ovascular disease (CVD) is the leading cause of death in
women worldwide and that women with CVD continue to
be underdiagnosed and undertreated due to misconcep-
tions and lack of awareness among both patients and
doctors [1]. The American Heart Association (2004) cre-
ated Go Red for Women, a social initiative designed to
empower women to take charge of their heart health,
by raising awareness of heart disease and stroke being
the number one killer of women [2]. Indeed, women
have unique female-specific risk factors for stroke, which
include pregnancy, history of preeclampsia/eclampsia or
gestational diabetes, duration of reproductive life, oral After a merry singing episode at a wedding ceremony in a
hotel setting, a 40-year-old woman felt unwell with
nausea. She had drunk five glasses of cocktail (with
alcohol) between 4 and 6 pm. She was thus accompanied
to the washroom. It seemed that she was having the
counter side of drinking on an empty stomach. After
vomiting, she felt weak and could not walk. She was
helped by two persons to lie down on a nearby mattress. Lying down on her left side, she looked grey. She com-
plained of having difficulty in breathing and was seen to
gasp intermittently. After some minutes, she looked slightly
better and managed to open her eyes. At one point, she
managed to move both hands by pointing her left and right
index fingers. The general medicine doctor on call who
arrived at 9 pm found the heartbeat was regular with
normal pulmonary auscultation, the blood pressure was
10/6, and the glucometer test was 6.1 mmol/L. * Corresponding author: Marie Chan Sun, World Heart Federation,
Geneva, Switzerland; Department of Medicine, Faculty of Medicine
and Health Sciences, University of Mauritius, 80837 Réduit,
Mauritius, e-mail: lan.sun@uom.ac.mu * Corresponding author: Marie Chan Sun, World Heart Federation,
Geneva, Switzerland; Department of Medicine, Faculty of Medicine
and Health Sciences, University of Mauritius, 80837 Réduit,
Mauritius, e-mail: lan.sun@uom.ac.mu 2
Marie Chan Sun References among women in the world [24], the need for examina-
tion of sex-specific risk factors is being highlighted for
timely diagnosis and proper treatment of women pre-
senting with CVD. [1]
World Heart Federation. Women & CVD [Internet]. World Heart
Federation; 2022 [cited 2022 Dec 28]. https://world-heart-
federation.org/what-we-do/women-cvd/. With the view to increase awareness of CVDs, we
highlight the public education message bearing the mne-
monic FAST, for Face droop, Arm weakness, Speech
difficulties and Time, which consists of three common
stroke warning signs (Face droop, Arm weakness, and
Speech problem) and one action (Time to dial emergency
number) in case symptoms occur [25]. The subsequent
stroke-screening scale, BE-FAST, acronym for Balance,
Eyes, Face, Arm, Speech, Time, was developed with the
inclusion of two additional criteria (Balance and Eye) to
enhance stroke detection [26]. Nonetheless, BE-FAST
and FAST scale accuracy for stroke identification was
comparable [27]. In this clinical case, the fact that the
person required help for walking was considered a weak-
ness in relation to the lack of food intake prior to the
alcohol intake and the subsequent vomiting. Indeed, FAST
public awareness campaigns should be undertaken to sen-
sitize the general population on the need to recognize stroke
signs for prompt diagnosis and medical intervention to
improve prognosis. [2]
American Heart Association. Go Red for Women [Internet]. www.goredforwomen.org. 2004 [cited 2022 Dec 28]. https://
www.goredforwomen.org/en/about-go-red-for-women. [3]
McGrath ER, Rexrode KM. Stroke risk factors in women. In:
O’Neal MA, editor. Neurology and Psychiatry of Women: A
Guide to Gender-based Issues in Evaluation, Diagnosis, and
Treatment [Internet]. Cham: Springer International Publishing;
2019 [cited 2022 Dec 26]. p. 205–11. doi: 10.1007/978-3-030-
04245-5_23. [3]
McGrath ER, Rexrode KM. Stroke risk factors in women. In:
O’Neal MA, editor. Neurology and Psychiatry of Women: A
Guide to Gender-based Issues in Evaluation, Diagnosis, and
Treatment [Internet]. Cham: Springer International Publishing;
2019 [cited 2022 Dec 26]. p. 205–11. doi: 10.1007/978-3-030-
04245-5_23. [4]
Agarwala A, Michos ED, Samad Z, Ballantyne CM, Virani SS. The use of sex-specific factors in the assessment of
women’s cardiovascular risk. Circulation. 2020 Feb
18;141(7):592–9. [5]
World Heart Federation. World Heart Day 2022 [Internet]. World Heart Day. 2022 [cited 2022 Dec 28]. https://world-
heart-federation.org/world-heart-day/about-whd/world-
heart-day-2022/. [6]
Hirose G. The Barrés test and Mingazzini test: Importance of
the original paper by Giovanni Mingazzini. Rinsho
Shinkeigaku. 2015;55(7):455–8. [7]
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Emerg Med. 1999 Apr 1;33(4):373–8. 5 Discussion Irrespective of the setting and the context, there is
a need to look for cardiovascular risk factors specific to
women during the history-taking of female patients. The
pertinent risk factor in this clinical case was the notion of
preeclampsia during the second pregnancy and the pre-
sence of migraine in episodes. Indeed, preeclampsia is a
sex-specific risk factor for future stroke that is likely
under-recognized [21]. As a matter of fact, women with
a history of preeclampsia have an 80% increased risk of
future ischemic stroke than their counterparts without
any such obstetrical history [21]. In addition, physicians
have to inquire about any history of infertility, which is
considered a potential sex-specific risk factor for cardio-
vascular events, despite the fact that the body of evidence
on infertility and CVD is inconclusive to date [22,23]. Nonetheless, with CVD being the leading cause of death In this clinical case, the initial evaluation and man-
agement of the patient as well as measures for the pre-
vention of stroke in this TIA patient were implemented as
per prescribed guidelines [13,14]. Brain imaging, namely
Computed Tomography Scan and Magnetic Resonance
Imaging, as an essential part of the diagnostic approach,
was performed within 24 h. Etiological investigations,
namely Holter electrocardiogram and carotid ultrasono-
graphy, were undertaken, in addition to blood analysis
and blood pressure monitoring. Screening for atrial fibril-
lation and carotid stenosis among other possible causes
turned out to be negative. This is in line with the sys-
tematic review by Sposato et al., who showed the overall
proportion of patients diagnosed with poststroke atrial
fibrillation was found to be 23.7% after all phases of 6 Conclusion [9]
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less likely to have imaging evidence of infarction than men:
Results from the Rhode Island Hospital Prospective TIA unit
cohort. Stroke. 2019;50(Suppl_1):ATP294. The clinical diagnosis was a TIA identified by a specialist
doctor at the clinic where the patient was admitted. The
emphasis is laid on the fact that there is a need for both
specialist and non-specialist doctors to search for sex-
specific cardiovascular risk factors and to exclude in
the first line a CVD irrespective of age, gender, and social
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advances in the management of transient ischemic attacks. Fac Rev. 2022;11:19. [11]
Lamme J, Sauvin M, Sztajzel R. Management of transient
ischemic attack. Rev Med Suisse. 2022 Sep 7;18(794):1671–6. [12] Mendelson SJ, Prabhakaran S. Diagnosis and management of
transient ischemic attack and acute ischemic stroke: a review. JAMA. 2021 Mar 16;325(11):1088–98. Acknowledgments: I am grateful to the World Heart
Federation for the support and training provided. [13] Bushnell C, McCullough LD, Awad IA, Chireau MV, Fedder WN,
Furie KL, et al. Guidelines for the prevention of stroke in
women: a statement for healthcare professionals from the
American Heart Association/American Stroke Association. Stroke. 2014 May;45(5):1545–88. Funding information: The author states no funding
involved. [14] Kernan WN, Ovbiagele B, Black HR, Bravata DM, Chimowitz MI,
Ezekowitz MD, et al. Guidelines for the prevention of stroke in
patients with stroke and transient ischemic attack: a guideline
for healthcare professionals from the American Heart
Association/American Stroke Association. Stroke. 2014
Jul;45(7):2160–236. Conflict of interest: The author states no conflict of
interest. Data availability statement: Data sharing is not applic-
able to this article as no datasets were generated or ana-
lyzed during the current study. [15] Sposato LA, Cipriano LE, Saposnik G, Ruíz Vargas E, Riccio PM,
Hachinski V. Diagnosis of atrial fibrillation after stroke and 4
Marie Chan Sun 4 encephalopathy syndrome and stroke. Stroke. 2018
Mar;49(3):524–30. transient ischaemic attack: a systematic review and meta-
analysis. Lancet Neurol. 2015 Apr;14(4):377–87. [16] Nadarajan V, Perry RJ, Johnson J, Werring DJ. Transient
ischaemic attacks: mimics and chameleons. Practical Neurol. 2014 Feb 1;14(1):23–31. [22] Gleason JL, Shenassa ED, Thoma ME. Self-reported infertility,
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implications for advocacy, policy, and research. Alcohol Clin
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Metcalfe A. Infertility and the risk of cardiovascular disease:
Findings from the study of women’s health across the nation
(SWAN). CJC Open. 2021 Apr;3(4):400–8. [18] Radcliffe S. Binge Drinking: What It Does to Your Body
[Internet]. Healthline; 2018 [cited 2022 Dec 28]. https://www. healthline.com/health-news/what-happens-to-your-body-
when-you-binge-drink. [24] Roth GA, Johnson C, Abajobir A, Abd-Allah F, Abera SF, Abyu G,
et al. Global, regional, and national burden of cardiovascular
diseases for 10 causes, 1990 to 2015. J Am Coll Cardiology. 2017 Jul 4;70(1):1–25. [19] Tasnim S, Tang C, Musini VM, Wright JM. Effect of alcohol on
blood pressure. Cochrane Database Syst Rev. 2020 Jul
1;7(7):CD012787. [25] Kleindorfer DO, Miller R, Moomaw CJ, Alwell K, Broderick JP,
Khoury J, et al. Designing a message for public education
regarding stroke. Stroke. 2007 Oct;38(10):2864–8. [20] Smyth A, O’Donnell M, Rangarajan S, Hankey GJ,
Oveisgharan S, Canavan M, et al. Alcohol intake as a risk
factor for acute stroke: The INTERSTROKE study. Neurol
[Internet]. 2022 Oct 11 [cited 2022 Dec 26]. https://n. neurology.org/content/early/2022/10/11/WNL. 0000000000201388. [26] Tan H, Alexander A, Gopalan A, Hannon C, Gunaga S, Patel J,
et al. The Efficacy Of BE-FAST In Identifying Strokes; 2019. https://scholarlycommons.henryford.com/merf2019clinres/17. [27] Pickham D, Valdez A, Demeestere J, Lemmens R, Diaz L,
Hopper S, et al. Prognostic value of BEFAST vs FAST to identify
stroke in a prehospital setting. Prehospital Emerg Care. 2019
Mar 4;23(2):195–200. [21] McDermott M, Miller EC, Rundek T, Hurn PD, Bushnell CD. Preeclampsia: association with posterior reversible
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Effects of Exhaustive Aerobic Exercise on Tryptophan-Kynurenine Metabolism in Trained Athletes
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RESEARCH ARTICLE a11111 Effects of Exhaustive Aerobic Exercise on
Tryptophan-Kynurenine Metabolism in
Trained Athletes Barbara Strasser1, Daniela Geiger2, Markus Schauer2, Hannes Gatterer3,
Martin Burtscher3, Dietmar Fuchs2* Barbara Strasser1, Daniela Geiger2, Markus Schauer2, Hannes Gatterer3,
Martin Burtscher3, Dietmar Fuchs2* 1 Medical University Innsbruck, Division of Medical Biochemistry, Biocenter, Innsbruck, Austria, 2 Medical
University Innsbruck, Division of Biological Chemistry, Biocenter, Innsbruck, Austria, 3 Department of Sport
Science, Medical Section, University of Innsbruck, Innsbruck, Austria * dietmar.fuchs@i-med.ac.at * dietmar.fuchs@i-med.ac.at * dietmar.fuchs@i-med.ac.at Abstract a1111 Exhaustive exercise can cause a transient depression of immune function. Data indicate
significant effects of immune activation cascades on the biochemistry of monoamines and
amino acids such as tryptophan. Tryptophan can be metabolized through different path-
ways, a major route being the kynurenine pathway, which is often systemically up-regu-
lated when the immune response is activated. The present study was undertaken to
examine the effect of exhaustive aerobic exercise on biomarkers of immune activation
and tryptophan metabolism in trained athletes. After a standardized breakfast 2 h prior to
exercise, 33 trained athletes (17 women, 16 men) performed an incremental cycle ergom-
eter exercise test at 60 rpm until exhaustion. After a 20 min rest phase, the participants
performed a 20 min maximal time-trial on a cycle ergometer (RBM Cyclus 2, Germany). During the test, cyclists were strongly encouraged to choose a maximal pedalling rate that
could be maintained for the respective test duration. Serum concentrations of amino acids
tryptophan, kynurenine, phenylalanine, and tyrosine were determined by HPLC and
immune system biomarker neopterin by ELISA at rest and immediately post exercise. Intense exercise was associated with a strong increase in neopterin concentrations
(p<0.001), indicating increased immune activation following intense exercise. Exhaustive
exercise significantly reduced tryptophan concentrations by 12% (p<0.001) and increased
kynurenine levels by 6% (p = 0.022). Also phenylalanine to tyrosine ratios were lower after
exercise as compared with baseline (p<0.001). The kynurenine to tryptophan ratio corre-
lated with neopterin (r = 0.560, p<0.01). Thus, increased tryptophan catabolism by indo-
leamine 2,3-dioxygenase appears likely. Peak oxygen uptake correlated with baseline
tryptophan and kynurenine concentrations (r = 0.562 and r = 0.511, respectively, both
p<0.01). Findings demonstrate that exhaustive aerobic exercise is associated with
increased immune activation and alterations in monoamine metabolism in trained athletes
which may play a role in the regulation of mood and cognitive processes. OPEN ACCESS Citation: Strasser B, Geiger D, Schauer M, Gatterer
H, Burtscher M, Fuchs D (2016) Effects of Exhaustive
Aerobic Exercise on Tryptophan-Kynurenine
Metabolism in Trained Athletes. PLoS ONE 11(4):
e0153617. doi:10.1371/journal.pone.0153617 Introduction Exhaustive exercise has been associated with a transient depression of immune function [1]. Already moderate physical activity significantly impacts on inflammation cascades that involve
several pro-inflammatory cytokines like interferon-y (IFN-y) and down-stream biochemical
pathways [2, 3]. Recently, two studies reported detailed insights into the physiological mecha-
nisms involved under extreme conditions [4, 5]. One study noted a significant increase of the
reactive oxygen species (ROS) production rate and oxidative damage after a mountain ultra-
marathon running [4]. This includes also changes in the concentrations of urinary neopterin,
which is predominantly produced by human monocytes/macrophages, and elevation of which
is often linked with conditions of immune activation and inflammation [6]. A second study
revealed changes in serum free amino acids during a half-ironman triathlon [5]. Further, data indicates significant effects of immune activation cascades on the biochemistry
of monoamines such as essential amino acid tryptophan (TRP), the precursor molecule of
5-hydroxy-tryptamine (5HT, serotonin) [7]. Serotonin plays a key role in signal transduction
between neurons, and exercise-induced changes in serotonin concentrations have been linked
to central fatigue [8]. However, TRP is not only precursor of the serotonin biochemical path-
way which is conducted by enzyme tryptophan 5-hydroxylase but is also the key element for
the formation of nicotinamide-adenine-dinucleotides NAD and NADH via the so-called
kynurenine (KYN) pathway [9]. The latter is either achieved by hepatic tryptophan 2,3-dioxy-
genase (TDO) or by indoleamine 2,3-dioxygenases 1 and 2 (IDO1 and IDO2). TDO is con-
trolled by TRP levels and can be also induced by steroid hormones like cortisol [10]. However,
pro-inflammatory stimuli like Th1-type cytokine IFN-y strongly induce IDO1 expression and
activity during cellular immune response, and accelerated TRP breakdown is indicated by an
increased KYN to TRP ratio (KYN/TRP) [11]. IFN-y is the most important stimulus of indoleamine 2,3-dioxygenase-1 (IDO1) activity
although other mainly pro-inflammatory stimuli can do the same [7]. In long distance runners,
significantly increased production of IFN-y and also other relevant cytokines was reported in urine
[2]. In this early study, IFN-y remained undetectable in serum, most probably because of lacking
sensitivities of available assays. However, the parallel increase of KYN/TRP and neopterin concen-
trations in our study further supports the involvement of IFN-y. Moreover, exercise increased
IDO1 activity of macrophages was reported in rats [12]. Editor: Gilles J. Guillemin, Macquarie University,
AUSTRALIA Editor: Gilles J. Guillemin, Macquarie University,
AUSTRALIA Editor: Gilles J. Guillemin, Macquarie University,
AUSTRALIA Copyright: © 2016 Strasser et al. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any
medium, provided the original author and source are
credited. Data Availability Statement: All relevant data are
within the paper and its Supporting Information files. Funding: The authors have no support or funding to
report. Competing Interests: The authors have declared
that no competing interests exist. Competing Interests: The authors have declared
that no competing interests exist. PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 1 / 10 Tryptophan-Kynurenine Metabolism to Intense Exercise PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Subjects Thirty-three healthy and trained athletes volunteered to participate in this study. Participants
were excluded if they had a previous history of muscle disorder, cardiac or kidney disease or
those taking medicine (including anti-inflammatory drugs, antibiotics, supplements), nicotine,
or consuming regularly alcohol (>10/20 g for women and men, respectively, per day). A ques-
tionnaire about their medical history and previous training was filled out by each participant. Selected baseline characteristics of the 33 eligible individuals (17 women, 16 men) in this inves-
tigation are presented in Table 1. Endurance training refers to training of the aerobic system, mainly continuous endurance
training at moderate intensity (60% to 80% peak oxygen uptake), covered per week in the
month prior to the test. Ethics Statement All the participants were informed of the risks and discomforts associated with the investiga-
tion and signed a written consent to participate. The study was approved by the Board for Ethi-
cal Questions in Science Ethics of the University Innsbruck according to the principles
expressed in the Declaration of Helsinki. Introduction Still until now there is no direct informa-
tion available on the effect of exercise on IFN-y and IDO1 activity in humans as any effect of other
pro- and anti-inflammatory cytokines by exercise on IDO1 activity remains to be examined. It should not be neglected that in addition, stimulation of TDO can increase KYN/TRP. However, a contribution of IDO1 rather than TDO to the enhanced TRP breakdown can be
substantiated when an association of KYN/TRP with a biomarker of immune activation, such
as neopterin, can be demonstrated [7], because in parallel to IDO1, IFN-y induces also GTP-
cyclohydrolase 1 (GCH1), the key enzyme for the production of neopterin [6]. Enhanced serum KYN/TRP levels are frequent in patients suffering from diseases which go
along with immune activation and inflammation such as infections, autoimmune syndromes or
malignant tumors [7]. Thereby, IDO1 catalyses the formation of KYN and, hence, limiting availabil-
ity of TRP for serotonin biosynthesis in the brain. Indeed, the probability of depression and fatigue
development is increased in such clinical conditions [13, 14]. Because of its immunotolerizing prop-
erties [15], the upregulation of IDO1 activity may also represent a key component of exercise-
induced immunosuppression in athletes, e.g., immune system disturbances following intense exer-
cise have been linked to an increased risk to upper-respiratory tract infections among athletes [16]. Another important aspect in the pathogenesis of depression and fatigue might represent the
phenylalanine–tyrosine pathway which is controlled by enzyme phenylalanine 4-hydroxylase 2 / 10 PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Tryptophan-Kynurenine Metabolism to Intense Exercise (PAH) and represents the starting point of the biosynthetic pathway of noradrenergic, adrener-
gic and dopaminergic neurotransmitters. The activity of the enzyme is reflected by the phenyl-
alanine (PHE) to tyrosine (TYR) ratio (PHE/TYR). Recently pro-inflammatory cascades were
found to be associated with a disturbed PAH activity [17, 18]. Moreover, in older adults with
chronic low-grade inflammation, not only disturbed TRP metabolism but also PHE and TYR
metabolism was associated with neuropsychiatric symptoms [19]. The aim of the present study was to investigate the effect of exhaustive aerobic exercise on
biomarkers of immune activation and metabolism of the amino acids TRP, KYN, PHE, and
TYR in trained athletes. We hypothesized that exhausting exercise is associated with an acceler-
ated TRP breakdown, shifts of PHE/TYR metabolism, and increased neopterin production,
indicating increased immune activation following intense exercise. PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Experimental Protocol In the morning of the exercise test a standardized breakfast was provided 2 hours prior to exer-
cise (379 kcal; 88 energy percent carbohydrates, 11 energy percent proteins, and 1 energy per-
cent fat). For eligibility testing all subjects performed an incremental cycle ergometer exercise Table 1. Baseline characteristics, aerobic fitness for the participants in this study. Variable
mean ± SD
n (males/females)
16/17
Age (yrs)
26.7 ± 3.7
BMI (kg/m2)
21.7 ± 2.0
Height (cm)
173 ± 8.1
Weight (kg)
65.5 ± 10.6
Endurance training (h/wk)
7.3 ± 3.5
Peak power output (W/kg)
4.6 ± 0.5
Time-trial mean power output (WTT)
205 ± 47
doi:10.1371/journal.pone.0153617.t001 Table 1. Baseline characteristics, aerobic fitness for the participants in this study. Variable
mean ± SD
n (males/females)
16/17
Age (yrs)
26.7 ± 3.7
BMI (kg/m2)
21.7 ± 2.0
Height (cm)
173 ± 8.1
Weight (kg)
65.5 ± 10.6
Endurance training (h/wk)
7.3 ± 3.5
Peak power output (W/kg)
4.6 ± 0.5
Time-trial mean power output (WTT)
205 ± 47
doi:10.1371/journal.pone.0153617.t001 able 1. Baseline characteristics, aerobic fitness for the participants in this study. Table 1. Baseline characteristics, aerobic fitness for the participants in this study. PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 3 / 10 PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Tryptophan-Kynurenine Metabolism to Intense Exercise test until exhaustion. The cycle ergometry was performed on an electronically braked ergome-
ter (Ergometrics 900, Ergoline, Germany) and started at a workload of 50/75 W (women/men)
for 5 minutes (warm up) with a following increase in workload of 25 W per minute until
exhaustion. Exhaustion was assumed when the pedalling rate became below 60 rpm. Heart rate
and ventilatory parameters were monitored continuously (Oxycon mobile, Jaeger, Germany). Peak power output (Wmax) was defined as the last completed workload rate plus the fraction of
time spent in the final uncompleted work rate multiplied by 25 W [20]. Peak oxygen uptake
(VO2max) was defined as the highest 30-second average during the test. After a 20 minutes resting period, athletes with a maximal aerobic capacity of 150% of ref-
erence values for cardiorespiratory response [21] performed a 20-minute maximal time-trial
on a cycle ergometer (RBM Cyclus 2, Germany) as described by Faulhaber and colleagues [20]. Briefly, the cycle ergometer was shifted to a fixed pedal force in which power output was
dependent on the pedalling rate. Blood measurements We conducted blood collections in supine position from a medial cubital vein before exercise
and within 5 min post exercise. After centrifugation for 10 minutes plasma was removed and
samples were frozen at −20°C until analysis. Neopterin concentrations were measured by
ELISA (BRAHMS Diagnostics, Hennigsdorf, Germany) following the manufacturer’s instruc-
tions. Serum concentrations of free TRP and KYN as well as concentrations of phenylalanine
(PHE) and tyrosine (TYR) were determined by high-performance liquid chromatography
(HPLC), as previously described [22, 23]. The ratios of KYN/TRP and PHE/TYR were calcu-
lated as indexes of TRP breakdown and PAH activity, respectively. Experimental Protocol Pedal force for each participant was set so that pedalling at
100 rpm would produce about 70% (rounded to 5 W) of peak power output, which was deter-
mined by the incremental cycle ergometry. During the test, cyclists were strongly encouraged
to choose a maximal pedalling rate that could be maintained for the respective test duration. The main outcome measurement was mean power output during the 20-min test, which was
automatically calculated by the software of the ergometer. The participants were allowed to
drink water ad libitum. Statistical Analysis Statistical analyses were conducted by SPSS (IBM SPSS Statistics Version 22). Normality in the
distribution of data was tested by the Kolmogorov-Smirnov's test. In dependency of Gaussian
distribution, the Student’s t test for paired samples or the Mann-Whitney-U-test was carried
out to assess significant differences in changes of the same variables before and after the inter-
vention (results of parametric and non-parametric calculations were very similar). Spearman’s
rank correlation was used to assess the association between two variables. A p-value of less
than 0.05 (two-tailed) was considered to indicate statistical significance. Data are presented by
mean values ± standard deviation (SD) for baseline characteristics, for biomarkers of immune
activation and metabolism of amino acids by mean values ± standard error of the mean (SEM). Tryptophan-Kynurenine Metabolism to Intense Exercise Table 2. Biological markers before (pre) and after (post) an exhaustive aerobic exercise in 33 athletes. Variable
Pre mean ± SEM
Post mean ± SEM
P-value
Neopterin (nmol/L)
6.4 ± 0.56
10.2 ± 0.97
< 0.001
Tryptophan (μmol/L)
65.1 ± 1.87
57.1 ± 1.65
< 0.001
Kynurenine (μmol/L)
1.88 ± 0.08
1.99 ± 0.09
0.022
KYN/TRP (μmol/mmol)
29.0 ± 1.07
35.1 ± 1.43
< 0.001
Tyrosine (μmol/L)
134 ± 4.66
141 ± 3.90
0.018
Phenylalanine (μmol/L)
69.3 ± 1.59
68.7 ± 1.28
n.s. PHE/TYR (μmol/μmol)
0.53 ± 0.01
0.49 ± 0.01
< 0.001
KYN/TRP kynurenine to tryptophan ratio
PHE/TYR phenylalanine to tyrosine ratio
n.s. not significant
doi:10.1371/journal.pone.0153617.t002 Table 2. Biological markers before (pre) and after (post) an exhaustive aerobic exercise in 33 athletes. At baseline, neopterin concentrations correlated with KYN/TRP levels (r = 0.608,
p<0.0001), a biomarker of TRP catabolism. However, there was only a moderate association
between neopterin and TRP and KYN concentrations (r = -0.297 and r = 0.306, respectively,
p>0.05). No such correlations existed between neopterin and PHE or TYR metabolism. p<0.0001), a biomarker of TRP catabolism. However, there was only a moderate association
between neopterin and TRP and KYN concentrations (r = -0.297 and r = 0.306, respectively,
p>0.05). No such correlations existed between neopterin and PHE or TYR metabolism. Exhausting exercise was associated with a strong increase in neopterin levels (by 159% of
baseline, p<0.001) and this increase was significantly influenced by endurance training volume
with a strong negative correlation between athletes' training status and concentrations of neop-
terin at exhaustion (r = -0.502, p = 0.006). p
In parallel, TRP breakdown was significantly induced by exercise as indicated by a decline
in TRP levels by 12% (p<0.001) and an increase of KYN levels by 6% (p<0.02), accompanied
by an elevation of KYN/TRP by 20% (p<0.001). Concentrations of neopterin still correlated
with KYN/TRP levels (r = 0.560, p = 0.001; Fig 1), Further, both TYR levels and PHE/TYR sig-
nificantly increased with intense exercise (p = 0.018 and p<0.001, respectively). There was no
significant change of PHE concentrations induced by exercise (Table 2). g
g
y
VO2max correlated significantly with baseline concentrations of TRP (r = 0.562, p = 0.001)
and KYN (r = 0.511, p = 0.002) and this relation remained significant for KYN after exercise
(r = 0.359, p = 0.04). Results Females had a lower body mass index (BMI), VO2max, and mean power output during the
20-min test (WTT) compared to male athletes, as KYN levels were lower in females (U = 2.342,
p = 0.019). None of the other parameters was influenced by gender. Concentrations of bio-
markers of immune activation and metabolism of amino acids following intense exercise are
listed in Table 2. 4 / 10 PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Discussion Results of this study show that exhaustive aerobic exercise in well trained athletes elicits signifi-
cant biochemical alterations which are related to inflammation and immune activation cas-
cades. They may also shed some new light on the shift of TRP levels between different body
compartments which has been documented recently by Areces et al. [5]. The decline of essen-
tial amino acid TRP in our study was much stronger as compared with the observations after
the half iron-marathon where the alteration of absolute TRP levels failed to reach statistical sig-
nificance, and only the decline of the ratio of TRP vs. branch chained amino acids (BCAA) was
significant [5]. Still the drastic decline of TRP in our study could relate to muscle fatigue experi-
enced after intense physical exercise. Our results indicate an involvement of IDO1 activation in the enhanced TRP catabolism
and KYN production following intense exercise, because a strong association was found
between neopterin and KYN/TRP levels throughout the study suggestive of IDO1-induced
TRP catabolism. This observation could be of special relevance for several pathophysiological
consequences that are induced by physical exercise. The decline of TRP by IDO1 is well estab-
lished as an immunoregulatory event [24] which is related to the development of regulatory T-
cells and dampening immune response as a kind of feedback control [15]. In the present study, the resulting low TRP levels followed by intense exercise may diminish
brain supply, thereby reducing TRP availability to the brain for serotonin production. The
decline of TRP was drastic and would certainly also diminish the TRP to BCAA ratio that is
most relevant for the transport of TRP into the brain via the leucine-preferring L1 transporter
system. Alternatively, down-stream catabolites of kynurenine like quinolinic acid and picolinic
acid are formed which could be of physiological importance especially regarding their possible
interference with neuroendocrine circuits like the interaction with the N-methyl-D-aspartate
(NMDA) receptor [25–28]. Thus, the stimulated TRP breakdown could relate to mood alter-
ations related to physical exercise and may contribute to a declining training adherence in
athletes. In our study, baseline TRP metabolism was found to be associated with aerobic fitness. Higher levels of both TRP and KYN at baseline were associated with a higher aerobic capacity
(VO2max). WTT correlated with KYN levels before (r = 0.534, p = 0.001) and after
exercise (r = 0.409, p = 0.018) but not with baseline and post-exercise TRP concentrations. No
significant associations existed between PHE-TYR metabolism and all sport physiological vari-
ables (S1 and S2 Files). Fig 1. Association between neopterin and kynurenine to tryptophan concentrations (KYN/TRP) in 33
trained athletes after exhaustive aerobic exercise. The correlation of neopterin with post-exercise KYN/
TRP ratio, a biomarker of tryptophan catabolism, was statistically significant (r = 0.560, p = 0.001). doi:10.1371/journal.pone.0153617.g001
PLOS ONE | DOI 10 1371/j
l
0153617
A
il 28 2016
5 / 10 Fig 1. Association between neopterin and kynurenine to tryptophan concentrations (KYN/TRP) in 33
trained athletes after exhaustive aerobic exercise. The correlation of neopterin with post-exercise KYN/
TRP ratio, a biomarker of tryptophan catabolism, was statistically significant (r = 0.560, p = 0.001). doi:10.1371/journal.pone.0153617.g001 Fig 1. Association between neopterin and kynurenine to tryptophan concentrations (KYN/TRP) in 33
trained athletes after exhaustive aerobic exercise. The correlation of neopterin with post-exercise KYN/
TRP ratio, a biomarker of tryptophan catabolism, was statistically significant (r = 0.560, p = 0.001). doi:10.1371/journal.pone.0153617.g001 PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 5 / 10 Tryptophan-Kynurenine Metabolism to Intense Exercise PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Discussion However, when physical
exercise or training is too heavy and exhaustive, tryptophan breakdown by IDO1 becomes too drastic and the
decline of tryptophan due to IDO1 activity can no longer be compensated by BH4, the life span of the latter is
decreased by ROS exposure. Thus, athletes may suffer from insufficient supply with neurotransmitters and
will experience low mood. doi:10 1371/journal pone 0153617 g002 Fig 2. Divergent effects of moderate vs. exhaustive physical exercise on the production of
neurotransmitters: Pro-inflammatory cytokines like interferon-y (IFN-y) stimulates several enzymes
including [a] indoleamine 2,3-dioxygenase-1 (IDO1), which degrades tryptophan (TRP) and serotonin (5HT),
[b] NADPH oxidase (NOx), which produces reactive oxygen species (ROS), and [c] GTP-cyclohydrolase-1
(GCH1), which in human macrophages forms neopterin and in other cells tetrahydrobiopterin (BH4), the
necessary cofactor of several amino acid hydroxylases, including tryptophan 5-hydroxylase (TH) for the
production of 5-hydroxytryptamine (5HT, serotonin) and phenylalanine 4-hydroxylase (PAH) for the
production of tyrosine, precursor of dopamine, adrenaline and noradrenaline. Physical exercise is followed by
a pro-inflammatory immune response which induces BH4, thereby upregulating several neurotransmitters
(see pathways marked in red), associated with mood enhancement and well-being. However, when physical
exercise or training is too heavy and exhaustive, tryptophan breakdown by IDO1 becomes too drastic and the
decline of tryptophan due to IDO1 activity can no longer be compensated by BH4, the life span of the latter is
decreased by ROS exposure. Thus, athletes may suffer from insufficient supply with neurotransmitters and
will experience low mood. doi:10.1371/journal.pone.0153617.g002 IDO1 will follow and slows down serotonin formation when TRP becomes diminished. Conse-
quently, BH4-dependent biosynthesis of several neurotransmitters will decline and the mood
enhancement after moderate exercise will be followed by a decline when sports become too
heavy or training intervals too intense. It could play a role in the onset of fatigue and sleep dis-
turbances [29, 32]. One may conclude that sports performed occasionally with two or three days interval exerts
a beneficial effect on general well-being as it is the case when it is performed as recreational
activity whereas intense training will more and more achieve adverse effects on both mood
[29] and immune system [33]. Prolonged bouts of strenuous exercise have been shown to result
in transient depression of white blood cell functions and it is suggested that such changes create
an “open window” of decreased host protection, increasing the risk of developing an infection. Discussion On the other hand, athletes with higher training volumes in the month prior to the
test demonstrated a smaller rise in neopterin concentrations to intense exercise, suggesting
lower immune activation with better training status. In addition, we found a significant decline
of PHE/TYR following exercise, suggesting a higher activity of PAH induced by exercise. Such
an increase may indicate a stimulated production of the noradrenergic, adrenergic and dopa-
minergic neurotransmitters, which could contribute to an improved neuropsychiatric presen-
tation [18]. Thus, the higher production rate of the neurotransmitters may compensate, at least
partly, the mood lowering effect of intense exercise [29], potentially caused by the decline of
TRP. The increase of PAH activity, indicated by PHE/TYR, could result from the increased
activity of GCH1, which is reflected by the increase of monocyte-macrophage-derived neop-
terin that is usually accompanied by a similar increase of 5,6,7,8-tetrahydrobiopterin (BH4),
the cofactor of PAH and also tyrosine hydroxylase, as well as tryptophan hydroxylase, enzymes
that are all important in the formation of relevant neurotransmitters [30] (Fig 2). With this respect it is important to note that moderate vs. intense physical activity might
have contrasting effects. On the one hand, physical exercise may enhance the production of
neurotransmitters by induction of BH4 biosynthesis and increasing neurotransmitter biosyn-
thesis which will contribute to mood enhancement (Fig 2). However, the pro-inflammatory
cascades that are initiated are accompanied by production of ROS, which may diminish the
endogenous antioxidant pools, including BH4 concentrations [31]. Breakdown of TRP by PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 6 / 10 Tryptophan-Kynurenine Metabolism to Intense Exercise Fig 2. Divergent effects of moderate vs. exhaustive physical exercise on the production of
neurotransmitters: Pro-inflammatory cytokines like interferon-y (IFN-y) stimulates several enzymes
including [a] indoleamine 2,3-dioxygenase-1 (IDO1), which degrades tryptophan (TRP) and serotonin (5HT),
[b] NADPH oxidase (NOx), which produces reactive oxygen species (ROS), and [c] GTP-cyclohydrolase-1
(GCH1), which in human macrophages forms neopterin and in other cells tetrahydrobiopterin (BH4), the
necessary cofactor of several amino acid hydroxylases, including tryptophan 5-hydroxylase (TH) for the
production of 5-hydroxytryptamine (5HT, serotonin) and phenylalanine 4-hydroxylase (PAH) for the
production of tyrosine, precursor of dopamine, adrenaline and noradrenaline. Physical exercise is followed by
a pro-inflammatory immune response which induces BH4, thereby upregulating several neurotransmitters
(see pathways marked in red), associated with mood enhancement and well-being. PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 Supporting Information Supporting Information
S1 File. (PDF)
S2 File. (PDF) Acknowledgments We thank all participants for their contribution to the study. Discussion The current investigation presents some limitations with respect to the experimental design
and the measurements obtained. Our findings are certainly limited by the relatively small sam-
ple size, so that we were not able to analyze specific subgroups. This would be especially impor-
tant when effects of gender could be of relevance, e.g., females presented with lower KYN levels
and VO2max compared to males. Still, peak power output (W/kg) was not influenced by gender
and, further, the effect of exercise on KYN/TRP or neopterin concentrations was seen in both
subgroups and, thus, independent of gender. Future studies should consider gender effects in a
larger population since an influence of sex steroid hormones like estrogen on TDO is well
established [10]. In a similar way, a potential influence of cortisol on TDO activity cannot be
ruled out [9]. Salivary and serum cortisol levels in humans and corticosterone in rats are
increased by exercise [34, 35] and this could induce liver TDO, as demonstrated in rats. The
combined effects of increased liver TRP and TDO induction can elevate serum KYN in humans
[36] and rats [12]. In both of these latter studies, increases in serum KYN after exercise have PLOS ONE | DOI:10.1371/journal.pone.0153617
April 28, 2016 7 / 10 Tryptophan-Kynurenine Metabolism to Intense Exercise been shown. Thus, also an additional role of TDO in the exercise-induced TRP breakdown
cannot be fully ruled out. Further, we did not measure NAD+ levels and downstream NAD+ related pathways in the
present study. Future work should address the regulatory influence of NAD+ on the expression
of sirtuins (SIRT) and PGC-1α because enhanced SIRT1 and PGC-1α activity is associated
with improved mitochondrial function and exercise performance [37], and protection against
obesogenic feeding [38]. Author Contributions Conceived and designed the experiments: BS DG MS HG MB DF. Performed the experiments:
BS DG MS HG MB DF. Analyzed the data: BS HG MB DF. Contributed reagents/materials/
analysis tools: HG MB DF. Wrote the paper: BS DG MS HG MB DF. Conclusions In summary, our study demonstrates a significant influence of exhaustive aerobic exercise on
biochemical pathways which are linked to inflammation and immune activation responses
induced by exercise. Intense exercise was associated with a strong increase in neopterin levels. In parallel, TRP breakdown was significantly induced by exercise, accompanied by an elevation
of the KYN/TRP ratio, suggestive of IDO1-induced TRP catabolism. The stimulated TRP
breakdown could relate to mood alterations related to physical exercise and may play a role in
the onset of fatigue. Supporting Information
S1 File. (PDF)
S2 File. (PDF) PLOS ONE | DOI:10.1371/journal.pone.0153617
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ANALYSIS OF MEDICAL AND STATISTICAL MEASURES OF MORBIDITY IN CONSCRIPTS OF THE NAVY AND GROUND FORCES OF THE RUSSIAN FEDERATION IN 2003–2018
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1 Медицинская служба Главного командования Военно-морского флота России
(Россия, Санкт-Петербург, Адмиралтейский проезд, д. 1); 2 Военно-медицинская академия им. С.М. Кирова (Россия, Санкт-Петербург, ул. Акад. Лебедева, д. 6);
3 Всероссийский центр экстренной и радиационной медицины им. А.М. Никифорова МЧС России
(Россия, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2) Актуальность. Комплектование частей Военно-морского флота (ВМФ) и Сухопутных войск России но
выми образцами вооружения определяет высокие требования к состоянию соматического и психическо
го здоровья призываемого контингента. д р
р
Цель – проанализировать показатели заболеваемости военнослужащих, проходящих службу
зыву, ВМФ России и Сухопутных войск России в 2003–2018 гг. Методология. Провели выборочный статистический анализ медицинских отчетов о состоянии здоро
вья личного состава и деятельности медицинской службы по форме 3/МЕД воинских частей, в которых
проходили службу около 60 % от общего числа военнослужащих по призыву Вооруженных сил России
в 2003–2018 гг. Проанализировали общепринятые медико-статистические показатели заболеваемости
по классам болезней Международной статистической классификации болезней и проблем, связанных
со здоровьем, 10-го пересмотра. р
р
р
Результаты и их анализ. Среднегодовые уровни общей заболеваемости военнослужащих по призыву
ВМФ России были (1833,3 ± 84,9) ‰, Сухопутных войск России – (2008,0 ± 102,4) ‰, первичной заболе
ваемости – (1019,9 ± 54,8) и (1014,0 ± 79,2) ‰ соответственно, нуждаемости в диспансерном наблюде
нии – (166,5 ± 19,0) и (128,2 ± 8,1) ‰, госпитализации – (968,5 ± 71,3) и (1033,5 ± 89,6) ‰, дней трудо
потерь – (13 166 ± 7,99) и (11 104 ± 595) ‰ (p < 0,01), увольняемости – (33,38 ± 1,79) и (18,28 ± 1,66) ‰,
смертности – (24,87 ± 5,12) и (50,67 ± 7,84) на 100 тыс. военнослужащих по призыву соответствующей
когорты (p < 0,05). При разных по значимости коэффициентах детерминации полиномиальные тренды
общей и первичной заболеваемости, госпитализации и дней трудопотерь показывают тенденции увели
чения данных, увольняемости и смертности – уменьшения показателей. Согласованность трендов прак
тически всех видов заболеваемости – умеренная и статистически достоверная, что может указывать на
влияние в их формировании однонаправленных факторов, например военно-профессиональных. Согла
сованность трендов смертности мужского населения России в возрасте 20–24 года и военнослужащих по
призыву ВМФ России и Сухопутных войск – функциональная (r = 0,83 и 0,87; p < 0,001), что свидетельству
ет о влиянии однонаправленных факторов, например, макросоциальных, поведенческих и пр. В.И. Евдокимов2, 3, И.Г. Мосягин1,
П.П. Сиващенко2, Н.А. Мухина3 УДК 613.67 : [355.511.512 + 355.336]
DOI 10.25016/2541-7487-2019-0-3-15-51 Евдокимов Владимир Иванович – д-р мед. наук проф., Всерос. центр экстрен. и радиац. медицины им. А.М. Ни
кифорова МЧС России (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2); Воен.-мед. акад. им. С.М. Киро
ва (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 6), e-mail: 9334616@mail.ru;
Мосягин Игорь Геннадьевич – д-р мед. наук проф., нач. мед. службы Главного командования Военно-морского
флота России, e-mail: mosyagin-igor@mail.ru;
Сиващенко Павел Павлович – канд. мед. наук доц., Воен.-мед. акад. им. С.М. Кирова (Россия, 194044, Санкт-Пе
тербург, ул. Акад. Лебедева, д. 6);
Мухина Наталия Александровна – канд. мед. наук, ст. науч. сотр., Всерос. центр экстрен. и радиац. медицины им.
А.М. Никифорова МЧС России (Россия, 190044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2), e-mail: nata26@inbox.ru иологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 иологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 УДК 613.67 : [355.511.512 + 355.336]
DOI 10.25016/2541-7487-2019-0-3-15-51 В.И. Евдокимов2, 3, И.Г. Мосягин1,
П.П. Сиващенко2, Н.А. Мухина3 Мухина Наталия Александровна – канд. мед. наук, ст. науч. сотр., Всерос. центр экстрен. и радиац. медицины им.
А.М. Никифорова МЧС России (Россия, 190044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2), e-mail: nata26@inbox.ru щенко Павел Павлович – канд. мед. наук доц., Воен.-мед. акад. им. С.М. Кирова (Россия, 194044, Санкт-Пе
л. Акад. Лебедева, д. 6); MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 АНАЛИЗ МЕДИКО-СТАТИСТИЧЕСКИХ ПОКАЗАТЕЛЕЙ ЗАБОЛЕВАЕМОСТИ
ВОЕННОСЛУЖАЩИХ ПО ПРИЗЫВУ ВОЕННО-МОРСКОГО ФЛОТА
И СУХОПУТНЫХ ВОЙСК РОССИЙСКОЙ ФЕДЕРАЦИИ В 2003–2018 ГГ. 1 Медицинская служба Главного командования Военно-морского флота России
(Россия, Санкт-Петербург, Адмиралтейский проезд, д. 1); Евдокимов Владимир Иванович – д-р мед. наук проф., Всерос. центр экстрен. и радиац. медицины им. А.М. Ни
кифорова МЧС России (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2); Воен.-мед. акад. им. С.М. Киро
ва (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 6), e-mail: 9334616@mail.ru;
М
И
Г
ф
б
Г
В осягин Игорь Геннадьевич – д-р мед. наук проф., нач. мед. службы Главного командования Военно-морско
России, e-mail: mosyagin-igor@mail.ru; Евдокимов Владимир Иванович – д-р мед. наук проф., Всерос. центр экстрен. и радиац. медицины им. А.М. Ни
кифорова МЧС России (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2); Воен.-мед. акад. им. С.М. Киро
ва (Россия, 194044, Санкт-Петербург, ул. Акад. Лебедева, д. 6), e-mail: 9334616@mail.ru;
Мосягин Игорь Геннадьевич – д-р мед. наук проф., нач. мед. службы Главного командования Военно-морского
флота России, e-mail: mosyagin-igor@mail.ru;
Сиващенко Павел Павлович – канд. мед. наук доц., Воен.-мед. акад. им. С.М. Кирова (Россия, 194044, Санкт-Пе
тербург, ул. Акад. Лебедева, д. 6); ф
,
y g
g
;
Сиващенко Павел Павлович – канд. мед. наук доц., Воен.-мед. акад. им. С.М. Кирова (Россия, 194044, Санкт-Пе
тербург, ул. Акад. Лебедева, д. 6);
Мухина Наталия Александровна – канд. мед. наук, ст. науч. сотр., Всерос. центр экстрен. и радиац. медицины им.
А.М. Никифорова МЧС России (Россия, 190044, Санкт-Петербург, ул. Акад. Лебедева, д. 4/2), e-mail: nata26@inbox.ru Vladimir Ivanovich Evdokimov – Dr. Med. Sci. Prof., Nikiforov Russian Center of Emergency and Radiation Medicine of
EMERCOM of Russia (4/2, Academica Lebedeva Str., St. Petersburg, 194044, Russia), e-mail: 9334616@mail.ru;
Igor' Gennad'evich Mosyagin – Dr. Med. Sci. Prof., Chief Medical Officer of the Main Headquerters of the Russian Navy (1,
Admiralteiskii proezd, St. Petersburg, 190098, Russia), e-mail: mosyagin-igor@mail.ru;
Pavel Pavlovich Sivashchenko – PhD Med. Sci. Associate Prof., Kirov Military Medical Academy (6, Academica Lebedeva
Str., St. Petersburg, 194044, Russia), e-mail: pavel-siv@yandex.ru
Nataliia Aleksandrovna Mukhina – PhD Med, senior research associate, Medical Register of EMERCOM of Russia, Nikiforov
Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia (4/2, Academica Lebedeva Str., St. Petersburg,
194044, Russia), e-mail: nata26@inbox.ru Abstract Relevance. The Navy of Russia and the Ground Forces of Russia recieve new kinds of weapons, hence high
requirements to the somatic and mental health of the conscripts. Intention: To analyze morbidity in conscripts of the Russian Navy and the Ground Forces in 2003–2018 Methodology. A selective statistical analysis was performed using medical reports on the state of health of
personnel and activities of the medical service according to Form 3 / MED in military units, in which about 60 %
of the total number of conscripts of the Armed Forces of Russia served in 2003–2018. Commonly accepted
medical and statistical morbidity indicators were analyzed by disease categories of the International Statistical
Classification of Diseases and Health Problems, 10th revision. Results and Discussion. The average annual morbidity rates were (1833.3 ± 84.9) ‰ in conscripts of the
Russian Navy and (2008.,0 ± 102.4) ‰ in conscripts of the Russian Ground Forces; primary morbidity rates
were (1019.9 ± 54,8) and (1014.0 ± 79.2) ‰, respectively; the need for case follow-up was (166.5 ± 19.0) and
(128.2 ± 8.1) ‰, respectively; hospital admissions were (968.5 ± 71.3) and (1033.5 ± 89.6) ‰, respectively;
work days lost (13,166 ± 7.99) and (11,104 ± 595) ‰, respectively (p < 0.01); dismissal rates were (33.38 ±
1.79) and (18.28 ± 1.66) ‰, respectively; mortality rates were (24.87 ± 5.12) and (50.67 ± 7.84) per 100 thou
sand conscripts of the respective cohort (p < 0.05). With determination coefficients of various significance, the
polynomial trends show an increase in primary and general morbidity, hospital admissions and work days lost, as
well as decrease in dismissal rates and mortality rates. The trends in almost all morbidity types are moderately
and statistically significantly consistent, suggesting the influence of uniderectional factors, for example, mili
tary-professional ones. There is a functional consistency between mortality trends in Russian male population
aged 20–24 and conscripts of the Russian Navy and Ground Forces (r = 0.83 and 0.87; p < 0.001), thus suggest
ing the influence of uniderectional factors, for example, of macro-social, behavioural or other nature. ANALYSIS OF MEDICAL AND STATISTICAL MEASURES OF MORBIDITY IN CONSCRIPTS
OF THE NAVY AND GROUND FORCES OF THE RUSSIAN FEDERATION IN 2003–2018 1 Medical Service of the Navy Headquarters of the Russian Federation
(1, Admiralteiskii proezd, St. Petersburg, 190098, Russia)
2 Kirov Military Medical Academy (6, St. Petersburg, 194044, Russia)
3 Nikiforov Russian Center of Emergency and Radiation medicine
(4/2, Academica Lebedeva Str., St. Petersburg, 194044, Russia) 1 Medical Service of the Navy Headquarters of the Russian Federation
(1, Admiralteiskii proezd, St. Petersburg, 190098, Russia)
2 Kirov Military Medical Academy (6, St. Petersburg, 194044, Russia)
3 Nikiforov Russian Center of Emergency and Radiation medicine
(4/2, Academica Lebedeva Str., St. Petersburg, 194044, Russia) (1, Admiralteiskii proezd, St. Petersburg, 190098, Russia)
2 Kirov Military Medical Academy (6, St. Petersburg, 194044, Russia)
3 Nikiforov Russian Center of Emergency and Radiation medicine
(4/2, Academica Lebedeva Str., St. Petersburg, 194044, Russia) 2 Kirov Military Medical Academy (6, St. Petersburg, 194044, Russia)
3 Nikiforov Russian Center of Emergency and Radiation medicine
(4/2, Academica Lebedeva Str., St. Petersburg, 194044, Russia) 1 Медицинская служба Главного командования Военно-морского флота России
(Россия, Санкт-Петербург, Адмиралтейский проезд, д. 1); В анализи
руемых когортах военнослужащих по призыву ведущие ранги военно-эпидемиологической значимости
классов болезней совпали: 1-й ранг составили болезни органов дыхания (X класс по МКБ‑10); 2-й – трав
мы и другие воздействия внешних причин (XIX класс); 3-й – психические расстройства и расстройства по
ведения (V класс); 4-й – болезни кожи и подкожной клетчатки (XII класс); 5-й – болезни органов пищеваре
ния (XI класс). Ведущие ранги военно-эпидемиологической значимости нозологий у военнослужащих по
призыву составили показатели острых респираторных инфекций верхних дыхательных путей (J00–J06 по
МКБ‑10), травм головы (S00–S09), травм, захватывающих несколько областей тела (T00–T07), болезней
пищевода, желудка и двенадцатиперстной кишки (K20–К31). У военнослужащих по призыву ВМФ Рос
сии высокую значимость имели данные невротических, связанных со стрессом, и соматоформных рас
стройств (F40–F48), Сухопутных войск – инфекций кожи и подкожной клетчатки (L00–L08). Заключение. Профилактика, лечение и реабилитация по ведущим нозологиям будут способствовать
повышению состояния здоровья военнослужащих по призыву, а учет распространенности и структуры
заболеваемости – совершенствовать силы и средства медицинской службы Вооруженных сил России. повышению состояния здоровья военнослужащих по призыву, а учет распространенности и структуры
заболеваемости – совершенствовать силы и средства медицинской службы Вооруженных сил России. Ключевые слова: экстремальная деятельность, военнослужащий, служба по призыву, солдат, мат
рос, сержант, старшина, заболеваемость, трудопотери, госпитализация, увольняемость, смертность,
Военно-морской флот, Сухопутные войска, Вооруженные силы России. Ключевые слова: экстремальная деятельность, военнослужащий, служба по призыву, солдат, мат
рос, сержант, старшина, заболеваемость, трудопотери, госпитализация, увольняемость, смертность,
Военно-морской флот, Сухопутные войска, Вооруженные силы России. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 15 иологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 V.I. Evdokimov2, 3, I.G. Mosyagin1,
P.P. Sivashchenko2, N.A. Mukhina3 UDC 613.67 : [355.511.512 + 355.336]
DOI 10.25016/2541-7487-2019-0-3-15-51 UDC 613.67 : [355.511.512 + 355.336]
DOI 10.25016/2541-7487-2019-0-3-15-51 6
MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Introduction Due to the general population decline,
the share of conscription troops in the
Armed Forces (AF) of Russia decreases. Large-scale studies have shown that more
than 40 % of young draftees are virtually
unable to adjust to the conditions of mili
tary service in terms of physical and mental
development [7, 11, 19]. For example, out
of 5,000 conscripts sent for recruiting units
and parts of the Baltic Fleet in 2011–2015,
only 49 to 67 % were considered fit for mili
tary service without restrictions [8]. На фоне общей депопуляции населения умень
шается доля призывного контингента в Воору
женные силы (ВС) России. Широкомасштаб
ные исследования показали, что более 40 %
юношей-призывников по показателям физическо
го и психического развития практически не спо
собны адаптироваться к условиям военной служ
бы [7, 11, 19]. Например, из 5 тыс. призывников,
направляемых для комплектования соединений
и частей Балтийского флота в 2011–2015 гг., при
знавались годными к военной службе без ограни
чения от 49 до 67 % [8]. В то же время, комплектование частей Воен
но-морского флота (ВМФ) и Сухопутных войск
России новыми образцами вооружения определя
ет высокие требования к состоянию соматическо
го и психического здоровья призываемого контин
гента [6, 13, 15]. At the same time, new types of weap
ons acquired by the Navy and the Ground
Forces of Russia set high demands on the
state of somatic and mental health of the
conscripts [6, 13, 15]. Health disorders in the conscripts are
attributed to adaptation as a result of a
decrease in cellular immunity and resis
tance of the body’s functional reserves [3,
10, 12]. Unusual living conditions, mental
stress, shortcomings of draft commissions,
poor organization of military labor deter
mine a high proportion of dismissals of con
scripts from the Russian Armed Forces due
to mental disorders [9, 14, 20]. Нарушения состояния здоровья у военнослужа
щих по призыву относятся к адаптационно обуслов
ленным в результате снижения клеточного иммуни
тета и резистентности функциональных резервов
организма [3, 10, 12]. Непривычные условия обита
ния, психический стресс, недочеты призывных ко
миссий военкоматов, низкая организация военного
труда определяют высокую долю увольнений воен
нослужащих по призыву из ВС России по причине
психических расстройств [9, 14, 20]. Initially, the professional activities of mil
itary personnel imply risks to health or even
life, which increase in case of disturbed hy
gienic conditions of accommodation, food,
organization of studies and especially when
performing military professional tasks in
the field [2, 5, 17]. Introduction Изначально профессиональная деятельность
военнослужащих предполагает риск ухудшения
здоровья, однако в бо́льшей степени это сказыва
ется при нарушении гигиенических условий раз
мещения, питания, организации учебы и особенно
при выполнении военно-профессиональных задач
в полевых условиях [2, 5, 17]. The intention was to compare medical
and statistical morbidity indicators by dis
ease categories of the International Statis
tical Classification of Diseases and Health
Problems, 10th revision (ICD‑10) and de
termine a role of specific diseases (cate
gories) in developing health disorders in
the conscripts of the Navy of Russia and
the Ground Forces of Russia over 16 years,
from 2003 to 2018. Цель – провести сравнительный анализ меди
ко-статистических показателей заболеваемости
по классам Международной статистической клас
сификации болезней и проблем, связанных со
здоровьем, 10-го пересмотра (МКБ‑10) и устано
вить роль конкретных нозологий (групп в классах)
в развитии нарушений состояния здоровья воен
нослужащих по призыву ВМФ России и Сухопут
ных войск России за 16 лет с 2003 по 2018 г. Abstract In the co
horts of conscripts under study, the leading disease categories from military-epidemiological point of view were
similar: respiratory diseases (ICD‑10 category X) ranked 1st; injury, poisoning and certain other consequences
of external causes (XIX) ranked 2nd; mental and behavioural disorders (V) ranked 3rd; diseases of the skin and
subcutaneous tissue (XII) ranked 4th; diseases of the digestive system (XI) ranked 5th. In conscripts, the leading
diseases from military-epidemiological point of view were as follows: acute respiratory infections of the upper
respiratory tract (J00–J06 by ICD‑10); head injuries (S00–S09); injuries to unspecified part of trunk, limb or
body region (T00–T07); diseases of oesophagus, stomach and duodenum (K20–К31). In the Navy conscripts,
neurotic and stress-related and somatoform disorders (F40–F48) played an important role; in conscripts of the
Ground Forces, infections of the skin and subcutaneous tissue (L00–L08). Conclusion. Prevention, timely treatment and rehabilitation will help improve the health status of conscripts. Taking into account the rates and structure of morbidity will optimize allocation of resources the medical service
of the Armed Forces of Russia. Keywords: extreme activity, serviceman, conscript, soldier, sailor, sergeant, morbidity, work days
pital admission, dismissal, mortality, Navy, Ground Forces, Russian Armed Forces. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 16 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Material and methods A selective statistical analysis was per
formed using medical reports on the state
of health of personnel and activities of the
medical service according to Form 3 / MED
in military units, in which about 60 % of the
total number of conscripts of the Armed
Forces of Russia served in 2003–2018 [16]. Провели выборочный статистический анализ
медицинских отчетов о состоянии здоровья лично
го состава и деятельности медицинской службы по
форме 3/МЕД воинских частей, в которых проходи
ли службу около 60 % от общего числа военнослу
жащих по призыву ВС России в 2003–2018 гг. [16]. Проанализировали общепринятые медико-ста
тистические
показатели
заболеваемости
[18]
военнослужащих по призыву ВС России, ВМФ Commonly accepted medical and sta
tistical morbidity indicators [18] were ana 17 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № и Сухопутных войск России по классам болезней
МКБ‑10 (табл. 1). По сложившейся традиции не
учитывали стоматологическую заболеваемость
(K00–K14 по МКБ‑10). lyzed in the conscripts of the Armed Forces
of Russia, the Navy of Russia and the Ground
Forces of Russia by ICD‑10 disease catego
ries (Table 1). As usual, stomatological dis
eases (K00–K14 by ICD‑10) were omitted. Данные заболеваемости военнослужащих по
призыву определяли без учета возраста, воен
ной специальности, срока службы, региона бази
рования, организации медицинского и санитар
но-гигиенического обеспечения частей и других
факторов влияния. Morbidity rates in conscripts were deter
mined regardless of age, military special
ty, experience, home region, organization
of medical and sanitary-hygienic support or
other factors. Morbidity rates were calculated per 1000
conscripts ( ‰). The data from conscripts of
the Russian Navy and Ground Forces were
compared with all the conscripts of the Rus
sian Armed Forces [4], an integral part of
which they are. A share of 5 % or more was
considered significant in the structure of mor
bidity rates by ICD‑10 chapters, and for code
blocks – 0.5 % or more. Code blocks under
the ICD‑10 headings “Other …” were exclud
ed from analysis. Code blocks with contribu
tion of less than 0.5 % and included into the
tables for comparison, were ranked “>”, since
their actual rank in the morbidity structure in
conscripts was not assessed. Показатели заболеваемости рассчитывали
на 1000 военнослужащих по призыву или в про
милле ( ‰). Данные военнослужащих по призыву
ВМФ России и Сухопутных войск России срав
нили с показателями у военнослужащих по при
зыву ВС России [4], понимая, что они являются
составной частью общих сведений. Material and methods Значимым
классом в структуре показателей заболевае
мости считали с долей 5 % и более, нозологий
(групп в классах) – 0,5 % и более. Не анализиро
вали показатели групп нозологий в классах, вхо
дящих в рубрики МКБ‑10 «Другие …». Нозоло
гиям (группам в классах), которые имели вклад
менее 0,5 % и были представлены в таблицах
для сравнения, присваивали ранг со значком
«более» (>), так как их реальный ранг в структуре
заболеваемости военнослужащих по призыву не
исследовали. The article presents the mean arithmetic
values and their statistical errors M ± m. Mor
bidity over time was assessed by analyzing
the time series and calculating second-order
polynomial trends [1]. Coefficients of deter
mination (R2) were used to show the relation В статье указаны средние арифметические
данные и их статистические ошибки – M ± m. Ди
намику и прогнозирование показателей заболе 18
MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3
Таблица 1. Классы болезней и причин смерти, принятых в МКБ‑10
Table 1. Categories of diseases and causes-of-death by ICD‑10
Класс /
Chapter
Наименование класса
Title
Код /
Blocks
I
Некоторые инфекционные и паразитарные
болезни
Certain infectious and parasitic diseases
A00–B99
II
Новообразования
Neoplasms
С00–D48
III
Болезни крови, кроветворных органов и от
дельные нарушения, вовлекающие иммун
ный механизм
Diseases of the blood and blood-forming organs
and certain disorders involving the immune
mechanism
D50–D89
IV
Болезни эндокринной системы, расстрой
ства питания и нарушения обмена веществ
Endocrine, nutritional and metabolic diseases
E00–E90
V
Психические расстройства и расстройства
поведения
Mental and behavioural disorders
F00–F99
VI
Болезни нервной системы
Diseases of the nervous system
G00–G99
VII
Болезни глаз и его придаточного аппарата
Diseases of the eye and adnexa
H00–H59
VIII
Болезни уха и сосцевидного отростка
Diseases of the ear and mastoid process
H60–H95
IX
Болезни системы кровообращения
Diseases of the circulatory system
I00–I99
X
Болезни органов дыхания
Diseases of the respiratory system
J00–J99
XI
Болезни органов пищеварения
Diseases of the digestive system
K00–K93
XII
Болезни кожи и подкожной клетчатки
Diseases of the skin and subcutaneous tissue
L00–L99
XIII
Болезни костно-мышечной системы и соеди
нительной ткани
Diseases of the musculoskeletal system and
connective tissue
M00–M99
XIV
Болезни мочеполовой системы
Diseases of the genitourinary system
N00–N99
XIX
Травмы, отравления и некоторые другие по
следствия воздействия внешних причин
Injury, poisoning and certain other consequen
ces of external causes
S00–T98 Таблица 1. Классы болезней и причин смерти, принятых в МКБ‑10
Table 1. Результаты и их анализ Общая заболеваемость. Уровень общей за
болеваемости в 2003–2018 гг. военнослужащих
по призыву ВМФ России был (1833,3 ± 84,9) ‰,
Сухопутных войск России – (2008,0 ± 102,4) ‰,
ВС России – (1839,1 ± 50,5) ‰, т. е. ежегодно
практически каждый военнослужащий по при
зыву обращался 2 раза за медицинской помо
щью (консультацией) в военно-медицинскую
организацию (табл. 2). Статистически достовер
ных различий в уровнях общей заболеваемости
в рассматриваемых когортах нет. A polynomial trend of general morbidity
resembles an inverted U-curve with very low
determination coefficient (R2 = 0.09) in Navy
conscripts, and shows an increase with low
determination coefficient (R2 = 0.34) in the
Ground Forces conscripts (Fig. 1). General
morbidity curves between Navy conscripts
and Ground Forces conscripts are moderately
and positively consistent (statistically signifi
cantly; r = 0.61; p < 0.05), thus suggesting a
role of unidirectional factors in general mor
bidity (see Fig. 1). Полиномиальный тренд общей заболевае
мости военнослужащих по призыву ВМФ Рос
сии при очень низком коэффициенте детерми
нации (R2 = 0,09) напоминает инвертированную
U-кривую, военнослужащих по призыву Сухо
путных войск России при низком коэффициенте
детерминации (R2 = 0,34) – показывает тенден
цию увеличения данных (рис. 1). Согласован
ность кривых показателей общей заболеваемо
сти военнослужащих по призыву ВМФ России
и Сухопутных войск России – умеренная, поло
жительная и статистически значимая (r = 0,61;
p < 0,05), что может указывать на участие в фор
мировании общей заболеваемости однонаправ
ленных факторов (см. рис. 1). The highest general morbidity rates in Navy
conscripts were in ICD‑10 categories X, XII,
I and XI: (974.7 ± 64.7), (284.7 ± 19.3), (113.5
± 9.2) and (102.1 ± 6.5) ‰, respectively; in
Ground Forces conscripts in ICD‑10 cate
gories X, XII, XI and I: (914.9 ± 87.9), (529.3
± 43.7), (107.2 ± 6.6) and (104.8 ± 8.1) ‰,
respectively (see Table 2). In the Navy con
scripts, general morbidity rates including
ICD‑10 categories II, III, V and XIX were sta
tistically significantly higher compared to the
Ground Forces conscripts and lower for cate
gories VI, VII, VIII and XII (see Table 2). Material and methods Categories of diseases and causes-of-death by ICD‑10 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 18 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № ваемости оценивали с помощью анализа ди
намических рядов и расчета полиномиального
тренда второго порядка [1]. Силу связи показа
телей полиномиального тренда исследовали при
помощи коэффициента детерминации (R2), кото
рый определял связь динамики показателей за
болеваемости с построенной кривой (трендом). Чем больше был R2 (максимальный показатель –
1,0), тем более объективно был построен тренд. Конгруэнтность (согласованность) сравнивае
мых трендов заболеваемости оценивали при по
мощи коэффициента корреляции Пирсона. При
показателях коэффициента корреляции 0,30–
0,70 связь считали умеренной, более 0,70 – силь
ной. Положительная статистически значимая
связь свидетельствовала об однонаправленном
влиянии одинаковых факторов, отрицательная
связь – о влиянии разных факторов или разнона
правленном действии одинаковых факторов. ваемости оценивали с помощью анализа ди
намических рядов и расчета полиномиального
тренда второго порядка [1]. Силу связи показа
телей полиномиального тренда исследовали при
помощи коэффициента детерминации (R2), кото
рый определял связь динамики показателей за
болеваемости с построенной кривой (трендом). Чем больше был R2 (максимальный показатель –
1,0), тем более объективно был построен тренд. ваемости оценивали с помощью анализа ди
намических рядов и расчета полиномиального
тренда второго порядка [1]. Силу связи показа
телей полиномиального тренда исследовали при
помощи коэффициента детерминации (R2), кото
рый определял связь динамики показателей за
болеваемости с построенной кривой (трендом). Чем больше был R2 (максимальный показатель –
1,0), тем более объективно был построен тренд. ship of the morbidity over time with the curve
(trend) presented. Higher R2 (the maximum
value is 1.0) suggests greater objectivity of
the trend. The congruence (consistency) of the mor
bidity trends compared was assessed using
the Pearson correlation coefficients. With
correlation coefficients of 0.30–0.70, the
correlation was considered moderate, more
than 0.70 – strong. A positive statistically sig
nificant relationship suggested the unidirec
tional influence of similar factors, a negative
relationship showed the influence of different
factors or the multidirectional action of similar
factors Конгруэнтность (согласованность) сравнивае
мых трендов заболеваемости оценивали при по
мощи коэффициента корреляции Пирсона. При
показателях коэффициента корреляции 0,30–
0,70 связь считали умеренной, более 0,70 – силь
ной. Положительная статистически значимая
связь свидетельствовала об однонаправленном
влиянии одинаковых факторов, отрицательная
связь – о влиянии разных факторов или разнона
правленном действии одинаковых факторов. Results and Discussion General morbidity. The average an
nual morbidity rates for the conscripts of
Russian Armed Forces in 2003–2018 were
(1839.1 ± 50.5) ‰, Navy (1833.3 ± 84.9) ‰,
Ground Forces (2008.0 ± 102.4) ‰, i. e. an
nually almost every conscript turned twice to
the military medical service for medical advice
(Table 2). There are no statistically significant
differences in the general morbidity rates be
tween the cohorts under study. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Наибольшие показатели общей заболевае
мости были у военнослужащих по призыву ВМФ
России с X, XII, I и XI классами болезней с уров
нем (974,7 ± 64,7), (284,7 ± 19,3), (113,5 ± 9,2)
и (102,1 ± 6,5) ‰ соответственно, у военнослу
жащих по призыву Сухопутных войск России –
с X, XII, XI и I классами болезней – (914,9 ± 87,9),
(529,3 ± 43,7), (107,2 ± 6,6) и (104,8 ± 8,1) ‰ со
ответственно (см. табл. 2). Оказалось, что у воен The leading diseases categories (general
morbidity structure contribution of 5 % and
more): in the Navy conscripts, X, XII, I and XI
(increasing share) with overall contribution of MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 19 иологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Таблица 2. Показатели общей заболеваемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 2. Результаты и их анализ General morbidity in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
89.0 ± 5.0
113.5 ± 9.2
6.19
3
104.8 ± 8.1
5.22
4
0.05
II
4.2 ± 0.2
6.6 ± 0.5
0.36
14
3.7 ± 0.3
0.18
14
0.001
0.001
III
1.0 ± 0.1
1.5 ± 0.1
0.08
15
1.1 ± 0.1
0.05
15
0.001
0.05
IV
45.8 ± 2.3
40.9 ± 5.9
2.23
8
43.4 ± 3.4
2.16
7
V
19.4 ± 0.9
41.9 ± 2.8
2.28
6
18.7 ± 1.1
0.93
12
0.001
0.001
VI
27.4 ± 0.9
22.2 ± 1.9
1.21
13
27.9 ± 1.7
1.39
10
0.05
0.05
VII
40.9 ± 2.0
33.3 ± 1.8
1.82
10
39.1 ± 2.2
1.95
8
0.05
0.05
VIII
42.6 ± 2.7
33.1 ± 1.6
1.81
11
47.0 ± 4.7
2.34
6
0.01
0.05
IX
34.0 ± 1.7
35.3 ± 1.7
1.92
9
36.5 ± 2.9
1.82
9
X
844.4 ± 47.7
974.7 ± 64.7
53.17
1
914.9 ± 87.9
45.55
1
XI
105.3 ± 4.3
102.1 ± 6.5
5.57
4
107.2 ± 6.6
5.34
3
XII
430.0 ± 29.3
284.7 ± 19.3
15.53
2
529.3 ± 43.7
26.36
2
0.001
0.001
XIII
98.7 ± 7.0
76.3 ± 5.5
4.16
5
92.5 ± 11.1
4.61
5
0.05
XIV
27.5 ± 1.0
26.2 ± 1.6
1.43
12
25.9 ± 1.4
1.29
11
XIX
28.9 ± 3.7
41.0 ± 6.1
2.24
7
16.3 ± 1.4
0.81
13
0.01
0.01
Общий /
Total
1839.1 ± 50.5 1833.3 ± 84.9
100.00
2008.0 ±
102.4
100.00
Здесь и в табл. 2–15: полужирным шрифтом выделены классы (нозологии) с 1–5-м рангом значимости в структуре. Tables 2–15: categories ranking 1–5 in the disease structure are shown in bold. Таблица 2. Показатели общей заболеваемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Здесь и в табл. 2–15: полужирным шрифтом выделены классы (нозологии) с
Tables 2–15: categories ranking 1–5 in the disease structure are shown in bold. Результаты и их анализ General morbidity structure (left) and general morbidity structure over time (right)
in conscripts of the Russian Navy (%). России – те же самые с общим вкладом 82,5 %
с иной долей значимости – X, XII, XI и I (рис. 3, слева). XI diseases, with relative stability of category
XII diseases (see Fig. 3, right). В динамике структуры общей заболеваемо
сти военнослужащих по призыву ВМФ России по
ведущим классам демонстрируется увеличение
доли с болезнями X класса и уменьшение – с бо
лезнями I, XI и XII классов (см. рис. 2, справа). Primary morbidity. Annual primary mor
bidity rates were (1,044.7 ± 50.6) ‰ in the
Armed Forces conscripts, (1,019.9 ± 54.8) ‰
in the Navy conscripts, (1,014.0 ± 79.2) ‰
in the Ground Forces conscripts, i. e. every
conscript newly diagnosed annually (Table
3). There were no statistically significant dif
ferences in primary morbidity rates between
military cohorts under study. В динамике структуры общей заболеваемости
военнослужащих по призыву Сухопутных войск
России по ведущим классам выявлены практиче
ски те же самые тенденции (см. рис. 3): увеличение
доли с болезнями X класса, уменьшение – с болез
нями I и XI классов и определенная стабильность –
с болезнями XII класса (см. рис. 3, справа). Primary morbidity polynomial trends for
the Navy and Ground Forces conscripts show
an increase with low determination coeffi
cients (R2 = 0.49 and R2 = 0.53, respective
ly) (Fig. 4). Primary morbidity data in Russian
Navy and Ground Forces conscripts are mod
erately and significantly consistent (r = 0.63;
p < 0.01), suggesting contribution of unidi
rectional factors (see Fig. 4). Первичная заболеваемость. Уровень пер
вичной заболеваемости в 2003–2018 гг. военно
служащих по призыву ВМФ России был (1019,9 ±
54,8) ‰, Сухопутных войск России – (1014,0 ±
79,2) ‰, ВС России – (1044,7 ± 50,6) ‰, т. е. еже
годно каждому военнослужащему по призыву вы
ставляли новый диагноз (табл. 3). Статистически
достоверных различий уровней первичной забо
леваемости в анализируемых когортах военно
служащих по призыву нет. The highest primary morbidity rates in
the Navy conscripts were in ICD‑10 catego
ries X, XII and I: (582.7 ± 46.7), (133.2 ± 6.0)
and (72.2 ± 4.7) ‰, respectively; in Ground
Forces conscripts in ICD‑10 categories X,
XII and I: (619.9 ± 69.3), (204.3 ± 12.9) and Полиномиальные тренды первичной заболе
ваемости военнослужащих по призыву ВМФ Рос Рис. 3. Результаты и их анализ Здесь и в табл. 2–15: полужирным шрифтом выделены классы (нозологии) с 1–5-м рангом значимости в структуре. Tables 2–15: categories ranking 1–5 in the disease structure are shown in bold. 80.5 % (Fig. 2, left); in Ground Forces con
scripts, similar categories with overall con
tribution of 82.5 % and different contribution
shares – X. XII, XI and I (Fig. 3, left). нослужащих по призыву ВМФ России уровень
общей заболеваемости с болезнями и травмами
II, III, V и XIX классов был статистически значимо
больше, чем у военнослужащих по призыву Су
хопутных войск России, и меньше – с VI, VII, VIII
и XII классов (см. табл. 2). The general morbidity structure in the Navy
conscripts over time shows category X dis
eases increase and categories I, XI and XII de
crease (see Fig. 2, right). Ведущими классами болезней, которые име
ли вклад в структуру общей заболеваемости 5 %
и более, у военнослужащих по призыву ВМФ Рос
сии были X, XII, I и XI (расположены по возраста
нию доли) с общим вкладом 80,5 % (рис. 2, слева),
у военнослужащих по призыву Сухопутных войск The general morbidity structure in the
Ground Forces conscripts over time shows
similar trends (see Fig. 3): increase in catego
ry X diseases and decrease in category I and Рис. 1. Динамика показателей общей заболеваемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 1. General morbidity in conscripts of Russian Navy (left) and Ground Forces (right) over time ( ‰). Ф с. 1. Динамика показателей общей заболеваемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 1. General morbidity in conscripts of Russian Navy (left) and Ground Forces (right) over time ( ‰). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 20 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Рис. 2. Структура показателей общей заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 2. General morbidity structure (left) and general morbidity structure over time (right)
in conscripts of the Russian Navy (%). Рис. 2. Структура показателей общей заболеваемости (слева) и динамика структуры (справ
у военнослужащих по призыву ВМФ России (%). Fig. 2. General morbidity structure (left) and general morbidity structure over time (right)
in conscripts of the Russian Navy (%). Fig. 2. Результаты и их анализ Primary morbidity profile in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
61.8 ± 2.9
72.2 ± 4.7
7.08
3
75.1 ± 5.6
6.63
3
II
2.3 ± 0.1
3.8 ± 0.3
0.38
14
2.0 ± ± 0.1
0.18
14
0.001
0.001
III
0.6 ± 0.0
0.9 ± 0.1
0.08
15
0.6 ± 0.1
0.05
15
0.001
0.01
IV
18.2 ± 0.7
13.5 ± 1.0
1.32
12
18.9 ± 1.0
1.67
8
0.01
0.01
V
12.6 ± 0.4
25.7 ± 1.6
2.52
6
13.1 ± 0.8
1.16
12
0.001
0.001
VI
13.8 ± 0.6
10.1 ± 0.9
0.99
13
14.7 ± 1.3
1.3
10
0.01
0.01
VII
21.3 ± 1.5
16.4 ± 1.2
1.61
10
20.1 ± 1.4
1.77
7
0.05
VIII
21.8 ± 1.0
17.4 ± 0.5
1.71
9
23.1 ± 1.6
2.04
6
0.01
0.01
IX
16.7 ± 0.8
17.9 ± 1.4
1.75
8
18.4 ± 1.5
1.63
9
X
569.3 ± 45.8
582.7 ± 46.7
57.15
1
619.9 ± 69.3
54.68
1
XI
52.1 ± 2.3
50.7 ± 3.1
4.97
4
54.0 ± 3.9
4.76
4
XII
176.5 ± 7.3
133.2 ± 6.0
13.06
2
204.3 ± 12.9
18.02
2
0.001
0.001
XIII
46.2 ± 4.1
36.3 ± 2.8
3.56
5
43.5 ± 5.4
3.84
5
XIV
14.5 ± 0.6
13.9 ± 1.0
1.36
11
13.8 ± 0.8
1.22
11
XIX
17.1 ± 1.8
25.1 ± 2.9
2.46
7
11.9 ± 1.0
1.05
13
0.05
0.001
0.05
Общий /
Total
1044.7 ± 50.6 1019.9 ± 54.8
100.00
1014.0 ± 79.2
100.00 Таблица 3. Показатели первичной заболеваемости военнослужащих по призыву ВС России, ВМФ России
и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 3. Результаты и их анализ Primary morbidity profile in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
61.8 ± 2.9
72.2 ± 4.7
7.08
3
75.1 ± 5.6
6.63
3
II
2.3 ± 0.1
3.8 ± 0.3
0.38
14
2.0 ± ± 0.1
0.18
14
0.001
0.001
III
0.6 ± 0.0
0.9 ± 0.1
0.08
15
0.6 ± 0.1
0.05
15
0.001
0.01
IV
18.2 ± 0.7
13.5 ± 1.0
1.32
12
18.9 ± 1.0
1.67
8
0.01
0.01
V
12.6 ± 0.4
25.7 ± 1.6
2.52
6
13.1 ± 0.8
1.16
12
0.001
0.001
VI
13.8 ± 0.6
10.1 ± 0.9
0.99
13
14.7 ± 1.3
1.3
10
0.01
0.01
VII
21.3 ± 1.5
16.4 ± 1.2
1.61
10
20.1 ± 1.4
1.77
7
0.05
VIII
21.8 ± 1.0
17.4 ± 0.5
1.71
9
23.1 ± 1.6
2.04
6
0.01
0.01
IX
16.7 ± 0.8
17.9 ± 1.4
1.75
8
18.4 ± 1.5
1.63
9
X
569.3 ± 45.8
582.7 ± 46.7
57.15
1
619.9 ± 69.3
54.68
1
XI
52.1 ± 2.3
50.7 ± 3.1
4.97
4
54.0 ± 3.9
4.76
4
XII
176.5 ± 7.3
133.2 ± 6.0
13.06
2
204.3 ± 12.9
18.02
2
0.001
0.001
XIII
46.2 ± 4.1
36.3 ± 2.8
3.56
5
43.5 ± 5.4
3.84
5
XIV
14.5 ± 0.6
13.9 ± 1.0
1.36
11
13.8 ± 0.8
1.22
11
XIX
17.1 ± 1.8
25.1 ± 2.9
2.46
7
11.9 ± 1.0
1.05
13
0.05
0.001
0.05
Общий /
Total
1044.7 ± 50.6 1019.9 ± 54.8
100.00
1014.0 ± 79.2
100.00 Таблица 3. Показатели первичной заболеваемости военнослужащих по призыву ВС России, ВМФ России
и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. сии и Сухопутных войск России при невысоких
коэффициентах детерминации (R2 = 0,49 и R2 =
0,53 соответственно) показывают тенденцию уве
личения показателей (рис. 4). Результаты и их анализ Структура показателей общей заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 3. General morbidity structure (left) and primary morbidity structure over time (right)
in conscripts of the Ground Forces (%). Рис. 3. Структура показателей общей заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 3. General morbidity structure (left) and primary morbidity structure over time (right)
in conscripts of the Ground Forces (%). Fig. 3. General morbidity structure (left) and primary morbidity structure over time (right)
in conscripts of the Ground Forces (%). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 21 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № сии и Сухопутных войск России при невысоких
коэффициентах детерминации (R2 = 0,49 и R2 =
0,53 соответственно) показывают тенденцию уве
личения показателей (рис. 4). Согласованность
кривых показателей первичной заболеваемости
военнослужащих по призыву ВМФ России и Су
хопутных войск России – умеренная и статисти
чески достоверная (r = 0,63; p < 0,01), что может
указывать на участие в формировании заболевае
мости однонаправленных факторов (см. рис. 4). Наибольшие показатели первичной заболе
ваемости были у военнослужащих по призыву
ВМФ России с X, XII и I классами болезней с уров
нем (582,7 ± 46,7), (133,2 ± 6,0) и (72,2 ± 4,7) ‰
соответственно, у военнослужащих по призыву
(75.1 ± 5.6) ‰, respectively (see Table 3). Pri
mary morbidity rates in ICD‑10 categories II, III,
V and XIX were statistically significantly higher
in the Navy conscripts compared to Ground
Forces conscripts, and lower in ICD‑10 cate
gories IV, VI, VIII and XII (see Table 3). The leading diseases in primary morbidity
structure for the Navy conscripts were from
categories X, XII, I and XI (share-related or
der) with overall contribution of 82.3 % (Fig. 5,
left); in Ground Forces conscripts the leading
disease categories were similar (except for
category XI injuries), with overall contribution
of 79.3 % to the primary morbidity structure
(Fig. 6, left). Таблица 3. Показатели первичной заболеваемости военнослужащих по призыву ВС России, ВМФ России
и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 3. Результаты и их анализ Согласованность
кривых показателей первичной заболеваемости
военнослужащих по призыву ВМФ России и Су
хопутных войск России – умеренная и статисти
чески достоверная (r = 0,63; p < 0,01), что может
указывать на участие в формировании заболевае
мости однонаправленных факторов (см. рис. 4). сии и Сухопутных войск России при невысоких
коэффициентах детерминации (R2 = 0,49 и R2 =
0,53 соответственно) показывают тенденцию уве
личения показателей (рис. 4). Согласованность
кривых показателей первичной заболеваемости
военнослужащих по призыву ВМФ России и Су
хопутных войск России – умеренная и статисти
чески достоверная (r = 0,63; p < 0,01), что может
указывать на участие в формировании заболевае
мости однонаправленных факторов (см. рис. 4). (75.1 ± 5.6) ‰, respectively (see Table 3). Pri
mary morbidity rates in ICD‑10 categories II, III,
V and XIX were statistically significantly higher
in the Navy conscripts compared to Ground
Forces conscripts, and lower in ICD‑10 cate
gories IV, VI, VIII and XII (see Table 3). The leading diseases in primary morbidity
structure for the Navy conscripts were from
categories X, XII, I and XI (share-related or
der) with overall contribution of 82.3 % (Fig. 5,
left); in Ground Forces conscripts the leading
disease categories were similar (except for
category XI injuries), with overall contribution
of 79.3 % to the primary morbidity structure
(Fig. 6, left). Наибольшие показатели первичной заболе
ваемости были у военнослужащих по призыву
ВМФ России с X, XII и I классами болезней с уров
нем (582,7 ± 46,7), (133,2 ± 6,0) и (72,2 ± 4,7) ‰
соответственно, у военнослужащих по призыву Рис. 4. Динамика показателей первичной заболеваемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 4. Primary morbidity in conscripts of Russian Navy (left) and Ground Forces (right) over time ( ‰). 4. Динамика показателей первичной заболеваемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 4. Primary morbidity in conscripts of Russian Navy (left) and Ground Forces (right) over time ( ‰). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 22 иологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2 Сухопутных войск России – также с X, XII и I клас
сами болезней – (619,9 ± 69,3), (204,3 ± 12,9)
и (75,1 ± 5,6) ‰ соответственно (см. табл. 3). Результаты и их анализ Уровень
первичной
заболеваемости
военно
служащих по призыву ВМФ России с болезнями
и травмами II, III, V и XIX классов был статистиче
ски значимо больше, чем у военнослужащих по
призыву Сухопутных войск России, и меньше –
с болезнями IV, VI, VIII и XII классов (см. табл. 3). Primary morbidity structure over time
shows increase in category X, decrease in
categories I and XII with certain stability in
category XI diseases among Navy conscripts
(see Fig. 5, right). Primary morbidity structure over time
also shows increase in category X diseases
and decrease in categories I and XII among
Ground Forces conscripts (see Fig. 6, right). Ведущими классами первичной заболевае
мости военнослужащих по призыву ВМФ России
стали X, XII, I и XI (расположены по величине доли)
с общим вкладом 82,3 % (рис. 5, слева), у воен
нослужащих по призыву Сухопутных войск Рос
сии ведущие классы болезней совпали (за исклю
чением травм XI класса), их суммарная доля была
79,3 % от структуры первичной заболеваемости
(рис. 6, слева). Table 4 shows ICD‑10 blocks with ≥
0.5 % share in the primary morbidity struc
ture. Among the Navy conscripts, there were
18 such blocks with overall contribution of
66.7 %; among the Ground Forces conscripts,
there were 14 such blocks with overall con
tribution of 79.5 %. ICD‑10 blocks with share
< 0.5 % were not included and were ranked
“>”. The leading 14 ICD‑10 blocks were sim
ilar in Navy and Ground Forces conscripts. В динамике структуры первичной заболевае
мости по ведущим классам наглядно видны уве
личение доли военнослужащих по призыву ВМФ
России с болезнями X класса, уменьшение – с бо
лезнями I и XII классов, определенная стабиль
ность – с болезнями XI класса (см. рис. 5, справа). В динамике структуры первичной заболевае
мости по ведущим классам наглядно видны уве
личение доли военнослужащих по призыву ВМФ
России с болезнями X класса, уменьшение – с бо
лезнями I и XII классов, определенная стабиль
ность – с болезнями XI класса (см. рис. 5, справа). В динамике структуры первичной заболевае
мости военнослужащих по призыву Сухопутных
войск России с ведущими классами также отмеча
ется увеличение доли болезней X класса, умень
шение – болезней I и XII классов (см. рис. 6, слева). В табл. 4 сведены нозологии (группы в классах),
доля которых в структуре первичной заболевае
мости была 0,5 % и более. Результаты и их анализ У военнослужащих по
призыву ВМФ России таких нозологий оказалось
18 с вкладом в структуру 66,7 %, у военнослужа
щих по призыву Сухопутных войск России – 14
с долей 79,5 %. Нозологии с вкладом менее 0,5 %
в ранжировании не участвовали, им присваивали
ранг со значком «более» (>). 14 ведущих нозоло
гий у военнослужащих по призыву ВМФ России
и Сухопутных войск России были одинаковыми. y
p
In the structure of primary morbidity
among the in Navy conscripts, acute upper
respiratory infections (J00–J06 by ICD‑10)
ranked 1st with share of 46.3 % and rate
(472.2 ± 41.1) ‰; infections of the skin and
subcutaneous tissue (L00–L08) ranked 2nd
(8.6 % and (87.2 ± 5.8) ‰, respectively); in
fluenza and pneumonia (J10–J18) ranked
3rd (4.6 % and (46.9 ± 4.6) ‰, respectively);
other acute lower respiratory infections (J20–
J22) ranked 4th (4.1 % and (41.5 ± 3.1) ‰,
respectively); viral infections characterized
by skin and mucous membrane lesions (B00–
B09) ranked 5th (3.2 % and (32.3 ± 4.2) ‰,
respectively) (see Table 4). These 5 code
blocks made up 66.7 % of the whole struc
ture of primary morbidity among the Navy
conscripts. В динамике структуры первичной заболевае
мости военнослужащих по призыву Сухопутных
войск России с ведущими классами также отмеча
ется увеличение доли болезней X класса, умень
шение – болезней I и XII классов (см. рис. 6, слева). В табл. 4 сведены нозологии (группы в классах),
доля которых в структуре первичной заболевае
мости была 0,5 % и более. У военнослужащих по
призыву ВМФ России таких нозологий оказалось
18 с вкладом в структуру 66,7 %, у военнослужа
щих по призыву Сухопутных войск России – 14
с долей 79,5 %. Нозологии с вкладом менее 0,5 %
в ранжировании не участвовали, им присваивали
ранг со значком «более» (>). 14 ведущих нозоло
гий у военнослужащих по призыву ВМФ России
и Сухопутных войск России были одинаковыми. In the structure of primary morbidity
among the Ground Forces conscripts, acute
upper respiratory infections (J00–J06 by
ICD‑10) ranked 1st with share of 39 % and
rate (441.6 ± 53.8) ‰; infections of the У военнослужащих по призыву ВМФ России
в структуре первичной заболеваемости 1-й ранг Рис. 5. Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 5. Primary morbidity structure (left) and primary morbidity structure over time (right) in conscripts of Russian Navy (%). Рис. 5. Результаты и их анализ Показатели указанных
5 нозологий в общей сложности составили 66,7 %
от всей структуры первичной заболеваемости
военнослужащих по призыву ВМФ России. занимали показатели острых респираторных
инфекций верхних дыхательных путей (J00–
J06) с долей 46,3 % и уровнем (472,2 ± 41,1) ‰,
2-й – инфекций кожи и подкожной клетчатки
(L00–L08) – 8,6 % и (87,2 ± 5,8) ‰ соответствен
но, 3-й – гриппа и пневмонии (J10–J18) – 4,6 %
и (46,9 ± 4,6) ‰ соответственно, 4-й – других
острых респираторных инфекций нижних дыха
тельных путей (J20–J22) – 4,1 % и (41,5 ± 3,1) ‰
соответственно, 5-й – вирусных инфекций, харак
теризующихся поражениями кожи и слизистых
оболочек (B00–B09),– 3,2 % и (32,3 ± 4,2) ‰ соот
ветственно (см. табл. 4). Показатели указанных
5 нозологий в общей сложности составили 66,7 %
от всей структуры первичной заболеваемости
военнослужащих по призыву ВМФ России. Compared to the Ground Forces con
scripts, the structure of primary morbidity
included statistically significantly more Navy
conscripts with neurotic, stress-related and
somatoform disorders (F40–F48), disorders
of adult personality and behaviour (F60–
F69), diseases of appendix (К35–К38) and
less with mycoses (B35–B49), malnutrition
(E40–E64), diseases of external ear (H60–
H62), hypertensive diseases (I10–I15), other
acute lower respiratory infections (J20–J22),
gastritis and duodenitis (K29), infections of
the skin and subcutaneous tissue (L00–L08),
deforming dorsopathies (M40–M43), in
cluding spinal osteochondrosis (M42) (see
Table 4). У военнослужащих по призыву Сухопутных
войск России в структуре первичной заболевае
мости 1-й ранг занимали показатели острых
респираторных
инфекций
верхних
дыхатель
ных путей (J00–J06) с долей 39 % и уровнем
(441,6 ± 53,8) ‰, 2-й – инфекций кожи и подкож
ной клетчатки (L00–L08) – 12,7 % и (144,1 ± 11,4) ‰
соответственно, 3-й – других острых респира
торных инфекций нижних дыхательных путей
(J20–J22) – 8,5 % и (96,6 ± 13,9) ‰ соответствен
но, 4-й – гриппа и пневмонии (J10–J18) – 4,3 %
и (49,1 ± 5,3) ‰ соответственно, 5-й – болезней
пищевода, желудка и двенадцатиперстной кишки
(K20–К31) – 3,3 % и (37,7 ± 3,1) ‰ соответствен
но (см. табл. 4). Показатели перечисленных 5 но
зологий в общей сложности составили 67,9 % от
всей структуры первичной заболеваемости воен
нослужащих по призыву Сухопутных войск России. Case follow-up. Annual need for case
follow-up in the conscripts of Russian Armed
Forces was (124.2 ± 7.3) ‰, Navy (166.5
± 19.0) ‰, Ground Forces (128.2 ± 8.1) ‰,
i. e. Результаты и их анализ Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Рис. 5. Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Рис. 5. Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig 5 Primary morbidity structure (left) and primary morbidity structure over time (right) in conscripts of Russian Navy (% imary morbidity structure (left) and primary morbidity structure over time (right) in conscripts of Russian Navy (%). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 23 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Рис. 6. Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 6. Primary morbidity structure (left) and primary morbidity structure over time (right)
in conscripts of Russian Ground Forces (%). ис. 6. Структура показателей первичной заболеваемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 6. Primary morbidity structure (left) and primary morbidity structure over time (right)
in conscripts of Russian Ground Forces (%). skin and subcutaneous tissue (L00–L08)
ranked 2nd (12.7 % and (144.1 ± 11.4) ‰,
respectively); other acute lower respiratory
infections (J20–J22) ranked 3rd (8.5 % and
(96.6 ± 13.9) ‰, respectively); influenza and
pneumonia (J10–J18) ranked 4th (4.3 % and
(49.1 ± 5.3) ‰, respectively); diseases of
oesophagus, stomach and duodenum (K20–
К31) ranked 5th (3.3 % and (37.7 ± 3.1) ‰,
respectively) (see Table 4). These 5 code
blocks made up 67.9 % of the whole struc
ture of primary morbidity among the Ground
Forces conscripts. занимали показатели острых респираторных
инфекций верхних дыхательных путей (J00–
J06) с долей 46,3 % и уровнем (472,2 ± 41,1) ‰,
2-й – инфекций кожи и подкожной клетчатки
(L00–L08) – 8,6 % и (87,2 ± 5,8) ‰ соответствен
но, 3-й – гриппа и пневмонии (J10–J18) – 4,6 %
и (46,9 ± 4,6) ‰ соответственно, 4-й – других
острых респираторных инфекций нижних дыха
тельных путей (J20–J22) – 4,1 % и (41,5 ± 3,1) ‰
соответственно, 5-й – вирусных инфекций, харак
теризующихся поражениями кожи и слизистых
оболочек (B00–B09),– 3,2 % и (32,3 ± 4,2) ‰ соот
ветственно (см. табл. 4). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ every year each 6th Navy conscript and
each 8th conscript of Russian Armed Forces
and Ground Forces was followed up (Table 5). There were no statistically significant differ
ences in case follow-up between the cohorts
of conscripts. У военнослужащих по призыву ВМФ России по
сравнению с военнослужащими по призыву Су
хопутных войск России уровень первичной забо
леваемости был статистически достоверно боль
ше с невротическими, связанными со стрессом,
и соматоформными расстройствами (F40–F48),
расстройствами поведения в зрелом возрас Polynomial trend of the need for case fol
low-up shows an increase in the Navy con 24 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № Таблица 4. Уровень первичной заболеваемости военнослужащих по призыву ВМФ России и Сухопутных войск России
по основным болезням (группам в классах) (‰)
Table 4. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ 2019. № 3 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № те (F60–F69), болезнями аппендикса (К35–К38)
и меньше – с микозами (B35–B49), недостаточ
ностью питания (E40–E64), болезнями наружного
уха (H60–H62), болезнями, характеризующими
ся повышенным кровяным давлением (I10–I15),
другими острыми респираторными инфекциями
нижних дыхательных путей (J20–J22), гастритом
и дуоденитом (K29), инфекциями кожи и подкож
ной клетчатки (L00–L08), деформирующими дор
сопатиями (M40–M43), в том числе остеохондро
зом позвоночника (M42) (см. табл. 4). scripts with low determination coefficient
(R2 = 0.10), and in the Ground Forces con
scripts shows a decrease with low determi
nation coefficient (R2 = 0.12) (Fig. 7). Consis
tency between curves of need for follow-up
among the Navy and Ground Forces con
scripts is low (r = 0.04; p > 0.05), suggesting
different factors and effects (see Fig. 7) The highest need for a case follow-up was
in the Navy conscripts: ICD‑10 categories X,
XI and I with (86.9 ± 11.0), (17.8 ± 2.1) and
(12.5 ± 1.7) ‰, respectively; in the Ground
Forces conscripts, ICD‑10 categories X, XI
and IV with (66.4 ± 7.0), (15.4 ± 1.1) and
(10.7 ± 1.0) ‰. Диспансерное наблюдение. Уровень дис
пансерного наблюдения в 2003–2018 гг. воен
нослужащих по призыву ВМФ России был
(166,5 ± 19,0) ‰, Сухопутных войск России –
(128,2 ± 8,1) ‰, ВС России – (124,2 ± 7,3) ‰, т. е. ежегодно каждый шестой военнослужащий по
призыву ВМФ России и каждый восьмой ВС Рос
сии и Сухопутных войск России находился под
диспансерным наблюдением (табл. 5). Статисти
чески значимых различий в показателях диспан
серного наблюдения между когортами военно
служащих по призыву нет. Case follow-up rates were statistically sig
nificantly higher among the Navy conscripts
with diseases from categories II, III, V and
XII and lower with diseases from category IV
than among the Ground Forces conscripts
(see Table 5). The leading disease categories for case
follow-up in the Navy conscripts were X, XI,
I and XII (share-related order) with overall
contribution of 79.9 % (Fig. 8, left); in the
Ground Forces conscripts, X, XI, VI and I with
overall contribution of 79.1 % (Fig. 9, left). Of
note, Armed Forces and Ground Forces con
scripts had similar most significant disease
categories ranking 1st, 2nd and 5th (see
Table 5). 26
MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Полиномиальный тренд нуждаемости в дис
пансерном наблюдении военнослужащих по при
зыву ВМФ России при низком коэффициенте де
терминации (R2 = 0,10) показывает увеличение
данных, военнослужащих по призыву Сухопутных
войск России при низком коэффициенте детер
минации (R2 = 0,12) – демонстрирует уменьше
ние показателей (рис. 7). Согласованность кри Таблица 5. Показатели нуждаемости в диспансерном наблюдении военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 5. Need for case follow-up in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
7.5 ± 0.9
12.5 ± 1.7
7.52
3
8.9 ± 1.6
6.97
4
0.05
II
0.5 ± 0.0
1.1 ± 0.2
0.64
14
0.4 ± 0.1
0.28
14
0.001
0.01
III
0.2 ± 0.0
0.4 ± 0.0
0.22
15
0.1 ± 0.0
0.11
15
0.001
0.001
IV
9.9 ± 0.8
7.3 ± 0.8
4.39
6
10.7 ± 1.0
8.34
3
0.05
0.05
V
2.2 ± 0.2
4.6 ± 0.7
2.79
8
2.2 ± 0.3
1.72
11
0.01
0.01
VI
2.8 ± 0.2
2.5 ± 0.4
1.50
10
2.6 ± 0.2
2.02
8
VII
1.7 ± 0.2
1.8 ± 0.2
1.06
13
1.7 ± 0.2
1.30
12
VIII
2.1 ± 0.1
2.5 ± 0.5
1.47
11
2.3 ± 0.2
1.77
10
IX
5.2 ± 0.4
5.4 ± 0.6
3.25
7
5.1 ± 0.5
4.06
6
X
62.6 ± 5.3
86.9 ± 11.0
52.19
1
66.4 ± 7.0
51.78
1
XI
15.8 ± 0.9
17.8 ± 2.1
10.66
2
15.4 ± 1.1
12.04
2
XII
4.7 ± 0.7
10.9 ± 2.8
6.55
4
3.3 ± 0.6
2.60
7
0.05
0.05
XIII
5.5 ± 0.4
8.0 ± 1.2
4.83
5
5.4 ± 0.8
4.20
5
XIV
2.4 ± 0.2
2.8 ± 0.5
1.70
9
2.6 ± 0.2
2.00
9
XIX
1.3 ± 0.2
2.1 ± 0.5
1.23
12
1.1 ± 0.2
0.86
13
Общий /
Total
124.2 ± 7.3
166.5 ± 19.0
100.00
128.2 ± 8.1
100.00 Таблица 5. Результаты и их анализ Primary morbidity rates in Russian Navy and Ground Forces conscripts (ICD‑10 blocks) ( ‰)
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Кишечные инфекции / Intestinal infectious diseases (A00–A09)
9.2 ± 0.8
0.90
11
7.8 ± 1.6
0.69
13
Другие бактериальные болезни (менингит, скарлатина) / Other bacterial diseases (meningitis, scarlatina)
(A30–A49)
5.2 ± 0.6
0.51
18
2.7 ± 0.3
0.24
>15
Вирусные инфекции, характеризующиеся поражениями кожи и слизистых оболочек / Viral infections
characterized by skin and mucous membrane lesions (B00–B09)
32.3 ± 4.2
3.17
5
36.5 ± 4.9
3.22
6
Микозы / Mycoses (B35–B49)
9.9 ± 0.8
0.98
9
19.2 ± 2.5
1.70
7
0.01
Недостаточность питания / Malnutrition (E40–E64)
11.9 ± 0.9
1.17
8
16.8 ± 1.0
1.48
8
0.01
Невротические, связанные со стрессом, и соматоформные расстройства / Neurotic, stress-related and
somatoform disorders (F40–F48)
18.3 ± 1.8
1.80
7
4.3 ± 0.5
0.38
>15
0.001
Расстройства поведения в зрелом возрасте / Disorders of adult personality and behaviour (F60–F69)
5.4 ± 0.7
0.53
17
2.8 ± 0.5
0.25
>15
0.01
Болезни век, слезных путей, глазницы, конъюнктивы / Disorders of eyelid, lacrimal system and orbit (H00–H11)
9.8 ± 0.7
0.96
10
8.5 ± 1.3
0.75
11
Болезни наружного уха / Diseases of external ear (H60–H62)
7.8 ± 0.4
0.77
13
14.1 ± 1.2
1.25
9
0.001
Болезни среднего уха и сосцевидного отростка / Diseases of middle ear and mastoid (H65–H74)
8.8 ± 0.4
0.86
12
7.8 ± 0.4
0.69
12
Болезни, характеризующиеся повышенным кровяным давлением / Hypertensive diseases (I10–I15)
2.3 ± 0.5
0.22
>19
7.1 ± 0.8
0.63
14
0.001
Острые респираторные инфекции верхних дыхательных путей / Acute upper respiratory infections (J00–J06)
472.2 ± 41.1 46.30
1
441.6 ± 53.8 38.96
1
Грипп и пневмония / Influenza and pneumonia (J10–J18)
46.9 ± 4.6
4.60
3
49.1 ± 5.3
4.33
4
Другие острые респираторные инфекции нижних дыхательных путей / Other acute lower respiratory infections
(J20–J22)
41.5 ± 3.1
4.07
4
96.6 ± 13.9
8.52
3
0.01
Болезни пищевода, желудка и двенадцатиперстной кишки (K20–К31), в том числе/ Diseases of oesophagus,
stomach and duodenum (K20–К31), incl:
31.4 ± 2.1
3.07
6
37.7 ± 3.1
3.33
5
гастрит и дуоденит / gastritis and duodenitis (K29)
16.1 ± 1.3
1.58
22.3 ± 1.8
1.97
0.05
Болезни аппендикса / Diseases of appendix (К35–К38)
5.4 ± 0.4
0.53
16
3.6 ± 0.2
0.32
>15
0.001
Инфекции кожи и подкожной клетчатки / Infections of the skin and subcutaneous tissue (L00–L08)
87.2 ± 5.8
8.55
2
144.1 ± 11.4 12.71
2
0.001
Деформирующие дорсопатии / Deforming dorsopathies (M40–M43), в том числе / incl.:
7.4 ± 0.8
0.73
14
13.9 ± 1.6
1.23
10
0.01
остеохондроз позвоночника / spinal osteochondrosis (M42)
3.6 ± 0.5
0.35
8.8 ± 1.1
0.78
0.001
Болезни мужских половых органов / Diseases of male genital organs (N40–N50)
6.0 ± 0.5
0.59
15
5.2 ± 0.4
0.46
>15 25 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Результаты и их анализ Annual hospital
admission rates were (946.2 ± 51.5),
(968.5 ± 71.3) and (1033.5 ± 89.6) ‰, re
spectively, in the conscripts of Russian
Armed Forces, Navy and Ground Forces (i. e. annually each conscript underwent in-hospi
tal treatment or examination) (Table 6). There
were no statistically significant differences in
hospital admission rates between cohorts of
conscripts. Уровень диспансерного наблюдения у воен
нослужащих по призыву ВМФ России с болезня
ми II, III, V и XII классов был статистически досто
верно больше, чем у военнослужащих по призыву
Сухопутных войск России, а с болезнями IV клас
са – меньше (см. табл. 5). Ведущими классами болезней, определивши
ми диспансерное наблюдение военнослужащих
по призыву ВМФ России, стали X, XI, I и XII (рас
положены по величине доли) с общим вкладом
79,9 % (рис. 8, слева), у военнослужащих по при
зыву Сухопутных войск России – X, XI, VI и I с до
лей 79,1 % от структуры (рис. 9, слева). Уместно
указать, что классы болезней с 1-, 2-м и 5-м ран
гом значимости у военнослужащих по призыву
ВМФ и Сухопутных войск России были одинако
выми (см. табл. 5). Polynomial trends in hospital admission of
the Navy and Ground Forces conscripts show
an increase with determination coefficients of
various significance (R2 = 0.34 and R2 = 0.60,
respectively) (Fig. 10). Consistency of trends between the Navy
and Ground Forces conscripts is moderately
positive and statistically significant (r = 0.58;
p < 0.05), suggesting a role of unidirectional
(similar) factors (see Fig. 10). У военнослужащих по призыву ВМФ России
в динамике структуры диспансерного наблюде
ния по ведущим классам отмечается увеличение
доли пациентов с болезнями X класса, уменьше
ние – с болезнями I и XI классов. С болезнями
XII класса выявлено уменьшение доли в середине
периода наблюдения и увеличение в последний
период (см. рис. 8, справа). In the Navy conscripts, the highest
rates of in-patient treatment and exam
ination for ICD‑10 categories X, XII and
I were (561.5 ± 51.5), (114.8 ± 6.7) and
(68.2 ± 6.0) ‰, respectively; in the Ground
Forces conscripts for the same ICD‑10 cat
egories – (620.9 ± 71.5), (139.0 ± 10.5) and 27
Medico Biological and Socio Psychological Problems of Safety in Emergency Situations 2019 N 3
Рис. 7. Динамика показателей нуждаемости в диспансерном наблюдении военнослужащих по призыву
ВМФ России (слева) и Сухопутных войск России (справа) ( ‰). Fig. 7. Результаты и их анализ Показатели нуждаемости в диспансерном наблюдении военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. T bl
5 N
d f
f ll
i
i t
f R
i
A
d F
N
d G
d F
b ICD 10 i
2003 2018 аблица 5. Показатели нуждаемости в диспансерном наблюдении военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. 26 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № вых показателей нуждаемости в диспансерном
наблюдении военнослужащих по призыву ВМФ
России и Сухопутных войск России – низкая
(r = 0,04; p > 0,05), что может указывать на влия
ние разных факторов в формировании диспан
серного наблюдения (см. рис. 7) вых показателей нуждаемости в диспансерном
наблюдении военнослужащих по призыву ВМФ
России и Сухопутных войск России – низкая
(r = 0,04; p > 0,05), что может указывать на влия
ние разных факторов в формировании диспан
серного наблюдения (см. рис. 7) Among the Navy conscripts, the structure
of case follow-up over time showed increase
in the diseases from ICD‑10 category X, de
crease in the categories I and XI. For catego
ry XII diseases, there was a decrease in the
middle of the observation period with most
recent increase (see Fig. 8, right). Наибольшие показатели диспансерного на
блюдения были у военнослужащих по призыву
ВМФ России с X, XI и I классами болезней с уров
нем (86,9 ± 11,0), (17,8 ± 2,1) и (12,5 ± 1,7) ‰
соответственно, у военнослужащих по призы
ву Сухопутных войск России – также с X, XI и IV
классами болезней – (66,4 ± 7,0), (15,4 ± 1,1)
и (10,7 ± 1,0) ‰ соответственно. Among the Ground Forces conscripts, the
structure of case follow-up over time showed
increase in the diseases from ICD‑10 catego
ries IV and XI with decrease in the categories
I and X (see Fig. 9, right). Hospital admission. Annual hospital
admission rates were (946.2 ± 51.5),
(968.5 ± 71.3) and (1033.5 ± 89.6) ‰, re
spectively, in the conscripts of Russian
Armed Forces, Navy and Ground Forces (i. e. annually each conscript underwent in-hospi
tal treatment or examination) (Table 6). There
were no statistically significant differences in
hospital admission rates between cohorts of
conscripts. Hospital admission. Результаты и их анализ Need for case follow-up in conscripts of Navy (left) and Ground Forces (right) over time ( ‰). Рис. 7. Динамика показателей нуждаемости в диспансерном наблюдении военнослужащих по призыву
ВМФ России (слева) и Сухопутных войск России (справа) ( ‰). Fig. 7. Need for case follow-up in conscripts of Navy (left) and Ground Forces (right) over time ( ‰). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 27 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Рис. 8. Структура показателей диспансерного наблюдения (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 8. Case follow-up structure (left) and case follow-up structure over time (right) in Navy conscripts (%). Рис. 8. Структура показателей диспансерного наблюдения (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). у военнослужащих по призыву ВМФ России (%). Fig. 8. Case follow-up structure (left) and case follow-up structure over time (right) in Navy conscripts (%). Fig. 8. Case follow-up structure (left) and case follow-up structure over time (right) in Navy conscripts (%). Fig. 8. Case follow-up structure (left) and case follow-up structure over time (right) in Navy conscripts (%). Рис. 9. Структура показателей диспансерного наблюдения (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 9. Case follow-up structure (left) and case follow-up structure over time (right) in Ground Forces conscripts (%).. Рис. 9. Структура показателей диспансерного наблюдения (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Рис. 9. Структура показателей диспансерного наблюдения (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 9. Case follow-up structure (left) and case follow-up structure over time (right) in Ground Forces conscripts (%).. у военнослужащих по призыву Сухопутных войск России (%). Fig. 9. Case follow-up structure (left) and case follow-up structure over time (right) in Ground Forces conscripts (% w-up structure (left) and case follow-up structure over time (right) in Ground Forces conscripts (%).. В динамике структуры диспансерного наблю
дения по ведущим классам у военнослужащих по
призыву Сухопутных войск России выявлено уве
личение доли болезней IV и XI классов, уменьше
ние – I и X классов (см. рис. 9, справа). (72.0 ± 5.4) ‰. The above categories includ
ed more than 2/3 of all the in-patient con
scripts – 76.9 and 80.5 %, respectively. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Hos
pital admission rates in the Navy conscripts
with diseases from categories II, III, V and
category XIX injuries were statistically signifi
cantly higher than in the conscripts of Ground
Forces, and with diseases from categories IV,
VI, VIII – lower (see Table 6). Госпитализация. Уровень госпитализации
в 2003–2018 гг. военнослужащих по призыву ВМФ
России был (968,5 ± 71,3) ‰, Сухопутных войск
России – (1033,5 ± 89,6) ‰, ВС России – (946,2 ±
51,5) ‰, т. е. ежегодно каждый военнослужащий
по призыву находился на стационарном лечении
или обследовании (табл. 6). Статистически значи
мых различий в показателях госпитализации меж
ду когортами военнослужащих по призыву нет. In the structure of hospital admissions,
most cases in the Navy conscripts were re
lated to categories X, XII, I and XI (according
to their significance) with overall contribution
of 82.4 % (Fig. 11, left), in the conscripts of
Ground Forces – with diseases from catego
ries X, XII and I and a structure share of 80.5 %
(Fig. 12, left). Полиномиальные тренды госпитализации
военнослужащих по призыву ВМФ России и Су
хопутных войск России при разных по значимо
сти коэффициентах детерминации (R2 = 0,34
и R2 = 0,60 соответственно) показывают рост дан
ных (рис. 10). In the structure of hospital admissions
over time for the Navy conscripts, diseases
from categories X and other increase, while
diseases from categories I, XI and XII de
crease (see Fig. 11, right). Согласованность
трендов
госпитализации
военнослужащих по призыву ВМФ России и Су
хопутных войск России – умеренная, положи
тельная и статистически достоверная (r = 0,58;
p < 0,05), что может указывать на участие в фор
мировании госпитализации однонаправленных
(одинаковых) факторов (см. рис. 10). In the structure of hospital admissions
over time for the Ground Forces conscripts,
diseases from category X increase, while dis
eases from categories I and XII decrease (see
Fig. 12, right). 28 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2 Таблица 6. Показатели госпитализации военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 6. Результаты и их анализ Hospital admissions in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
59.6 ± 2.7
68.2 ± 6.0
7.04
3
72.0 ± 5.4
6.97
3
II
2.1 ± 0.1
4.2 ± 0.7
0.43
14
1.8 ± 0.1
0.17
14
0.01
0.01
III
0.5 ± 0.0
0.9 ± 0.1
0.10
15
0.5 ± 0.1
0.05
15
0.001
0.01
IV
13.4 ± 1.1
11.7 ± 0.8
1.20
12
15.3 ± 1.2
1.48
9
0.05
V
12.4 ± 0.5
26.1 ± 1.5
2.70
6
12.9 ± 0.7
1.25
10
0.001
0.001
VI
11.4 ± 0.5
9.2 ± 0.8
0.95
13
12.2 ± 1.2
1.18
11
0.05
0.05
VII
14.7 ± 1.0
13.0 ± 1.0
1.34
11
15.5 ± 1.4
1.50
8
VIII
19.4 ± 1.1
15.2 ± 0.7
1.56
9
21.6 ± 1.6
2.09
6
0.01
0.01
IX
15.6 ± 0.7
19.3 ± 1.4
2.00
8
17.1 ± 1.4
1.66
7
0.05
X
562.1 ± 45.8
561.5 ± 51.5
57.99
1
620.9 ± 71.5
60.08
1
XI
48.6 ± 2.0
53.9 ± 3.5
5.56
4
49.6 ± 3.8
4.80
4
XII
126.4 ± 5.8
114.8 ± 6.7
11.85
2
139.0 ± 10.5
13.45
2
XIII
32.2 ± 3.3
35.0 ± 4.2
3.61
5
31.3 ± 4.6
3.03
5
XIV
12.7 ± 0.5
14.4 ± 1.1
1.49
10
12.2 ± 0.6
1.18
12
XIX
15.1 ± 1.4
21.1 ± 1.8
2.18
7
11.4 ± 0.9
1.10
13
0.05
0.001
0.05
Общий /
Total
946.2 ± 51.5
968.5 ± 71.3
100.0
1033.5 ± 89.6
100.00 Таблица 6. Показатели госпитализации военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Результаты и их анализ Наибольшие показатели стационарного лече
ния и обследования были у военнослужащих по
призыву ВМФ России с болезнями X, XII и I клас
сов – (561,5 ± 51,5), (114,8 ± 6,7) и (68,2 ± 6,0) ‰
соответственно, у военнослужащих по призы
ву Сухопутных войск России с теми же клас
сами болезней – (620,9 ± 71,5), (139,0 ± 10,5)
и (72,0 ± 5,4) ‰ соответственно. Перечисленные
болезни были у 2/3 госпитализированных воен
нослужащих по призыву – 76,9 и 80,5 % соответ
ственно. Уровень госпитализации военнослужа
щих по призыву ВМФ России с болезнями II, III,
V классов и травмами XIX класса был статистиче
ски достоверно больше, чем военнослужащих по
призыву Сухопутных войск России, а с болезня
ми IV, VI, VIII классов – меньше (см. табл. 6). In the Navy conscripts, there were 18
ICD‑10 code blocks with ≥ 0.5 % share in the
structure with overall contribution of 80.5 %;
in the Ground Forces conscripts, there were
14 such code blocks with overall contribu
tion of 82 %. Thirteen leading diseases were
similar in the conscripts of the cohorts under
study (Table 7). In the structure of hospital admissions for
the Navy conscripts, acute upper respiratory
infections ranked 1st (share 46.9 % and rate
(454.2 ± 43.4) ‰); infections of the skin and
subcutaneous tissue (L00–L08) ranked 2nd
(7.6 % and (73.5 ± 3.6) ‰, respectively); in
fluenza and pneumonia ranked 3rd (4.8 %
and (46.2 ± 4.8) ‰); other acute lower respi 29
Medico Biological and Socio Psychological Problems of Safety in Emergency Situations 2019 N 3
Рис. 10. Динамика показателей госпитализации военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 10. Hospital admissions of Navy (left) and Ground Forces conscripts (right) over time ( ‰).. Fig. 10. Hospital admissions of Navy (left) and Ground Forces conscripts (right) over time ( ‰).. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 29 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2 У военнослужащих по призыву ВМФ России
в структуре госпитализации ведущие места за
нимали пациенты с болезнями X, XII, I и XI классов
(указаны по значимости) с общим вкладом 82,4 %
(рис. Результаты и их анализ 11, слева), у военнослужащих по призы
ву Сухопутных войск России – с болезнями X, XII
и I классов с долей 80,5 % от структуры (рис. 12,
слева). ratory infections (J20–J22) ranked 4th (4.3 %
and (42.0 ± 3.4) ‰); diseases of oesopha
gus, stomach and duodenum (K20–К31)
ranked 5th (3.5 % and (34.0 ± 2.3) ‰). The
above 5 disease blocks made up 67.1 % of
the structure of hospital admissions for the
Navy conscripts (see Table 7). In the structure of hospital admissions for
the Ground Forces conscripts, four most sig
nificant diseases were similar to the Navy con
scripts, e. g. acute upper respiratory infec
tions ranked 1st (share 43.2 % and rate (446.1
± 56.0) ‰); infections of the skin and subcu
taneous tissue (L00–L08) ranked 2nd (9.6 %
and (99.0 ± 8.7) ‰, respectively); other acute
lower respiratory infections (J20–J22) ranked
3rd (9.5 % and (98.3 ± 13.8) ‰); influenza
and pneumonia ranked 4th (4.8 % and (49.5 ±
5.4) ‰); viral infections characterized by skin
and mucous membrane lesions (B00–B09)
ranked 5th (3.6 % and (36.8 ± 4.9) ‰). The
above 5 disease blocks made up 70.6 % of
the structure of hospital admissions for the
Ground Forces conscripts (see Table 7). В динамике структуры госпитализации по
ведущим классам отмечается увеличение доли
военнослужащих по призыву ВМФ России с бо
лезнями X класса и прочих, уменьшение – с бо
лезнями I, XI и XII классов (см. рис. 11, справа). В динамике структуры госпитализации по
ведущим классам выявлено увеличение доли
военнослужащих по призыву Сухопутных войск
России с болезнями X класса, уменьшение – с бо
лезнями I и XII классов (см. рис. 12, справа). У военнослужащих по призыву ВМФ России
ведущих нозологий (групп в классах), доля ко
торых в структуру госпитализации была 0,5 %
и более, оказалось 18 с суммарным вкладом
в структуру 80,5 %, у военнослужащих по призы
ву Сухопутных войск России – 14 с долей 82 %. 13 ведущих нозологий в сравниваемых когортах
военнослужащих по призыву совпали (табл. 7). Compared to the Ground Forces con
scripts, hospital admissions for the Navy Рис. 11. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 11. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Navy (%). 30
MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3
Рис. 11. Результаты и их анализ Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 11. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Navy (%). Рис. 12. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 12. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Ground Forces (%) Рис. 11. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Рис. 11. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 11. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Navy (%). Fig. 11. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Navy (%). Рис. 12. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 12. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Ground Forces (%) Рис. 12. Структура показателей госпитализации (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 12. Hospital admissions structure (left) and hospital admissions structure over time (right)
in conscripts of Russian Ground Forces (%) MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 30 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № Таблица 7. Уровень госпитализации военнослужащих по призыву ВМФ России и Сухопутных войск России по основным болезням (группам в классах) ( ‰)
Table 7. Результаты и их анализ 2019. № В структуре госпитализации военнослужащих
по призыву ВМФ России 1-й ранг составили по
казатели острых респираторных инфекций верх
них дыхательных путей с долей 46,9 % и уровнем
(454,2 ± 43,4) ‰, 2-й ранг – инфекций кожи и под
кожной клетчатки (L00–L08) – 7,6 % и (73,5 ± 3,6) ‰
соответственно, 3-й – гриппа и пневмонии – 4,8 %
и (46,2 ± 4,8) ‰ соответственно, 4-й – других
острых респираторных инфекций нижних дыха
тельных путей (J20–J22) – 4,3 % и (42,0 ± 3,4) ‰
соответственно, 5-й – болезней пищевода, желуд
ка и двенадцатиперстной кишки (K20–К31) – 3,5 %
и (34,0 ± 2,3) ‰ соответственно. Данные пере
численных 5 нозологий в сумме составили 67,1 %
от структуры госпитализации военнослужащих по
призыву ВМФ России (см. табл. 7). conscripts with mycoses (B35–B49), mal
nutrition (E40–E64), diseases of external ear
(H60–H62), hypertensive diseases (I10–I15),
other acute lower respiratory infections (J20–
J22), infections of the skin and subcutaneous
tissue (L00–L08), deforming dorsopathies
(M40–M43), including spinal osteochon
drosis (M42), were less frequent, and with
neurotic, stress-related and somatoform
disorders (F40–F48), other cardiac diseases
(endocarditis, myocarditis) (I30–I52), diseas
es of appendix (К35–К38), diseases of male
genital organs (N40–N50) were more fre
quent (see Table 7). Work days lost. Annual morbidity with
temporary disability in the Russian Armed
Forces conscripts was (10,299 ± 340) ‰,
Navy conscripts – (13,166 ± 7.99) ‰, Ground
Forces conscripts – (11,104 ± 595) ‰; i.e. every year each conscript had 10–13 work
days lost (Table 8). The Navy conscripts had
almost 1.2-fold more work days lost than
Ground Forces conscripts (p < 0.01). Work days lost. Annual morbidity with
temporary disability in the Russian Armed
Forces conscripts was (10,299 ± 340) ‰,
Navy conscripts – (13,166 ± 7.99) ‰, Ground
Forces conscripts – (11,104 ± 595) ‰; i.e. every year each conscript had 10–13 work
days lost (Table 8). The Navy conscripts had
almost 1.2-fold more work days lost than
Ground Forces conscripts (p < 0.01). Результаты и их анализ В структуре госпитализации военнослужащих
по призыву Сухопутных войск России болезни,
включенные в первые четыре ранга значимости,
были одинаковыми, например, 1-й ранг занимали
данные острых респираторных инфекций верх
них дыхательных путей с долей 43,2 % и уровнем
(446,1 ± 56,0) ‰, 2-й – инфекций кожи и подкож
ной клетчатки (L00–L08) – 9,6 % и (99,0 ± 8,7) ‰
соответственно, 3-й – других острых респиратор
ных инфекций нижних дыхательных путей (J20–
J22) – 9,5 % и (98,3 ± 13,8) ‰ соответственно,
4-й – гриппа и пневмонии – 4,8 % и (49,5 ± 5,4) ‰
соответственно, 5-й – вирусных инфекций, ха
рактеризующихся поражением кожи и слизистых
оболочек (B00–B09),– 3,6 % и (36,8 ± 4,9) ‰ соот
ветственно. В сумме показатели указанных 5 но
зологий составили 70,6 % от структуры госпита
лизации военнослужащих по призыву Сухопутных
войск России (см. табл. 7). Polynomial trends for work days lost in the
Navy and Ground Forces conscripts show
a decrease with low determination coeffi
cients (Fig. 13). Trends are moderately and
statistically significantly consistent (r = 0.58;
p < 0.05), suggesting a role of unidirectional
(similar) factors. The highest numbers of work days lost
in the Navy conscripts were related to the
diseases from ICD‑10 categories X, XII
and I ((6,052 ± 537), (1,500 ± 72) and
(1042 ± 81) ‰, respectively) with overall
contribution of 65.3 %; in the conscripts
of Ground Forces ICD‑10 categories were
similar ((6,060 ± 573), (1,483 ± 134) and
(855 ± 66) ‰, respectively) with overall con
tribution of 75.6 %. The Navy conscripts had
less work days lost due to diseases from most
categories (II, III, V, VI, VII, IX, XI, XIII, XIV) and
category XIX injuries compared to Ground
Forces conscripts (see Table 8). (
)
В структуре госпитализации военнослужа
щих по призыву ВМФ России было меньше, чем
военнослужащих по призыву Сухопутных войск
России, пациентов с микозами (B35–B49), не
достаточностью питания (E40–E64), болезнями
наружного уха (H60–H62), болезнями, характери
зующимися повышенным кровяным давлением
(I10–I15), другими острыми респираторными ин
фекциями нижних дыхательных путей (J20–J22),
инфекциями кожи и подкожной клетчатки (L00–
L08), деформирующими дорсопатиями (M40–
M43), в том числе с остеохондрозом позво
ночника (M42), и больше – с невротическими,
связанными со стрессом, и соматоформными
расстройствами (F40–F48), другими болезнями
сердца (эндокардитом, миокардитом) (I30–I52),
болезнями аппендикса (К35–К38), болезнями
мужских половых органов (N40–N50) (см. табл. 7). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Hospital admissions in Russian Navy and Ground Forces conscripts by main diseases (ICD‑10 blocks) ( ‰)
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Кишечные инфекции / Intestinal infectious diseases (A00–A09)
8.8 ± 0.9
0.91
10
8.4 ± 1.7
0.81
11
Другие бактериальные болезни (менингит, скарлатина) / Other bacterial diseases (meningitis, scarlatina)
(A30–A49)
8.9 ± 4.6
0.92
9
4.2 ± 2.1
0.41
>15
Вирусные инфекции, характеризующиеся поражениями кожи и слизистых оболочек / Viral infections
characterized by skin and mucous membrane lesions (B00–B09)
30.5 ± 4.6
3.15
6
36.8 ± 4.9
3.56
5
Микозы / Mycoses (B35–B49)
8.3 ± 0.7
0.86
12
15.4 ± 2.4
1.49
7
0.05
Недостаточность питания / Malnutrition (E40–E64)
10.0 ± 0.8
1.03
8
14.2 ± 1.2
1.37
8
0.05
Невротические, связанные со стрессом, и соматоформные расстройства / Neurotic, stress-related and
somatoform disorders (F40–F48)
18.3 ± 1.7
1.89
7
4.3 ± 0.6
0.42
>15
0.001
Болезни век, слезных путей, глазницы, конъюнктивы / Disorders of eyelid, lacrimal system and orbit
(H00–H11)
7.8 ± 0.6
0.81
13
6.7 ± 1.0
0.65
14
Болезни наружного уха / Diseases of external ear (H60–H62)
5.7 ± 0.4
0.59
15
12.6 ± 1.1
1.22
9
0.001
Болезни среднего уха и сосцевидного отростка / Diseases of middle ear and mastoid (H65–H74)
8.7 ± 0.5
0.9
11
8.0 ± 0.5
0.77
12
Болезни, характеризующиеся повышенным кровяным давлением / Hypertensive diseases (I10–I15)
2.1 ± 0.5
0.21
>19
6.9 ± 0.7
0.66
13
0.001
Другие болезни сердца (эндокардит, миокардит) / Other cardiac diseases (endocarditis, myocarditis) (I30–I52)
5.0 ± 0.4
0.51
18
1.7 ± 0.2
0.17
>15
0.001
Острые респираторные инфекции верхних дыхательных путей / Acute upper respiratory infections (J00–J06)
454.2 ± 43.4 46.89
1
446.1 ± 56.0 43.16
1
Грипп и пневмония / Influenza and pneumonia (J10–J18)
46.2 ± 4.8
4.77
3
49.5 ± 5.4
4.79
4
Другие острые респираторные инфекции нижних дыхательных путей/ Other acute lower respiratory infections
(J20–J22)
42.0 ± 3.4
4.34
4
98.3 ± 13.8
9.51
3
0.01
Болезни пищевода, желудка и двенадцатиперстной кишки (K20–К31), в том числе / Diseases of oesophagus,
stomach and duodenum (K20–К31), incl:
34.0 ± 2.3
3.51
5
34.8 ± 3.1
3.37
6
гастрит и дуоденит / gastritis and duodenitis (K29)
19.7 ± 1.4
2.03
22.2 ± 2.1
2.14
Болезни аппендикса / Diseases of appendix (К35–К38)
5.4 ± 0.5
0.56
17
3.5 ± 0.2
0.34
>15
0.01
Инфекции кожи и подкожной клетчатки / Infections of the skin and subcutaneous tissue (L00–L08)
73.5 ± 3.6
7.59
2
99.0 ± 8.7
9.57
2
0.05
Деформирующие дорсопатии / Deforming dorsopathies (M40–M43), в том числе / incl:
5.7 ± 0.7
0.59
16
10.6 ± 1.4
1.03
10
0.01
остеохондроз позвоночника / spinal osteochondrosis (M42)
3.0 ± 0.5
0.31
7.2 ± 1.0
0.69
0.01
Болезни мужских половых органов / Diseases of male genital organs (N40–N50)
6.2 ± 0.4
0.64
14
4.7 ± 0.4
0.45
>15
0.05 31 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Результаты и их анализ Полиномиальные тренды дней трудопотерь
у военнослужащих по призыву ВМФ России и Су
хопутных войск России при невысоких коэффи
циентах детерминации показывают увеличение
данных (рис. 13). Согласованность трендов –
умеренная и статистически достоверная (r = 0,58;
p < 0,05), что может указывать на участие в фор
мировании трудопотерь однонаправленных (оди
наковых) факторов. In the structure of work days lost for the
Navy conscripts, acute upper respiratory in
fections ranked 1st (share 32.2 % and rate
(4,245 ± 419) ‰); influenza and pneumo
nia (J10–J18) ranked 2nd (7.1 % and (937 ±
95) ‰); infections of the skin and subcutane
ous (L00–L08) ranked 3rd (6.9 % and (911 ±
40) ‰); neurotic, stress-related and somato
form disorders (F40–F48) ranked 4th (4.7 %
and (618 ± 58) ‰); other acute lower respi
ratory infections (J20–J22) ranked 5th (4.6 %
and (601 ± 41) ‰) (see Table 9). The above 5
code blocks made up 55.5 % of the structure
of work days lost for the Navy conscripts. Наибольшие показатели дней трудопотерь
были у военнослужащих по призыву ВМФ Рос
сии с болезнями X, XII и I классов – (6052 ± 537),
(1500 ± 72) и (1042 ± 81) ‰ соответственно
с вкладом в структуру 65,3 %, у военнослужащих
по призыву Сухопутных войск России – точно
с такими же классами болезней – (6060 ± 573),
(1483 ± 134) и (855 ± 66) ‰ соответственно с
вкладом в структуру 75,6 %. Уровень дней трудо
потерь у военнослужащих по призыву ВМФ Рос
сии с болезнями большинства классов (II, III, V, VI,
VII, IX, XI, XIII, XIV) и травмами XIX класса был боль
ше, чем у военнослужащих по призыву Сухопут
ных войск России (см. табл. 8). In the structure of work days lost for the
Ground Forces conscripts, four most signi
ficant disease categories were similar, e. g. 33
Medico Biological and Socio Ps chological Problems of Safet in Emergenc Sit ations 2019 N 3
Таблица 8. Показатели дней трудопотерь у военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 8. Результаты и их анализ В структуре госпитализации военнослужа
щих по призыву ВМФ России было меньше, чем
военнослужащих по призыву Сухопутных войск
России, пациентов с микозами (B35–B49), не
достаточностью питания (E40–E64), болезнями
наружного уха (H60–H62), болезнями, характери
зующимися повышенным кровяным давлением
(I10–I15), другими острыми респираторными ин
фекциями нижних дыхательных путей (J20–J22),
инфекциями кожи и подкожной клетчатки (L00–
L08), деформирующими дорсопатиями (M40–
M43), в том числе с остеохондрозом позво
ночника (M42), и больше – с невротическими,
связанными со стрессом, и соматоформными
расстройствами (F40–F48), другими болезнями
сердца (эндокардитом, миокардитом) (I30–I52),
болезнями аппендикса (К35–К38), болезнями
мужских половых органов (N40–N50) (см. табл. 7). Трудопотери
Уровень
дней
трудопотерь In the structure of work days lost for the
Navy conscripts, the leading diseases were
from categories X, XII, I, V, XI and XIII (according
to their significance) with overall contribution
of 84.3 % (Fig. 14, left); in the Ground Forces
conscripts – from categories X, XII and XI with
overall contribution of 79.7 % (Fig. 15, left). The structure of work days lost in the Navy
conscripts over time shows increase in dis
eases from ICD‑10 categories X and XIII and
decrease in diseases from other leading cat
egories (see Fig. 14, right). Трудопотери. Уровень дней трудопотерь
у военнослужащих по призыву ВМФ России был
(13 166 ± 7,99) ‰, Сухопутных войск России – 32 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуация (11 104 ± 595) ‰, ВС России – (10 299 ± 340) ‰,
т. е. ежегодно каждый военнослужащий по при
зыву имел по 10–13 дней трудопотерь (табл. 8). Уровень заболеваемости с временными трудопо
терями у военнослужащих по призыву ВМФ Рос
сии был почти в 1,2 раза больше по сравнению с
уровнем у военнослужащих по призыву Сухопут
ных войск России (p < 0,01). The structure of work days lost in the
Ground Forces conscripts over time shows
increase in diseases from ICD‑10 category
X and decrease in diseases from categories
I and XII, with certain stability for category XI
(see Fig. 15, right). In the Navy conscripts, there were 26
ICD‑10 code blocks with ≥ 0.5 % contribution
to the structure of work days lost, with over
all contribution of 78 %; in the Ground Forces
conscripts, there were 19 such ICD‑10 code
blocks with contribution of 80.6 %. Eighteen
leading disease categories were similar (Ta
ble 9). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Work days lost in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
727 ± 30
1042 ± 81
7.91
3
855 ± 66
7.70
3
0.01
II
35 ± 3
63 ± 6
0.48
14
35 ± 7
0.32
14
0.001
0.01
III
11 ± 1
29 ± 3
0.22
15
9 ± 1
0.08
15
0.001
0.001
0.05
IV
172 ± 11
216 ± 14
1.64
11
186 ± 15
1.67
10
0.05
V
359 ± 15
921 ± 56
6.99
4
325 ± 22
2.93
6
0.001
0.001
VI
151 ± 7
188 ± 9
1.43
13
147 ± 13
1.32
13
0.01
0.05
VII
158 ± 6
197 ± 12
1.50
12
159 ± 9
1.43
12
0.01
0.05
VIII
204 ± 9
218 ± 13
1.65
10
217 ± 13
1.96
9
IX
225 ± 10
397 ± 29
3.02
8
221 ± 17
1.99
8
0.001
0.001
X
5385 ± 359
6052 ± 537
45.97
1
6060 ± 573
54.58
1
XI
626 ± 25
904 ± 51
6.86
5
617 ± 47
5.56
4
0.001
0.001
XII
1323 ± 70
1500 ± 72
11.39
2
1483 ± 134
13.36
2
XIII
410 ± 29
675 ± 77
5.13
6
373 ± 39
3.36
5
0.01
0.01
XIV
188 ± 7
271 ± 22
2.06
9
169 ± 8
1.52
11
0.01
0.001
XIX
324 ± 29
494 ± 36
3.75
7
247 ± 19
2.22
7
0.001
0.05
Общий /
Total
10 299 ± 340
13 166 ± 7.99
100.00
11 104 ± 595
100.0
0.01 Таблица 8. Показатели дней трудопотерь у военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. 33 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Рис. 13. Динамика показателей дней трудопотерь у военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 13. Результаты и их анализ Work days lost in conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). ис. 13. Динамика показателей дней трудопотерь у военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 13. Work days lost in conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). У военнослужащих по призыву ВМФ России
в структуре дней трудопотерь ведущими оказа
лись показатели болезней X, XII, I, V, XI и XIII клас
сов (указаны по значимости) с общим вкладом
84,3 % (рис. 14, слева), у военнослужащих по при
зыву Сухопутных войск России – болезней X, XII
и XI классов с долей 79,7 % от структуры (рис. 15,
слева). acute upper respiratory infections ranked
1st (share 33.1 % and rate (3,677 ± 409) ‰);
other acute lower respiratory infections (J20–
J22) ranked 2nd (10.5 % and (1161 ± 171) ‰);
infections of the skin and subcutaneous (L00–
L08) ranked 3rd (9.6 % and (1,064 ± 118) ‰);
influenza and pneumonia (J10–J18) ranked
4th (8.6 % and (956 ± 100) ‰); diseases of
oesophagus, stomach and duodenum (K20–
К31) ranked 5th (4.0 % and (445 ± 44) ‰) acute upper respiratory infections ranked
1st (share 33.1 % and rate (3,677 ± 409) ‰);
other acute lower respiratory infections (J20–
J22) ranked 2nd (10.5 % and (1161 ± 171) ‰);
infections of the skin and subcutaneous (L00–
L08) ranked 3rd (9.6 % and (1,064 ± 118) ‰);
influenza and pneumonia (J10–J18) ranked
4th (8.6 % and (956 ± 100) ‰); diseases of
oesophagus, stomach and duodenum (K20–
К31) ranked 5th (4.0 % and (445 ± 44) ‰) В динамике структуры трудопотерь по веду
щим классам отмечается увеличение доли воен Рис. 14. Структура показателей дней трудопотерь (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 14. Work days lost structure (left) and work days lost structure over time (right) in conscripts of Russian Navy (%). Рис. 14. Структура показателей дней трудопотерь (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). у военнослужащих по призыву ВМФ России (%). Work days lost structure (left) and work days lost structure over time (right) in conscripts of Russian Navy (%). у
у
р
у
(
)
Fig. 14. Work days lost structure (left) and work days lost structure over time (right) in conscripts of Russian Navy (% Fig. 14. Результаты и их анализ У военнослужащих по призыву ВМФ России ве
дущих нозологий (групп в классах), доля которых
в структуру дней трудопотерь была 0,5 % и более,
оказалось 26 с суммарным вкладом в структуру
78 %, у военнослужащих по призыву Сухопутных
войск России – 19 с долей 80,6 %. 18 ведущих но
зологий в анализируемых когортах военнослужа
щих по призыву были одинаковыми (табл. 9). Dismissal. Annual dismissal rates for
health reasons were (17.57 ± 1.19) ‰ in the
Armed Forces conscripts, (33.38 ± 1.79) ‰
in the Navy conscripts and (18.28 ± 1.66) ‰
in the Ground Forces conscripts (Table 10). Dismissal rates in the Navy conscripts with
most diseases were statistically significantly
higher compared to all the conscripts of Rus
sian Armed Forces and Ground Forces (see
Table 10); hence, general dismissal rates
were almost 2-fold higher in the Navy con
scripts (p < 0.001). В структуре дней трудопотерь у военнослу
жащих по призыву ВМФ России 1-й ранг соста
вили показатели острых респираторных инфек
ций верхних дыхательных путей с долей 32,2 %
и уровнем (4245 ± 419) ‰, 2-й ранг – гриппа
и пневмонии (J10–J18) – 7,1 % и (937 ± 95) ‰ со
ответственно, 3-й ранг – инфекций кожи и под
кожной клетчатки (L00–L08) – 6,9 % и (911 ± 40) ‰
соответственно, 4-й – невротических, связанных
со стрессом, и соматоформных расстройств
(F40–F48) – 4,7 % и (618 ± 58) ‰ соответствен
но, 5-й – других острых респираторных инфек
ций нижних дыхательных путей (J20–J22) – 4,6 %
и (601 ± 41) ‰ соответственно (см. табл. 9). По
казатели указанных 5 нозологий в общей сложно
сти составили 55,5 % от структуры дней трудопо
терь у военнослужащих по призыву ВМФ России. Polynomial trends of dismissal rates for
the Navy and Ground Forces conscripts show
a decrease with low determination coeffi
cients (Fig. 16). Dismissal trends for the Navy
and Ground Forces conscripts are moder
ately consistent and approach a statistically
significant difference (r = 0.49; p < 0.1), sug
gesting unidirectional factors, e. g. organiza
tional ones. The Navy conscripts with diseases from
categories V, IX and XI had the highest dis
missal rates (17.69 ± 1.14), (4.87 ± 0.39) and
(1.93 ± 0.14) ‰, respectively); in Ground
Forces conscripts the highest dismissal rates
were related to the same categories ((9.24
± 0.77), (2.69 ± 0.35) and (1.11 ± 0.14) ‰)
(see Table 10). Результаты и их анализ Work days lost structure (left) and work days lost structure over time (right) in conscripts of Russian Navy (%). Рис. 15. Структура показателей дней трудопотерь (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 15. Working days lost structure (left) and working days lost structure over time (right)
in conscripts of Russian Ground Forces (%) Рис. 15. Структура показателей дней трудопотерь (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 15. Working days lost structure (left) and working days lost structure over time (right)
in conscripts of Russian Ground Forces (%) Рис. 15. Структура показателей дней трудопотерь (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig 15 Working days lost structure (left) and working days lost structure over time (right) Fig. 15. Working days lost structure (left) and working days lost structure over time (right)
in conscripts of Russian Ground Forces (%) MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 34 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № нослужащих по призыву ВМФ России с болезня
ми X и XIII классов, уменьшение – с болезнями
других ведущих классов (см. рис. 14, справа). (see Table 9). The above 5 code blocks made
up 65.8 %% of the structure of work days lost
for the Ground Forces conscripts. (see Table 9). The above 5 code blocks made
up 65.8 %% of the structure of work days lost
for the Ground Forces conscripts. Compared to Ground Forces conscripts,
the Navy conscripts had less work days lost
due to diseases of external ear (H60–H62),
hypertensive diseases (I10–I15), other acute
lower respiratory infections (J20–J22) and
more work days lost due to other leading dis
eases (see Table 9). В динамике структуры дней трудопотерь по ве
дущим классам отмечается увеличение доли воен
нослужащих по призыву Сухопутных войск России
с болезнями X класса, уменьшение – с болезнями
I и XII классов и определенная стабильность – с бо
лезнями XI класса (см. рис. 15, справа). Compared to Ground Forces conscripts,
the Navy conscripts had less work days lost
due to diseases of external ear (H60–H62),
hypertensive diseases (I10–I15), other acute
lower respiratory infections (J20–J22) and
more work days lost due to other leading dis
eases (see Table 9). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Dismissal rates in the Navy
conscripts with most disease categories were
statistically significantly higher compared to
Ground Forces conscripts (see Table 10). В структуре дней трудопотерь у военнослужа
щих по призыву Сухопутных войск России первые
четыре ранга значимости занимали те же самые
нозологии, например, 1-й ранг составили показа
тели острых респираторных инфекций верхних ды
хательных путей с долей 33,1 % и уровнем (3677 ±
409) ‰, 2-й – других острых респираторных инфек
ций нижних дыхательных путей (J20–J22) – 10,5 %
и (1161 ± 171) ‰ соответственно, 3-й – инфекций
кожи и подкожной клетчатки (L00–L08) – 9,6 %
и (1064 ± 118) ‰ соответственно, 4-й – гриппа
и пневмонии (J10–J18) – 8,6 % и (956 ± 100) ‰ со
ответственно, 5-й – болезней пищевода, желудка
и двенадцатиперстной кишки (K20–К31) – 4,0 %
и (445 ± 44) ‰ соответственно (см. табл. 9). В сум
ме доля указанных 5 нозологий составила 65,8 %
от структуры трудопотерь у военнослужащих по
призыву Сухопутных войск России. Russian Navy conscripts with diseases
from categories V, XI and IX (according to
their significance, > 5 % share each) made
up 73.4 % of the structure (Fig. 17, left);
Ground Forces conscripts with diseases from
categories V, XI, IX and I made up 76.5 % of
the structure (Fig. 18, left). The structure of dismissal for the Navy
conscripts over time shows an increase in
category V diseases, a decrease in category
XI diseases, with certain stability for category
IX diseases (see Fig. 17, right). По сравнению с военнослужащими по призы
ву Сухопутных войск России уровень дней тру
допотерь у военнослужащих ВМФ России был
меньше с болезнями наружного уха (H60–H62), The structure of dismissal for the Ground
Forces conscripts over time shows an in 35 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № д
ц
р
р
у ц
Таблица 9. Уровень дней трудопотерь у военнослужащих по призыву ВМФ России
и Сухопутных войск России по основным болезням (группам в классах) ( ‰)
Table 9. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Результаты и их анализ Work days lost in Russian Navy and Ground Forces conscripts by main diseases (ICD‑10 blocks) ( ‰)
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Кишечные инфекции / Intestinal infectious diseases (A00–A09)
104 ± 11
0.79
15
88 ± 19
0.80
13
Туберкулез / Tuberculosis (A15–A19)
71 ± 10
0.54
23
55 ± 9
0.50
19
Другие бактериальные болезни (менингит, скарлатина) / Other bacterial diseases (meningitis, scarlatina) (A30–
A49)
69 ± 8
0.53
24
37 ± 4
0.33
>20
0.01
Вирусные инфекции, характеризующиеся поражениями кожи и слизистых оболочек / Viral infections
characterized by skin and mucous membrane lesions (B00–B09)
383 ± 53
2.91
7
401 ± 61
3.61
6
Вирусный гепатит / Viral hepatitis (В15–В19)
81 ± 22
0.61
19
37 ± 14
0.34
>20
Микозы / Mycoses (B35–B49)
143 ± 14
1.09
10
159 ± 19
1.43
8
Недостаточность питания / Malnutrition (E40–E64)
177 ± 13
1.34
9
172 ± 15
1.55
7
Невротические, связанные со стрессом, и соматоформные расстройства / Neurotic, stress-related and
somatoform disorders (F40–F48)
618 ± 58
4.69
4
94 ± 11
0.84
11
0.001
Расстройства поведения в зрелом возрасте / Disorders of adult personality and behaviour (F60–F69)
225 ± 19
1.71
8
88 ± 15
0.79
14
0.001
Поражения отдельных нервов, нервных корешков и сплетений / Nerve, nerve root and plexus disorders (G50–
G58)
77 ± 5
0.58
20
53 ± 4
0.47
>20
0.01
Болезни век, слезных путей, глазницы, конъюнктивы / Disorders of eyelid, lacrimal system and orbit (H00–H11)
107 ± 7
0.81
14
61 ± 8
0.55
17
0.001
Болезни наружного уха / Diseases of external ear (H60–H62)
82 ± 10
0.62
18
114 ± 8
1.03
10
0.05
Болезни среднего уха и сосцевидного отростка / Diseases of middle ear and mastoid (H65–H74)
123 ± 8
0.94
12
90 ± 5
0.81
12
0.01
Болезни, характеризующиеся повышенным кровяным давлением / Hypertensive diseases (I10–I15)
38 ± 8
0.29
>27
77 ± 7
0.70
15
0.01
Другие болезни сердца (эндокардит, миокардит) / Other cardiac diseases (endocarditis, myocarditis) (I30–I52)
139 ± 14
1.06
11
32 ± 4
0.29
>20
0.001
Острые респираторные инфекции верхних дыхательных путей / Acute upper respiratory infections (J00–J06)
4245 ± 419
32.24
1
3677 ± 409
33.12
1
Грипп и пневмония / Influenza and pneumonia (J10–J18)
937 ± 95
7.11
2
956 ± 100
8.61
4
Другие острые респираторные инфекции нижних дыхательных путей / Other acute lower respiratory infections
(J20–J22)
601 ± 41
4.57
5
1161 ± 171
10.45
2
0.01
Болезни пищевода, желудка и двенадцатиперстной кишки (K20–К31), в том числе / Diseases of oesophagus,
stomach and duodenum (K20–К31), incl:
592 ± 35
4.50
6
445 ± 44
4.00
5
0.05
язва двенадцатиперстной кишки / duodenal ulcer (K26)
125 ± 12
0.95
54 ± 17
0.49
0.01
гастрит и дуоденит / gastritis and duodenitis (K29)
323 ± 22
2.45
267 ± 19
2.40
Болезни аппендикса / Diseases of appendix (К35–К38)
74 ± 5
0.56
21–22
45 ± 2
0.41
>20
0.001
Инфекции кожи и подкожной клетчатки / Infections of the skin and subcutaneous tissue (L00–L08)
911 ± 40
6.92
3
1064 ± 118
9.58
3
Деформирующие дорсопатии / Deforming dorsopathies (M40–M43), в том числе / incl:
115 ± 18
0.88
13
133 ± 16
1.20
9
остеохондроз позвоночника / spinal osteochondrosis (M42)
68 ± 15
0.52
93 ± 14
0.84 36 едикобиологические и социальн
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Болезни мужских половых органов / Diseases of male genital organs (N40–N50)
100 ± 8
0.76
16
62 ± 4
0.56
16
0.001
Травмы головы / Injury to the head (S00–S09)
85 ± 11
0.65
17
56 ± 7
0.51
18
0.05
Травмы запястья и кисти / Injuries to the wrist and hand (S60–S69)
74 ± 7
0.56
21–22
33 ± 2
0.30
>20
0.001
Травмы колена и голени / Injuries to the knee and lower leg (S80–S89)
68 ± 4
0.51
25–26
34 ± 3
0.30
>20
0.001
Последствия травм, отравлений и других воздействий внешних причин / Sequelae of injuries, of poisoning and
of other consequences of external causes (T90–T98)
67 ± 7
0.51
25–26
8 ± 2
0.07
>20
0.001 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Окончание таблицы 9 болезнями, характеризующимися повышенным кровяным
давлением (I10–I15), другими острыми респираторными ин
фекциями нижних дыхательных путей (J20–J22) и больше –
с другими ведущими нозологиями (см. табл. 9). Увольняемость. Уровень увольняемости по состоянию
здоровья в 2003–2018 гг. военнослужащих по призыву ВМФ
России был (33,38 ± 1,79) ‰, Сухопутных войск России –
(18,28 ± 1,66) ‰, ВС России – (17,57 ± 1,19) ‰ (табл. 10). Уровень увольняемости военнослужащих по призыву ВМФ
России с большинством классов болезней был статисти
чески значимо больше, чем военнослужащих по призыву
ВС России и Сухопутных войск России (см. табл. 10). Само
собой разумеется, общий уровень увольняемости военно
служащих по призыву ВМФ России оказался почти в 2 раза
больше (p < 0,001). Полиномиальные тренды увольняемости военнослужа
щих по призыву ВМФ России и Сухопутных войск России при
низких коэффициентах детерминации показывают уменьше
ние данных (рис. 16). Согласованность трендов увольняемос
ти военнослужащих по призыву ВМФ России и Сухопутных
войск России – умеренная и приближается к статистически
значимой (r = 0,49; p < 0,1), что может указывать на тенден
цию участия в формировании увольняемости однонаправ
ленных факторов, например организационных. Наибольшие показатели увольняемости военнослужа
щих по призыву ВМФ России были с болезнями V, IX и XI
классов – (17,69 ± 1,14), (4,87 ± 0,39) и (1,93 ± 0,14) ‰, воен
нослужащих Сухопутных войск России – с болезнями анало
гичных классов – (9,24 ± 0,77), (2,69 ± 0,35) и (1,11 ± 0,14) ‰
(см. табл. 10). Уровень увольняемости военнослужащих
по призыву ВМФ России оказался статистически значимо
больше, чем военнослужащих по призыву Сухопутных войск
России по большинству классов болезней (см. табл. 10). С долей более 5 % в структуру увольняемости вошли
военнослужащие по призыву ВМФ России с болезнями V,
XI и IX классов (указаны по значимости) с общим вкладом
73,4 % (рис. 17, слева), военнослужащие по призыву Сухо
путных войск России – с болезнями V, XI, IX и I классов с до
лей 76,5 % (рис. 18, слева). В динамике структуры увольняемости по ведущим
классам отмечается увеличение доли военнослужащих по
призыву ВМФ России с болезнями V класса, уменьшение –
с болезнями XI класса, определенная стабильность – с бо
лезнями IX класса (см. рис. 17, справа). В динамике структуры увольняемости по ведущим
классам отмечается увеличение доли военнослужащих по
призыву ВМФ России с болезнями V класса, уменьшение –
с болезнями XI класса, определенная стабильность – с бо
лезнями IX класса (см. рис. 17, справа). Результаты и их анализ 2019. № Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Болезни мужских половых органов / Diseases of male genital organs (N40–N50)
100 ± 8
0.76
16
62 ± 4
0.56
16
0.001
Травмы головы / Injury to the head (S00–S09)
85 ± 11
0.65
17
56 ± 7
0.51
18
0.05
Травмы запястья и кисти / Injuries to the wrist and hand (S60–S69)
74 ± 7
0.56
21–22
33 ± 2
0.30
>20
0.001
Травмы колена и голени / Injuries to the knee and lower leg (S80–S89)
68 ± 4
0.51
25–26
34 ± 3
0.30
>20
0.001
Последствия травм, отравлений и других воздействий внешних причин / Sequelae of injuries, of poisoning and
of other consequences of external causes (T90–T98)
67 ± 7
0.51
25–26
8 ± 2
0.07
>20
0.001
Окончание таблицы 9 Окончание таблицы 9 Dismissal profiles in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
0.87 ± 0.14
1.50 ± 0.27
4.50
5
0.94 ± 0.14
5.16
4
0.05
0.05
II
0.11 ± 0.01
0.17 ± 0.02
0.51
13
0.12 ± 0.02
0.63
13
0.05
0.001
III
0.09 ± 0.01
0.16 ± 0.03
0.47
14
0.07 ± 0.01
0.4
15
0.05
0.001
IV
0.16 ± 0.01
0.39 ± 0.06
1.17
11
0.14 ± 0.01
0.77
12
0.01
0.05
V
8.69 ± 0.50
17.69 ± 1.14
52.98
1
9.24 ± 0.77
50.58
1
0.001
VI
0.55 ± 0.06
0.79 ± 0.07
2.37
9
0.55 ± 0.06
2.99
9
0.05
0.05
VII
0.22 ± 0.03
0.29 ± 0.04
0.87
12
0.19 ± 0.03
1.01
11
0.001
VIII
0.09 ± 0.02
0.15 ± 0.02
0.45
15
0.08 ± 0.02
0.46
14
0.05
0.001
IX
1.09 ± 0.11
1.93 ± 0.14
5.77
3
1.11 ± 0.14
6.05
3
0.001
0.001
X
0.33 ± 0.03
0.89 ± 0.09
2.65
7
0.29 ± 0.05
1.56
10
0.001
0.01
XI
2.62 ± 0.23
4.87 ± 0.39
14.59
2
2.69 ± 0.35
14.72
2
0.001
0.01
XII
0.67 ± 0.09
1.37 ± 0.18
4.09
6
0.68 ± 0.11
3.72
6
0.01
XIII
0.92 ± 0.08
1.56 ± 0.13
4.68
4
0.90 ± 0.12
4.92
5
0.001
XIV
0.54 ± 0.04
0.77 ± 0.07
2.30
10
0.61 ± 0.06
3.36
8
0.05
0.001
XIX
0.62 ± 0.07
0.87 ± 0.07
2.60
8
0.67 ± 0.11
3.67
7
0.05
Общий /
Total
17.57 ± 1.19
33.38 ± 1.79
100.00
18.28 ± 1.66
100.0
0.001
Рис. 16. Динамика показателей увольняемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 16. Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). Таблица 10. Показатели увольняемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Рис. 16. Окончание таблицы 9 В динамике структуры увольняемости по ведущим клас
сам выявлено увеличение доли военнослужащих по призыву
Сухопутных войск России с болезнями V класса, уменьше
ние – с болезнями I класса, определенная стабильность –
с болезнями IX и XI классов (см. рис. 18, справа). В динамике структуры увольняемости по ведущим клас
сам выявлено увеличение доли военнослужащих по призыву
Сухопутных войск России с болезнями V класса, уменьше
ние – с болезнями I класса, определенная стабильность –
с болезнями IX и XI классов (см. рис. 18, справа). У военнослужащих по призыву ВМФ России ведущих но
зологий (групп в классах), доля которых в структуру уволь
няемости была 0,5 % и более, оказалось 13 с суммарным
вкладом в структуру 78 %, у военнослужащих по призыву
Сухопутных войск России – 15 с долей 67,1 % (табл. 11). У военнослужащих по призыву ВМФ России ведущих но
зологий (групп в классах), доля которых в структуру уволь
няемости была 0,5 % и более, оказалось 13 с суммарным
вкладом в структуру 78 %, у военнослужащих по призыву
Сухопутных войск России – 15 с долей 67,1 % (табл. 11). В структуре увольнений военнослужащих по призыву
ВМФ России 1-й ранг занимали сведения о невротических, В структуре увольнений военнослужащих по призыву
ВМФ России 1-й ранг занимали сведения о невротических, 37 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2 Медико биологические и социально психологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3
Таблица 10. Показатели увольняемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 10. Окончание таблицы 9 Dismissal profiles in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
0.87 ± 0.14
1.50 ± 0.27
4.50
5
0.94 ± 0.14
5.16
4
0.05
0.05
II
0.11 ± 0.01
0.17 ± 0.02
0.51
13
0.12 ± 0.02
0.63
13
0.05
0.001
III
0.09 ± 0.01
0.16 ± 0.03
0.47
14
0.07 ± 0.01
0.4
15
0.05
0.001
IV
0.16 ± 0.01
0.39 ± 0.06
1.17
11
0.14 ± 0.01
0.77
12
0.01
0.05
V
8.69 ± 0.50
17.69 ± 1.14
52.98
1
9.24 ± 0.77
50.58
1
0.001
VI
0.55 ± 0.06
0.79 ± 0.07
2.37
9
0.55 ± 0.06
2.99
9
0.05
0.05
VII
0.22 ± 0.03
0.29 ± 0.04
0.87
12
0.19 ± 0.03
1.01
11
0.001
VIII
0.09 ± 0.02
0.15 ± 0.02
0.45
15
0.08 ± 0.02
0.46
14
0.05
0.001
IX
1.09 ± 0.11
1.93 ± 0.14
5.77
3
1.11 ± 0.14
6.05
3
0.001
0.001
X
0.33 ± 0.03
0.89 ± 0.09
2.65
7
0.29 ± 0.05
1.56
10
0.001
0.01
XI
2.62 ± 0.23
4.87 ± 0.39
14.59
2
2.69 ± 0.35
14.72
2
0.001
0.01
XII
0.67 ± 0.09
1.37 ± 0.18
4.09
6
0.68 ± 0.11
3.72
6
0.01
XIII
0.92 ± 0.08
1.56 ± 0.13
4.68
4
0.90 ± 0.12
4.92
5
0.001
XIV
0.54 ± 0.04
0.77 ± 0.07
2.30
10
0.61 ± 0.06
3.36
8
0.05
0.001
XIX
0.62 ± 0.07
0.87 ± 0.07
2.60
8
0.67 ± 0.11
3.67
7
0.05
Общий /
Total
17.57 ± 1.19
33.38 ± 1.79
100.00
18.28 ± 1.66
100.0
0.001
Рис. 16. Динамика показателей увольняемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 16. Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). Таблица 10. Показатели увольняемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 10. Окончание таблицы 9 Dismissal profiles in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10 in 2003–2018
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
0.87 ± 0.14
1.50 ± 0.27
4.50
5
0.94 ± 0.14
5.16
4
0.05
0.05
II
0.11 ± 0.01
0.17 ± 0.02
0.51
13
0.12 ± 0.02
0.63
13
0.05
0.001
III
0.09 ± 0.01
0.16 ± 0.03
0.47
14
0.07 ± 0.01
0.4
15
0.05
0.001
IV
0.16 ± 0.01
0.39 ± 0.06
1.17
11
0.14 ± 0.01
0.77
12
0.01
0.05
V
8.69 ± 0.50
17.69 ± 1.14
52.98
1
9.24 ± 0.77
50.58
1
0.001
VI
0.55 ± 0.06
0.79 ± 0.07
2.37
9
0.55 ± 0.06
2.99
9
0.05
0.05
VII
0.22 ± 0.03
0.29 ± 0.04
0.87
12
0.19 ± 0.03
1.01
11
0.001
VIII
0.09 ± 0.02
0.15 ± 0.02
0.45
15
0.08 ± 0.02
0.46
14
0.05
0.001
IX
1.09 ± 0.11
1.93 ± 0.14
5.77
3
1.11 ± 0.14
6.05
3
0.001
0.001
X
0.33 ± 0.03
0.89 ± 0.09
2.65
7
0.29 ± 0.05
1.56
10
0.001
0.01
XI
2.62 ± 0.23
4.87 ± 0.39
14.59
2
2.69 ± 0.35
14.72
2
0.001
0.01
XII
0.67 ± 0.09
1.37 ± 0.18
4.09
6
0.68 ± 0.11
3.72
6
0.01
XIII
0.92 ± 0.08
1.56 ± 0.13
4.68
4
0.90 ± 0.12
4.92
5
0.001
XIV
0.54 ± 0.04
0.77 ± 0.07
2.30
10
0.61 ± 0.06
3.36
8
0.05
0.001
XIX
0.62 ± 0.07
0.87 ± 0.07
2.60
8
0.67 ± 0.11
3.67
7
0.05
Общий /
Total
17.57 ± 1.19
33.38 ± 1.79
100.00
18.28 ± 1.66
100.0
0.001 Таблица 10. Показатели увольняемости военнослужащих по призыву ВС России,
ВМФ России и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. Table 10. Окончание таблицы 9 связанных со стрессом, и соматоформных рас
стройствах (F40–F48) с долей 32,2 % и уровнем
(10,75 ± 1,44) ‰, 2-й – расстройствах пове
дения в зрелом возрасте (F60–F69) – 16,9 %
и (5,63 ± 0,72) ‰ соответственно, 3-й ранг – бо
лезнях пищевода, желудка и двенадцатиперст
ной кишки (K20–К31) – 13,5 % и (4,51 ± 0,40) ‰
соответственно, 4-й ранг – вирусном гепатите
(В15–В19) – 2,9 % и (0,95 ± 0,22) ‰ соответ
ственно, 5-й – других болезнях сердца (эндо
кардите, миокардите) (I30–I52) – 2,8 % и (0,92 ±
0,09) ‰ соответственно. Показатели 5 нозо
логий в общей сложности составили 68,2 % от
всей структуры увольняемости военнослужа
щих по призыву ВМФ России (см. табл. 11). In the Navy conscripts, there were 13 ICD‑10
code blocks with ≥ 0.5 % share in the dismiss
al structure with overall contribution of 78 %; in
the Ground Forces conscripts, there also were
15 such ICD‑10 code blocks with overall contri
bution of 67.1 % (Table 11). In the dismissal structure for the Navy con
scripts, neurotic, stress-related and somato
form disorders (F40–F48) ranked 1st (share
32.2 % and rate (10.75 ± 1.44) ‰); disorders
of adult personality and behaviour (F60–F69)
ranked 2nd (16.9 % and (5.63 ± 0.72) ‰); dis
eases of oesophagus, stomach and duode
num (K20–К31) ranked 3rd (13.5 % and (4.51
± 0.40) ‰); viral hepatitis (В15–В19) ranked
4th (2.9 % and (0.95 ± 0.22) ‰); other cardiac
diseases (endocarditis, myocarditis) (I30–I52)
ranked 5th (2.8 % and (0.92 ± 0.09) ‰). The
above 5 disease categories made up 68.2 % of
the dismissal structure for the Navy conscripts
(see Table 11). В структуре увольнений военнослужащих
по призыву Сухопутных войск России 1-й ранг
занимали сведения о невротических, связан
ных со стрессом, и соматоформных расстрой
ствах (F40–F48) с долей 14,8 % и уровнем
(2,70 ± 0,42) ‰, 2-й – расстройствах поведения
в зрелом возрасте (F60–F69) – 14,5 % и (2,65 ±
0,48) ‰ соответственно, 3-й – болезнях пище
вода, желудка и двенадцатиперстной кишки
(K20–К31) – 13,4 % и (2,44 ± 0,34) ‰ соответ
ственно, 4-й – органических, включая симпто
матические, психических расстройствах (F01–
F09) – 8,6 % и (1,56 ± 0,31) ‰ соответственно,
5-й – туберкулезе (A15–A19) – 3,4 % и (0,61 ±
0,08) ‰ соответственно (см. табл. 11). В общей
сложности показатели перечисленных 5 нозо
логий составили 54,6 % от всей структуры
увольняемости военнослужащих по призыву
Сухопутных войск России (см. табл. 11). Окончание таблицы 9 Динамика показателей увольняемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig 16 Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right) over time (‰) Рис. 16. Динамика показателей увольняемости военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) ( ‰). Fig. 16. Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). Fig. 16. Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right Fig. 16. Dismissal rates for conscripts of the Russian Navy (left) and Ground Forces (right) over time ( ‰). Рис. 17. Структура показателей увольняемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 17. Dismissal structure (left) and dismissal structure over time (right) in conscripts of Russian Navy (%). льняемости (слева) и динамика структуры (справа)
х по призыву ВМФ России (%) Рис. 17. Структура показателей увольняемости (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). у военнослужащих по призыву ВМФ России (%). Fig. 17. Dismissal structure (left) and dismissal structure over time (right) in conscripts of Russian Navy (%) MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 38 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Рис. 18. Структура показателей увольняемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 18. Dismissal structure (left) and dismissal structure over time (right) in conscripts of Russian Ground Forces (%). Рис. 18. Структура показателей увольняемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Рис. 18. Структура показателей увольняемости (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 18. Dismissal structure (left) and dismissal structure over time (right) in conscripts of Russian Ground Forces (%). Fig. 18. Dismissal structure (left) and dismissal structure over time (right) in conscripts of Russian Grou crease in category V diseases, a decrease in
category I diseases, with certain stability for
category IX and XI diseases (see Fig. 18, right). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Окончание таблицы 9 In the dismissal structure for the Ground
Forces conscripts, neurotic, stress-related and
somatoform disorders (F40–F48) ranked 1st
(share 14.8 % and rate (2.70 ± 0.42) ‰); dis
orders of adult personality and behaviour (F60–
F69) ranked 2nd (14.5 % and (2.65 ± 0.48) ‰);
diseases of oesophagus, stomach and duode
num (K20–К31) ranked 3rd (13.4 % and (2.44
± 0.34) ‰); organic, including symptomatic,
mental disorders (F01–F09) ranked 4th (8.6 %
and (1.56 ± 0.31) ‰); tuberculosis (A15–A19)
ranked 5th (3.4 % and (0.61 ± 0.08) ‰) (see Ta
ble 11). The above 5 disease categories made
up В 54.6 % of the dismissal structure in the
Ground Forces conscripts (see Table 11). Уровень увольнений военнослужащих по
призыву Сухопутных войск России был стати
стически значимо меньше, чем военнослужа
щих по призыву ВМФ России с вирусным гепа
титом (В15–В19), невротическими, связанными
со стрессом, и соматоформными расстрой
ствами (F40–F48), расстройствами поведения Compared to the Navy conscripts, the
Ground Forces conscripts were statistically sig 39 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations 2019 N 3
Таблица 11. Уровень увольняемости военнослужащих по призыву ВМФ России и Сухопутных войск России по основным болезням (группам в классах) ( ‰)
Table 11. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Окончание таблицы 9 Mortality trends be
tween the Navy and Ground Forces conscripts
are highly consistent and statistically signifi
cant (r = 0.87; p < 0.05), suggesting the role of
various factors, e. g. military professional ones. Полиномиальные
тренды
смертности
военнослужащих по призыву ВМФ России
и Сухопутных войск России при высоких ва
риабельности показателей и коэффициен
тах детерминации показывают уменьшение
данных (рис. 19). Согласованность трендов
смертности военнослужащих по призыву ВМФ
России и Сухопутных войск России – силь
ная и статистически достоверная (r = 0,87; The highest mortality rates were observed in
the Navy conscripts with injures and diseases Таблица 12. Показатели смертности военнослужащих по призыву ВС России, ВМФ России
и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. (на 100 тыс. военнослужащих
по призыву соответствующей когорты)
Table 12. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Окончание таблицы 9 Dismissal rates in Russian Navy and Ground Forces conscripts by main diseases (ICD‑10 blocks) ( ‰)
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Туберкулез / Tuberculosis (A15–A19)
0.46 ± 0.06
1.36
8
0.61 ± 0.08
3.36
5
Вирусный гепатит / Viral hepatitis (В15–В19)
0.95 ± 0.22
2.86
4
0.26 ± 0.06
1.43
9
0.01
Органические, включая симптоматические, психические расстройства / Organic, including symptomatic,
mental disorders (F01–F09)
0.39 ± 0.05
1.16
10
1.56 ± 0.31
8.57
4
0.01
Невротические, связанные со стрессом, и соматоформные расстройства / Neurotic, stress-related and
somatoform disorders (F40–F48)
10.75 ± 1.44 32.21
1
2.70 ± 0.42
14.77
1
0.001
Расстройства поведения в зрелом возрасте / Disorders of adult personality and behaviour (F60–F69)
5.63 ± 0.72
16.87
2
2.65 ± 0.48
14.52
2
0.001
Поражения отдельных нервов, нервных корешков и сплетений / Nerve, nerve root and plexus disorders (G50–
G58)
0.14 ± 0.03
0.42
>14
0.16 ± 0.03
0.87
10
Болезни, характеризующиеся повышенным кровяным давлением / Hypertensive diseases (I10–I15)
0.39 ± 0.05
1.16
9
0.36 ± 0.09
1.99
7
Другие болезни сердца (эндокардит, миокардит) / Other cardiac diseases (endocarditis, myocarditis) (I30–I52)
0.92 ± 0.09
2.77
5
0.36 ± 0.07
1.98
8
0.001
Хронические болезни нижних дыхательных путей / Chronic lower respiratory diseases (J40–J47)
0.73 ± 0.09
2.19
6
0.16 ± 0.02
0.87
11
0.001
Болезни пищевода, желудка и двенадцатиперстной кишки (K20–К31), в том числе/ Diseases of oesophagus,
stomach and duodenum (K20–К31), incl:
4.51 ± 0.40
13.51
3
2.44 ± 0.34
13.39
3
0.01
язва желудка / stomach ulcer (K25)
0.39 ± 0.06
1.18
0.72 ± 0.10
3.92
0.05
язва двенадцатиперстной кишки / duodenal ulcer (K26)
4.03 ± 0.35
12.07
1.63 ± 0.26
8.96
0.001
Болезни желчного пузыря, желчевыводящих путей и поджелудочной железы / Disorders of gallbladder, biliary
tract and pancreas (К80–К86)
0.17 ± 0.02
0.51
13
0.08 ± 0.01
0.43
>16
0.01
Деформирующие дорсопатии / Deforming dorsopathies (M40–M43), в том числе / incl.:
0.36 ± 0.05
1.09
11
0.43 ± 0.06
2.35
6
остеохондроз позвоночника / spinal osteochondrosis (M42)
0.16 ± 0.03
0.48
0.24 ± 0.04
1.30
Мочекаменная болезнь / Urolithiasis (N20–N23)
0.31 ± 0.03
0.92
12
0.13 ± 0.02
0.74
13
0.001
Травмы головы / Injury to the head (S00–S09)
0.11 ± 0.01
0.32
>14
0.15 ± 0.03
0.83
12
Травмы живота, нижней части спины, поясничного отдела позвоночника и таза / Injuries to the abdomen, lower
back, lumbar spine and pelvis (S30–S39)
0.12 ± 0.02
0.36
>14
0.13 ± 0.02
0.71
15
Последствия травм, отравлений и других воздействий внешних причин / Sequelae of injuries, of poisoning and
of other consequences of external causes (T90–T98)
0.46 ± 0.06
1.39
7
0.13 ± 0.04
0.72
14
0.001 40 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Окончание таблицы 9 2019. № в зрелом возрасте (F60–F69), другими болез
нями сердца (эндокардитом, миокардитом)
(I30–I52), хроническими болезнями нижних ды
хательных путей (J40–47), язвой двенадцати
перстной кишки (K26), мочекаменной болезнью
(N20–N23), последствиями травм и отравлений
и других воздействий внешних причин (T90–
T98) и больше с органическими, включая симп
томатические, психическими расстройствами
(F01–F09) и язвой желудка (K25) (см. табл. 11). в зрелом возрасте (F60–F69), другими болез
нями сердца (эндокардитом, миокардитом)
(I30–I52), хроническими болезнями нижних ды
хательных путей (J40–47), язвой двенадцати
перстной кишки (K26), мочекаменной болезнью
(N20–N23), последствиями травм и отравлений
и других воздействий внешних причин (T90–
T98) и больше с органическими, включая симп
томатические, психическими расстройствами
(F01–F09) и язвой желудка (K25) (см. табл. 11). nificantly less frequently dismissed due to viral
hepatitis (В15–В19), neurotic, stress-related
and somatoform disorders (F40–F48), disor
ders of adult personality and behaviour (F60–
F69), other cardiac diseases (endocarditis,
myocarditis) (I30–I52), chronic lower respirato
ry diseases (J40–J47), duodenum ulcer (K26),
urolithiasis (N20–N23), sequelae of injuries, of
poisoning and of other consequences of ex
ternal causes (T90–T98) and more frequently
due to organic, including symptomatic, mental
disorders (F01–F09) and stomach ulcer (K25)
(see Table 11). Смертность. Среднегодовой
уровень
смертности военнослужащих по призыву ВС
России составил (42,26 ± 6,01), ВМФ Рос
сии – (24,87 ± 5,12), Сухопутных войск России –
(50,67 ± 7,84) на 100 тыс. соответствующих ко
горт военнослужащих по призыву (табл. 12). Уровень смертности военнослужащих по при
зыву ВМФ России оказался самым низким по
сравнению с показателями у военнослужащих
по призыву ВС России и Сухопутных войск Рос
сии (p < 0,05 для обеих когорт). Mortality. Annual mortality rates were
(42.26 ± 6.01), (24.87 ± 5.12), (50.67 ± 7.84)
per 100 thousand conscripts of the Armed
Forces, Navy and Ground Forces (Table 12). Mortality rates in the Navy conscripts were the
lowest compared to that in the Armed Forces
and Ground Forces (p < 0.05 for both cohorts). Mortality. Annual mortality rates were
(42.26 ± 6.01), (24.87 ± 5.12), (50.67 ± 7.84)
per 100 thousand conscripts of the Armed
Forces, Navy and Ground Forces (Table 12). Mortality rates in the Navy conscripts were the
lowest compared to that in the Armed Forces
and Ground Forces (p < 0.05 for both cohorts). Polynomial trends for mortality among the
Navy and Ground Forces conscripts show a de
crease with high variability and high determina
tion coefficients (Fig. 19). Окончание таблицы 9 Mortality rates were lower in
the Navy conscripts with diseases from ICD‑10
categories IX, XI and XIV compared to Ground
Forces conscripts (see Table 12). p < 0,05), что может указывать на участие в раз
витии смертности военнослужащих по призыву
однонаправленных факторов, например воен
но-профессиональных. Самые высокие показатели смертности
были у военнослужащих по призыву ВМФ Рос
сии с травмами и болезнями XIX, I и IX клас
сов – (20,81 ± 4,37), (1,03 ± 0,38) и (0,83 ± 0,42)
на 100 тыс. военнослужащих по призыву со
ответственно, у военнослужащих по призыву
Сухопутных войск России – XIX, IX и II классов –
(43,76 ± 7,63), (2,18 ± 0,45) и (1,11 ± 0,31) на 100
тыс. соответственно (см. табл. 12). Уровень
смертности военнослужащих по призыву ВМФ
России с болезнями IX, XI и XIV классов был
меньше, чем военнослужащих по призыву Су
хопутных войск России (см. табл. 12). The mortality structure for the Navy con
scripts included diseases and injures from
categories XIX, I, IX and II (according to their
significance, share > 2 % each) with overall
contribution of 93.5 % (Fig. 20, left); for the
Ground Forces conscripts – diseases from cat
egories XIX, IX, II and X with overall contribution
of 94.7 % (Fig. 21, left). В структуру с долей около 2 % вошли пока
затели смертности военнослужащих по призы
ву ВМФ России с болезнями и травмами XIX, I,
IX и II классов (указаны в порядке значимости)
с общим вкладом 93,5 % (рис. 20, слева), воен
нослужащих по призыву Сухопутных войск Рос
сии – XIX, IX, II и X классов с вкладом 94,7 % от
структуры смертности (рис. 21, слева). The mortality structure for the Navy con
scripts over time shows a decrease in injuries
and leading diseases with increase in other dis
orders (see Fig. 20, right). The mortality structure for the Ground For
ces conscripts shows an increase in category II,
IX and X diseases with decrease in category XIX
injuries (see Fig. 21, right). В динамике структуры смертности отме
чается уменьшение доли военнослужащих по
призыву ВМФ России по причине травм и бо
лезней ведущих классов и увеличение доли
прочих расстройств (см. рис. 20, справа). Окончание таблицы 9 Mortality in conscripts of Russian Armed Forces, Navy and Ground Forces by ICD‑10
in 2003–2018 (per 100,000 conscripts)
Класс по
МКБ-10 /
ICD-10
Chapter
(1) ВС России /
Armed Forces,
уровень / rates,
(M ± m) ‰
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p <
(2) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
(3) уровень /
rates,
(M ± m) ‰
структура /
structure, %
ранг /
rank
1/2
2/3
1/3
I
1.01 ± 0.17
1.03 ± 0.38
4.14
2
0.68 ± 0.15
1.35
5
II
1.21 ± 0.20
0.59 ± 0.28
2.36
4
1.11 ± 0.31
2.18
3
III
0.13 ± 0.05
0.28 ± 0.28
1.13
8
0.17 ± 0.09
0.34
9
IV
0.03 ± 0.02
0.15 ± 0.15
0.62
9
0.03 ± 0.03
0.06
12
V
0.38 ± 0.15
0.39 ± 0.39
1.57
6
0.49 ± 0.25
0.97
7
VI
0.33 ± 0.10
0.33 ± 0.33
1.32
7
0.55 ± 0.17
1.09
6
VII
0.00 ± 0.00
0.00 ± 0.00
0.00 ± 0.00
VIII
0.01 ± 0.01
0.00 ± 0.00
0.03 ± 0.03
0.05
13–14
IX
2.20 ± 0.21
0.83 ± 0.42
3.35
3
2.18 ± 0.45
4.29
2
0.05
0.05
X
0.89 ± 0.14
0.45 ± 0.25
1.82
5
0.98 ± 0.21
1.92
4
XI
0.33 ± 0.09
0.00 ± 0.00
0.44 ± 0.15
0.87
8
0.01
0.01
XII
0.04 ± 0.02
0.00 ± 0.00
0.08 ± 0.05
0.17
11
XIII
0.01 ± 0.01
0.00 ± 0.00
0.02 ± 0.02
0.05
13–14
XIV
0.11 ± 0.04
0.00 ± 0.00
0.14 ± 0.08
0.28
10
0.05
0.05
XIX
35.56 ± 5.95
20.81 ± 4.37
83.69
1
43.76 ± 7.63
86.38
1
Общий /
Total
42.26 ± 6.01
24.87 ± 5.12
100.0
50.67 ± 7.84
100.00
0.05
0.05 Таблица 12. Показатели смертности военнослужащих по призыву ВС России, ВМФ России
и Сухопутных войск России по классам МКБ‑10 в 2003–2018 гг. (на 100 тыс. военнослужащих
по призыву соответствующей когорты) 41 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № from ICD‑10 categories XIX, I and IX ((20.81 ±
4.37), (1.03 ± 0.38) and (0.83 ± 0.42) per 100
thousand conscripts; in Ground Forces con
scripts with diseases from ICD‑10 categories
XIX, IX and II ((43.76 ± 7.63), (2.18 ± 0.45) and
(1.11 ± 0.31) per 100 thousand conscripts)
(see Table 12). Окончание таблицы 9 According to the Federal State Statistics
Service (Rosstat), mortality rates in Russian
male population in 2003–2018 were (126.6 ±
7.4) for men aged 15–19, (266.3 ± 21.1) for
men aged 20–24, and (448.8 ± 35.8) for men
aged 25–29 per 100 thousand men of respec
tive age [http://www.gks.ru/]. В динамике структуры смертности военно
служащих по призыву Сухопутных войск России
выявлено увеличение доли болезней II, IX и X
классов, уменьшение – с травмами XIX класса
(см. рис. 21, справа). Mortality rates in conscripts of the Russian
Armed Forces, Navy and Ground Forces were
6.3, 10.7 and 5.3 times lower compared to Rus
sian men aged 20–24. Mortality trends between По данным Росстата, смертность мужчин
России в возрасте 15–19 лет в 2003–2018 гг. Рис. 19. Динамика показателей смертности военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) (на 100 тыс. военнослужащих по призыву соответствующей когорты). Fig. 19. Mortality in conscripts of Russian Navy (left) and Ground Forces (right) over time
(per 100,000 conscripts of the respective cohort). Рис. 19. Динамика показателей смертности военнослужащих по призыву ВМФ России (слева)
и Сухопутных войск России (справа) (на 100 тыс. военнослужащих по призыву соответствующей когорты). Fig. 19. Mortality in conscripts of Russian Navy (left) and Ground Forces (right) over time
(per 100,000 conscripts of the respective cohort). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 42 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № Russian men aged 20–24 and conscripts of the
Russian Armed Forces, Navy and Ground Forc
es are functionally consistent (r = 0.90, 0.83
and 0.87; p < 0.001), suggesting influence of
unidirectional factors, e. g. macrosocial ones. Therefore, injury incidence among conscripts
is not as high as is commonly believed. составила (126,6 ± 7,4), в возрасте 20–24 года –
(266,3 ± 21,1), в возрасте 25–29 лет – (448,8 ± 35,8)
на 100 тыс. мужчин соответствующего возраста
[http://www.gks.ru/]. Уровень смертности военнослужащих по
призыву ВС России, ВМФ России и Сухопутных
войск России был в 6,3, 10,7 и 5,3 раза мень
ше, чем мужчин России в возрасте 20–24 года. Окончание таблицы 9 Согласованность смертности мужчин России
в этом возрасте и военнослужащих по призы
ву ВС России, ВМФ России и Сухопутных войск
России – функциональная (r = 0,90, 0,83 и 0,87
соответственно; p < 0,001), что может указывать
на участие в развитии смертности однонаправ
ленных факторов, например макросоциальных. Эти данные разрушают существующие домыс
лы о высоком уровне травматизма среди воен
нослужащих по призыву. According to the Rosstat, mortality rates
due to injury, poisoning and certain other con
sequences of external causes (ICD‑10 chapter
XIX) in 2011-2018 were (183.2 ± 13.3) per 100
thousand Russian men aged 20–24, (24.8 ±
3.1) in conscripts of the Russian Armed Forces,
(11.6 ± 3.9) in conscripts of the Russian Navy
and (14.1 ± 3.0) in conscripts of the Russian
Ground Forces per 100 thousand conscripts
from respective cohorts (p < 0.001 for all three
cohorts). In the Navy conscripts, there were 13
ICD‑10 code blocks with ≥ 0.5 % share in the
cause-of-death structure, with overall contri
bution of 50.3 %; for the Ground Forces con
scripts, there also were 13 such code blocks
with overall contribution of 66.8 %. Eleven
leading diseases in the cohorts under study
were similar (Table 13). По данным Росстата, в 2011–2018 гг. уро
вень смертности мужчин России в возрас
те 20–24 года по причине травм, отравлений
и других внешних причин (XIX класс) составил
(183,2 ± 13,3) на 100 тыс. мужчин соответствую
щего возраста, военнослужащих по призыву ВС
России – (24,8 ± 3,1), ВМФ России – (11,6 ± 3,9),
Сухопутных войск России – (14,1 ± 3,0) на 100 тыс. Рис. 20. Структура показателей смертности (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig 20 Mortality structure (left) and mortality structure over time (right) in conscripts of the Russian Navy (%) Рис. 20. Структура показателей смертности (слева) и динамика структуры (справа)
у военнослужащих по призыву ВМФ России (%). Fig. 20. Mortality structure (left) and mortality structure over time (right) in conscripts of the Russian Navy (%). Fig. 20. Mortality structure (left) and mortality structure over time (right) in conscripts of the Russian Navy (%). Рис. 21. Структура показателей смертности (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). Fig. 21. Mortality structure (left) and mortality structure over time (right) in conscripts of Russian Ground Forces (%). мертности (слева) и динамика структуры (справа)
ризыву Сухопутных войск России (%). Окончание таблицы 9 i
( i h ) i
i
f R
i
G
d F
(%) Рис. 21. Структура показателей смертности (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). G Рис. 21. Структура показателей смертности (слева) и динамика структуры (справа)
у военнослужащих по призыву Сухопутных войск России (%). у военнослужащих по призыву Сухопутных войск России (%). Fig. 21. Mortality structure (left) and mortality structure over time (right) in conscripts of Russian Ground Fo ality structure (left) and mortality structure over time (right) in conscripts of Russian Ground Forces (%). MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 43 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № военнослужащих по призыву соответствующей
когорты (p < 0,001 для всех трех когорт). In the cause-of-death structure for the
Navy conscripts, injury to the head (S00–S09)
ranked 1st (share 14.2 % and rate (4.26 ± 1.24)
per 100 thousand conscripts); injuries involv
ing multiple body regions (T00–T07) ranked
2nd (13.4 % and (3.33 ± 1.00)); injuries to the
abdomen, lower back, lumbar spine and pel
vis (S30–S39) ranked 3rd (3.47 % and (0.86 ±
0,64)); other cardiac diseases (endocarditis,
myocarditis) (I30–I52) ranked 4th (2.5 % and
(0.63 ± 0.36)); injury to the chest (S20–S29)
ranked 5th (2.0 % and (0.50 ± 0.23)) (see Ta
ble 13). The above 5 code blocks made up
38.5 % of the cause-of-death structure for the
Navy conscripts. военнослужащих по призыву соответствующей
когорты (p < 0,001 для всех трех когорт). У военнослужащих по призыву ВМФ России
ведущих нозологий (групп в классах), ставших
причинами смертности с долей 0,5 % и более,
оказалось 13 с суммарным вкладом в структу
ру 50,3 %, у военнослужащих по призыву Сухо
путных войск России – также 13 с долей 66,8 %. 11 ведущих нозологий в сравниваемых когортах
военнослужащих по призыву были одинаковы
ми (табл. 13). (
)
В структуре смертности военнослужащих
по призыву ВМФ России 1-й ранг состави
ли данные о травмах головы (S00–S09) с до
лей 14,2 % и уровнем (4,26 ± 1,24) на 100 тыс. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Окончание таблицы 9 военнослужащих по призыву, 2-й – травмах,
захватывающих несколько областей тела (T00–
T07),– 13,4 % и (3,33 ± 1,00) соответственно,
3-й – травмах живота, нижней части спины,
поясничного отдела позвоночника и таза (S30–
S39) – 3,47 % и (0,86 ± 0,64) соответственно,
4-й – других болезнях сердца (эндокардите,
миокардите) (I30–I52) – 2,5 % и (0,63 ± 0,36)
соответственно, 5-й – травмах грудной клетки
(S20–S29) – 2,0 % и (0,50 ± 0,23) соответствен
но (см. табл. 13). Показатели указанных 5 нозо
логий в общей сложности составили 38,5 % от
структуры смертности военнослужащих по при
зыву ВМФ России. In the cause-of-death structure for the
Ground Forces conscripts, injury to the head
(S00–S09) ranked 1st (share 23.0 % and rate
(11.65 ± 2.62) per 100 thousand conscripts);
injuries involving multiple body regions (T00–
T07) ranked 2nd (16.8 % and (8.51 ± 2.10));
injury to the neck (S10–S19) ranked 3rd (8.0 %
and (4.07 ± 1.22)); injury to the chest (S20–
S29) ranked 4th (6.5 % and (3.29 ± 0.82)); inju
ries to the abdomen, lower back, lumbar spine
and pelvis (S30–S39) ranked 5th (6.5 % and
(3.29 ± 0.82) (see Table 13). The leading 5 code
blocks made up 57.5 % of the cause-of-death
structure for the Ground Forces conscripts. В структуре смертности военнослужащих
по призыву Сухопутных войск России 1-й ранг
занимали показатели травм головы (S00–S09)
с долей 23,0 % и уровнем (11,65 ± 2,62) на
100 тыс. военнослужащих по призыву, 2-й ранг –
травм, захватывающих несколько областей тела
(T00–T07),– 16,8 % и (8,51 ± 2,10) соответствен
но, 3-й – травм шеи (S10–S19) – 8,0 % и (4,07 ±
1,22) соответственно, 4-й – травм грудной клет
ки (S20–S29) – 6,5 % и (3,29 ± 0,82) соответ
ственно, 5-й – травм живота, нижней части спи
ны, поясничного отдела позвоночника и таза
(S30–S39) – 6,5 % и (3,29 ± 0,82) соответственно
(см. табл. 13). Ведущие 5 причин смертности
военнослужащих по призыву Сухопутных войск
России составили 57,5 % от структуры. В структуре смертности военнослужащих
по призыву Сухопутных войск России 1-й ранг
занимали показатели травм головы (S00–S09)
с долей 23,0 % и уровнем (11,65 ± 2,62) на
100 тыс. Окончание таблицы 9 военнослужащих по призыву, 2-й ранг –
травм, захватывающих несколько областей тела
(T00–T07),– 16,8 % и (8,51 ± 2,10) соответствен
но, 3-й – травм шеи (S10–S19) – 8,0 % и (4,07 ±
1,22) соответственно, 4-й – травм грудной клет
ки (S20–S29) – 6,5 % и (3,29 ± 0,82) соответ
ственно, 5-й – травм живота, нижней части спи
ны, поясничного отдела позвоночника и таза
(S30–S39) – 6,5 % и (3,29 ± 0,82) соответственно
(см. табл. 13). Ведущие 5 причин смертности
военнослужащих по призыву Сухопутных войск
России составили 57,5 % от структуры. Compared to the Ground Forces conscripts,
the cause-of-death structure for the Navy
conscripts included statistically significantly
less injuries to the head (S00–S09), injuries
to the neck (S10–S19), injuries to the chest
(S20–S29) and injuries involving multiple body
regions (T00–T07) (see Table 13). Assessments of military epidemiologi
cal significance of diseases. Taking into
consideration significance of medical and
statistical groups of diseases, contributions
of 80 ICD‑10 diseases categories and code
blocks into morbidity measures were calcula
ted. Table 14 presents a structure of perceived
military and epidemiological significance of
specific disease categories for the cohorts of
conscripts under study. Уровень смертности военнослужащих по
призыву ВМФ России был статистически досто
верно меньше, чем военнослужащих по призы
ву Сухопутных войск России с травмами головы
(S00–S09), травмами шеи (S10–S19), травмами
грудной клетки (S20–S29) и травмами, захваты
вающими несколько областей тела (T00–T07)
(см. табл. 13). For the conscripts of the Russian Armed
Forces, diseases of the respiratory system
(ICD‑10 chapter X) were the most significant in
terms of health effects, injury, poisoning and
certain other consequences of external caus
es (XIX) ranked 2nd, mental and behavioural
disorders (V) ranked 3rd, diseases of the skin
and subcutaneous tissue (XII) ranked 4th, dis
eases of the digestive system (XI) ranked 5th
(see Table 14). Оценка военно-эпидемиологической зна-
чимости заболеваний. С учетом значимости
медико-статистических видов заболеваемости Оценка военно-эпидемиологической зна-
чимости заболеваний. С учетом значимости
медико-статистических видов заболеваемости 44 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Таблица 13. Уровень смертности военнослужащих по призыву ВМФ России и Сухопутных войск России по основным болезням (группам в классах) (на 100 тыс. военнослужащих по призыву соответствующей когорты)
Table 13. Окончание таблицы 9 For the conscripts of the
Russian Armed Forces, there were 25 ICD‑10
code blocks with ≥ 0.5 % share, for the Navy and
Ground Forces conscripts there were 21 and 24
such code blocks (see Table 15). Code blocks
with > 0.5 % contribution were marked with “>”. Для военнослужащих по призыву ВМФ Рос
сии и Сухопутных войск России ведущие ранги
значимости классов болезней совпали с ранга
ми общей когорты военнослужащих по призыву
ВС России (см. табл. 14). В сумме доля перечис
ленных классов в структуре оценки военно-эпи
демиологической значимости классов для рас
стройств здоровья военнослужащих по призыву
ВС России, ВМФ России и Сухопутных войск
России составила 79,7, 80,3 и 80,6 % соответ
ственно. For the Navy conscripts the leading signifi
cant code blocks were as follows (see Table 15):
1) acute upper respiratory infections (J00–J06
by ICD‑10); 2) neurotic, stress-related and so
matoform disorders (F40–F48); 3) injury to the
head (S00–S09); 4) diseases of oesophagus,
stomach and duodenum (K20–К31); 5) inju
ries involving multiple body regions (T00–T07). Overall contribution of the above diseases to
the structure of military and epidemiological
significance in terms of health effects in the
Navy conscripts was 42 %. В табл. 15 представлена структура сформи
рованной оценки военно-эпидемиологической
значимости отдельных болезней (групп в клас
сах) для сравниваемых когорт военнослужащих
по призыву. У военнослужащих по призыву ВС
России имели долю 0,5 % и более в обобщенной
оценке 25 нозологий (групп в классах), у воен
нослужащих по призыву ВМФ России – 21,
у военнослужащих по призыву Сухопутных войск
России – 24 (см. табл. 15). Нозологиям, у кото
рых вклад был менее 0,5 %, присвоили ранг со
значком «более» (>). For the Ground Forces conscripts the lead
ing significant code blocks were as follows (see
Table 15): 1) acute upper respiratory infections
(J00–J06 by ICD‑10); 2) injury to the head (S00–
S09); 3) infections of the skin and subcutane
ous tissue (L00–L08); 4) diseases of oesoph
agus, stomach and duodenum (K20–К31); 5)
injuries involving multiple body regions (T00–
T07). Overall contribution of the above diseases
to the structure of military and epidemiological
significance in terms of health effects in the
Ground Forces conscripts was 39.4 %. У военнослужащих по призыву ВМФ России
(см. табл. Окончание таблицы 9 15) 1-й ранг составили показатели
острых респираторных инфекций верхних дыха
тельных путей (J00–J06 по МКБ‑10), 2-й – нев
ротических, связанных со стрессом, и сомато
формных расстройств (F40–F48), 3-й – травм
головы (S00–S09), 4-й – болезней пищевода, же
лудка и двенадцатиперстной кишки (K20–К31),
5-й – травм, захватывающих несколько обла
стей тела (T00–T07). В сумме указанные нозоло
гии составили 42 % от структуры оценки военно-
эпидемиологической значимости нозологий для
развития расстройств здоровья военнослужа
щих по призыву ВМФ России. Inr the Navy conscripts vs Armed Forc
es and Ground Forces conscripts, neurot
ic, stress-related and somatoform disorders
(F40–F48) were more common (7.7 vs 4.1 or
3.4 %, respectively) (see Table 15). Окончание таблицы 9 2019. № 3 кобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № рассчитали вклад в показатели заболеваемости
классов болезней и 80 нозологий (групп в клас
сах). В табл. 14 представлена структура сформи
рованной оценки военно-эпидемиологической
значимости классов болезней для сравниваемых
когорт военнослужащих по призыву. рассчитали вклад в показатели заболеваемости
классов болезней и 80 нозологий (групп в клас
сах). В табл. 14 представлена структура сформи
рованной оценки военно-эпидемиологической
значимости классов болезней для сравниваемых
когорт военнослужащих по призыву. For the conscripts of the Navy and Ground
Forces, the most significant disease catego
ries were similar compared to the conscripts
of the Russian Armed Forces (see Table 14). Overall contributions of the above diseases in
the structure of military and epidemiological
significance in terms of health effects were
79.7, 80.3 and 80.6 % in the conscripts of
the Russian Armed Forces, Navy and Ground
Forces, respectively. Оказалось, что наибольшую значимость по
классам болезней или 1-й ранг для здоровья
военнослужащих по призыву ВС России соста
вили болезни органов дыхания (X класс), 2-й –
травмы и другие воздействия внешних причин
(XIX класс), 3-й – психические расстройства
и расстройства поведения (V класс), 4-й – бо
лезни кожи и подкожной клетчатки (XII класс),
5-й – болезни органов пищеварения (XI класс)
(см. табл. 14). ,
p
y
Table 15 shows such a structure of military
and epidemiological significance for separate
diseases (code blocks) for the cohorts of con
scripts under study. For the conscripts of the
Russian Armed Forces, there were 25 ICD‑10
code blocks with ≥ 0.5 % share, for the Navy and
Ground Forces conscripts there were 21 and 24
such code blocks (see Table 15). Code blocks
with > 0.5 % contribution were marked with “>”. For the Navy conscripts the leading signifi
cant code blocks were as follows (see Table 15):
1) acute upper respiratory infections (J00–J06
by ICD‑10); 2) neurotic, stress-related and so
matoform disorders (F40–F48); 3) injury to the
head (S00–S09); 4) diseases of oesophagus,
stomach and duodenum (K20–К31); 5) inju
ries involving multiple body regions (T00–T07). Overall contribution of the above diseases to
the structure of military and epidemiological
significance in terms of health effects in the
Navy conscripts was 42%. Table 15 shows such a structure of military
and epidemiological significance for separate
diseases (code blocks) for the cohorts of con
scripts under study. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Окончание таблицы 9 Mortality in the Russian Navy and Ground Forces conscripts by main diseases (ICD‑10 blocks) (per 100 thousand conscripts of respective cohorts)
Нозология, группа в классе (код по МКБ-10) /
ICD-10 blocks
ВМФ России / Navy
Сухопутные войска России /
Ground Forces
p 1–2 <
(1) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
(2) уровень /
rates,
(M ± m) ‰
%
ранг /
rank
Другие бактериальные болезни (менингит, скарлатина) / Other bacterial diseases (meningitis, scarlatina) (A30–
A49)
0.42 ± 0.23
1.69
8
0.66 ± 0.15
1.30
8
Вирусный гепатит / Viral hepatitis (В15–В19)
0.30 ± 0.30
1.20
12
0.00 ± 0.00
0.00
>14
Злокачественные новообразования / Malignant neoplasms (C00–C80)
0.10 ± 0.10
0.40
>14
0.35 ± 0.10
0.70
12
Злокачественные новообразования лимфоидной и кроветворной и родственных им тканей / Malignant
neoplasms, stated or presumed to be primary, of lymphoid, haematopoietic and related tissue (C81–C96)
0.28 ± 0.19
1.12
13
0.55 ± 0.22
1.08
9
Органические, включая симптоматические, психические расстройства / Organic, including symptomatic,
mental disorders (F01–F09)
0.39 ± 0.36
1.57
9
0.25 ± 0.13
0.50
13
Демиелинизирующие болезни центральной нервной системы / Malignant neoplasms, stated or presumed to be
primary, of lymphoid, haematopoietic and related tissue (G35–G37)
0.33 ± 0.33
1.32
11
0.00 ± 0.00
0.00
>14
Другие болезни сердца (эндокардит, миокардит) / Other cardiac diseases (endocarditis, myocarditis) (I30–I52)
0.63 ± 0.36
2.54
4
1.14 ± 0.29
2.25
6
Цереброваскулярные болезни / Cerebrovascular disorders (I60–I69)
0.10 ± 0.10
0.42
>14
0.40 ± 0.12
0.80
10
Грипп и пневмония / Influenza and pneumonia (J10–J18)
0.45 ± 0.25
1.82
6
0.92 ± 0.22
1.81
7
Травмы головы / Injury to the head (S00–S09)
4.26 ± 1.24
17.14
1
11.65 ± 2.62 22.98
1
0.05
Травмы шеи / Injury to the neck (S10–S19)
0.33 ± 0.23
1.34
10
4.07 ± 1.22
8.04
3
0.01
Травмы грудной клетки / Injury to the chest (S20–S29)
0.50 ± 0.23
2.00
5
3.29 ± 0.82
6.50
4
0.05
Травмы живота, нижней части спины, поясничного отдела позвоночника и таза / Injuries to the abdomen, lower
back, lumbar spine and pelvis (S30–S39)
0.86 ± 0.64
3.47
3
1.63 ± 0.49
3.23
5
Травмы, захватывающие несколько областей тела / Injuries involving multiple body regions (T00–T07)
3.33 ± 1.00
13.38
2
8.51 ± 2.10
16.80
2
0.05
Последствия травм, отравлений и других воздействий внешних причин / Sequelae of injuries, of poisoning and
of other consequences of external causes (T90–T98)
0.43 ± 0.24
1.74
7
0.39 ± 0.16
0.77
11 45 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Conclusion Russian Navy conscripts had statistically
significantly more work days lost (p < 0.01),
dismissal for health reasons (p < 0.001) and
lower mortality rates (p < 0.05) compared to
Ground Forces conscripts. Other medical and
statistical measures in conscripts were similar. У военнослужащих по призыву Сухопутных
войск России (см. табл. 15) 1-й ранг состави
ли показатели острых респираторных инфек 46 Таблица 14. Классы болезней, имеющие важное военно-эпидемиологическое значение
для военнослужащих по призыву
Table 14. MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Conclusion Disease categories of military-epidemiological importance for conscripts
Класс по МКБ-10 / ICD-10 category
ВС России /
Armed Forces
ВМФ России /
Navy
Сухопутные
войска России /
Ground Forces
%
ранг /
rank
%
ранг /
rank
%
ранг /
rank
I
Некоторые инфекционные и паразитарные болезни / Certain
infectious and parasitic diseases
4.72
6
5.72
6
4.79
6
II
Новообразования / Neoplasms
1.12
14
1.04
12
0.89
14
III
Болезни крови, кроветворных органов и отдельные нару
шения, вовлекающие иммунный механизм / Diseases of the
blood and blood-forming organs and certain disorders involving
the immune mechanism
0.23
15
0.50
15
0.22
15
IV
Болезни эндокринной системы, расстройства питания
и нарушения обмена веществ / Endocrine, nutritional and
metabolic diseases
1.73
9
1.50
9
1.70
9
V
Психические расстройства и расстройства поведения /
Mental and behavioural disorders
11.04
3
12.80
3
11.21
3
VI
Болезни нервной системы / Diseases of the nervous system
1.63
10
1.48
10
1.65
10
VII
Болезни глаз и его придаточного аппарата / Diseases of the
eye and adnexa
1.13
12–13
0.91
13–14
1.00
13
VIII
Болезни уха и сосцевидного отростка / Diseases of the ear
and mastoid process
1.13
12–13
0.91
13–14
1.13
12
IX
Болезни системы кровообращения / Diseases of the
circulatory system
3.95
7
3.35
7
3.61
7
X
Болезни органов дыхания / Diseases of the respiratory system
27.26
1
27.72
1
27.55
1
XI
Болезни органов пищеварения / Diseases of the digestive
system
6.68
5
6.28
5
6.46
5
XII
Болезни кожи и подкожной клетчатки / Diseases of the skin
and subcutaneous tissue
7.82
4
6.66
4
8.17
4
XIII
Болезни костно-мышечной системы и соединительной тка
ни / Diseases of the musculoskeletal system and connective
tissue
3.22
8
3.07
8
2.90
8
XIV
Болезни мочеполовой системы / Diseases of the genitourinary
system
1.49
11
1.26
11
1.48
11
XIX
Травмы, отравления и некоторые другие последствия воз
действия внешних причин / Injury, poisoning and certain other
consequences of external causes
26.85
2
26.80
2
27.24
2
Сумма вклада классов / Total
100.0
100.0
100.0 ций верхних дыхательных путей (J00–J06
по МКБ‑10), 2-й – травм головы (S00–S09),
3-й – инфекций кожи и подкожной клетчатки
(L00–L08), 4-й – болезней пищевода, желуд
ка и двенадцатиперстной кишки (K20–К31),
5-й – травм, захватывающих несколько облас
тей тела (T00–T07). Conclusion В сумме указанные но
зологии составили 39,4 % от структуры оцен
ки военно-эпидемиологической значимости
нозологий для развития расстройств здоро
вья военнослужащих по призыву Сухопутных
войск России. ций верхних дыхательных путей (J00–J06
по МКБ‑10), 2-й – травм головы (S00–S09),
3-й – инфекций кожи и подкожной клетчатки
(L00–L08), 4-й – болезней пищевода, желуд
ка и двенадцатиперстной кишки (K20–К31),
5-й – травм, захватывающих несколько облас
тей тела (T00–T07). В сумме указанные но
зологии составили 39,4 % от структуры оцен
ки военно-эпидемиологической значимости
нозологий для развития расстройств здоро
вья военнослужащих по призыву Сухопутных
войск России. In the Russian Navy and Ground Forces con
scripts, polynomial trends of general and prima
ry morbidity, hospital admissions and work days
lost show an increase, while dismissal and mor
tality trends show a decrease with determination
coefficients of varying significance. For almost all
morbidity categories, the trends are moderately
consistent and statistically significant, suggest
ing unidirectional factors, e. g. military and pro
fessional ones. Mortality trends between Russian male popu
lation aged 20–24 and Russian Navy and Ground
Forces conscripts are functionally consistent
(r = 0.83 and 0.87; p < 0.001), suggesting unidi
rectional factors, e. g. macrosocial or bevavioral
ones. По сравнению с когортами военнослужа
щих по призыву ВС России и Сухопутных войск
у военнослужащих по призыву ВМФ России
выявлен бо҂льший вклад в структуру здоровья
невротических, связанных со стрессом, и со
матоформных расстройств (F40–F48) – 4,1,
3,4 и 7,7 % соответственно (см. табл. 15). In the Russian Navy and Ground Forces con
scripts, most significant from military and epide
miological point of view diseases were similar: 47 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 Таблица 15. Нозологии (группы в классах), имеющие важное военно-эпидемиологическое значение для военнослужащих по призыву
Table 15. References 1. Afanas'ev V.N., Yuzbashev M.M. Analiz vremen
nykh ryadov i prognozirovanie [Time Series Analysis
and Forecasting]. Moskva. 2001. 228 p. (In Russ.) 2. Gurevitch K.G., Karazhelyaskov O.P. Vliyanie
vneshnikh faktorov na zabolevaemost' voennoslu
zhashchikh [The Influence of External Factors on the
Morbidity of Military Personnel]. Vestnik novykh med-
itsinskikh tekhnologii [Journal of New Medical Tech
nologies]. 2015. Vol. 22, N 4. Pp. 83–89. (In Russ.) 2. Gurevitch K.G., Karazhelyaskov O.P. Vliyanie
vneshnikh faktorov na zabolevaemost' voennoslu
zhashchikh [The Influence of External Factors on the
Morbidity of Military Personnel]. Vestnik novykh med-
itsinskikh tekhnologii [Journal of New Medical Tech
nologies]. 2015. Vol. 22, N 4. Pp. 83–89. (In Russ.) а росоц ал
, о еде
ес
р
У военнослужащих по призыву ВМФ Рос
сии и Сухопутных войск России ведущие ран
ги военно-эпидемиологической значимости
классов болезней совпали: 1-й ранг состави
ли болезни органов дыхания (X класс); 2-й –
травмы и другие воздействия внешних причин
(XIX класс); 3-й – психические расстройства
и расстройства поведения (V класс); 4-й – бо
лезни кожи и подкожной клетчатки (XII класс);
5-й – болезни органов пищеварения (XI класс). Ведущими рангами военно-эпидемиологи
ческой значимости нозологий оказались пока
затели острых респираторных инфекций верх
них дыхательных путей (J00–J06 по МКБ‑10),
травм головы (S00–S09), травм, захватываю
щих несколько областей тела (T00–T07), болез
ней пищевода, желудка и двенадцатиперстной
кишки (K20–К31). У военнослужащих по при
зыву ВМФ России высокую значимость име
ли также данные невротических, связанных
со стрессом, и соматоформных расстройств
(F40–F48), Сухопутных войск – инфекций кожи
и подкожной клетчатки (L00–L08). У военнослужащих по призыву ВМФ Рос
сии и Сухопутных войск России ведущие ран
ги военно-эпидемиологической значимости
классов болезней совпали: 1-й ранг состави
ли болезни органов дыхания (X класс); 2-й –
травмы и другие воздействия внешних причин
(XIX класс); 3-й – психические расстройства
и расстройства поведения (V класс); 4-й – бо
лезни кожи и подкожной клетчатки (XII класс);
5-й – болезни органов пищеварения (XI класс). 3. Davydova T.E. Sostoyanie zdorov'ya i kachestvo
zhizni grazhdan, prokhodyashchikh voennuyu sluzhbu
po prizyvu [The state of health and the quality of life of
citizens serving in conscription]: Abstract dissertation
PhD Med. Sci. Moskva. 2016. 22 p. (In Russ.) 4. Evdokimov V.I., Sivashchenko P.P. Pokazate
li zabolevaemosti voennosluzhashchikh po prizyvu
Vooruzhennykh sil Rossiiskoi Federatsii (2003–2016
gg.) [Indicators of morbidity among conscripts in
Armed Forces of the Russian Federation (2003–2016)]. Sankt-Peterburg. 2018. 76 p. (Seriya «Zabolevaemost'
voennosluzhashchikh» ["Morbidity in servicemen" Se
ries]. Issue 4). (In Russ.) 4. Заключение 1) diseases of the respiratory system (ICD‑10
chapter X); 2) injury, poisoning and certain other
consequences of external causes (XIX); 3) men
tal and behavioural disorders (V); 4) diseases of
the skin and subcutaneous tissue (XII); 5) disea
ses of the digestive system (XI). Проведенные исследования выявили у воен
нослужащих по призыву ВМФ России по срав
нению с военнослужащими по призыву Сухо
путных войск статистически значимо бо҂льшие
уровни дней трудопотерь (p < 0,01) и уволь
няемости по состоянию здоровья (p < 0,001),
и меньший уровень смертности (p < 0,05). Остальные медико-статистические показатели
заболеваемости военнослужащих по призыву
статистически достоверно не различались. Most significant ICD‑10 code blocks in terms
of the military epidemiology were as follows:
acute upper respiratory infections (J00–J06 by
ICD‑10), injury to the head (S00–S09), injuries
involving multiple body regions (T00–T07), dis
eases of oesophagus, stomach and duodenum
(K20–К31). In the Navy and Ground Forces con
scripts, neurotic, stress-related and somatoform
disorders (F40–F48) and infections of the skin
and subcutaneous tissue, (L00–L08) were of ad
ditional importance, respectively. У военнослужащих по призыву ВМФ России
и Сухопутных войск при разных по значимости
коэффициентах детерминации полиномиаль
ные тренды общей и первичной заболевае
мости, госпитализации и дней трудопотерь
показывают тенденции увеличения данных,
увольняемости и смертности – уменьшения
показателей. Согласованность трендов прак
тически всех видов заболеваемости – уме
ренная и статистически достоверная, что
может указывать на влияние в их формирова
нии однонаправленных факторов, например
военно-профессиональных. Prevention, timely treatment and rehabilitation
will help improve the health status of conscripts. Taking into account the rates and structure of
morbidity will optimize allocation of resources
of the medical service of the Armed Forces of
Russia. Согласованность трендов смертности муж
ского населения России в возрасте 20–24 года
и военнослужащих по призыву ВМФ России
и Сухопутных войск – функциональная (r = 0,83
и 0,87; p < 0,001), что свидетельствует о влия
нии однонаправленных факторов, например,
макросоциальных, поведенческих и пр. Conclusion 2019. № 3 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. 2019. № 3 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 Conclusion Diseases (ICD‑10 code blocks) of military-epidemiological importance for conscripts
Нозология, группа в классе (код по МКБ-10) /
ICD-10 code blocks
ВС России /
Armed Forces
ВМФ России /
Navy
Сухопутные
войска / Ground
Forces
%
ранг /
rank
%
ранг /
rank
%
ранг /
rank
Кишечные инфекции / Intestinal infectious diseases (A00–A09)
0.39
>26
0.51
21
0.51
23
Туберкулез / Tuberculosis (A15–A19)
0.62
21
0.48
>22
0.76
19
Другие бактериальные болезни (менингит, скарлатина) / Other bacterial diseases (meningitis, scarlatina) (A30–A49)
0.72
19
0.79
16
0.58
21–22
Вирусные инфекции, характеризующиеся поражениями кожи и слизистых оболочек / Viral infections characterized by
skin and mucous membrane lesions (B00–B09)
1.31
15
1.45
11
1.39
14
Вирусный гепатит / Viral hepatitis (В15–В19)
0.55
24
1.09
13
0.41
>25
Микозы / Mycoses (B35–B49)
0.65
20
0.49
>22
0.72
20
Недостаточность питания / Malnutrition (E40–E64)
1.36
13–14
0.94
15
1.40
13
Органические, включая симптоматические, психические расстройства / Organic, including symptomatic, mental
disorders (F01–F09)
1.56
11
0.26
>22
1.95
12
Невротические, связанные со стрессом, и соматоформные расстройства / Neurotic, stress-related and somatoform
disorders (F40–F48)
4.13
6
7.65
2
3.26
8
Расстройства поведения в зрелом возрасте / Disorders of adult personality and behaviour (F60–F69)
2.79
9
3.69
7
3.09
9
Поражения отдельных нервов, нервных корешков и сплетений / Nerve, nerve root and plexus disorders (G50–G58)
0.58
22
0.36
>22
0.50
24
Болезни наружного уха / Diseases of external ear (H60–H62)
0.54
25
0.30
>22
0.58
21–22
Болезни среднего уха и сосцевидного отростка / Diseases of middle ear and mastoid (H65–H74)
0.48
>26
0.53
20
0.44
>25
Болезни, характеризующиеся повышенным кровяным давлением / Hypertensive diseases (I10–I15)
0.81
18
0.37
>22
0.90
18
Другие болезни сердца (эндокардит, миокардит) / Other cardiac diseases (endocarditis, myocarditis) (I30–I52)
1.36
13–14
1.65
10
1.19
16
Острые респираторные инфекции верхних дыхательных путей / Acute upper respiratory infections (J00–J06)
17.68
1
19.80
1
16.79
1
Грипп и пневмония / Influenza and pneumonia (J10–J18)
3.88
7
4.04
6
4.32
7
Другие острые респираторные инфекции нижних дыхательных путей / Other acute lower respiratory infections (J20–J22)
3.5
8
2.17
9
4.40
6
Хронические болезни нижних дыхательных путей / Chronic lower respiratory diseases (J40–J47)
0.35
>26
0.55
19
0.28
>25
Болезни пищевода, желудка и двенадцатиперстной кишки (K20–К31),
в том числе / Diseases of oesophagus, stomach and duodenum (K20–К31), incl:
5.30
3
4.99
4
5.13
4
язва желудка / stomach ulcer (K25)
0.63
0.26
0.90
язва двенадцатиперстной кишки / duodenal ulcer (K26)
2.14
2.63
1.90
гастрит и дуоденит / gastritis and duodenitis (K29)
1.84
1.45
1.70
Инфекции кожи и подкожной клетчатки / Infections of the skin and subcutaneous tissue (L00–L08)
4.77
3.60
8
5.29
3
Деформирующие дорсопатии / Deforming dorsopathies (M40–M43), в том числе / incl.:
1.06
17
0.64
18
1.20
15
остеохондроз позвоночника / spinal osteochondrosis (M42)
0.63
0.32
0.77
Травмы головы / Injury to the head (S00–S09)
6.52
2
5.50
3
7.13
2
Травмы шеи / Injury to the neck (S10–S19)
1.81
10
0.42
>22
2.40
10
Травмы грудной клетки / Injury to the chest (S20–S29)
1.55
12
0.65
17
1.96
11
Травмы живота, нижней части спины, поясничного отдела позвоночника и таза / Injuries to the abdomen, lower back,
lumbar spine and pelvis (S30–S39)
1.08
16
1.17
12
1.12
17
Травмы, захватывающие несколько областей тела / Injuries involving multiple body regions (T00–T07)
5.27
4
4.09
5
5.03
5
Последствия травм, отравлений и других воздействий внешних причин / Sequelae of injuries, of poisoning and of other
consequences of external causes (T90–T98)
0.56
23
0.95
14
0.39
>25 48 Медикобиологические и социальнопсихологические проблемы безопасности в чрезвычайных ситуациях. Литература 1. Афанасьев В.Н., Юзбашев М.М. Анализ вре
менных рядов и прогнозирование. М.: Финансы
и статистика, 2001. 228 с. 2. Гуревич К.Г., Каражелясков О.П. Влияние
внешних факторов на заболеваемость военнослу
жащих // Вестн. новых мед. технологий. 2015. Т. 22,
№ 4. С. 83–89. 8. Kaprun N.A., Sosnovskiy A.A., Gasparian O.V.,
Pukhova N.M. Analiz sostoyaniya zdorov'ya grazhdan,
pribyvayushchikh dlya komplektovaniya soedinenii
i chastei Baltiiskogo flota [Assessment of health
conditions of servicemen recruited to Baltic fleet
units and formations]. Morskaya meditsina [Marine
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Поступила 30.07.2019 г. Для цитирования. Евдокимов В.И., Мосягин И.Г., Сиващенко П.П., Мухина Н.А. Анализ медико-статистических
показателей заболеваемости военнослужащих по призыву Военно-морского флота и Сухопутных войск Российской
Федерации в 2003–2018 гг. // Мед.-биол. и соц.-психол. пробл. безопасности в чрезв. ситуациях. 2019. № 3. С. 15–51. DOI 10.25016/2541-7487-2019-0-3-15-51 The authors declare the absence of obvious and potential conflicts of interest related to the publication of this article. Received 30.07.2019 For citing: Evdokimov V.I., Mosyagin I.G., Sivashchenko P.P., Mukhina N.A. Analiz mediko-statisticheskikh pokazatelei
zabolevaemosti voennosluzhashchikh po prizyvu Voenno-morskogo flota i Sukhoputnykh voisk Rossiiskoi Federatsii v 2003–
2018 gg. Mediko-biologicheskie i sotsial'no-psikhologicheskie problemy bezopasnosti v chrezvychaynykh situatsiyakh. 2019. N 3. Pp. 15–51. (In Russ.) Evdokimov V.I., Mosyagin I.G., Sivashchenko P.P., Mukhina N.A. Analysis of medical and statistical measures of morbidity in
conscripts of the Navy and Ground Forces of the Russian Federation in 2003–2018. Medicо-Biological and Socio-Psychological
Problems of Safety in Emergency Situations. 2019. N 3 Pp. 15–51. DOI 10.25016/2541-7487-2019-0-3-15-51 Evdokimov V.I., Mosyagin I.G., Sivashchenko P.P., Mukhina N.A. Analysis of medical and statistical measures of morbidity in
conscripts of the Navy and Ground Forces of the Russian Federation in 2003–2018. Medicо-Biological and Socio-Psychological
Problems of Safety in Emergency Situations. 2019. N 3 Pp. 15–51. DOI 10.25016/2541-7487-2019-0-3-15-51 MedicoBiological and SocioPsychological Problems of Safety in Emergency Situations. 2019. N 3 51
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The influence of trash minerals and agglomerate particles on spiral separation performance
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Journal of the Southern African Institute of Mining and Metallurgy/Journal of the South African Institute of Mining and Metallurgy
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* Exxaro Metallurgical Services, Pretoria, South
Africa.
† Ex-Mente, Pretoria, South Africa.
© The Southern African Institute of Mining and
Metallurgy, 2017. ISSN 2225-6253. This paper
was first presented at the Heavy Minerals
Conference ‘Expanding the Horizon’, 16–17
August 2016, Sun City, South Africa. http://dx.doi.org/10.17159/2411-9717/2017/v117n5a5 http://dx.doi.org/10.17159/2411-9717/2017/v117n5a5 :<=8437<>8: Heavy mineral sands deposits generally
consist of naturally fine-grained, free-flowing
sand due to the way in which the deposit was
formed. The size of the heavy minerals market
is roughly 8 Mt/a of zircon and titanium
combined (Iluka, 2013). If an average
combined feed grade of 5% zircon and
titanium and an average combined overall
product recovery of 80% are assumed, the
resulting run-of-mine feed tons that would
require primary spiral beneficiation is 200
Mt/a. The number of spirals required for the
processing of this material, assuming 6500
operating hours per annum, an average of 2
t/h per start, and the same number of rougher
spirals as other duty spirals, is over 30 000
spirals for the heavy mineral sands business
alone. These numbers clearly illustrate the
importance of the spiral in the heavy mineral
industry. /:809>9 Understanding the influence of feed material mineral assemblage on spiral
separation is paramount for the successful and consistent operation of a
spiral plant. Plant stability is required to maintain high recovery of
valuable heavy minerals (VHM). Standard material analysis methods used
in the heavy mineral industry to evaluate spiral separation behaviour are
unable to quantify performance differences in the case of large variations
in feed assemblage. This study illustrates a systematic approach to
evaluate spiral separation performance. This technique was applied on a
four-stage spiral plant. The performance information was used in a circuit
simulation to clearly illustrate the importance of correct setting of the
spiral splitter position and resulting spiral mass loading to ensure optimal
circuit stability and maximum VHM recovery. Consistently achieving a high-grade heavy
mineral concentrate at high valuable mineral
recovery is paramount to business success. The forecast VHM recovery (especially of
zircon) is an important input into monthly
and annual budgets. A 10% decline in zircon
recovery over a month equates to significant
revenue losses for a medium-sized operation,
without considering other valuable mineral
losses. The quick investigation and
understanding of the reasons for a decline in
zircon recovery and an appropriate response
with corrective actions can save the business
millions of dollars. The purpose of this article
is to illustrate the systematic approach that
was followed to identify recovery losses of a
typical four-stage spiral circuit and the
corrective actions taken to minimize the
impact. A?/'8=49 A?/'8=49
spiral separation, zircon, recovery, optimization. spiral separation, zircon, recovery, optimization. The influence of trash minerals and
agglomerate particles on spiral
separation performance by J. Grobler* and J. Zietsman† free-flowing sand), the feed is properly
deslimed (-45 µm fraction removed to less
than 5% of the solids mass), and the spirals
are fed within design solids concentration
(water to solids ratio) and design spiral load
(dry tons per hour per start). The influence of trash minerals and agglomerate particles four-hourly basis. This study was initiated after a 10%
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@$"&@!@.>446>:1@43</(@ @$ &@!@97;*?:1?=@43</(@
$ &@!@76?;:?=@43</%@ 0>=;6@78:7?:<=;<?@!@7(@90>=;6@.>446>:1@!@.(@90>=;6
<;>6@!@< ->13=?@+ 5; >:1@<;,6?@2=;7<>8:;<>8:@'><5@5>15@<=;95@78:<?:< four-hourly basis. This study was initiated after a 10%
decline in zircon recovery that persisted for a considerable
period. This decline was correlated with changes in plant feed
material. However, since there were large volumes of this
feed material, the plant had to process it at the highest
achievable zircon recovery. second band constituted the ‘other’ coloured heavy minerals,
the third band the fine quartz, and the last band the coarse
tail particles. Figure 2 illustrates the coloured bands on the
shaking table during fractionation for a condition of low
trash, while Figure 3 illustrates the coloured bands for high
trash content. Each mass fraction was assayed by X-ray
fluorescence (XRF) and a recovery-yield curve was drawn
(Figure 4). The shaking table result presents the best case, or
theoretical maximum recovery that can be achieved for a
specific feed material, since the shaking table provides
sufficient separation space and separation time for each
particle to reach its equilibrium position from a particle
density, size, and shape perspective. The following procedure was applied to minimize the
zircon losses while maintaining a reasonable zircon product
grade for the downstream process. ®
Theoretical maximum recovery: determine how much of
the zircon in the feed is recoverable by spiral
separation ®
Feed mineral assemblage impact: determine the
mineralogical composition of the feed material and
assess what the impact is on the individual spiral
separation Figure 4 illustrates a 91% zircon recovery at 21% mass
yield, which implies that 9% of the zircon is trapped in the
remaining 79% of the mass. This zircon recovery is quite low,
and further investigation indicated that fine agglomerates
(less than 500 μm) were present in the feed. These
agglomerates trapped zircon particles (and other VHM) in a
low-density clay and quartz matrix, causing them to be
misplaced to the tail fraction. The same feed sample was
attrition-scrubbed in a laboratory unit and processed on the
shaking table after desliming. The result from the scrubbed
feed fractionation is presented in Figure 5. The influence of trash minerals and agglomerate particles A clear step
change (5–10%) in recovery is apparent for all three of the
heavy elements (Zr, Fe, and Ti), which shows that the VHM
particles were liberated by the scrubbing process. Detailed
QEMSCAN analyses of some of these agglomerates are
illustrated in Figure 6. The spiral circuit is unable to recover
the zircon contained in these agglomerates due to their p
®
Spiral circuit performance assessment: evaluate the
performance of the spiral circuit by sampling the four
duties. Apply mass balance techniques using the spiral
duty mass splits and stream analyses ®
Corrective measures: implement corrective measures
based on the interpretation of the circuit performance
results. Each step is presented in the next four sections with a
method description, followed by illustration and discussion of
the results. =87?43=? The spiral plant that was evaluated in this
study has four separation stages, as indicated
in Figure 1. The first three duties are all
equipped with Mineral Technology MG6.3
spirals and the final cleaner stage utilizes
Mineral Technology HG10S spirals. The
primary performance indicator of the plant is
zircon recovery, which is determined by the
two-product formula (using XRF assay of
ZrO2 in the feed, concentrate, and combined
tail stream). Three Vezin samplers take
composite samples of these three streams on a The aim of a spiral plant in the primary
processing value chain of a heavy mineral
process is to reject non-valuable quartz (a
light mineral) from various valuable heavy
minerals (VHM). The product from the spiral
plant is a heavy mineral concentrate that has
a reasonable grade (more than 90% heavy
minerals) at a reasonable heavy mineral
recovery (greater than 90%). This benchmark
is fairly easy to achieve if all the mineral
particles are well liberated (as in the case of VOLUME 117
A spiral circuit feed sample was fractionated on a laboratory
shaking table under controlled feed conditions. The sample
was divided into four mass fractions based on the band
colours. The first band constituted the ‘clear black band’, the L
436
VOLUME 117 L
436
VOLUME 117 The influence of trash minerals and agglomerate particles ->13=?@+ ?78*?=/#/>?64@=?6;<>8:95>0@82@<5?@283=@.;>:@?6?.?:<9@>:
90>=;6@2??4@2=;7<>8:;<?4@8:@;@95; >:1@<;,6? ->13=?@
@@1168.?=;<?@7=899#9?7<>8:9@,63?@0;=<>76?9@;=?@ >=78:(@1=??:
>9@76;/@.;<=>(@6>15<@0>: @0;=<>76?9@;=?@3;=< %@5?@=83:4@0;=<>76?(
9?78:4@2=8.@<5?@6?2<(@>9@"@.@>:@4>;.?<?= ->13=?@
@@1168.?=;<?@7=899#9?7<>8:9@,63?@0;=<>76?9@;=?@ >=78:(@1=??:
>9@76;/@.;<=>(@6>15<@0>: @0;=<>76?9@;=?@3;=< %@5?@=83:4@0;=<>76?(
9?78:4@2=8.@<5?@6?2<(@>9@"@.@>:@4>;.?<?=
Zircon I was used as an indicator particle class to
compare its recovery at different trash contents. The
pyroxene particle class was selected as the most predominant
trash mineral particle, and quartz was selected as the most
predominant light mineral particle. Since there is sufficient separation space on a shaking
table the trash minerals can easily achieve their respective
equilibrium positions. However, on a spiral the separation
space is limited, and the residence time for separation is short
(less than 15 seconds). Trash minerals will concentrate
towards the middle of the spiral, between the heavy minerals
and light minerals (Figure 8). This implies that there is
probably a threshold of trash mineral content that can be
accommodated on the spiral, beyond which separation will be
adversely affected. Figure 9 illustrates the three trash content scenarios. The
enhanced Holland-Batt equation was used to fit the test work
data points. This fitting technique for spiral data was
described by Grobler, Naude, and Zietsman (2016). In all
three feed scenarios the Zircon I recovery was above 95% for
the shaking table. However, the Zircon I recovery for the
spiral decreased significantly with increasing trash (from
95% to 83% to 78% at 20% mass yield to concentrate –
Figure 9). It is well known that particle size can also play a role in
spiral separation (Grobler and Bosman, 2011). To illustrate
that trash content had a more pronounced effect than particle
size distribution, the particle size distributions of the three
feed materials from Figure 9 were drawn up using the
QEMSCAN size distribution data from the Zircon I and
Pyroxene particle classes. Figure 10 illustrates that the
particle size distributions were similar for the three trash
content scenarios, while the spiral separation response
(Figure 9) varied significantly due to the trash content itself. y
Literature reports on the influence of trash minerals on
spiral separation are limited, since most mineral sands
deposits contain low trash concentrations, seldom exceeding
5% of the mineral assemblage. In this investigation the trash
mineral content was as high as 25%. To measure the impact
of trash on spiral separation, shaking table separation
response (ideal separation) was compared with spiral
separation response (on the same spiral profile) for
completely liberated particles with increasing trash content
(three trash content scenarios, based on actual plant feed
variation). QEMSCAN particle mineral analysis (PMA)
enables the classification of particles into different quality
classes.
The feed mineral assemblage, especially the concentration of
near-density heavy mineral particles, can have a significant
impact on spiral recovery. The heavy mineral industry gets its
name from the particle class ‘heavy minerals’. Traditionally
this class contains mineral particles that sink in an organic
medium (usually tetrabromoethane or bromoform) with a
liquid density of 2.96 g/cm3, and which constitute total heavy
minerals (THM). The THM thus have a density greater than
2.96 g/cm3. These heavy mineral particles are further
classified. Valuable heavy minerals (VHM, for example
zircon, rutile, and ilmenite) usually have a density greater
than 4 g/cm3. The term ‘trash’ is used to describe a particle
class with a specific density range. In this article, that range
is considered as being between 3 g/cm3 and 4 g/cm3, and
therefore this material has an intermediate density between
‘light’ particles and ‘heavy’ particles. It is called trash because
it contains no valuable or saleable minerals, but the minerals
are considered as ‘heavy’ since their density is greater than
2.96 g/cm3. This paper makes reference to three distinct
particle classes, heavy minerals (>4 g/cm3), trash minerals
(3–4 g/cm3), and light minerals (<3 g/cm3). To illustrate the
difference in separation response between these different
mineral groupings, a shaking table fractionation was carried ->13=?@+ ?78*?=/#/>?64@=?6;<>8:95>0@82@<5?@283=@.;>:@?6?.?:<9@>:
90>=;6@2??4@2=;7<>8:;<?4@8:@;@95; >:1@<;,6? ->13=?@ + ?78*?=/#/>?64@=?6;<>8:95>0@82@<5?@283=@.;>:@?6?.?:<9@>:
90>=;6@2??4@2=;7<>8:;<?4@8:@;@95; >:1@<;,6?@@,?28=?@;:4@;2<?=@;<<=><>8:
97=3,,>:1 ->13=?@+ 5; >:1@<;,6?@4?:9></@2=;7<>8:;<>8:@82@76?;:?=@90>=;6@2??4%
3.,?=9@78==?908:4@'><5@=;: >:1@>:@;,6?@%@7<3;6@4;<;@08>:<9@$:8<
958':&@'?=?@78::?7<?4@'><5@9<=;>15<@6>:?9@@:8@73=*?@9.88<5>:1@';9
;006>?4 ->13=?@ + ?78*?=/#/>?64@=?6;<>8:95>0@82@<5?@283=@.;>:@?6?.?:<9@>:
90>=;6@2??4@2=;7<>8:;<?4@8:@;@95; >:1@<;,6?@@,?28=?@;:4@;2<?=@;<<=><>8:
97=3,,>:1 combined low density, larger size, and irregular shape. This
material needs to be liberated before it reaches the spiral
circuit, and the spiral separation performance should be
measured against a new ‘baseline’. That is, a theoretical
maximum recovery as achieved by the shaking table; 91% in
the case of Figure 4. The results indicated in Figure 5 can be
achieved only with improved feed preparation in the plant
(milling and/or scrubbing) to liberate VHM from
agglomerates prior to spiral separation, with significant
power input. ->13=?@+ 5; >:1@<;,6?@4?:9></@2=;7<>8:;<>8:@82@76?;:?=@90>=;6@2??4%
3.,?=9@78==?908:4@'><5@=;: >:1@>:@;,6?@%@7<3;6@4;<;@08>:<9@$:8<
958':&@'?=?@78::?7<?4@'><5@9<=;>15<@6>:?9@@:8@73=*?@9.88<5>:1@';9
;006>?4
VOLUME 117 437 The influence of trash minerals and agglomerate particles Table I
>:?=;681>7;6@78.089><>8:@;:4@76;99>2>7;<>8:@82@<5?@9;.06?@39?4@28=@<5?@4?:9></@2=;7<>8:;<>8:@958':@>:
->13=?@ Table I
>:?=;681>7;6@78.089><>8:@;:4@76;99>2>7;<>8:@82@<5?@9;.06?@39?4@28=@<5?@4?:9></@2=;7<>8:;<>8:@958':@>:
->13=?@ able I
>:?=;681>7;6@78.089><>8:@;:4@76;99>2>7;<>8:@82@<5?@9;.06?@39?4@28=@<5?@4?:9></@2=;7<>8:;<>8:@958':@>:
>13=?@ out on the cleaner spiral feed. The fractions were analysed by
QEMSCAN to quantify the minerals present in the
assemblage. Figure 7 illustrates the three density response
bands, and Table I lists the mineral types and percentages in
each band. than 66%.
This analytical technique has been effectively applied
in many other density fractionation studies (Grobler and
Bosman, 2011). One of the PMA mineral classes is Zircon I, a
clean, completely liberated particle with ZrO2 content greater The impact of unliberated zircon particles as well as trash
content has been illustrated for the spiral in isolation; the
next step is to determine the impact on the spiral circuit as a
whole.
The influence of trash minerals and agglomerate particles The influence of trash minerals and agglomerate particles
->13=?@+>?64#=?78*?=/@=?6;<>8:95>09@28=@<5=??@4>22?=?:<@<=;95@78:<?:<@97?:;=>89%@$;&@
%@<=;95(@$,&@)"%@<=;95(@$7&@")%)@<=;95& he influence of trash minerals and agglomerate particles ->13=?@+>?64#=?78*?=/@=?6;<>8:95>09@28=@<5=??@4>22?=?:<@<=;95@78:<?:<@97?:;=>89%@$;&@
%@<=;95(@$,&@)"%@<=;95(@$7&@")%)@<=;95& middling duty tail, and scavenger duty tail in relation to the
mass yield to concentrate. Refer to Figure 1 for the spiral
circuit flow sheet. Case 1: the circuit was sampled under normal feed
conditions when the circuit zircon recovery was at
acceptable levels (above 90%). The following three feed scenarios were used to illustrate
the impact of trash and agglomerates on the spiral circuit
performance. Case 2: the circuit was sampled after the feed had
changed and the zircon recovery declined by more
than 15%, creating an unacceptable performance
condition.
->13=?@)+;=<>76?@9> ?@4>9<=>,3<>8:@4;<;@28=@>=78:@@;:4@/=8?:?@>:
90>=;6@2??4@28=@<5=??@<=;95@78:<?:<@97?:;=>89 Case 3: the circuit was sampled after corrective
measures were employed on the new feed condition
to minimize zircon losses. The three feed conditions are depicted in Table II. The
trash content in the feed increased in cases 2 and 3, while the
ZrO2 grade decreased. The most significant difference is
shown in the zircon recovery for the three cases. Each of the
three cases is discussed in the next three sections. Table III
gives the normalized mass yield to the different tail and
concentrate streams. The design load for middling and
scavenger spirals is 2 t/h per start, and for the cleaner 1.85
t/h per start. Figure 11 gives the zircon recovery to
concentrate response, and Figure 12 the zircon grade to
concentrate response, over the entire mass yield spectrum.
Figure 13 compares the ideal zircon recovery with spiral
circuit zircon recovery. The ideal recovery was determined to
be 94%, indicating that 6% of the zircon is not available for
The four-stage spiral circuit was sampled by selecting an
individual spiral column in each spiral duty and taking a
timed sample of the feed, concentrate, middling, and tail
streams. Feed distribution checks were done beforehand to
confirm that the selected spiral column was representative of
the entire spiral duty. The feed to the rougher spiral bank
was fractionated into four samples on the laboratory shaking
table to determine the theoretical maximum recovery. The
four spiral duty sample sets with the four shaking table
fractions resulted in 20 samples, which were deslimed and
assayed by XRF. No sink-float analyses were done. The XRF
assays were used in a mass balance model using the Limn
flow sheet processor (Excel add-in) to determine the mass-
balanced zircon recovery considering the main elements in
the mass balance solver. The circuit mass balance calculated
the total mass yield contributions of the rougher duty tail, ->13=?@+89><>8:9@82@4>22?=?:<@.>:?=;6@4?:9></@76;99?9@8:@<5?@90>=;6
7=899#9?7<>8: ->13=?@+89><>8:9@82@4>22?=?:<@.>:?=;6@4?:9></@76;99?9@8:@<5?@90>=;6
7=899#9?7<>8: L
438
VOLUME 117 438 VOLUME 117 The influence of trash minerals and agglomerate particles The influence of trash minerals and agglomerate particles
Figure 13 compares the ideal zircon recovery with spiral
circuit zircon recovery. The ideal recovery was determined to
be 94%, indicating that 6% of the zircon is not available for ->13=?@)+;=<>76?@9> ?@4>9<=>,3<>8:@4;<;@28=@>=78:@@;:4@/=8?:?@>:
90>=;6@2??4@28=@<5=??@<=;95@78:<?:<@97?:;=>89 439
L VOLUME 117 439
The influence of trash minerals and agglomerate particles spiral concentration due to lack of liberation (indicated by the
red arrow in Figure 13). The circuit zircon recovery response
was 90%, which implies that 96% of the available zircon was
successfully recovered by the four-stage spiral circuit. The
trash content was at the low end at 11.1% (Table II). The
downstream spiral loading (midd, scavenger, and cleaner)
was acceptably low. Case 1 demonstrates an acceptable circuit
separation response with high zircon recovery (96% with
consideration of irrecoverable zircon) to concentrate, and the
bulk of the zircon losses occurring in the rougher tail stream. spiral concentration due to lack of liberation (indicated by the
red arrow in Figure 13). The circuit zircon recovery response
was 90%, which implies that 96% of the available zircon was
successfully recovered by the four-stage spiral circuit. The
trash content was at the low end at 11.1% (Table II). The
downstream spiral loading (midd, scavenger, and cleaner)
was acceptably low. Case 1 demonstrates an acceptable circuit
separation response with high zircon recovery (96% with
consideration of irrecoverable zircon) to concentrate, and the
bulk of the zircon losses occurring in the rougher tail stream.
The influence of trash minerals and agglomerate particles The influence of trash minerals and agglomerate particles gg
p
spiral concentration due to lack of liberation (indicated by the
ed arrow in Figure 13). The circuit zircon recovery response
was 90%, which implies that 96% of the available zircon was
successfully recovered by the four-stage spiral circuit. The
rash content was at the low end at 11.1% (Table II). The
d
i
l l
di
idd
d l
(indicated by the red arrow in Figure 14). The circuit zircon
recovery response was 72%, which implies that 83% of the
available zircon was successfully recovered by the four-stag
spiral circuit. The trash content was on the high end at 17.2
(Table II). The downstream spiral loading (midd, scavenger
and cleaner) was unacceptably high, with every spiral duty
exceeding the design loading. Case 2 illustrates the impact
that high feed trash content has on the spiral circuit Firstly
Table II
-??4@;:4@7>=73><@=?908:9?@28=@<5?@<5=??@7;9?9
Table III
8=.;6> ?4@.;99@,;6;:7?@;:4@7>=73><@68;4>:1@93..;=/@28=@<5?@<5=??@7;9?9
->13=?@))+ ?78*?=/#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9%@5?@=?90?7<>*?@/>?649@<8@<5?@4>22?=?:<@?><>:1@0=87?99
9<=?;.9@;=?@>:4>7;<?4@,/@<5?@283=@58=> 8:<;6@6>:?9@$28=@ ;9?@"@8:6/&
->13=?@)"+ =;4?#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9 Table II
-??4@;:4@7>=73><@=?908:9?@28=@<5?@<5=??@7;9?9
Table III
8=.;6> ?4@.;99@,;6;:7?@;:4@7>=73><@68;4>:1@93..;=/@28=@<5?@<5=??@7;9?9
->13=?@))+ ?78*?=/#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
->13=?@)"+ =;4?#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9 8=.;6> ?4@.;99@,;6;:7?@;:4@7>=73><@68;4>:1@93..;=/@28=@<5?@<5=??@7;9?9 ->13=?@))+ ?78*?=/#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9%@5?@=?90?7<>*?@/>?649@<8@<5?@4>22?=?:<@?><>:1@0=87?99
9<=?;.9@;=?@>:4>7;<?4@,/@<5?@283=@58=> 8:<;6@6>:?9@$28=@ ;9?@"@8:6/& ->13=?@))+ ?78*?=/#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9%@5?@=?90?7<>*?@/>?649@<8@<5?@4>22?=?:<@?><>:1@0=87?99
9<=?;.9@;=?@>:4>7;<?4@,/@<5?@283=@58=> 8:<;6@6>:?9@$28=@ ;9?@"@8:6/& ->13=?@)"+ =;4?#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9 ->13=?@)"+ =;4?#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9 ->13=?@)"+ =;4?#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9 ->13=?@))+ ?78*?=/#/>?64@=?6;<>8:95>0@28=@<5?@<5=??@90>=;6@7>=73><
9?0;=;<>8:@7;9?9%@5?@=?90?7<>*?@/>?649@<8@<5?@4>22?=?:<@?><>:1@0=87?99
9<=?;.9@;=?@>:4>7;<?4@,/@<5?@283=@58=> 8:<;6@6>:?9@$28=@ ;9?@"@8:6/& (indicated by the red arrow in Figure 14). The circuit zircon
recovery response was 72%, which implies that 83% of the
available zircon was successfully recovered by the four-stage
spiral circuit. The trash content was on the high end at 17.2%
(Table II). The downstream spiral loading (midd, scavenger,
and cleaner) was unacceptably high, with every spiral duty
exceeding the design loading. Case 2 illustrates the impact
that high feed trash content has on the spiral circuit. Firstly,
it decreases the separation space and reduces the recovery
potential on the spiral. Secondly, it increases the loading
(tons per hour per start) on the downstream spiral duty,
causing a further decrease in zircon recovery. These are the
two main reasons for the drop in zircon recovery of 13%,
from 96% in Case 1 to 83% in Case 2. Case 2 demonstrates
an unacceptable circuit separation response with low zircon
recovery to concentrate (only 83% with consideration of
irrecoverable zircon), and zircon losses occurring in all three
tail streams. A high zircon loss in the scavenger tail stream is
a clear indication of overload in the spiral circuit. The influence of trash minerals and agglomerate particles ->13=?@)+ ;9?@)@>4?;6@=?78*?=/ 90>=;6@7>=73><@=?78*?=/ measures could be applied to the ideal zircon recovery, since
this depends on the nature of the feed material entering the
spiral circuit and can only be addressed by upgrades in the
feed preparation circuits, which was not part of this
investigation. The trash content was at the high end (19%)
as indicated in Table II. The feed trash content can be diluted
by blending with other low-trash mining zones, but this was
not part of this investigation since this feed condition
persisted for a considerable period due to large volumes of
this type of material being mined. The negative impact of
trash on individual spiral performance has to be tolerated
(Figure 9). However, the increased loading on the
downstream spiral duties can be managed by the operational
personnel. In Case 3 the tail splitters on the rougher and midd duties
were opened ‘abnormally’ wide to reject mass to the rougher
tail and midd tail streams. The rougher and midd duty outer
splitter position was changed (set inward) by more than 60
mm due to the mass flow shift that occurred with the high
percentage of trash in the feed. A splitter template was
designed and implemented to set every spiral splitter in each
duty at exactly the same position in the spiral trough. This
counter-intuitive operational change rejected 85% of the feed
mass to the rougher tail and midd tail steams (Case 3)
instead of only 71% in Case 2, as shown in Table III. Table III
further shows that the mass loading per spiral duty reduced
to more acceptable levels for Case 3, with a significant
decrease in recirculating loads. ->13=?@)+ ;9?@)@>4?;6@=?78*?=/ 90>=;6@7>=73><@=?78*?=/ ->13=?@)+ ;9?@)@>4?;6@=?78*?=/ 90>=;6@7>=73><@=?78*?=/ ->13=?@)+ ;9?@"@>4?;6@=?78*?=/@ 90>=;6@7>=73><@=?78*?=/ After this intervention the overall circuit zircon recovery
response was 83%, which implies that 93% of the recoverable
zircon was successfully recovered to concentrate by the four-
stage spiral circuit (indicated by the blue arrow in Figure 15). The change in spiral control approach resulted in a recovery
increase of 10% (from Case 2 to Case 3, with consideration of
irrecoverable zircon) while at the same time increasing the
concentrate grade by more than 1% ZrO2 as indicated in
Figure 12. VOLUME 117
441
L
->13=?@) + ;9?@@>4?;6@=?78*?=/@ 90>=;6@7>=73><@=?78*?=/ ->13=?@) + ;9?@@>4?;6@=?78*?=/@ 90>=;6@7>=73><@=?78*?=/ ->13=?@)+ ;9?@"@>4?;6@=?78*?=/@ 90>=;6@7>=73><@=?78*?=/ ->13=?@)+ ;9?@"@>4?;6@=?78*?=/@ 90>=;6@7>=73><@=?78*?=/
Figure 14 compares the ideal zircon recovery with spiral
circuit zircon recovery. The ideal zircon recovery was
determined to be 87%, indicating that 13% of the zircon is
not available for spiral concentration due to lack of liberation
L
440
VOLUME 117 L
440
VOLUME 117 VOLUME 117 The influence of trash minerals and agglomerate particles The influence of trash minerals and agglomerate particles to reduce this impact, other than reducing the spiral loading
to below design feed capacity. 8:7639>8:9 The application of attrition scrubbing to liberate
agglomerates and QEMSCAN to define the content of the
agglomerates is an excellent example of this, resulting in a
clear explanation of the ‘irrecoverable’ zircon phenomenon. Since Case 2 resulted in an unacceptable zircon recovery,
corrective measures were implemented in Case 3. The feed
composition cannot be changed, therefore the only remaining
control parameters are the operating conditions, total feed to
the plant, and more specifically the spiral splitter positions.
Figure 15 compares the ideal zircon recovery with spiral
circuit zircon recovery. The ideal zircon recovery was
determined to be 90%, indicating that 10% of the zircon is
not available for spiral concentration due to lack of liberation
(indicated by the red arrow in Figure 15). No corrective 8:7639>8:9 Trash mineral content in the feed to spiral plants is increasing
as more ‘difficult’ mineral deposits are being beneficiated. It
is therefore important to understand the impact of increased
trash mineral content and unliberated minerals on an
individual spiral profile, as well as on the spiral circuit as a
whole. An increase in feed trash content was shown to adversely
impact the zircon recovery of the total circuit by increasing
the load to the downstream circuits and increasing
recirculating load, since a large portion of the mass moved to
the middle of the spiral trough. This zircon recovery loss can
be minimized by adjusting the spiral splitter to reject as much
mass as possible (more than 85%) in the first two separation
stages, resulting in a recovery improvement for recoverable
zircon of more than 10%. QEMSCAN analyses provided important insight for
differentiating between trash minerals and valuable heavy
minerals, which can easily be misinterpreted when
considering heavy mineral content (typical sink-float results)
in new deposits or new mining areas of existing deposits. The
characterization of the separation response of ideal feed
material using a laboratory shaking table assisted
considerably in identifying the portion of the recovery
problem that addressed by the spiral operator, and the
portion that is not. The mass balance of the spiral circuit enabled the
measurement of the total circuit response. Although some
effort is required to configure the calculations initially, it
provides invaluable insight into the recirculating loads and
interaction between different spiral separation duties. It is
important to verify the mass balanced information with all
available measurement data. p
Using different techniques simultaneously to measure the
same initial observation can speed up understanding of the
problem. The application of attrition scrubbing to liberate
agglomerates and QEMSCAN to define the content of the
agglomerates is an excellent example of this, resulting in a
clear explanation of the ‘irrecoverable’ zircon phenomenon. An increase in feed trash content was shown to adversely
impact zircon recovery by decreasing the separation space on
the spiral trough. An increase of trash from 6% to 21%
resulted in a decrease in liberated zircon particle recovery
from 95% to 80%. Limited operational control can be applied Using different techniques simultaneously to measure the
same initial observation can speed up understanding of the
problem. ILUKA. 2013. Mineral Sands Industry. Fact Book. http://www.zircon-
association.org/Websites/zircon/images/The_Mineral_Sands_Industry
_Factbook.pdf [accessed 4 January 2015]. N GROBLER, J.D. and BOSMAN, J.B. 2011. Gravity separator performance evaluation
using Qemscan® particle mineral analysis. Journal of the Southern African
Institute of Mining and Metallurgy, vol. 111. pp. 401–408. GROBLER, J.D., NAUDE, N., and ZIETSMAN, J.H. 2016. Enhanced Holland-Batt
spline for describing spiral concentrator performance. Minerals
Engineering, vol. 92. pp. 189-195.
?2?=?:7?9 GROBLER, J.D. and BOSMAN, J.B. 2011. Gravity separator performance evaluation
using Qemscan® particle mineral analysis. Journal of the Southern African
Institute of Mining and Metallurgy, vol. 111. pp. 401–408. GROBLER, J.D. and BOSMAN, J.B. 2011. Gravity separator performance evaluation
using Qemscan® particle mineral analysis. Journal of the Southern African
Institute of Mining and Metallurgy, vol. 111. pp. 401–408. GROBLER, J.D., NAUDE, N., and ZIETSMAN, J.H. 2016. Enhanced Holland-Batt
spline for describing spiral concentrator performance. Minerals
Engineering, vol. 92. pp. 189-195. An increase in feed trash content was shown to adversely
impact zircon recovery by decreasing the separation space on
the spiral trough. An increase of trash from 6% to 21%
resulted in a decrease in liberated zircon particle recovery
from 95% to 80%. Limited operational control can be applied ILUKA. 2013. Mineral Sands Industry. Fact Book. http://www.zircon-
association.org/Websites/zircon/images/The_Mineral_Sands_Industry
_Factbook.pdf [accessed 4 January 2015]. N L
442 VOLUME 117 442
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Flexible and Scalable Transform-Domain Codebook for High Bit Rate CELP Coders
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Zenodo (CERN European Organization for Nuclear Research)
| 2,018
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cc-by
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2017 25th European Signal Processing Conference (EUSIPCO) 2017 25th European Signal Processing Conference (EUSIPCO) Flexible and Scalable Transform-Domain
Codebook for High Bit Rate CELP Coders Václav Eksler, Bruno Bessette, Milan Jelínek, Tommy Vaillancourt
University of Sherbrooke, VoiceAge Corporation
Montreal, QC, Canada Abstract—The Code-Excited Linear Prediction (CELP) model
is very efficient in coding speech at low bit rates. However, if the
bit rate of the coder is increased, the CELP model does not gain
in quality as
quickly as other
approaches. Moreover,
the computational complexity of the CELP model generally
increases significantly at higher bit rates. In this paper we focus
on a technique that aims to overcome these limitations by means
of a special transform-domain codebook within the CELP model. We show by the example of the AMR-WB codec that the CELP
model with the new flexible and scalable codebook improves
the quality at high bit rates at no additional complexity cost. (above approximately 16 kbps) obtained by using more pulses
in an ACELP innovation codebook is not as large as the gain
in dB/bit/sample of transform coding and vector quantization. At lower bit rates (below 12 kbps), the ACELP model captures
quickly the essential components of the excitation. However, at
higher bit rates, higher granularity and, in particular, better
control over how the additional bit budget is spent
across different frequency components of the signal are useful. (above approximately 16 kbps) obtained by using more pulses
in an ACELP innovation codebook is not as large as the gain
in dB/bit/sample of transform coding and vector quantization. At lower bit rates (below 12 kbps), the ACELP model captures
quickly the essential components of the excitation. However, at
higher bit rates, higher granularity and, in particular, better
control over how the additional bit budget is spent
across different frequency components of the signal are useful. Then, with increased bit rate, more pulses are searched
within very large codebooks and the complexity of the ACELP
search
algorithm
becomes
too
high
for
practical
implementations. Though many ACELP search algorithms
have been proposed that address this problem [3][4],
the computational complexity of these algorithms still
represents a substantial part of the overall codec complexity. Consequently,
careful
control
over
the
computational
complexity is needed, which in turn limits the coding
efficiency of these algorithms. Keywords—Speech coding, ACELP, AMR-WB I.
INTRODUCTION The Code-Excited Linear Prediction (CELP) model [1] is
widely used to encode speech signals at low bit rates. In CELP,
the speech signal is synthesized by filtering an excitation signal
through an all-pole digital synthesis filter where the excitation
parameters are optimized in a perceptually weighted synthesis
domain in a closed-loop manner. The filter is estimated by
means of linear prediction (LP) and it represents short-term
correlations between speech signal samples. In this paper we show how the traditional CELP model can
be extended at high bit rates to overcome the limitations of
scalability and complexity. While the presented technique can
be implemented in any CELP-based coder, we have chosen
the AMR-WB codec to demonstrate the performance of the
proposed technique. The rest of the paper is organized as
follows. In Section II, the AMR-WB codec is described with
emphasis on the innovation codebook used. In Section III we
extend the traditional CELP model by a new special transform-
domain codebook. Section IV shows the performance of the
presented
technique. In
Section V
we
discuss
some
implementation considerations of the model before concluding
the paper in Section VI. The excitation signal is typically composed of two parts
searched sequentially. The first part of the excitation, known
as a long-term prediction (LTP), is usually selected from an
adaptive codebook to exploit the quasi-periodicity of voiced
speech. This is done by searching in the past excitation the
segment most similar to the segment being currently encoded. The second part of the excitation is selected from an innovation
codebook and it models the evolution (difference) between the
past segment and the currently encoded segment. The innovation codebook can be designed using many
structures and constraints. However, in modern speech coding
systems, the Algebraic CELP (ACELP) model [2] is often
used. The codevectors in an algebraic innovation codebook
contain only few non-zero pulses while the number of the
pulses depends on the bit rate of the codebook. The task of the
ACELP coder is to search the pulse positions and their signs to
minimize a mean square error criterion [2]. While the ACELP
coder is very efficient at low bit rates, it faces some difficulties
at high bit rates. A. Innovation Codebook 0
2
4
6
8
10
12
14
6.6
8.85
12.65 14.25
15.85
18.25 19.85
23.05
bit rate [kbps]
segmental SNR
0
10
20
30
40
50
60
complexity
segSNR [dB]
complexity [WMOPS]
bit rate [kbps]
original
relaxed
original
relaxed
6.6
8.85
12.65 14.25
15.85
18.25 19.85
23.05
Fig. 1. Performance of the original and the relaxed innovation codebook
search in the AMR-WB encoder. In modern speech coding, very large codebooks are needed
in order to guarantee a high subjective quality. In the
AMR-WB codec an algebraic innovation codebook structure
with codebooks as large as 88 bits is used. The codebook
structure is based on interleaved single-pulse permutation
design [5] where 64 positions (corresponding to 5 ms subframe
at 12.8 kHz sampling rate) are divided into 4 tracks of
interleaved positions, 16 positions in each track. Different
codebooks at different bit rates are constructed by placing
a certain number of signed pulses in the tracks, from 1 to 6
pulses per track. The codebook index, or codeword, represents
the pulse positions and signs in each track. In order to keep the computational complexity reasonable,
a fast procedure known as a depth-first tree search [6] is used. The search begins with subset #1 and proceeds with subsequent
subsets according to a tree structure while only 2 pulses are
determined at each tree level. Obviously, the complexity
increases when more pulses are searched. To reduce
complexity
while
testing the
possible
combinations of two pulses at each tree level, a limited number
of potential positions of the first pulse is tested (typically 8 or
fewer positions out of 16). Further, in case of large number
of pulses, some pulses in the higher levels of the search tree are
fixed based on a pulse-position likelihood estimate [7]. Fig. 1. Performance of the original and the relaxed innovation codebook
search in the AMR-WB encoder. used to control the complexity of AMR-WB. At the highest bit
rate with the blind BWE (23.05 kbps), there is a gain of 0.168
dB in segSNR for 20 WMOPS complexity increase when the
relaxed codebook search is used. If we wanted to extend the bit
rates of the AMR-WB codec even higher, the traditional CELP
model would become even less efficient or enormously
complex. III.
TRANSFORM-DOMAIN CODEBOOK All these different constraints ensure that the complexity
does not explode but rather saturates at higher bit rates. This is
however at a price of limited quality gain when the bit rate
increases, as will be shown in the next subsection. In order to overcome the trade-off between the
effectiveness and the complexity of the traditional CELP, we
propose an efficient, flexible and scalable extended CELP
model [9]. This model introduces a transform-domain
codebook that is incorporated into the traditional CELP and
can be seen as a pre-quantizer of the innovation codebook. The
parameters of the new codebook are set at the encoder in such
a way that the subsequent innovation codebook search is
applied to a target signal which has – in the residual domain –
less pronounced spectral dynamics than the target after the
adaptive codebook only. A. Innovation Codebook Then the search algorithm starts with placing two first
pulses at two consecutive tracks and this process is iterated
four times by assigning first two pulses to different starting
tracks at each iteration. However, in order to further reduce the
complexity of the search algorithm the number of the iterations
in the AMR-WB codec is reduced to 3 (at 18.25 and 19.85
kbps) resp. 2 (at 23.05 kbps). II.
AMR-WB CODEC BACKGROUND The AMR-WB codec [5] is currently widely deployed
in mobile communications. It delivers wideband speech with
audio bandwidth of 50–7000 Hz at one of the nine specified
constant bit rates: 23.85, 23.05, 19.85, 18.25, 15.85, 14.25,
12.65, 8.85 and 6.6 kbps. In the AMR-WB codec, the input
audio signal is processed in frames of 20 ms while each frame
is further divided into four subframes. The codec employs
a band-split processing. The low band is coded by the ACELP
model at the internal sampling rate of 12.8 kHz and covers
frequencies up to 6.4 kHz while a band-width extension
(BWE) is used to cover the rest of the spectrum. A blind BWE
is used at all bit rates except of 23.85 kbps, where a guided
(16 bits/frame) BWE is employed. First, the ACELP model does not gain in quality as quickly
as other approaches such as transform coding and vector
quantization when increasing the innovation codebook size. When measured in dB/bit/sample, the gain at higher bit rates ISBN 978-0-9928626-7-1 © EURASIP 2017 1659 2017 25th European Signal Processing Conference (EUSIPCO) B. Transform-Domain Codebook Gain The inner mechanism of the AVQ scales down in
amplitude
the
DCT
coefficients
[10]. Consequently,
the transform-domain codebook gain, β3, is estimated to
compensate for the AVQ scaling as follows Fig. 2. Proposed
transform-domain
codebook
in
the
extended
ACELP encoder. [
]
[
]
−
=
−
=
′
⋅
′
′
⋅
′
=
1
0
1
0
3
)
(
)
(
)
(
)
(
K
k
K
k
in
k
Q
k
Q
k
Q
k
Q
β
(3) )
(
)
(
)
(
1
n
v
n
r
n
qin
⋅
−
=
β
(1) (1) (3) where n = 0,…, N–1 is the time-domain sample index and
N = 64. Further, r(n) is the adaptive codebook search target
vector in the residual domain, i.e., signal x(n) filtered through
1/H(z) with zero states. where k = 0, …, K–1 is the transform-domain coefficient index
and K = 64 is the number of the DCT coefficients in the current
subframe. Then the transform-domain codebook target vector in the
residual domain qin(n) is pre-emphasized with a filter F(z) to
amplify lower frequencies of this vector: Subsequently, the gain is quantized. In our experimental
implementation a 6-bit scalar quantizer is used whereby the
quantization levels are uniformly distributed in the log domain. The index of the quantized gain is transmitted to the decoder
once per subframe as another transform-domain codebook
parameter. Subsequently, the gain is quantized. In our experimental
implementation a 6-bit scalar quantizer is used whereby the
quantization levels are uniformly distributed in the log domain. )1
(
)
(
)
(
−
′
⋅
+
=
′
n
q
n
q
n
q
in
in
in
α
(2) (2) q
y
g
The index of the quantized gain is transmitted to the decoder
once per subframe as another transform-domain codebook
parameter. where the coefficient α = 0.3 controls the level of the pre-
emphasis. Next, a Discrete Cosine Transform (DCT) is applied to the
pre-emphasized
vector
q′in(n)
using
a
rectangular
non-overlapping window resulting in 8 DCT blocks, each
covering one 800 Hz DCT band. These blocks of DCT
coefficients Q′in(k) are then quantized using a vector quantizer. However, depending on the bit budget, some of the blocks
might not be transmitted. Those blocks, corresponding usually
to higher frequencies, are set to zero. C. Reconstruction To obtain the target vector for the innovation codebook
search, a time-domain contribution from the transform-domain
codebook, q(n), is reconstructed as follows. First, the quantized
DCT coefficients Q′(k) are scaled up using the quantized gain
β3. Next, the scaled DCT coefficients are inverse transformed
using inverse DCT (iDCT). Finally, a de-emphasis filter 1/F(z)
is applied to obtain the time-domain contribution q(n). The
same operations are also done in the decoder. A. DCT Quantization In our implementation we have chosen the Algebraic
Vector Quantizer (AVQ) [10] to quantize the DCT coefficients
Q′in(k). The AVQ encoder thus produces quantized DCT
coefficients Q′(k) and the AVQ indices are transmitted as
transform-domain codebook parameters to the decoder. 2017 25th European Signal Processing Conference (EUSIPCO) 2017 25th European Signal Processing Conference (EUSIPCO) Fig. 2. Proposed
transform-domain
codebook
in
the
extended
ACELP encoder. In every subframe, the bit budget allocated to the AVQ
is composed of a fixed bit budget and a floating number of bits. The fixed AVQ bit budget is directly derived from the codec
bit rate. Giving the scalability of the AVQ, practically arbitrary
bit budget can be allocated to the transform-domain codebook. The effectiveness of the codebook scales up with a higher
bit budget as both the number of the quantized DCT blocks and
consequently the SNR increase. In other words, with increasing
the bit budget the level of the coding noise (shaped to follow
the frequency response of the inverse of the weighting filter)
decreases. Then, depending on the used AVQ sub-quantizers [10], the
AVQ usually does not consume all of the allocated bit budget,
leaving a small variable number of bits available in each
subframe. These bits are floating bits employed in the
following subframe within the same frame. The floating
number of bits is equal to 0 in the first subframe, and
the floating bits resulting from the AVQ in the last subframe
in a given frame remain unused or could be used by another
coding module. B. Relaxed innovation codebook To illustrate the impact of the constraints in the innovation
codebook search as described in the previous subsection,
we have conducted the following experiment. We have
modified the innovation codebook search algorithm in the
AMR-WB codec such that at least 8 potential positons are
tested for each pulse and the number of iterations is fixed to
4 at all bit rates. The results showing the difference between
the original AMR-WB algorithm and the relaxed (less
constrained) algorithm are shown in Fig. 1. The principle of the proposed extended ACELP encoder is
depicted in Fig. 2. In contrast to the traditional ACELP where
the excitation is composed of the adaptive excitation vector
v(n) and the innovation excitation vector c(n) only, the
extended model introduces a third part of the excitation,
namely the transform-domain excitation vector q(n). In Fig. 2,
β1 and β2 are the adaptive and innovation codebook gains, x(n)
and x1(n) are the targets for the adaptive and innovation
codebook search, and H(z) denotes the weighted synthesis
filter, which is the cascade of the LP synthesis filter 1/A(z) and
the perceptual weighting filter W(z). In this experiment, a 188 s database consisting of
40 sentences was used. The database contained clean and noisy
speech samples and was sampled at 16 kHz. The segmental
SNR (segSNR) was measured in the perceptually weighted
speech domain and only in the low band. The worst case (WC)
complexity
is
measured
in
terms
of
Weighted
Million Operations Per Second (WMOPS) using the ITU-T
complexity evaluation tool [8]. It can be seen from Fig. 1 that
as the bit rate increases, the differences between both variants
of the codec get larger, which means that more constraints are In a given subframe, the target in the residual domain after
subtracting the adaptive codebook contribution is computed as 1660 ISBN 978-0-9928626-7-1 © EURASIP 2017 D. Target Vectors for Innovation Codebook Search It was experimentally found that the transform-domain
codebook contribution can be used to refine the previously
computed adaptive codebook gain, β1, to enhance the coding 1661 ISBN 978-0-9928626-7-1 © EURASIP 2017 2017 25th European Signal Processing Conference (EUSIPCO) efficiency. Consequently, the adaptive codebook gain is
recomputed as -3.5
-3.0
-2.5
-2.0
-1.5
-1.0
-0.5
0.0
15
20
25
30
35
40
45
50
55
PEAQ
bit rate [kbps]
extended model
DI
ODG
degradation [–]
8
12
16
20
24
15
20
25
30
35
40
45
50
55
segSNR [dB]
bit rate [kbps]
segmental SNR
extended model
Fig. 3. Performance of the transform-domain codebook implemented in the
AMR-WB codec. -3.5
-3.0
-2.5
-2.0
-1.5
-1.0
-0.5
0.0
15
20
25
30
35
40
45
50
55
PEAQ
bit rate [kbps]
extended model
DI
ODG
degradation [–]
8
12
16
20
24
15
20
25
30
35
40
45
50
55
segSNR [dB]
bit rate [kbps]
segmental SNR
extended model
Fig. 3. Performance of the transform-domain codebook implemented in the
AMR-WB codec. {
}
[
]
( )
( )
[
]
−
=
−
=
⋅
⋅
−
=
′
1
0
1
0
1
)
(
)
(
)
(
N
n
N
n
n
y
n
y
n
y
n
w
n
x
β
(4) (4) where w(n) is the filtered transform-domain codebook
contribution, i.e., the zero-state response of the weighted
synthesis filter H(z) to the vector q(n). Similarly y(n) is the
filtered adaptive codebook contribution. Note that in traditional
ACELP the vector x(n) is used instead of {
})
(
)
(
n
w
n
x
−
in (4). Then the computation of the target vector for innovation
codebook search, x1(n), is done using ). (
)
(
)
(
)
(
1
1
n
w
n
y
n
x
n
x
−
⋅′
−
=
β
(5) (5) Finally, the innovation codebook search is performed using
a moderate innovation codebook size. We have experimentally
found that it is advantageous to allocate more bits to the AVQ
quantization of the transform-domain codebook than to the
innovation codebook. On the other hand, a too small
innovation codebook would not be able to capture all the
remaining essential components of the residual signal. IV. PERFORMANCE As mentioned previously, we have implemented the
transform-domain codebook as described in Section III in the
AMR-WB codec. We have chosen several bit rates starting
from 30 kbps that extend the AMR-WB bit rate coverage and
tested if the performance scales-up proportionally with
increased bit rate. Our goal was also to optimize the model
such that the complexity at the extended bit rates does not
increase the WC complexity of the AMR-WB encoder. In our
experimental implementation the traditional AMR-WB internal
sampling rate of 12.8 kHz and the blind BWE were used. Another example is the gain quantization. In AMR-WB,
a 7-bit vector quantizer (VQ) is used to quantize the adaptive
and innovation codebook gains β1 and β2. While this VQ works
sufficiently well at AMR-WB bit rates, its performance is not
good enough at the extended bit rates. It would thus be
advantageous to increase the bit budget of the gain quantizer. One needs to be also very careful about enhancers and post-
processing techniques used in a codec, e.g., the enhancement
of the excitation signal used in AMR-WB [7]. At the extended
bit rates these techniques are usually not needed and could
actually have a negative effect on the synthesised speech
quality. In general, it thus seems beneficial to disable them. Using the same database as described in subsection II.B,
we have computed complexity, segSNR and Perceptual
Evaluation of Audio Quality (PEAQ) scores. The PEAQ [11]
is an objective metric that automatically assesses the audio
quality degradation in terms of the Objective Difference Grade
(ODG) and the Distortion Index (DI). The results are shown
in Fig. 3. They also contain the original higher AMR-WB
bit rates to better observe the trends. V. OTHER CONSIDERATIONS In Section IV we have demonstrated how the new
transform-domain codebook enhances the performance of
a CELP codec by increasing the segSNR and PEAQ scores
without affecting the WC complexity. While the codec
performance scales effectively at the extended bit rates, it is not
possible to reach transparent quality without introducing
further tunings into the codec. An example would be a need of
an improved coding of the high band either by increasing the
internal sampling rate or by introducing a high-quality BWE. D. Target Vectors for Innovation Codebook Search Consequently, the 36-bits innovation codebook was found as
the best choice, and it was used in our experimental
implementation within the AMR-WB codec, regardless of the
codec bit rate. Note that the 36-bits codebook is used in
AMR-WB at the 12.65-kbps bit rate. It places 8 pulses in a
given subframe and it has a computational complexity of about
4 WMOPS below the worst case complexity of the complete
encoder (see Fig. 1). This difference in complexity can be thus
spent to perform the transform-domain codebook search
without affecting the encoder’s WC complexity. bit rate [kbps] Fig. 3. Performance of the transform-domain codebook implemented in the
AMR-WB codec. both in terms of segSNR and PEAQ. We have also verified that
the encoder’s complexity at the extended bit rates indeed does not
exceed the WC complexity of the original AMR-WB encoder. REFERENCES [1] M. R. Schroeder and B. S. Atal, "Code-excited linear prediction
(CELP): High-quality speech at very low bit rates," in Proc. IEEE Int. Conf. on Acoust., Speech and Signal Process. (ICASSP), Tampa, FL,
1985, pp. 937–940, vol. 10. [2] J. P. Adoul, P. Mabilleau, M. Delprat, and S. Morissette, "Fast CELP
coding based on algebraic codes," in Proc. IEEE Int. Conf. on Acoust.,
Speech and Signal Process. (ICASSP), Dallas, TX, 1987, pp. 1957–1960. [3] R. Salami, C. Laflamme, J. -P. Adoul, and D. Massaloux, "A toll quality
8 kb/s speech codec for the personal communications system (PCS),"
IEEE Trans. on Vehicular Technology, vol.43, no.3, pp. 808-816,
August 1994. [4] E.-D. Lee, M. S. Lee, an D. Y. Kim, "Global pulse replacement method
for fixed codebook search of ACELP speech codec," in Proc. IASTED
CIIT 2003, Scottsdale, AZ, 2003, pp. 372-375. [5] B. Bessette, R. Salami, R. Lefebvre, M. Jelinek, J. Rotola-Pukkila,
J. Vainio, H. Mikkola, K. Jarvinen, "The Adaptive Multirate Wideband
Speech Codec (AMR-WB)," IEEE Trans. Speech Audio Process.,
vol. 10, no. 8, pp. 620–636, November 2002. [6] R. Salami, C. Laflamme, B. Bessette, and J. -P. Adoul, "ITU-T G.729
Annex A: reduced complexity 8 kb/s CS-ACELP codec for digital
simultaneous voice and data," IEEE Communication Magazine, vol. 35,
no. 9, pp. 56-63, September 1997. If the transform-domain codebook is applied only in low
frequencies, a larger algebraic innovative codebook is
preferable to get a sufficient quality in higher frequencies. The
64-bits innovation codebook (with 16 pulses per subframe)
has been found perceptually optimal. Based on the fact that
the CELP model was theoretically imagined to produce a
spectrally flat excitation, a high emphasis in the filter F(z)
may be seen inadequate. However in practice, the ACELP,
with its ability to permute pulses, is very efficient to produce
an emphasized excitation. In fact, also the LTP conditions the
target in the same way as the transform-domain codebook
does when the prediction is limited to the low frequencies. [7] 3GPP TS 26.190: Speech codec speech processing functions; Adaptive
Multi-Rate – Wideband (AMR-WB) speech codec; Transcoding
functions. [8] Recommendation ITU-T G.191: Software Tools for Speech and Audio
Coding Standardization (STL), 2010. [9] B. Bessete, "Flexible and scalable combined innovation codebook for
use in CELP coder and decoder," US Patent 9,053,705, June 2015. [10] M. Xie and J.-P. Adoul, "Embedded algebraic vector quantization
(EAVQ) with application to wideband audio coding," In Proc. A. Variable Bit Rate Audio Coding In Section IV the performance of the proposed model has
been assessed for the case of constant bit rate coding. This
is typical in low delay applications such as telephony where
the total bit budget is usually fixed. In higher delay
applications like audio streaming, a bit reservoir is often used It can be seen from the results that the performance of the
AMR-WB codec scales effectively at the extended bit rates, ISBN 978-0-9928626-7-1 © EURASIP 2017 1662 2017 25th European Signal Processing Conference (EUSIPCO) 2017 25th European Signal Processing Conference (EUSIPCO) in conjunction with a variable bit rate encoder. In those
applications, the proposed technology can be further tuned to
profit from the variable bit allocation. Consequently, the audio
quality can be perceptually further improved as the bit
allocation can be source controlled. proposed technology. A bit reservoir was used so that the
model was fully flexible and source controlled in order to
obtain the desired SNR in low frequencies. We have tested
a database consisting of speech, speech over music and music
items where the segSNR was measured only in the frames
operating in the time-domain coding mode. Objectively the
overall performance of the tuned transform-domain codebook
was similar to what is presented in Fig. 3. However, an
informal subjective assessment showed a clear advantage in
controlling the quality in the context of source controlled
variable bit rate coding of a general audio content. It is known that coding of lower frequencies is more
critical for the overall perceptual quality, and thus a higher
SNR is usually targeted in lower frequencies than in higher
frequencies. In the CELP model, this higher SNR is mainly
achieved by the LTP. However, the LTP is not efficient in
every case, such as in voice onsets, transients or in the context
of background noise or speech over music. These scenarios
require a much better control of quality which cannot be
simply achieved by either the LTP or the innovation
codebook. To guarantee a uniform quality in case of a variable
bit rate coding, we have experimented with different tunings
of the transform-domain codebook from those described in
Section III. VI. CONCLUSION A new transform-domain codebook implemented in the
CELP model was introduced. The new codebook is efficient,
flexible and scalable and can easily extend any existing CELP
codec to provide coding at high bit rates. We demonstrated by
the example of the AMR-WB codec that the proposed model
gains in quality at the extended bit rates while not increasing
the computational complexity of the codec. To apply the technology in the context of a variable bit
rate coding, the coefficient α of the pre-emphasis filter F(z)
could be increased in order to emphasize even more the low
frequencies. E.g., α = 0.9 can be chosen to obtain an emphasis
around 20 dB at 20 Hz, 11 dB at 800 Hz and 3 dB at 2400 Hz. Relative to the coded bandwidth (6400 Hz), this area contains
70% of the auditory critical bands [12]. This configuration has
thus the benefit to control the SNR in the area which is
perceptually most important and contributes in the frequency
range which is more likely to be profitable by the LTP. For
example, many vowels are voiced (predictable) in low
frequencies and noisy (unpredictable) in higher frequencies. Using this configuration, the DCT spectrum is quantized only
in the concerned area (≤ 2400 Hz) meaning that only 3 DCT
blocks are quantized in every subframe. In low frequencies,
the transform-domain codebook gain then controls the SNR,
and its bit consumption depends on the targeted SNR and the
LTP efficiency. The weaker the LTP contribution is, the
stronger and richer is the transform-domain codebook
contribution and vice versa. REFERENCES IEEE Int. Conf. on Acoust., Speech and Signal Process. (ICASSP), Atlanta, GA,
1996, pp. 240-243, vol 1. In order to assess the quality of the tuned transform-
domain codebook as described in this subsection, we have
implemented it into the MPEG USAC codec [13]. USAC is
a hybrid audio codec, which consists of a time-domain coding
mode and a frequency-domain coding mode. The time-domain
coding mode is in general used to encode speech segments
(with or without music), transients and attacks. It has a similar
structure as the AMR-WB codec and was extended by the [11] Recommendation ITU-R BS.1387: Method for objective measurements
of perceived audio quality (PEAQ), 2001. [12] J. D. Johnston, "Transform Coding of Audio Signals Using Perceptual
Noise Criteria," IEEE Journal on Selected Areas in Communications,
vol. 6, no. 2, pp. 314-323, February 1988. [13] M. Neuendorf et al., "MPEG Unified Speech and Audio Coding - The
ISO/MPEG Standard for High-Efficiency Audio Coding of All Content
Types," in Proc. 132nd AES Convention, Budapest, Hungary, 2012,
pp. 248–269. 1663 ISBN 978-0-9928626-7-1 © EURASIP 2017
|
https://openalex.org/W4210372449
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|
English
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Additions to the Bombyliid fauna of South Africa as represented in the South African Museum
|
Annals of the South African Museum
| 1,921
|
public-domain
| 5,123
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( 469 ) ( 469 ) 7.—Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera),
as
represented in the South African Musewm.—By Prof. M. Buzz. 7.—Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera),
as
represented in the South African Musewm.—By Prof. M. Buzz. THE Director of the South African Museum has sent me a small
number
of
Bombyliidae
from
South-West
Africa,
obtained
after
the
publication, on February 21st, 1921, of my memoir. These are now
described. I have also added a few others, sent me by Dr. H. Brauns,
that do not figure in my first list. Some of these will be more fully
described in my forthcoming paper on the African Bombylidae belong-
ing to the Hungarian Museum at Budapest. BomBynius PLAGIATUS, N. Sp. Closely allied to B. acroleucus, Bezzi, and belonging to the same
group, but at once distinguishable by the more slender third antennal
joint
and
by
the
wings
having
the
discal
cross-vein
placed
much
beyond the middle of the discoidal cell, and having moreover a broad
fuscous patch ending in a truncate form at the discal cross-vein. Closely allied to B. acroleucus, Bezzi, and belonging to the same
group, but at once distinguishable by the more slender third antennal
joint
and
by
the
wings
having
the
discal
cross-vein
placed
much
beyond the middle of the discoidal cell, and having moreover a broad
fuscous patch ending in a truncate form at the discal cross-vein. Type Q, one specimen from Delarey (W. Transvaal), January, 1917
(Dr. H. Brauns). (
)
Length of body 9 mm.; of wing 10 mm. Length of body 9 mm.; of wing 10 mm. g
y
g
Head and appendages exactly as in B. acroleucus; the third joint
of the antennae is distinctly more slender at its broadest portion and
not broader than the second joint. Thorax as in B. acroleucus, like-
wise with three equal longitudinal stripes of black hairs on the back ;
bristles
entirely
black. Scutellum,
halteres
and
squamae
as
in
B. acroleucus; abdomen of a like pattern, consisting of two broad round
white spots on the sides of second and third segment and a smaller
one in the middle; the terminal segments have the median spots more
developed, being thus almost entirely white ; the terminal bristles are
black
at
base
and
white
at
end. Legs
as
in
B. acroleucus;
the
wings have the same nervation, but are longer, and therefore the dis-
coidal cell is elongate and the second and third posterior cells shorter ;
the discal cross-vein is placed on the terminal third of the discoidal cell. There is, moreover, a well-developed pattern, the extreme base to the
basal
cross-veins
being
black
as
in
B. acroleucus;
following
is
a
yellowish patch extending from the costa to the whole of the second
basal cell and filling up the whole of the costal cells; then a broad 30 of
the
South
African
Museum. ADELIDEA
(SOBARUS)
AND
SOSIOMYIA. It seems that the species of this peculiar South African group are
more numerous than was at present believed. There can be no doubt
that Adelidea fuscipennis, Macquart, is the same insect as Sobarus
anomalus,
Wiedemann
and
Loew,
and
even
as
Cyllenia
longirostris,
Wiedemann; all three are one species. I have seen, however, two
other species which are certainly different, and therefore I will give
the following key for all the known species. 1 (2). Third antennal joint beset with long bristles above; antennae entirely
black; femora black; tibiae with partly black spicules; wings with a
rounded fuscous spot at end of the anal cell . comata, Bezzi, 1921. 1 (2). Third antennal joint beset with long bristles above; antennae entirely
black; femora black; tibiae with partly black spicules; wings with a
rounded fuscous spot at end of the anal cell . comata, Bezzi, 1921. Bezzi, loc. cit., p. 3. Bezzi, loc. cit., p. 3. 2 (1). Third antennal joint quite bare above; legs entirely reddish with
quite reddish spicules ; no fuscous spot at end of anal cell. 3 (4). Basal joints of the antennae red ; wings with the anterior half yellowish
and with fuscous spots on cross-veins ; : ruficornis, nN. sp. 3 (4). Basal joints of the antennae red ; wings with the anterior half yellowish
and with fuscous spots on cross-veins ; : ruficornis, nN. sp. 4 (8). Antennae entirely black; wings with the anterior half infuscated or
entirely fuscous. 5 (6). Wings with the anterior half infuscated and with fuscous spots on
cross-veins ; discoidal cell much longer than the second posterior cell. anomalaWied * ,
6 (5). Wings equally and intensively infuscated, not spotted; discoidal cell
only a little longer than the second posterior cell braunsi, n. sp. ,
6 (5). Wings equally and intensively infuscated, not spotted; discoidal cell
only a little longer than the second posterior cell braunsi, n. sp. 6 (5). Wings equally and intensively infuscated, not spotted; discoidal cell
only a little longer than the second posterior cell braunsi, n. sp. BomBynius PLAGIATUS, N. Sp. 470 fuscous patch of rectangular shape, extending fromthe face of the second
longitudinal vein to the discal cross-vein and ending there in a straight
line; this fuscous patch extends below a little into the upper part of
the discoidal cell and into the apical part of the second basal cell and
also along the fifth longitudinal vein. SOSIOMYA,
Bezzi. Ann. 8. Afr. Mus., 8, 1921, p. 67. 2. Length of body and wing 8 mm. Wings yellowish at base and on middle of fore border,
grey on the remainder; the dark spots are on the praefurca, on the
discal cross-vein (the largest of all), on base of cubital fork and on
the apical cross-vein of the second basal cell; 2 less developed spots
are on the apical cross-vein of the discoidal cell and on the external
inner angle of the third posterior cell; there is no trace of fuscous
spot at end of the anal cell. Venation as in A. anomala, with the Head yellowish, clothed with greyish tomentum, which is darker on
the occiput ; ocellar tubercle blackish; frons about as broad as one
eye,
clothed
with
short
pale
yellowish
hairs,
and
having
4-5
long
yellowish
bristles
on
each
side;
face
with
short
whitish
hairs,
and
with a long mystax of yellowish, bristle-like hairs at mouth border ;
peristome
narrow
and
whitish;
beard
white;
upper
occipital
hairs
yellowish. Antennae
with
the
2
basal
joints
red;
the
first
joint
is
slender and long, about three times as long as the second joint,
clothed above with short and below with long yellowish hairs; third
joint black, slender, more slender than the first, as long as the 2 first
joints together, with a short, hair-like terminal style. Palpi blackish,
short;
proboscis
black,
4
mm. long. Thorax
black,
clothed
with
golden tomentum on the back, and with whitish tomentum on the
pleurae;
humeri
and
notapleural
region
reddish;
a
short
stripe
of
white hairs on the sides above the root of the wings; lateral bristles
long, strong, rather numerous, of a reddish colour, but the postalar
ones are in part blackish; mesopleura with long white hairs and with
a
row
of
yellowish
bristles
at
the
hind
border;
sternopleura
with
short white hairs; metapleura bare. Scutellum entirely black, clothed
with golden tomentum, and having 2-3 rows of long blackish bristles
at the hind border. Squamae and halteres whitish, the former with
white frmge. Abdomen black, densely clothed with golden tomentum,
the last segments reddish at hind border; the hairs are short, scarce
and yellowish, but all the segments except the first have a row of
erect
blackish
bristles
at
the
hind
border. Venter
reddish,
clothed
with whitish tomentum and white hairs, which are denser towards the
sides, forming a white longitudinal stripe on each side. 2. Length of body and wing 8 mm. 2. Length of body and wing 8 mm. 2. Length of body and wing 8 mm. Head yellowish, clothed with greyish tomentum, which is darker on
the occiput ; ocellar tubercle blackish; frons about as broad as one
eye,
clothed
with
short
pale
yellowish
hairs,
and
having
4-5
long
yellowish
bristles
on
each
side;
face
with
short
whitish
hairs,
and
with a long mystax of yellowish, bristle-like hairs at mouth border ;
peristome
narrow
and
whitish;
beard
white;
upper
occipital
hairs
yellowish. Antennae
with
the
2
basal
joints
red;
the
first
joint
is
slender and long, about three times as long as the second joint,
clothed above with short and below with long yellowish hairs; third
joint black, slender, more slender than the first, as long as the 2 first
joints together, with a short, hair-like terminal style. Palpi blackish,
short;
proboscis
black,
4
mm. long. Thorax
black,
clothed
with
golden tomentum on the back, and with whitish tomentum on the
pleurae;
humeri
and
notapleural
region
reddish;
a
short
stripe
of
white hairs on the sides above the root of the wings; lateral bristles
long, strong, rather numerous, of a reddish colour, but the postalar
ones are in part blackish; mesopleura with long white hairs and with
a
row
of
yellowish
bristles
at
the
hind
border;
sternopleura
with
short white hairs; metapleura bare. Scutellum entirely black, clothed
with golden tomentum, and having 2-3 rows of long blackish bristles
at the hind border. Squamae and halteres whitish, the former with
white frmge. Abdomen black, densely clothed with golden tomentum,
the last segments reddish at hind border; the hairs are short, scarce
and yellowish, but all the segments except the first have a row of
erect
blackish
bristles
at
the
hind
border. Venter
reddish,
clothed
with whitish tomentum and white hairs, which are denser towards the
sides, forming a white longitudinal stripe on each side. Legs quite
reddish, the last tarsal joint only black at end; coxae white tomentose,
and white pilose; all the bristles of the femora and the spicules of
tibiae are reddish or yellowish, the hind femora having a row of 4-5
below near the end, and another row of 2-3 on the outer side at end. Claws black and thin, little curved ; pulvilli whitish and shorter than
the claws. ADELIDEA
RUFICORNIS,
0.
sp. Closely allied to A. anomala and possibly the unknown female of
this species, but distinguished by the red basal joints of the antennae
and by the not infuscated fore half of wings. * Of this species I have seen only a rather old male specimen in the British
Museum, 471 Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera). One
female
specimen
from
Willowmore
(Cape),
November,
1920
(Dr. H. Brauns). One
female
specimen
from
Willowmore
(Cape),
November,
1920
(Dr. H. Brauns). 2. Length of body and wing 8 mm. Legs quite
reddish, the last tarsal joint only black at end; coxae white tomentose,
and white pilose; all the bristles of the femora and the spicules of
tibiae are reddish or yellowish, the hind femora having a row of 4-5
below near the end, and another row of 2-3 on the outer side at end. Claws black and thin, little curved ; pulvilli whitish and shorter than
the claws. Wings yellowish at base and on middle of fore border,
grey on the remainder; the dark spots are on the praefurca, on the
discal cross-vein (the largest of all), on base of cubital fork and on
the apical cross-vein of the second basal cell; 2 less developed spots
are on the apical cross-vein of the discoidal cell and on the external
inner angle of the third posterior cell; there is no trace of fuscous
spot
at
end
of
the
anal
cell
Venation
as
in
A
anomala
with
the of
the
South
African
Museum. 472 discoidal cell much longer than the second posterior cell; discal cross-
vein on the last third of the discoidal cell; first posterior cell narrowed
at end, about as broad as the anal cell at end. Basal hook yellowish ;
basal
comb
small
and
clothed
with
yellowish
hairs;
alula
rounded,
yellowish, not fringed. Veins reddish at base, blackened at end. * The above described species have nothing to do with the recently erected
genus Conophorina, Becker, ‘ Entom. Mitteil.,’ ix, 1920, pp. 181-184, figs. 1-3, the
type species of which, C. biceliaris, Becker, was likewise collected near Willow-
more (Cape), by Dr. H. Brauns; it has an incrassated first antennal joint and
only 2 submarginal cells on the wings. Ann. 8. Afr. Mus., xviii, p. 84. In the male specimen the anal cell is closed and shortly stalked,
while in the female the same cell is broadly open. This makes the
distinction
of
H. soror,
Bezzi,
1921,
very. doubtful. Mafa
(Ovambo
Land),
February,
1921
(K. H. Barnard). PHTHIRIINAK. GONARTHRUS,
Bezzi. Ann. 8. Afr. Mus., xviii, p. 88. PHTHIRIINAK. GONARTHRUS,
Bezzi. Ann. 8. Afr. Mus., xviii, p. 88. ADELIDEA BRAUNSI, N. sp. A small species, at once distinguished by the equally infuscated,
not spotted wings, which have, moreover, a short discoidal cell. Two
2
specimens
from
Willowmore
(Cape),
April,
1920
(collected
by Dr. H. Brauns, in whose honour the species is named). The present species cannot be the A. fuscipennis of Macquart as
one
may
conclude
from
the
name,
the
wings
being
described
as
yellowish at base and dark spotted. y
p
2. Length
of
body
4-45
mm.;
of
wing
5-6
mm. Head
black,
densely clothed with dark grey dust; occiput with dense yellowish hairs ;
frons broader than one eye, with short black bristles on the sides. Antennae
quite
black,
the
first
jomt
with
scarce
black
hairs;
third
joint as long as the 2 first joints together, with a short black terminal
style. Face
rounded,
convex,
blackish
brown,
with
short
blackish
hairs
at
sides
and
at
mouth
border. Palpi
black,
long;
proboscis
black,
3-3°5
mm. long;
beard
scarce
and
yellowish. Thorax
entirely
black, with reddish humeri; on the back it is clothed with yellowish
tomentum, and has numerous yellowish bristles at sides, the supra-
alar ones being darker; on the pleurae the tomentum is more grey,
and there are yellowish hairs only on the meso- and sternopleura, the
rest
being
bare. Squamae
blackish,
with
scarce
yellowish
fringe
;
halteres
whitish. Scutellum
entirely
black
and
rather
shiny,
clothed,
like the back of the mesonotum, with several rows of blackish bristles
at hind border. Abdomen entirely black, rather shiny and yellowish
tomentose; first segment with dense white hairs at sides; a row of
blackish thin bristles at hind border of the segments. Venter entirely
black,
yellowish
tomentose,
clothed
with
dark
hairs. Legs
reddish,
with black coxae and black tarsi except at base; the femora are
blackened
near
the
base;
all
the
bristles
and
spicules
reddish
or
yellowish ; hind femora below without bristles, but with a row of 5-6
at
the
outer
side
near
the
end. Claws
reddish,
with
black
tips;
pulvilli short, yellowish. Wings equally and rather intensely infuscated,
almost
blackish,
without
darker
spots
on
cross-veins,
but
with
a
broad whitish praediscoidal spot, which is very striking against the
infuscation. Basal comb not distinct, alula rounded, blackish, without
fringe. Venation as in two preceding species, but the first posterior Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera). CorsoMYZA
SIMPLEX,
Wiedemann,
1820. A couple of specimens from Willowmore (Cape), September, 1920,
Dr. H. Brauns. The
female
greatly
resembles my
Megapalpus fulvi-
ceps and may even be the same. A couple of specimens from Willowmore (Cape), September, 1920,
Dr. H. Brauns. The
female
greatly
resembles my
Megapalpus fulvi-
ceps and may even be the same. ADELIDEA BRAUNSI, N. sp. 473 cell not narrowed at end, the discal cross-vein placed near the middle
of the discoidal cell, this last cell not much longer than the second
posterior cell, anal cell very broadly open. All the veins are black,
except the first, which is yellowish.* PHTHIRIINAK. GONARTHRUS CHIONEUS, Bezzi. One specimen from Willowmore (Cape), Dr. H. Brauns. Note.—In my paper, loc cit., p. 4, in the key of genera the name
Pseudoamictus, Big., is used, while on p. 94 I employed the name
Pseudempis. The two names are synonyms, and that of Bigot must
be used because it is a valid one (even if not described), its type-
species having been indicated. of
the
South
African
Museum. 474 TOMOMYZA,
Wied. ToMOMYZA PICTIPENNIS, Bezzi, n. sp. A very distinct species, resembling a small specimen of Henica, but
belonging to the gen. Tomomyza in its true sense, as indicated in my
key of the genera (‘ Ann.S. Afr. Mus.,’ xvii, p.5). The Mediterranean
species, attributed wrongly to the present genus, must be removed to
Stomylomyia, Big., and belongs to the Lomatiinae. Several specimens from Willowmore (Cape), December and January,
Dr. H. Brauns ; the species will be more fully described in my paper
on the Bombyliidae of the Hungarian Museum. PANTOSTOMUS GIBBIVENTRIS, Bezzi, n. sp. A species very distinct on account of its gibbose abdominal seg-
ments. Some
specimens
from
Willowmore
(Cape),
December
and
January, Dr. H. Brauns. The new gen. Pantostomus of the Cyllentinae
agrees with Tomomyza in the absence of ocelli, in the hairy metapleura
and
in
other
characters;
but
it
is
at
once
distinguished
by
the
face
being
practically
wanting,
the
mouth-opening
extending
to
the
base of the antennae. Both the new genus and the new species will be
fully described in my forthcoming paper on the Hungarian Museum
material. THYRIDANTHRAX
LUTULENTUS,
0 Near
7. leucoproctus,
but
distinguished
by
the
greater
size,
the
dense tufts of yellow hairs on the sides of thorax and by the more
yellowish base of the wings. Type
2,
a
single
specimen
from
8.W. Protectorate,
Otjiverongo,
April, 1921 (J. S. Brown). Type
2,
a
single
specimen
from
8.W. Protectorate,
Otjiverongo,
April, 1921 (J. S. Brown). Q. Length
of
body
11
mm.;
of
wing
10
mm. Head’
entirely
black
;
occiput
with
white
scales
at
eye
border,
chiefly
near
the
indentation. Frons
clothed
with
short
black
hairs,
more
densely
on the anterior half. Face bluntly convex, not much, yet distinctly
prominent, with black hairs at mouth border and with yellowish ones
at sides. Antennae quite black, with the third joint elongate conical. 475 Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera). Proboscis
black,
not
projecting
beyond
the
mouth. Thorax
entirely
black,
clothed
on
the
back
with
shining
scales,
which
in
certain
lights are whitish, and seems to be disposed in front on three broad
longitudinal
stripes;
the
collar
and
the
sides
along
the
notopleural
lime bear long and dense tufts of golden-yellow hairs; pleurae black,
with
like
tufts
on
pro-,
meso-
and
metapleura. Scutellum
black,
clothed,
like
the
back
of
mesonotum,
with
black
bristles
at
hind
border. Squamae
brown,
with
white
fringe;
halteres
whitish. Abdomen entirely. black; the hairs of sides are white at base and
black on the remainder; the third segment has a broad, complete,
transverse band of white scales at.the base, and similar bands are to
be seen on the sixth and-seventh segment, the remainder being clothed
with
black
scales. Venter
black;
spines
of
the
ovipositor
reddish
brown. Legs black, with smooth front tibiae; front tarsi with short
thin hairs ; hind femora with a complete row of bristles below ; spicules
of the 4 posterior tibiae black and long ; hairs of coxae black like all the
others
;
claws
simple;
no
pulvilli. Wings
proportionately
long
and
broad ; they are hyaline, iridescent, with yellowish base and with a
faint but distinct yellowish tint along the costal cells and into the
basal cells. The veins are yellow, blackened to the terminal half, but
the costa is entirely black. Basal hook slender, long, curved, black ;
basal comb golden, with black bristles at border. THYRIDANTHRAX
LUTULENTUS,
0 Second longitudinal
vein originating opposite to the discal cross-vein, with broad and flat
loop at end; upper branch of the cubital fork much retreating at base ;
discal cross-vein placed before the middle of the discoidal cell; first
posterior cell long and narrow and slightly but distinctly narrowed at
end; second posterior cell at end three times broader than the pre-
ceding one and as broad as the following; fourth posterior cell as
broad as the two preceding cells taken together, its basal contact with
the discoidal cell short, being only a quarter of that of the preceding
cell
with
the
same
cell;
anal
cell
broadly
open. Alula
rounded,
yellowish, with white fringe; axillary lobe broad, hyaline iridescent. Exoprosopa
(HXOPROSOPA)
ACRODISCOIDES,
0.
sp. A species of the seniculus group, near morosa and ignava, but
distinct from both on account of the more lightly coloured body and
wings, and of the quite whitish metapleural tuft. Type Q from Nomtele (Ovamboland), February, 1921, and a badly
preserved
male
from
Mafa
(Ovamboland),
February,
1921
(K. H. Barnard). ; )
3,
2. Length
of
body
12-15
mm.;
of
wing
13-16
mm;
spread
-31-37
mm. Occiput
black,
whitish
tomentose
at
border
of
eyes. Frons of the male only a little more broad than that of the female; it
is black on distal half and red on anterior half above the antennae,
but more broadly in the female than in the male. The face is likewise
more broadly red in the female with a black median spot, while in the
male it is black with a red spot on each side. Frons with black hairs on
distal half and with yellowish tomentum in front, like the face, which
has whitish hairs on the sides and is of conical shape ; mouth borders
pale yellowish. The basal joints of the antennae are reddish below
and black above, and are darker in the male; third joint wanting in
ease of types. Proboscis black, not projecting. Thorax black, clothed
with yellowish scaly tomentum on the back, and with tufts of whitish
hairs
on
the
sides;
bristles
black;
pleurae
reddish,
black
spotted,
with whitish hairs even on the metapleurae. Scutellum reddish, with
a narrow black base, clothed, like the back of the mesonotum, with
black bristles at hind border. Squamae brown with whitish fringe ;
halteres
yellowish. Abdomen
black,
with
reddish
sides
and
hind
border of segments, in the male almost red, with black spots at base
of each segment; the hairs of sides are white near the base, yellowish
on the rest with a few black ones; it is clothed above with yellowish
scales like those of the back of thorax, but at the base of the second
segment and, as it seems, on the second to third terminal segments, there
are transverse bands of whitish scales. Venter red, unspotted in the
male, with broad black bands in the female, both with whitish tomen-
tum. Spines of the ovipositor reddish. Legs black, but with a reddish-
yellow appearance on account of the dense vestiture of scales; spines
and spicules black; front tibiae long and smooth; front tarsi shiny )
3,
2. LITORRHYNCHUS
DiILaTATus,
Bezzi
(1921). Very like a small specimen of ZL. mawrus and likewise with black
and black-fringed squamae, but belonging to the section of the genus
in which the second posterior cell at end is not or only a little more
narrow than the third, the vein between them being less twisted. One
2
specimen
from
S.W,
Protectorate,
Otjiverongo,
April,
1921
(J. S. Brown). One
2
specimen
from
S.W,
Protectorate,
Otjiverongo,
April,
1921
(J. S. Brown). 476 of
the
South
African
Museum. The species was described from Central Africa, but seems to be
widely spread. In the present specimen the central hyaline spot of
the discoidal cell is rather broad, and the anal cell is narrowly hyaline
at end. Exoprosopa
(HXOPROSOPA)
ACRODISCOIDES,
0.
sp. Length
of
body
12-15
mm.;
of
wing
13-16
mm;
spread
-31-37
mm. Occiput
black,
whitish
tomentose
at
border
of
eyes. Frons of the male only a little more broad than that of the female; it
is black on distal half and red on anterior half above the antennae,
but more broadly in the female than in the male. The face is likewise
more broadly red in the female with a black median spot, while in the
male it is black with a red spot on each side. Frons with black hairs on
distal half and with yellowish tomentum in front, like the face, which
has whitish hairs on the sides and is of conical shape ; mouth borders
pale yellowish. The basal joints of the antennae are reddish below
and black above, and are darker in the male; third joint wanting in
ease of types. Proboscis black, not projecting. Thorax black, clothed
with yellowish scaly tomentum on the back, and with tufts of whitish
hairs
on
the
sides;
bristles
black;
pleurae
reddish,
black
spotted,
with whitish hairs even on the metapleurae. Scutellum reddish, with
a narrow black base, clothed, like the back of the mesonotum, with
black bristles at hind border. Squamae brown with whitish fringe ;
halteres
yellowish. Abdomen
black,
with
reddish
sides
and
hind
border of segments, in the male almost red, with black spots at base
of each segment; the hairs of sides are white near the base, yellowish
on the rest with a few black ones; it is clothed above with yellowish
scales like those of the back of thorax, but at the base of the second
segment and, as it seems, on the second to third terminal segments, there
are transverse bands of whitish scales. Venter red, unspotted in the
male, with broad black bands in the female, both with whitish tomen-
tum. Spines of the ovipositor reddish. Legs black, but with a reddish-
yellow appearance on account of the dense vestiture of scales; spines
and spicules black; front tibiae long and smooth; front tarsi shiny Additions
to
the
Bombyliid
Fauna
of
South
Africa
(Diptera). 477 pilose;
claws
with
long
and
acute
basal
tooth. Exoprosopa
(HXOPROSOPA)
ACRODISCOIDES,
0.
sp. Wings
whitish
hyaline; the fuscous pattern is faintly developed, but ends distinctly
in a truncate form into the marginal cell opposite to the marginal
cross-vein; the basal and the discoidal cell are less infuscated, and
therefore there is a fuscous patch at the end of the discoidal cell
which seems sometimes to be isolated as in the species of the EH. heros-
group. First
posterior
cell
narrowed
at
end,
being
only
a
little
broader than the anal cell at end; the terminal cross-vein of the
discoidal
cell
is
horizontal
and
somewhat
S-shaped;
moreover,
the
discoidal cell has a projection into the third posterior cell of a rather
angular shape and appendiculate as in the subgenus Acrodisca, from
which the present species is distinguished im having elongate and not
spiculose front tibiae. Exoprosopa
(ExOPROSOPA)
ATRINASIS,
Speis
(1910). A species of proportionately greater size, very distinct on account of
its large wings, which are infuscated along the fore border, and have
the cross-vein edged with fuscous, but not in the shape of rounded
spots as in the balioptera-group. In the key of the species (‘ Ann. S. Afr. Mus.,’ xvii, p. 151) the present species should follow No. 39,
being
distinct
from
Juteicosta
owing
to
the
more
developed
wing
pattern. 1° from Pemba, N. Rhodesia (Father Cassett), and another female
from
Otjiverongo,
S.W. Protectorate,
February,
1921
(x. H. Barnard). The species is known from East Africa and Abyssinia, but was also
found in Nyassaland ; it is pre-eminently a centro-oriental form. ADDITIONS
AND
CorrReEcTIONS
TO
MonoGrapH
oF
SourH
AFRICAN
Bowsriiup#
(Vou. XVIII,
Pr. I,
pp. 1—180). ADDITIONS
AND
CorrReEcTIONS
TO
MonoGrapH
oF
SourH
AFRICAN
Bowsriiup#
(Vou.
XVIII,
Pr.
I,
pp.
1—180). p
(
,
,
pp
)
P. 4, 1. 87, and p. 178, 1. 34, read Pseudoamictus instead of Pseudowmictus and
Pseudomnictus. P. 4, 1. 87, and p. 178, 1. 34, read Pseudoamictus instead of Pseudowmictus and
Pseudomnictus. . 5, 1. 12, and p. 179, 1. 6, read Tomomyza instead of Tomomyra. . 9, after Bombylius bombiformis add spec. nov. . 52, after Anastoechus macrophthalmus add spe p
p
69, 1. 19, and p. 176, 1. 17, read comata instead of carnata. p
p
69, 1. 19, and p. 176, 1. 17, read comata instead of carnata. 69, 1. 19, and p. 176, 1. 17, read comata ins . 78, after Corsomyza anceps add spec. nov. . 78, after Corsomyza anceps add spec. nov. 88, after Gonarthrus add gen. nov. HHH 88, after Gonarthrus add gen. nov. HHH 99, after Geron barbatus add spec. nov. HHH 105, after Toxophora punctipennis add spec. nov. 130, after Oestranthrax delete (1912) and add gen. nov. . 134, after Thyridanthrax transiens add spec. nov. . 136, after Thyridanthraz ternarius add spec. noy. . 189, after Metapenta add sub. gen. nov. dl of
the
South
African
Museum. 478 P. 149, after Pterobates add sub. gen. nov. P. 155, after Exoprosopa infumata add spec. nov. P. 159, after Ezoprosopa perpulchra add spec. nov. P. 160, after Exoprosopa pediformis add spec. nov. INDEX. PAGE. |
PAGE. A. |
H. acrodiscoides
(Exoprosopa)
. 476)
Hyperusia
. :
:
:
. 473
Adelidea
. 5
,
:
. 470
|
L
anomala
(Adelidea)
. ;
|
ATO
were
;
atrinasis
(Exoprosopa)
;
ATT
ee
ry
eee
;
inte
longirostris
(Cyllenia)
;
. 470
B. lutulentus
(Thyridanthrax)
. 474
bicellaris
(Conophorina)
. . 473
M. BoMBYLIIDAE. :
;
. 469
|
Megapalpus
;
:
:
. 473
Bombylius
. ,
3
;
. 469
minor
(Hyperusia)
. :
. 4783
braunsi
(Adelidea)
. :
. 472
A
C. |
Pantostomus
:
:
:
. ATA
chioneus
(Gonarthrus)
:
. 473
|
Ist
Satna
? :
:
Bell
Conophorina
;
_
473
|
Pictipennis
(Tomomyza)
. . A474
Coromyza
-
_
473
|
plagiatus
(Bombylius)
:
. 469
Cyllenia
en
ARO
|
Pseudempis
. ;
:
:
. AT3
CrmmmNgeyamies
wk
ATA
EEE
Tee
via
Carel
|
RB. D. |
ruficornis
(Adelidea)
. 5
. 470
dilatatus (Litorrhynchus) . . ATS |
|
S. E. |
simplex
(Corsomyza)
. a
Las
EXOPROSOPINAE
. 5
:
. 474
|
Sobarus
:
:
‘
;
. ATO
Exoprosopa
. :
:
. 476
|
soror
(Hyperusia)
:
:
. 413
|
Sosiomya
. 3
. :
. 470
F. nt
fulviceps
(Megapalpus)
. . 473
‘Titanate
it
474
G
Tomomyza
. :
:
5
. 474
gibbiventris
(Pantostomus)
. ATA
U. Gonarthrus
. :
5
:
. 473
|
Ustrnar
i
;
:
:
. A738 1921. "Additions to the Bombyliid fauna of South Africa as represented in the
South African Museum." Annals of the South African Museum. Annale van die
Suid-Afrikaanse Museum 18, 469–478. https://doi.org/10.5962/bhl.part.8028. This file was generated 1 April 2024 at 09:24 UTC Copyright & Reuse Copyright & Reuse
Copyright Status: Public domain. The BHL considers that this work is no longer under
copyright protection. This document was created from content at the Biodiversity Heritage Library, the world's
largest open access digital library for biodiversity literature and archives. Visit BHL at
https://www.biodiversitylibrary.org. This file was generated 1 April 2024 at 09:24 UTC
|
https://openalex.org/W2049779453
|
https://europepmc.org/articles/pmc2978097?pdf=render
|
English
| null |
Inhibiting α-Synuclein Oligomerization by Stable Cell-Penetrating β-Synuclein Fragments Recovers Phenotype of Parkinson's Disease Model Flies
|
PloS one
| 2,010
|
cc-by
| 11,484
|
Introduction substantia
nigra
pars
compacta
with
presence
of
eosinophillic,
intracytoplamic, proteinaceous inclusions termed Lewy bodies
(LB) and dystrophic Lewy neuritis (LN) in surviving neurons. Among the clinical features of PD are motor impairments
involving resting tremor, bradykinesia, postural instability and
rigidity along with non-motoric symptoms such as autonomic,
cognitive and psychiatric problems. The cause of these patholog-
ical characteristics is not yet fully understood but it is believed that
environmental factors as well as a genetic causation or a
combination of the two might result in the abovementioned
clinical syndromes [17]. It is now known that less than 10% of the
PD cases have a strict familial etiology while the majority of cases
are sporadic [18]. Among the mutations associated with familial
PD, three missense mutations in the a-synuclein (a-syn) gene
termed A53T, A30P and E46K have been widely characterized
[19,20]. The
role
of
protein
oligomerization
and
deposition
in
neurodegenerative
diseases
has
become
evident
in
diverse
disorders
including
Alzheimer’s
disease
(AD),
Huntington’s
disease, Parkinson’s disease (PD), Creutzfeldt-Jacob disease, prion
disease, and type II diabetes [1–8]. In each case of these diseases, a
different endogenous protein self-assembles into highly ordered
fibrillar structures. Though there is no specific sequence homology
among these proteins, they are all thought to involve important
conformational changes in the corresponding protein, usually the
production of b-sheet structures which have a strong tendency to
aggregate into water-insoluble fibrous polymers [5,9]. It is
currently debated whether the precipitated insoluble fibrils or
actually soluble oligomers are the cytotoxic aggregative elements
that are associated with the pathologies of each of these diseases
[1,10–16]. Moreover, it is also not yet clear whether the soluble
oligomers are formed in an on-pathway fashion as intermediates
eventually leading to the non-soluble fibrils and plaques or formed
off- pathway by a separate mechanism. The 140 amino acid a-syn protein is a small, highly charged,
natively unfolded protein, first identified in 1977 as the major
component of LB and LN [21,22]. This was followed by the
discovery of the point mutations in its gene [23]. Besides its role in
PD, the protein is associated with pathological inclusions of several
other neurodegenerative diseases including dementia associated
with Lewy bodies, LB variant of AD, and multiple system atrophy
[24]. Abstract The intracellular oligomerization of a-synuclein is associated with Parkinson’s disease and appears to be an important target
for disease-modifying treatment. Yet, to date, there is no specific inhibitor for this aggregation process. Using unbiased
systematic peptide array analysis, we indentified molecular interaction domains within the b-synuclein polypeptide that
specifically binds a-synuclein. Adding such peptide fragments to a-synuclein significantly reduced both amyloid fibrils and
soluble oligomer formation in vitro. A retro-inverso analogue of the best peptide inhibitor was designed to develop the
identified molecular recognition module into a drug candidate. While this peptide shows indistinguishable activity as
compared to the native peptide, it is stable in mouse serum and penetrates a-synuclein over-expressing cells. The
interaction interface between the D-amino acid peptide and a-synuclein was mapped by Nuclear Magnetic Resonance
spectroscopy. Finally, administering the retro-inverso peptide to a Drosophila model expressing mutant A53T a-synuclein in
the nervous system, resulted in a significant recovery of the behavioral abnormalities of the treated flies and in a significant
reduction in a-synuclein accumulation in the brains of the flies. The engineered retro-inverso peptide can serve as a lead for
developing a novel class of therapeutic agents to treat Parkinson’s disease. Citation: Shaltiel-Karyo R, Frenkel-Pinter M, Egoz-Matia N, Frydman-Marom A, Shalev DE, et al. (2010) Inhibiting a-Synuclein Oligomerization by Stable Cell-
Penetrating b-Synuclein Fragments Recovers Phenotype of Parkinson’s Disease Model Flies. PLoS ONE 5(11): e13863. doi:10.1371/journal.pone.0013863 Editor: Mark R. Cookson, National Institutes of Health, United States of America Received May 4, 2010; Accepted October 15, 2010; Published November 10, 2010 Copyright: 2010 Shaltiel-Karyo et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by funding from the Israeli Science Foundation (ISF) and the German-Israeli Project Cooperation (D
no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. supported by funding from the Israeli Science Foundation (ISF) and the German-Israeli Project Cooperation (DIP) to UG. The funders had
data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: vpr@tauex.tau.ac.il Ronit Shaltiel-Karyo1, Moran Frenkel-Pinter1, Nirit Egoz-Matia1, Anat Frydman-Marom1, Deborah E.
Shalev2, Daniel Segal1, Ehud Gazit1* 1 Department of Molecular Microbiology and Biotechnology, Tel Aviv University, Tel Aviv, Israel, 2 Wolfson Centre for Applied Structural Biology, Hebrew University of
Jerusalem, Jerusalem, Israel ogy and Biotechnology, Tel Aviv University, Tel Aviv, Israel, 2 Wolfson Centre for Applied Structural Biology, Hebrew University of of Molecular Microbiology and Biotechnology, Tel Aviv University, Tel Aviv, Israel, 2 Wolfson Centre for Applied Structural Biology, He
usalem, Israel PLoS ONE | www.plosone.org Inhibiting a-Synuclein Oligomerization by Stable Cell-
Penetrating b-Synuclein Fragments Recovers Phenotype
of Parkinson’s Disease Model Flies Ronit Shaltiel-Karyo1, Moran Frenkel-Pinter1, Nirit Egoz-Matia1, Anat Frydman-Marom1, Deborah E. Shalev2, Daniel Segal1, Ehud Gazit1* Expression and purification of a-syn The protein was expressed in pT7-7 BL21 E. coli bacteria. For
over expression, bacterial cultures were grown to logarithmic
stage in the presence of ampicillin (100 mg/l) and protein
expression was induced using IPTG (1 mM) for 3 hours. The
bacterial pellet was re-suspended in TEN buffer (50 mM Tris
pH 8.0, 10 mM EDTA, 150 mM NaCl), and frozen at 280uC
until purification. The sequence of the three synuclein protein is highly conserved,
especially within their N terminal domains. When the sequences of
a-syn and b-syn are compared, a major difference within the
hydrophobic central domain is apparent: b-syn, a 134 amino acid
protein, lacks the NAC region of a-syn and, in contrast to the
latter, does not aggregate to form amyloid fibrils under various
stress conditions such as free radicals or increased concentration
[9,28]. Moreover, b-syn was shown to inhibit the aggregation of a-
syn in vitro in a dose-dependent manner [29,30]. Uversky et al. proposed that this inhibition occurs due to the incorporation of b-
syn into oligomeric intermediates which in turn leads to their
stabilization. It is hypothesized that b-syn interacts with a putative
transient oligomeric intermediate of a-syn, normally on its way to
form fibrils, resulting in the stabilization of the a-syn intermediate
and blocking its conversion into fibrils [31]. Protein purification was accomplished using a non-chromato-
graphic method as described by Volles and Lansbury [33]. Briefly,
after boiling and centrifugation of the bacterial pellet, the
supernatant was removed to a fresh tube and streptomycin sulfate
(136 ml of a 10% solution/ml supernatant) and acetic acid (glacial,
228 ml/ml supernatant) were added, followed by additional
centrifugation for two minutes. The supernatant was removed
again and then precipitated with ammonium sulfate (saturated
ammonium sulfate at 4uC was used 1:1 v:v with supernatant). The
precipitated protein was collected by centrifugation, and the pellet
was washed once with 1 ml of 50% ammonium sulfate solution
(4uC; 1:1 v:v saturated ammonium sulfate (4uC):water). The
washed pellet was re-suspended in 900 ml of 100 mM ammonium
acetate (to form a cloudy solution) and precipitated by adding an
equal volume of 100% ethanol at room temperature. Ethanol
precipitation was repeated once more, followed by a final re-
suspension in 100 mM ammonium acetate, overnight dialysis to
water at 4uC, freezing in liquid nitrogen, and lyophilization. Expression and purification of a-syn Based on the finding that b-syn inhibits the aggregation of a-
syn, Windisch and colleagues further examined the inhibitory
potential of several deletion mutants of b-syn, focusing on the N-
terminal 1–15 amino acids of the protein. The researchers
generated a peptide library containing different variations of
amino acid composition derived from this specific sequence of b-
syn, with the aim of finding a peptide that could be used for
therapeutic applications, or could serve as a basis for developing
peptidomimetic small molecules [32]. In the present work, we systematically mapped the entire
sequence of b-syn in order to identify other domains that have the
potential to facilitate the molecular recognition between b-syn and
a-syn. Based on these findings, we rationally designed modified
metabolically-stable peptides and analyzed their interaction with
a-syn using chemical and biophysical methods. These peptides
could be internalized into cultured cells and caused a marked
amelioration of the behavioral impairments in a PD fly model. For validation, the a-syn protein was later purified by an
additional method using chromatography and was tested for
aggregation inhibition using the same peptides. Briefly, the
bacterial pellet was re-suspended in 50 mM Tris, 50 mM KCl,
5 mM MgAc, 0.1% NaN3, pH 8.5 and was supplemented with
300 mM PMSF inhibitors. The solution was sonicated and
ultracentrifuged for 30 minutes at 14,000 RPM at 4uC. The
resulting supernatant was boiled for 15 minutes in a water bath. To separate the precipitated proteins from the solution, the sample
was centrifuged for 20 minutes (7,000 RPM at 4uC). The
supernatant was filtered through a 0.45 mm filter and was kept
at 4uC for further purification. Introduction Three major regions are recognized in a-syn: the amino PD, the second most common form of neurodegenerative
diseases after AD, is a devastating neurological disease without
cure, affecting 1–2% of the elderly population. It is characterize by
loss of neuromelanin-containing dopaminergic neurons in the PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 1 b-Syn Fragments Inhibit PD terminal region containing several imperfect repeats of the
KTKEGV sequence, a hydrophobic central domain called the
non-amyloid component (NAC) region, and the carboxy terminal
region characterized by its highly negatively charged amino acids
[25,26]. The a-syn protein is predominantly expressed in neurons
of the central nervous system (CNS), where it is localized at pre-
synaptic termini in close proximity to synaptic vesicles, and can
associate with lipid membranes by forming amphipathic a-helices
[24]. contained libraries of non-relevant peptides. These libraries were
supplied and synthesized by JPT company and were used as a
negative control. After blocking, the membrane was incubated in the presence of
N-terminal histidine-tagged recombinant a-syn, followed by
treatment with HRP-conjugated anti-His monoclonal antibody. In order to identify the strongest binding sites, an additional
membrane containing b-syn decamer peptides with a 9 amino acid
overlap between each sequential peptide along the protein was
used under more stringent conditions; this time we used a smaller
amount of a-syn reacting with the membrane and half the
concentration of the secondary antibody. Once b-syn binding sites
were identified, small b-syn derived peptides were synthesized. a-syn is a member of the family of synuclein proteins, along with
b-synuclein (b-syn) and c-synuclein (c-syn) [27]. a-syn and b-syn
are found primarily in brain tissues located mainly in the pre-
synaptic nerve terminals, while c-syn is found primarily in the
peripheral nervous system and the retina, although it has also been
observed to be highly expressed in some tumor tissues, including
breast, ovarian and bladder tissues [24]. PLoS ONE | www.plosone.org Peptide internalization into SH-SY5Y cells The retro-inverso b-syn 36 peptide internalization into SH-
SY5Y cells was visualized by immunocytochemistry staining. 104
cells were seeded on a cover slip coated with poly-L-lysine (0.1%)
in a 24-well plate and underwent differentiation as described. Differentiated cells were incubated with the peptide in the cells
growth medium for 30 minutes to 4 hours at 37uC. Cells were
washed with PBS and fixed with 4% paraformaldehyde in PBS for
30 minutes at room temperature and then washed twice with PBS
and permeabilized with 0.1% Triton in PBS for 2 minutes. Following two PBS washes, cells were blocked with 10% normal
goat serum in 3% BSA for 30 minutes and incubated with anti a-
syn antibody (Santa Cruz Biotechnology) diluted 1:1000 and
Phalloidin 4 mg/ml (Sigma) for one hour, followed by an Determination of soluble oligomer formation Monomeric a-syn was dissolved to a concentration of 200 mM
in 100 mM Tris buffer (pH 7.4) and was immediately mixed with
or without different b-syn-derived peptides at a 1:1 ratio to a final
concentration of 100 mM, as was described above for the ThT
fluorescence assay. After the samples were agitated at 37uC for
several days, 10 ml of the protein were centrifuged at 13,000 RPM
for 10 minutes, the supernatant was collected and was electro-
phoresed in acrylamide gel using native loading buffer without b-
mercaptoethanol or boiling. The gel was washed three times in
ddH2O and samples were transferred to the nitrocellulose
membrane using a semi-dry blot technique, applying 323 mA
current for 30 minutes. The membrane was blocked for one hour
using 5% milk diluted in TBS (0.3% Tween) while shaking. Anti a-
syn antibody, diluted 1:1,000 (Santa Cruz Biotechnology) in 5%
milk in TBS (0.3% Tween), was added to the membrane for two
hours of incubation followed by several washes with TBS (0.3%
Tween). Rabbit anti-mouse IgG (Fc specific)-HRP-conjugated
antibody diluted 1:5,000 in 5% milk diluted in TBS (0.3% Tween)
was added for one hour at room temperature while shaking. Blots
were developed after thorough TBS (0.3% Tween) washes, using
an Enhanced Chemiluminescence System (ECL) according to the
manufacturer’s manual. In vitro assay of peptide stability Peptides were dissolved to give a 1 mM solution in 50 mM Tris
buffer (pH 7.6). 120 ml of the peptide solution was diluted into a
10% freshly-made homogenate of mouse brain excluding the
cerebellum (in 16Tris buffer and 0.5% Triton X-100). A mixture
containing 20% peptide solution and 80% mouse brain homog-
enate was incubated at 37uC with delicate shaking for two hours. The enzyme reaction was stopped by adding 0.1 M HCl solution,
followed by denaturation of the protein using methanol and
incubation at 20uC for one hour. The precipitated proteins were
centrifuged at 29,0006g for 20 minutes at 4uC and the
supernatant containing the peptide was concentrated under
vacuum and separated using a C18 HPLC column. The area of
the peak (UV absorbance at 280 nm) corresponding to the intact
peptide was compared with an equivalent sample incubated in
50 mM Tris buffer. Transmission electron microscopy Samples (10 ml) from the a-syn ThT fluorescence assay (with
and without inhibitors) were placed on 400-mesh copper grids
covered by carbon-stabilized Formvar film (SPI Supplies, West
Chester, PA). After 1.5 minutes, excess fluid was removed, and the
grids were negatively stained with 10 ml of 2% uranyl acetate
solution for two minutes. Finally, excess fluid was removed and the
samples were viewed by a JEOL 1200EX electron microscope
operating at 80 kV. Cell line SH-SY5Y cells (kindly provided by Prof. Daniel Offen, Tel Aviv
University),
stably
transfected
with
wild
type
a-syn,
were
maintained with DMEM: F12 1:1 containing 5% fetal bovine
serum, 2 mM L-glutamine, 1000 U/ml penicillin-G sodium,
1 mg/ml streptomycin sulfate and 1 mM sodium pyruvate under
selective conditions with 100 mM G-418 at 37uC with 5% CO2. Cells underwent differentiation with 10 mM retinoic acid (Sigma)
in complete growth medium, replaced every two days for a period
of eight days. ThT fluorescence assay of fibril formation ThT fluorescence assay of fibril formation a-syn was dissolved to a concentration of 200 mM in 100 mM
Tris buffer (pH 7.4). In order to obtain the monomeric fraction, the
protein was filtered through a 100 kDa centricon. Since a-syn is not
a globular but rather a natively unfolded protein, only monomers
and some dimers pass through the membrane. The monomeric
protein was immediately mixed with or without different b-syn-
derived peptides at a 1:1 ratio to a final concentration of 100 mM. The samples were incubated at 37uC with agitation of 850 RPM as
described by Tsigelny, et al. [34] and the rate of fibrillogenesis was
monitored using the thioflavin T (ThT) fluorescence assay
(excitation at 450 nm, 2.5 nm slit, and emission at 480 nm, 5 nm
slit), after three days of incubation. ThT was added to a 500-fold
diluted sample and fluorescence was measured using a Jobin Yvon
Horiba Fluoromax 3 fluorometer. To assure that b-syn-derived
peptides don’t aggregate, their fluorescence was measured as control
and subtracted from the test samples. shielded xyz-gradient coil. The transmitter frequency was set on
the HDO signal, which was calibrated according to temperature
(4uC–4.974 ppm;
10uC–4.821 ppm;
15uC–4.773 ppm;
37uC–
4.658 ppm). TOSCY [35,36] and NOESY [36,37] experiments
were acquired for each temperature and sample set. Spectra were processed and analyzed with TOPSPIN software
(Bruker Analytische Messtechnik GmbH). Zero filling in the
indirect dimension, and data apodization with a shifted squared
sine bell window functions in both dimensions, were applied prior
to Fourier transformation for optimal resolution. The baseline was
further corrected in the direct dimension with a quadratic
polynomial function. Resonance assignment was done according to the sequential
assignment methodology developed by Wu¨thrich [John Wiley &
Sons, 38] based on the TOCSY and NOESY spectra measured
under identical experimental conditions. Chemical shift deviations
of the HN-Ha peaks were read from carefully calibrated, highly
resolved, strongly apodized overlaid 2D spectra. Peptide array The molecular mapping of b-syn fragments binding a-syn was
performed using peptide array technology. Decamer peptides
corresponding to overlapping sequences of the full length, 134
amino acids, b-syn protein were synthesized on a cellulose
membrane matrix (JPT Peptide Technologies GmbH, Germany). The first 15 amino acids were synthesized with 9 amino acids
overlap with a shift of one amino acid from one peptide to its
subsequent. The
importance
of
this
region
was
suggested
previously as a binding domain [32]. The rest of the peptide
was synthesized with five amino acids overlap. The peptides were
covalently bound to a Whatman 50 cellulose support via the C-
terminal amino acid residues. Two spots on the membrane The filtrated supernatant was applied on a HiPrep 16/10 QFF
anion exchange column. At 20 mM Tris, pH 8, a-syn has a net
negative charge, and can therefore interact with the positively
charged moieties (NH+) of the column. The protein was eluted
using 30–40% of 20 mM Tris and 1 M NaCl solution (pH 8) and
was subjected to a HiLoad 26/20 superdex 200 size exclusion
column. Anion exchange fractions were loaded with 50 mM Tris/
HCl and 150 mM NaCl buffer (pH 7.5), and the monomer
fraction was analyzed by SDS-PAGE and was collected, dialyzed
overnight to ddH2O at 4uC, frozen in liquid nitrogen, and
lyophilized. November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 2 b-Syn Fragments Inhibit PD b-Syn Fragments Inhibit PD The NMR experiments were performed on a Bruker Avance
600 MHz DMX spectrometer operating at the proton frequency
of 600.13 MHz using a 5 mm selective probe equipped with a self-
shielded xyz-gradient coil. The transmitter frequency was set on
the HDO signal, which was calibrated according to temperature
(4uC–4.974 ppm;
10uC–4.821 ppm;
15uC–4.773 ppm;
37uC–
4.658 ppm). TOSCY [35,36] and NOESY [36,37] experiments
were acquired for each temperature and sample set. The NMR experiments were performed on a Bruker Avance
600 MHz DMX spectrometer operating at the proton frequency
of 600.13 MHz using a 5 mm selective probe equipped with a self- NMR analysis Peptide samples were prepared from peptides stored in
lyophilized
form,
dissolved
in
a
solution
containing
10%
deuterium oxide in 20 mM phosphate buffer and 50 mM NaCl
in ddH2O. Samples of peptides complexed with a-syn were
prepared from stock solutions of a-syn in 20 mM phosphate
solution to which NaCl and deuterium oxide were added to
achieve the above-mentioned concentrations, and the lyophilized
peptide was added at the designated molar ratio. pH was
measured for all samples. Samples were prepared in Shigemi
tubes with a final volume of 260 ml. PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 3 b-Syn Fragments Inhibit PD beginning of their eclosion at 25uC in order to avoid offspring
from the next generation (F2). beginning of their eclosion at 25uC in order to avoid offspring
from the next generation (F2). additional hour of incubation with Cy5-conjugated goat anti-
rabbit IgG (Jackson ImmunoResearch). After being thoroughly
washed with PBS, cells were mounted using ProLong Antifade
(Invitrogene). Images were taken with LSM510 confocal micro-
scope (Zeiss). Fly crossing Male
flies
carrying
the
driver
elavc155-Gal4
on
their
X
chromosome were crossed to females carrying the UAS-regulated
A53T a-syn transgene located on their X chromosome (kindly
provided by Prof. Mel Feany, Harvard Medical School). This
resulted in first generation (F1) female offspring expressing A53T a-
syn in their nervous system which served as our PD fly model. In
parallel, male flies carrying the driver elavc155-Gal4 on their X
chromosome were crossed to wild type (Oregon-R) females and the
resultant F1, carrying only the elavc155-Gal4 driver served as control Fly keeping Flies were reared on standard cornmeal-molasses medium and
were kept at 25uC. As Drosophila females can store sperm cells in
their bodies, crosses were conducted using virgin females collected
no longer than eight hours after eclosion at 25uC or 18 hours after
eclosion at 18uC. The crosses were performed at 29uC. Adult
offspring (F1) from the crosses were collected up to 9 days after the Figure 1. Mapping of the interaction areas between b-syn and a-syn. (A) Schematic illustration of interactions between b-syn and a-syn; a-
syn monomers (red) aggregate to form fibrils and oligomers. (B) b-syn (grey-blue) is known to inhibit the aggregation of a-syn. (C). Potential b-syn-
derived peptides (blue) as inhibitors of a-syn aggregation. (D) Molecular mapping of b-syn sequences that bind a-syn using peptide-array analysis. (E)
Screen for a-syn binding sequences within the b-syn protein. Decamer peptides corresponding to consecutive 9 residue overlapping sequences
whithin the first 15 amino acids of b-syn and 5 residue overlapping sequences of the rest of the protein were synthesized on a cellulose membrane
and incubated with His-labeled a-syn. (F) An additional screen was performed using more stringent conditions and a 9 residue overlapping along the
entire protein. doi:10.1371/journal.pone.0013863.g001 Figure 1. Mapping of the interaction areas between b-syn and a-syn. (A) Schematic illustration of interactions between b-syn and a-syn; a-
syn monomers (red) aggregate to form fibrils and oligomers. (B) b-syn (grey-blue) is known to inhibit the aggregation of a-syn. (C). Potential b-syn-
derived peptides (blue) as inhibitors of a-syn aggregation. (D) Molecular mapping of b-syn sequences that bind a-syn using peptide-array analysis. (E)
Screen for a-syn binding sequences within the b-syn protein. Decamer peptides corresponding to consecutive 9 residue overlapping sequences
whithin the first 15 amino acids of b-syn and 5 residue overlapping sequences of the rest of the protein were synthesized on a cellulose membrane
and incubated with His-labeled a-syn. (F) An additional screen was performed using more stringent conditions and a 9 residue overlapping along the
entire protein. doi:10.1371/journal.pone.0013863.g001 Figure 1. Mapping of the interaction areas between b-syn and a-syn. (A) Schematic illustration of interactions between b-syn and a-syn; a-
syn monomers (red) aggregate to form fibrils and oligomers. (B) b-syn (grey-blue) is known to inhibit the aggregation of a-syn. (C). Potential b-syn-
derived peptides (blue) as inhibitors of a-syn aggregation. (D) Molecular mapping of b-syn sequences that bind a-syn using peptide-array analysis. Immuno-staining of adult fly brains Fourteen-day old adult flies were dissected and their brains were
removed. After incubation in Paraformaldehyde the whole brains
were transferred to formic acid and blocked with 0.5% Triton in
5% BSA. Following an overnight incubation with anti a-syn
antibody (Santa Cruz Biotechnology) diluted 1:200, the brains Statistical analysis P-values
were calculated
for
comparison of female flies
expressing A53T a-syn reared on regular medium with female
flies expressing A53T a-syn on medium supplemented with the
tested compound and with control flies using one tail ANOVA test
with a value of 0.05. P,0.05 was considered significant. The peptides containing amino acids 36–39 of b-syn were of
special interest due to the presence of tyrosine. We and others have
previously identified an important role of aromatic amino acids in
the
acceleration
of
amyloid
fibrils
formation
[5,6,9,39,40]. Therefore, the presence of tyrosine within these peptides might
be useful in the inhibition of the self-assembly of a-syn. Locomotive (climbing) assay Vials of each of the following four F1 classes: 1) Females
expressing A53T a-syn reared on regular medium; 2) Females
expressing A53T a-syn reared on medium supplemented with the
tested compound; 3) Control females carrying only the elavc155-
Gal4 driver, reared on regular medium; 4) Control females
carrying
only
the
elavc155-Gal4
driver,
reared
on
medium
supplemented with the tested compound, each containing 10 flies
were tapped gently on the table and were let to stand for 20
seconds. The percentage of flies which climbed along the test tube
was monitored over time. Each class had four independent vial
repeats. Special fly feeding The retro-inverso b-syn 36 peptide was added to standard
cornmeal-molasses medium, upon cooking, at a concentration of
0.75 mg/ml. The peptide was mixed thoroughly into the medium
when it was at 40uC and the mixture was aliquoted into fly rearing
vials. The vials were kept at 4uC until use. Crosses were performed
either on regular Drosophila medium (control) or on medium
supplemented with the peptide. Animals fed on the appropriate
medium from the beginning of the larval stage. After eclosion,
adult offspring were transferred into a fresh vial containing regular
Drosophila medium on top of which a solution of 0.75 mg/ml
peptide was dripped every other day. Identification of the recognition domains between b-syn
and a-syn The full length b-syn has been shown to inhibit the aggregation
of a-syn in a dose-dependent manner. Therefore the recognition
modules within the b-syn protein were systematically mapped
(illustrated in Figure 1A–D). An overlapping peptide array technique was used for fine
molecular mapping using 9 amino acids overlapping decamer
peptides for the first 15 amino acids of the protein and 5 amino
acids overlapping decamer peptides for the rest of the protein
(Figure 1E). For validation, an additional set of decamer peptides
overlapping in 9 amino acids were synthesized on a membrane
and their binding detection to a-syn was performed under more
stringent conditions. A few binding regions were detected in the
first screen but only some of them appeared under more stringent
binding conditions (Figure 1F). These results lead to the synthesis
of seven candidate decamers for further analysis. These peptides
were termed b-syn 6, 14, 36, 37, 38, 39, 77, and 78 according to
the location of their first amino acid (Table 1). A septamer,
containing amino acids 9–15 termed b-syn 6short (Table 1), that
was shown to display significant inhibition of a-syn aggregation
[32] was used as positive control. Fly keeping (E)
Screen for a-syn binding sequences within the b-syn protein. Decamer peptides corresponding to consecutive 9 residue overlapping sequences
whithin the first 15 amino acids of b-syn and 5 residue overlapping sequences of the rest of the protein were synthesized on a cellulose membrane
and incubated with His-labeled a-syn. (F) An additional screen was performed using more stringent conditions and a 9 residue overlapping along the
entire protein. doi:10.1371/journal.pone.0013863.g001 Figure 1. Mapping of the interaction areas between b-syn and a-syn. (A) Schematic illustration of interactions between b-syn and a-syn; a-
syn monomers (red) aggregate to form fibrils and oligomers. (B) b-syn (grey-blue) is known to inhibit the aggregation of a-syn. (C). Potential b-syn-
derived peptides (blue) as inhibitors of a-syn aggregation. (D) Molecular mapping of b-syn sequences that bind a-syn using peptide-array analysis. (E)
Screen for a-syn binding sequences within the b-syn protein. Decamer peptides corresponding to consecutive 9 residue overlapping sequences
whithin the first 15 amino acids of b-syn and 5 residue overlapping sequences of the rest of the protein were synthesized on a cellulose membrane
and incubated with His-labeled a-syn. (F) An additional screen was performed using more stringent conditions and a 9 residue overlapping along the
entire protein. doi:10 1371/journal pone 0013863 g001 p
doi:10.1371/journal.pone.0013863.g001 November 2010 | Volume 5 | Issue 11 | e13863 November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 4 b-Syn Fragments Inhibit PD were stained with Cy5-conjugated goat anti-rabbit IgG (Jackson
ImmunoResearch). After being thoroughly washed with PBS, cells
were mounted using ProLong Antifade (Invitrogene). Images were
taken with LSM510 confocal microscope (Zeiss). November 2010 | Volume 5 | Issue 11 | e13863 Inhibition of formation of a-syn amyloid fibrils by b-syn-
derived peptides Thioflavin-T (ThT) binding assay was used to examine and
quantify the inhibitory effect of the b-syn peptides on a-syn fibril Table 1. b-syn derived peptides. Peptide name
Sequence
Position within b-syn protein sequence
b-syn 6short (control)
SMAKEGV
9–15
b-syn 14
GVVAAAEKTK
14–23
b-syn 36
GVLYVGSKTR
36–45
b-syn 37
VLYVGSKTRE
37–46
b-syn 38
LYVGSKTREG
38–47
b-syn 39
YVGSKTREGV
39–48
b-syn 77
AAATGLVKRE
77–86
b-syn 78
AATGLVKREE
78–87
b-syn 36 modifications (all aa are D enantiomers)
b-syn 36D
GVLYVGSKTR
36–45
Retro inverso b-syn 36
RTKSGVYLVG
36–45
Acetylated and amidated retro inverso b-syn 36
Acetyl-RTKSGVYLVG-amide
36–45
b-syn 36D short
YVGSKTR
39–45
Y39W b-syn 36D
GVLWVGSKTR
36–45
doi:10.1371/journal.pone.0013863.t001
PLoS ONE | www plosone org
5
November 2010 | Volume 5 | Issue 11 | e13863 Table 1. b-syn derived peptides. November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 5 b-Syn Fragments Inhibit PD b-Syn Fragments Inhibit formation. This method provides quantitative information on
tion process was monitored for three days. Figure 2A demonstrates
Figure 2. In vitro analysis of a-syn aggregation. (A) In vitro inhibition of a-syn fibrillar assembly. b-syn peptides were screened for inhibiting a-
syn aggregation using ThT (molar ratio 20:1 respectively). (B) Kinetic analysis of the aggregation of a-syn in the presence of b-syn peptides 36 and 39. Control a-syn (¤), peptide b-syn 39 with a-syn (m), peptide b-syn 36 with a-syn (&). (C) Dose dependent inhibition effect of peptide b-syn 36 on a-
syn aggregation. (D-K) TEM images of a-syn fibrils with several peptides; a-syn alone, a-syn with b-syn 6short, 36, 37, 38, 39, 77, 78. 6 short, 37 and 38;
bar = 500 nM. Rest of the peptides bar = 1 mM. *, P,0.08. doi:10.1371/journal.pone.0013863.g002
b Syn Fragments Inhibit PD Figure 2. In vitro analysis of a-syn aggregation. (A) In vitro inhibition of a-syn fibrillar assembly. b-syn peptides were screened for inhibiting a-
syn aggregation using ThT (molar ratio 20:1 respectively). (B) Kinetic analysis of the aggregation of a-syn in the presence of b-syn peptides 36 and 39. Control a-syn (¤), peptide b-syn 39 with a-syn (m), peptide b-syn 36 with a-syn (&). (C) Dose dependent inhibition effect of peptide b-syn 36 on a-
syn aggregation (D-K) TEM images of a-syn fibrils with several peptides; a-syn alone a-syn with b-syn 6short 36 37 38 39 77 78 6 short 37 and 38; Figure 2. In vitro analysis of a-syn aggregation. Inhibition of formation of a-syn amyloid fibrils by b-syn-
derived peptides (A) In vitro inhibition of a-syn fibrillar assembly. b-syn peptides were screened for inhibiting a-
syn aggregation using ThT (molar ratio 20:1 respectively). (B) Kinetic analysis of the aggregation of a-syn in the presence of b-syn peptides 36 and 39. Control a-syn (¤), peptide b-syn 39 with a-syn (m), peptide b-syn 36 with a-syn (&). (C) Dose dependent inhibition effect of peptide b-syn 36 on a-
syn aggregation. (D-K) TEM images of a-syn fibrils with several peptides; a-syn alone, a-syn with b-syn 6short, 36, 37, 38, 39, 77, 78. 6 short, 37 and 38;
bar = 500 nM. Rest of the peptides bar = 1 mM. *, P,0.08. doi:10.1371/journal.pone.0013863.g002 tion process was monitored for three days. Figure 2A demonstrates
that the formation of amyloid fibrils was significantly reduced in the
presence of several of the peptide inhibitors. A kinetic analysis was
performed for the two potential inhibitors b-syn 36 and b-syn 39 formation. This method provides quantitative information on
amyloid fibril growth. a-syn was incubated for several days at
37uC with vigorous shaking to allow the formation of amyloid fibrils
with or without the different b-syn-derived peptides. The fibrilliza- November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 6 b-Syn Fragments Inhibit PD Figure 3. (A) Aggregation of a-syn over time. Soluble oligomers were formed while shaking at 37uC. Detection was carried using
western blot analysis. (B) Inhibition of oligomer assembly using b-syn peptides following 24 hours of incubation. doi:10.1371/journal.pone.0013863.g003 Figure 3. (A) Aggregation of a-syn over time. Soluble oligomers were formed while shaking at 37uC. Detection was carried using
western blot analysis. (B) Inhibition of oligomer assembly using b-syn peptides following 24 hours of incubation. doi:10.1371/journal.pone.0013863.g003 (Figure 2B) as these two peptides represent the first and last peptides
screened within the aromatic region (Table 1). b-syn 36 showed
better inhibition than b-syn 39 and dose-dependent inhibition tests
revealed that b-syn 36 is still efficient at an excess of 10:1 (b-syn 36:
a-syn) molar ratio but is less efficient at of 5:1 ratio (Figure 2C). degradation by tissue and serum proteases and peptidases. b-syn
36 is a decamer peptide comprising natural L-amino acids hence is
susceptible to proteolytic degradation. Inhibition of formation of a-syn amyloid fibrils by b-syn-
derived peptides Three modified, more
stable versions of b-syn 36 peptide were designed: the first
composed of D instead of L-amino acids, termed b-syn 36D; the
second was an analogue with retro-inversion of the amino acids
sequence, composed of D-amino acids assembled in the reverse
order from that of the parent L-sequence, termed retro-inverso b-
syn 36. To negate the terminal negative and positive charges of the
retro-inverso peptide, a third modified N-terminal acetylated and
C-terminal amidated peptide was synthesized, termed acetylated
and amidated retro-inverso b-syn 36. The modifications of these
peptides were suspected to alter the efficacy of the lead peptide
without harming its biological activity. Indeed, when tested for
their ability to inhibit a-syn fibril formation, using the ThT
binding assay with a molar ratio of 1:20 or 1:10 in favor of the
peptides, all three modified versions demonstrated a similar
biological effect as b-syn 36 (Figure 4). The t-test showed P,0.08. TEM analysis of a-syn aggregation Transmission electron microscopy (TEM) analysis was per-
formed on samples of a-syn incubated with and without the b-syn
inhibitor peptides. The samples were taken from the ThT
experiment (presented in Figure 2A). The t-test showed P,0.08. While the fibrils formed by a-syn alone were large, broad and
had a ribbon-like shape (Figure 2D), only short or no fibrils were
detected in the presence of the b-syn peptides. b-syn 36 almost
completely inhibited fibril formation (Figure 2F). These results are
highly correlated with the results of the ThT assay. Transmission electron microscopy (TEM) analysis was per-
formed on samples of a-syn incubated with and without the b-syn
inhibitor peptides. The samples were taken from the ThT
experiment (presented in Figure 2A). The t-test showed P,0.08. While the fibrils formed by a-syn alone were large, broad and
had a ribbon-like shape (Figure 2D), only short or no fibrils were
detected in the presence of the b-syn peptides. b-syn 36 almost
completely inhibited fibril formation (Figure 2F). These results are
highly correlated with the results of the ThT assay. Screening for inhibition of a-syn oligomer formation To examine the ability of the peptides to inhibit the early stage
of a-syn aggregation, soluble fractions of a-syn were collected after
incubation with and without the b-syn peptides. The reaction
mixtures were separated by SDS-PAGE followed by western blot
analysis using a specific anti a-syn antibody (Santa-Cruz). A
preliminary test of a-syn oligomer formation over time in the
absence of b-syn-derived peptides revealed that high oligomers
could be detected after ,20 hours of incubation (Figure 3A). Therefore samples incubated with b-syn-derived peptides were
collected after this period of time. Given the current debate
regarding the identity of the toxic species in the process of a-syn
aggregation, it was interesting to note that some b-syn-derived
peptides affected the formation of a-syn fibrils while others
affected the formation of oligomers (Figure 3B). The b-syn 6short
peptide (our positive control peptide) and the b-syn 37 peptide
inhibited formation of mature fibrils but not oligomers, while the
b-syn 78 peptide inhibited oligomers but not fibril formation. The
b-syn 36 peptide inhibited almost completely the formation of
both aggregation types (Figure 2F, 3B). Figure 4. Inhibition of a-syn fibrillar assemblies using ThT
fluorescence assay. Three modified peptides of b-syn 36 were
screened for their inhibition on a-syn aggregation: a-syn alone (black),
b-syn 36 with a-syn (grey), and the modified peptides with a-syn: b-syn
36D (white), retro-inverso b-syn 36 (cross-hatch), acetylated and
amidated retro inverso b-syn 36 (dots). The molar ratios of a-
syn:peptide were 1:20 and 1:10. *, P,0.08. doi:10.1371/journal.pone.0013863.g004 Figure 4. Inhibition of a-syn fibrillar assemblies using ThT
fluorescence assay. Three modified peptides of b-syn 36 were
screened for their inhibition on a-syn aggregation: a-syn alone (black),
b-syn 36 with a-syn (grey), and the modified peptides with a-syn: b-syn
36D (white), retro-inverso b-syn 36 (cross-hatch), acetylated and
amidated retro inverso b-syn 36 (dots). The molar ratios of a-
syn:peptide were 1:20 and 1:10. *, P,0.08. doi:10.1371/journal.pone.0013863.g004 b-Syn Fragments Inhibit PD Figure 5. Serum stability of b-syn 36 peptides. Brain extract of
wild type ICR white mouse was incubated with the peptides b-syn 36
and retro-inverso b-syn 36 for two hours and loaded on a C18 column
for serum stability analysis. (A) Spectra of the b-syn 36 peptide (blue)
and b-syn 36 peptide with brain extract (red). (B) Spectra of the retro-
inverso b-syn 36 peptide (blue) and retro-inverso b-syn 36 peptide with
brain extract (red). (C) Quantification of the area under the peak. The
area of the peptide in time ‘0’ was determined as 100%. The left
columns represent the peptide (black) and the right columns represent
the peptide with the brain extract (grey). doi:10.1371/journal.pone.0013863.g005 spectrum that is distinct from that of a-syn. The residues that
participate in the intermolecular interactions of the retro-inverso
b-syn 36 with a-syn were determined by following changes in their
amide and Ha chemical shifts upon binding. The spectrum of the
peptide in the absence and in the presence of a-syn was
determined under several sets of conditions: Molar ratios of 1:1
and 1:5 peptide to a-syn; peptide concentrations of 160 and
400 mM; temperatures of 4uC, 10uC, 20uC, 25uC, and 37uC; and
using Y39W b-syn 36D analog. All residues of the retro-inverso b-syn 36 peptide were identified
and assigned (Figure S2). R45 and T44 were not detected in the
amide region, but were identified in the aliphatic region. In all
samples some of the N-terminal region amide signals were lost
between G41 and R45; however, all samples showed the general
trend found for the sample of retro-inverso b-syn 36 with a-syn at
a 1:1 molar ratio, 160 mM, 4uC, at pH 6.7 (Figure 6A and B-E
and Table 1). The amide resonances showed stronger HN and Ha
deviations in residues K43, V40, L38 and G36. The participation of
L38 and G36 in binding was ascertained using an additional
truncated peptide, b-syn 36D short, which lacks the first three
amino acids, G36, V37 and L38, and showed no inhibition of a-syn
aggregation in ThT assay (Figure 6F). These results suggest that
K43, V40, L38 and G36 participate in the binding interaction and
that the large chemical shift deviation of L38 and G36 is not only
due to their position at the peptide terminus. The effect of the retro-inverso b-syn 36 peptide in vivo The effect of the retro-inverso b-syn 36 peptide in vivo
The effect of the retro-inverso b-syn 36 peptide on a-syn
aggregation in the living organism was assessed using a Drosophila
model of PD. These transgenic flies over- express the mutated
A53P a-syn in their nervous system, via the Gal4-UAS system. A
common behavioral phenotype of these flies is age-dependent
defective locomotion: While normal flies tend to climb up the side
of the rearing tube, A53T a-syn-expressing flies remain at the
bottom [42]. Crossing male flies carrying the pan-neuronal elav-
Gal4 driver (on their X chromosome) to females carrying the
UAS-regulated A53T a-syn transgene resulted in female offspring
expressing A53T a-syn in their nervous system. This cross was
performed either on regular Drosophila medium or on medium
supplemented with 0.75 mg/mL of the retro-inverso b-syn 36
peptide. The climbing ability of the flies was monitored for 27 days
(Figure 8). Locomotion of flies expressing A53T a-syn in their To evaluate the binding of the peptides to a-syn, increasing
amounts of a-syn monomers were titrated into a solution of the
modified b-syn 36 peptide and anisotropy was determined (Figure
S1). The affinity constant was determined to be ,1 mM. Modified versions of b-syn 36 peptide inhibit the
aggregation of a-syn and have increased serum sta gg
g
y
y
For using the b-syn 36 peptide as potential drug it is desirable to
increase its half-life in the patient’s serum, by reducing its PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 7 b-Syn Fragments Inhibit PD PLoS ONE | www.plosone.org Internalization of b-syn-derived peptides into
mammalian cells To evaluate the efficacy of the b-syn-derived peptide in
mammalian cell culture, which serves as a model for PD, SH-
SY5Y5 cells over-expressing wild type a-syn were used. They were
first subjected to retinoic acid treatment to induce cell differen-
tiation [41]. After eight days of treatment, cells were cultured into
fresh growing media for 0.5–4 hours followed by incubation with
FITC-conjugated retro-inverso b-syn 36 peptide (green fluores-
cence). After incubation, cells were washed, fixed, and permea-
bilized. Overexpression of wild type a-syn in these cells was
verified using cy5-conjugated goat anti-rabbit antibody (purple
fluorescence) and the cell membrane was marked using Phalloidin
reagent (red fluorescence) (Figure 7). The two concentrations of b-
syn-derived peptide, 50 mM and 250 mM, showed similar results. After 30 minutes of incubation the peptide did not enter into the
cells. No attachment to the cells was noted. After two hours of
incubation the peptide was detected mostly on the cell surface. Small amount was detected inside the cells at the concentration of
250 mM. After four hours of incubation, the peptide was clearly
visible inside the cells (Figure 7). Figure 5. Serum stability of b-syn 36 peptides. Brain extract of
wild type ICR white mouse was incubated with the peptides b-syn 36
and retro-inverso b-syn 36 for two hours and loaded on a C18 column
for serum stability analysis. (A) Spectra of the b-syn 36 peptide (blue)
and b-syn 36 peptide with brain extract (red). (B) Spectra of the retro-
inverso b-syn 36 peptide (blue) and retro-inverso b-syn 36 peptide with
brain extract (red). (C) Quantification of the area under the peak. The
area of the peptide in time ‘0’ was determined as 100%. The left
columns represent the peptide (black) and the right columns represent
the peptide with the brain extract (grey). doi:10.1371/journal.pone.0013863.g005 Next, the serum stability of the unmodified b-syn 36 peptide and
the modified retro-inverso peptide was compared by incubating
them for two hours in mouse brain homogenate to model serum
degradation. The results showed that the modified peptide is more
stable than the unmodified b-syn 36 (Figure 5). NMR analysis of the interaction of b-syn peptides with
a-syn Studying interactions between a-syn and b-syn by NMR is
severely limited by signal overlap due to the similarity in
their
sequences. This
was
overcome
by
using
the
all-D
chirality retro-inverso b-syn 36 peptide (retro-inverso b-syn 36:
R45T44K43S42G41V40Y39L38V37G36),
which
gives
a
resolved PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 8 b-Syn Fragments Inhibit PD Figure 6. NMR analysis. (A) Chemical shift deviations of HN (black) and Ha (grey) backbone atoms of the b-syn 36 retro-inverso peptide upon
binding a-syn. (B) Ha-HN region of TOCSY spectra overlay of the b-syn 36 retro-inverso peptide (red) and a-syn (blue) upon binding (green), (C–E)
Expansions of all the retro-inverso peaks that showed deviations upon binding. (F) ThT analysis of fibril formation by a-syn with and without b-syn
36D and its truncated version which lacks amino acids G36, V37 and L38. d i 10 1371/j
l
0013863 006 Figure 6. NMR analysis. (A) Chemical shift deviations of HN (black) and Ha (grey) backbone atoms of the b-syn 36 retro-inverso peptide upon
binding a-syn. (B) Ha-HN region of TOCSY spectra overlay of the b-syn 36 retro-inverso peptide (red) and a-syn (blue) upon binding (green), (C–E)
Expansions of all the retro-inverso peaks that showed deviations upon binding. (F) ThT analysis of fibril formation by a-syn with and without b-syn
36D and its truncated version which lacks amino acids G36, V37 and L38. doi:10.1371/journal.pone.0013863.g006 (Figure 9A), the brains of the treated flies exhibited a greatly
reduced a-syn staining (Figure 9B). Quantification of 8 treated
brains and 7 untreated brains was performed and t-test analysis
showed **, P,0.0001 (Figure 9C). nervous system deteriorated significantly from day 13 onwards. The untreated flies became almost immobile by day 27, while the
control classes were very active at this time. In contrast, the treated
flies displayed dramatic improvement, behaving almost identical to
the control classes, presenting an increase of nearly 86% in their
climbing ability in comparison with the untreated group. The
peptide had no significant effect on locomotion of the control flies. These results indicate significant phenotypic recovery of the A53T
a-syn flies by the retro-inverso b-syn 36 peptide. One tail ANOVA
statistics showed P,0.05. nervous system deteriorated significantly from day 13 onwards. The untreated flies became almost immobile by day 27, while the
control classes were very active at this time. NMR analysis of the interaction of b-syn peptides with
a-syn In contrast, the treated
flies displayed dramatic improvement, behaving almost identical to
the control classes, presenting an increase of nearly 86% in their
climbing ability in comparison with the untreated group. The
peptide had no significant effect on locomotion of the control flies. These results indicate significant phenotypic recovery of the A53T
a-syn flies by the retro-inverso b-syn 36 peptide. One tail ANOVA
statistics showed P,0.05. PLoS ONE | www.plosone.org Discussion The pathological hallmark of PD is the occurrence of insoluble
intracellular inclusions termed LB and LN within the surviving
dopaminergic neurons of the substantia nigra and several other brain
regions, with a-syn as the major component of these inclusions
[21]. The first indication that a-syn could be involved in
neurodegenerative diseases came from work on the NAC region
isolated from brain amyloid preparations of AD patients [43]. Two
different point mutations, resulting in amino acid substitutions To evaluate the effect of the retro-inverso b-syn 36 peptide on
a-syn accumulation in the brains of these flies, adult flies were fed
or unfed with the peptide, and their brains were immunostained
with anti a-syn antibody. While a massive accumulation of a-syn
aggregates was detected in the brains of the untreated flies PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 November 2010 | Volume 5 | Issue 11 | e13863 9 b-Syn Fragments Inhibit PD b-Syn Fragments Inhibit PD Figure 7. Internalization of retro-inverso b-syn 36 into the cells. Differentiated SH-SY5Y cells over expressing wild type a-syn were incubated
with 50 mM and 250 mM of FITC-conjugated retro-inverso b-syn 36 peptide for periods of half an hour, two hours and four hours. After fixation, the
presence of the FITC-conjugated peptide (green) was monitored inside the cells. Cellular a-syn was detected using cy5-conjugated antibody (purple)
and the cell membrane was marked with Phalloidin (red). (A) After 30 minutes at 37uC there was no peptide staining (green). (B–C) After two hours of
incubation, no or little amount of peptide was detected inside the cells (green). (D–E) After 4 hours of incubation, the peptide was clearly detected
inside the cells. Peptide localization was visualized using an LSM-510 Zeiss confocal microscope. doi:10.1371/journal.pone.0013863.g007 Figure 7. Internalization of retro-inverso b-syn 36 into the cells. Differentiated SH-SY5Y cells over expressing wild type a-syn were incubated
with 50 mM and 250 mM of FITC-conjugated retro-inverso b-syn 36 peptide for periods of half an hour, two hours and four hours. After fixation, the
presence of the FITC-conjugated peptide (green) was monitored inside the cells. Cellular a-syn was detected using cy5-conjugated antibody (purple)
and the cell membrane was marked with Phalloidin (red). (A) After 30 minutes at 37uC there was no peptide staining (green). (B–C) After two hours of
incubation, no or little amount of peptide was detected inside the cells (green). PLoS ONE | www.plosone.org Discussion (D–E) After 4 hours of incubation, the peptide was clearly detected
inside the cells. Peptide localization was visualized using an LSM-510 Zeiss confocal microscope. doi:10.1371/journal.pone.0013863.g007 A53T
and
A30P,
have
firmly
established
the
pathogenic
importance of a-syn when they were indentified as a cause of
rare, inherited forms of PD [19]. Although the etiology of
idiopathic PD is unknown, it is likely to be multifactorial. A
presumptive feature leading to PD is the aggregation of a-syn,
which can be triggered by a range of factors, both environmental
and endogenous [31]. Uversky et al. suggested that the cellular
levels of b-syn and c-syn could constitute such an endogenous
factor [31]. identify all the domains that have the potential to mediate
molecular recognition events between b-syn and a-syn. Using the
peptide array technique several interaction modules were
identified and some of them were verified by a second round
under more stringent interaction conditions. Decamer peptides
corresponding to the interaction regions were synthesized and
their ability to inhibit the aggregation of a-syn was tested using
ThT and TEM assays. Both methods showed consistent results,
indicating an inhibitory effect of the peptides on fibril formation
by a-syn. Due to the importance of a-syn aggregation in the pathology of
PD, a useful strategy in drug development is to design short
peptides, or small molecules, that can block, slow down, or reverse
a-syn aggregation, particularly at its early stages [27]. The fact
that a-syn is a natively unfolded protein, makes the rational design
of compounds that can stabilize the native, nontoxic conformation
of a-syn a challenging task [27]. y
y
Recent studies reveal that early ‘‘soluble oligomers’’ rather than
mature amyloid fibrils are likely the pathogenic species that drive
neurodegeneration and neuronal cell death [44,45]. It is believed
that the conversion of a-syn into these oligomers, and then into
insoluble fibrils, is initiated by conformational changes from
random coil to b-sheet structure [4,27,46,47]. This suggests that a
candidate anti-amyloid compound should block the very early
stages of amyloid oligomerization. Consequently, we tested the
ability of the interacting b-syn-derived peptides to inhibit the
formation of the soluble fraction of a-syn, and found several A recently proposed approach is to use the properties of the b-
syn protein as a natural inhibitor of the aggregation of a-syn [32]. November 2010 | Volume 5 | Issue 11 | e13863 Discussion In the present work we applied this approach and for the first
time systematically mapped the entire sequence of b-syn to November 2010 | Volume 5 | Issue 11 | e13863 November 2010 | Volume 5 | Issue 11 | e13863 10 b-Syn Fragments Inhibit PD b-Syn Fragments Inhibit PD Figure 8. Effect of b-syn-derived peptide on locomotive
behavior of PD model flies. Four classes of flies, each containing
five tubes of ten flies were analyzed using the climbing assay: Flies
expressing a-syn A53T grown on a regular medium (black); flies
expressing a-syn A53T grown on medium containing 0.75 mg/mL b-syn
retro inverso peptide (grey); Control flies, not expressing a-syn A53T,
grown on a regular medium (cross-hatch); Control flies, not expressing
a-syn A53T, grown on medium containing 0.75 mg/mL b-syn retro
inverso peptide (white). Results show the percent of flies which climbed
along the test tube after 20 seconds. *, P,0.05. doi:10.1371/journal.pone.0013863.g008 peptides that were able to do so, while the positive control peptide
appeared to inhibit only the formation of the insoluble fibrils. While several peptides such as b-syn 6short and b-syn 37 inhibit
fibril formation but not oligomer formation, b-syn 78 inhibits
oligomer formation but not fibril formation. These results support
the latest theory of a two-pathway mechanism of amyloids
aggregation. A similar hypothesis was demonstrated on b-amyloid
in AD [48]. Our best inhibitor candidate was the b-syn 36 peptide which
inhibited both the soluble and insoluble fibrillar aggregates of a-
syn. In addition to its inhibitory properties, this peptide showed no
toxicity towards cultured cells (PC12) when added, at concentra-
tions up to 1 mM, to the their growing medium and incubated for
twenty four hours (data not shown). The sequence of the b-syn 36 peptide has two interesting
features. First, it contains part of the repetitive domain of
KTKEGV (with a substitution of K for R), which has been
suggested to represent an evolutionary balance between the
functional conformer of a-syn (a-helix and/or random coil) and its
pathogenic b-sheet conformation [49]. Second, the b-syn 36
peptide contains the aromatic amino acid tyrosine, which
according to the ‘‘aromatic stacking’’ hypothesis previously
suggested by us [6], may contribute to the binding energetics as
well as induce order and directionality to the self-assembly of
amyloid structures. These two characteristics may play a role in
the inhibitory potential of this peptide. Figure 8. Discussion Effect of b-syn-derived peptide on locomotive
behavior of PD model flies. Four classes of flies, each containing
five tubes of ten flies were analyzed using the climbing assay: Flies
expressing a-syn A53T grown on a regular medium (black); flies
expressing a-syn A53T grown on medium containing 0.75 mg/mL b-syn
retro inverso peptide (grey); Control flies, not expressing a-syn A53T,
grown on a regular medium (cross-hatch); Control flies, not expressing
a-syn A53T, grown on medium containing 0.75 mg/mL b-syn retro
inverso peptide (white). Results show the percent of flies which climbed
along the test tube after 20 seconds. *, P,0.05. doi:10.1371/journal.pone.0013863.g008 Figure 9. Immunostaining of PD model flies brains. Brains of fourteen-day flies were extracted and immunostained with anti a-syn
antibody followed by a Cy5-conjugated antibody. a-syn is detected in green. (A) Staining of untreated flies expressing a-syn A53T grown on a
regular medium. (B) Staining of flies expressing a-syn A53T treated with b-syn retro inverso peptide. (C) Quantification of treated and untreated
brains. The t-test analysis showed **, P,0.0001. doi:10.1371/journal.pone.0013863.g009 Figure 9. Immunostaining of PD model flies brains. Brains of fourteen-day flies were extracted and immunostained with anti a-syn
antibody followed by a Cy5-conjugated antibody. a-syn is detected in green. (A) Staining of untreated flies expressing a-syn A53T grown on a
regular medium. (B) Staining of flies expressing a-syn A53T treated with b-syn retro inverso peptide. (C) Quantification of treated and untreated
brains. The t-test analysis showed **, P,0.0001. doi:10.1371/journal.pone.0013863.g009 November 2010 | Volume 5 | Issue 11 | e13863 PLoS ONE | www.plosone.org 11 b-Syn Fragments Inhibit PD Two problems commonly associated with peptides in therapy
are their high sensitivity to proteolytic degradation and lack of
permeation into the cells [50]. We attempted to overcome the first
obstacle by using modified peptides containing D amino acids and
by reversing the peptide sequence. Both modifications retained the
inhibitory effect of the peptides. In addition to its excellent
inhibition ability, the retro-inverso b-syn 36 modified peptide
displayed increased stability towards proteolytic degradation. Concerning the cell permeability, we showed that the stable
retro-inverso
b-syn
36
peptide
can penetrate
differentiated
SHSY5Y cells without adding any internalization motif. unbound structure, which was not significantly changed upon
interacting
with
a-syn. The
two
neighboring
hydrophobic
residues, L38 and V40, may reside within a hydrophobic binding
interface with a-syn. Supporting Information Figure S1
The affinity of the modified b-syn 36 containing
tryptophan instead of tyrosine towardsa-syn monomers was
examined using fluorescent anisotropy. Kd = 1 mM. Found
at:
doi:10.1371/journal.pone.0013863.s001
(0.02
MB
TIF) Figure S1
The affinity of the modified b-syn 36 containing
tryptophan instead of tyrosine towardsa-syn monomers was
examined using fluorescent anisotropy. Kd = 1 mM. Found
at:
doi:10.1371/journal.pone.0013863.s001
(0.02
MB
TIF) Figure S1
The affinity of the modified b-syn 36 containing
tryptophan instead of tyrosine towardsa-syn monomers was
examined using fluorescent anisotropy. Kd = 1 mM. Figure S1
The affinity of the modified b-syn 36 containing
tryptophan instead of tyrosine towardsa-syn monomers was
examined using fluorescent anisotropy. Kd = 1 mM. Found
at:
doi:10.1371/journal.pone.0013863.s001
(0.02
MB
TIF) Figure S2
NMR assignment spectra of b-syn 36 retro-inverso
peptide. Overlay of HN-Ha interaction regions of TOCSY (red)
and NOESY (green) spectra of taken under identical conditions
according to which assignment was performed. Found
at:
doi:10.1371/journal.pone.0013863.s002
(8.35
MB
TIF) Acknowledgments We thank Yaacov Delarea for help with electron microscopy experiments;
Prof. Mel Feany from Harvard Medical School for supplying us with fly
stocks; Prof. Hilal Lashuel and Dr. Diana Olschewski from Ecole
Polytechnique Federale de Lausanne for helping with the protein
purification; Prof. Danny Offen and Dr. Nirit Lev for supplying us with
the cell cultures; and the Gazit research group for fruitful discussions. The backbone chemical shift deviations of the retro-inverso b-
syn 36 peptide upon binding to a-syn, as determined by NMR, are
largest for the hydrophobic residues L38 and V40, in addition to the
more flexible terminal residues K43 and G36. Residues L38 and V40
surround the aromatic Y39, previously shown to participate in the
oligomeric interaction [5,6], although Y39 itself showed little
change in chemical shift upon binding a-syn. It is possible that the
bulky side chain of Y39 stabilized a local conformation in the Discussion The chemical environment of this region,
reflected in the deviations in chemical shift, changes from a
monomeric, exposed conformation to a more shielded, hydropho-
bic environment upon interacting with a-syn. Taken together, our results support the idea of using the
naturally occurring b-syn protein for the development of inhibitors
of a-syn aggregation. We have mapped a decamer peptide in b-
syn which is able to inhibit both the early and late stages of a-syn
aggregation and we were able to stabilize it using several
modifications. The modified decamer peptide showed a significant
amelioration of PD-associated defects in a transgenic fly model. g
y
In this context, it is worth mentioning that although a-syn is a
typical cytoplasmic protein, a small amount of both monomeric
and aggregated forms were reported to be secreted from cells and
exist in human body fluids, such as cerebrospinal fluid [51,52]. Furthermore, extracellular a-syn was shown to be capable of
entering cells, and the mechanism of its internalization was
suggested to depend on the assembly state of the protein:
aggregated forms of a-syn, both fibrils and oligomers, enter into
cells via receptor-mediated endocytosis, while monomeric a-syn
diffuses passively across the plasma membrane [53]. It is worth
noting that there is high homology between a-syn and b-syn,
especially in the N terminal region, and the sequence of the b-syn
36 peptide can be detected within the sequence of a-syn at the
same position with one amino acid substitution of lysine to
arginine. We therefore speculate that the identified peptide either
diffuses passively across the plasma membrane or enters into cells
via
the
same
receptor-mediated
endocytosis
as
a-syn. In
accordance with the in vitro results, the retro-inverso b-syn 36
modified peptide alleviated a-syn engendered symptoms in a
transgenic fly model of PD and significantly reduced the
accumulation of aggregated a-syn in their brains. Despite its
specificity to a-syn, it will be interesting to examine the efficacy of
the peptide in other fly models of amyloidogenic proteins such as
polyQ and b-amyloid. Author Contributions Conceived and designed the experiments: RSK DES DS EG. Performed
the experiments: RSK MFP NEM AFM DES. Analyzed the data: RSK
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Biol 40: 1835–1849. PLoS ONE | www.plosone.org November 2010 | Volume 5 | Issue 11 | e13863 13
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Publisher pdf DOI for this version doi:10.1002/1873-3468.13081 doi:10.1002/1873-3468.13081 Edited by Wilhelm Just Edited by Wilhelm Just Keywords: integrin; mechanobiology; talin FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies
This is an open access article under the terms of the Creative Commons Attribution License, which permits use,
distribution and reproduction in any medium, provided the original work is properly cited. ABS, actin-binding site; DD, dimerisation domain; ECM, extracellular matrix; FA, focal adhesion; FERM, 4.1 protein, ezrin, radixin, moesin;
IBS, integrin-binding site; IF, intermediate filament; PTM, post-translational modification; VBS, vinculin-binding site. Integrin adhesions: linking the cell to
the extracellular matrix components that couple adhesions to numerous sig-
nalling cascades. These enable diverse intracellular
responses, a process often referred to as ‘outside-in’
signalling. These signalling hubs regulate a multitude
of cellular processes including cytoskeletal dynamics
and cell motility, cell growth, survival and the cellular
response to the local environment. Unsurprisingly,
numerous diseases arise from the defects in compo-
nents of the integrin adhesome [3]. Further analysis of
integrin adhesome datasets collected under different
conditions has revealed the dynamic nature of these
complexes, and the functional diversity that can derive
from the same building blocks. What emerges is a con-
sensus adhesome of ~ 60 proteins centred around four
axes comprising; ILK-PINCH-kindlin, FAK-paxillin,
talin-vinculin and a-actinin-zyxin-VASP, although it
seems likely that all of these axes are linked to talin in
some way. As well as 24 different integrins, vertebrates
also have two major talin isoforms: talin1 and talin2. Integrin-mediated adhesions to the extracellular matrix
(ECM) are found in nearly all cell types and mediate a
diverse
range
of
functions. There
are
24
ab
heterodimeric integrins, which show distinct patterns of
cell-type and tissue-specific expression, and support dif-
ferent forms of cell-ECM and cell-cell attachment. Inte-
grins connect to the ECM via their large extracellular
domains but, in contrast, the cytoplasmic domains, the
‘integrin tails’, are generally short (~ 40–60 amino
acids). Despite this diminutive size, large multiprotein
complexes assemble on the cytoplasmic face of integrins,
providing linkages to the cell cytoskeleton and to
numerous intracellular signalling pathways. The complexity of cell-matrix adhesions has been
highlighted by the analysis of the ‘integrin adhesome’
using mass spectrometry on multiple integrin adhesion
complexes. This identified a network of > 240 proteins
[1,2], and additional adhesome proteins are constantly
being discovered, many of which are cytoplasmic Most of the attention on talin has focused on talin1,
primarily due to its essential role in mediating cell The tale of two talins – two isoforms to fine-tune integrin
signalling
Rosemarie E. Gough and Benjamin T. Goult
School of Biosciences, University of Kent, Canterbury, UK Correspondence
B. T. Goult, School of Biosciences,
University of Kent, Canterbury CT2 7NJ, UK
Tel: +44 1227 816142
E-mail: b.t.goult@kent.ac.uk
(Received 16 March 2018, revised 12 April
2018, accepted 26 April 2018, available
online 18 May 2018)
doi:10.1002/1873-3468.13081
Edited by Wilhelm Just Talins are cytoplasmic adapter proteins essential for integrin-mediated cell
adhesion to the extracellular matrix. Talins control the activation state of inte-
grins, link integrins to cytoskeletal actin, recruit numerous signalling molecules
that mediate integrin signalling and coordinate recruitment of microtubules to
adhesion sites via interaction with KANK (kidney ankyrin repeat-containing)
proteins. Vertebrates have two talin genes, TLN1 and TLN2. Although talin1
and talin2 share 76% protein sequence identity (88% similarity), they are not
functionally redundant, and the differences between the two isoforms are not
fully understood. In this Review, we focus on the similarities and differences
between the two talins in terms of structure, biochemistry and function, which
hint at subtle differences in fine-tuning adhesion signalling. Correspondence
B. T. Goult, School of Biosciences,
University of Kent, Canterbury CT2 7NJ, UK
Tel: +44 1227 816142
E-mail: b.t.goult@kent.ac.uk (Received 16 March 2018, revised 12 April
2018, accepted 26 April 2018, available
online 18 May 2018) Enquiries If you have questions about this document contact ResearchSupport@kent.ac.uk. Please include the URL of the record
in KAR. If you believe that your, or a third party's rights have been compromised through this document please see
our Take Down policy (available from https://www.kent.ac.uk/guides/kar-the-kent-academic-repository#policies). REVIEW ARTICLE Abbreviations ABS, actin-binding site; DD, dimerisation domain; ECM, extracellular matrix; FA, focal adhesion; FERM, 4.1 protein, ezrin, radixin, moesin;
IBS, integrin-binding site; IF, intermediate filament; PTM, post-translational modification; VBS, vinculin-binding site. 2108 R. E. Gough and B. T. Goult Tale of two talins Fig. 1. Talin at the core of the adhesion. A cartoon of the core of
integrin adhesions, highlighting talins central role. Talin coordinates
both the actin cytoskeleton, and through the interaction with KANK
proteins, the microtubule cytoskeleton at adhesion sites. Once the
adhesion core is assembled, talin serves as a scaffold to recruit many
other proteins in order to form all the many different types of
adhesive structures (focal adhesions, podosomes, invadopodia, etc.). adhesion as shown by studies on talin1 knockout [4]
and talin1-depleted cells [5–7]. Talin1 knockout is also
embryonic lethal in mice due to arrested gastrulation,
indicating a key role in early development [4]. In con-
trast talin2, which was only discovered following pub-
lication of the human genome sequence [8,9] has
received less attention, and the fact that talin2 knock-
out mice are viable and fertile [10] suggests isoform
redundancy. However, talin2 knockout mice display a
mild dystrophic phenotype and the variability in the
number of pups surviving to adulthood suggest under-
lying defects [10]. Therefore, it appears that talin2
plays important roles in development although many
of its functions can be compensated for by talin1. This Review aims to summarise what is currently
known about the structural, biochemical and func-
tional differences between the two talin isoforms. Evo-
lutionary
genomics has been
used to study talin
isoforms in the past, and here, we combine genetic
analysis with the recent wealth of structural informa-
tion to highlight the emerging functions of the two tal-
ins as signalling platforms. Fig. 1. Talin at the core of the adhesion. A cartoon of the core of
integrin adhesions, highlighting talins central role. Talin coordinates
both the actin cytoskeleton, and through the interaction with KANK
proteins, the microtubule cytoskeleton at adhesion sites. Once the
adhesion core is assembled, talin serves as a scaffold to recruit many
other proteins in order to form all the many different types of
adhesive structures (focal adhesions, podosomes, invadopodia, etc.). Integrin–talin–actin: the core of
cell-matrix adhesions Strikingly, despite the structural complexity of cell-
ECM adhesions, the majority of these dynamic adhe-
sion complexes comprise a simple and robust core of
three proteins: talin which binds to and activates inte-
grins and couples them to the actin cytoskeleton
(Fig. 1). All of the other components can be assembled
on to this framework to give rise to various types of
adhesive structures. Once formed, the protein vinculin
is recruited to the complex to stabilise the connection
to actin, a process which is regulated by an elegant
feedback mechanism. Thus, vinculin is only recruited
when talin experiences mechanical force, and force will
only be exerted when talin is successfully bound to an
integrin and coupled to actin. If these conditions are
not met, then nascent adhesions will not experience
sufficient force to recruit vinculin and will disassemble. However, once vinculin is recruited to nascent adhe-
sions, it crosslinks talin to actin, and the core, linkages
are stabilised. How such complexes mature depends on
multiple variables including cell type, ECM composi-
tion, matrix stiffness, integrin subtype, mechanical sig-
nals, etc. This leads to the development of a variety of
adhesion complexes including nascent adhesions, focal
adhesions
(FAs),
fibrillar
adhesions,
podosomes,
invadopodia, etc., all of which have at their core the
same integrin–talin–actin connection. In their recent
Review, Klapholz and Brown eloquently describe the myriad of different roles that talin plays in adhesion,
and provocatively call talin ‘the master of integrin
adhesions’, a view we share [11]. Tale of two talins Tale of two talins small introns resulting in a gene of ~ 30 kb, talin2 is
much bigger (~ 190 kb), due to the presence of much
larger introns. Moreover, initial studies suggest that
multiple talin2 isoforms are generated via differential
splicing [36]. While the function of these isoforms is
currently unknown, the expression pattern of each is
distinct. Testes, kidney and brain express short C-
terminal proteins lacking the FERM domain [36] rais-
ing the possibility that such variants might function
independent of integrins, although they do contain the
integrin-binding
site
located
in
the
rod
domain. Intriguingly, expression of a C-terminal talin1 frag-
ment resembling the testes-specific talin2 isoform was
sufficient to rescue cell cycle progression in talin1-
depleted cells suggesting a role in cellular signalling
[37,38]. Tissue expression and cellular localisation vary con-
siderably between the two isoforms; talin1 is expressed
in all tissues. In contrast, talin2 expression is more
variable, and it is absent entirely from some cell types,
for example, no talin2 is present in endothelial cells
possibly via silencing of the Tln2 gene by promoter
methylation [26,27]. The Human Protein Atlas [28]
shows the near ubiquitous expression of talin1 in all
cell types in all tissues, whereas the high levels of tal-
in2 are found mainly in the brain, particularly the
cerebral cortex, heart muscle and the kidney. There is clear interplay in the expression of the two
talin isoforms, although the mechanism for this is not
known. Talin2 expression is rapidly upregulated fol-
lowing knockout of talin1, both transiently [26] and
also in Tln1-knockout mouse embryonic fibroblasts [7]
leading to rescue of many of the consequences of the
loss of talin1. However, knockout of both talin iso-
forms completely ablates cell-ECM adhesion [5], con-
firming the essential role of talins in integrin biology. The ancestral Tln gene appears to have undergone
duplication in chordates with the emergence of verte-
brates to give rise to talin1 and talin2 [39]. Inverte-
brates and simple chordates have a single talin gene;
vertebrates have two. Chordates can be divided into
three major groups: Craniata (including the verte-
brates), Cephalochordata (including the lancelets) and
Tunicata (including sea squirts). Since the original
publication on talin evolution [39], the genomes of
Petromyzon marinus (one of the Cyclostomata, a jaw-
less vertebrate) [40] and Branchiostoma floridae (a lan-
celet, one of the Cephalochordata) [41] have been
published. Tale of two talins Strikingly, the Branchiostoma has only a
single talin, whereas the Petromyzon genome encodes
two. This confirms and extends the original conclu-
sions about talin evolution [9,39] and suggests that the
genome duplication leading to present day talin1 and
talin2 took place before the divergence of jawed and
jawless vertebrates, but after (or with) the divergence
of the craniates from other chordates. The acquisition
of two different copies of talin appears to be beneficial
to
an
organism;
talin
gene
duplication
has
also
occurred in Amboebozoa and in Dictyostelium dis-
coideum. The Dictyostelium talin genes, TalA and
TalB, encode proteins with distinct functions, with
TalA required for cell–substrate adhesion, phagocyto-
sis and cytokinesis, and TalB required for the force
transmission required to support morphogenetic move-
ments during differentiation [42]. g
g
gy
In fibroblasts, both talins localise to FAs, and talin1
is recruited directly to the leading edge, via proteins
like the Rap1 effector RIAM (Rap1-interacting adap-
ter molecule) [29,30] and FAK (focal adhesion kinase)
[31]. In contrast, less is known about talin2 recruit-
ment. Although talin2 binds to RIAM, talin2-specific
antibodies reveal that talin2 forms diffuse aggregates
throughout the cell, which overtime coalesce to form
larger complexes, either at focal or fibrillar adhesion
sites [25]. For the most part, only talin2 is found at
fibrillar adhesions in the centre of the cell [25]. This
localisation positions talin2 at sites of fibronectin
secretion and assembly [32] and also to formation of
invadopodia [33]. Although much less is known about
the role of talin2, it has recently been the subject of
increased interest and isoform-specific functionalities
have been reported. For example, talin2 has been
reported to be indispensable for the generation of trac-
tion force and invadopodium-mediated matrix degra-
dation
required
for
invadopodia
formation
[33]. Furthermore, talin2 has been shown to be able to
recruit vinculin in the absence of mechanical force sug-
gesting different mechanical properties [34]. In sum-
mary, the relative roles of talin1 and talin2 remain to
be fully elucidated. Talin domain structure Remarkably, despite millions of years of evolutionary
time since talin first appeared and since the two talins
diverged, the length of both the major talin isoforms
has remained almost identical (talin1: 2541aa; talin2:
2540aa). Furthermore,
both
major
isoforms
have The talins Talin is a large 270 kDa actin-binding protein that was
first discovered in 1983 as a component of FAs and ruf-
fling membranes [12]. Talin comprises an N-terminal
FERM domain (the head) coupled to a flexible talin
rod. Since then, it has been shown to be a key compo-
nent of integrin adhesions with roles in integrin activa-
tion [13], the molecular clutch that couples integrins to
cytoskeletal actin [14], FA assembly and the recruitment
of numerous signalling molecules [15]. Talins also inter-
act with the KANK family of adapter proteins [16,17]
which target microtubules to adhesion sites, stimulating
FA turnover [18]. As well as transmitting forces between
integrins and the actin cytoskeleton, the length of talin
has been shown to define the geometry of the adhesion
[19], and talin plays a key role as a mechanosensitive
adapter, undergoing force-dependent conformational
transitions in its 13 rod domains [20–24] that modulate
binding interactions with mechanosensitive ligands. Given the above, it is unsurprising that talin1 knockout
in mice is embryonic lethal [4,25]. 2109 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies Gene structure and splice variation in talins The two talins are encoded by separate genes, Tln1
and Tln2, which have conserved intron–exon bound-
aries [9,35]. However, whereas talin1 has relatively 2110 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins identical domain structure and contain 18 domains. This invariability is in stark contrast to many other
multidomain proteins such as titin, spectrin and fil-
amin – these have varied in length, increasing and
decreasing in size through evolution until reaching the
length we see today [43–45]. This unvarying domain
arrangement in all available talin sequences suggests
that
each
domain
has
a
role
that
is
universally
required (it is worth mentioning that in some nonver-
tebrate organisms, including Dictyostelium and Droso-
phila, talin has acquired additional C-terminal residues
that extend beyond the universal domain arrangement
[39,46]). The following discussion of talin domain
structure, therefore, applies to both talin1 and talin2. plasma membrane, which applies torque on the inte-
grin to stabilise the active conformation [52,56,57]. In
addition,
the
F1
subdomain
contains
a
large
(~ 30aa) unstructured insertion, the F1-loop, which,
via a cluster of positively charged residues, interacts
with PIP2 and is essential for integrin activation
[58]. The F0 subdomain has been shown to bind the
membrane-tethered small GTPase, Rap1 [58–60] and
this interaction has been implicated in membrane
targeting
of
talin
to
the
plasma
membrane
[59]. Interestingly, the additional F0 subdomain and the
F1-loop elements of the talin FERM domain are
also found in the kindlin family of proteins [58,61]
which synergise with talin to activate integrins [62]. These
features
are
not
found
in
other
FERM
domain proteins and are unique to integrin-activating
FERM domain proteins. The talin head Talins consists of an atypical N-terminal FERM (4.1
protein, ezrin, radixin, moesin) domain, known as
the talin head containing four subdomains F0–F3
[47,48] rather than the three subdomains (F1–F3)
found in most other FERM domain proteins. More-
over,
the
crystal
structure
of
the
talin1
FERM
domain shows a linear domain structure [47] rather
than the cloverleaf structure found in other FERM
domain proteins. The structure of the talin2 FERM
domain
confirms
this
linear
domain
arrangement
(our unpublished data). Extensive studies show that
it is the talin F3 subdomain that directly engages
the beta-integrin cytoplasmic tail via the first (mem-
brane proximal) of two NPxY motifs in the tail [49]. The
integrin-binding
interfaces
have
been
charac-
terised in both the talin1 and talin2 F3 subdomains,
and this has revealed that conserved residue changes
in the binding surfaces tune the affinities of the two
talins for different integrin tails [50]. For example,
the ubiquitous beta1a-integrin was recently shown to
bind preferentially to talin2 [51] whereas the muscle-
specific beta1d-integrin has a threefold higher prefer-
ence for talin2 over talin1 [33,52,53]. This provides
selectivity for different talin and integrin complexes,
and different couplings are likely to regulate differ-
ent cellular functions [54]. As well as binding to integrins, Rap1 and the mem-
brane, the talin head (via the F3 subdomain) has been
shown to bind to PIP kinase gamma [63], which is
thought to generate the PIP2 required to support inte-
grin activation [64]. Beyond this, the F3 subdomain
has emerged as showing remarkable ligand-binding
plasticity and has been linked to binding FAK [31],
TIAM1 (T-cell lymphoma invasion and metastasis 1)
[65], layilin [66], Ga13 (G-protein subunit Galpha13)
and RIAM [67] all via the same site. The hierarchy of
these interactions, that are presumably mutually exclu-
sive with integrin binding and each other, is not yet
fully understood. Talin contains three actin-binding
sites (ABS1-3) [68]. ABS1 is in F2-F3 in the talin head
[69] and has recently been shown to be important for
capping actin filaments to block actin polymerisation
[70]. FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies The talin rod The talin head is connected, via an 82-amino acid
unstructured [71] calpain-sensitive linker [72,73] to the
large 2000 residue talin rod that is made up of 62 a-
helices. We have recently determined the boundaries
and structures of the talin1 rod domains showing it
contains 13 domains (R1–R13) [24] organised into two
functionally distinct regions, a linear C-terminal rod-
like region comprised of 5-helix bundles and a com-
pact N-terminal region where three 4-helix bundles
(R2–R4) are inserted into the series of 5-helix bundles
(Fig. 2b). However, whilst F3 is the only talin head subdo-
main that engages the integrin, F3 in isolation is not
very effective at activating integrins and the other
head subdomains are also required to make an effec-
tive ‘integrin activation lock’ and maintain the inte-
grin in the active, high-affinity conformation [55]. The other head subdomains achieve this by interac-
tion
with
phosphoinositides
such
as
PtdIns(4,5)P2
(PIP2) in the plasma membrane; a basic surface on
the
F2
subdomain
mediates
interaction
with
the Structural analysis of the talin rod was complicated
as only two regions of the rod have sequence homol-
ogy
to
other
proteins:
R13,
which
contains
an
I/LWEQ domain [8,74], and the central region of the
rod (resolved to be R7–R8) which has homology to a 2111 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins protein of unknown function, MESDC1 [75]. The rest
of the rod lacks homology to other proteins meaning
possible. Part of the reason for the lack of homolog
of the talin rod to other proteins turned out to be th
Fig. 2. Structure and domain map of the two talin isoforms. (A) Structural model of talin showing the domain arrangement of talin. Vincul
binding sites are shown in red. The N-terminal talin head comprising F0–F3 and the talin rod domains R1–R13 are shown. (B) Schema
representation of the talin domain structures coloured by sequence identity between the two isoforms. The domain boundaries are giv
for mouse talin1 (UniProt: P26039) and talin2 (UniProt: B2RY15). Provided that these boundaries are used, it is possible to make any ta
fragment or delete any talin domain while maintaining the structural integrity of the protein. (C) The locations of many of the talin ligan
binding sites are shown, as are the calpain cleavage sites. Fig. 2. The talin rod Structure and domain map of the two talin isoforms. (A) Structural model of talin showing the domain arrangement of talin. Vincu
binding sites are shown in red The N terminal talin head comprising F0 F3 and the talin rod domains R1 R13 are shown (B) Schem Fig. 2. Structure and domain map of the two talin isoforms. (A) Structural model of talin showing the domain arrangement of talin. Vinculin-
binding sites are shown in red. The N-terminal talin head comprising F0–F3 and the talin rod domains R1–R13 are shown. (B) Schematic
representation of the talin domain structures coloured by sequence identity between the two isoforms. The domain boundaries are given
for mouse talin1 (UniProt: P26039) and talin2 (UniProt: B2RY15). Provided that these boundaries are used, it is possible to make any talin
fragment or delete any talin domain while maintaining the structural integrity of the protein. (C) The locations of many of the talin ligand-
binding sites are shown, as are the calpain cleavage sites. possible. Part of the reason for the lack of homology
of the talin rod to other proteins turned out to be that
8 of the 13 talin rod domains – R1, R5–R7, R9–R12 – protein of unknown function, MESDC1 [75]. The rest
of the rod lacks homology to other proteins meaning
prediction of the domain boundaries a priori was not possible. Part of the reason for the lack of homology
of the talin rod to other proteins turned out to be that
8 of the 13 talin rod domains – R1, R5–R7, R9–R12 – 2112 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins contain a 5-helix bundle fold, the ‘talin rod fold’, that
has so far only been recognised in talin. with the two talin heads buried inside. Activation of tal-
ins to a more open-active conformation requires a vari-
ety of activators. Integrin As well as interacting with the talin head, the beta-
integrin tail also interacts with the R11–R12 domains
of the talin rod [integrin binding site 2 (IBS2)] via a
structurally undefined mechanism [79–81]. Unlike IBS1
where the integrin binds to the folded F3 subdomain,
integrin binding to IBS2 appears to involve some inter-
mediate conformation of the rod domains (integrin
does not bind folded or unfolded R11–R12). The role
of IBS2 in flies [79,82] has been well established, but
its role in mammals is less clear, although it has been
linked to nascent adhesion formation [83]. Ligand-binding sites in the talin rod Elucidation of the domain structure of the talin rod
has enabled the precise mapping of established ligand-
binding sites, and the location of these sites is shown
in Fig. 2. Talin1 and talin2 rod interactions Although 4-helix bundles are common in nature (the
4-helix up–down bundle present in R2, R3, R4 and R8
is a common fold (SCOP 47161 [76])), 5-helix bundles
are unusual. At the core of the talin rod fold is a com-
mon 4-helix up–down, left-handed twist topology as
seen in numerous 4-helix bundles. However, in the
talin rod, this fold is augmented by an extra N-term-
inal helix, connected by a long (~ 9 residue) loop that
allows the first helix to pack against helices 3 and 4 of
the bundle (Fig. 2) to form a 5-helix bundle. This
addition of an extra helix to the talin rod domains has
profound effects on talin function, not least because 5-
helix bundles, where the N- and C-termini are located
at opposite ends of the bundle are optimal for forming
a rod-like arrangement (Fig. 2). The linear rod-like
region is perfectly designed to transmit forces, which
act on the compact N-terminal region. Furthermore,
the additional helix significantly enhances the thermal
and mechanical stability of the domains [22,77], help-
ing provide different mechanical responses for each
domain [21] and restricting access to the cryptic vin-
culin-binding sites (VBS) buried inside many of the
domains. As a result, each rod domain has its own
unique properties, and this is central to talins role as a
mechanosensor (see next section). Unfolding 5-helix
bundles by pulling on the termini positioned at oppo-
site ends of the bundle is restricted by extensive con-
tacts throughout the length of the helices and requires
a gradual breaking of hydrogen bonds. In 4-helix bun-
dles, the termini are at the same end, and applied force
acts on the weak hydrophobic contacts, peeling helices
away from the bundle [22]. To date, all the ligands that bind to talin1 have been
shown to bind talin2 although the affinities for the dif-
ferent isoforms can be markedly different. Binding
partners can interact with the talin rod domains via a
number of different modes, that is, to the folded rod
domains, to the unfolded rod domains or to some
strained conformation between these two extremes. Mechanical force can drive transitions between these
conformations and so dramatically alter the binding
affinities of different ligands. FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies Vinculin A common mechanism for talin rod-binding proteins
is via helix addition, whereby a helix from a ligand
packs against the side of a talin rod domain. A num-
ber of talin ligands have now been identified that con-
tain
an ‘LD-motif’
[92] that mediates
such helix
addition. First identified in paxillin [93], amphipathic
LD-motif helices bind via the aspartate (D) which
forms an initial salt bridge with a basic residue at the
beginning of the furrow between two adjacent helices
of the interacting bundle. Specificity is then encoded
by residues downstream of the ‘LD’ interaction site. This mode of binding to talin was initially identified
from work on the tumour suppressor protein deleted
in liver cancer 1 (DLC1) [94,95]. This led on to the
identification of the talin-binding sequence in RIAM
[24,29] as an LD-motif, and the identification of pax-
illin as a novel talin ligand [94]. More recently, the
KANK (kidney ankyrin repeat-containing) proteins
have been identified as LD-motif-containing ligands
[16,17], binding to a conserved face on the R7 5-helix
bundle. The
ability
of
5-helix
bundles
to
bind Vinculin, discovered in 1979 [87], is another key talin
interactor, and has been shown to bind to at least 11 of
the 62 talin1 helices [88]. The vinculin-binding determi-
nants lay on one side of each vinculin-binding helix [89]. However, the VBS are buried within the rod domains
and are only exposed by mechanical force (Fig. 3),
enabling vinculin to bind and strengthen the actin con-
nection. It has been shown that exposed talin VBS can
activate vinculin [90], and active vinculin has been
shown to be able to activate talin [91]. The 11 VBS in
talin2 are all conserved and so it is likely that the talin2
rod will also engage vinculin in a similar fashion. Talin dimers The talin rod contains two actin-binding sites, ABS2
(R4–R8) [68,84,85] and ABS3 (R13-DD) [8,86] which
play different roles in adhesion. The current model of
talin function envisages the C-terminal ABS3 [74] as
responsible for the initial force exerted on talin that
leads to unfolding of the mechanosensitive talin rod
domain, R3. This triggers vinculin interactions and
leads to adhesion maturation [23]. In contrast, ABS2,
in the centre of the rod provides the tension-bearing
actin connection [84,85]. As with the integrin connec-
tions, the actin-binding sites in talin2 bind more tightly
to actin than the equivalent regions in talin1 ([35] and
our unpublished data). Full-length talin is dimeric, and helix 62 [dimerisation
domain (DD)] forms an antiparallel dimer with another
talin molecule [74]. In all our experiments to date, we
see talin as a constitutive dimer when the DD is present;
however, a calpain cleavage site immediately prior to
the DD means it can be cleaved to yield monomeric
talin [71]. Interestingly, the DD in talin2 is conserved
with talin1, and structural predictions suggest it should
be able to form heterodimers. However, to our knowl-
edge, heterodimers have not been described in the litera-
ture. Dimeric full-length talin1 can adopt a compact
autoinhibited conformation in the cytosol [78] where the
two rod domains wrap around to form a ‘double donut’ 2113 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins R. E. Gough and B. T. Goult FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies Other interactors In the extended conformation in the absence of force, all the domains are still folded, and additional binding sites are exposed (IBS1, IBS2,
ABS3, plus the sites for those ligands that require folded-rod domains) (Right). The exposure of IBS1 and ABS3 facilitates adhesion
formation, and by activating integrins and crosslinking them to the actin cytoskeleton, a nascent adhesion can form. As force is exerted on
talin, another layer of autoinhibition is uncovered (Bottom). As talin domains unfold, starting with R3, the initial mechanosensor in talin
[21,23,24], vinculin-binding sites are exposed and talin:vinculin interactions can now occur. R3 unfolding also reveals the high affinity actin-
binding site in talin, ABS2 that can then activate tension-bearing actin connections [84,85]. As domains unfold, the binding sites for ligands
that engage the folded rod domains are destroyed, as is the case for RIAM binding to R3. A remarkable feature of talins conformational
plasticity is that, in the absence of other factors, talin can readily refold to its default low-force state. Fig. 3. Layers and layers of autoinhibition. A striking feature of talins is their remarkable conformational plasticity that enables different
ligands to engage the same platform in different conditions; part of this flexibility emerges from autoinhibition. (Left) In the closed
autoinhibited form, all of the domains are folded, and many of the ligand-binding sites for actin, integrin and vinculin are thought to be
cryptic. Some binding sites may face outwards and remain accessible; for instance, RIAM is able to bind to the inactive conformation [127]. In the extended conformation in the absence of force, all the domains are still folded, and additional binding sites are exposed (IBS1, IBS2,
ABS3, plus the sites for those ligands that require folded-rod domains) (Right). The exposure of IBS1 and ABS3 facilitates adhesion
formation, and by activating integrins and crosslinking them to the actin cytoskeleton, a nascent adhesion can form. As force is exerted on
talin, another layer of autoinhibition is uncovered (Bottom). As talin domains unfold, starting with R3, the initial mechanosensor in talin
[21,23,24], vinculin-binding sites are exposed and talin:vinculin interactions can now occur. R3 unfolding also reveals the high affinity actin-
binding site in talin, ABS2 that can then activate tension-bearing actin connections [84,85]. As domains unfold, the binding sites for ligands
that engage the folded rod domains are destroyed, as is the case for RIAM binding to R3. Other interactors While the integrin-, actin- and vinculin-binding inter-
actions define the primary adapter function of talin,
there are an increasing number of additional ligands
that bind to the talin rod that contribute to its
mechanosignalling capabilities. These are summarised
below. Fig. 3. Layers and layers of autoinhibition. A striking feature of talins is their remarkable conformational plasticity that enables different
ligands to engage the same platform in different conditions; part of this flexibility emerges from autoinhibition. (Left) In the closed
autoinhibited form, all of the domains are folded, and many of the ligand-binding sites for actin, integrin and vinculin are thought to be
cryptic. Some binding sites may face outwards and remain accessible; for instance, RIAM is able to bind to the inactive conformation [127]. In the extended conformation in the absence of force, all the domains are still folded, and additional binding sites are exposed (IBS1, IBS2,
ABS3, plus the sites for those ligands that require folded-rod domains) (Right). The exposure of IBS1 and ABS3 facilitates adhesion
formation, and by activating integrins and crosslinking them to the actin cytoskeleton, a nascent adhesion can form. As force is exerted on
talin, another layer of autoinhibition is uncovered (Bottom). As talin domains unfold, starting with R3, the initial mechanosensor in talin
[21,23,24], vinculin-binding sites are exposed and talin:vinculin interactions can now occur. R3 unfolding also reveals the high affinity actin-
binding site in talin, ABS2 that can then activate tension-bearing actin connections [84,85]. As domains unfold, the binding sites for ligands
that engage the folded rod domains are destroyed, as is the case for RIAM binding to R3. A remarkable feature of talins conformational
plasticity is that, in the absence of other factors, talin can readily refold to its default low-force state. Fig. 3. Layers and layers of autoinhibition. A striking feature of talins is their remarkable conformational plasticity that enables different
ligands to engage the same platform in different conditions; part of this flexibility emerges from autoinhibition. (Left) In the closed
autoinhibited form, all of the domains are folded, and many of the ligand-binding sites for actin, integrin and vinculin are thought to be
cryptic. Some binding sites may face outwards and remain accessible; for instance, RIAM is able to bind to the inactive conformation [127]. The talin-moesin-NHE-1 complex and pH modulation of
adhesion sites The talin-moesin-NHE-1 complex and pH modulation of
adhesion sites The C-terminal part of the talin rod has also been
shown to bind directly to the FERM domain of moe-
sin, an interaction that is required to recruit the
sodium/hydrogen exchanger (NHE-1) to adhesion sites
[96]. This recruitment of a proton exchanger to adhe-
sions and the resulting localised alterations of intracel-
lular pH has a dramatic effect on adhesions. Small
changes in local pH can result in protonation/deproto-
nation of side chains, particularly histidines, and this
can directly alter interactions in a similar fashion to
phosphorylation. In many ways, protonation can be
considered
a
post-translational
modification
[97]. Many important protein:protein interactions have been
shown to be regulated by pH in this way, including
the interaction of talin ABS3 with actin [98], and it is
likely that local fluctuations in pH will alter the proto-
nation state of many other important interaction sites. Other interactors A remarkable feature of talins conformational
plasticity is that, in the absence of other factors, talin can readily refold to its default low-force state. 2114 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins LD-motif proteins greatly expands the number of
potential ligand-binding sites in talin. LD-motif proteins greatly expands the number of
potential ligand-binding sites in talin. The talin–alpha-synemin connection – a link to
intermediate filaments? Another ligand that has been linked to talin which has
the potential to have a significant impact on our view
of adhesions is alpha-synemin [99], an intermediate fil-
ament (IF) protein expressed in skeletal muscle. This
suggests that talin has the potential to coordinate
interactions between the actin, microtubule and IF net-
works. Mechanotransduction: force driving changes in
biological signalling The mutually exclusive interactions between talin and
RIAM and talin and vinculin provide the perfect
example of how talin can convert mechanical forces
into
biological
signalling
responses. The
initial
mechanosensitive domain in talin has been shown to
be R3 [23,24], which binds RIAM but also contains
two VBS. However, vinculin and RIAM have funda-
mentally different modes of binding. Talin VBS are
buried within the talin rod domains and are only
exposed when mechanical force unfolds that domain,
allowing vinculin to bind. In contrast, the talin-binding
sites in RIAM are single helices that interact only with
folded
talin
rod
domains
(Fig. 2). The
exquisite
mechanosensitivity of R3 is due to the presence of a
destabilising cluster of threonine residues buried in its
hydrophobic core [24]. This means that the R3 domain
is the first to unfold when talin experiences force, driv-
ing the transition between folded and unfolded R3
(this is one of the exciting aspects of structural
mechanobiology in that the precise structural basis of
a mechanosensitive event can be pinpointed to specific
amino acids that encode the mechanosensitivity). This
conformational change in R3 drives a change in bio-
logical signalling, displacing RIAM and thus the link
to the Rap1 signalling pathways. Simultaneously, R3
unfolding leads to the recruitment of vinculin and
strengthening of the connection to actin. This allows
two different ligands to engage the same talin domain
under different conditions and explains the different
localisation of RIAM and vinculin in cells [100]. In the
case of R3, a force of ~ 5 pN is required for it to
unfold, disrupting the RIAM-binding sites and recruit-
ing vinculin, driving the maturation of nascent adhe-
sions into FAs. This 5 pN force is roughly the force of
a single actomyosin contraction, leading to an attrac-
tive hypothesis that talin only experiences this force
threshold when it binds to an integrin and simultane-
ously connects to the actin cytoskeleton. Only when
these two criteria are met will the R3 domain unfold
and trigger adhesion maturation. Talin: layers and layers of autoinhibition An interesting feature of talins is that the binding sites
described above are not all accessible all of the time. Talin activity is regulated by multiple layers of autoin-
hibition where binding sites are masked, and only
made available for binding in response to different sig-
nals. Talin autoinhibition mediated via the interaction
between the integrin-binding site in F3 and the talin
rod domain R9 maintains talin in a compact cytosolic
form [104–107]. The F3-binding surface on talin2 R9
is
highly
conserved
with
only
subtle
conservative
changes, and as such autoinhibition is likely common
to both talins. Multiple factors (e.g. PIP2, FAK, Vin-
culin, RIAM, etc.) have been implicated in relieving
talin
autoinhibition,
most
recently,
the
G-protein
Ga13 which binds F3, displacing the R9 rod domain,
has emerged as an important talin regulator [108]. Once autoinhibition is relieved, it is likely that some of
talins functionalities are exposed, such as the integrin-
and membrane-binding sites on the talin head [52,109],
and the C-terminal actin-binding site ABS3 [8,74]. However, other functions are still autoinhibited; for
example, the VBS remain inaccessible, buried in the
hydrophobic core of the rod domains. As mechanical
force is exerted on talin, its rod domains can unfold,
exposing VBS and simultaneously destroying the bind-
ing
sites
for
folded
rod
binders,
enabling
mechanochemical switching of binding. In this scenar-
io, once RIAM has served its purpose and helped
translocate talin to the plasma membrane, its binding
to talin is no longer required and so those domains are
repurposed
for
alternative
functions. Furthermore,
high affinity actin binding is mediated via the central
actin-binding site (ABS2; R4–R8) which is maintained
in an inactive conformation via the inhibitory effects
of the adjacent R3 and R9 domains [84]. As a result
of this stratified nature of talin autoinhibition, the
same protein scaffold can coordinate many different
processes. There are likely numerous other talin func-
tions tightly regulated by talin conformation in a simi-
lar fashion. The R8 domain is a hotspot for protein interactions,
suggesting it represents a major signalling hub. R8 is a
4-helix bundle, uniquely protected from mechanical
force by being inserted into the loop of a 5-helix bun-
dle (R7), creating a novel 9-helix module, and a
branch in the talin rod [21,75] (Fig. 2). By being posi-
tioned outside of the force-bearing region, R8 remains
folded whilst talin is under force and maintains its
ligand-binding surface. Talin: layers and layers of autoinhibition A striking feature of the talin rod’s response to force
is that even after complete unfolding, the removal of
force leads to refolding to the original native state,
and this response is maintained through multiple
cycles of extension and relaxation. The robustness of
the mechanical response of the talin rod is perfectly
suited to its role as a mechanosensor; when the
mechanical force is relieved, the sensor reverts back to
its original state. Together, these features suggest that talins can sense
and respond to mechanical forces with remarkable ver-
satility. Depending on the applied force, different
domains will unfold, and depending on the repertoire
of expressed ligands, different signals will be gener-
ated. Depending on the mechanism of linkage to actin
(e.g. via ABS2 vs. ABS3), or to microtubules via
KANK, different regions of talin will be under ten-
sion. This network of protein interactions thus pro-
vides a mechanism for context and force-dependent
regulation of multiple signalling pathways. It will be important to characterise the mechanical
response
of
talin2
as
differences
in
mechanical
responses of individual rod domains might help pin-
point sites of functional divergence. Recent work has
shown that the two talins provide different mechanical
linkages in cells [34], with talin2 able to engage vin-
culin in the absence of mechanical force, suggesting
that the two proteins respond to forces differently. Talin2 is expressed at high levels in cardiac and skele-
tal muscle [103] where presumably its higher affinity The mechanical properties of talin The mechanosensing abilities of talin rely on its force-
dependent interactions with its binding partners. Some
ligands (i.e. RIAM, KANK, DLC1, actin) bind only
to folded talin domains, whereas vinculin is known to
require domain unfolding and exposure of cryptic
VBS. Force-induced
talin
domain
unfolding
will
therefore release binding partners that bind to folded
talin and stimulate binding of vinculin, triggering
mechanosensing
signals. In vivo,
talin
is
initially
extended by actin retrograde flow and then by acto-
myosin contractility and the resulting forces exerted
on the talin rod drive structural transitions. Depend-
ing on the precise mechanical environment, individual
talin molecules will experience different forces, and
the
different
conformations
may
engage
different
ligands. Expanding this to the rest of the talin rod, it seems
likely that each of the talin rod domains can also serve
as mechanochemical switches, and under different con-
ditions, individual talin rod domains can adopt differ-
ent conformations that support different signalling
pathways. In vivo measurements of talin extension
have shown that talin length is normally between 90
and 250 nm [101] (compared with a folded talin length
of 50–60 nm in vitro [102]), suggesting that between
two and eight talin rod domains are unfolded at any 2115 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins for integrin beta1d and actin may serve to create more
resilient adhesive connections. time [21]. Using single molecule analysis with ultra-
stable magnetic tweezers, we recently characterised the
mechanical response of talin1 [21–23]. Stretching the
whole
talin
rod
revealed
quantised
mechanical
responses with all 13 rod domains exhibiting switch-
like behaviour at different force thresholds. These
unfolding responses range from ~ 5 to 25 pN and are
all rapidly reversible when force is removed. This
reveals
a
spectrum
of
mechanosensitive
switching
events, turning on and off distinct effector functions in
a force-dependent manner. This stochastic force-depen-
dent folding and refolding of talin also make talin an
effective
force
buffer
protecting
adhesions
against
excessive force [21]. Talin2 in disease and development The roles of talin2 during embryogenesis and develop-
ment are not fully understood, but studies of the two
talin isoforms in the heart reveal that they are tightly
regulated [5,111]. Both isoforms are highly expressed
in cardiomyocytes, but during maturation, and in the
mature heart, talin2 becomes the major isoform, local-
ising to the costameres [111]. Indeed, cardiac-specific
talin1 knockout mice show normal basal cardiac func-
tion. Interestingly, talin1 is upregulated in the failing
human heart, and studies in mice show that an abla-
tion of cardiac talin1 blunts the hypertrophic response
and improves cardiac function [106]. The mechanisms
behind isoform switching in heart remain to be eluci-
dated, but the data clearly indicate that the two talin
isoforms play distinct roles in cardiac muscle. Further
evidence of the importance of talin2 in development
comes from the exome sequencing-based identification
of a mutation (S339L in F3) in the Tln2 gene that
causes fifth finger Camptodactyly [112]. Given that tal-
in2 is not an essential gene, it seems likely that whole
exome sequencing will reveal further disease-associated
mutations in the Tln2 gene, and these will provide fur-
ther insights into its functions. We used BlastP to align the sequences of the corre-
sponding domains from each isoform to establish the
identity and similarity of each domain and to look for
local variations. Interestingly, although the sequence
identity between the two talins is 76%, sequence iden-
tity at the domain level shows much greater variation. The F2 (86%), F3 (89%) and R13 (92%) are highly
conserved between isoforms, supporting their role in
assembly of the core adhesion complex. In contrast,
other regions of the talin rod show considerable varia-
tion in conservation between isoforms with R5 (60%)
being the most divergent. Using a sample set of vertebrates, we compared the
conservation of each individual talin domain between
species and between isoforms. Regions where sequence
conservation is low within and between the individual
isoforms likely represent regions of less functional
importance. In contrast, regions that are highly con-
served
within
an
isoform
but
are
less
conserved
between isoforms might indicate regions of functional
divergence. This structure-oriented conservation analy-
sis reveals that for some talin rod domains, the bind-
ing
surfaces
are
completely
conserved
between
isoforms. For example, talin rod domains, R7 and R8,
both contain binding sites for LD-motif-containing
proteins, and the R7- and R8-binding surfaces on both
talin isoforms are identical. Conserved differences between the two isoforms The two talins are highly conserved (76% identical),
and it is likely that identical regions between the two
isoforms carry out equivalent functions. What has not
been explored in detail is the 24% of the sequence that
is not identical – it is here that differences in isoform
function might be found. In particular, we sought to
identify conserved differences between the isoforms in
these divergent regions as these might provide the key
to understand the differences in isoform function. We
set out to look at differences in ligand specificity, affin-
ity tuning, tertiary structure and the conservation of
post-translational modification sites (PTMs). Comparison of the talin1 and talin2 domains Due to the high homology between talin1 and talin2,
we used Modeller [110] to generate structural models
of the talin2 domains using the talin1 structures as
templates. Validation of the conserved hydrophobic
cores
of
these
domains
and
comparison
of
the 2116 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies R. E. Gough and B. T. Goult Tale of two talins modelled F2F3 region of talin2 with the known struc-
ture [52] confirmed the reliability of this modelling
approach. The domain boundaries of talin1 and talin2
are shown in Fig. 2. From this bioinformatics analysis,
we have designed and validated expression constructs
to express and purify each of the talin2 domains (de-
posited in Addgene www.addgene.org/ben_goult). We
have recently solved the structures of a number of
these talin2 rod domains and it is striking how struc-
turally
similar
they
are
to
the
equivalent
talin1
domains (our unpublished data). similar binding constants (Kd) of these domains for
their respective ligands, that is, the KANK 1 and
KANK 2 binding site on R7 and the RIAM- and
DLC1-binding site on R8. These proteins bind in the
same manner and with the same affinity to both iso-
forms ([16] and our own unpublished data). In contrast, the R5 domain, for which ligands have
yet to be identified, has a highly conserved surface in
talin2 with the characteristics of an LD-motif-binding
domain, but this surface is markedly different in tal-
in1. Based on this analysis, we suggest that regions of
divergence between isoforms that are well conserved
within each isoform likely encode regions that define
the subtle differences in isoform functionalities. Talin2 in disease and development This is reflected by the Interestingly, the Tln2 gene also includes a highly
conserved microRNA, miR-190, situated in intron 51
[10], which has been implicated as a modulator in mul-
tiple
signalling
pathways. Moreover,
talin2
has
appeared in a number of screens as a protein regulated
by microRNAs whose expression is perturbed in cancer
[53,113]. Thus, the humanised antibody trastuzumab,
which recognises the extracellular domain of HER2,
upregulates miR-194 expression in two HER2-positive
breast cancer cell lines [113], and miR-194 suppresses
cell migration reportedly via downregulation of talin2. Talin2 is also downregulated by miR-132, but miR-132
expression is itself suppressed by promoter methylation 2117 FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies Tale of two talins R. E. Gough and B. T. Goult PTMs. To explore this, we took all reported phosphory-
lation and acetylation sites for talin1 and found that
the majority were conserved in talin2 (Table 1). Mass
spectrometric
analysis
of
calyculin-treated
platelets
identified numerous talin1 phosphorylation sites, with
the three most abundant sites being T144 and T150 in
the F1-loop (phosphorylation appears to negatively reg-
ulate integrin activation [58,115]) and S446 [115] in the
linker between the head and rod domains. Phosphoryla-
tion of S446 is believed to be important in the regulation
of the calpain cleavage between the head and the rod
domain and is important for regulating FA turnover
[71,117], a process that has recently been shown to be
essential for adhesion development and rigidity sensing
[118]. In addition, S425, which is also in the linker, is in prostate cancer cells. This correlates with a worse
prognosis, and the authors speculate that elevated tal-
in2 levels may suppress cell death and increase metasta-
sis [114]. Talin2 upregulation has also been implicated
in breast cancer tumorigenesis and metastasis [33,53]
driving more aggressive cell invasion. Table 1. Post-translational modifications in talin1 and talin2. Summary of the identified talin phosphorylation [115,116], acetylation [128],
arginylation [121], glycosylation [123] and methylation sites [122]. For each PTM, the modified residue, the domain it is located, and the
conservation between isoforms are shown. Residue numbering is for mouse talin1 and talin2. FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies 2118
FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies Post-translational modification of
talins phosphorylated by CDK5, and this phosphorylation
has been shown to enhance talin activity and increase
integrin activation [119]. Calpain cleavage of talin is a permanent PTM, and
three cleavage sites have been identified in talin1. The
best characterised is that within the linker between the
talin head and rod (between residues Q433 and Q434)
[73], but there is a second site immediately prior to the
DD (between residues K2493 and M2494) [71]. Both
of these cleavage sites are present in talin2 [71]. Cal-
pain cleavage of the neck exposes a recognition site for
the E3 ligase SMURF1 that leads to ubiquitination of
the liberated talin1 head [120]. A third force-dependent
calpain cleavage site in the talin1 rod has also been
identified [121]. This cleavage occurs between residue
P1902 and A1903 (a site which is normally buried in
the folded R10 domain) and is likely to only be acces-
sible when talin is under force. This cleavage appears
to be regulated by arginylation (a PTM that only
occurs on the N-term residue of proteins [121]). It is
not yet known whether this force-dependent cleavage
site is also present in talin2, but the region is well
conserved between both isoforms. Many different adhesive structures form on talins
which signal in a highly reproducible manner. Precisely
how talins’ mechanosignalling capabilities are inte-
grated with the more classical signalling pathways to
give rise to these robust metastable cellular responses
that facilitate all our cellular processes remain to be
determined. The signalling pathways that regulate talin
function, localisation, post-translational modifications,
etc. coupled with the forces that define the conforma-
tional status of its rod domains, which cumulatively
lead to the correct cellular responses are still poorly
understood. For talin to generate robust, reproducible
signalling responses and specialised adhesive structures
in response to such diverse, multiple inputs suggest
that there must be a code underpinning talins mechan-
otransductive response. Deciphering this ‘talin code’ is
the next major challenge. Finally, a number of additional PTMs have also been
identified in talin1. For instance, the affinity of the tal-
in1:actin connection is controlled via the methyltrans-
ferase Ezh2 which methylates talin at lysine K2454 in
ABS3 [122]. This PTM site is completely conserved in
talin2. Talin1 is also modified by glycosylation [123] at
sites in R8 and R10. Post-translational modification of
talins Talin has been shown to be regulated by post-transla-
tional modification and the phosphorylation sites in pla-
telet talin1 have been mapped [115]. Proteomics studies
on the ‘adhesome’ also show phosphorylation of talin in
adhesions [116], but there is much less data on talin2 Talin1 phosphorylation
site
Domain of talin1
Site conserved in talin2
Talin1 phosphorylation site
Domain of talin1
Site conserved in talin2
S5
F0
Yes
S677
R2
No
Y26
F0
Yes
S729
R2
Yes
Y70
F0
Yes
S815
R3
Yes
T78
F0
No
S940
R4
No
T96
F1
No
S979/S981
R4
No/Yes
T114
F1
Yes
S1021
R4
Yes
Y127
F1
Yes
Y1116
R5
Yes
S128
F1
Yes
T1142
R5
Yes
T144
F1
Yes
S1201
R5
No
T150
F1
Yes
S1225
R6
No
T167
F1
Yes
T1263
R6
No
T190
F1
Yes
S1323
R6
Yes
S311
F3
Yes
S1508
R8
No
S405
LINKER
Yes
S1641
R7
Yes
S425
LINKER
Yes
S1684
R9
Yes
S429/T430
LINKER
Yes
S1849
R10
No
Y436
LINKER
No
T1855
R10
Yes
S446
LINKER
Yes
S1878
R10
No
S455/S458
LINKER
Yes
S2040
R11
No
S467
LINKER
Yes
S2127
R11
Yes
S620
R1
Yes
S2338
R12
No
Y2530
DD
Yes
S2535
DD
No
Talin2 phosphorylation
site
Domain of talin2
Site conserved in talin1
Talin1 PTM
Domain of talin1
Site conserved in talin2
Y1665
R9
No
K1544 (acetylation)
R8
Yes
T1843
R10
No
K2031 (acetylation)
R11
Yes
K2115 (acetylation)
R11
Yes
A1903 (arginylation)
R10
Yes
T1487 (glycosylation)
R8
No
T1890 (glycosylation)
R10
Yes
K2454 (methylation)
R13
Yes FEBS Letters 592 (2018) 2108–2125 ª 2018 Federation of European Biochemical Societies 2118 R. E. Gough and B. T. Goult Tale of two talins other cell types. Recent work has shown that the two
talins provide different mechanical linkages in cells
[34], and the ability to measure talins mechanical
response both at the single molecule level [21], in cells
using genetically encoded tension sensors [34,85,125],
and in silico with force extension molecular dynamics
simulations [126] provide the tools to understand how
talin signalling varies with mechanical forces. Detailed
structural and biochemical characterisation of talin
interactions
is
enabling
targeted
mutations
to
be
designed that specifically disrupt individual talin func-
tions, which in conjunction with the aforementioned
technical advances should enable the study of talin
function in unprecedented detail. It is likely that fur-
ther novel talin-mediated cell functions will be identi-
fied as additional binding partners of the two talins are
discovered. Post-translational modification of
talins Interestingly, the glycosylation sites
are not conserved between talin1 and talin2, suggesting
that if talin2 is glycosylated, then it is at different sites
and linked to different functions. Acknowledgements Gene duplication is often viewed as an evolutionarily
advantageous process, with duplicated genes giving rise
to two proteins that can acquire distinct or completely
new functions (subfunctionalisation or neofunctionali-
sation). Gene duplication may also allow more complex
patterns of gene expression in different cell types and
tissues [124]. Furthermore, differences in PTM sites as
reported here for the two talins may enable new modes
of regulation at the protein level. Although both talin
isoforms have maintained their ancestral properties
relating to cell adhesion, it seems likely that the two
isoforms have undergone some neofunctionalisation to
generate nuanced, isoform-specific regulation of sig-
nalling in cell adhesion. We imagine a scenario whereby
talin2 plays a central role in some tissues, such as car-
diac muscle, the brain and kidney, but then also a more
global role in fine-tuning the adhesive response in many We thank David Critchley for stimulating discussions
and critical reading of the manuscript and for all his
support over the years. We also thank Alex Carisey
(Baylor College of Medicine) for the fantastic artwork. We thank the members of the Goult laboratory for
discussions and reading of the manuscript. BTG is
funded by BBSRC grant (BB/N007336/1) and HFSP
grant (RGP00001/2016). REG is funded by a Univer-
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The influence of parenting, self-efficacy, and entrepreneurial interest toward the learning motivation of creative products and entrepreneurship of culinary management vocational school students
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*Corresponding Author:
Naning Yulianti Antonia
naningyulianti.2018@student.uny.ac.id
Department of Technology and Vocational
Education, Graduate School, Universitas Negeri
Yogyakarta
Jl. Colombo No. 1, Karangmalang, Depok,
Sleman, Yogyakarta 55281, Indonesia THE INFLUENCE OF PARENTING, SELF-EFFICACY, AND
ENTREPRENEURIAL INTEREST TOWARD THE LEARNING
MOTIVATION OF CREATIVE PRODUCTS AND ENTREPRENEURSHIP
OF CULINARY MANAGEMENT VOCATIONAL SCHOOL STUDENTS Naning Yulianti Antonia1*, Badraningsih Lastariwati1
1Universitas Negeri Yogyakarta
Jl. Colombo No. 1, Karangmalang, Depok, Sleman, Yogyakarta 55281, Indonesia INTRODUCTION Adolescence is a transitional period from childhood to adulthood, that is, children do not
want to be treated as children, but physically they cannot be said as adults. Adolescence may be a
defenseless period, particularly the issue of juvenile delinquency. The causes of juvenile delinquen-
cy include heredity, identity problems, community influence and family experiences. As an illustra-
tion within the case of a young battle in Magelang, approximately 40 vocational students were in-
cluded in a brawl (Fitriana, 2019). This case should be a family concern, how to reduce juvenile de-
linquency at this time with legitimate care. q
y
g
The role of parents is crucial. Based on Law of the Republic of Indonesia No. 23 of 2002
article 26 concerning Child Protection, that parents have the obligation and responsibility to care
for, nurture and educate, protect children, foster children according to their abilities, talents and in-
terests and prevent child marriages. The role of parents can be well-applied if the parental care has
a good influence too, especially in the children’s development. One of the problems which is cur-
rently being debated in some research that focuses on family matters is how to attain optimal par-
enting in educating children. One of the positive supports that can be given by the family is to mo-
tivate the children, particularly to provide motivation in learning. According to Oyeku et al. (2014) in a research article about the influence of family, moti-
vation and personality towards entrepreneurial interest through self-efficacy reveal that the variable
that can foster motivation or interest is self-efficacy (Kurniawan, et al., 2016, p. 100). Self-efficacy
is a person’s belief to accomplish the task. Self-efficacy is influenced by the conceptual factors
such as education and past experiences. The level of efficacy will affect a person’s enthusiasm; the
low level of efficacy considers someone incapable to do something around him whereas someone
with high level of efficacy in any situation will try hard to overcome the challenges (Ghufron &
Risnawita, 2012, p. 76). The level of student’s success depends on the enthusiasm and motivation
(self-efficacy) within the students themselves. This will have a huge impact in learning process,
when the learning motivation is high then the learning outcome will be high too, however if the
learning motivation is low, the learning outcome will be low. INTRODUCTION Self-efficacy has an effect on entre-
preneurial interest, research revealed by Farida and Nurkhin (2016, p. 273). The interest arises when someone has the feeling of like and dislike to do the accomplished
task. Interest is not carried from birth, interest is influenced by physical factors, psychological fac-
tors and environmental factors. The interest to be researched is the level of entrepreneurial interest
of the XII graders vocational school in Magelang. The way to foster entrepreneurial interest is by
developing within them an interest in entrepreneurship. With an interest in them, students are en-
couraged to learn knowledge related to entrepreneurship for the better. It is expected that students
will have self-awareness and enthusiasm to foster entrepreneurial attitudes. The entrepreneurial interest from the respondents, especially students of class XII Catering
Services Department in Magelang, which is one of the tourist cities that is quite crowded with tour-
ists. Business opportunities in the culinary field are the main attraction, so this is a very good op-
portunity to develop. This research is expected to draw a clear picture of how to see the influence
of parenting, self-efficacy or students' self-confidence as well as their inner interests so that they
can have the learning motivation, especially entrepreneurial learning and creative products of XII
graders of Catering Service Department in Magelang. Abstract This study aims to reveal the effects of (1) parenting, (2) self-efficacy, (3) entrepreneurial interest,
and (4) parenting, self-efficacy, and interest in entrepreneurship simultaneously on the motivation
to learn creative products and entrepreneurship in class XII vocational school of Catering Services
Department in Magelang. This research is ex-post facto research. The research population was
201 students of the Catering Services Department from two vocational schools in Magelang. The
sample consisted of 127 students who were taken using the proportional random sampling tech-
nique. The data analysis technique used descriptive analysis and regression analysis. The results
showed that: Parenting, self-efficacy, and interest in entrepreneurship were able to predict learn-
ing motivation with an effective contribution to parenting (5.15%, high category); self-efficacy
(3.63, low category); interest in entrepreneurship (32.80%, very high category). The coefficient of
determination (R2) is 0.416 (41.6%), which means that 41.6% of students' motivation to learn cre-
ative products and entrepreneurship in class XII of the Catering Services Department are influ-
enced by parenting, self-efficacy, and interest in entrepreneurship, while the remaining 58.4% is
influenced by other variables that are not examined in this study. K
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lf ffi Keywords: interest in entrepreneurship, learning motivation, parenting, self-efficacy How to cite: Antonia, N., & Lastariwati, B. (2020). The influence of parenting, self-
efficacy, and entrepreneurial interest toward the learning motivation of creative products
and entrepreneurship of culinary management vocational school students. Jurnal
Pendidikan Vokasi, 10(3), 315-323. doi:https://doi.org/10.21831/jpv.v10i3.32428 *Corresponding Author:
Naning Yulianti Antonia
naningyulianti.2018@student.uny.ac.id
Department of Technology and Vocational
Education, Graduate School, Universitas Negeri
Yogyakarta
Jl. Colombo No. 1, Karangmalang, Depok,
Sleman, Yogyakarta 55281, Indonesia This is an open access article under the CC-BY-SA license. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Self Efficacy The definition of self-efficacy according to Bandura (1997, p. 3) is a belief about the ability
to organize and take a series of actions needed to achieve their beliefs. Meanwhile, according to
Woolfolk (2007) in Marini and Hamidah (2014), self-efficacy is a person's assessment of himself
or his level of confidence regarding how much his or her ability to do a certain task to achieve cer-
tain results. There are four sources of self-efficacy: (1) mastery experience, it will foster self-effi-
cacy in oneself, while failure can be overcome with certain efforts that can strengthen self-motiva-
tion if someone can find meaning from the experience. (2) Vicarious experience is the experience
of the success of others with comparable abilities in doing tasks that will increase individual self-
efficacy. (3) Verbal persuasion is more directed by providing advice and guidance so it can in-
crease confidence in one's abilities. (4) Physiological state is a physical tension in a stressful situ-
ation, seen as a sign of incapacity due to weakening of individual work. In addition, self-efficacy is
divided into three dimensions: (1) level, related to the level of difficulty experienced by individuals
in completing tasks, with different ability limits for each individual; (2) strength, related to strength
or belief about the abilities that people have - weak expectations are easily shaken by unsupportive
experiences, while steady expectations will encourage individuals to stay in higher dimensions; (3)
generality, related to the broad field of behavior where individuals feel confident in their abilities. 317 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 317 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 also popularized by Marcoby and Martin (1983) in Sutanto and Andriyani (2020, p. 108), includ-
ing: (a) authoritarian care, where the child must obey all the rules set by the parents. Parents will
apply high discipline to their children. (b) Permissive care, where parents give complete freedom to
their children, without giving any rules to family members, including children. (c) Democratic/
authoritative care, where parents act as the determinants of rules, but children are still given the
opportunity to know and understand the rules that have been made by parents. (d) Neglect, that is,
parenting style that is too indulgent, where parents are not very involved in the child's life. Pradani
(2017) in Sutanto and Andriyani (2020, p. 15) claims that parents should apply the positive parent-
ing style. Parental care is said to be positive when parents can think positively about their children. This type of parenting will foster self-concept and positive thinking in children. Negative parenting
style is parenting style that often performs negative actions in parenting, for example hitting, ignor-
ing, humiliating oneself, being unfair, and so on. The characteristics of positive parenting style ac-
cording to Faber (1980), Hansen (1982), Jams (1985), and Gordon (2000) in Sunarty (2015, pp. 17-
24) are reasonable parents, encouraging parents, con-sistent parents and responsible parents. Ac-
cording to González et al. (2016), with positive parenting, parents show the better control as shown
by quarrels, fights, and reprimand against children decreased significantly. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Parenting Parenting is a style of taking care applied by parents in various ways and understandings. The definition of parenting based on Indonesian Dictionary is the process, method, action, and par-
enting style. Meanwhile, Masud Hoghughi, Professor of the Faculty of Psychology, University of
Hull America in Nefrijanti (2018) assumes that parenting is a relationship between parents and
children that is multi-dimensional and can grow steadily and it also includes a variety of activities
with various goals, namely, the child can develop optimally and survive well, therefore, parenting
style includes physical care, emotional care and social care. y
p y
According to Baumrind (1971) in Santrock (2007, p. 167), parenting has four styles formed
from two dimensions: dimensions of responsiveness and demandingness. This parenting style is Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Entrepreneurial Interest The definition of interest, according to Slameto in Marini and Hamidah (2014, p. 201), is
that there is a feeling of preference and interest in something or an activity, without being asked. According to Winkel and Hastuti (2004), it is a rather sedentary tendency for someone to feel at-
tracted to a certain field and feel happy to be involved in various activities in that field. Daryanto
(2012, p. 7) believes that entrepreneurship is an effort to create added value by combining sources
through new and different ways to win the competition. Meanwhile, according to Zimmerereal
(2008) in Wijaya (2017, p. 21), an entrepreneur is someone who creates a new business by facing
uncertainty and risk to achieve profits and business growth by identifying significant opportunities
and using the required resources. Entrepreneurial interest is the desire or interest and ability of students in entrepreneurship,
especially the students of XII grade of Catering Services Department in Magelang. According to
Meredith in Marini and Hamidah (2014), there are six characteristics of entrepreneurship: having
self-confidence, being task-oriented, courageous to take risks, socializing with others, innovative
and creative and future-oriented. These interests will grow when there is motivation. Learning mo-
tivation needs to be cultivated as an effort to support learning, especially learning entrepreneurship
and creative products. Learning motivation is students’ willingness in the learning process, espe-
cially in entrepreneurship lessons and creative products. Learning creative products and entrepre- 318 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 neurship is a learning process in creating new ideas as a form of business in a new work that is oc-
cupied. There are many factors that can influence the interest in entrepreneurship, one of which is
developing entrepreneurship education and teachers as role models for building interest in entre-
preneurship, as indicated by a research conducted by San-Martín et al. (2019). This is in line with
research disclosed by Iwu et al. (2019) which also explains that entrepreneurship education can
influence entrepreneurial interest in South Africa. Entrepreneurial interest is a very important part
of building learning motivation which is also related to entrepreneurship education. Research Setting This research was conducted on February 2020 till April 2020. The research subjects were
XII graders of vocational school of Catering Service Department in Magelang, namely, Sekolah
Menengah Kejuruan (SMK) Negeri 3 Magelang and SMK Pius X Magelang. As shown in Table 1, the research population was 201 students from grade XII of Catering
Service Department in Magelang. The sample in this study was 127 students collected using pro-
portional random sampling technique, where each school with a diverse number of respondents was
tested based on the extent of the number of respondents in each of these schools (Sugiyono, 2017,
p. 120). The determination of the number of samples was based on Isaac and Michael's table at an
error rate of 5%. Table 1. Population Data and Research Sampling
Schools’ Name
Population
Sample
SMK N 3 MGL
108
68
SMK Pius X MGL
93
59
Total
201
127 This study used questionnaires and document review as the data collection techniques. The
questionnaire with the consideration that the number of respondents was quite large and spread
across several schools in Magelang was used. The validity test used content validation and con-
struct validity. Reliability test was using Alpha Cronbach formula. The instrument can be said to be
reliable if the Cronbach Alpha coefficient is greater than 0.70 (Mardapi, 2008, p. 125). The data analysis technique used descriptive analysis and regression analysis. Descriptive
analysis provided an overview of the data seen from the mean, median, mode, standard deviation,
maximum, minimum, range using SPSS 20.0 for windows software to be analyzed. Entrepreneurial Interest Entrepreneurial interest is generally influenced by (1) the ideals and aspirations of students,
in this case the aspirations will strengthen motivation; (2) abilities coupled with student desires; (3)
the student's condition is good (physically and mentally healthy) so that it is easy to focus attention
in learning; and (4) students' environmental conditions will increase children's interest or enthusi-
asm in learning. Therefore, this study aims to reveal the effects of: (1) parenting, (2) self-efficacy,
(3) interest in entrepreneurship, and (4) parenting, self-efficacy and interest in entrepreneurship si-
multaneously on the motivation to learn creative products and entrepreneurship in class XII voca-
tional school of Catering Services Department in Magelang. RESEARCH METHOD This study is an ex-post facto research, since it only reveals the symptoms that occur as they
were and reveals the factors that influence the independent variable on the dependent variable. This
research is a descriptive correlational study according to the problem with a quantitative approach. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 319 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 gory, and six students (4.7%) had a very low category. In term of the parenting variable, the mean
(105.59) result of the descriptive analysis is compared to the category distribution, so the mean par-
enting lies in the high value range. Thus, it is stated that the respondents’ parenting is in the high
category. The results of data analysis illustrated by the frequency histogram is shown in Figure 1. Figure 1. Parenting Frequency Histograms of XII Grade of Catering Service Department of
Vocational School Students in Magelang Figure 1. Parenting Frequency Histograms of XII Grade of Catering Service Department of
Vocational School Students in Magelang The high parenting tendency implies that parenting style has provided positive motivation in
learning. This support includes examples of exemplary action, being consistent with responsibility
and obeying common provisions, positive habituation, and building communication with children
without violence. This parenting has a good impact on children, so they are motivated to learn well. This research is supported by previous research conducted by Delima and Usman (2019). This
study confirms that there is a positive and significant relationship between interest in learning, pa-
rental attention and learning motivation towards student achievement in class XI SMA in Jakarta. It
is also in line with the research revealed by Kaukab (2016, p. 72) that parental involvement posi-
tively increases children's learning motivation for the better. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Parenting The results of the parenting analysis show that 20 students (15.7%) were in the very high
category, 65 students (51.2%) were in the high category, 36 students (28.3%) were in the low cate- Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Entrepreneurial Interest The results of data analysis on entrepreneurial interest showed that there were 68 students
(53.5%) in the very high category; as many as 44 students (34.6%) in the high category; as many as
13 students (10.2%) in the low category and two students (1.6%) in the very low category. The
mean (51.00) result of the descriptive analysis compared to the distribution of categories lies at a
very high value. Thus, it can be stated that the interest in entrepreneurship among the respondents
is in the very high category. The results of data analysis illustrated by the frequency histogram are
presented in Figure 3. Figure 3. Histogram of Frequency of Entrepreneurial Interest in XII Grade of Catering Service
Department of Vocational School Students in Magelang Figure 3. Histogram of Frequency of Entrepreneurial Interest in XII Grade of Catering Service
Department of Vocational School Students in Magelang The tendency of very high interest in entrepreneurship from the respondents has very high
curiosity about entrepreneurship and willingness to be entrepreneurial. This study supports the
previous research conducted by Widjaja (2019). It is also in line with research conducted by
Nurmaliza et al. (2018, p. 43) which confirmed that entrepreneurship education and the family en-
vironment have a positive and significant influence on entrepreneurial interest. The influence of
interest in entrepreneurship in this study is learning motivation, where in addition to learning moti-
vation, entrepreneurship education also has a role in developing interest. 320 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 320 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 that exist. In addition, Bandura (1997) argues that the characteristics of individuals who have high
self-efficacy are the individuals feel confident that they feel able to deal effectively with the events
or situations they are facing, are diligent in completing tasks, believe in their own abilities and per-
ceive difficulties as a challenge. The results of this study reveal that the average student still did not
have the confidence to motivate themselves, especially in terms of learning. This study supports the
previous research conducted by Robih et al. (2017) that self-efficacy significantly affects learning
motivation. It is also in line with Kuo et al. (2017, p. 131) who assert that “students with higher
levels of research self-efficacy also tend to have higher research productivity”. that exist. In addition, Bandura (1997) argues that the characteristics of individuals who have high
self-efficacy are the individuals feel confident that they feel able to deal effectively with the events
or situations they are facing, are diligent in completing tasks, believe in their own abilities and per-
ceive difficulties as a challenge. The results of this study reveal that the average student still did not
have the confidence to motivate themselves, especially in terms of learning. This study supports the
previous research conducted by Robih et al. (2017) that self-efficacy significantly affects learning
motivation. It is also in line with Kuo et al. (2017, p. 131) who assert that “students with higher
levels of research self-efficacy also tend to have higher research productivity”. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Self Efficacy The results of the self-efficacy data analysis show that there were 18 students (14.2%) in the
very high category, 37 students (29.1%) in the high category, 46 students (36.2%) in the low cate-
gory and 26 students (20.5%) in very low category. The mean value of 46.51 results of descriptive
analysis is compared with the distribution of categories, so self-efficacy lies in the range of values
in the low category. Thus, it is stated that the self-efficacy of the respondents is included in the low
category. The results of data analysis illustrated by the frequency histogram presented in Figure 2. Figure 2. Histogram of Frequency of Self Efficacy of XII Grade of Catering Service Department of
Vocational School Students in Magelang Figure 2. Histogram of Frequency of Self Efficacy of XII Grade of Catering Service Department of
Vocational School Students in Magelang The tendency of self-efficacy from the respondents has a low category, meaning that the be-
lief students have is less confident in their ability to motivate themselves, especially in term of
learning, confidence in self-strength and self-ability to motivate learning with all the challenges Results of Hypotheses The results of the first hypothesis test with simple regression analysis showed that the par-
enting variable had an effect on the motivation to learn entrepreneurship and the creative products
of the respondents. The results of data analysis show that the significance value is as large as 0.000
<0.05 at the 5% significance level. The results of the analysis also show that the correlation value
(r) is positive, which is 0.403, so it can be stated that the parenting variable has a positive effect on
entrepreneurial learning motivation and creative products of the respondents. This is in line with
the research conducted by Delima and Usman (2019). The results of this study confirm that there is
a positive and significant influence between classroom learning, parental attention, learning moti-
vation, and learning outside the classroom using a mentor at SMK Jakarta, the effect is 82.7%. Par-
enting in the family is a key element in motivating children's learning as well as shaping their per-
sonality traits. Motivation to learn will grow along with the parenting patterns that are applied. Po-
sitive parenting according to Sutanto and Adriyani (2020, p. 23) is care that is based on love, mutu-
al respect, and stimulates children's growth and development. One example of giving rewards to
children who get good grades, this is one way to motivate children to learn. g
g
g
y
The results of the second hypothesis with simple regression analysis indicate that the self-
efficacy variable has a positive effect on the motivation to learn entrepreneurship and the creative
products of the respondents. It can be read from the results of data analysis that the significance
value is 0.000 <0.05 at the 5% significance level. The results of the analysis also show that the val-
ue of the correlation coefficient (r) is positive, which is 0.454, so it can be stated that the self-effi-
cacy variable has a positive effect on the learning motivation of the respondents. Someone who has
high self-efficacy will also have high learning motivation. A research by Robih et al. (2017) show
that self-efficacy affects the motivation to learn office automation skills for class X administrative
competence at private vocational schools in North Surabaya. Self-efficacy is a belief in yourself,
when you do not have confidence in yourself, you cannot achieve your existing desires. Learning Motivation The results of the data analysis on entrepreneurial learning motivation and creative products
showed that there were two students (1.6%) in the very high category; as many as 17 students
(13.4%) in the high category; as many as 90 students (70.9%) in the low category and 18 students
(14.2%) in the very low category. The mean value (52.12) of the results of the descriptive analysis
compared to the distribution of categories lies in the low value range. Thus, it can be stated that the
motivation to learn entrepreneurship and the creative products of respondents is in the low cate-
gory. The data analysis is illustrated through the frequency histogram, presented in Figure 4. The low tendency of motivation to learn entrepreneurship and creative products from the re-
spondents means that they are lacking, in terms of students' lack of knowledge, less desire and curi-
osity, encouragement of the need and persistence in learning entrepreneurship and products crea-
tive is also lacking. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 321 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428
Figure 4. Histogram of Frequency of Entrepreneurship Learning Motivation and Creative Products
of XII Grade of Catering Service Department of Vocational School Students in Magelang 321 – Naning Yulianti Antonia & Badraningsih Lastariwati 321 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 321 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 Figure 4. Histogram of Frequency of Entrepreneurship Learning Motivation and Creative Products
of XII Grade of Catering Service Department of Vocational School Students in Magelang Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 CONCLUSION Based on the results of data analysis and the discussion previously presented, there are four
conclusions in this study. Partially that: (1) parenting has an effective contribution of 5.18% in the
high category, (2) self-efficacy has an effective contribution of 36.22% in the low category, (3) in-
terest in entrepreneurship has an effective contribution of 53.54 % is in the very high category, (4)
motivation to learn entrepreneurship and creative products has an effective contribution of 70.87%
in the low category. g
y
Based on the results of the hypothesis test, it can be concluded as follows: there is a positive
and significant influence of parenting on entrepreneurial learning motivation and creative products,
with the correlation coefficient (rx1, y) is 0.403 and p <0.05; There is a positive and significant ef-
fect of self-efficacy on motivation to learn entrepreneurship and creative products, with a correla-
tion coefficient (rx2, y) of 0.454 and p <0.05; there is a positive and significant effect of interest in
entrepreneurship, there is motivation to learn entrepreneurship and creative products, with the cor-
relation coefficient (rx3, y) is 0.626 and p <0.05; and there is a positive and significant effect to-
gether from parenting (parenting), self-efficacy, interest in entrepreneurship, there is entrepreneur-
ial learning motivation and creative products with a correlation coefficient value (Rx1, x2, x3, y) is
0.645. The strong influence of parenting variables, self-efficacy and interest in entrepreneurship is
indicated by the magnitude of the R2 value, namely 0.416. Thus, it can be said that the effective
contribution of the three independent variables to student learning motivation is 41.58%. 322 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 The results of the multiple regression analysis show that parenting (parenting), self-efficacy,
and interest in entrepreneurship have a positive effect together on entrepreneurial learning motiva-
tion and creative products of the respondents. This can be read from the results of the F test which
shows a value of 29.183 compared to the value of 2.68 at the 5% significance level. The results of
the analysis also show that the value of the correlation coefficient (r) is positive, which is equal to
0.416, thus, it can be stated that parenting variables, self-efficacy and interest in entrepreneurship
have a positive effect on the learning motivation of the respondents. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 Results of Hypotheses The results of the third hypothesis test with simple regression analysis show that the variable
interest in entrepreneurship has a positive effect on the motivation to learn entrepreneurship and the
creative products of the respondents. It can be read from the results of data analysis that the signi-
ficance value is 0.000 <0.05 at the 5% significance level. The results of the analysis also show that
the correlation coefficient (r) is positive, which is 0.626, so it can be stated that the Entrepreneurial
Interest variable has a positive effect on entrepreneurial learning motivation and creative products
of the respondents. Entrepreneurial interest grows because of inner motivation, Hurlock (1978) in
Farida and Nurkhin (2016, p. 277) emphasizes that interest is a strong motivation to learn. With the
interest in entrepreneurship, students are able to work independently by creating their own jobs. According to Widjaja (2019, p. 46), there is a significant influence between learning motivation
and interest in entrepreneurship in XYZ school, West Jakarta. It is also in line with a research con-
ducted by Hutagalung et al. (2017, p. 345) showing that partially, the variables of entrepreneurship
education and family environment (care) have a positive value and have a significant impact on
students' entrepreneurial motivation. Copyright © 2020, Jurnal Pendidikan Vokasi, 10(3), 2020
ISSN (print) 2088-2866 | ISSN (online) 2476-9401 323 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 323 – Naning Yulianti Antonia & Badraningsih Lastariwati
https://doi.org/10.21831/jpv.v10i3.32428 Iwu, C. G., Opute, P. A., Nchu, R., Eresia-Eke, C., Tengeh, R. K., Jaiyeoba, O., & Aliyu, O. A. (2019). Entrepreneurship education, curriculum and lecturer-competency as antecedents of
student entrepreneurial intention. The International Journal of Management Education,
19(1), 100295. https://doi.org/10.1016/j.ijme.2019.03.007 Kaukab, S. R. (2016). The impact of parent/family involvement on student learning outcomes. International Journal of Research – Granthaalayah, 4(10), 72-81. https://doi.org/10.5281/
zenodo.164925 Kuo, P. B., Woo, H., & Bang, N. M. (2017). Advisory relationship as a moderator between
research self-efficacy, motivation, and productivity among counselor education doctoral
students. Counselor Education and Supervision, 56(2), 130-144. https://doi.org/10.1002/
ceas.12067 Kurniawan, A., Khafid, M., & Pujiati, A. (2016). Pengaruh lingkungan keluarga, motivasi, dan
kepribadian terhadap minat wirausaha melalui self efficacy. Journal of Economic Education,
5(1), 100-109. Retrieved from https://journal.unnes.ac.id/sju/index.php/jeec/article/view/
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tawuran-yang-tewaskan-1-siswa-3-pelajar-smk-diamankan-polisi?page=all Ghufron, M. N., & Risnawita, R. (2012). Teori-teori psikologi. Ar-Ruzz Media. González, R. A. M., Ruiz, B. R., Blanco, L. Á., & Vázquez, C. B. (2016). Evidence in promoting
positive parenting through the Program-Guide to Develop Emotional Competences. Psychosocial Intervention, 25(2), 111-117. https://doi.org/10.1016/j.psi.2016.04.001 Hutagalung, B., Dalimunthe, D. M. J., Pambudi, R., Hutagalung, A. Q., & Muda, I. (2017). The
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Pustaka Baru Press. Widjaja, W. (2019). Pengaruh motivasi belajar terhadap minat berwirausaha siswa SMA XYZ di
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English
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The impact of housing payment problems on health status during economic recession: A comparative analysis of longitudinal EU SILC data of 27 European states, 2008–2010
|
SSM, population health
| 2,016
|
cc-by
| 8,982
|
Amy Clair a,n, Rachel Loopstra a, Aaron Reeves a, Martin McKee b, Danny Dorling c,
David Stuckler a a Department of Sociology, University of Oxford, Manor Road Building, Manor Road, Oxford OX1 3UQ, UK a Department of Sociology, University of Oxford, Manor Road Building, Manor Road, Oxford OX1 3UQ, UK
b European Observatory on Health Systems and Policies, London School of Hygiene and Tropical Medicine, 15-17 Tavistock Place, London WC1H 9SH, UK
c School of Geography and the Environment, University of Oxford, Oxford, Oxford University Centre for the Environment, University of Oxford, South Parks
Road, Oxford OX1 3QY, UK SSM -Population Health journal homepage: www.elsevier.com/locate/ssmph a r t i c l e i n f o Taylor, & Todd, 2008; Thomson & Thomas, 2015; Thomson, Tho-
mas, Sellstrom, & Petticrew, 2013; Winslow, 1937) and the large
number of families who have fallen into rent or mortgage arrears
during the recession, estimated to be around 3.5 million people
across Europe. Contents lists available at ScienceDirect Contents lists available at ScienceDirect n Corresponding author. Tel.: þ44 1865 274443.
E-mail address: amy.clair@sociology.ox.ac.uk (A. Clair). a r t i c l e i n f o Although the recent Great Recession had its origins in the housing sector, the short-term health impact of
the housing crisis is not well understood. We used longitudinal data to evaluate the impact of housing
payment problems on health status among home-owners and renters in 27 European states. Multi-level
and fixed-effects models were applied to a retrospective cohort drawn from the EU Statistics on Income
and Living Conditions survey of employed persons, comprising those without housing arrears in the base
year 2008 and followed through to 2010 (n¼45,457 persons, 136,371 person-years). Multi-variate
models tested the impact of transitioning into housing payment arrears on self-reported health (0-worst
to 4-best), adjusting for confounders including age, sex, baseline health, and individual fixed effects. Transitioning into housing arrears was associated with a significant deterioration in the health of renters
(0.09 units, 95% CI 0.05 to 0.13), but not owners (0.00, 95% CI 0.05 to 0.06), after adjusting for
individual fixed effects. This effect was independent of and greater than the impact of job loss for the full
sample (0.05, 95% CI 0.002 to 0.09). The magnitude of this association varied across countries; the
largest adverse associations were observed for renters in Belgium, Austria, and Italy. There was no ob-
served protective association of differing categories of social protection or of the housing regulatory
structure for renters. Women aged 30 and over who rented appeared to have worse self-reported health
when transitioning into arrears than other groups. Renters also fared worse in those countries where
house prices were escalating. We therefore find that housing payment problems are a significant risk
factor for worse-self reported health in persons who are renting their homes. Future research is needed
to understand potential sources of health resilience among renters, especially at a time when housing
prices are rising in many European states. Article history:
Received 25 January 2016
Received in revised form
13 April 2016
Accepted 11 May 2016
Keywords:
Housing
Arrears
Multi-level modelling
Fixed effects
Tenure
Comparative Article history:
Received 25 January 2016
Received in revised form
13 April 2016
Accepted 11 May 2016 Keywords:
Housing
Arrears
Multi-level modelling
Fixed effects
Tenure
Comparative & 2016 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license
(http://creativecommons.org/licenses/by/4.0/). http://dx.doi.org/10.1016/j.ssmph.2016.05.006
2352-8273/& 2016 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). p
ors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/ The impact of housing payment problems on health status during
economic recession: A comparative analysis of longitudinal EU SILC
data of 27 European states, 2008–2010 r a,n, Rachel Loopstra a, Aaron Reeves a, Martin McKee b, Danny Dorling c,
kl
a Amy Clair a,n, Rachel Loopstra a, Aaron Reeves a, Martin McKee b, Danny Dorlin
David Stuckler a SSM -Population Health 2 (2016) 306–316 SSM -Population Health 2 (2016) 306–316 http://dx.doi.org/10.1016/j.ssmph.2016.05.006
2352-8273/& 2016 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). Table 1 A growing body of scholarship
indicates that people who experience housing insecurity, in-
dependent of other financial difficulties, experience declines in
mental health (Gili, Roca, Basu, McKee, & Stuckler, 2012; Keene,
Cowan & Baker, 2015; Meltzer, Bebbington, Brugha, Farrell & Jen-
kins, 2013; Meltzer et al., 2011; Nettleton & Burrows, 1998). In
Australia, analysis of the longitudinal HILDA dataset found that
those in lower income households who had moved into un-
affordable housing experienced a worsening in mental health
(Bentley, Baker, Mason, Subramanian, & Kavanagh, 2011), with
male renters faring worse (Bentley, Baker & Mason, 2012; Mason,
Baker, Blakely, & Bentley, 2013). Similarly, research comparing US
cities found that foreclosures were associated with greater rates of
hospital visits (Currie & Tekin, 2011). A qualitative study in five US
cities, which included 14 focus groups of low- and mid-income
homeowners facing foreclosure as well as foreclosure avoidance
professionals, identified housing payment difficulties as a major
trigger of psychological feelings of insecurity and worse mental
health (Libman, Fields, & Saegert, 2012). Evidence from the US also
suggests that foreclosure and eviction during the housing crisis
was associated with increased suicide rates (Fowler, Gladden, Vagi,
Barnes, & Frazier, 2015; Houle & Light, 2014). alongside the ‘hard’, physical impacts of the infrastructure itself,
such as damp, mould, and cold. A growing body of scholarship
indicates that people who experience housing insecurity, in-
dependent of other financial difficulties, experience declines in
mental health (Gili, Roca, Basu, McKee, & Stuckler, 2012; Keene,
Cowan & Baker, 2015; Meltzer, Bebbington, Brugha, Farrell & Jen-
kins, 2013; Meltzer et al., 2011; Nettleton & Burrows, 1998). In
Australia, analysis of the longitudinal HILDA dataset found that
those in lower income households who had moved into un-
affordable housing experienced a worsening in mental health
(Bentley, Baker, Mason, Subramanian, & Kavanagh, 2011), with
male renters faring worse (Bentley, Baker & Mason, 2012; Mason,
Baker, Blakely, & Bentley, 2013). Similarly, research comparing US
cities found that foreclosures were associated with greater rates of
hospital visits (Currie & Tekin, 2011). A qualitative study in five US
cities, which included 14 focus groups of low- and mid-income
homeowners facing foreclosure as well as foreclosure avoidance
professionals, identified housing payment difficulties as a major
trigger of psychological feelings of insecurity and worse mental
health (Libman, Fields, & Saegert, 2012). Table 1 Evidence from the US also
suggests that foreclosure and eviction during the housing crisis
was associated with increased suicide rates (Fowler, Gladden, Vagi,
Barnes, & Frazier, 2015; Houle & Light, 2014). Using the only source of longitudinal data from 27 European
nations which covers both housing and health data, we test the
hypotheses that transitioning into housing arrears worsens peo-
ple's health and that this effect is worse for renters than owners,
varies across countries and according to individual characteristics. Table 1 The health impacts of economic shocks such as job loss have
been shown to vary across countries dependent on social support
and protection, a stark example being the difference between
Iceland and Greece's responses and resulting health issues during
the Great Recession (Stuckler & Basu, 2013). Housing market
characteristics, such as prices and the ownership rate, vary con-
siderably across Europe, as do levels of social protection and in-
formal social support, such as social capital. Here we investigate
whether experiencing housing payment problems has different
consequences, including health impacts, depending on where a
person lives and what differences in social support may best ex-
plain this variation. Both owners and renters risk housing payment problems, with
potentially differing consequences for health. Renters are more
likely to experience homelessness as a result of arrears, whereas
those who have mortgages face the potential additional loss of
substantial accumulated capital. However, owners may also have
greater resilience, resulting from greater financial resources and
housing market options. To our knowledge, while it is very plau-
sible that renters may be more vulnerable than those with a
mortgage, few studies have sought to differentiate these groups
(Mason et al., 2013; Pollack, Griffin, & Lynch, 2010). The causes and consequences of transitioning into housing
payment arrears are similarly likely to vary according to individual
characteristics and life stages. Women, who often still take on the
bulk of caring duties, may experience a greater reduction in health
when facing payment problems than men, for example, although
Australian evidence suggests that men may fare worse and evi-
dence from Britain indicates men are more likely to seek medical
support (Nettleton & Burrows, 1998). Older people who fall into
arrears may have fewer options available to them, increasing the
risk of health consequences. In contrast, younger adults may have
fewer resources available, having had less time to accumulate
savings for example, but may have parental support. Notes: Data from Eurostat (n.d.). The average across the 27 countries was 0.7, as
such 13 countries are above average and 13 are below, with the Netherlands re-
porting the average number of arrears. alongside the ‘hard’, physical impacts of the infrastructure itself,
such as damp, mould, and cold. Introduction The recent Great Recession had its origins in the U.S. housing
market, as growing numbers of people fell into rent or mortgage
arrears. However, research on the health effects of the recent crisis
has focused primarily upon the consequences of job loss and, to a
lesser extent, on homelessness, with relatively little attention paid
to impact of housing payment problems (Burgard & Kalousova,
2015). This is perhaps surprising given the importance of housing
as a determinant of health (Britten, 1934; Muir Gray, 1978; Pevalin, Yet as shown in Table 1, not all countries fared equally during
the Great Recession. Some countries, such as Italy, saw a slight
decline in arrears, falling from 4.3% in 2008 to 4.2% in 2010, while
Greece (among others) experienced a substantial rise, from 5.5% to
10.2% during this same period. These marked differences create a
unique opportunity to investigate how housing payment problems
influence health. n Corresponding author. Tel.: þ44 1865 274443. E-mail address: amy.clair@sociology.ox.ac.uk (A. Clair). Housing arrears is one of the so-called ‘soft’ ways in which
housing influences health (Shaw, 2004), especially mental health, A. Clair et al. / SSM -Population Health 2 (2016) 306–316 307 Table 1
Prevalence of housing payment arrears in Europe, 2008 and 2010. 2008
2010
Change 2008–2010
Iceland
5.5
11.2
5.7
Greece
5.5
10.2
4.7
Slovakia
3.0
6.8
3.8
Latvia
3.2
5.8
2.6
Cyprus
3.4
5.6
2.2
Portugal
2.8
4.8
2.0
Spain
4.5
6.4
1.9
Hungary
3.8
5.6
1.8
Denmark
1.1
2.7
1.6
Estonia
1.1
2.7
1.6
Czech Republic
2.3
3.5
1.2
United Kingdom
3.7
4.8
1.1
Lithuania
0.5
1.3
0.8
Netherlands
2.4
3.1
0.7
EU 27
3.3
4.0
0.7
Sweden
1.7
2.3
0.6
Poland
0.6
1.0
0.4
France
5.8
6.1
0.3
Luxembourg
1.1
1.4
0.3
Finland
4.4
4.7
0.3
Bulgaria
1.5
1.7
0.2
Belgium
3.3
3.4
0.1
Austria
3.9
3.9
0.0
Romania
0.6
0.6
0.0
Italy
4.3
4.2
0.1
Norway
5.0
4.8
0.2
Slovenia
2.7
2.4
0.3
Malta
1.5
1.1
0.4
Notes: Data from Eurostat (n.d.). The average across the 27 countries was 0.7, as
such 13 countries are above average and 13 are below, with the Netherlands re-
porting the average number of arrears. outcomes by tenure have been identified in other nations includ-
ing Australia and the USA (Bentley et al., 2011; Bentley et al., 2012;
Mason et al., 2013), here we investigate whether the same is true
in Europe. Data sources We use the 2010 longitudinal EU-SILC dataset to construct a
retrospective cohort of employed persons who had no housing
payment arrears in 2008. Details of EU SILC have been described
elsewhere (Arora et al., 2015; Europa, 2015; Iacovou, Kaminska, &
Levy, 2012), but briefly it uses a four-year rotating panel sample
with 25% of the sample being replaced in each wave. The precise
methods of data collection vary across the (then) 27 Member
States, although standards are set centrally. Following previous
analyses of effects of the Great Recession (Gili et al., 2012; Reeves,
Basu, Meissner, McKee, & Stuckler, 2013; Reeves, McKee, & Stuck-
ler, 2015; Stuckler, Basu, Meissner, Fishback, & McKee, 2012;
Stuckler, Basu, Suhrcke, Coutts, & McKee, 2009), we selected the
2010 dataset and used 2008 as the baseline year as, empirically, it
corresponds with the onset of the recession and associated rise in
housing payment arrears across Europe. Selecting this 3 year
period means that our analysis can cover the period of the re-
cession while ensuring a decent longitudinal sample size as fewer Here we seek to expand this literature by investigating how
housing payment arrears are related to people's health in Europe,
and, based on previous findings, how this impact varies across
countries, tenures and individual characteristics. The relationship
between housing payment arrears and health will likely vary ac-
cording to housing tenure given the different circumstances of
renters and owners in many nations. Renters tend to have fewer
savings, including a lack of equity associated with home owner-
ship, which may render housing hardship more hazardous (Ke-
meny, 1978; Lowe, 2004; Mason et al., 2013). Such differences in A. Clair et al. / SSM -Population Health 2 (2016) 306–316 308 respondents (Arora et al., 2015). Population registers and censuses
were used to assess the representativeness of the samples (Euro-
stat, 2012). respondents (Arora et al., 2015). Population registers and censuses
were used to assess the representativeness of the samples (Euro-
stat, 2012). sample rotations are lost than if a 4 year dataset were used. Only
those respondents that participated in all 3 surveys were included,
as limiting the sample in this way better enables us to look at the
effects of the transition into housing payment arrears including
through the use of fixed-effects models, giving us health and ar-
rears information at 3 time points. Table 2 Descriptive statistics for outcome and predictor variables in retained sample. 2008 (%)
2009 (%)
2010 (%)
Data source
Individual variables
Good Health (0 worst-4 best)
3.12
3.09
3.07
EU SILC
Housing payment arrears
No arrears
45 457 (100%)
39 654 (96.3%)
39 991 (96.0%)
Arrears
0
1508 (3.66%)
1672 (4.01%)
Tenure
Owner-occupier
34 226 (75.3%)
34 864 (76.8%)
35 326 (77.8%)
Private rent
7967 (17.5%)
7534 (16.6%)
7282 (16.0%)
Reduced rent
3256 (7.16%)
3021 (6.65%)
2794 (6.15%)
Age
41.4
42.4
43.4
Gender
Male
24 111 (53.0%)
24 111 (53.0%)
24 111 (53.0%)
Female
21 346 (47.0%)
21 346 (47.0%)
21 346 (47.0%)
Marital status
Married
28 613 (63.0%)
29 067 (64.1%)
29 463 (64.9%)
Never married
12 607 (27.8%)
11 993 (26.4%)
11 504 (25,4%)
Separated/divorced
3594 (7.92%)
3706 (8.17%)
3756 (8.28%)
Widowed
579 (1.28%)
607 (1.34%)
654 (1.44%)
Education level
Primary only
2411 (5.37%)
2359 (5.24%)
2337 (5.19%)
Secondary only
26 007 (57.9%)
25 872 (57.5%)
25 840 (57.36%)
Post-secondary
16 472 (36.7%)
16 761 (37.3%)
16 874 (37.5%)
Disposable income (1000 s)
43.7
44.5
44.5
Chronic illness at baseline
No
29 658 (80.3%)
–
–
Yes
7293 (19.7%)
–
–
Limiting illness at baseline
No
32 393 (87.7%)
–
–
Yes
3760 (10.2%)
–
–
Yes, strongly limiting
781 (2.1%)
–
–
Economic activity
Employed
45 457 (100%)
41 637 (92.0%)
40 528 (89.6%)
Unemployed
–
1530 (3.38%)
1812 (4.01%)
Retired
–
750 (1.66%)
1343 (2.97%)
Other inactive
–
1312 (2.90%)
1549 (3.42%)
Population level variables
Housing
EEE (Eurostat, 2015)
Home-ownership rate
75.6%
73.5%
73.1%
Change in house prices
2.55
5.10
1.72
Arrears rate
3.39%
4.06%
4.47%
Macroeconomic
Change in GDP
142
2330
1220
Interest rate
4.53%
4.23%
3.78%
Social spending (Purchasing Power Standard per inhabitant, divided by 100)
Disability
5.27
5.11
5.15
Total welfare
58.27
58.53
59.48
Old age
22.79
22.99
23.79
Survival benefits
3.41
3.33
3.37
Health
16.99
16.66
16.63
Family and child
5.30
5.15
5.18
Unemployment
2.54
3.27
3.31
Housing
0.78
0.79
0.76
Social Capital
(EVS, 2008)
Most people can be trusted
34.31
–
–
People helpful most of the time
36.78
–
–
Sample countries: Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Greece, Spain, Finland, France, Hungary, Iceland, Italy, Lithuania, Luxemburg, Latvia,
Malta, The Netherlands, Norway, Poland, Portugal, Romania, Sweden, Slovenia, Slovakia, and the UK. Sample countries: Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Greece, Spain, Finland, France, Hungary, Iceland, Italy, Lithuania, Luxemburg, Latvia,
Malta, The Netherlands, Norway, Poland, Portugal, Romania, Sweden, Slovenia, Slovakia, and the UK.
Housing payment arrears and economic activity other than employed are zero in 2008 due to sample constraints. House price data are missing for Norway and Iceland.
Interest rates missing for Norway, Iceland and Estonia. Social capital data missing for Iceland, Spain and the Czech Republic. Rental regulation information is missing for
Cyprus, Czech Republic, Estonia, Hungary, Iceland, Malta, Norway, Romania, and Slovenia. Data sources Respondents were selected for analysis on the basis of being
aged 16 or over, not living in rent free accommodation and being
both employed and not in housing payment arrears in 2008. These
restrictions were applied to ensure that the analysis would capture
those most affected by the recession rather than those already
experiencing difficulty. This resulted in a sample size of 136,371
person/year responses (45,457 individuals) with country sample
sizes varying between 294 person/years for Bulgaria and over
14,000 for France and The Netherlands. Longitudinal data were available for 25 of the then 27 EU
member states, the exceptions being Germany and Ireland. In
addition, we included data for Iceland and Norway, yielding a total
of 27 nations. The overall individual non-response rate (account-
ing for multiple interviews in the 2010 dataset) varies across
countries, from around 10% in Cyprus to over 40%, the maximum
non-response permitted by Eurostat, in Denmark and Norway
(Eurostat, 2012). EU-SILC requires representative probabilistic
samples, which was achieved in all the countries with the ex-
ception of Spain, which used substitution methods to replace non- Transitioning into housing payment arrears was a dichotomous
measure of whether a household entered into arrears across sur-
vey waves, as assessed by the following question: “In the last
twelve months, has the household been in arrears, i.e. has been Table 2
Descriptive statistics for outcome and predictor variables in retained sample. Table 2
Descriptive statistics for outcome and predictor variables in retained sample. Table 2 H
i
d
i
i i
h
h
l
d
i
2008 d
l
i
H
i
d
i
i
f
N
d I
l
d Sample countries: Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Greece, Spain, Finland, France, Hungary, Iceland, Italy, Lithuania, Luxemburg, Latvia,
Malta, The Netherlands, Norway, Poland, Portugal, Romania, Sweden, Slovenia, Slovakia, and the UK. Housing payment arrears and economic activity other than employed are zero in 2008 due to sample constraints. House price data are missing for Norway and Iceland. Interest rates missing for Norway, Iceland and Estonia. Social capital data missing for Iceland, Spain and the Czech Republic. Rental regulation information is missing for
Cyprus, Czech Republic, Estonia, Hungary, Iceland, Malta, Norway, Romania, and Slovenia. A. Clair et al. / SSM -Population Health 2 (2016) 306–316 309 unable to pay on time due to financial difficulties for: (a) rent
(b) mortgage repayment, for the main dwelling?” * po 0.05.
** p o 0.01.
*** p o 0.001. Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level
account for within-individual (time), household and country-level clustering. * po 0.05.
** p o 0.01. Statistical models We use individual fixed effect models to investigate the re-
lationship between housing payment arrears and health, as fol-
lows: Following convention we assessed self-reported health using a
scale from 0 – very bad to 4 – very good (Singh-Manoux et al.,
2006). It is well-known that self-reported health is subject to
perceptual and cultural biases (Jürges, 2007; Zimmer, Natividad,
Lin, & Chayovan, 2000). However it is a widely used indicator of
individual health and correlates strongly with mortality in many
countries, provides a summary of overall health problems, and,
when measured repeatedly in the same individuals, avoids po-
tential cross-cultural biases (Ganna & Ingelsson, 2015; Jylhä, 2009). = α + β
+ β
+ β
+ β
+γ +ε
( )
Health
Arrears
Age
Income
DEM
1
i j k
i j k
i j k
i j k
i j k
i j k
, ,
0
1
, ,
2
, ,
3
, ,
n
, ,
i
, , ( )
1 Here i is individual, j is country, and k is year. γi represents the
individual fixed effects and εi,j,k the individual error term. DEM Table 3
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Multilevel models
account for within-individual (time), household and country-level clustering.
* ndividual (time), household and country-level clustering. te: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variable
ount for within-individual (time), household and country-level clustering.
* po 0 05 Table 3 using payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed. Table 3
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individua Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed. Full sample
Owners
Renters
Covariate
Unadjusted multilevel
Adjusted multilevel model
Individual Fixed effects
Individual Fixed effects
Individual Fixed effects
Housing payment arrears
0.10***
0.07***
0.06**
0.00
0.09***
(0.01)
(0.01)
(0.02)
(0.03)
(0.02)
Age
-
0.03***
0.02
0.02
0.01
(0.00)
(0.02)
(0.03)
(0.02)
Age2
-
0.00***
0.00
0.00
0.00***
(0.00)
(0.00)
(0.00)
(0.00)
Female
-
0.04***
-
-
-
(0.00)
Marital status
Married
-
0.01
0.04
0.01
0.15**
(0.01)
(0.03)
(0.03)
(0.05)
Separated or divorced
-
0.01
0.04
0.01
0.05
(0.01)
(0.04)
(0.04)
(0.05)
Widowed
-
0.08***
0.10
0.18
0.37
(0.02)
(0.12)
(0.10)
(0.21)
Never married
-
Ref. Ref. Ref. Ref. Education level
Post-secondary
-
0.21***
0.03
0.03
0.05
(0.01)
(0.08)
(0.08)
(0.11)
Secondary
-
0.10***
0.02
0.00
0.03
(0.01)
(0.10)
(0.09)
(0.10)
Primary
-
Ref. Ref. Ref. Ref. Tenure
Private rent
-
0.06***
0.03
-
-
(0.01)
(0.02)
Reduced rent
-
0.06***
0.01
-
-
(0.01)
(0.02)
Owner occupier
Ref. Ref. -
-
Disposable income (1000 s)
-
0.00***
0.00
0.00
0.00
(0.00)
(0.00)
(0.00)
(0.00)
Chronic Illness at baseline year (yes)
-
0.41***
-
-
-
(0.01)
Limiting illness at baseline year
Yes, strongly limiting
-
0.82***
-
-
-
(0.02)
Yes
-
0.41***
-
-
-
(0.01)
No
-
Ref. -
-
-
Economic activity
Lost job
–
0.09***
0.05*
0.03
0.04
(0.01)
(0.02)
(0.03)
(0.05)
Retired
–
0.05**
0.02
0.05
0.00
(0.02)
(0.04)
(0.04)
(0.07)
Other inactive
–
0.12***
0.04
0.03
0.05
(0.01)
(0.03)
(0.03)
(0.04)
Employed
–
Ref. Ref. Ref. Ref. Individual fixed effects
N
N
Y
Y
Y
Country-level variables
N
N
Y
Y
Y
Number of individual-years
103041
101155
91547
65264
26285
Countries in sample
27
27
24
24
24
Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Multilevel models
account for within individual (time) household and country level clustering A. Clair et al. / SSM -Population Health 2 (2016) 306–316 310 In subsequent models we evaluated a range of potential mod-
ifying factors using a series of interaction terms. Table 4 Association of transitioning into housing payment arrears on self-reported health
among renters. Age
Gender
Women
Men
All
Under 30
0.08
0.06
0.00
(0.05)
(0.08)
(0.05)
30–59
0.17***
0.06
0.11***
(0.04)
(0.04)
(0.03)
60 and over
1.12***
0.31
0.40
(0.04)
(0.18)
(0.22)
All
0.12**
0.08*
0.09***
(0.04)
(0.03)
(0.02)
Notes: Results presented from 12 separate fixed effects statistical models. Robust
standard errors in parentheses. Table 3 These potential
modifiers included country-level macroeconomic risks, social ca-
pital and support, and social protection expenditure, as well as age
and gender. All models were performed using Stata 13 (xtreg and
xtmixed commands). Table 2 provides descriptive statistics for the
variables used in the study. Results Association of transitioning into housing payment arrears and health
status Table 3 shows the results of a series of multi- and single-level
statistical models progressively incorporating control variables. As
shown in the first column, the unadjusted association of transi-
tioning into housing payment arrears with worse health, con-
trolling for household and country clustering, is 0.10 units (95%
CI: 0.07 to 0.12). After adjusting for baseline health, education,
and potential confounding factors, the association was slightly
attenuated to 0.07 (95% CI: 0.05 to 0.10). Including in-
dividual fixed effects yielded the most conservative estimated
association of 0.06 (95% CI 0.02 to 0.10). * po 0.05,
** p o 0.01,
*** po 0.001. * po 0.05,
** p o 0.01,
*** po 0.001. represents a vector of control variables, including marital status,
education level, and housing tenure. All models are clustered by
country to reflect non-independence of sampling. All regression
models were weighted by person weights, to represent popula-
tions. Multilevel analysis with household and country level clus-
tering is also conducted. To facilitate interpretation of results we
standardised the health outcome for analysis. represents a vector of control variables, including marital status,
education level, and housing tenure. All models are clustered by
country to reflect non-independence of sampling. All regression
models were weighted by person weights, to represent popula-
tions. Multilevel analysis with household and country level clus-
tering is also conducted. To facilitate interpretation of results we
standardised the health outcome for analysis. We then disaggregated these associations by owners and ren-
ters, proceeding with the individual fixed effects regression Fig. 1. Estimated impact of housing payment arrears on self-reported health with 95% confidence intervals, 2008–2010. Fig. 1. Estimated impact of housing payment arrears on self-reported health with 95% confidence intervals, 2008–2010. Fig. 1. Estimated impact of housing payment arrears on self-reported health with 95% confidence intervals, 2008–2010. A. Clair et al. / SSM -Population Health 2 (2016) 306–316 311 hown in Table 3, there was no significant association
using payment arrears and health among owners
0.05 to 0.06), but there was among renters (0.09,
5 to 0.13). Next we further investigated whether these impacts w
ferent by age and sex, focusing on renters. As shown in
presenting 12 separate fixed effects models, we observ
associations were greatest among women aged 30 to 59
Fig. 2. Table 5 Table 5
Interaction of potential modifying factors with the housing payment arrears-health
association, baseline sample of individuals not in arrears and employed, multilevel
analysis. Next we sought to investigate potential factors which may
exacerbate or mitigate this impact, given the marked variation
across countries. First we investigated characteristics of the
housing market using a series of interaction terms. Table 5 shows
the results. We observed that in countries with greater prevalence
of homeownership, both owners and renters appeared to have
slightly attenuated health risks associated with experiencing
housing payment arrears. Additionally, rising housing prices ap-
peared to exacerbate the estimated impact of housing payment
arrears on health status. Modifying factor
Owners
Renters
Macroeconomic
Home ownership rate
0.006**
0.006**
(0.002)
(0.002)
House price change
0.004*
0.006*
(0.002)
(0.003)
GDP change
0.00
0.002*
(0.00)
(0.001)
Interest rate
0.00
0.02*
(0.01)
(0.01)
Country arrears rate
0.02**
0.02*
(0.01)
(0.01)
Social Capital
People are helpful most of the time
0.00
0.00
(0.00)
(0.00)
Most people can be trusted
0.00
0.00
(0.00)
(0.00)
Social protection expenditure
Disability spending
0.01
0.01
(0.00)
(0.01)
Total social protection spending
0.001*
0.00
(0.001)
(0.00)
Old age social protection spending
0.004*
0.005*
(0.002)
(0.002)
Survival benefits spending
0.01
0.01
(0.01)
(0.01)
Health spending
0.005*
0.00
(0.002)
(0.00)
Family and child spending
0.01
0.01
(0.00)
(0.01)
Unemployment spending
0.00
0.01
(0.01)
(0.01)
Housing spending
0.05**
0.01
(0.02)
(0.02)
Rental market regulations
Rent controls
0.04
0.01
(0.04)
(0.04)
Tenant-landlord relationship
0.03
0.02
(0.04)
(0.04)
Note: Models include all individual level control variables as used in the previous
table, separate models run for each potential interaction. Standard errors in par-
entheses
***po0.001. * po0 .05. ** po 0.01. Modifying factor
Owners
Renters We then tested whether greater rental market regulation was
protective, using binary measures taken from a European Com-
mission Study (Cuerpo, Kalantaryan, & Pontuch, 2014). These
binary measures capture whether a country has a higher level of
tenancy protection, either through rent controls or favourable
positioning of tenants in negotiations with landlords. None found
a significant interaction with housing payment arrears. Finally we included a set of formal and informal social support
measures. The formal ones included alternative categories of social
protection
expenditure,
including
housing
support,
old-age
spending, healthcare, family support and unemployment benefits. The informal ones involved social capital factors, the proportion in
a country responding that people are helpful most of the time and
whether people can be trusted. Cross-national differences in the impact of housing payment arrears Having observed that renters had worse health, we next in-
vestigated differing impacts across countries. Figs. 1 and 2 plot the
country-specific associations of transitioning into housing pay-
ment arrears with self-reported health, both overall (Fig. 1), and
disaggregated by tenure (Fig. 2). The estimated impact varied
considerably. In some countries, there was no effect of housing
payment arrears, while in others, such as Poland, the negative
effect of arrears on health is considerably larger than average
(0.36 compared to the average 0.07, both po0.05). Although
there are a few exceptions, we observed that in the majority of
countries, including those diverse as Belgium, Iceland, Austria, and
Sweden, renters are much worse off than owners when facing Table 5 None of the many variables tested
had a significant protective effect on the housing payment arrears-
health status association. Robustness checks Thus far we have treated the health outcomes as a scale vari-
able for ease of interpretation. However treating health in this way
may have affected our results. We check this by re-running the
models shown in Table 3 using ordinal analyses, the results of
which are given in Appendix Table A1. The results of these models
are consistent with the linear approach, finding that housing
payment arrears is associated with deterioration in health for the
full sample, which reflects a large effect for renters and a non-
significant effect for owners when disaggregated by tenure. As a further check we run a number of longitudinal linear
probability models where the self-reported health outcome has
been transformed into a binary variable (Good/very good health
coded as 0, fair/bad/very bad as 1). The results of these models are
given in Appendix Table A2. The findings are equivalent to our
previous models, finding that housing payment arrears is asso-
ciated with an increased risk of reporting poor health, which when
disaggregated by tenure reflects a large risk for renters and no
statistically significant risk for owners. 95% CI 0.09 to 0.26) and particularly women aged 60 and over
(1.12, 95% CI 1.03 –to 1.21). These results are in contrast to
results from Australia, which indicated that male renters fared
worse when facing housing payment difficulties (Bentley et al.,
2011, 2012). 95% CI 0.09 to 0.26) and particularly women aged 60 and over
(1.12, 95% CI 1.03 –to 1.21). These results are in contrast to
results from Australia, which indicated that male renters fared
worse when facing housing payment difficulties (Bentley et al.,
2011, 2012). In order to further investigate the differences found across te-
nures we investigate whether the differing characteristics of
owners and renters may account for some of the variation found. Web Appendix Table A3 gives the descriptives for the sample
disaggregated by tenure. As would be expected there are differ-
ences in the characteristics of individuals in these different te-
nures. We ran fixed-effects interaction models (equivalent to the
third model in Table 3) for all of the time-varying individual-level
predictors to investigate whether any differences in sample char-
acteristics may help us to further understand the health con-
sequences of housing payment arrears. Only one significant in-
teraction was found, indicating that widow(er)s report higher
health when living in reduced rate rented homes. Robustness checks Cross-national differences in the impact of housing payment arrears Cross-national differences in the impact of housing payment arrears Results Estimated impact of housing payment arrears on self-reported health with 95% confidence intervals, 2008–2010, by tenure. Fig. 2. Estimated impact of housing payment arrears on self-reported health with 95% confidence intervals, 2008–2010, by tenure. models. As shown in Table 3, there was no significant association
between housing payment arrears and health among owners
(0.00, 95% CI: 0.05 to 0.06), but there was among renters (0.09,
95% CI: 0.05 to 0.13). models. As shown in Table 3, there was no significant association
between housing payment arrears and health among owners
(0.00, 95% CI: 0.05 to 0.06), but there was among renters (0.09,
95% CI: 0.05 to 0.13). models. As shown in Table 3, there was no significant association
between housing payment arrears and health among owners
(0.00, 95% CI: 0.05 to 0.06), but there was among renters (0.09,
95% CI: 0.05 to 0.13). Next we further investigated whether these impacts were dif-
ferent by age and sex, focusing on renters. As shown in Table 4,
presenting 12 separate fixed effects models, we observed that
associations were greatest among women aged 30 to 59 (0.17, A. Clair et al. / SSM -Population Health 2 (2016) 306–316 312 housing payment arrears. housing payment arrears. Discussion Our analysis demonstrates that transitioning into housing
payment arrears is a significant risk factor for worsening health A. Clair et al. / SSM -Population Health 2 (2016) 306–316 313 that, in contrast to the general trend in Europe, owners in Poland
had greater risk of health problems than did renters. This may
reflect exposure to foreign currency debt, used in some nations to
purchase homes. Approximately 60% of Polish mortgages were
denominated in foreign currency, often from Swiss banks, expos-
ing them to exchange rate fluctuations that may not have been
anticipated (Miles & Pillonca, 2008). In Austria and Belgium ren-
ters who experience housing payment arrears appeared to fare
particularly poorly compared with owners and the rest of Europe. Such unanticipated shocks, which occur outside the control of
individuals, may be particularly deleterious. This is also consistent
with our observation that escalating housing prices in a nation
appeared to exacerbate the impacts of arrears on health status. status across Europe. We found that these negative associations
were concentrated in renters, especially women aged 30 and over. However, in most countries, a short term period of housing arrears
had no identifiable impact on owners’ health status. Escalating
house prices appeared to render renters more vulnerable to the
health consequences of payment problems. As with all observational studies, our analysis has several lim-
itations. First, reflecting the availability of comparative data, we
focused on the financial aspects of housing. This does not reflect
the many aspects of housing difficulties, including low quality or
substandard housing. Second, although our analysis utilised the
longest series of available comparative longitudinal data for a large
number of European countries, it still is limited to a relatively
short duration. This makes the effects observed more likely to
reflect mental, rather than physical health, many of which would
require longer time periods to develop. Further research is also
needed to investigate the longer-term consequences and potential
scarring effects of housing insecurity. Third, we failed to identify
statistically significant protective factors, whether from social
protection or social capital. This may reflect a low degree of sta-
tistical power when these protective factors are observed at the
country level or the crude nature of such measures which do not
capture how, as well as how much, money is spent. Discussion Given the
number of countries included in our study and the complexity and
variety that characterises housing policy, it has not been possible
to thoroughly investigate the relative protections afforded to
owners and renters across countries. Additional studies are nee-
ded to understand the reasons for the considerable differences
across nations in the risks faced by renters. Fourth, one strength of
our study is that we were able to adjust for baseline health and
identify changes in persons who experienced new payment pro-
blems. While we adjust for baseline health it remains possible that
people's health may influence their likelihood of falling to housing
payment arrears given the period of time between data collection
points. However we believe this unlikely given our analytical ap-
proach. The controls for individual fixed effects may also yield
conservative estimates of the association of housing with worse
health status. Fifth, again reflecting the lack of available com-
parative data for a large number of EU countries, we were unable
to disaggregate those facing housing payment problems of differ-
ent levels of severity. In some European countries, especially parts of the UK, housing
prices have risen and are projected to remain high. While policies
are in place in some countries to subsidise and support people
experiencing arrears, less attention has been paid to intervening
directly in the market to render housing more affordable. In some
cases, policies designed to facilitate home ownership have per-
versely increased prices, so worsening the risk to health. For ex-
ample in the UK (particularly England) official policies have the
effect of maintaining or increasing already high house prices, ei-
ther directly or through encouraging the perception of housing as
an asset, with examples including ‘Buy to Let’ and ‘Help to Buy’,
although April 2016 tax changes have made the former less at-
tractive for landlords, with as yet unknown consequences. Overall our results suggest that, similar to job loss, housing
payment arrears is a major public health concern especially in
times of economic downturn and hardship. As well as seeking to
reduce instances of housing payment arrears, research should in-
vestigate how to mitigate the impacts of arrears on health, and
understand why some groups are more severely affected than
others. Acknowledgements Our study points to several directions for future research,
which should be a priority given that difficulties in the housing
sector are likely to persist in Europe. One is to account for not only
demand-side factors, which shape people's ability to afford
housing, but also supply-side factors, which may help contain
escalating housing costs, such as rent controls and provision of
social housing. Further research is needed to develop and use
better measures of rental controls and negotiating power of ren-
ters, likely necessitating new data collection across Europe. This work was supported by the European Research Council
[grant number 313590-HRES] (AC and DS) and a Wellcome Trust
Investigator Award (DS). Conflicts of interest None declared. Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Multilevel models
account for within-individual (time), household and country-level clustering. Log odds coefficients. Fixed effects models not available, all models unweighted as weighting
t
t d * p o 0.05.
** p o 0.01.
*** p o 0.001. Appendix A See Tables A1–A3. See Tables A1–A3. See Tables A1–A3. Several observations of outliers are noteworthy. We observed A. Clair et al. / SSM -Population Health 2 (2016) 306–316 314 Table A1
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed, ordinal
models. Covariate
Full sample
Owners
Renters
Unadjusted multilevel
Adjusted multilevel model
Longitudinal ordinal RE
Longitudinal ordinal RE
Longitudinal ordinal RE
Housing payment arrears
0.42***
0.31***
0.21*
0.02
0.43***
(0.06)
(0.05)
(0.09)
(0.10)
(0.08)
Age
–
0.13***
0.13***
0.12***
0.15***
(0.01)
(0.02)
(0.02)
(0.02)
Age2
–
0.00***
0.00***
0.00*
0.00***
(0.00)
(0.00)
(0.00)
(0.00)
Female
–
0.18***
0.20***
0.14*
0.32***
(0.02)
(0.04)
(0.06)
(0.04)
Marital status
Married
–
0.04
0.10
0.08
0.10
(0.03)
(0.05)
(0.05)
(0.09)
Separated or divorced
–
0.02
0.05
0.11
0.01
(0.05)
(0.07)
(0.09)
(0.11)
Widowed
–
0.27**
0.26*
0.32*
0.13
(0.10)
(0.11)
(0.13)
(0.17)
Never married
–
Ref. Ref. Ref. Ref. Education level
Post-secondary
–
0.94***
1.17***
1.16***
1.18***
(0.05)
(0.17)
(0.20)
(0.14)
Secondary
–
0.44***
0.61***
0.63**
0.55***
(0.05)
(0.17)
(0.19)
(0.14)
Primary
–
Ref. Ref. Ref. Ref. Tenure
Private rent
–
0.27***
0.12
-
-
(0.03)
(0.07)
Reduced rent
–
0.31***
0.25**
-
-
(0.04)
(0.07)
Owner occupier
–
Ref. Ref. -
-
Disposable income (1000 s)
–
0.004***
0.01***
0.00***
0.01**
(0.000)
(0.00)
(0.00)
(0.00)
Chronic Illness at baseline year
(yes)
–
1.81***
1.85***
1.90***
1.70***
(0.04)
(0.11)
(0.10)
(0.13)
Limiting illness at baseline year
Yes, strongly limiting
–
3.09***
3.13***
3.18***
3.05***
(0.09)
(0.19)
(0.20)
(0.27)
Yes
–
1.63***
1.72***
1.77***
1.66***
(0.05)
(0.09)
(0.09)
(0.14)
No
–
Ref. Ref. Ref. Ref. Economic activity
Lost job
–
0.35***
0.39***
0.43**
0.31**
(0.05)
(0.10)
(0.14)
(0.12)
Retired
–
0.18*
0.16
0.08
0.30
(0.08)
(0.10)
(0.14)
(0.16)
Other inactive
–
0.40***
0.39***
0.40***
0.35**
(0.06)
(0.08)
(0.10)
(0.10)
Employed
–
Ref. Ref. Ref. Ref. Individual fixed effects
N
N
N
N
N
Country-level variables
N
N
Y
Y
Y
Number of individual-years
103041
101155
92931
66026
26907
Countries in sample
27
27
24
24
24
Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Multilevel models
account for within-individual (time), household and country-level clustering. Log odds coefficients. Fixed effects models not available, all models unweighted as weighting
not supported Table A1 Table A1
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed, ordinal
models. Table A1
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individ
models Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Multilevel models
account for within-individual (time), household and country-level clustering. Log odds coefficients. Fixed effects models not available, all models unweighted as weighting
not supported A. Clair et al. / SSM -Population Health 2 (2016) 306–316 315 Table A2
Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed, longitudinal
linear probability models with fixed effects. Covariate
Full sample
Owners
Renters
Unadjusted
longitudinal
Adjusted longitudinal
Longitudinal with country
vars
Longitudinal with country
vars
Longitudinal with country
vars
Housing payment arrears
0.05***
0.04**
0.04**
0.00
0.07***
(0.01)
(0.01)
(0.01)
(0.01)
(0.01)
Age
–
0.01
0.01
0.01
0.02
(0.01)
(0.01)
(0.01)
(0.02)
Age2
–
0.00*
0.00*
0.00
0.00
(0.00)
(0.00)
(0.00)
(0.00)
Female
–
–
–
–
–
Marital status
Married
–
0.01*
0.01
0.01
0.06**
(0.01)
(0.01)
(0.01)
(0.01)
Separated or divorced
–
0.04**
0.04**
0.02
0.07*
(0.01)
(0.01)
(0.02)
(0.03)
Widowed
–
0.01
0.00
0.04
0.05
(0.06)
(0.07)
(0.05)
(0.11)
Never married
–
Ref. Ref. Ref. Ref. Education level
Post-secondary
–
0.03
0.02
0.02
0.09
(0.04)
(0.04)
(0.04)
(0.05)
Secondary
–
0.00
0.01
0.02
0.05
(0.01)
(0.05)
(0.04)
(0.06)
Primary
–
Ref. Ref. Ref. Ref. Tenure
Private rent
–
0.00
0.00
–
–
(0.01)
(0.01)
Reduced rent
–
0.02
0.01
–
-
(0.01)
(0.01)
Owner occupier
–
Ref. Ref. –
–
Disposable income (1000 s)
–
0.00
0.00
0.00
0.00
(0.00)
(0.00)
(0.00)
(0.00)
Chronic Illness at baseline
year (yes)
–
–
–
–
–
Limiting illness at baseline year
Yes, strongly limiting
–
–
–
–
–
Yes
–
–
–
–
–
No
–
Ref. Ref. Ref. Ref. Economic activity
Lost job
–
0.03**
0.03*
0.02
0.02
(0.01)
(0.01)
(0.02)
(0.01)
Retired
–
0.01
0.01
0.03
0.02
(0.03)
(0.03)
(0.02)
(0.05)
Other inactive
–
0.01
0.01
0.02
0.01
(0.02)
(0.02)
(0.02)
(0.02)
Employed
–
Ref. Ref. Ref. Ref. Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Good/very good health
coded as 0, fair/bad/very bad as 1. Fixed effects with clustered standard errors.
*
0 05 Table A1 Individual fixed effects
N
N
N
N
N
Country-level variables
N
N
Y
Y
Y
Number of individual-years
101483
99749
91547
65264
26285
Countries in sample
27
27
24
24
24
Note: Renters include those who rent at a reduced rate and market rate. Country-level variables are those population level variables given in Table 2. Good/very good health
coded as 0, fair/bad/very bad as 1. Fixed effects with clustered standard errors. * p o0 .05. References p
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Estimated impact of transitioning into housing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed, longitudinal
linear probability models with fixed effects. ing payment arrears on self-reported health, 2008–2010, baseline sample of individuals not in arrears and employed, longitudinal
s A. Clair et al. / SSM -Population Health 2 (2016) 306–316 316 Table A3
Sample descriptives disaggregated by tenure. Owners
Renters
Statistical test
(difference)
Housing payment
arrears
1.95%
4.09%
p o0 .05
Age
43.04 (S.D. 10.70)
40.39 (S.D. 11.74)
p o 0.05
Marital status
Married 69.58%
45.80%
p o 0.05
Single 22.51%
39.62%
Divorced 6.65%
12.94%
Widowed 1.26%
1.64%
Education
Primary 3.84%
9.93%
p o 0.05
Secondary 56.43%
61.44%
Post-secondary
39.73%
28.63%
Disposable income
47665.13
(36255.76)
33083.15
(25852.57)
p o 0.05
Chronic illness
19.60%
20.12%
p o 0.05
Limiting illness
No 88.29%
86.04%
p o 0.05
Yes 9.76%
11.38%
Strongly limiting
1.95%
2.58%
Economic activity
Employed 94.54%
91.80%
p o 0.05
Unemployed 1.99%
4.01%
Retired 1.52%
1.62%
Inactive 1.96%
2.57% Table A3
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English
| null |
Combined direct-sun ultraviolet and infrared spectroscopies at Popocatépetl volcano (Mexico)
|
Frontiers in earth science
| 2,023
|
cc-by
| 16,135
|
OPEN ACCESS OPEN ACCESS
EDITED BY
Ryunosuke Kazahaya,
National Institute of Advanced Industrial
Science and Technology (AIST), Japan
REVIEWED BY
Ben Esse,
The University of Manchester,
United Kingdom
Jean-François Smekens,
University of Oxford, United Kingdom
*CORRESPONDENCE
N. Taquet,
noemi.taquet@gmail.com
SPECIALTY SECTION
This article was submitted to
Volcanology, a section of the journal
Frontiers in Earth Science
RECEIVED 06 October 2022
ACCEPTED 09 March 2023
PUBLISHED 06 April 2023
CITATION
Taquet N, Rivera Cárdenas C,
Stremme W, Boulesteix T, Bezanilla A,
Grutter M, García O, Hase F and
Blumenstock T (2023), Combined direct-
sun ultraviolet and infrared
spectroscopies at Popocatépetl
volcano (Mexico). Front. Earth Sci. 11:1062699. doi: 10.3389/feart.2023.1062699 EDITED BY
Ryunosuke Kazahaya,
National Institute of Advanced Industrial
Science and Technology (AIST), Japan
REVIEWED BY
Ben Esse,
The University of Manchester,
United Kingdom
Jean-François Smekens,
University of Oxford, United Kingdom
*CORRESPONDENCE
N. Taquet,
noemi.taquet@gmail.com
SPECIALTY SECTION
This article was submitted to
Volcanology, a section of the journal
Frontiers in Earth Science
RECEIVED 06 October 2022
ACCEPTED 09 March 2023
PUBLISHED 06 April 2023
CITATION
Taquet N, Rivera Cárdenas C,
Stremme W, Boulesteix T, Bezanilla A,
Grutter M, García O, Hase F and
Blumenstock T (2023), Combined direct-
sun ultraviolet and infrared
spectroscopies at Popocatépetl
volcano (Mexico). Front. Earth Sci. 11:1062699. doi: 10.3389/feart.2023.1062699 N. Taquet1*, C. Rivera Cárdenas1, W. Stremme1, T. Boulesteix2,
A. Bezanilla1, M. Grutter1, O. García3, F. Hase4 and
T. Blumenstock4 1Instituto de Ciencias de la Atmósfera y Cambio Climático, Universidad Nacional Autónoma de México,
Mexico City, Mexico, 2Volcanology Research Group, Department of Life and Earth Sciences, Instituto de
Productos Naturales Y Agrobiología (IPNA-CSIC), San Cristóbal de La Laguna, Spain, 3Centro de
Investigación Atmosférica de Izaña (CIAI), Agencia Estatal de Meteorología (AEMet), Santa Cruz de
Tenerife, Spain, 4Karlsruhe Institute of Technology (KIT), Institute of Meteorology and Climate Research
(IMK-ASF), Karlsruhe, Germany Volcanic plume composition is strongly influenced by both changes in magmatic
systems
and
plume-atmosphere
interactions. Understanding
the
degassing
mechanisms controlling the type of volcanic activity implies deciphering the
contributions of magmatic gases reaching the surface and their posterior chemical
transformations in contact with the atmosphere. Remote sensing techniques based on
direct solar absorption spectroscopy provide valuable information about most of the
emitted magmatic gases but also on gas species formed and converted within the
plumes. TYPE Original Research
PUBLISHED 06 April 2023
DOI 10.3389/feart.2023.1062699 TYPE Original Research
PUBLISHED 06 April 2023
DOI 10.3389/feart.2023.1062699 solar absorption spectroscopy, volcanic degassing, Popocatépetl, FTIR, DOAS, Pandora,
dome cycle OPEN ACCESS In this study, we explore the procedures, performances and benefits of
combining
two
direct
solar
absorption
techniques,
high
resolution
Fourier
Transform
Infrared
Spectroscopy
(FTIR)
and
Ultraviolet
Differential
Optical
Absorption Spectroscopy (UV-DOAS), to observe the composition changes in the
Popocatépetl’s plume with high temporal resolution. The SO2 vertical columns
obtained from three instruments (DOAS, high resolution FTIR and Pandora) were
found similar (median difference <12%) after their intercalibration. We combined them
to determine with high temporal resolution the different hydrogen halide and halogen
species to sulfur ratios (HF/SO2, BrO/SO2, HCl/SO2, SiF4/SO2, detection limit of HBr/
SO2) and HCl/BrO in the Popocatépetl’s plume over a 2.5-years period (2017 to mid-
2019). BrO/SO2, BrO/HCl, and HCl/SO2 ratios were found in the range of (0.63 ±
0.06 to 1.14 ± 0.20) × 10−4, (2.6 ± 0.5 to 6.9 ± 2.6) × 10−4, and 0.08 ± 0.01 to 0.21 ±
0.01 respectively, while the SiF4/SO2 and HF/SO2 ratios were found fairly constant at
(1.56 ± 0.25) × 10−3 and 0.049 ± 0.001. We especially focused on the full growth/
destruction cycle of the most voluminous lava dome of the period that took place
between February and April 2019. A decrease of the HCl/SO2 ratio was observed with
the decrease of the extrusive activity. Furthermore, the short-term variability of BrO/
SO2 is measured for the first time at Popocatépetl volcano together with HCl/SO2,
revealing different behaviors with respect to the volcanic activity. More generally,
providing such temporally resolved and near-real-time time series of both primary and
secondary volcanic gaseous species is critical for the management of volcanic
emergencies, as well as for the understanding of the volcanic degassing processes
and their impact on the atmospheric chemistry. COPYRIGHT
© 2023 Taquet, Rivera Cárdenas,
Stremme, Boulesteix, Bezanilla, Grutter,
García, Hase and Blumenstock. This is an
open-access article distributed under the
terms of the Creative Commons
Attribution License (CC BY). The use,
distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication
in this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms. COPYRIGHT
© 2023 Taquet, Rivera Cárdenas,
Stremme, Boulesteix, Bezanilla, Grutter,
García, Hase and Blumenstock. This is an
open-access article distributed under the
terms of the Creative Commons Attribution License (CC BY). COPYRIGHT
© 2023 Taquet, Rivera Cárdenas,
Stremme, Boulesteix, Bezanilla, Grutter,
García, Hase and Blumenstock. This is an
open-access article distributed under the
terms of the Creative Commons
Attribution License (CC BY). The use,
distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication
in this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms. Attribution License (CC BY). The use,
distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication
in this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms. KEYWORDS 1 Introduction (Rüdiger et al., 2021) and provide valuable information about the
lifetimes and conversion rates of the primary (HCl, HF, HBr) and
secondary (BrO, ClO, OClO, BrOH, Br2, BrCl, BrONO2) halogen
species. However, the lack of simultaneous measurements of the
different halogens species leaves some input parameters poorly
constrained,
and
the
influence
of
plume-mixing
processes,
relative humidity, presence of aerosols, photochemistry and/or
redox conditions on the conversion rates remains still not fully
understood. A review of the main recent findings on this topic is
given in Gutmann et al. (2018). Some typical behaviors of the BrO/
SO2 ratios with respect to the volcanic activity were described at
Mount Etna (Bobrowski and Giuffrida, 2012), Nevado del Ruiz
(Lübcke et al., 2014), Tungurahua (Warnach et al., 2019) and
Cotopaxi (Dinger et al., 2018), with low ratios prior to and
during eruptions and/or explosive activity, while higher ratios
occurred at the end of the eruptions or during passive degassing. On the other hand, Aiuppa et al. (2005) observed a coherent
geochemical behaviour of halogen (F, Cl, Br, I) to sulfur ratios in
the Mount Etna’s plume before (increase of the ratio) and during
(decrease of the ratio) the 2004 eruption. Unless Cl and F partition
early in water/brines (Aiuppa, 2009; Shinohara, 2009), these species
are generally exsolved from magma shallower than sulfur. The
conditions of bromine exsolution from magma are not yet well
understood, and both shallow and deep origins are still considered
(Gutmann et al., 2018), complicating the interpretation of the BrO/
SO2 ratios in terms of magmatic processes. Volcanic plume composition measurements allow detection of
significant changes in the magmatic processes from the magma
chamber to the most superficial levels (Francis et al., 1995;
Oppenheimer
and
McGonigle,
2004)
and
contribute
to
determining the degassing balance during eruptions (Gerlach and
Graeber, 1985). Among the main magmatic volatile species involved
in volcanic eruptions, halogens (F, Cl, Br, I) play a key role in the
extrusive processes, affecting the physico-chemical properties of
magmas (Dingwell and Mysen, 1985; Aiuppa, 2009), and having
major implications for their transport towards the surface, their
ascension speed, their potential to segregate a gas phase, their
eruptability and the eruption dynamics/explosivity. 1 Introduction In particular,
based on their contrasted solubility in magma compared to sulfur,
the temporal variability of halogen-to-sulfur ratios in plumes
allowed
deciphering
the
magmatic
processes
and
eruptive
dynamics (Mori et al., 1993; Burton et al., 2007; Schipper et al.,
2019; Aiuppa et al., 2021). For example, an increase of Cl/S ratios was observed during the
eruptive periods with respect to quiescent periods, at, e.g., Mount
Etna during the 2001 and 2002–2003 eruptions (Aiuppa et al., 2002;
2004; Notsu and Mori, 2010), which was interpreted in terms of
solubility-controlled fractionation. The variability of the HCl/SO2,
CO2/SO2 in different eruptive contexts provides insights on the
transition between quiet and explosive regimes, and on explosion
mechanisms (Burton et al., 2007; Woitischek et al., 2020; Aiuppa
et al., 2021). Edmonds et al. (2002) observed an increase of the HCl/
SO2 ratio at Soufrière Hills Volcano during lava extrusion,
illustrating the shallow decompressional degassing of Cl. SiF4 is a byproduct of the reaction of HF with silicates either
within the conduit or during the ash transport in the plume (Love
et al., 1998; de Hoog et al., 2005). FTIR spectroscopy is the only
method allowing its quantification together with HF (Mori et al.,
2002). This compound has been detected at a few volcanoes such as
Mount Etna and Satsuma-Iwojima, but especially at Popocatépetl
(Love et al., 1998; Stremme et al., 2011; Stremme et al., 2012; Taquet
et al., 2017; Taquet et al., 2019) where frequent ash-laden plumes
and vigorous degassing through successive dome extrusions favors
the formation of SiF4 (de Hoog et al., 2005). The previous studies
evidenced that both formation modes occur at Popocatépetl (Taquet
et al., 2017; Taquet et al., 2019) and that the SiF4/SO2 ratio increases
several minutes preceding (Love et al., 1998) and during dome-
related explosions (Stremme et al., 2011; Taquet et al., 2017). g
p
g
g
Some of these halogen derivatives are reactive in volcanic
plumes,
obscuring
the
relationships
between
the
plume
composition and the magmatic/eruptive processes, which require
both primary (magmatic) and secondary (in-plume) species to be
accounted for in degassing balances. In-plumes reactions include
SO2 and HCl scavenging on volcanic ash (Martin et al., 2012;
Delmelle et al., 2018), uptake by sulfate aerosols (Edmonds et al.,
2003; Rodriguez et al., 2008; Rüdiger et al., 2021), and halogen
chemistry (Bobrowski et al., 2007; Rudiger et al., 2021). OPEN ACCESS The use,
distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication
in this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms. Frontiers in Earth Science 01 frontiersin.org Taquet et al. Taquet et al. 10.3389/feart.2023.1062699 Frontiers in Earth Science frontiersin.org 1 Introduction Its manifestation at
the surface consisted in a “Strombolian-type” episode with sustained
ash-venting, ejecting incandescent blocks on the external slopes of
the cone. On 19 February 2019, an overflight confirmed the presence
of a voluminous dome inside the crater with 200 m diameter
(i.e., ~1.3 Mm3 according to Valade et al., submitted). Similar
3–9 h-long tremor segments were recorded over the next days
until day 27 summing to approximately 120 h. Another overflight
on 28 February confirmed that the dome 82 was still occupying the
crater. Between 26 February and 10 March 2019, 48 explosions were
reported by CENAPRED (Figure 5), some of which (e.g., 1 and
9 March 2019) expelled incandescent blocks on the external slopes of
the cone hundreds of meters downslope. This explosive activity
resulted in 69 VAAC alerts, 29 of which occurred between
5–8 March. Strong degassing during the overflight on 7 March
did not allow observations of the dome destruction. An image of plumes, mainly in open-path configuration, which requires an
intense IR source (lava fountains and lakes or artificial), limiting
its applicability. Solar occultation FTIR spectroscopy, using direct
sunlight
overcomes
some
of
these
limitations,
allowing
measurements
during
passive
and
active
degassing
phases
(Francis et al., 1998; Oppenheimer et al., 1998; Burton et al.,
2001; Stremme et al., 2011; Taquet et al., 2019). Few studies
(Love et al., 1998; Love et al., 2000; Smekens et al., 2023) present
simultaneous FTIR and UV measurements of volcanic plumes
composition using thermal emission (in the LWIR range) and
the solar radiation scattered by the atmosphere or clouds behind
volcanic plumes. Thermal emission measurements have been
performed recently at Popocatépetl (Taquet et al., 2017) and have
proven to complement direct solar absorption measurements. Using
thermal emission allows measuring with high frequency by just
pointing towards the volcanic plume, but solving the radiative
transfer and deriving the composition remain challenging (Love
et al., 2000; Platt et al., 2018). Butz et al. (2017) combined for the first
time direct-sun DOAS and FTIR measurements using portable
instruments
(EM27/SUN
FTIR
and
UV
spectrometers)
to
simultaneously measure SO2, HCl, HF, CO2, BrO species in
Mount Etna’s volcanic plume. In such configuration, the different
instruments share the same line of sight and thus the same airmass. 1 Introduction Based on CENAPRED reports, following this
explosion and before the onset of the dome growth episode, daily
averages of ~15 min of tremor, of 1.3 Volcano Tectonic (VT)
earthquakes and of 150 exhalations (local term for low intensity
explosions with little impulsive character) were recorded. On
15 February 2019, at approximately 2:55 UTC (14 February at
20:55, local time), the first 6 h-long LPs-tremor-explosions
segment of this dome growth episode started. Its manifestation at
the surface consisted in a “Strombolian-type” episode with sustained
ash-venting, ejecting incandescent blocks on the external slopes of
the cone. On 19 February 2019, an overflight confirmed the presence
of a voluminous dome inside the crater with 200 m diameter
(i.e., ~1.3 Mm3 according to Valade et al., submitted). Similar
3–9 h-long tremor segments were recorded over the next days
until day 27 summing to approximately 120 h. Another overflight
on 28 February confirmed that the dome 82 was still occupying the
crater. Between 26 February and 10 March 2019, 48 explosions were
reported by CENAPRED (Figure 5), some of which (e.g., 1 and
9 March 2019) expelled incandescent blocks on the external slopes of
the cone hundreds of meters downslope. This explosive activity
resulted in 69 VAAC alerts, 29 of which occurred between
5–8 March. Strong degassing during the overflight on 7 March
did not allow observations of the dome destruction. An image of
8 March in Google Earth offers a clear view on more than half of the
crater floor, showing a stepped profile of the walls and clear marks of
several explosions, confirming that the dome was destroyed during
this explosion sequence. Eleven additional and sometimes large y
g
g
y
p
p
In this contribution, we present 2.5 years (2017 to mid-2019)
of high quality multispecies compositional data of Popocatépetl’s
plume combining ground-based DOAS and FTIR direct-sun
measurements, with a focus over the most voluminous dome
growth and destruction episode (February–April 2019) of the
study period. The measurements were performed from the
Altzomoni
(ALTZ)
observatory,
one
of
the
few
stations
pertaining to the Network for the Detection of Atmospheric
Composition Change (NDACC) and to the Pandonia Gobal
Network (PGN) located near a persistently degassing volcano. The site instruments (a high resolution (HR)-FTIR coupled with
a DOAS and a Pandora spectrometers) and measurements benefit
from standardized protocols, intercalibrations and comply with
the
measurement
quality
requirements. 1 Introduction We
studied
the
variability of both the primary and secondary species and of
their ratios (i.e., BrO/SO2, BrO/HCl, HCl/SO2, HF/SO2, SiF4/
SO2) in the volcanic plume during different phases of the dome
growth and relate it with the degassing processes and in-plume
reactions. Frontiers in Earth Science 1 Introduction The
conversion of SO2 to sulfate aerosols can be significant over
long-term
observation
periods
under
humid
conditions
(Rodriguez et al., 2008), but seems to play a minor role in ash
poor-plumes over a period of minutes to hours (McGonigle et al.,
2004; Voigt et al., 2014). On the other hand, the characterization of
the halogen activation mechanisms in volcanic plumes and their
reaction kinetics is critical for both long and short-term studies,
affecting the variability of primary species ratios and degassing
balances. Many primary magmatic species (H2O, CO2, SO2, HCl, HF,
HBr, CO, OCS, H2S) and their derivatives (BrO, OClO, SiF4) can be
remotely measured using satellite and ground-based spectroscopic
techniques (from ultraviolet to thermal infrared) in both active and
passive configurations. A complete review of the different ground-
based techniques used to date for measuring and imaging the
different species in volcanic plumes is given in Platt et al. (2018). The use of DOAS instruments to measure SO2 and reactive
halogen species (BrO, OClO) has proven to be simple and effective,
providing the possibility to automatize current instrumentation and
create robust monitoring networks (Galle et al., 2010) such as the
Network for Observation of Volcanic and Atmospheric Change
(NOVAC). BrO/SO2 ratios are measured by DOAS instruments
using scattered sunlight (Gutmann et al., 2018) and Gliß et al. (2015)
additionally measured the OClO species in the volcanic plume of
Mount Etna
using
Multi-AXis DOAS
(MAX-DOAS). FTIR
spectroscopy has long been used to study the compositional
variability (SO2, HCl, HF, SiF4, CO2, OCS, CO, H2S) of volcanic Among the halogen secondary species, BrO was measured for
the first time in a volcanic plume at Soufrière Hills (Montserrat) by
Bobrowski et al. (2003), and then at a number of other volcanoes
(Oppenheimer et al., 2006; Bobrowski and Platt, 2007; Kern et al.,
2009; Heue et al., 2011; Rix et al., 2012; Donovan et al., 2014; Gliß
et al., 2015; Dinger et al., 2018; Warnach et al., 2019; Dinger et al.,
2021) including Popocatépetl (Boichu et al., 2011; Bobrowski and
Giuffrida, 2012; Platt and Bobrowski, 2015; Fickel and Delgado
Granados, 2017). Recent models succeeded in reproducing some
observations of plume compositions at different plume ages 02 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 10.3389/feart.2023.1062699 degassing (Taquet et al., 2019). 1 Introduction The long duration of the ongoing
eruption (>28 years at the time of writing) and its relatively high
degassing rate are ideal to explore the link between the volcanic
activity and the variability of the plume composition, which is one of
the few observable manifestations of the magmatic processes. The
previously reported short- or long-term time series of the HCl, HF,
SO2, CO2, SiF4 species abundance and of their ratios in the
Popocatépetl’s plume (Goff et al., 1998; Love et al., 1998; Goff
et al., 2001; Gerlach, 2004; Stremme et al., 2011; Stremme et al.,
2012; Taquet et al., 2017; Taquet et al., 2019) gave valuable
information on the behavior of these species with respect to the
volcanic activity. However, the optimal combination of eruptive and
meteorological conditions for measurements is seldom met multiple
times during a single dome growth episode. We focused here on a
2.5-month time-series spanning the full cycle of construction and
destruction of dome 82 (according to reports of the local institution
in charge of the monitoring, i.e., Centro Nacional de Prevención de
Desastres or CENAPRED). Following a relatively intense extrusive
activity during September–December 2018, with the succession of
the
dome
emplacement-destruction
cycles
#80
and
#81
(CENAPRED daily reports; Caballero Jimenez et al., 2019),
January 2019 is marked by a powerful explosion on day 22,
apparently without a dome occupying the crater. Such explosions
are rare, but they became more frequent following the earthquakes
of 2017 (Boulesteix et al., 2022). A crater overflight was operated on
day 27, which confirmed that the crater was still empty and that the
conduit was permeable. Caballero and Valderrama (2020) indeed
report the observation of a thermal anomaly coinciding with that of
Sentinel-2 data (see, e.g., Mounts processing, Valade et al., 2019) and
with the emission source of passive degassing plumes, as already
proposed in previous thermal imaging studies at Popocatépetl (e.g.,
Wright et al., 2002). Based on CENAPRED reports, following this
explosion and before the onset of the dome growth episode, daily
averages of ~15 min of tremor, of 1.3 Volcano Tectonic (VT)
earthquakes and of 150 exhalations (local term for low intensity
explosions with little impulsive character) were recorded. On
15 February 2019, at approximately 2:55 UTC (14 February at
20:55, local time), the first 6 h-long LPs-tremor-explosions
segment of this dome growth episode started. 1 Introduction This allows straightforward comparisons of the retrieved vertical
columns (VCs) and robust calculation of multiple volcanic species
ratios, making this method one of the most reliable and adaptable for
studying long-term variability of the volcanic plume composition. degassing (Taquet et al., 2019). The long duration of the ongoing
eruption (>28 years at the time of writing) and its relatively high
degassing rate are ideal to explore the link between the volcanic
activity and the variability of the plume composition, which is one of
the few observable manifestations of the magmatic processes. The
previously reported short- or long-term time series of the HCl, HF,
SO2, CO2, SiF4 species abundance and of their ratios in the
Popocatépetl’s plume (Goff et al., 1998; Love et al., 1998; Goff
et al., 2001; Gerlach, 2004; Stremme et al., 2011; Stremme et al.,
2012; Taquet et al., 2017; Taquet et al., 2019) gave valuable
information on the behavior of these species with respect to the
volcanic activity. However, the optimal combination of eruptive and
meteorological conditions for measurements is seldom met multiple
times during a single dome growth episode. We focused here on a
2.5-month time-series spanning the full cycle of construction and
destruction of dome 82 (according to reports of the local institution
in charge of the monitoring, i.e., Centro Nacional de Prevención de
Desastres or CENAPRED). Following a relatively intense extrusive
activity during September–December 2018, with the succession of
the
dome
emplacement-destruction
cycles
#80
and
#81
(CENAPRED daily reports; Caballero Jimenez et al., 2019),
January 2019 is marked by a powerful explosion on day 22,
apparently without a dome occupying the crater. Such explosions
are rare, but they became more frequent following the earthquakes
of 2017 (Boulesteix et al., 2022). A crater overflight was operated on
day 27, which confirmed that the crater was still empty and that the
conduit was permeable. Caballero and Valderrama (2020) indeed
report the observation of a thermal anomaly coinciding with that of
Sentinel-2 data (see, e.g., Mounts processing, Valade et al., 2019) and
with the emission source of passive degassing plumes, as already
proposed in previous thermal imaging studies at Popocatépetl (e.g.,
Wright et al., 2002). frontiersin.org 2 Popocatépetl volcano and
measurement site Popocatépetl is an andesitic-dacitic stratovolcano of the trans-
Mexican volcanic belt, located 70 km SE of Mexico City. Its past
activity comprises destructive episodes of lateral flank collapses and
plinian phases. Its historical activity is characterized by cycles of
extrusion
of
lava
domes
and
their
destruction
through
a
combination of compaction and explosions (Campion et al.,
2018), interspersed with quiet periods with continuous passive 03 frontiersin.org 10.3389/feart.2023.1062699 Taquet et al. beam and reflects it into the container (see, e.g., Plaza-Medina et al.,
2017; Taquet et al., 2019). A part of the solar beam is directed by
mirrors toward the entrance of the S2000 DOAS optical fiber and the
other part towards the FTIR spectrometer aperture (Figure 1). beam and reflects it into the container (see, e.g., Plaza-Medina et al.,
2017; Taquet et al., 2019). A part of the solar beam is directed by
mirrors toward the entrance of the S2000 DOAS optical fiber and the
other part towards the FTIR spectrometer aperture (Figure 1). explosions occurred until the end of March, with a large outreach of
the blocks that set fire to the highest altitude vegetation on 27 March
2019. The explosions ceased during April and the activity declined
significantly with, e.g., a threefold decrease of the number of
exhalations. Dome forming activity only returned in mid-July
(Caballero and Valderrama, 2020). p
p
p
g
The Bruker IFS-125HR FTIR spectrometer can provide a
maximal
optical
path
difference
(OPDmax)
of
257 cm
corresponding
to
a
spectral
resolution
of
0.0035
cm−1
(resolution = 0.9/OPDmax). The spectrometer operates with
either KBr or CaF2 beam splitters, three different detectors
(MCT, InSb and InGaAs) and a set of seven optical filters
installed in a rotating wheel. A remote control liquid-nitrogen
filling system assures that the MCT and InSb detectors are
properly cooled. The measurement routine was programmed to
acquire high (0.005 cm−1), moderate (0.02 cm−1) and low (0.1 cm−1)
resolution spectra in the near-infrared (NIR) and mid-infrared
(MIR) spectral ranges using the different filters. The complete
normal-mode sequence lasts ~20 min. Every time the volcanic
plume is observed in the optical path of the instrument, the
acquisition is switched to a faster sequence, keeping only the NIR
and the low resolution acquisitions with the NDACC filters three
and 6 (~5 min for a complete sequence). The field of view varies
between 2 and 4 mrad, depending on the used optical filter and
measurement configuration. 2 Popocatépetl volcano and
measurement site Currently, there are several strategies to remotely monitor the
emission of gases from Popocatépetl volcano, both in mobile
(Campion et al., 2018) and static configurations (Taquet et al., 2017;
Schiavo et al., 2019; Taquet et al., 2019; Arellano et al., 2021). Since 2012,
the installation of the ALTZ Atmospheric Observatory in the Izta-Popo-
Zoquiapan National Park, 11 km from the Popocatépetl crater, allowed
the
ICAyCC-UNAM
team
to
perform
not
only
atmospheric
measurements, but also volcanic plume measurements using solar,
lunar absorption (Stremme et al., 2012; Krueger et al., 2013; Taquet
et al., 2019) and thermal emission (Stremme et al., 2011; Stremme et al.,
2012; Taquet et al., 2017) FTIR spectroscopic techniques. This site
benefits from a number of advantages such as open and direct view
to the volcano, a high altitude to minimize the interferences from water
vapor and anthropogenic pollutants, a safe distance from the crater and
from abundant ash-falls, and the availability of operational services
(power and communication). Apart from the HR-FTIR spectrometer (a
Bruker IFS-125HR), the station was also equipped with meteorological
instruments, visible cameras, in-situ gas analyzers for atmospheric and
environmental monitoring, among other instrumentation. Additionally,
remote sensing equipments including a UV camera, a Pandora, another
DOAS spectrometer coupled with the HR-FTIR instrument and a low-
resolution FTIR spectrometer (a Bruker EM27-MIR), occasionally allow
measuring
Popocatépetl’s
plume
composition
with
different
configurations (solar absorption, thermal emission, UV scattered-light
mode). For the DOAS measurements, spectra were collected using an
Ocean Optics S2000 spectrometer (spectral resolution of ~0.6 nm)
combined with an optical fiber and a band-pass filter (Hoya U330)
which blocks visible light with wavelength between 360 and 640 nm. S2000 DOAS spectra were acquired using the DOASIS software
(Kraus, 2006) adjusting automatically the exposure time to the Sun
intensity. Collected spectra were averaged so that a final collection
time of 1 min was achieved. The container is thermally stabilized to provide optimal
operating conditions for the different instruments. A remote-
controlled dome cover and a microwave communication system
(50 km line of sight to the university campus) allow us to fully
control and perform the measurements remotely. 3.2 The Pandora measurements A Pandora instrument (number 65) was implemented in the ALTZ
station in 2018, and the data products have been available since January
2019. The Pandora head sensor is installed on the rooftop of the
container, and points accurately to the Sun with a 2.5° field of view
(FWHM SunFOV) using its own solar tracking system (SciGlob/NASA
tracker). The head sensor is connected through an optical fiber to an
Avantes spectrometer located in the thermally stabilized FTIR container. The spectrometer (model Avantes AvaSpec ULS 2048x64) has a 0.6 nm
resolution (FWHM) covering the spectral range from 280 to 530 nm. Different bandpass and neutral density filters can be chosen and
combined inside the optical head by two filter wheels, so that a large
range of intensity is covered by the instrument and direct and indirect
Sun or Moon light can be recorded. The measurement routine starts
automatically in the morning, collecting direct solar spectra, but also
hourly indirect sunlight spectra (zenith sky measurements, pointing
above the crater and MAXDOAS-type measurements). During night
time, lunar spectra are recorded, if possible. Spectra are sent to the central
server of the PANDONIA Global Network (PGN, https://www. 3 Instrumental set-up and analysis
procedures In
this
study,
we
present
direct
solar
absorption
measurements performed from the ALTZ station using the
coupled
HR-FTIR/DOAS
set-up,
and
the
Pandora
spectrometer over a period of 2.5 years (2017-mid 2019). Plume measurements with this configuration are possible on
days with clear sky conditions and only during favorable wind
conditions, at times when the volcanic plume and the instrument
are aligned with the Sun. Since both the DOAS instrument and
Pandora are based on the DOAS technique, we hereafter referred
to the DOAS integrated in the coupled HR-FTIR/DOAS set-up as
“S2000 DOAS,” and as “Pandora” for the Pandora instrument. In
the same way, the DOAS measurements made by the Pandora will
be referred to as “Pandora measurements” while the DOAS
measurements of the HR-FTIR/DOAS set-up will be referred
to as “S2000 DOAS measurements.” 3.1 The combined FTIR-DOAS solar
absorption measurements A solar tracker (Camtracker; Gisi et al., 2011; Gisi et al., 2012;
Gisi, 2012) located on the roof of the station collects the direct solar Frontiers in Earth Science 04 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 1
Set-up of the Pandora, HR-FTIR and S2000 DOAS instruments at the ALTZ station. (A) Global view of the instruments installed on the roof of the
ALTZ station. (B) Instrumental set-up for the combined DOAS-FTIR measurements. FIGURE 1 FIGURE 1
Set-up of the Pandora, HR-FTIR and S2000 DOAS instruments at the ALTZ station. (A) Global view of the instruments installed on the roof of the
ALTZ station. (B) Instrumental set-up for the combined DOAS-FTIR measurements. TABLE 1 Retrieval settings used for the S2000 DOAS and Pandora spectra analysis. Instrument/Target
species
Spectral
windows (nm)
Interference
species
Cross sections
S2000 DOAS
SO2
312.0–326.8
O3
SO2 at 298 K (Vandaele et al., 2009), O3 at 221 K (Burrows et al., 1999)
BrO
330.6–356.3
SO2, O3, O4, NO2, OClO
BrO (Wilmouth et al., 1999), SO2 at 298 K (Hermans et al., 2009), O3 at 221 K
(Burrows et al., 1999), O4 (Hermans et al., 2003), NO2 at 293 K (Voigt et al.,
2002), OClO at 293 K (Bogumil et al., 2003)
Pandora
SO2
306.0–326.4
O3, NO2, HCHO,
HONO, BrO
SO2 (Vandaele et al., 2009), O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al.,
1998), HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000) and BrO
(Wilmouth et al., 1999)
BrO (from the PGN HCHO
algorithm “fus5”)
322.5–359.2
SO2, O3, NO2, HCHO,
HONO
O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al., 1998), SO2 (Vandaele et al.,
2009), HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000), and BrO
(Wilmouth et al., 1999) pandonia-global-network.org/) where the data are processed and
analyzed. 3.3 Data analysis
3.3.1 UV analysis
3.3.1.1 Pandora data processing
In this study, we utilized the L2-rsus1p1-8 products available in
the PGN server, in which the SO2 total columns are retrieved using
the code “sus1” of the BlickSFA algorithm (Cede et al., 2021). The
PGN retrievals follow the Cede et al. (2021) Total Optical
Absorption Spectroscopy approach. It is based on non-linear
least square fitting of the absorption cross-sections of multiple
atmospheric absorbers to the spectra, and uses a synthetic solar
absorption reference spectrum, constructed from the measurements
of the same spectrometer. 3.1 The combined FTIR-DOAS solar
absorption measurements SO2 is fitted in the 306–326.4 nm spectral
range (Table 1) with a fourth order smoothing polynomial function
and includes the following cross sections: SO2 (Vandaele et al.,
2009), O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al., 1998),
HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000) and
BrO (Wilmouth et al., 1999). Additionally, we explored the BrO
TABLE 1 Retrieval settings used for the S2000 DOAS and Pandora spectra analysis. Instrument/Target
species
Spectral
windows (nm)
Interference
species
Cross sections
S2000 DOAS
SO2
312.0–326.8
O3
SO2 at 298 K (Vandaele et al., 2009), O3 at 221 K (Burrows et al., 1999)
BrO
330.6–356.3
SO2, O3, O4, NO2, OClO
BrO (Wilmouth et al., 1999), SO2 at 298 K (Hermans et al., 2009), O3 at 221 K
(Burrows et al., 1999), O4 (Hermans et al., 2003), NO2 at 293 K (Voigt et al.,
2002), OClO at 293 K (Bogumil et al., 2003)
Pandora
SO2
306.0–326.4
O3, NO2, HCHO,
HONO, BrO
SO2 (Vandaele et al., 2009), O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al.,
1998), HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000) and BrO
(Wilmouth et al., 1999)
BrO (from the PGN HCHO
algorithm “fus5”)
322.5–359.2
SO2, O3, NO2, HCHO,
HONO
O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al., 1998), SO2 (Vandaele et al.,
2009), HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000), and BrO
(Wilmouth et al., 1999) TABLE 1 Retrieval settings used for the S2000 DOAS and Pandora spectra analysis. PGN retrievals follow the Cede et al. (2021) Total Optical
Absorption Spectroscopy approach. It is based on non-linear
least square fitting of the absorption cross-sections of multiple
atmospheric absorbers to the spectra, and uses a synthetic solar
absorption reference spectrum, constructed from the measurements
of the same spectrometer. SO2 is fitted in the 306–326.4 nm spectral
range (Table 1) with a fourth order smoothing polynomial function
and includes the following cross sections: SO2 (Vandaele et al.,
2009), O3 (Serdyuchenko et al., 2013), NO2 (Vandaele et al., 1998),
HCHO (Meller and Moortgat, 2000), HONO (Stutz et al., 2000) and
BrO (Wilmouth et al., 1999). Additionally, we explored the BrO Frontiers in Earth Science 3.3.1 UV analysis
3.3.1.1 Pandora data processing In this study, we utilized the L2-rsus1p1-8 products available in
the PGN server, in which the SO2 total columns are retrieved using
the code “sus1” of the BlickSFA algorithm (Cede et al., 2021). The Frontiers in Earth Science 05 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 2
Example of fit of SO2 analyzed in the 312.0–326.8 nm spectral window from the S2000 DOAS measurements at ALTZ station performed on
20 February 2019 using (A) the measured reference (Rmeas) and (C) the RSAO reference. The corresponding residues are presented in (B,D), respectively. The retrieved slant column density (SCD, in molec/cm2) is shown in the legend. FIGURE 2
Example of fit of SO2 analyzed in the 312.0–326.8 nm spectral window from the S2000 DOAS measurements at ALTZ station performed on
20 February 2019 using (A) the measured reference (Rmeas) and (C) the RSAO reference. The corresponding residues are presented in (B,D), respectively. The retrieved slant column density (SCD, in molec/cm2) is shown in the legend. band extinction. Intensity offset, non-linear wavelength shift and
first order stretching corrections were included in the DOAS fit
settings. In contrast to the scattered light radiance measurements,
the direct-sun configuration is not affected by the Ring effect
(Herman et al., 2009), so that we did not include it in the
retrieval. The retrieval of SO2 was performed applying two
different
strategies
based
on
two
different
references. The
“DOAS_v1” product is produced using the high resolution solar
reference spectrum from Chance and Kurucz (2010) (“sao2010_
solref_air.dat”: hereafter “RSAO”), and the “DOAS_v2” product
obtained with a measured clear sky reference spectrum (hereafter
“Rmeas”) acquired on 17 February 2019 with a solar zenith angle
(SZA) close to zero. Fits and residuals obtained from the two
strategies are presented in Figure 2. The use of a measured
reference significantly improves the SO2 fits, limiting the effect of
interfering atmospheric gases and avoiding structures of the solar
spectrum. To correct the I0 effect (Platt and Stutz, 2008), QDOAS
performs an on-line approximation by correcting the cross sections
with the high resolution solar Fraunhofer spectrum, considering a
fixed predefined SCDI0. 3.3.1 UV analysis
3.3.1.1 Pandora data processing The I0 effect was corrected as following for
the full time series: six SO2 retrievals were performed using the
QDOAS I0-correction algorithm for fixed SO2 slant column values
of 0.0, 1.0 × 1018, 2.0 × 1018, 3.0 × 1018, 4.0 × 1018, 5.0 × 1018 molec/
cm2 (the latter is close to the maximum uncorrected slant slant columns density (SCD) product, one of the outputs of the PGN
“fus5” algorithm, officially used to retrieve the HCHO species in the
322.5–359.2 nm spectral range. In this algorithm, all of the O3, NO2,
SO2, HCHO, HONO, BrO species are fitted using the cross sections
reported in Table 1. We converted the BrO SCDs to VCs to be
compared with the S2000 DOAS products. Further details about the
fitting procedures and calibrations can be found in Cede et al. (2021). Frontiers in Earth Science 3.3.2 FTIR analysis column). For each uncorrected datum, the most adequate I0-
correction was interpolated from the five retrievals, depending
on their slant column values. Only one iteration was required,
since the difference between the slant columns obtained after
the second and third iteration was lower than 1% of the retrieval
error, so that the convergence was achieved after the second
iteration. The O3 cross section was also I0-corrected using the
fixed slant column value of 1.0 × 1019 molec/cm2, measured at
Altzomoni by Plaza-Medina et al. (2017). SO2, HCl, HF, and SiF4 are retrieved from the high, medium and
low resolution FTIR spectra using the PROFFIT9.6 retrieval code
(Hase et al., 2004). Retrieval settings and strategies are detailed in
Taquet et al. (2019) and summarized in Table 2. Examples of SO2,
HCl, HF, and SiF4 fits are given in supplementary material
(Supplementary Figures S3–S7). Importantly, this set-up allows
the sequential analysis of the following pairs of gas species: SO2
and HCl (InSb detector, NDACC filter 3), HF and HCl (InGaAs
detector, Open) and SO2 and SiF4 (MCT detector, NDACC Filter 6),
corresponding to different spectral regions (see Table 2). The
different SO2 (and HCl) products obtained using the different
spectral ranges were calibrated and combined according to our
previous study (Taquet et al., 2019). The only change with
respect to Taquet et al. (2019) is the inclusion of the new
spectral parameters for SiF4 measured by Boudon et al. (2020),
allowing retrievals of SiF4 from both high (0.005 cm−1) and low
(0.1 cm−1) resolution spectra. An estimation of the total error in the
retrieved total columns is calculated by the PROFFIT code for all
retrievals and includes systematic and statistical errors from
different sources, such as the noise, baseline with channeling,
Instrumental
Line
Shape
(ILS),
Line
Of
Sight
(LOS)
and
spectroscopic parameters. The error analysis for this ALTZ
instrument is described in more detail by Plaza-Medina et al. (2017). The channeling error (frequencies and amplitudes) for
the instrument has been recently characterized and described by
Blumenstock et al. (2020). BrO was analyzed in the 330.6–356.3 nm spectral window
(Table 1) according to Gliß et al. 3.3.1.2 S2000 DOAS data processing The UV spectra from the S2000 DOAS spectrometer were
analyzed using the QDOAS software version 2.111 (Danckaert
et al., 2013), developed by BIRA-IASB, which retrieves the SCDs
of trace gases from the measured spectra, using a Marquardt
Levenberg Non-Linear Least Squares (NLLS) Fitting algorithm. Wavelength calibration and slit function files were created with a
low-density mercury lamp and were used as inputs in the QDOAS
retrieval. An electronic offset correction was also measured and
applied in the spectra analysis. SO2 was retrieved in the
312.0–326.8 nm spectral window (Table 1) following the Butz
et al. (2017) procedure. In the retrieval, two cross sections were
used: SO2 at 298 K (Vandaele et al., 2009) and O3 at 221 K (Burrows
et al., 1999) as an interfering gas and a fifth-order polynomial
function was included in the fitting routine to remove broad Frontiers in Earth Science 06 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 3
Example of fit of BrO analyzed in the (330.6–356.3 nm) spectral window from the DOAS measurements at ALTZ station performed on 20 February
2019. The retrieved slant column density (SCD, in molec/cm2) is shown in the legend. FIGURE 3
Example of fit of BrO analyzed in the (330.6–356.3 nm) spectral window from the DOAS measurements at ALTZ station performed on 20 February
2019. The retrieved slant column density (SCD, in molec/cm2) is shown in the legend. 3.3.2 FTIR analysis Fit parameters are reported in Table 3. Spectral windows (cm–1), (NDACC filter number, detector)
Spectral resolution (cm–1)
Retrieval strategy
SO2
(2480.0–2520.0), (# 3, InSb)
0.1 and 0.005
Sc
(1120.0–1180.0), (# 6, MCT)
HCl
(2727.0–2728.5); (2775.0–2776.50); (2818.75–2820.35); (2820.75–2822.35); (2843.0–2844.4);
0.1 and 0.005
TP
(2903.35–2904.85); (2923.0–2924.50); (2925.0–2926.75); (2942.0–2943.5); (2960.3–2961.825);
(2962.3–2964.0); (2995.0–2996.5), (# 3, InSb)
(5738.0–5740.0); (5767.0–5767.8); (5779.2–5779.9), (Open, InGaAs)
0.02
HF
(3999.0–4003.5); (4036.5–4041.0), (Open, InGaAs)
0.1, 0.0075 and 0.02
TP
SiF4
(1015.0–1035.0), (#6, MCT)
0.1 and 0.005
Sc FIGURE 4
(A) Correlation plot of SO2 VCs obtained from S2000 DOAS, Pandora, and FTIR instruments. (B) Determination of calibration factors with respect to
the FTIR SO2 product. Fit parameters are reported in Table 3. G FIGURE 4
(A) Correlation plot of SO2 VCs obtained from S2000 DOAS, Pandora, and FTIR instruments. (B) Determination of calibration factors with respect to
the FTIR SO2 product. Fit parameters are reported in Table 3. TABLE 3 Linear fit parameters obtained from the intercomparison of the FTIR, S2000 DOAS and Pandora SO2 products. R2 stands for the determination coefficient. TABLE 3 Linear fit parameters obtained from the intercomparison of the FTIR, S2000 DOAS and Pandora SO2 products. R2 stands for the determination coefficient. Products
Regression label
Slopea
Intercept (molec/cm2)
R2
FTIR vs. DOAS_v1
y1
1.08 ± 0.02
(−4.48 ± 1.05) × 1016
0.90
Pandora vs. DOAS_v1
y2
0.92 ± 0.01
(−5.56 ± 0.91) × 1016
0.94
DOAS_v2 vs. DOAS_v1
y3
0.85 ± 0.01
(−5.34 ± 0.16) × 1016
0.99
Pandora vs. FTIR
y4
0.88 ± 0.02
(−8.17 ± 4.04) × 1015
0.90
DOAS_v2 vs. FTIR
y5
0.82 ± 0.01
(−1.13 ± 0.40) × 1015
0.90
aErrors in the slope and intercept are given with a confidence interval of 0.95. meters obtained from the intercomparison of the FTIR, S2000 DOAS and Pandora SO2 products. R2 stands for the determination coefficient The so-generated airmass-dependent function was used for the
whole time series and the intraday variability of SiF4 was
removed. Apart from increasing the number of SiF4 data in the
time series, the SiF4 analysis from the high resolution spectra allows
obtaining much lower total errors in the VCs than that for the low
spectral resolution. For SiF4 total columns higher than 2 ×
1015 molec/cm2, the SiF4 VCs total errors are lower than 20%,
while those obtained from the low resolution spectra occasionally
reached 60%. 3.3.2 FTIR analysis (2015), including the following
cross sections: BrO at 298 K (Wilmouth et al., 1999), SO2 at 298 K
(Hermans et al., 2009), O3 at 221 K (Burrows et al., 1999), O4
(Hermans et al., 2003), NO2 at 293 K (Voigt et al., 2002), OClO at
293 K (Bogumil et al., 2003) and a third-order polynomial function
was included in the fitting routine. The I0 correction was applied to
the O3 cross-section. For the reference spectra, the same measured
spectrum (Rmeas) as for the SO2 retrieval was used. OClO was
tentatively quantified from the same spectral window and settings,
but it was not detected during the study period. Figure 3 shows an
example of a BrO fit from DOAS measurements at ALTZ station. Data affected by the presence of clouds or without volcanic plume
contribution
were
discarded
using
the
SO2
slant
column
uncertainties calculated by the QDOAS algorithm (based on the
fit uncertainties) from the SO2 analysis. We only consider data for
which the relative SO2 retrieval uncertainties were ≤20%. DOAS SO2
and BrO SCDs were then converted into VCs to be compared with
the FTIR and Pandora products, by dividing them by the SZA-
dependent air mass factor (1/cos (SZA)). To evaluate the bias resulting from the use of the new
spectroscopy in the SiF4 analysis, we re-evaluated the SiF4 VCs Frontiers in Earth Science 07 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 TABLE 2 Spectral ranges and retrieval strategies (detailed in Taquet et al., 2019) applied for the SO2, HCl, HF, and SiF4 analysis from the FTIR data. TP refers to a
profile retrieval with a Tikhonov-Phillips regularization, and Sc to a scaling retrieval. p
g
g
(
q
,
) pp
2,
,
,
4
y
profile retrieval with a Tikhonov-Phillips regularization, and Sc to a scaling retrieval. Spectral windows (cm–1), (NDACC filter number, detector)
Spectral resolution (cm–1)
Retrieval strategy
SO2
(2480.0–2520.0), (# 3, InSb)
0.1 and 0.005
Sc
(1120.0–1180.0), (# 6, MCT)
HCl
(2727.0–2728.5); (2775.0–2776.50); (2818.75–2820.35); (2820.75–2822.35); (2843.0–2844.4);
0.1 and 0.005
TP
(2903.35–2904.85); (2923.0–2924.50); (2925.0–2926.75); (2942.0–2943.5); (2960.3–2961.825);
(2962.3–2964.0); (2995.0–2996.5), (# 3, InSb)
(5738.0–5740.0); (5767.0–5767.8); (5779.2–5779.9), (Open, InGaAs)
0.02
HF
(3999.0–4003.5); (4036.5–4041.0), (Open, InGaAs)
0.1, 0.0075 and 0.02
TP
SiF4
(1015.0–1035.0), (#6, MCT)
0.1 and 0.005
Sc
FIGURE 4
(A) Correlation plot of SO2 VCs obtained from S2000 DOAS, Pandora, and FTIR instruments. (B) Determination of calibration factors with respect to
the FTIR SO2 product. Frontiers in Earth Science 3.3.2 FTIR analysis from the most significant SiF4 events recorded at ALTZ on 06 March
2015 (Taquet et al., 2019). The re-analysis of the low resolution
spectra provided SiF4 columns slightly lower than previously
published, by a factor of (0.94 ± 0.03) with a determination
coefficient (R2) of 0.96. The spectral fits are only slightly
improved, but still a systematic artefact depending on the SZA,
likely due to spectral interferences, was observed in the new SiF4
time series. An empirical post-correction was applied to reduce this
effect. A mean SiF4 VC was calculated for all SZA (using bins of 5°). Frontiers in Earth Science 08 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 5
SO2, HCl, HF, BrO, and SiF4 VCs measured using S2000 DOAS, Pandora and FTIR instruments during the February–April 2019 period. The daily
number of measurement hours and the number of hours affected by the presence of the volcanic plume are presented in the upper panel. FIGURE 5
SO2, HCl, HF, BrO, and SiF4 VCs measured using S2000 DOAS, Pandora and FTIR instruments during the February–April 2019 period. The daily
number of measurement hours and the number of hours affected by the presence of the volcanic plume are presented in the upper panel. SO2, HCl, HF, BrO, and SiF4 VCs measured using S2000 DOAS, Pandora and FTIR instruments during the February–April 2019 period. The daily
number of measurement hours and the number of hours affected by the presence of the volcanic plume are presented in the upper panel. The quality control procedure described in Taquet et al. (2019) is
applied for the different target species and an additional filter
selecting SO2 columns with relative total error <50% discards
data with important uncertainties (presence of clouds, low
signal, etc.). Table
3. SO2
vertical
columns
obtained
from
the
DOAS
measurements using the two different strategies described in
Section 3.3.1 (DOAS_v1 vs. DOAS_v2) are perfectly correlated
(Figure 4A, in green) with a slope of 0.85 ± 0.01 (y3) and a
determination coefficient of 0.99. In comparison with the DOAS
products (measured with the highest temporal frequency), a good
correlation (R2 ≥0.90) is observed in Figure 4A (Table 3) for the
FTIR (y1) and Pandora (y2) SO2 VCs products. The DOAS_v2 and
the calibrated official Pandora SO2 products are consistent, with
respective bias of 0.85 ± 0.01 (R2 = 0.99) and 0.92 ± 0.01 (R2 = 0.94)
to the DOAS_v1 product. 3.3.2 FTIR analysis Using the pre-calibrated RSAO reference
(DOAS_v1) in the QDOAS retrieval instead of a “measured
reference” allows obtaining “absolute” vertical columns directly
comparable
with
those
obtained
from
other
instruments
(Herman et al., 2009). However, the use of a measured reference
(DOAS_v2 product) has the advantage to considerably reduce the
noise in the absorbance spectrum (Figure 2) improving the fits and
minimizing the total errors. The possible contamination of the Frontiers in Earth Science 4.1.2 BrO VCs from S2000 DOAS and Pandora The Pandora and the DOAS BrO detections occurred on the
same days (on the 16, 18 and 20/02/2019) during our 2.5-month
time series. At the intraday scale (Figure 6), the BrO VCs from the
two instruments agree within their uncertainties. However, the two
time series are not well correlated (R2 of 0.74) and show a slope
DOAS/Pandora around 1.2. The PGN “fus5” retrieval is indeed not
optimized for recovering BrO which is not a PGN official product, so
that we will not consider it further for combination with the other
products. Because of our longtime record of FTIR measurements at the
ALTZ station (Stremme et al., 2009; Stremme et al., 2011; Stremme
et al., 2012; Taquet et al., 2017; Taquet et al., 2019), the FTIR SO2
product is hereafter used as our absolute reference, and the
calibration factors defined in Figure 4B and reported in Table 3
were applied to the Pandora and DOAS_v2 products. The
intercomparison was also examined at the intraday scale for
different days, as shown in Figures 6A, B and Figures 7A, B,
capturing explosive and passive degassing events during the
dome growth episode of February 2019. The superimposition of
the SO2 VCs (median difference <12%) from the three instruments
highlights the possibility to combine them, increasing the time Frontiers in Earth Science 4.1 Intercomparison between the different
instruments 4.1.1 SO2 VCs from S2000 DOAS, FTIR, and
Pandora 4.1.1 SO2 VCs from S2000 DOAS, FTIR, and
Pandora The intercomparison between the SO2 VCs obtained from the
FTIR, S2000 DOAS, and Pandora instruments is shown in Figure 4. Data pairs were chosen with a coincidence criterium equal to 1 min. Equations of regression lines (including uncertainties) are given in Frontiers in Earth Science 09 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 6
SO2, HCl, HF, BrO, SiF4 VCs intraday time series captured on the (A) 16 February 2019 and (B) 18 February 2019 using the S2000 DOAS, FTIR and
Pandora instruments. For SiF4, open and full circles correspond to the low and high resolution products, respectively. The BrO from the Pandora
instrument [third (A) and (B) panels, in green] corresponds to a non-official PNG product and is only indicative of the S2000 DOAS and Pandora BrO VCs
agreement. The lower panel shows the time series of the HCl/SO2, HF/SO2 (×5) and BrO/SO2 ratios. See text for more details. FIGURE 6
SO2, HCl, HF, BrO, SiF4 VCs intraday time series captured on the (A) 16 February 2019 and (B) 18 February 2019 using the S2000 DOAS, FTIR and
Pandora instruments. For SiF4, open and full circles correspond to the low and high resolution products, respectively. The BrO from the Pandora
instrument [third (A) and (B) panels, in green] corresponds to a non-official PNG product and is only indicative of the S2000 DOAS and Pandora BrO VCs
agreement. The lower panel shows the time series of the HCl/SO2, HF/SO2 (×5) and BrO/SO2 ratios. See text for more details. measured reference, however, has to be evaluated. For this, the
measured reference used in the DOAS_v2 retrieval was pre-
evaluated regarding its SO2 content using a literature solar
background
spectrum
as
a
Fraunhofer
reference
spectrum
(Chance and Kurucz, 2010) which was convolved with the
instrumental line spread function. The resulting SO2 abundance
of 9.1 × 1016 molec/cm2 is of the same order of magnitude as the
mean fit error (5.0 ± 4.0 × 1016 molec/cm2) of the SCDs and can be
considered negligible in our measurement conditions. resolution of the SO2 time series and providing more pairs of
coincident data for the calculation of the ratios. 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume The combination of the DOAS-FTIR instruments allowed the
joint detection of more than 80 volcanic events during the whole
measurement period (01/2017–05/2019), but the most significant 10 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 7
Same as Figure 6 for (A) 20 february 2019 and (B) 11 March 2019. FIGURE 7
Same as Figure 6 for (A) 20 february 2019 and (B) 11 March 2019. dome destruction; and P3 (16 March–18 April 2019) with low
volcanic activity and without dome occupying the crater. SO2, HCl, HF, BrO, and SiF4 columns were recorded during the
main dome growth episodes. BrO was only detected during two
(December 2017 and February 2019) of the 11 dome growth and
destruction episodes that occurred during our 2.5-year time
series (public CENAPRED annual and daily reports: https://
www.cenapred.unam.mx/reportesVolcanGobMX/). The Appreciable SO2 columns were detected from 6 February 2019,
at the same time as a small HCl and HF enhancement, unfortunately
difficult to fully characterize on this day because of an unexpected
interruption of the FTIR measurements. After that, even if small SO2
short-term events were occasionally captured during the following
days, the hydrogen halide and active halogen species VCs remained
close to their detection limits until the first Strombolian-type
episode started (on 15 February 2019). High SO2, HCl, HF and
BrO VCs were recorded during both the P1 and P2 periods, while
significant columns of SiF4 were only detected during P1. During P1,
we captured volcanic plumes emitted between tremor episodes,
sometimes accompanied with Strombolian-type phases (on 16,
18, 20, 21, and 22 February 2019). Two of these detections were
facilitated by optimal northward wind conditions, driving the plume
toward the ALTZ station for several hours. During P2, we captured
an explosion plume (on 7 March 2019) and passive degassing (on 6,
11, and 15 March 2019). Examples of intraday time series
corresponding to the different phases of the dome growth cycle
are presented in Figures 6, 7. Intraday ratios are also calculated
“point by point” from the measurement pairs to show the intraday
variability of each species with respect to that of SO2. Frontiers in Earth Science 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume by the relatively constant HCl and HF to sulfur ratios of 0.20,
0.05 respectively. Although the intraday variability of the BrO VCs is
the same order of magnitude as its total error, the BrO/SO2 is less
affected and allows observing a progressive decrease between 07:
30 and 09:30. This decrease is mainly controlled by an increase in
SO2 columns during this period. Although the error of SiF4 VCs is
also important, its time series follows the general trend of the other
species. and/or built upon species acquired within 1 min intervals, may be
affected by noise errors and shall only be considered as indicative. For the longer-term comparison of the plume chemistry, we prefer
the ratio obtained from daily correlation plots, averaging the noise
and very short-term variations. Plume ages (±1 min) were estimated
from the ALTZ webcams, capturing images every minute. All
mentioned times are local winter times (UTC-6). On 16 February 2019, our measurements take place during a
passive degassing episode, a few hours after a ~3-h long
Strombolian-type episode. An abrupt increase of SO2 VCs was
detected at 11:30, recording the arrival of the plume in the FTIR
line of sight. This plume, aged of ~20 min and free of ash,
corresponds to a continuous degassing period between two
Strombolian-type episodes (from ~1:30 to ~4:30 and from ~16:
00 to ~20:30), in the early phases of the dome construction. BrO was
initially detected at the same time as SO2, and abruptly increased
around 12:05, as well as HF and HCl VCs. SO2 and BrO are well
correlated after this event (regression y1c in Table 4; Figure 8). The
intraday BrO/SO2 time series shows a small but statistically
significant increase (>2 × Ratio error) between 11:45 and 13:00,
coinciding with an apparent increase in the degassing (station’s
webcams). Although the time resolution of the FTIR time series is
less than that of DOAS, HCl/SO2 seems to show similar variations. The SiF4 species is also detected on this day but with a very low
abundance, close to its detection limit (VCs <1σ). On 20 February 2019 (Figure 7A), the measurements started
during an ~11-h long tremor episode and captured an ash-free
plume, corresponding to continuous degassing. The wind conditions
in the early morning drove the plume towards the ENE reaching the
instruments field of view in about 21 min. 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume Such ratios,
occasionally combining VCs from different instruments (HF/SO2) p
p
)
February-March 2019 dome growth episode is the only one
during which we capture the volcanic plume over several
consecutive days with the three (FTIR, S2000 DOAS and
Pandora) instruments, with optimal meteorological conditions. Indeed over the 77 days of the period February–April 2019,
67 allowed measuring at least 1 h with the solar absorption
configuration, cumulating a total of 558 h of measurements. The volcanic plume was captured by the different instruments
over 118 h, distributed over 33 days. Figure 5 reports the SO2,
HCl, HF, BrO, SiF4 time series during this crisis, concurrently
with the tremor duration (from the daily CENAPRED reports),
generally associated with short-term (several hours) extrusive
periods. During this episode, we distinguished three main activity
periods (see Introduction): P1 (15 February–26 February 2019),
dominated
by
the
dome
growth
manifestations
such
as
“Strombolian-type
activity”
accompanied
by
spasmodic
tremor; P2 (26 February–16 March 2019) characterized by the
occurrence of strong explosions accompanying and following the 11 Frontiers in Earth Science frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 TABLE 4 Linear fit parameters obtained from the correlation plots as presented in Figures 8 and 9. R2 stands for the determination coefficient. Species ratio
Regression label; Date
Slope
Intercept
(molec/cm2)
R2
BrO/SO2
y1a; 24/12/2017
(1.14 ± 0.2) × 10−4
(9.45 ± 1.70) × 1013
0.83
y1b; 20/02/2019
(8.70 ± 0.82) × 10−5
(3.86 ± 0.84) × 1013
0.66
y1c; 16/02/2019
(6.28 ± 0.65) × 10−5
(4.46 ± 9.93) × 1012
0.86
BrO/HCl
y2a; 24/12/2017
(6.94 ± 2.67) × 10−4
(6.10 ± 4.13) × 1013
0.89
y2b; 20/02/2019
(3.89 ± 0.33)× 10−4
(3.59 ± 0.76) × 1013
0.84
y2c; 16/02/2019
(2.60 ± 0.55) × 10−4
(0.6 ± 1.7) × 1013
0.92
HCl/SO2
y3a; from 16/02 to 25/02/2019
(0.21 ± 0.01)
(1.90 ± 2.20) × 1014
0.98
y3b; from 07/03 to 15/03/2019
(0.16 ± 0.01)
(1.34 ± 1.27) × 1015
0.96
y3c; from 15/04 to 18/04/2019
(0.08 ± 0.01)
(1.42 ± 0.12) × 1015
0.92
HF/SO2
y4; from 16/02 to 18/04/2019
(0.049 ± 0.001)
(1.45 ± 1.44) × 1014
0.96
SiF4/SO2
Figure 9
(1.56 ± 0.25) × 10−3
(2.08 ± 1.87) × 1014
0.96
aErrors in the slope and intercept are given with a confidence interval of 0.95. rrors in the slope and intercept are given with a confidence interval of 0.95. Frontiers in Earth Science 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume At 07:52, a puff occurred
(Supplementary Figure S2, supplementary material), and the plume
reached the measurement line of sight at ~ 08:11. At ~8:45, the wind
field changes towards the NE accompanying the rising Sun and the
plume was progressively driven towards the station. A small puff
occurred at 10:10, and an apparently more condensed plume was
observed
from
10:40
to
12:00
(Supplementary
Figure
S2,
supplementary material). Despite the low time resolution of the
FTIR measurements, SO2, HCl and HF time series have a similar
behaviour, with coincident peaks at 08:20 and 11:25. BrO VCs are on
average about a factor of 1.5 higher than that observed on
18 February 2019, and although its variability is different from
that of the other species, it shows the same significant peaks at 08:
20 and 11:25. The peak observed at 08:20 coincides with the
arrival of the first puff plume in the optical path of the
instrument. The BrO/SO2 linearly decreases with time until
~9:30 when it reaches a plateau, increases again from 10:15 to
10:50 and finally drops again until the end of the measurements. HCl/SO2 and HF/SO2 oscillate around their average ratio, except
for the slight decrease of HF/SO2 at the beginning of the time
series until ~9:00. SiF4 is detected over a short period (<30 min,
two measurements: 7.2 × 1015 molec/cm2) with VCs much higher
than those of the other presented days, closer to the values
recorded on 6 March 2015 (1.2 × 1016 molec/cm2; Taquet
et al., 2019). The BrO/SO2 peak at 10:50 coincides with the On 18 February 2019, we captured an ash-poor plume from
continuous degassing during several hours, immediately following
the end of a weak Strombolian-type episode (~21:30 to ~7:30 from
CENAPRED reports) with few extra-crater projections. After a
strong drop of ~30 min duration in the degassing activity (from
~6:30), an explosion occurred at 07:04, preceding the beginning of
our measurements. Our measurements captured the return to a
higher degassing regime. Wind speed was evaluated to more or less
20 m/s this day (from the GDAS/NOAA model), and the plume age
can be estimated to be ~15 min using the station webcam. The HF,
HCl, and SO2 column variations remained perfectly correlated
between them during the whole measured period as highlighted Frontiers in Earth Science 12 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 FIGURE 8
Correlation plots of (A) BrO vs. SO2, (B) BrO vs. 4.2 SO2, HCl, HF, BrO and SiF4 VCs
measurements in the Popocatépetl’s plume HCl, (C) HCl vs. SO2 and (D) HF vs. SO2 VCs combining DOAS and FTIR data on 24 December 2017 and
during the February–April 2019 dome cycle. The BrO/SO2, BrO/HCl, HCl/SO2 and HF/SO2 ratios were determined from the slopes of the correlation
plots. FIGURE 8
Correlation plots of (A) BrO vs. SO2, (B) BrO vs. HCl, (C) HCl vs. SO2 and (D) HF vs. SO2 VCs combining DOAS and FTIR data on 24 December 2017 and
during the February–April 2019 dome cycle. The BrO/SO2, BrO/HCl, HCl/SO2 and HF/SO2 ratios were determined from the slopes of the correlation
plots. FIGURE 9
SiF4/SO2 correlation plot for the 2017–2019 measurement
period. The slope is found to be (1.56 ± 0.25) × 10−3. Error bars
represent the total errors of the FTIR VCs measurements, detailed in
the text. destruction. The plume entered the instruments line of sight around
10:30 until 15:00. The SO2 peak at 11:46 corresponds to a plume
coming from a small puff detected on the visible cameras above the
crater at 11:30. HCl and SO2 vary in time with the same magnitude,
such that the HCl/SO2 ratio is found to be almost constant. BrO is
also detected, but contrary to the other days, BrO/SO2 does not
present a specific trend, oscillating around its mean value. Frontiers in Earth Science 4.3 BrO/SO2, HCl/SO2, HF/SO2, and BrO/HCl
ratios at Popocatépetl BrO/SO2, HCl/SO2, HF/SO2, and BrO/HCl ratios have been
calculated from the intraday correlation plots from February to
April 2019 and reported in Figure 8. For the calculation of the HF/
SO2 (HF and SO2 measured with two different NDACC filters) and
BrO/HCl ratios, we interpolated the SO2 and BrO S2000 DOAS time
series (which have the highest time resolution) at the time of the
FTIR measurements (HF, HCl). The interpolation is performed only
if there are coincident data within an interval of 1 min for SO2 or
2 min for BrO. An evaluation of the bias generated by the use of the
SO2 columns from the different instruments for the calculation of
HF/SO2 and BrO/SO2 ratios is shown in supplementary material,
Supplementary Figure S1. A maximum bias of 3% is found, making
possible the combination of the different SO2 products to increase arrival of the more condensed plume (after the puff at 10:10) in
the optical path of the instrument. On 11 March 2019 (Figure 7B), we captured a northward blown
plume from passive degassing a few days after the end of the dome Frontiers in Earth Science 13 frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 10.3389/feart.2023.1062699 10.3389/feart.2023.1062699 In our opinion, these two types of observation support the
occurrence at Popocatépetl of both SiF4 generation mechanisms,
as previously reported (Love et al., 1998; Taquet et al., 2017), i.e.,
1) the HF interaction with silica in the closing connected
porosity of a mature dome and 2) the in-plume reaction of
HF with ash. BrO/SO2 and BrO/HCl have similar slopes on
16 and 20 February 2019, but further data are needed to explore
its variability as a function of the dome growth phases. Finally,
we compared our bromine to sulfur ratios with some of the
meteorological parameters of interest available at the ALTZ
station. The lowest ratio (6.3 ± 0.6 × 10−5), obtained on
16 February 2019 is recorded on the day with the highest RH
(>48%). On 18 February 2019, the RH even falls under 10% over
the measurement period with a mid-range ratio (Figure 6). The
highest ratios were obtained on 24 December 2017 and
20 February 2019, 2 days characterized with low humidity
conditions <24% and <34%, respectively, which do not favor
the formation of BrO (Oppenheimer et al., 2006). These
observations suggest that atmospheric humidity might not be
the
main
control
on
BrO
conversion
at
Popocatépetl. 5 Discussion and conclusion In this contribution, we explored the feasibility and
performance of combining direct-sun measurements from
S2000 DOAS, Pandora and HR-FTIR instruments to study
the
short-term
variability
of
Popocatépetl’s
plume
composition. The SO2 vertical columns obtained from the
three
instruments
were
found
similar
after
their
intercalibration (median difference <12%), making it possible
to combine the different products to increase the number of
coincident data pairs in the ratios calculation. Although the
DOAS instrument is generally used in “scattering” mode to
continuously measure BrO/SO2 in volcanic plumes, there are
important advantages of direct-sun such as the straightforward
conversion from slant to vertical columns using a simple
geometric
determination
of
the
airmass
factor,
and
the
absence of ring effect, thus considerably reducing the total
errors. Moreover, this configuration is one of the most
reliable allowing direct comparison with VCs from other
techniques (FTIR) and determining ratios of species that are
not detectable with the same technique. FTIR SiF4 retrieval was
improved compared to that published in Taquet et al. (2019)
using the new spectroscopy from Boudon et al. (2020) and
allowing the treatment of high resolution spectra. The VCs
total errors obtained from the low resolution products are
only slightly improved, but the results obtained from the
high resolution spectra are significantly more precise with
total error <20% for total columns >2.0 × 1015 molec/cm2
when those from the low resolution spectra occasionally
reached 60%. HF/SO2 (Figure 8D) correlation plot shows a more important
dispersion, and only one trend line can be identified over the
2019 dome cycle, with a HF/SO2 ratio around 0.049 ± 0.001. This value is in accordance with the previous values measured
since 2012 (from 0.01 to 0.04) (Taquet et al., 2019). Our previous studies on the long-term variability of the
Popocatépetl’s plume composition showed a link between the
HCl/SO2 and the main long-term changes in the deep and
shallow magmatic processes (intrusions, long-term cessation of
extrusive activity, etc). In this study we examined the variability
of this ratio during a complete dome growth cycle (timescale of
several months), and found a clear increase of HCl with respect to
sulfur during the period of dome growth. It decreased during the
period of destruction and reached very small values when no dome
sits on the floor of the crater. HF/SO2 does not show the same
behavior and remains almost constant during the whole cycle. 4.3 BrO/SO2, HCl/SO2, HF/SO2, and BrO/HCl
ratios at Popocatépetl Furthermore, on 20 February 2019, the highest ratios are
measured at ~7:46, during sunrise, with a solar irradiance as
low as 133 W/m2, which appears sufficient to allow the
conversion of HBr into detectable amounts of BrO. the number of coincident pairs, in particular for the HF/SO2 ratio. S2000 DOAS and FTIR SO2 products were therefore combined
before the calculation of the different ratios, presented in Figure 8. Because of the rare occurrence of the BrO and SiF4 species in our 2.5-
month time series, we added the event of 24 December 2017
(Figure 8 in green) which was one of the most significant events
of our 2.5-year time series to give an idea of their long-time
variability. BrO/SO2 ratios (Figure 8A) are found in the range of [0.6–1.1] ×
10−4, with a significantly higher ratio on 24 December 2017 than
those reported for 2019. This falls in the higher range of the few
ratios previously reported for the Popocatépetl volcano between
0.04 × 10−4 and 1.4 × 10−4 (Boichu et al., 2011; Bobrowski and
Giuffrida, 2012; Platt and Bobrowski, 2015; Fickel and Delgado
Granados, 2017) and compiled in Gutmann et al. (2018). BrO/HCl (Figure 8B) ranges between 2.6 and 6.9 × 10−4 and
shows similar slopes (~3 × 10−4) during the 2019 dome growth phase
(on 16 and 20 February 2019) but different offsets. HCl/SO2 (Figure 8C) ranges between 0.08 ± 0.01 and 0.21 ±
0.01 and decreases with time during the 2019 dome growth cycle. Three different trend lines can be distinguished corresponding to
three periods, from 15 to 25 February 2019, from 7 to 15 March
2019 and after this. The first two trend lines coincide with the
dome growth (p1), and destruction (p2) periods described in
Section
4.2,
respectively. The
third
trend
line
(y3c),
corresponding to the period p3, with very low HCl column
abundances and low HCl/SO2 ratios characterizes a new phase
without extrusive activity. 10−3; Taquet et al., 2019). 5 Discussion and conclusion Its
high intraday variability is likely due to in-plume reaction during the
transport to the measurement locus. Two types of SiF4 detections occurred over the 2.5 years survey,
either 1) as transient peaks generally occurring during explosions
with ash-rich plumes, too short-termed for correlations with the SO2
or HF to be clearly established, or 2) as several hour long
enhancements with clear correlations with the other volcanic gas
species, occurring only at the end of the dome growth episodes. Over
the complete 2.5-years time series, only 6 days (Figure 9) allowed
calculating an average SiF4/SO2 ratio, found equal to (1.56 ± 0.25) ×
10−3, similar to the lowest values found in 2012–2016 (1.0 −9.9 ×
10 3 T
l 2019) 10−3; Taquet et al., 2019). Frontiers in Earth Science 14 frontiersin.org Taquet et al. Taquet et al. 10.3389/feart.2023.1062699 10.3389/feart.2023.1062699 Oppenheimer et al., 2006; Kern et al., 2009). Since the BrO
content in the Popocatépetl’s plume may not be mainly
controlled
by
the
meteorological
conditions
or
sunlight
intensity, the impact of plume aerosol content has to be
explored in future studies. BrO/HCl was quantified for the
first time at Popocatépetl (from 2.6 to 6.9 × 10−4). It is found
almost constant at the intraday scale but with significant day to
day variability. Considering the BrO as full conversion from HBr,
this is already a factor of 1.5–3.5 higher than the lower bound of
Gerlach. (2004) Br/Cl range, and thus constitutes a first
minimum
constraint
on
this
ratio
at
Popocatépetl. The
maximum conversion rates ever described (65% at Ambrym,
Vanuatu; Bani et al., 2009; Gutman et al., 2018) would provide a
higher bound for this ratio at Popocatépetl of 1.08 × 10−3. The BrO/SO2 ratio was successfully quantified at a distance of
11 km from the crater, found between 0.6 and 1.1 × 10−4, in the
range of the few values previously reported (Bobrowski and
Giuffrida,
2012;
Platt
and
Bobrowski,
2015;
Fickel
and
Delgado Granados, 2017). However, HBr analysis from the
high resolution FTIR spectra is not straightforward at this
distance from the crater, and so as in Taquet et al. (2019),
only a detection limit of 5.0 × 1014 molec/cm2 was calculated. 5 Discussion and conclusion Following this study and assuming the average Br/Cl ratio for arc
volcanoes (2.2 ± 2.0 × 10−3; Gerlach, 2004), we calculate a
speculative primitive HBr/SO2 for the surveyed period from
the average HCl/SO2 ratio (0.21 ± 0.01) of (4.6 ± 4.2) × 10−4. Assuming the maximum value (8.8 × 10−4) within the uncertainty
interval for this (HBr/SO2) ratio, our measured BrO/SO2 ratios
would thus correspond to minimum HBr conversion rates of the
order of 10%, which is in the range for plumes with an age equal
to or higher than 15 min (Rüdiger et al., 2021). Using this
detection limit, the maximum HBr/BrO ratio would then be
between 0.11 and 0.19. This study is a new step towards the establishment of a
comprehensive highly time-resolved database of volcanic gas
ratios for Popocatépetl by integrating multiple spectroscopic
techniques. Such database should provide insight for, e.g., 1)
delineating the degassing processes at Popocatépetl occurring
during both passive degassing and extrusive phases, 2) refining
the degassing balance by taking into account the in-plume
interactions, and
3) comparing
the degassing
processes
observed at Popocatépetl with those of analogue volcanoes. For the first time, the BrO/SO2, HCl/SO2, HF/SO2, BrO/HCl,
and SiF4/SO2 ratios have been simultaneously measured in the
Popocatépetl’s plume and their behavior with respect to the
volcanic activity was explored at the scale of a dome growth
cycle (several weeks). The decrease of the HCl/SO2 ratio after the
dome destruction at the end of February 2019 suggests that halogen
emissions are higher during the dome growth periods, i.e., during
sustained ascension of the magma column, confirming the long-
term observations reported in Taquet et al. (2019). HF/SO2 ratios
apparently
remain
constant
during
the
February
to
April
2019 period and show a high intraday variability. SiF4/SO2 was
quantified on various occasions during the dome cycle period and
apparently remains constant, equal to (1.56 ± 0.25) × 10−3
compatible with previous results (Goff et al., 1998; Love et al.,
1998; Stremme et al., 2012; Taquet et al., 2017; Taquet et al.,
2019). Considering a HF/SO2 average ratio of 0.049 ± 0.001 over
the dome cycle period, we determine an average SiF4/HF ratio of
0.031 ± 0.001. Our
study
shows
the
benefits
and
performance
of
implementing a UV-Vis spectrometer on an existing HR-FTIR
NDACC instrument to simultaneously measure the main
primary
and
secondary
volcanic
gaseous
species. Data availability statement The raw data supporting the conclusion of this article will be
made available by the authors, without undue reservation. 5 Discussion and conclusion Such
implementation
does
not
either
disturb
or
interrupt
the
continuous
solar
absorption
atmospheric
measurements
performed
for
the
networks. This
experiment
could
be
reproduced
at
any
other
NDACC
stations
located
near
volcanoes, such as, e.g., the Izaña Observatory which recently
recorded atmospheric data during the 2021-La Palma (Canary
Islands, Spain) eruption (Garcia et al., 2022), at Maïdo
observatory (La Réunion Island, France), or Mauna
Loa
Observatory (Hawaii), providing quasi-near-real-time critical
data for the management of volcanic emergencies. High
spectral resolution spectroscopic data also have the advantage
to provide simultaneous atmospheric vertical profiles of both
atmospheric and some volcanic species, giving new insight into
the effect of volcanic emissions on the atmospheric chemistry and
climate change. BrO/SO2 and HCl/SO2 apparently show different long-term
variability, at least for the three examples presented here (on
24 December 2017 and on 16 and 20 February 2019), for which
two significantly different BrO/SO2 ratios were found while the HCl/
SO2 was found to be constant (Figure 8). On a few occasions, clear patterns appear in the BrO/SO2
intraday time series (Figures 6, 7A) while the HCl/SO2 remains
almost constant (flat baseline). These patterns, generally a few
hours long, consisted either in decreasing trends (e.g., the first
hours of 18 and 20 February) or increase/decrease sequences
(e.g., 10:00–12:00 on 20 February). In particular, 2 days (16 and
20 February 2019) show significant BrO/SO2 variations, which
are not dominated by the SO2 columns variability. The most
significant variations (20 February 2019 between 10:50 and 12:
00) occur coincidentally with a change in the aspect of the plume
and might be related to changes in the plume moisture content,
although further studies are needed to explore this aspect in more
details. Surprisingly, the highest average BrO/SO2 ratio (recorded
on 20 February 2019) is not observed during meteorological and
sunlight intensity conditions (RH <34%, and solar irradiance
~133 W/m2)
that
favour
HBr
conversion
to
BrO
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and F degassing as an indicator of volcanic dynamics: The 2001 eruption of Mount Etna. Geophys. Res. Lett. 29, 1559. doi:10.1029/2002GL015032 Boulesteix, T., Legrand, D., Taquet, N., Coppola, D., Laiolo, M., Valade, S., et al. (2022). Modulation of Popocatépetl’s activity by regional and worldwide earthquakes. Bull. Volcanol. 84, 80. doi:10.1007/s00445-022-01584-2 Aiuppa, A., Federico, C., Franco, A., Giudice, G., Gurrieri, S., Inguaggiato, S., et al. (2005). Emission of bromine and iodine from Mount Etna volcano. Geochem. Geophys. 6 (8). doi:10.1029/2005GC000965 Burrows, J. P., Richter, A., Dehn, A., Deters, B., Himmelmann, S., Voigt, S., et al. (1999). Atmospheric remote-sensing reference data from GOME: Part 2. Temperature-
Dependent absorption cross-sections of O3 in the 231-794 nm range. J. Quant. Spectrosc. Radiat. Transf. 61, 509–517. doi:10.1016/S0022-4073(98)00037-5 Arellano, S., Galle, B., Apaza, F., Avard, G., Barrington, C., Bobrowski, N., et al. (2021). Synoptic analysis of a decade of daily measurements of SO2 emission in the
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Volcanic and Atmospheric Change. Earth Syst. Sci. Data 13 (3), 1167–1188. doi:10. 5194/essd-13-1167-2021 Burton, M., Allard, P., Muré, F., and La Spina, A. (2007). Acknowledgments We thank the two reviewers for their very constructive
comments, which help to significantly improve the manuscript. We thank R. Campion for fruitfull discussions on the degassing
and activity of the Popocatépetl. We thank all the personnel of the
CENAPRED for realizing the Popocatépetl activity reports. We
thank the PGN team for the Pandora data processing. The PGN Supplementary material The Supplementary Material for this article can be found online
at: https://www.frontiersin.org/articles/10.3389/feart.2023.1062699/
full#supplementary-material Publisher’s note We acknowledge financial support from grants UNAM-PAPIIT
IA101620 and IN111521. NT and TBo also thank the stipend given by
the Mexican Foreign Affairs Department (Secretaría de Relaciones
Exteriores) and its AMEXCID program. Financial support from
Conacyt-AEM through grant No. 275239 is acknowledged. All claims expressed in this article are solely those of the authors
and
do
not
necessarily
represent
those
of
their
affiliated
organizations, or those of the publisher, the editors and the
reviewers. Any product that may be evaluated in this article, or
claim that may be made by its manufacturer, is not guaranteed or
endorsed by the publisher. Author contributions All the co-authors contributed in the development and the discussion
of concepts, and to the preparation of the manuscript. NT, CR, and WS
were responsible of the DOAS and FTIR measurements and the data
analysis. MG and WS lead the ALTZ station development and its
long-term operation. TBo participated to the measurements and to
the interpretation. AB was responsible of the maintenance of the Frontiers in Earth Science 15 frontiersin.org frontiersin.org Taquet et al. 10.3389/feart.2023.1062699 instruments at the Altzomoni station. OG contributes for the FTIR/
DOAS intercomparison and discussions. TBl leads at KIT the German-
Mexican collaboration for the deployment of the high resolution FTIR
spectrometer and supports its long-term operation as part of NDACC. FH has helped in the design and setup of the spectrometer and solar
tracker before it was shipped to Mexico. He has developed the retrieval
code PROFFIT and gives continuously support to the UNAM group for
its use and in operating the spectrometer. is a bilateral project supported with funding from NASA and ESA. We thank to C. Fayt, M. Van Roozendael, and A. Merlaud for their
help to use the QDOAS software. Conflict of interest The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be
construed as a potential conflict of interest. Cede, A., Tiefengraber, M., Gebetsberger, M., and Spinei Lind, E. (2021). Pandonia
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Popocatépetl volcano between 2007 and 2008: FTIR spectroscopic measurements of an 18 Frontiers in Earth Science frontiersin.org 18
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Plackett-Burman Design and Response Surface Optimization of Medium Trace Nutrients for Glycolipopeptide Biosurfactant Production
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ABSTRACT ABSTRACT
Background: A glycolipopeptide biosurfactant produced by Pseudomonas aeruginosa strain IK
surface tension of fermentation broth from 71.31 to 24.62 dynes/cm at a critical micelle conce
mg/L. The compound proved suitable for applications in emulsion stabilization in food, as well a
pharmaceutical formulations. Methods: In the present study, Plackett-Burman design (PBD) and
method (RSM) were employed to screen and optimize concentrations of trace nutrients in
medium, to increase surfactant yield. Results: The PBD selected 5 significant trace nutrien
screened. The RSM, on the other hand, resulted in the production of 84.44 g glycolipopeptide/L
medium containing 1.25 mg/L nickel, 0.125 mg/L zinc, 0.075 mg/L iron, 0.0104 mg/L boron,
copper. Conclusion: Significant second-order quadratic models for biomass (P<0.05; adjusted
biosurfactant (R2=99.44%) responses suggest excellent goodness-of-fit of the models. However
non-significant lack-of-fit (Biomass: F=1.28; P=0.418; Biosurfactant: F=1.20; P=0.446) test
their adequacy to explain data variations in the experimental region. The glycolipopeptide
for the formulation of inexpensive pharmaceutical products that require surface-act
DOI: 10.18869/acadpub.ibj.21.4.249
Keywords: Pseudomonas aeruginosa, Surface-active agents, Fermentation, Nickel, Copper
Corresponding Author: Maurice George Ekpenyong
Environmental Microbiology and Biotechnology Unit, Department of Microbiology, Faculty of Biological Sciences, University of
Calabar, Nigeria. Tel.: (+234-70)38218972; E-mail: maurygg2002@yahoo.com or maurygg2002@unical.edu.ng Background: A glycolipopeptide biosurfactant produced by Ps
surface tension of fermentation broth from 71.31 to 24.62 dyn
mg/L. The compound proved suitable for applications in emulsi
pharmaceutical formulations. Methods: In the present study, P
method (RSM) were employed to screen and optimize conce
medium, to increase surfactant yield. Results: The PBD sele
screened. The RSM, on the other hand, resulted in the product
medium containing 1.25 mg/L nickel, 0.125 mg/L zinc, 0.075
copper. Conclusion: Significant second-order quadratic mode
biosurfactant (R2=99.44%) responses suggest excellent goodne
non-significant lack-of-fit (Biomass: F=1.28; P=0.418; Biosu
their adequacy to explain data variations in the experiment
for the formulation of inexpensive pharmaceutical prod
DOI: 10.18869/acadpub.ibj.21.4.249
Keywords: Pseudomonas aeruginosa, Surface-active agents, Fermentation,
Corresponding Author: Maurice George Ekpenyong
Environmental Microbiology and Biotechnology Unit, Department of Microbiology
Calabar, Nigeria. Tel.: (+234-70)38218972; E-mail: maurygg2002@yahoo.com or m Background: A glycolipopeptide biosurfactant produced by Pseudomonas aeruginosa strain IKW1 reduced the
surface tension of fermentation broth from 71.31 to 24.62 dynes/cm at a critical micelle concentration of 20.80
mg/L. The compound proved suitable for applications in emulsion stabilization in food, as well as in cosmetic and
pharmaceutical formulations. FULL LENGTH FULL LENGTH Iranian Biomedical Journal 21(4): 249-260 July 2017 ABSTRACT Methods: In the present study, Plackett-Burman design (PBD) and response surface
method (RSM) were employed to screen and optimize concentrations of trace nutrients in the fermentation
medium, to increase surfactant yield. Results: The PBD selected 5 significant trace nutrients out of the 12
screened. The RSM, on the other hand, resulted in the production of 84.44 g glycolipopeptide/L in the optimized
medium containing 1.25 mg/L nickel, 0.125 mg/L zinc, 0.075 mg/L iron, 0.0104 mg/L boron, and 0.025 mg/L
copper. Conclusion: Significant second-order quadratic models for biomass (P<0.05; adjusted R2=94.29%) and
biosurfactant (R2=99.44%) responses suggest excellent goodness-of-fit of the models. However, their respective
non-significant lack-of-fit (Biomass: F=1.28; P=0.418; Biosurfactant: F=1.20; P=0.446) test results indicate
their adequacy to explain data variations in the experimental region. The glycolipopeptide is recommended
for the formulation of inexpensive pharmaceutical products that require surface-active compounds. DOI: 10.18869/acadpub.ibj.21.4.249 Keywords: Pseudomonas aeruginosa, Surface-active agents, Fermentation, Nickel, Copper Corresponding Author: Maurice George Ekpenyong
Environmental Microbiology and Biotechnology Unit, Department of Microbiology, Faculty of Biological Sciences, University of Calabar, P.M.B.1115
Calabar, Nigeria. Tel.: (+234-70)38218972; E-mail: maurygg2002@yahoo.com or maurygg2002@unical.edu.ng Plackett-Burman Design and Response
Surface Optimization of Medium Trace Nutrients for
Glycolipopeptide Biosurfactant Production
Maurice George Ekpenyong*1, Sylvester Peter Antai1, Atim
David Asitok1 and Bassey Offiong Ekpo2,3 1Environmental Microbiology and Biotechnology Unit, Department of Microbiology, Faculty of Biological
Sciences, University of Calabar, P.M.B.1115 Calabar, Nigeria; 2Exploration, Research and Services Section,
Research and Development (R&D) Division, Nigerian National Petroleum Corporation (NNPC),
Port-Harcourt, Nigeria; 3Environmental Geochemistry Unit, Department of Pure and Applied Chemistry, Faculty
of Physical Sciences, University of Calabar, P.M.B. 1115, Calabar, Nigeria Received 29 August 2016; revised 8 October 2016; accepted 10 October 2016 Optimization experiments
Screening trace elements g
The PBD incorporated into MINITAB 17 statistical
software (trial version) was used to screen 12 trace
nutrients in 20 randomized experimental runs. The
nutrients were purchased from Sigma Aldrich, USA
and included NaCl, KCl, CaCl2, MgSO4.7H2O,
CuSO4.5H2O,
NiCl2.6H2O,
FeCl3,
ZnCl2.7H2O,
K3BO3, MoNa2O4.2H2O, CoCl2, and MnSO4.4H2O. Each was tested only at two levels, low and high. Biosurfactant concentration (g/L) served as the only
response
variable. Fermentation
medium
was
formulated according to the experimental design and
based on 50 mL of each formulation dispensed into
250-mL Erlenmeyer flask. Phosphorus (1.0 g/L
(NH4)2HPO4/NH4H2PO4) and carbon (5% v/v waste
frying sunflower oil with composition of [% w/w]
stearic acid [2.21], palmitic acid [6.11], volatile
fractions (16.23), oleic acid [22.34], and linoleic acid
[50.76]; saponification value 76 and density [30°C]
283.3 kg/m3) sources were added to the media, and
flasks were corked and sterilized by autoclaving at
121°C for 15 min. Upon cooling, filter-sterilized urea
(1.19 g/L) and inoculum (10% v/v-108 cells/mL) were
added to the medium, and flasks were incubated on a
rotary shaker agitating at 150 rpm at room temperature
for 72 h. All arrangements were made in triplicates. Optimization of fermentation media involving major
and trace nutrients, as well as environmental and
fermentation conditions has been documented[14-16]. A
good number of biosurfactant fermentation media
compositions incorporating both major and minor
nutrients have been reported without information on
source of trace element composition. A remarkable
study on trace mineral composition of a biosurfactant
medium has been reported by Joshi et al.[17]. p
y
Trace elements are required in small amounts and
play essential roles in cellular metabolism, mostly as
enzyme co-factors and/or side groups in some
microbial metabolites[18]. Selection of significant trace
elements for incorporation into fermentation media is
an uphill task from the viewpoint of variable size. A
classical method for screening large variables is the use
of Plackett-Burman design (PBD). It is a small-sized
two-level factorial experimental design programmed to
identify critical physicochemical parameters from N
number of variables in N+1experiments without
recourse to the interaction effects between and among
the variables. Since the sample size is traditionally
small, the interaction effects are completely shrouded
in the main effects. PBD therefore simply screens the
design space to detect large main effects[19]. The
selected parameters are further optimized by the means
of an appropriate design technique of a response
surface method (RSM). INTRODUCTION Microbial surfactants are of diverse chemical nature,
including glycolipids[3], flavolipids[4], lipopeptides[5],
and glycolipopeptides[6-8]. Each biosurfactant performs
specialized function(s) in their respective producing
organisms. Such functions include solubilization,
emulsification, pathogenesis, antibiosis, swarming
motility, as well as wetting of and attachment to
surfaces[9]. B iosurfactants are amphiphilic compounds that
reduce the free surface enthalpy per unit area of
surfaces and interfaces[1]. They are derived
from biological sources including plants, animals, and
microorganisms; however, world commercial interest
has focused, principally, on microbial derivatives from
bacteria, yeasts, and molds[2]. B Commercial interest in biosurfactant production has Iran. Biomed. J. 21 (4): 249-260 249 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. 30°C for 36 h. The bacterium was passed a second time
through the reactivation step in the same medium for
24 h, after which a loop-full of culture was used to
inoculate freshly prepared Luria broth in 50-mL
Erlenmeyer flask containing 10 mL of medium. Flasks
were incubated on a rotary shaker at 150 rpm at room
temperature (28±2°C) for 18 h. heightened over the years owing to their increasingly
widening applications. Surface-active compounds are
applied for different purposes in pharmaceutical,
detergent, food, cosmetic, petroleum, and agricultural
sectors, as well as the environment[10,11]. A threatening
bottleneck in microbial production of biosurfactants is
the economics of production[1]. Several strategies exist
to increase biosurfactant yield, that include the use of
low-cost substrates, medium optimization, strain
improvement, and development of fermentation
conditions[12,13]. Response surface design and fermentation studies Response surface design and fermentation studies
The experimental design employed to fit the multiple
regression models of the fermentation study was a 25-1
half-fractional factorial central composite rotatable
design. The test variables were Ni2+, Zn2+, Fe3+, B3+,
and Cu2+; each at five levels. The selection of factor
levels for RSM followed, on one hand, the method of
the path of steepest ascent, a procedure that moves
nutrient levels sequentially in the direction of
maximum increase in the response investigated, and on
the other, a consideration for the introduction of
biomass concentration as a second response variable to
facilitate the determination of biosurfactant production Optimization experiments
Screening trace elements This method is a collection of
statistical techniques that uses design of experiments to
build models, evaluate the effect of factors and predict
optimum conditions for the factors[20]. The objective of
the present study was to optimize the conditions of
trace nutrients in a fermentation medium using PBD
and RSM to improve biosurfactant yield. Biosurfactant quantification was performed as
described elsewhere[6]. Data were analyzed using the
same statistical software that generated the design. Significant trace elements were selected for RSM and
optimization. Statistical analysis All data generated from the factorial experiment
were subjected to multiple regression analysis using
least squares to build the regression models. A second-
order (quadratic) function was used to fit the data
generated. Experimental
design,
data
analysis,
interaction plotting, and optimization of factor
conditions were done with MINITAB 17 statistical
software, while Excel 2007 was used for confirmation Table 1. Producing organism The bacterium, Pseudomonas aeruginosa strain
IKW1, was earlier isolated[6,21]. The stock culture of
the bacterium was retrieved and subcultured in Tryptic
Soy Agar by the quadrant-streak plate technique at 250 Iran. Biomed. J. 21 (4): 249-260 Ekpenyong et al. Nutrient Optimization for Biosurfactant Production Nutrient Optimi
Ekpenyong et al. oil was subsequently added to each flask at 5% (v/v)
concentration and flasks, without nitrogen source, were
sterilized by autoclaving at 121°C for 15 min. Upon
cooling, filter-sterilized urea (1.19 g/L) and inoculum
(10% v/v–108 cells/mL) were introduced into flasks
prepared in triplicates, and then the flasks were
incubated on rotary shakers agitating at 150 rpm at
room temperature for 72 h. yield, Yp/x. Results of preliminary screening for the
effects of increasing concentrations of metals on the
growth of the bacterium were also considered in the
selection of factor levels of significant trace nutrients
from PBD for biomass and biosurfactant formation. Actual levels of each factor were calculated using the
equation of Myers and Montgomery[20]. Determination of biomass (Y1) and biosurfactant
(Y2) concentrations Small portions of 72-h fermentation broth from each
experimental setting (10 mL) were centrifuged at 8,000
g for 10 min. Cell-free supernatants were collected,
and the cell pellets were washed twice in de-
mineralized water. The supernatants were subjected to
0.45-µM and 0.2-µM Millipore membrane filtrations
and subsequently to acid precipitation with 6N HCl,
pH 2.0. Biomass and biosurfactant concentrations were
determined from cell pellets and acid precipitates of
supernatants, respectively as described previously[6]. The coded levels were determined as follows:
X1=(Ni2+-0.75)/0.25,
X2=(Zn2+–0.075)/0.025,
X3=
(Fe3+–0.075)/0.025, X4=(B3+–0.03)/0.01, and X5=(Cu2+- The coded levels were determined as follows:
X1=(Ni2+-0.75)/0.25,
X2=(Zn2+–0.075)/0.025,
X3=
(Fe3+–0.075)/0.025, X4=(B3+–0.03)/0.01, and X5=(Cu2+-
0.075)/0.025. The response variables were biomass Y1
(g/L) and biosurfactant Y2 (g/L) concentrations. The
experimental design required 32 experimental runs,
which were set up in 250-mL Erlenmeyer flasks, each
containing 50 mL of fermentation medium composed
(g/L) of Na2HPO4/KH2PO4 (2:1; 4.5), MgSO4.7H2O
(0.2),
NaCl
(0.5),
CaCl2
(0.5),
KCl
(0.5),
MoNa2O4.2H2O (0.05 mg/L), CoCl2.6H2O (0.05
mg/L), and MnSO4.4H2O (0.05 mg/L). Different
concentrations of the five significant trace elements
obtained from PBD screening were then added to the
flasks according to the actual values of the factor levels
specified by the coded values in Table 1. Medium pH
was adjusted to 7.0 with KOH pellets[4]. Waste frying Statistical analysis Placket-Burman design matrix (randomized) for trace element contribution to biosurfactant formation in coded units
Run
A
B
C
D
E
F
G
H
J
K
L
M
BSC (g/L)
1
-1
-1
-1
1
-1
1
-1
1
1
1
1
-1
23.58
2
1
1
1
1
-1
-1
1
1
-1
1
1
-1
25.96
3
-1
-1
1
1
-1
1
1
-1
-1
-1
-1
1
24.21
4
1
-1
-1
1
1
-1
1
1
-1
-1
-1
-1
21.43
5
-1
1
1
-1
-1
-1
-1
1
-1
1
-1
1
24.19
6
1
1
-1
1
1
-1
-1
-1
-1
1
-1
1
28.96
7
-1
1
1
1
1
-1
-1
1
1
-1
1
1
31.73
8
1
1
-1
-1
-1
-1
1
-1
1
-1
1
1
32.75
9
-1
-1
-1
-1
-1
-1
-1
-1
-1
-1
-1
-1
17.45
10
1
-1
1
-1
1
1
1
1
-1
-1
1
1
36.02
11
1
-1
1
1
1
1
-1
-1
1
1
-1
1
24.72
12
-1
1
-1
1
-1
1
1
1
1
-1
-1
1
23.48
13
1
1
1
-1
-1
1
1
-1
1
1
-1
-1
22.31
14
1
-1
1
1
-1
-1
-1
-1
1
-1
1
-1
27.08
15
-1
-1
-1
-1
1
-1
1
-1
1
1
1
1
33.27
16
-1
1
-1
1
1
1
1
-1
-1
1
1
-1
25.31
17
-1
1
1
-1
1
1
-1
-1
-1
-1
1
-1
28.78
18
1
1
-1
-1
1
1
-1
1
1
-1
-1
-1
21.08
19
1
-1
-1
-1
-1
1
-1
1
-1
1
1
1
33.10
20
-1
-1
1
-1
1
-1
1
1
1
1
-1
-1
23.29
A, boron; B, calcium, C, cobalt; D, copper; E, iron; F, potassium; G, magnesium; H, manganese; J, molybdenum; K, sodium; L,
nickel; M, zinc; ‘1’, high value; ‘-1‘, low value; BSC, biosurfactant concentration 9-260
251 251
Iran. Biomed. J. 21 (4): 249-260 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. (v/v) waste frying sunflower oil as carbon source. Cooled sterilized media were supplemented with filter-
sterilized urea (1.19 g/L) and then inoculated with 10%
(v/v–108
cells/mL)
inoculum
of
Pseudomonas
aeruginosa strain IKW1. Statistical analysis Flasks were incubated as
earlier described and biomass and biosurfactant
concentrations determined as described in Ekpenyong
et al.[6]. Means of triplicate determinations of
concentrations of biomass and biosurfactant from
corroborating experiments were compared with those
predicted by the regression models. of model fits where predicted responses were plotted
against experimentally-derived data. All hypotheses
were tested at 95% confidence level. For the five factors considered in the optimization
experiment, the quadratic model took the form below: where b0 is the value of the fixed response at the
central point (0, 0, 0, 0, 0); b1, b2, b3, b4, and b5 the
coefficients of the linear terms; b11, b22, b33, b44, and
b55 the coefficients of the quadratic terms; b12, b13,
b14, b15, b23, b24, b25, b34, b35, and b45 are the
coefficients of the cross products (interactive terms). where b0 is the value of the fixed response at the
central point (0, 0, 0, 0, 0); b1, b2, b3, b4, and b5 the
coefficients of the linear terms; b11, b22, b33, b44, and
b55 the coefficients of the quadratic terms; b12, b13,
b14, b15, b23, b24, b25, b34, b35, and b45 are the
coefficients of the cross products (interactive terms). Plackett-Burman design The design matrix of the PBD for the effects of 12
trace nutrients on biosurfactant production and their
responses are shown in Table 1. Results showed the
highest biosurfactant concentration of 36.02 g/L in run
10. However, the results of the trace element modeling
experiment by PBD revealed that only 5 out of 12
nutrient
elements
significantly
influenced
the
glycolipopeptide production. The non-selection of the
remaining
seven
elements
suggests
their
non-
significant (P>0.05) contributions to the response
under investigation at the confidence level selected for
the study. Verification experiments Studies to confirm the validity of the results of the
optimization experiment were done by setting up
glycolipopeptide production in the optimized medium. The medium had the same composition as that
described
in
Materials
and
Methods
with
concentrations of nickel, zinc, iron, boron, and copper
incorporated in accordance with the results of the
various optimized conditions. Fermentation was
conducted in 250-mL Erlenmeyer flasks containing 50
mL of fermentation medium supplemented with 5% Fig. 1. Normal plot of standardized effects of significant trace nutrients of a Plackett-Burman design for glycolipoeptide-
biosurfactant production. Bo is used loosely to indicate boron and not as a chemical symbol. Fig. 1. Normal plot of standardized effects of significant trace nutrients of a Plackett-Burman design for glycolipoeptide-
biosurfactant production. Bo is used loosely to indicate boron and not as a chemical symbol. Iran. Biomed. J. 21 (4): 249-260 252 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. Fig. 2. Main effects plots of contributions of significant trace elements to glycolipopeptide-biosurfactant production by
Pseudomonas aeruginosa strain IKW1. BSC, biosurfactant concentration Fig. 2. Main effects plots of contributions of significant trace elements to glycolipopeptide-biosurfactant production by
Pseudomonas aeruginosa strain IKW1. BSC, biosurfactant concentration Results of the trace element screen test by PBD
indicated the same results as trace element modeling
experiment. Figure 1 is the normal plot of the
standardized effect of the significant nutrients, showing
the magnitude and direction of their significant effects. The Figure reveals that nickel has the highest
significant positive effect on biosurfactant production
by the bacterium since its effect is positioned the
furthest to the right of the response line. Other
nutrients with significant enhancement effect on
biosurfactant production were zinc, iron, and boron,
which is in agreement with the Joshi et al.'s study[17]. However, the Figure reveals a significant reductive
effect of copper on glycolipopeptide production by the
bacterium since its effect is positioned to the left of the
biosurfactant response line. Figure 2 shows the main
effects plots of significant trace nutrients on the
response variable and confirms the results displayed in
Figure 1. Nickel has been shown to make the highest
contribution to biosurfactant production, whereas
copper exerts a negative effect on biosurfactant
production. The analysis of variance (ANOVA) result for
significant trace elements is demonstrated in Table 2
and confirms nickel as having the most significant
(P<0.05)
enhancement
effect
on
biosurfactant
production given by its very large F value. Iran. Biomed. J. 21 (4): 249-260 Verification experiments The linear
regression coefficient of determination, adjusted R2 of
91.81%, indicates that the model equation (below),
given in un-coded units, was significant and could
explain 91.81% of the variability in the response data. The equation reveals that nickel has the largest
coefficient that is preceded by a positive sign,
confirming once again its strong enhancement effect on
biosurfactant formation. Iran. Biomed. J. 21 (4): 249-260 Response surface optimization
Experimental data of the response surface method Table 3 shows the actual factor levels corresponding
to coded factor levels for a 25-1 half-fractional factorial, Table 2. Analysis of variance (ANOVA) of the regression model from the Plackett-Burman
design for trace element contribution to biosurfactant formation in un-coded units
Source
DF
Adj. SS
Adj. MS
F value
P value
Model
5
428.38
85.677
43.61
0.000
Linear
5
428.38
85.677
43.61
0.000
Bo
1
16.42
16.417
8.36
0.012
Cu
1
12.45
12.450
6.34
0.025
Fe
1
20.97
20.972
10.67
0.006
Ni
1
220.85
220.847
112.41
0.000
Zn
1
157.70
157.697
80.27
0.000
Error
14
27.51
1.965
Total
19
455.89
Model Summary: S, 1.40166; R2, 93.97%; adjusted R2, 91.81%; predicted R2,
87.69%; P<0.05, 5% significance level. Bo is used loosely to indicate boron and not as a
chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of squares. Table 2. Analysis of variance (ANOVA) of the regression model from the Plackett-Burman
design for trace element contribution to biosurfactant formation in un-coded units 253 Iran. Biomed. J. 21 (4): 249-260 Iran. Biomed. J. 21 (4): 249-260 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. Table 3. Actual factor levels corresponding to coded factor levels for 25-1 half-fractional factorial
central composite rotatable design of response surface method Table 3. Actual factor levels corresponding to coded factor levels for 25-1 half-fractional factorial
central composite rotatable design of response surface method central composite rotatable design of response surface method
Variable (mg/L)
Actual values
Code
-2
-1
0
1
2
NiCl2.6H2O
X1
0.250
0.50
0.750
1.00
1.250
ZnSO4.7H2O
X2
0.025
0.05
0.075
0.10
0.125
FeCl3
X3
0.025
0.05
0.075
0.10
0.125
K3BO3
X4
0.010
0.02
0.030
0.04
0.050
CuSO4.5H2O
X5
0.025
0.05
0.075
0.10
0.125 Table 4. Response surface optimization
Experimental data of the response surface method Actual factor levels corresponding to coded factor levels for the CCRD of the response surface optimization showing biomass
and biosurfactant concentrations
Run
order
X1
X2
X3
X4
X5
NiCl2
(mg/L)
ZnSO4
(mg/L)
FeCl3
(mg/L)
K3BO3
(mg/L)
CuSO4
(mg/L)
BMC
(g/L)
BSC
(g/L)
1
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
16.86
41.03
2
-2
0
0
0
0
0.250
0.075
0.075
0.030
0.075
14.36
29.55
3
-1
-1
-1
1
-1
0.500
0.050
0.050
0.040
0.050
8.96
36.04
4
1
-1
1
1
-1
1.000
0.050
0.100
0.040
0.050
12.98
38.32
5
0
0
0
2
0
0.750
0.075
0.075
0.050
0.075
13.52
28.39
6
0
0
0
-2
0
0.750
0.075
0.075
0.010
0.075
18.52
36.57
7
0
0
0
0
-2
0.750
0.075
0.075
0.030
0.025
15.39
46.47
8
-1
1
-1
-1
-1
0.500
0.100
0.050
0.020
0.050
11.35
38.32
9
-1
1
1
1
-1
0.500
0.100
0.100
0.040
0.050
18.26
35.31
10
1
-1
-1
-1
-1
1.000
0.050
0.050
0.020
0.050
10.62
32.19
11
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
18.76
41.22
12
0
-2
0
0
0
0.750
0.025
0.075
0.030
0.075
9.67
36.82
13
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
18.87
41.46
14
-1
1
-1
1
1
0.500
0.100
0.050
0.040
0.100
13.28
27.54
15
-1
-1
1
1
1
0.500
0.050
0.100
0.040
0.100
13.41
26.79
16
0
0
2
0
0
0.750
0.075
0.125
0.030
0.075
16.59
40.45
17
1
1
-1
1
-1
1.000
0.100
0.050
0.040
0.050
12.38
31.07
18
1
1
-1
-1
1
1.000
0.100
0.050
0.020
0.100
16.58
30.01
19
0
0
-2
0
0
0.750
0.075
0.025
0.030
0.075
8.95
28.12
20
0
2
0
0
0
0.750
0.125
0.075
0.030
0.075
15.38
46.05
21
1
1
1
1
1
1.000
0.100
0.100
0.040
0.100
8.47
39.67
22
0
0
0
0
2
0.750
0.075
0.075
0.030
0.125
11.04
37.58
23
1
-1
1
-1
1
1.000
0.050
0.100
0.020
0.100
11.94
41.40
24
-1
-1
1
-1
-1
0.500
0.050
0.100
0.020
0.050
21.07
29.56
25
1
1
1
-1
-1
1.000
0.100
0.100
0.020
0.050
19.53
57.21
26
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
18.94
42.08
27
-1
1
1
-1
1
0.500
0.100
0.100
0.020
0.100
17.39
37.99
28
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
19.04
41.68
29
-1
-1
-1
-1
1
0.500
0.050
0.050
0.020
0.100
9.47
31.07
30
0
0
0
0
0
0.750
0.075
0.075
0.030
0.075
18.83
40.78
31
1
-1
-1
1
1
1.000
0.050
0.050
0.040
0.100
9.57
30.24
32
2
0
0
0
0
1.250
0.075
0.075
0.030
0.075
12.35
39.00
X1, nickel; X2, zinc; X3, iron; X4, boron; X5, copper; BMC, biomass concentration; BSC, biosurfactant concentration Table 4. Regression model for biomass concentration, Y1 a central composite rotatable design of RSM. The
experimental responses from the 32 experimental runs
of the surface methodology are presented in Table 4. Maximum biomass concentration of 21.07 g/L was
obtained at conditions set at (X1, X2, X3, X4, X5)=(-1,
-1, 1, -1, -1) corresponding to 0.5 mg/L nickel, 0.05
mg/L zinc, 0.1 mg/L iron, 0.02 mg/L boron, and 0.05
mg/L copper. In these conditions, biosurfactant
concentration obtained was 29.56 g/L. However, the
highest biosurfactant concentration of 57.21 g/L was
acquired at conditions set at (X1, X2, X3, X4, X5)=(1, 1,
1, -1, -1) corresponding to 1 mg/L nickel, 0.1 mg/L
zinc, 0.1 mg/L iron, 0.02 mg/L boron, and 0.05 mg/L
copper and a corresponding biomass concentration of
19.53 g/L. Glycolipopeptide production yield, Yp/x,
under these conditions would be 2.93. The attainment
of peak biomass and biosurfactant concentrations at
different experimental runs suggest the requirement of
different trace nutrient conditions for cellular growth
and metabolite synthesis, especially metabolites
obtained during or near idiophasic metabolism of
organisms. Regression model for biomass concentration, Y1
The data presented in Table 4 was subjected to
multiple regression analyses using least squares
regression to fit a second-order (quadratic) regression
model for biomass concentration, Y1. The model
stipulated 20 predictors; however, the result of
ANOVA of the model in Table 5 reveals that 5
interactive terms were removed by stepwise selection,
since their contributions were not significant at P=0.05. The T values of the predictor coefficients (data not
shown) suggest that all model predictors, except the
linear terms of zinc and iron, made significant negative
contributions to the model, implying their reductive
effects on biomass formation. The model equation is
presented in un-coded units as equation 4 below: Regression model for biomass concentration, Y1
The data presented in Table 4 was subjected to
multiple regression analyses using least squares
regression to fit a second-order (quadratic) regression
model for biomass concentration, Y1. The model
stipulated 20 predictors; however, the result of
ANOVA of the model in Table 5 reveals that 5
interactive terms were removed by stepwise selection,
since their contributions were not significant at P=0.05. Table 5. Analysis of variance of the 25-1 half-fractional factorial central composite rotatable design of an response surface
method for biomass regression model in un-coded units Table 5. indicate boron and not as a chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of squares Response surface optimization
Experimental data of the response surface method Actual factor levels corresponding to coded factor levels for the CCRD of the response surface optimization showing biomass
and biosurfactant concentrations els corresponding to coded factor levels for the CCRD of the response surface optimization showing biomass
rations Iran. Biomed. J. 21 (4): 249-260 254 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. ary: S, 0.909022; R2, 97.05%; adjusted R2, 94.29%; Predicted R2, 86.75%; P<0.05, 5% significance level. Bo is used Model Summary: S, 0.909022; R , 97.05%; adjusted R , 94.29%; Predicted R , 86.75%; P<0.05, 5% significance level. Bo is used
loosely to indicate boron and not as a chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of squares Nutrient Optimization for Biosurfactant Production Ekpenyong et al. was included in the model, to increase biosurfactant
concentration as long as the adjusted R2 of the model
was not negatively affected. The model equation, in
un-coded units, is therefore presented as equation 5
below. The metal, iron, is described as the only macro-bio-
element of the heavy metals and the most biologically
relevant trace nutrient[18]. The effect of its linear term
on biomass formation was understandably pronounced,
and the significant reductive effect of its quadratic term The metal, iron, is described as the only macro-bio-
element of the heavy metals and the most biologically
relevant trace nutrient[18]. The effect of its linear term
on biomass formation was understandably pronounced,
and the significant reductive effect of its quadratic term
suggests possible toxicity at high concentrations and
therefore its trace requirement. Abalos et al.[14] have
reported a similar result trend during production of
rhamnolipid from a strain of Pseudomonas aeruginosa. suggests possible toxicity at high concentrations and
therefore its trace requirement. Abalos et al.[14] have
reported a similar result trend during production of
rhamnolipid from a strain of Pseudomonas aeruginosa. The requirement of zinc for biomass formation
derives from its role in DNA-binding proteins and a
variety of enzymes. Zinc is also known to act as a
Lewis base needed to activate water for involvement in
aqueous reactions[6]. The significance of a model is given by its goodness-
of-fit test, often expressed as the coefficient of
determination R2, which is the percentage of the
variations in the response that can be explained by
independent factors and their interactions. In this study,
the biomass regression model (Table 5) was highly
significant (F=35.12; P=0.000) with an adjusted R2 of
94.29%, indicating that only 5.71% of the variability in
the response is not explainable by the model. 2 The model was shown to have an adjusted R2 value
of 99.44%, suggesting that only 0.56% of the
variability in biosurfactant responses could not be
explained by it, and that the observed variations were
due to the factor effects and not due to noise. To
confirm the adequacy of the biosurfactant model in
explaining
the
variations
about
the
data,
an
examination of the lack-of-fit test result in the
ANOVA table was made and demonstrated a non-
significant (F=1.20; P=0.446) lack-of-fit, indicating
the adequacy of the model to explain data in the
experimental region. Nutrient Optimization for Biosurfactant Production A well-fitted model estimated by adjusted R2 might
not adequately explain data variations in the region of
experimentation. The lack-of-fit test is therefore
frequently used as a support test for adequacy of the
fitted model. The ANOVA table for the biomass
regression model in this study shows a non-significant
lack-of-fit (F=1.28; P=0.418) for the model, which
suggests its adequacy for the explanation of data
variations in the region of experimentation. A plot of experimental biosurfactant concentrations
against predicted concentrations obtained by solving
equation 5 is presented in Figure 4 as the final
ratification of the fit of the biosurfactant regression
model and confirms a goodness-of-fit, R2 (unadjusted)
value of 99.97%. The regression plot of experimental against predicted
biomass responses is presented as Figure 3 and reveals
a coefficient of determination, R2 of 0.9705, which is in
agreement with the R2 for the model in the ANOVA
table. Fig. 3. Experimental biomass concentration plotted against
biomass concentration predicted by the fitted model. y = 0.9707x + 0.4223
R² = 0.9705
0
5
10
15
20
25
0
5
10
15
20
25
Experimental biomass concentration (g/L)
Predicted biomass concentration (g/L) y = 0.9707x + 0.4223
R² = 0.9705
0
5
10
15
20
25
0
5
10
15
20
25
Experimental biomass concentration (g/L)
Predicted biomass concentration (g/L) Regression model for biomass concentration, Y1 Analysis of variance of the 25-1 half-fractional factorial central composite rotatable design of an response surface
method for biomass regression model in un-coded units Table 5. Analysis of variance of the 25-1 half-fractional factorial central composite rotatable design of an response surface
method for biomass regression model in un-coded units method for biomass regression model in un-coded units
Source
DF
Adjusted SS
Adjusted MS
F value
P value
Model
15
435.332
29.0222
35.12
0.000
Linear
5
199.895
39.9790
48.38
0.000
Ni
1
9.551
9.5508
11.56
0.004
Zn
1
39.117
39.1171
47.34
0.000
Fe
1
88.627
88.6273
107.26
0.000
Bo
1
39.117
39.1171
47.34
0.000
Cu
1
23.483
23.4828
28.42
0.000
Square
5
134.701
26.9401
32.60
0.000
Ni*Ni
1
35.179
35.1787
42.57
0.000
Zn*Zn
1
49.773
49.7729
60.23
0.000
Fe*Fe
1
45.202
45.2022
54.70
0.000
Bo*Bo
1
5.395
5.3951
6.53
0.021
Cu*Cu
1
37.463
37.4633
45.34
0.000
Two-way interaction
5
100.737
20.1473
24.38
0.000
Ni*Fe
1
33.931
33.9306
41.06
0.000
Ni*Bo
1
6.126
6.1256
7.41
0.015
Zn*Fe
1
7.182
7.1824
8.69
0.009
Fe*Bo
1
10.530
10.5300
12.74
0.003
Fe*Cu
1
42.968
42.9680
52.00
0.000
Error
16
13.221
0.8263
Lack-of-fit
11
9.748
0.8862
1.28
0.418
Pure error
5
3.473
0.6946
Total
31
448.554
Model Summary: S, 0.909022; R2, 97.05%; adjusted R2, 94.29%; Predicted R2, 86.75%; P<0.05, 5% significance level. Bo is used
loosely to indicate boron and not as a chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of squares Iran. Biomed. J. 21 (4): 249-260 255 Nutrient Optimization for Biosurfactant Production Regression model for biosurfactant concentration, Analysis of variance of the 25-1 half fractional factorial central composite rotatable design (CCRD
surface method for biosurfactant regression model in un-coded units Model Summary: S, 0.499416; R2, 99.71%; adjusted R2, 99.44%; predicted R2, 98.77%; P<0.05, 5% significance level. Bo is
used loosely to indicate boron and not as a chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of
squares. boron, with the concentrations of nickel, zinc, and
iron supplied at high levels. Regression model for biosurfactant concentration, The ANOVA for the biosurfactant regression model
presented in Table 6 indicated that the model was
highly significant (F=371.07; P=0.000). All the linear
predictor terms and only the square terms of nickel,
iron, and boron were significant. Six of the interactive
predictor terms were also significant; however, Fe*Cu
interaction, although included in the model, was not
significant (P=0.051). The magnitude and direction of
significant effect of model predictors are given by the
T values of the coefficient estimates of the predictors
(data not shown) and reveal that the linear, but not the
quadratic terms of nickel, zinc, and iron, significantly
enhanced biosurfactant formation. It is probably
because of the enhancing effect of Fe*Cu that the term Experimental biomass concentration (g/L) Fig. 3. Experimental biomass concentration plotted against
biomass concentration predicted by the fitted model. Iran. Biomed. J. 21 (4): 249-260 256 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. Table 6. Analysis of variance of the 25-1 half fractional factorial central composite rotatable design (CCRD) of an response
surface method for biosurfactant regression model in un-coded units
Source
DF
Adjusted SS
Adjusted MS
F value
P value
Model
15
1388.26
92.551
371.07
0.000
Linear
5
676.69
135.338
542.62
0.000
Ni
1
132.49
132.493
531.21
0.000
Zn
1
104.04
104.042
417.14
0.000
Fe
1
230.83
230.826
925.46
0.000
Bo
1
100.57
100.573
403.23
0.000
Cu
1
108.76
108.758
436.05
0.000
Square
3
326.18
108.726
435.92
0.000
Ni*Ni
1
105.31
105.312
422.24
0.000
Fe*Fe
1
105.03
103.033
421.11
0.000
Bo*Bo
1
161.50
161.502
647.52
0.000
2-Way Interaction
7
385.39
55.056
220.74
0.000
Ni*Fe
1
198.88
198.881
797.38
0.000
Ni*Bo
1
6.57
6.566
26.33
0.000
Zn*Fe
1
84.23
84.227
337.69
0.000
Zn*Bo
1
45.93
45.935
184.17
0.000
Zn*Cu
1
25.23
25.226
101.14
0.000
Fe*Bo
1
23.45
23.450
94.02
0.000
Fe*Cu
1
1.11
1.108
4.44
0.051
Error
16
3.99
0.249
Lack-of-Fit
11
2.90
0.263
1.20
0.446
Pure Error
5
1.09
0.219
Total
31
1392.25
Model Summary: S, 0.499416; R2, 99.71%; adjusted R2, 99.44%; predicted R2, 98.77%; P<0.05, 5% significance level. Bo is
used loosely to indicate boron and not as a chemical symbol. DF, degrees of freedom; SS, sum of squares; MS, mean sum of
squares. Table 6. Contour and surface plots for biomass (Y1) and
biosurfactant (Y2) concentrations Boron,
a
micro-nutrient
with
intermediary
properties of metals and non-metals, is known as
a regulator of certain pathways that require
serine proteases or oxidoreductases involving co-
enzymes like nicotinamide adenine dinucleotide and Biomass response plots were made with the vertical
axis representing biomass concentration (Y1) and two
horizontal axes representing the most significant
two- way interaction (X3, X5)=(Fe, Cu). The plots
led to maximal biomass formation with the remaining
three factors (X1, X2, X4)=(Ni, Zn, Bo) held at their
optimum levels. Biomass concentration under this
condition was in excess of 20 g/L. The contour and
surface plots displayed in Figure 5 reveal that
maximal biomass would be accumulated under the
high levels of iron and low levels of copper. y = 0.9972x + 0.1022
R² = 0.9971
0
10
20
30
40
50
60
70
0
20
40
60
80
Predicted glycolipopeptide
concentration (g/L)
Experimental glycolipopeptide
concentration (g/L) y = 0.9972x + 0.1022
R² = 0.9971
0
10
20
30
40
50
60
70
0
20
40
60
80
Predicted glycolipopeptide
concentration (g/L)
E
i
l l
li
id g
pp
The biosurfactant response plots (Fig. 6) were made
with the vertical axis representing biosurfactant
concentration
(Y2)
and
two
horizontal
axes
representing the two most significant variables (X4,
X5)=(Bo, Cu) that led to maximal biosurfactant
formation with the remaining three factors (X1, X2,
X3)=(Ni, Zn, Fe) held at their optimum levels. Maximal biosurfactant concentration under this
condition was in excess of 80 g/L. The contour and
surface plots reveal that the highest biosurfactant
concentration would be obtained when fermentation
medium contains low levels of both copper and Experimental glycolipopeptide
concentration (g/L) Experimental glycolipopeptide
concentration (g/L) Fig. 4. Experimental biosurfactant concentrations versus
theoretical values predicted by the regression model. Iran. Biomed. J. 21 (4): 249-260 257 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. Fig. 5. Contour (A) and surface (B) plots of two-way
interactions of independent variables for maximal biomass
production. Bo is used loosely to indicate boron and not as a
chemical symbol. Contour and surface plots for biomass (Y1) and
biosurfactant (Y2) concentrations BMC, biomass concentration
2
0
0
10
-2
0
-2
2
20
C
M
B
u
C
e
F
urface pl
S
t for bi
o
mass concentr
o
(A)
(B) urface pl
S
t for bi
o
mass concentr
o
(A) biosurfactant production by Pseudomonas aeruginosa
strain IKW1 (unpublished data) demystifies the
significant
enhancement
effect
of
nickel
on
biosurfactant production and suggests a correlation
between urease activity and biosurfactant production
in the bacterium. Selection of optimum conditions for response
variables For situations where biomass accumulation is
desired instead of the surface-active compound,
which was not the case in this study, the highest
predictable biomass concentration of 26.55 g/L was
obtained at trace mineral conditions set at (X1, X2,
X3, X4, X5)=(-1.0707, -0.0202, 2.0, -2.0, -1.8788)
corresponding to 0.482 mg/L nickel, 0.074 mg/L
zinc, 0.125 mg/L iron, 0.010 mg/L boron, and 0.028
mg/L copper. Ni 0
Zn 0
Bo 0
Hold Values
on 2
0
0
10
-2
0
-2
2
20
C
M
B
u
C
e
F
(B) (B) g
pp
For tertiary oil recovery and spilled oil remediation
where maximum concentrations of both responses Fig. 6. Contour (A) and surface (B) plots of two-way
interactions
of
independent
variables
for
maximal
glycolipopeptide production. Bo is used loosely to indicate
boron and not as a chemical symbol. BSC, biosurfactant
concentration
Bo
Cu
2
1
0
-1
-2
2
1
0
-1
-2
p
40
60
2
-
0
2
-
2
0
2
80
C
S
B
u
C
o
B
urface plot
S
or biosu
f
factant concen
r
(B)
(A)
Ni 2
Zn 2
Fe 2
Hold Values
Bo
2
1
0
-1
-2
2
1
0
-1
-2
>
–
–
–
–
–
< 30
30
40
40
50
50
60
60
70
70
80
80
BSC
Contour plot for biosurfactant concentration Bo
Cu
2
1
0
-1
-2
2
1
0
-1
-2
urface plot
S
or biosu
f
factant conce
r
(A)
Ni 2
Zn 2
Fe 2
Hold Values
>
–
–
–
–
–
< 30
30
40
40
50
50
60
60
70
70
80
80
BSC
ontour plot for biosurfactant concentration 0
e
F 2 Fig. 5. Contour (A) and surface (B) plots of two-way
interactions of independent variables for maximal biomass
production. Bo is used loosely to indicate boron and not as a
chemical symbol. BMC, biomass concentration nicotinamide adenine dinucleotide phosphate for
activity[22]. These activities of boron in bacteria could
very well be the reason for its very little requirement
both for cellular growth and secondary metabolism. nicotinamide adenine dinucleotide phosphate for
activity[22]. These activities of boron in bacteria could
very well be the reason for its very little requirement
both for cellular growth and secondary metabolism. Selection of optimum conditions for response
variables Copper, on the other hand, is mostly important in
cytochrome C oxidases; oxygen-dependent terminal
oxidases in the electron transport chain of aerobic
organisms[23]. The interaction therefore between
boron and copper has to be very important in
biosurfactant synthesis since the whole process is
energy driven; copper to generate energy and boron
to inhibit the process. 40
60
2
-
0
2
-
2
0
2
80
C
S
B
u
C
o
B
(B)
Bo
2
1
0
-1
2 (B) Copper, on the other hand, is mostly important in
cytochrome C oxidases; oxygen-dependent terminal
oxidases in the electron transport chain of aerobic
organisms[23]. The interaction therefore between
boron and copper has to be very important in
biosurfactant synthesis since the whole process is
energy driven; copper to generate energy and boron
to inhibit the process. Nickel is a transition metal useful only in a few
selected reactions. It associates with iron in NiFe
hydrogenases; an enzyme system that splits molecular
hydrogen into protons and electrons. The metal is
also intrinsically bound to urease; an enzyme that
catalyses the splitting of urea into carbon dioxide and
ammonia thus supplying nitrogen in its available
form for cellular metabolism[18,24]. The preference of
urea as a nitrogen source for glycolipopeptide 0
o
B Fig. 6. Contour (A) and surface (B) plots of two-way
interactions
of
independent
variables
for
maximal
glycolipopeptide production. Bo is used loosely to indicate
boron and not as a chemical symbol. BSC, biosurfactant
concentration Fig. 6. Contour (A) and surface (B) plots of two-way
interactions
of
independent
variables
for
maximal
glycolipopeptide production. Bo is used loosely to indicate
boron and not as a chemical symbol. BSC, biosurfactant
concentration 258 Iran. Biomed. J. 21 (4): 249-260 Nutrient Optimization for Biosurfactant Production Ekpenyong et al. Table 7. Selection of optimum conditions for response
variables Design codes, actual values, experimental and predicted responses of validation
experiments of an RSM for glycolipopeptide production
Parameters
BMC
maximized
BSC
maximized
BMC and BSC
maximized
X1
-1.0707
2
0.2626
X2
-0.0202
2
0.9495
X3
2
2
2
X4
-2
-1.9596
-2
X5
-1.8788
-2
-2
Ni (mg/L)
0.482
1.25
0.816
Zn (mg/L)
0.074
0.125
0.099
Fe (mg/L)
0.125
0.125
0.125
Bo (mg/L)
0.010
0.010
0.010
Cu (mg/L)
0.028
0.025
0.025
eBMC (g/L
25.74
19.14
25.14
pBMC (g/L)
26.55
NP
23.40
eBSC (g/L)
25.96
84.44
56.83
pBSC (g/L)
NP
81.92
57.55
X1, nickel (Ni); X2, zinc (Zn); X3, iron (Fe); X4, boron (Bo); X5, copper (Cu); eBMC,
experimental biomass concentration; pBMC, predicted biomass concentration; eBSC,
experimental biosurfactant concentration; pBSC, predicted biosurfactant concentration; NP,
not predicted. Bo is used loosely to indicate boron and not as a chemical symbol. Table 7. Design codes, actual values, experimental and predicted responses of validation
experiments of an RSM for glycolipopeptide production 1
(
)
2
(
)
3
(
)
4
(
)
pp
(
)
experimental biomass concentration; pBMC, predicted biomass concentration; eBSC,
experimental biosurfactant concentration; pBSC, predicted biosurfactant concentration; NP,
not predicted. Bo is used loosely to indicate boron and not as a chemical symbol. are desired, trace mineral conditions set at (X1, X2,
X3, X4, X5)=(0.2626, 0.9495, 2.0, -2.0, -2.0)
corresponding to 0.816 mg/L nickel, 0.099 mg/L zinc,
0.125 mg/L iron, 0.01 mg/L boron, and 0.025
mg/L copper, which would lead to 23.4 g/L biomass
and 57.55 g/L biosurfactant. concentration of 84.44 g/L, when compared to 23.86
g/L obtained from a previous (control) experiment
(unpublished data), reveals a ~3.54fold increase in
biosurfactant concentration, thereby making separate
optimization
experiments
for
trace
nutrients
appropriate for developing the fermentation media for
the production of microbial metabolites. are desired, trace mineral conditions set at (X1, X2,
X3, X4, X5)=(0.2626, 0.9495, 2.0, -2.0, -2.0)
corresponding to 0.816 mg/L nickel, 0.099 mg/L zinc,
0.125 mg/L iron, 0.01 mg/L boron, and 0.025
mg/L copper, which would lead to 23.4 g/L biomass
and 57.55 g/L biosurfactant. Finally,
for
detergent,
food,
cosmetic,
and
pharmaceutical applications where maximum surface-
active compound and zero microbial cells are desired,
the response optimizer set conditions for maximum
biosurfactant concentration of 81.92 g/L at (X1, X2,
X3, X4, X5)=(2.0, 2.0, 2.0, -1.9596, -2) corresponding
to 1.25 mg/L nickel, 0.125 mg/L zinc, 0.125 mg/L iron,
0.0104 mg/L boron, and 0.025 mg/L copper. Verification experiments p
Table
7
shows
that
maximum
biosurfactant
concentration obtained in the validation experiments
under conditions (X1, X2, X3, X4, X5) =(2, 2, 2, -
1.9596, -2) corresponding to 1.25 mg/L nickel, 0.125
mg/L zinc, 0.125 mg/L iron, 0.0104 mg/L boron, and
0.025
mg/L
copper
was
84.44
g/L
with
a
corresponding biomass concentration of 19.14 g/L,
giving a production yield (Yp/x) of 4.41. When
fermentation conditions were set to maximize both
responses, biosurfactant yield of 2.26 was achieved. Compared to the 81.92 g/L biosurfactant concentration
predicted by the optimizer, the concentration obtained
in the confirmation experiment was only 0.03% higher,
thus validating the prediction of the response
optimizer. A
glycolipopeptide
biosurfactant Selection of optimum conditions for response
variables PBD selected nickel, zinc, iron, boron, and copper as
the most significant (P<0.05; adjusted R2=91.82%)
trace minerals for glycolipopeptide biosurfactant
production. Optimization of the nutrients by RSM
resulted in 84.44 g/L of the biosurfactant under
conditions set at (X1, X2, X3, X4, X5)=(2, 2, 2, -
1.9596, -2). These conditions corresponded to 1.2500
mg/L nickel, 0.125 mg/L zinc, 0.125 mg/L iron, 0.0104
mg/L boron, and 0.025 mg/L copper, giving a
production yield, Yp/x of 4.41. In conclusion, PBD and
RSM are dependable tools for selecting and optimizing
conditions of nutrients for biosurfactant production. REFERENCES 14. Abalos A, Maximo F, Manresa MA, Bastida J. Utilization of response surface methodology to optimize
the culture media for the production of rhamnolipids by
Pseudomonas aeruginosa AT10. Journal of chemical
technology and biotechnology 2002; 77(7): 777-784. 1. Desai JD, Banat IM. Microbial production of surfactants
and their commercial potential. Microbiology and
molecular biology reviews 1997; 61(1): 47-64. 2. Abouseoud M, Maachi R, Amrane A, Boudergua S,
Nabi A. Evaluation of different carbon and nitrogen
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Bramhachari
PV. Optimization
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16. Rodrigues L, Teixeira J, Oliveira R, van der Mei H C. Response
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components for the production of biosurfactants by
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10. 4. Bodour AA, Drecs KP, Maier RM. Distribution of
biosurfactant-producing bacteria in undisturbed and
contaminated arid southwestern soils. Applied and
environmental microbiology 2003; 69(6): 3280-3287. 17. Joshi S, Yadav S, Nerurkar A, Desai AJ. Statistical
optimization of medium components for the production
of biosurfactant by Bacillus licheniformis K51. Journal
of microbiology and biotechnology 2007; 17(2): 313-
319. 5. Morikawa M, Daido H, Takao T, Murata S, Shimonishi
Y, Imanaka T. A new lipopeptide biosurfactant
produced by Arthrobacter sp. strain MIS38. Journal of
bacteriology 1993; 175(20): 6459-6466. 6. Ekpenyong M, Antai SP, Asitok, A. A Pseudomonas
aeruginosa strain IKW1 produces an unusual polymeric
surface-active compound in waste frying oil-minimal
medium. International journal of sciences 2016; 5(6):
108-123. 18. Nies DH. Microbial heavy-metal resistance. Applied
microbiology and biotechnology 1999; 51(6): 730-750. 19. Cavazzuti M. Optimization Methods: from Theory to
Design, London: Springer, 2013. 20. Myers R H, Montgomery D C. Response Surface
Methodology: Product and Process Optimization Using
Designed Experiments. Second ed. New York: John
Wiley & Sons, 2002. 7. Ilori MO, Amund DI. Production of a peptidoglycolipid
bioemulsifier from Pseudomonas aeruginosa grown on
hydrocarbon. Zeitschrift für naturforschung c 2001;
56(7-8): 547-552. 21. Raorane CJ, Gavimath CC, Kukreja GP, Kalsekar DP,
Kulkarni SM, Ravishankar, BE, Hooli RS. ACKNOWLEDGEMENTS The authors are grateful to the government of Cross
River
State,
Nigeria
for
the
research
grant
(CRS/SB/P/149/30) awarded for this work through
their Ministry of Education, Scholarship and Grant
unit. Sincere gratitude also goes to Mr. Obi Oyeka-
Director of Bez Pharmacy and Stores, Calabar, Nigeria
and Dr. Samuel Ekpata for their financial assistance
towards this research. 259 Iran. Biomed. J. 21 (4): 249-260 Ekpenyong et al. Nutrient Optimization for Biosurfactant Production CONFLICT OF INTEREST. None declared. 13. Thavasi R, Subramanyam Nambaru VRM, Jayalakshmi
S, Balasubramanian T, Banat IM. Biosurfactant
production by Pseudomonas aeruginosa from renewable
resources. Indian journal of microbiology 2011; 51(1):
30-36. REFERENCES Isolation,
identification and phylogenetic analysis of phthalic acid-
degrading bacteria. International journal of advanced
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production
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Corynebacterium
kutscheri from waste motor lubricant oil and peanut oil
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Cameotra
SS. Environmental
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biosurfactants: recent advances. International journal of
molecular sciences 2011; 12(1): 633-654. 23. Michel H, Behr J, Harrenga A, Kannt A. Cytochrome C
oxidase: structure and spectroscopy. Annual review of
biophysics and biomolecular structure 1998; 27: 329-
356. 11. Sachdev
DP,
Cameotra
SS. Biosurfactants
in
agriculture. Applied microbiology and biotechnology
2013; 97(3): 1005-1016. 24. Maier T, Jacobi A, Sauter M, Bock A. The product of
the hypB gene, which is required for nickel
incorporation into hydrogenases, is a novel guanine
nucleotide-binding protein. Journal of bacteriology
1993; 175(3): 630-635. 12. Sekhon KK, Khanna S, Cameotra SS. Enhanced
biosurfactant production through cloning of three genes
and role of esterase in biosurfactant release. Microbial
cell factories 2011; 10: 49. Iran. Biomed. J. 21 (4): 249-260 260
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English
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Measurement of prompt D0 and <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" altimg="si1.svg"><mml:msup><mml:mrow><mml:mover accent="true"><mml:mrow><mml:mi mathvariant="normal">D</mml:mi></mml:mrow><mml:mo>‾</mml:mo></mml:mover></mml:mrow><mml:mrow><mml:mn>0</mml:mn></mml:mrow></mml:msup></mml:math> meson azimuthal anisotropy and search for strong electric fields in PbPb collisions at <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" altimg="si2.svg"><mml:msqrt><mml:mrow><mml:…
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Physics letters. B
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cc-by
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1. Introduction pected to receive important contributions from medium energy
loss and coalescence effects [17,18]. In ultrarelativistic heavy ion collisions, very strong and tran-
sient (∼10−1 fm/c) magnetic and electric fields are expected to
be induced by the collision spectators and participants [19]. Such
electromagnetic (EM) fields are predicted to produce a difference
in the vn harmonics for positively and negatively charged parti-
cles [19]. In such a picture, the magnetic field is mainly responsible
for splitting the rapidity (y)-odd directed flow (v1) [19,20]. The
electric field is predicted to induce a charge-dependent splitting in
the v2 coefficient and in the average transverse momentum (⟨pT⟩)
values of the emitted particles [19]. As charm quarks are expected
to be created very early in the collision, they have a higher proba-
bility of interacting with this strong EM field than the light flavor
quarks [20,21]. The observation of a strongly-coupled quark-gluon plasma
(QGP), a state of matter composed of deconfined quarks and glu-
ons, was established by experiments investigating ultrarelativistic
heavy ion collisions at the BNL RHIC [1–4] and CERN LHC [5,6]. The azimuthal particle correlations constitute an effective tool to
probe the properties of the QGP [1–9]. These correlations are pa-
rameterized by a Fourier expansion [10–12], with the magnitude
of the Fourier coefficients, vn, providing information about the
initial collision geometry and its fluctuations [12]. The second-
(v2) and third- (v3) order Fourier coefficients are referred to as
“elliptic” and “triangular” flow harmonics, respectively. Measuring
these coefficients for particle species with different quark compo-
sition provides additional information about this hot and dense
medium [13]. Because of their large mass, charm and bottom
quarks are produced earlier in the collisions than the light quarks
(up and down) [14,15]. In addition, the charm and bottom quarks
have masses many times larger than the typical temperatures in
the QGP [16]. These heavy quarks experience the full evolution of
the medium until the hadronization phase. As a consequence, the
vn of charmed D0 (uc) and D
0 (uc) mesons (henceforth referred
to as D0 mesons, except where explicitly stated otherwise) are ex- In this letter, measurements of the v2 and v3 coefficients as
functions of D0 meson rapidity, pT, and lead-lead (PbPb) collision
centrality are presented. a r t i c l e
i n f o Article history:
Received 29 September 2020
Received in revised form 21 February 2021
Accepted 25 March 2021
Available online 29 March 2021
Editor: M. Doser
Keywords:
CMS
Heavy-flavor
Charm
Electromagnetic fields The strong Coulomb field created in ultrarelativistic heavy ion collisions is expected to produce a rapidity-
dependent difference (v2) in the second Fourier coefficient of the azimuthal distribution (elliptic flow, Article history:
Received 29 September 2020
Received in revised form 21 February 2021
Accepted 25 March 2021
Available online 29 March 2021
Editor: M. Doser The strong Coulomb field created in ultrarelativistic heavy ion collisions is expected to produce a rapidity-
dependent difference (v2) in the second Fourier coefficient of the azimuthal distribution (elliptic flow,
v2) between D0 (uc) and D
0 (uc) mesons. Motivated by the search for evidence of this field, the CMS
detector at the LHC is used to perform the first measurement of v2. The rapidity-averaged value is
found to be ⟨v2⟩ = 0.001 ±0.001 (stat)±0.003 (syst) in PbPb collisions at √
sNN = 5.02 TeV. In addition,
0 v2) between D0 (uc) and D
0 (uc) mesons. Motivated by the search for evidence of this field, the CMS
detector at the LHC is used to perform the first measurement of v2. The rapidity-averaged value is
found to be ⟨v2⟩ = 0.001 ±0.001 (stat)±0.003 (syst) in PbPb collisions at √
sNN = 5.02 TeV. In addition,
0 the influence of the collision geometry is explored by measuring the D0 and D
0mesons v2 and triangular
flow coefficient (v3) as functions of rapidity, transverse momentum (pT), and event centrality (a measure
of the overlap of the two Pb nuclei). A clear centrality dependence of prompt D0 meson v2 values is
observed, while the v3 is largely independent of centrality. These trends are consistent with expectations
of flow driven by the initial-state geometry. Keywords:
CMS
Heavy-flavor
Charm
Electromagnetic fields © 2021 The Author. Published by Elsevier B.V. This is an open access article under the CC BY license
(http://creativecommons.org/licenses/by/4.0/). Funded by SCOAP3. Measurement of prompt D0 and D
0 meson azimuthal anisotropy and
search for strong electric fields in PbPb collisions at √sNN = 5.02 TeV Measurement of prompt D0 and D
0 meson azimuthal anisotropy and
search for strong electric fields in PbPb collisions at √sNN = 5.02 TeV .The CMS Collaboration ⋆ CERN, Switzerland CERN, Switzerland Contents lists available at ScienceDirect Contents lists available at ScienceDirect Physics Letters B 816 (2021) 136253 Physics Letters B 816 (2021) 136253 ⋆E-mail address: cms -publication -committee -chair @cern .ch.
considered as a systematic uncertainty. Using the data recorded in
PbPb collisions during the 2018 LHC run period, corresponding to
https://doi.org/10.1016/j.physletb.2021.136253
0370-2693/© 2021 The Author. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). Funded by
SCOAP3. ⋆E-mail address: cms -publication -committee -chair @cern .ch. ed by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). Funded by 1. Introduction The collision centrality bins are given in
percentage ranges of the total inelastic hadronic cross section, with
the 0–10% centrality bin corresponding to the 10% of collisions
having the largest overlap of the two nuclei. The flow harmonics
are measured using the scalar product method [22,23]. In this anal-
ysis, the selection of D0 mesons uses multivariate methods [24]
for selecting D0 candidates and their antiparticles. The contamina-
tion from nonprompt D0 candidates, arising from B meson decay, is
considered as a systematic uncertainty. Using the data recorded in
PbPb collisions during the 2018 LHC run period, corresponding to The CMS Collaboration Physics Letters B 816 (2021) 136253 of tracks from oppositely charged particles and requiring an in-
variant mass (minv) within a ±200 MeV/c2 window of the world-
average D0 meson mass of (1864.83 ± 0.05) MeV/c2 [33]. For each
pair of selected tracks, two possible candidates for D0 and D
0
mesons are considered by assuming one of the tracks has the pion
mass, while the other track has the kaon mass, and vice versa. Kinematic vertex fits are performed to reconstruct the secondary
vertices of D0 candidate decays. 0.58 nb−1 of integrated luminosity, the flow coefficients are mea-
sured within the rapidity range |y| < 2, which is twice as large as
achieved in previous CMS measurements [25]. The extension of the
measurements to this larger rapidity range, together with smaller
statistical uncertainties provided by a larger data set, furnish im-
portant inputs for a better understanding of the three-dimensional
evolution of the QGP formed in heavy ion collisions. Measurements
of the v2 difference between D0 and D
0 mesons, v2, as a func-
tion of rapidity are presented as a method to probe possible effects
originating from the Coulomb fields. of tracks from oppositely charged particles and requiring an in-
variant mass (minv) within a ±200 MeV/c2 window of the world-
average D0 meson mass of (1864.83 ± 0.05) MeV/c2 [33]. For each
0 pair of selected tracks, two possible candidates for D0 and D
0
mesons are considered by assuming one of the tracks has the pion
mass, while the other track has the kaon mass, and vice versa. Kinematic vertex fits are performed to reconstruct the secondary
vertices of D0 candidate decays. 4. Analysis technique The elliptic and triangular flow coefficients of D0 mesons are
extracted using the scalar product (SP) method, similarly to what
was done in a previous CMS publication [25]. In this method, the
vn coefficients of D0 candidates (including backgrounds) are mea-
sured using Events from Monte Carlo (MC) simulations are used to study
both prompt and nonprompt D0 meson processes. The events are
generated using an embedding procedure, in which D0 mesons
generated by pythia 8.212 [29] (tune CP5 [30]) are embedded into
MB events from hydjet 1.9 [31]. A full simulation of the CMS de-
tector is performed using Geant4 [32]. The prompt D0 meson MC
simulation is employed to define signal selections and measure ef-
ficiency corrections, while the nonprompt D0 meson MC sample is
used to estimate systematic uncertainties coming from nonprompt
D0 contamination. vn{SP} ≡
⟨Q D0
n Q ∗
nA⟩
⟨Q nA Q ∗
nB⟩⟨Q nA Q ∗
nC⟩
⟨Q nB Q ∗
nC⟩
,
(1) (1) with the Q -vectors expressed as Q n ≡M
j=1 wjeinφj, where the
sum is over the total number (M) of HF towers above a certain
energy threshold (with the weights wj taken as the energy de-
posited in the HF tower at azimuthal angle φj), of tracks with pT
above a certain threshold (with wj taken as track pT in φj angle),
or of selected D0 meson candidates (with wj taken equal to 1). 2. Experimental apparatus and data sample The central feature of the CMS apparatus is a superconduct-
ing solenoid of 6 m internal diameter, providing a magnetic field
of 3.8 T. Within the solenoid volume, there are four primary
subdetectors including a silicon pixel and strip tracker detector,
a lead tungstate crystal electromagnetic calorimeter, and a brass
and scintillator hadron calorimeter, each composed of a barrel
and two endcap sections. Iron and quartz-fiber Cherenkov hadron
forward (HF) calorimeters cover the pseudorapidity range 2.9 <
|η| < 5.2. The HF calorimeters are segmented to form 0.175×0.175
(η×φ) towers. Muons are measured in gas-ionization detec-
tors embedded in the steel flux-return yoke outside the solenoid. The silicon tracker measures charged particles within the range
|η| < 2.5. A detailed description of the CMS detector, together with
a definition of the coordinate system used and the relevant kine-
matic variables, can be found in Ref. [26]. The analysis presented in this letter uses approximately 4.27 ×
109 minimum bias (MB) PbPb collision events collected by the CMS
experiment during the 2018 LHC run. The MB events are triggered
by requiring signals in both forward and backward sides of the HF
calorimeters [27]. Further selections are applied offline to reject
events from background processes (beam-gas interactions and non-
hadronic collisions), see Ref. [28] for details. Events are required to
have at least one interaction vertex, reconstructed based on two
tracks or more, and with a distance of less than 15 cm from the
center of the nominal interaction point along the beam axis. The
primary interaction vertex is defined as the one with the high-
est track multiplicity in the event. The shapes of the clusters in
the pixel detector have to be compatible with those expected from
particles produced at the primary vertex location. The PbPb colli-
sion events are also required to have at least two calorimeter tow-
ers in each HF detector with energy deposits of more than 4 GeV
per tower. These criteria select (99 ±2)% of inelastic hadronic PbPb
collisions. The possibility to have values higher than 100% reflects
the possible presence of ultra-peripheral (nonhadronic) collisions
in the selected event sample. 1. Introduction y
After the D0
candidate reconstruction, a selection using
a boosted decision tree (BDT) algorithm from the tmva pack-
age [24] is employed. For the BDT training, misidentified D0 candi-
dates in data events, where pion and kaon have the same charge,
are used to mimic the combinatorial background. The signal candi-
dates are taken from MC simulations of prompt D0 mesons and are
required to match D0 particles at the generator level. The variables
related to D0 mesons used to discriminate the signal from the
background are: χ 2 probability for the D0 vertex fit, 3D distance
between the secondary and primary vertices and its significance,
the decay length significance projected in the xy-plane, and the
angle in two and three dimensions between the momentum of the
D0 meson candidate and the line connecting the primary and the
secondary vertices (pointing angle). Related to the decay products
of the D0 meson candidate, the variables used are: the uncertainty
in pT returned by the track fitting procedure, the significance of
the z and the xy distances of closest approach to the primary ver-
tex, and the number of hits in the tracker detector. These variables
are chosen by analyzing their BDT ranking (variables more fre-
quently used in the decision tree) and correlation matrix among
all variables. Different BDT boost algorithms are tested, choosing
the adaptive boost algorithm [24] as default. Overtraining checks
are done for all analysis bins by comparing the BDT distributions
from training and testing D0 meson samples. In addition, a BDT cut
optimization is performed in bins of centrality, pT, and rapidity,
doing a scan in different BDT scores and finding the one resulting
in maximal D0 mesons signal significance for each analysis bin. Compared to a cutoff-based procedure, this BDT selection almost
doubles the signal significance for D0 mesons in 1 < |y| < 2, and
increases the signal significance by 30% for D0 mesons in |y| < 1,
for events with collision centrality in the range 0–30%. For the
remaining analysis bins a similar performance of BDT and cutoff-
based methods is observed. 5. Systematic uncertainties vsig+bkg
n
(minv) = α(minv)vsig
n + [1 −α(minv)]vbkg
n (minv). (3) (3) The sources of systematic uncertainties include the D0 identi-
fication requirements (BDT selection); the probability distribution
function (PDF) for modeling the background in the invariant mass
fit; the impact of acceptance and efficiency of the D0 meson yield;
the variation of the PDF for modeling the background vn; and the
remaining nonprompt D0 contamination. With the exception of the
last component, the uncertainties are quoted as absolute values
of vn and vn after comparing the default analysis configuration
with the variations. To diminish the influence of statistical fluctua-
tions, after observing no special trends in the deviations from the
default measurements, the systematic uncertainties are evaluated
by averaging the deviations with a constant fit as a function of the
analysis bins. The α(minv) parameter, which characterizes the signal fraction as
a function of mass, is defined as follows: The α(minv) parameter, which characterizes the signal fraction as
a function of mass, is defined as follows: α(minv) = [S(minv) + SW (minv)]/[S(minv) + SW (minv) + B(minv)]
= αsignal(minv) + αswap(minv). (4 (4) For extracting the difference vsig
n , the following expression is em-
ployed: For extracting the difference vsig
n , the following expression is em-
ployed: vsig+bkg
n
(minv) = vsig
n (αsignal(minv) −αswap(minv)) + const. vsig+bkg
n
(minv) = vsig
n (αsignal(minv) −αswap(minv)) + const. In order to take into account the systematic uncertainty associ-
ated with the BDT selection, the BDT cut is varied up and down by
the maximal deviation between the BDT optimized selection based
on MC simulations and data. The BDT cuts (and variations for sys-
tematic uncertainties) are defined in bins of collision centrality, pT,
and rapidity, ranging from 0.28 to 0.47 (±0.02–0.03). Regarding
the effect of the background mass modeling, either an exponential
function together with a second order polynomial, or just a second
order polynomial, are considered instead of the default fit function
using a third-order polynomial. To fit vn as a function of mass, the
default configuration using a linear function is replaced by either
a constant or a second order polynomial. Although the D0 meson
selection efficiency essentially cancels in vn measurements, a sys-
tematic uncertainty is assigned by comparing the results with and
without applying corrections based on MC simulations in bins of
pT and rapidity. 3. Reconstruction and selection of D0 mesons The Q -vectors related to HF and the tracker are measured and
corrected for detector irregularities by applying a flattening and a
recentering procedure [12,34]. The Q nA and Q nB are defined using
the event-plane measurements from the negative (−5 < η < −3,
HF−) and the positive (3 < η < 5, HF+) sides of HF, and Q nC is
measured using the tracker information in the region of |η| < 0.75, Prompt D0 mesons are reconstructed from the decay D0 →
π+ + K−and D
0 →π−+ K+ with a branching fraction of (3.94 ±
0.04)%, using selected tracks with pT > 1.0 GeV/c and within the
acceptance of |η| < 2.4. Candidates are formed by combining pairs 2 The CMS Collaboration Physics Letters B 816 (2021) 136253 allowing to minimize the correlations among the three regions,
with a gap of more than two units of rapidity. The Q D0
n
vector
is defined for each D0 meson candidate. The averages ⟨Q nA Q ∗
nB⟩,
⟨Q nA Q ∗
nC⟩, and ⟨Q nB Q ∗
nC⟩are made considering all selected events,
while the average ⟨Q D0
n Q ∗
nA⟩is made considering all D0 meson
candidates in all selected events. To avoid autocorrelations, the
terms ⟨Q D0
n Q ∗
nA⟩and ⟨Q nA Q ∗
nB⟩use A = HF−(HF+) when the D0
meson candidate is at positive (negative) pseudorapidity. The nonprompt D0 meson contamination is taken into account as a
systematic uncertainty, by checking that the nonprompt D0 meson
fraction is always smaller than 12% (i.e., comparable to the un-
certainties in the reconstructed D0 meson yield). This implies that
the central values of vn will not be considerably affected by this
component, being compatible within statistical uncertainties. Such
a low fraction arises from the use of prompt D0 meson signals in
the BDT training, together with variables that are highly correlated
with the distance of closest approach (DCA) to the primary ver-
tex, which is defined as the flight distance of the D0 particle times
the sine of the pointing angle in three dimensions. Additional DCA
selection and dedicated training, involving prompt and nonprompt
D0 meson signals, do not bring considerable improvements in per-
formance. The prompt and nonprompt D0 meson fractions are ob-
tained using the DCA variable. 3. Reconstruction and selection of D0 mesons For prompt D0 mesons, the nonzero
DCA corresponds to the detector resolution, and is expected to be
concentrated around zero. For nonprompt D0 mesons, larger val-
ues of DCA result from the B meson decay. To extract the prompt
and nonprompt D0 meson fractions, a fit to the DCA distributions
is performed in data considering DCA shapes from MC simulations
for prompt and nonprompt D0 meson components. The nonprompt
D0 meson vn is estimated by considering two regions in the DCA:
one with very low fraction (2.7–8.0%) of nonprompt D0 particles
(DCA < 0.012 cm), and one with a high fraction (62.0–88.0%) of
nonprompt D0 particles (DCA > 0.012 cm). Using this information
together with Eq. (6), it is possible to estimate vnonprompt
n
by solv-
ing a system of two equations from the two DCA regions. In the
current analysis this procedure can only be done in wide pT, cen-
trality, and rapidity bins, because of the limited amount of data
available in the region with DCA > 0.012 cm. One goal of this analysis is to measure the difference (vn)
between D0 and D
0meson flow coefficients, vn, as a function of
rapidity, to probe effects from EM fields. The difference vn is
measured as: vn{SP} ≡⟨Q D0
n Q ∗
nA⟩−⟨Q D
0
n Q ∗
nA⟩
⟨Q nA Q ∗
nB⟩⟨Q nA Q ∗
nC⟩
⟨Q nB Q ∗
nC⟩
. (2) (2) The vn and vn of D0 meson candidates are first measured as
a function of their minv. The extraction of the D0 mesons signal
vn (vn), vsig
n
(vsig
n ), is performed via a simultaneous binned
χ 2 fit of the minv distribution and of vn (vn). The minv distri-
bution is fit with three components: a third-order polynomial to
model the combinatorial background, B(minv); two Gaussians with
the same mean but different widths to describe the minv in dif-
ferent kinematic regions for the D0 mesons signal, S(minv); and
one additional Gaussian distribution for the swap component cor-
responding to the incorrect mass assignment for the assumed pion
and kaon particles, SW (minv). The width of SW (minv) and the ra-
tio between the yields of SW (minv) and S(minv) are fixed by the
values extracted from MC simulations. In this case, the following
expression can be used for extracting vsig
n : vsig
n = fpromptvprompt
n
+ (1 −fprompt)vnonprompt
n
. Table 1 Table 1
Summary of systematic uncertainties in absolute values for v2, v3, and v2. Ranges of the variation of uncertainties for all the bins are presented. The cells filled with “—”
refer to the cases where the uncertainty cancels out. Systematic sources
pT bins
y bins
Centrality bins
v2
BDT selection
0.002–0.014
0.0065
0.005
Bkg. mass PDF
0.0002–0.0017
0.0007–0.0015
0.0007–0.0011
Bkg. vn PDF
0.01–0.05
0.004–0.007
0.003–0.005
D0 efficiency correction
—
0.004–0.007
0.0040–0.0045
Nonprompt D0 meson contamination
0.0002–0.0077
0.004
0.002–0.005
v3
BDT selection
0.002–0.023
0.001–0.009
0.002–0.006
Bkg. mass PDF
0.0001–0.0040
0.0005–0.0008
0.0012–0.0040
Bkg. vn PDF
0.01–0.05
0.003–0.004
0.0011
D0 efficiency correction
—
0.002–0.004
0.003–0.005
Nonprompt D0 meson contamination
0.0001–0.0090
0.0010–0.0015
0.0001–0.0008
v2
BDT selection
0.001–0.009
Bkg. mass PDF
0.00015–0.00030
D0 efficiency correction
0.001–0.004
Nonprompt D0 meson contamination
0.00002–0.00010 Table 1
Summary of systematic uncertainties in absolute values for v2, v3, and v2. Ranges of the variation of uncertainties for all the bins are presented. The cells filled with “—”
refer to the cases where the uncertainty cancels out. lute values for v2, v3, and v2. Ranges of the variation of uncertainties for all the bins are presented. The cells filled with “—”
cels out certainties in absolute values for v2, v3, and v2. Ranges of the variation of uncertainties for all the bins are presented. The cells fi
e uncertainty cancels out matic uncertainties in absolute values for v2, v3, and v2. Ranges of the variation of uncertainties for all the bins are presented. Th
where the uncertainty cancels out. BDT selection
Bkg. mass PDF
D0 efficiency correction
Nonprompt D0 meson contamination Calculations from theoretical models at midrapidity (|y| < 1)
are also presented. These models use different assumptions of the
QGP properties, for example in the thermal evolution of the colli-
sion system and in the initial-state conditions before the formation
of the QGP. In addition, different mechanisms are considered re-
garding the interaction of heavy quarks with the medium and
for the hadronization process. Results from the models LBT [36],
CUJET 3.0 [37], and SUBATECH [38] include collisional and ra-
diative energy losses, while those from the models TAMU [39],
PHSD [15], and TAMU SMCs [40] include only collisional energy
loss. Initial-state fluctuations are included in the calculations by
LBT, SUBATECH, and PHSD, and calculations for the v3 coefficient
are only available from these three models. Table 1 Coalescence mecha-
nisms are also included in LBT, SUBATECH, TAMU, PHSD, and TAMU
SMCs. While most models seem to capture the qualitative trend
of the data (except for the v2 description provided by TAMU in
the 10–50% centrality range), most of the models do not provide a
quantitative description over the full range, except for TAMU SMCs. The TAMU SMCs version improves the TAMU model by implement-
ing event-by-event space-momentum correlations (SMCs) between
charm quarks and the high-flow partons in the QGP medium [40]. Since it does not include initial-state fluctuations, TAMU SMCs
does not provide v2 calculations for centrality values between
0–10%. This puts more stringent constraints on the development
of the collective flow for charm quarks in the QGP medium, giving meson vn in wide bins of pT, rapidity, and centrality. A relative
systematic uncertainty is obtained by comparing vn from mixed
prompt and nonprompt D0 mesons to the vn derived from non-
prompt D0 mesons. Table 1 summarizes the estimates of systematic uncertainties in
absolute values for v2, v3, and v2. The ranges of variation of the
uncertainties are presented for each binning. 5. Systematic uncertainties The D0 meson selection efficiency times accep-
tance varies from 0.5 to 12.5% in the pT range of 1.0–8.0 GeV/c,
reaching a plateau of approximately 17.0% for pT > 15.0 GeV/c. (5) The term vbkg
n (minv) from Eq. (3) is modeled with a linear function,
while the constant parameter const in Eq. (5) is added to account
for possible fluctuations in the background vn component. The rel-
evance of this const parameter was investigated by redoing vn
measurements in MC simulation (without azimuthal correlations
or effects from EM fields), indicating that this parameter improves
the fit quality and does not introduce artificial signals. A cross-
check is performed by redoing the measurements using a linear
function instead of a constant. No significant changes in the cen-
tral values of v2 and on their uncertainties are observed. Fig. 1
shows an example of a simultaneous fit for v2 and v2. After performing the fits for extracting the signal vn, there is
still a sizable fraction of nonprompt D0 mesons embedded in vsig
n . The extracted vn can be written as The systematic uncertainties regarding contamination from
nonprompt D0 mesons are estimated by measuring nonprompt D0 (6) 3 Physics Letters B 816 (2021) 136253 The CMS Collaboration Fig. 1. Simultaneous fit of the πK invariant mass (left) and v2 (v2) as function of invariant mass (right) for 3.0 < pT < 3.5 GeV/c, centrality 20–70%, and −0.6 < y < 0.0. Fig. 1. Simultaneous fit of the πK invariant mass (left) and v2 (v2) as function of invariant mass (right) for 3.0 < pT < 3.5 GeV/c, Fig. 1. Simultaneous fit of the πK invariant mass (left) and v2 (v2) as function of invariant mass (right) for 3.0 < pT < 3.5 GeV/c, centrality 20–70%, and −0.6 < y < 0.0. 6. Results Results for prompt D0 meson v2 and v3 anisotropic flow co-
efficients, obtained with 2018 PbPb data, as functions of pT and
for |y| < 1, are shown in Fig. 2 for three centrality ranges: 0–10%,
10–30%, and 30–50%. The results extend previously published data
from CMS [25], by extending the high-pT coverage to ∼60.0 GeV/c
and by providing finer pT bins. These high-precision data are com-
patible with previous measurements from Ref. [25], and a clear
trend of rise and fall from low to high pT is observed for both
v2 and v3 across the full centrality range. This behavior is simi-
lar to that observed for inclusive charged particles [35] for |η| <
1.0, also shown in Fig. 2. For noncentral collisions (i.e., central-
ity 10–50%), values of prompt D0 meson v2 are positive up to
pT ∼30.0–40.0 GeV/c, whereas the v3 values become consistent
with zero at pT ∼10.0 GeV/c. 4 Physics Letters B 816 (2021) 136253 The CMS Collaboration Fig. 2. Prompt D0 meson and charged particle flow coefficients v2 (upper) and v3 (lower) at midrapidity (|y| < 1.0 for prompt D0 mesons and |η| < 1.0 for charged
particles) for the centrality classes 0–10% (left), 10–30% (middle), and 30–50% (right). The vertical bars and open boxes represent the statistical and systematic uncertainties,
respectively. The horizontal bars represent the width of each pT bin. Theoretical calculations for vn coefficients of prompt D0 mesons are also plotted for comparison:
LBT [36], CUJET 3.0 [37], SUBATECH [38], TAMU [39], PHSD [15]. The TAMU SMCs model [40] is available only in the 10–50% centrality bins. Fig. 2. Prompt D0 meson and charged particle flow coefficients v2 (upper) and v3 (lower) at midrapidity (|y| < 1.0 for prompt D0 mesons and |η| < 1.0 for charged
particles) for the centrality classes 0–10% (left), 10–30% (middle), and 30–50% (right). The vertical bars and open boxes represent the statistical and systematic uncertainties,
respectively. The horizontal bars represent the width of each pT bin. Theoretical calculations for vn coefficients of prompt D0 mesons are also plotted for comparison:
LBT [36], CUJET 3.0 [37], SUBATECH [38], TAMU [39], PHSD [15]. The TAMU SMCs model [40] is available only in the 10–50% centrality bins. Fig. 3. 6. Results 4), and can be understood in terms of colli-
sion geometry and viscosity effects. In particular, a faster increase
of v2 is observed from central to peripheral collisions for charged
particles compared to prompt D0 mesons. This feature was also ob-
served when comparing v2 of low-pT J/ψ with charged pions [42],
where it is claimed that this could be understood in terms of two
phenomena: one, associated with transport models predicting an
increasing fraction of regenerated J/ψ at low-pT, when going from
peripheral to central collisions; the other, not related to regener-
ation, is associated with a possible partial or later thermalization
of charm quarks compared to light quarks [42]. The v3 shows no
centrality dependence, which is also consistent with expectations
from collision geometry fluctuations [43]. Fig. 4 (right) presents results for the rapidity dependence of
prompt D0 meson v2 and v3, for centrality 20–70%, averaged over
2.0 < pT < 8.0 GeV/c. A weak rapidity dependence of v2 and v3 is
observed in the data. Fig. 5. Prompt D0 meson v2 as a function of rapidity, for 2.0 < pT < 8.0 GeV/c
and centrality 20–70%. The vertical bars represent statistical uncertainties and open
boxes represent systematic uncertainties. The horizontal bars represent the width
of each bin. Finally, to search for effects of strong EM fields, the difference
v2 between the v2 values of D0 and D
0mesons is measured. These results are presented in Fig. 5, as a function of rapidity, av-
eraged over 2.0 < pT < 8.0 GeV/c and for centrality 20–70%. For all
rapidity bins, the v2 values are compatible with zero. The aver-
age over the full rapidity region is ⟨v2⟩= 0.001 ± 0.001 (stat) ±
0.003 (syst). In Ref. [19], the predicted v2 splitting for inclusive
charged particles due to electric fields is ∼0.001 at the LHC ener-
gies. While quantitative predictions for v2 splitting of D0 mesons
are not yet available, they are expected to be much larger than
those for inclusive charged particles. In the case of v1, the ALICE
collaboration reported results about three orders of magnitude
larger than measurements for charged hadrons [44], although the
uncertainties prevent a clear conclusion. 6. Results The main reason is that
heavy-flavor quarks are usually produced much earlier than light-
flavor quarks, the former being predominantly produced soon after
the collision takes place, when the EM field strength is several or- ders of magnitude stronger [20]. The results presented here pose
constraints on possible EM effects on charm quarks. 6. Results Prompt D0
meson flow coefficients v2 (upper) and v3 (lower) at midrapidity (|y| < 1, red open circles) and forward rapidity (1 < |y| < 2, blue open diamonds) for
the centrality classes 0–10% (left), 10–30% (middle), and 30–50% (right). The vertical bars and open boxes represent the statistical and systematic uncertainties, respectively. The horizontal bars represent the width of each pT bin. Fig. 3. Prompt D0
meson flow coefficients v2 (upper) and v3 (lower) at midrapidity (|y| < 1, red open circles) and forward rapidity (1 < |y| < 2, blue open diamonds) for
the centrality classes 0–10% (left), 10–30% (middle), and 30–50% (right). The vertical bars and open boxes represent the statistical and systematic uncertainties, respectively. The horizontal bars represent the width of each pT bin. further inputs for understanding heavy-quark interactions with the
medium (for example, energy loss and coalescence mechanisms). observation is similar to that for inclusive charged-hadron mea-
surements [41]. In Fig. 4 (left), results for prompt D0 mesons v2 and v3, av-
eraged over 2.0 < pT < 8.0 GeV/c, for |y| < 1 and 1 < |y| < 2, are
presented as a function of collision centrality. This pT range is cho-
sen in order to cover the widest possible pT range, while maximiz-
ing the D0 meson signal yield significance. These pT- and rapidity- Results for the rapidity dependence of heavy-flavor collective
flow are presented for the first time for prompt D0 meson v2 and
v3 as functions of pT, both at midrapidity (|y| < 1) and in the
forward (1 < |y| < 2) region, as shown in Fig. 3. No clear rapidity
dependence is observed for both v2 and v3 as functions of pT. This 5 Physics Letters B 816 (2021) 136253 The CMS Collaboration The CMS Collaboration
Physics Letters B 816 (2021) 136253
Fig. 4. Prompt D0 meson v2 and v3 as functions of centrality, for 2.0 < pT < 8.0 GeV/c and for rapidity ranges |y| < 1 and 1 < |y| < 2. The results are compared with charged
particle v2 and v3 in the same pT range and with |η| < 1 (left). Prompt D0 v2 and v3 as functions of rapidity, for 2.0 < pT < 8.0 GeV/c and for centrality 20–70% (right). The vertical bars represent statistical uncertainties and open boxes represent systematic uncertainties. The horizontal bars represent the width of each bin. Fig. 6. Results 4. Prompt D0 meson v2 and v3 as functions of centrality, for 2.0 < pT < 8.0 GeV/c and for rapidity ranges |y| < 1 and 1 < |y| < 2. The results are compared with charged
particle v2 and v3 in the same pT range and with |η| < 1 (left). Prompt D0 v2 and v3 as functions of rapidity, for 2.0 < pT < 8.0 GeV/c and for centrality 20–70% (right). The vertical bars represent statistical uncertainties and open boxes represent systematic uncertainties. The horizontal bars represent the width of each bin. Fig. 5. Prompt D0 meson v2 as a function of rapidity, for 2.0 < pT < 8.0 GeV/c
and centrality 20–70%. The vertical bars represent statistical uncertainties and open
boxes represent systematic uncertainties. The horizontal bars represent the width
of each bin. integrated results include an additional centrality bin (50–70%),
which has an insufficient number of events for the full differential
analysis. For both mid- and forward-rapidity regions, the v2 results
show a clear increase from the most central to mid-central events,
and then a declining trend toward the most peripheral events. This
trend is similar to that observed for inclusive charged particles
(also shown in Fig. 4), and can be understood in terms of colli-
sion geometry and viscosity effects. In particular, a faster increase
of v2 is observed from central to peripheral collisions for charged
particles compared to prompt D0 mesons. This feature was also ob-
served when comparing v2 of low-pT J/ψ with charged pions [42],
where it is claimed that this could be understood in terms of two
phenomena: one, associated with transport models predicting an
increasing fraction of regenerated J/ψ at low-pT, when going from
peripheral to central collisions; the other, not related to regener-
ation, is associated with a possible partial or later thermalization
of charm quarks compared to light quarks [42]. The v3 shows no
centrality dependence, which is also consistent with expectations
from collision geometry fluctuations [43]. integrated results include an additional centrality bin (50–70%),
which has an insufficient number of events for the full differential
analysis. For both mid- and forward-rapidity regions, the v2 results
show a clear increase from the most central to mid-central events,
and then a declining trend toward the most peripheral events. This
trend is similar to that observed for inclusive charged particles
(also shown in Fig. References [1] I. Arsene, et al., BRAHMS, Quark-gluon plasma and color glass condensate at
RHIC? The perspective from the BRAHMS experiment, Nucl. Phys. A 757 (2005)
1, https://doi .org /10 .1016 /j .nuclphysa .2005 .02 .130, arXiv:nucl -ex /0410020. [2] B.B. Back, et al., PHOBOS, The PHOBOS perspective on discoveries at RHIC, Nucl. Phys. A 757 (2005) 28, https://doi .org /10 .1016 /j .nuclphysa .2005 .03 .084, arXiv:
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PhysRevLett .105 .252302, arXiv:1011.3914. [8] ATLAS Collaboration, Measurement of the pseudorapidity and transverse mo-
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produced in PbPb collisions at √
sNN = 2.76 TeV, Phys. Rev. C 87 (2013) 014902,
https://doi .org /10 .1103 /PhysRevC .87.014902, arXiv:1204 .1409. Individuals have received support from the Marie-Curie pro-
gram and the European Research Council and Horizon 2020 Grant,
contract Nos. 675440, 752730, and 765710 (European Union);
the Leventis Foundation; the A.P. 7. Summary Measurements of the elliptic (v2) and triangular (v3) flow coef-
ficients of prompt D0 mesons are presented as functions of trans-
verse momentum (pT), rapidity, and collision centrality, in PbPb
collisions at √
sNN = 5.02 TeV. The results improve previously pub-
lished CMS data by extending the pT and rapidity coverage and by
providing more differential information in pT, rapidity, and cen-
trality. A clear centrality dependence of prompt D0 meson v2 is
observed, while v3 is largely centrality independent. These trends
are consistent with the expectation that v2 and v3 are driven by
initial-state geometry. A weak rapidity dependence of prompt D0 6 The CMS Collaboration Physics Letters B 816 (2021) 136253 FIA research grants 123842, 123959, 124845, 124850, 125105,
128713, 128786, and 129058 (Hungary); the Council of Science
and Industrial Research, India; the HOMING PLUS program of the
Foundation for Polish Science, cofinanced from European Union,
Regional Development Fund, the Mobility Plus program of the
Ministry of Science and Higher Education, the National Science
Center (Poland), contracts Harmonia 2014/14/M/ST2/00428, Opus
2014/13/B/ST2/02543, 2014/15/B/ST2/03998, and 2015/19/B/ST2/
02861, Sonata-bis 2012/07/E/ST2/01406; the National Priorities Re-
search Program by Qatar National Research Fund; the Ministry of
Science and Higher Education, project no. 02.a03.21.0005 (Russia);
the Tomsk Polytechnic University Competitiveness Enhancement
Program; the Programa Estatal de Fomento de la Investigación
Científica y Técnica de Excelencia María de Maeztu, grant MDM-
2015-0509 and the Programa Severo Ochoa del Principado de
Asturias; the Thalis and Aristeia programs cofinanced by EU-ESF
and the Greek NSRF; the Rachadapisek Sompot Fund for Postdoc-
toral Fellowship, Chulalongkorn University and the Chulalongkorn
Academic into Its 2nd Century Project Advancement Project (Thai-
land); the Kavli Foundation; the Nvidia Corporation; the SuperMi-
cro Corporation; the Welch Foundation, contract C-1845; and the
Weston Havens Foundation (USA). meson v2 and v3 is observed. When comparing various theoret-
ical calculations to the data at midrapidity, no model is able to
describe the data over the full centrality and pT ranges. Motivated by the search for evidence of the strong electric field
expected in PbPb collisions, a first measurement of the v2 flow
coefficient difference (v2) between D0 and D
0mesons as a func-
tion of rapidity is presented. The rapidity-averaged v2 difference is
measured to be ⟨v2⟩ = 0.001 ±0.001 (stat)±0.003 (syst). Declaration of competing interest The authors declare that they have no known competing finan-
cial interests or personal relationships that could have appeared to
influence the work reported in this paper. 7. Summary This in-
dicates that there is no evidence that charm hadron collective flow
is affected by the strong Coulomb field created in ultrarelativistic
heavy ion collisions. Future comparisons of theoretical models with
these results may provide constraints on the electric conductivity
of the quark-gluon plasma. Acknowledgements We congratulate our colleagues in the CERN accelerator depart-
ments for the excellent performance of the LHC and thank the
technical and administrative staffs at CERN and at other CMS in-
stitutes for their contributions to the success of the CMS effort. In addition, we gratefully acknowledge the computing centers and
personnel of the Worldwide LHC Computing Grid for delivering so
effectively the computing infrastructure essential to our analyses. Finally, we acknowledge the enduring support for the construc-
tion and operation of the LHC and the CMS detector provided by
the following funding agencies: BMBWF and FWF (Austria); FNRS
and FWO (Belgium); CNPq, CAPES, FAPERJ, FAPERGS, and FAPESP
(Brazil); MES (Bulgaria); CERN; CAS, MOST, and NSFC (China);
COLCIENCIAS (Colombia); MSES and CSF (Croatia); RIF (Cyprus);
SENESCYT (Ecuador); MoER, ERC IUT, PUT and ERDF (Estonia);
Academy of Finland, MEC, and HIP (Finland); CEA and CNRS/IN2P3
(France); BMBF, DFG, and HGF (Germany); GSRT (Greece); NK-
FIA (Hungary); DAE and DST (India); IPM (Iran); SFI (Ireland);
INFN (Italy); MSIP and NRF (Republic of Korea); MES (Latvia); LAS
(Lithuania); MOE and UM (Malaysia); BUAP, CINVESTAV, CONACYT,
LNS, SEP, and UASLP-FAI (Mexico); MOS (Montenegro); MBIE (New
Zealand); PAEC (Pakistan); MSHE and NSC (Poland); FCT (Portu-
gal); JINR (Dubna); MON, ROSATOM, RAS, RFBR, and NRC KI (Rus-
sia); MESTD (Serbia); SEIDI, CPAN, PCTI, and FEDER (Spain); MoSTR
(Sri Lanka); Swiss Funding Agencies (Switzerland); MST (Taipei);
ThEPCenter, IPST, STAR, and NSTDA (Thailand); TÜBITAK and TAEK
(Turkey); NASU (Ukraine); STFC (United Kingdom); DOE and NSF
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Institute for Nuclear Problems, Minsk, Belarus V. Chekhovsky, A. Litomin, V. Makarenko, J. Suarez Gonzalez
Institute for Nuclear Problems, Minsk, Belarus V. Chekhovsky, A. Litomin, V. Makarenko, J. Suarez Gonzalez
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Université Libre de Bruxelles, Bruxelles, Belgium T. Cornelis, D. Dobur, I. Khvastunov 3, M. Niedziela, C. Roskas, K. Skovpen, M. Tytgat, W. Verbeke,
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Ghent University, Ghent, Belgium G. Bruno, F.J.J. Bury, C. Caputo, P. David, C. Delaere, M. Delcourt, I.S. Donertas, A. Giammanco,
V. Lemaitre, J. Prisciandaro, A. Saggio, A. Taliercio, M. Teklishyn, P. Vischia, S. Wuyckens, J. Zobec
Université Catholique de Louvain, Louvain-la-Neuve, Belgium G.A. Alves, G. Correia Silva, C. Hensel, A. Moraes
Centro Brasileiro de Pesquisas Fisicas, Rio de Janeiro, Brazil W.L. Aldá Júnior, E. Belchior Batista Das Chagas, W. Carvalho, J. Chinellato 4, E. Coelho, E.M. Da Costa,
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E.J. Tonelli Manganote 4, F. Torres Da Silva De Araujo, A. Vilela Pereira
Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil C.A. Bernardes a, L. Calligaris a, T.R. Fernandez Perez Tomei a, E.M. Gregores b, D.S. Lemos a,
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Institute for Nuclear Problems, Minsk, Belarus Padula a
a Universidade Estadual Paulista, São Paulo, Brazil
b Universidade Federal do ABC, São Paulo, Brazil A. Aleksandrov, G. Antchev, I. Atanasov, R. Hadjiiska, P. Iaydjiev, M. Misheva, M. Rodozov, M. Shopova,
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Institute for Nuclear Research and Nuclear Energy, Bulgarian Academy of Sciences, Sofia, Bulgaria Institute for Nuclear Research and Nuclear Energy, Bulgarian Academy of Sciences, Sofia, Bulgaria Institute for Nuclear Research and Nuclear Energy, Bulgarian Academy of Sciences, Sofia, Bulgaria M. Bonchev, A. Dimitrov, T. Ivanov, L. Litov, B. Pavlov, P. Petkov, A. Petrov
University of Sofia, Sofia, Bulgaria W. Fang 2, X. Gao 2, Q. Guo, H. Wang, L. Yuan
Beihang University, Beijing, China M. Ahmad, Z. Hu, Y. Wang
Department of Physics, Tsinghua University, Beijing, China E. Chapon, G.M. Chen 8, H.S. Chen 8, M. Chen, C.H. Jiang, D. Leggat, H. Liao, Z. Liu, A. Spiezia, J. Tao,
J. Wang, E. Yazgan, H. Zhang, S. Zhang 8, J. Zhao
Institute of High Energy Physics, Beijing, China A. Agapitos, Y. Ban, C. Chen, G. Chen, A. Levin, J. Li, L. Li, Q. Li, X. Lyu, Y. Mao, S.J. Qian, D. Wang,
Q. Wang, J. Xiao State Key Laboratory of Nuclear Physics and Technology, Peking University, Beijing, China State Key Laboratory of Nuclear Physics and Technology, Peking University, Beijing, China Z. You Sun Yat-Sen University, Guangzhou, China 9 9 The CMS Collaboration Physics Letters B 816 (2021) 136253 M. Xiao Zhejiang University, Hangzhou, China J. Mejia Guisao, J.D. Ruiz Alvarez, C.A. Salazar González, N. Vanegas Arbelaez
Universidad de Antioquia, Medellin, Colombia Z. Antunovic, M. Kovac
University of Split, Faculty of Science, Split, Croatia E. Carrera Jarrin S. Bhowmik, A. Carvalho Antunes De Oliveira, R.K. Dewanjee, K. Ehataht, M. Kadastik, M. Raidal,
C. Veelken
National Institute of Chemical Physics and Biophysics, Tallinn, Estonia P. Eerola, L. Forthomme, H. Kirschenmann, K. Osterberg, M. Voutilainen
Department of Physics, University of Helsinki, Helsinki, Finland E. Brücken, F. Garcia, J. Havukainen, V. Karimäki, M.S. Kim, R. Kinnunen, T. Lampén, K. Lassila-Perini,
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Helsinki Institute of Physics, Helsinki, Finland M. Besancon, F. Couderc, M. Dejardin, D. Denegri, J.L. Faure, F. Ferri, S. Ganjour, A. Givernaud, P. Gras,
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IRFU, CEA, Université Paris-Saclay, Gif-sur-Yvette, France 10 Physics Letters B 816 (2021) 136253 The CMS Collaboration S. Ahuja, C. Amendola, F. Beaudette, M. Bonanomi, P. Busson, C. Charlot, O. Davignon, B. Diab,
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Laboratoire Leprince-Ringuet, CNRS/IN2P3, Ecole Polytechnique, Institut Polytechnique de Paris, Palaiseau, France J.-L. Agram 15, J. Andrea, D. Bloch, G. Bourgatte, J.-M. Brom, E.C. Chabert, C. Collard, J.-C. Fontaine 15,
D. Gelé, U. Goerlach, C. Grimault, A.-C. Le Bihan, P. Van Hove
Université de Strasbourg, CNRS, IPHC UMR 7178, Strasbourg, France E. Asilar, S. Beauceron, C. Bernet, G. Boudoul, C. Camen, A. Carle, N. Chanon, R. Chierici, D. Contardo,
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Université de Lyon, Université Claude Bernard Lyon 1, CNRS-IN2P3, Institut de Physique Nucléaire de Lyon, Villeurbanne, France R. Aggleton, S. Bein, L. Benato, A. Benecke, K. De Leo, T. Dreyer, A. Ebrahimi, F. Feindt, A. Fröhlich,
C. Garbers, E. Garutti, D. Gonzalez, P. Gunnellini, J. Haller, A. Hinzmann, A. Karavdina, G. Kasieczka,
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A. Nigamova, K.J. Pena Rodriguez, A. Reimers, O. Rieger, P. Schleper, S. Schumann, J. Schwandt,
D. Schwarz, J. Sonneveld, H. Stadie, G. Steinbrück, B. Vormwald, I. Zoi A. Khvedelidze 10 Georgian Technical University, Tbilisi, Georgia Georgian Technical University, Tbilisi, Georgia L. Feld, K. Klein, M. Lipinski, D. Meuser, A. Pauls, M. Preuten, M.P. Rauch, J. Schulz, M. Teroerde
RWTH Aachen University, I. Physikalisches Institut, Aachen, Germany D. Eliseev, M. Erdmann, P. Fackeldey, B. Fischer, S. Ghosh, T. Hebbeker, K. Hoepfner, H. Keller,
L. Mastrolorenzo, M. Merschmeyer, A. Meyer, P. Millet, G. Mocellin, S. Mondal, S. Mukherjee, D. Noll,
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RWTH Aachen University, III. Physikalisches Institut A, Aachen, Germany C. Dziwok, G. Flügge, W. Haj Ahmad 16, O. Hlushchenko, T. Kress, A. Nowack, C. Pistone, O. Pooth, D. Roy,
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RWTH Aachen University, III. Physikalisches Institut B, Aachen, Germany H. Aarup Petersen, M. Aldaya Martin, P. Asmuss, I. Babounikau, S. Baxter, K. Beernaert, O. Behnke,
A. Bermúdez Martínez, A.A. Bin Anuar, K. Borras 18, V. Botta, D. Brunner, A. Campbell, A. Cardini,
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Th. Müller, M. Musich, G. Quast, K. Rabbertz, J. Rauser, D. Savoiu, D. Schäfer, M. Schnepf, M. Schröder,
I. Shvetsov, H.J. Simonis, R. Ulrich, M. Wassmer, M. Weber, C. Wöhrmann, R. Wolf, S. Wozniewski
Karlsruher Institut fuer Technologie, Karlsruhe, Germany G. Anagnostou, P. Asenov, G. Daskalakis, T. Geralis, A. Kyriakis, D. Loukas, G. Paspalaki, A. Stakia
Institute of Nuclear and Particle Physics (INPP), NCSR Demokritos, Aghia Paraskevi, Greece M. Diamantopoulou, D. Karasavvas, G. Karathanasis, P. Kontaxakis, C.K. Koraka, A. Manousakis-katsikakis,
A. Panagiotou, I. Papavergou, N. Saoulidou, K. Theofilatos, K. Vellidis, E. Vourliotis
National and Kapodistrian University of Athens, Athens, Greece G. Bakas, K. Kousouris, I. Papakrivopoulos, G. Tsipolitis, A. Zacharopoulou
National Technical University of Athens, Athens, Greece G. Bakas, K. Kousouris, I. Papakrivopoulos, G. Tsipolitis, A. Zacharopoulou
National Technical University of Athens, Athens, Greece I. Evangelou, C. Foudas, P. Gianneios, P. Katsoulis, P. Kokkas, S. Mallios, K. Manitara, N. Manthos,
I. Papadopoulos, J. Strologas, F.A. Triantis, D. Tsitsonis
University of Ioánnina, Ioánnina, Greece M. Bartók 23, R. Chudasama, M. Csanad, M.M.A. Gadallah 24, P. Major, K. Mandal, A. Mehta, G. Pasztor,
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MTA-ELTE Lendület CMS Particle and Nuclear Physics Group, Eötvös Loránd University, Budapest, Hungary G. Bencze, C. Hajdu, D. Horvath 25, F. Sikler, V. Veszpremi, G. Vesztergombi †
Wigner Research Centre for Physics, Budapest, Hungary N. Beni, S. Czellar, J. Karancsi 23, J. Molnar, Z. Szillasi, D. Teyssier
Institute of Nuclear Research ATOMKI, Debrecen, Hungary P. Raics, Z.L. Trocsanyi, B. Ujvari
Institute of Physics, University of Debrecen, Debrecen, Hungary T. Csorgo, S. Lökös 26, F. Nemes, T. Novak
Eszterhazy Karoly University, Karoly Robert Campus, Gyongyos, Hungary S. Choudhury, J.R. Komaragiri, D. Kumar, L. Panwar, P.C. Tiwari
Indian Institute of Science (IISc), Bangalore, India S. Bahinipati 27, C. Kar, P. Mal, T. Mishra, V.K. Muraleedharan Nair Bindhu, A. Nayak 28, D.K. Sahoo 27,
N. Sur, S.K. H. Bakhshiansohi 34 Department of Physics, Isfahan University of Technology, Isfahan, Iran S. Chenarani 35, S.M. Etesami, M. Khakzad, M. Mohammadi Najafabadi, M. Naseri
Institute for Research in Fundamental Sciences (IPM), Tehran, Iran M. Abbrescia a,b, R. Aly a,b,36, C. Calabria a,b, A. Colaleo a, D. Creanza a,c, N. De Filippis a,c, M. De Palma a,b,
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P. Verwilligen a
a INFN Sezione di Bari, Bari, Italy
b Università di Bari, Bari, Italy
c Politecnico di Bari, Bari, Italy P. Verwilligen a
a INFN Sezione di Bari, Bari, Italy
b Università di Bari, Bari, Italy
c Politecnico di Bari, Bari, Italy
G. Abbiendi a, C. Battilana a,b, D. Bonacorsi a,b, L. Borgonovi a,b, R. Campanini a,b, P. Capiluppi a,b,
A. Castro a,b, F.R. Cavallo a, C. Ciocca a, M. Cuffiani a,b, G.M. Dallavalle a, T. Diotalevi a,b, F. Fabbri a,
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G.P. Siroli a,b, N. Tosi a
a INFN Sezione di Bologna, Bologna, Italy
b Università di Bologna, Bologna, Italy
S. Albergo a,b,38, S. Costa a,b, A. Di Mattia a, R. Potenza a,b, A. Tricomi a,b,38, C. Tuve a,b
a INFN Sezione di Catania, Catania, Italy
b Università di Catania, Catania, Italy
G. Barbagli a, A. Cassese a, R. Ceccarelli a,b, V. Ciulli a,b, C. Civinini a, R. D’Alessandro a,b, F. Fiori a,
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a INFN Sezione di Firenze, Firenze, Italy G. Abbiendi a, C. Battilana a,b, D. Bonacorsi a,b, L. Borgonovi a,b, R. Campanini a,b, P. Capiluppi a,b,
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National Institute of Science Education and Research, HBNI, Bhubaneswar, India S. Bansal, S.B. Beri, V. Bhatnagar, S. Chauhan, N. Dhingra 29, R. Gupta, A. Kaur, A. Kaur, S. Kaur, P. Kumari,
M. Lohan, M. Meena, K. Sandeep, S. Sharma, J.B. Singh, A.K. Virdi
Panjab University, Chandigarh, India A. Ahmed, A. Bhardwaj, B.C. Choudhary, R.B. Garg, M. Gola, S. Keshri, A. Kumar, M. Naimuddin,
P. Priyanka, K. Ranjan, A. Shah, R. Sharma
University of Delhi, Delhi, India 12 Physics Letters B 816 (2021) 136253 The CMS Collaboration M. Bharti 30, R. Bhattacharya, S. Bhattacharya, D. Bhowmik, S. Dutta, S. Ghosh, B. Gomber 31, M. Maity 32,
K. Mondal, S. Nandan, P. Palit, A. Purohit, P.K. Rout, G. Saha, S. Sarkar, M. Sharan, B. Singh 30, S. Thakur 30
Saha Institute of Nuclear Physics, HBNI, Kolkata, India P.K. Behera, S.C. Behera, P. Kalbhor, A. Muhammad, R. Pradhan, P.R. Pujahari, A. Sharma, A.K. Sikdar
Indian Institute of Technology Madras, Madras, India D. Dutta, V. Jha, D.K. Mishra, K. Naskar 33, P.K. Netrakanti, L.M. Pant, P. Shukla
Bhabha Atomic Research Centre, Mumbai, India T. Aziz, M.A. Bhat, S. Dugad, R. Kumar Verma, U. Sarkar
Tata Institute of Fundamental Research-A, Mumbai, India S. Banerjee, S. Bhattacharya, S. Chatterjee, P. Das, M. Guchait, S. Karmakar, S. Kumar, G. Majumder,
K. Mazumdar, S. Mukherjee, D. Roy, N. Sahoo
Tata Institute of Fundamental Research-B, Mumbai, India S. Dube, B. Kansal, A. Kapoor, K. Kothekar, S. Pandey, A. Rane, A. Rastogi, S. Sharma
Indian Institute of Science Education and Research (IISER), Pune, India L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy M. Bozzo a,b, F. Ferro a, R. Mulargia a,b, E. Robutti a, S. Tosi a,b M. Bozzo a,b, F. Ferro a, R. Mulargia a,b, E. Robutti a, S. Tosi a,b M. Bozzo a,b, F. Ferro a, R. Mulargia a,b, E. Robutti a, S. Tosi a,b
a INFN Sezione di Genova, Genova, Italy
b Università di Genova, Genova, Italy a INFN Sezione di Genova, Genova, Italy
b Università di Genova, Genova, Italy A. Benaglia a, A. Beschi a,b, F. Brivio a,b, F. Cetorelli a,b, V. Ciriolo a,b,17, F. De Guio a,b, M.E. Dinardo a,b,
P. Dini a, S. Gennai a, A. Ghezzi a,b, P. Govoni a,b, L. Guzzi a,b, M. Malberti a, S. Malvezzi a, D. Menasce a,
F. Monti a,b, L. Moroni a, M. Paganoni a,b, D. Pedrini a, S. Ragazzi a,b, T. Tabarelli de Fatis a,b,
D. Valsecchi a,b,17, D. Zuolo a,b
a INFN Sezione di Milano-Bicocca, Milano, Italy
b Università di Milano-Bicocca, Milano, Italy S. Buontempo a, N. Cavallo a,c, A. De Iorio a,b, F. Fabozzi a,c, F. Fienga a, G. Galati a, A.O.M. Iorio a,b,
L. Layer a,b, L. Lista a,b, S. Meola a,d,17, P. Paolucci a,17, B. Rossi a, C. Sciacca a,b, E. Voevodina a,b
a INFN Sezione di Napoli, Napoli, Italy
b Università di Napoli ‘Federico II’, Napoli, Italy
c Università della Basilicata, Potenza, Italy
d Università G. Marconi, Roma, Italy c Università della Basilicata, Potenza, Italy d Università G. Marconi, Roma, Italy P. Azzi a, N. Bacchetta a, D. Bisello a,b, A. Boletti a,b, A. Bragagnolo a,b, R. Carlin a,b, P. Checchia a,
P. De Castro Manzano a, T. Dorigo a, U. Dosselli a, F. Gasparini a,b, U. Gasparini a,b, S.Y. Hoh a,b,
M. Margoni a,b, A.T. Meneguzzo a,b, M. Presilla b, P. Ronchese a,b, R. Rossin a,b, F. Simonetto a,b, G. Strong,
A. Tiko a, M. Tosi a,b, M. Zanetti a,b, P. Zotto a,b, A. Zucchetta a,b, G. Zumerle a,b
a INFN Sezione di Padova, Padova, Italy
b Università di Padova, Padova, Italy
c Università di Trento, Trento, Italy A. Braghieri a, S. Calzaferri a,b, D. Fiorina a,b, P. Montagna a,b, S.P. Ratti a,b, V. Re a, M. Ressegotti a,b,
C. Riccardi a,b, P. Salvini a, I. Vai a, P. H. Bakhshiansohi 34 Giacomelli a, L. Giommi a,b, C. Grandi a, L. Guiducci a,b, F. Iemmi a,b,
S. Lo Meo a,37, S. Marcellini a, G. Masetti a, F.L. Navarria a,b, A. Perrotta a, F. Primavera a,b, T. Rovelli a,b,
G.P. Siroli a,b, N. Tosi a
a INFN Sezione di Bologna, Bologna, Italy
b U i
ità di B l
B l
It l S. Albergo a,b,38, S. Costa a,b, A. Di Mattia a, R. Potenza a,b, A. Tricomi a,b,38, C. Tuve a,b
a INFN Sezione di Catania, Catania, Italy
b Università di Catania, Catania, Italy G. Barbagli a, A. Cassese a, R. Ceccarelli a,b, V. Ciulli a,b, C. Civinini a, R. D’Alessandro a,b, F. Fiori a,
E. Focardi a,b, G. Latino a,b, P. Lenzi a,b, M. Lizzo a,b, M. Meschini a, S. Paoletti a, R. Seidita a,b,
G. Sguazzoni a, L. Viliani a
a INFN Sezione di Firenze, Firenze, Italy 13 Physics Letters B 816 (2021) 136253 The CMS Collaboration b Università di Firenze, Firenze, Italy b Università di Firenze, Firenze, Italy b Università di Firenze, Firenze, Italy L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy Vitulo a,b
a INFN Sezione di Pavia, Pavia, Italy
b Università di Pavia, Pavia, Italy A. Braghieri a, S. Calzaferri a,b, D. Fiorina a,b, P. Montagna a,b, S.P. Ratti a,b, V. Re a, M. Ressegotti a,b,
C. Riccardi a,b, P. Salvini a, I. Vai a, P. Vitulo a,b
a INFN S
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di P
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It l M. Biasini a,b, G.M. Bilei a, D. Ciangottini a,b, L. Fanò a,b, P. Lariccia a,b, G. Mantovani a,b, V. Mariani a
M. Menichelli a, A. Rossi a,b, A. Santocchia a,b, D. Spiga a, T. Tedeschi a,b
a INFN Sezione di Perugia, Perugia, Italy
b Università di Perugia, Perugia, Italy
K. Androsov a, P. Azzurri a, G. Bagliesi a, V. Bertacchi a,c, L. Bianchini a, T. Boccali a, R. Castaldi a,
M.A. Ciocci a,b, R. Dell’Orso a, M.R. Di Domenico a,b, S. Donato a, L. Giannini a,c, A. Giassi a, M.T. Gri
F. Ligabue a,c, E. Manca a,c, G. Mandorli a,c, A. Messineo a,b, F. Palla a, A. Rizzi a,b, G. Rolandi a,c,
S. Roy Chowdhury a,c, A. Scribano a, N. Shafiei a,b, P. Spagnolo a, R. Tenchini a, G. Tonelli a,b, N. Turi
A. Venturi a, P.G. Verdini a
a INFN Sezione di Pisa, Pisa, Italy
b Università di Pisa, Pisa, Italy
c Scuola Normale Superiore di Pisa, Pisa, Italy
F. Cavallari a, M. Cipriani a,b, D. Del Re a,b, E. Di Marco a, M. Diemoz a, E. Longo a,b, P. Meridiani a,
G. Organtini a,b, F. Pandolfia, R. Paramatti a,b, C. Quaranta a,b, S. Rahatlou a,b, C. Rovelli a,
F. Santanastasio a,b, L. Soffia,b, R. Tramontano a,b
a INFN Sezione di Roma, Rome, Italy
b Sapienza Università di Roma, Rome, Italy M. Biasini a,b, G.M. Bilei a, D. Ciangottini a,b, L. Fanò a,b, P. Lariccia a,b, G. Mantovani a,b, V. Mariani a,b,
M. Menichelli a, A. Rossi a,b, A. Santocchia a,b, D. Spiga a, T. Tedeschi a,b
a INFN Sezione di Perugia, Perugia, Italy
b Università di Perugia, Perugia, Italy K. Androsov a, P. Azzurri a, G. Bagliesi a, V. Bertacchi a,c, L. Bianchini a, T. Boccali a, R. Castaldi a,
M.A. Ciocci a,b, R. Dell’Orso a, M.R. Di Domenico a,b, S. Donato a, L. Giannini a,c, A. Giassi a, M.T. Grippo a,
F. Ligabue a,c, E. Manca a,c, G. Mandorli a,c, A. Messineo a,b, F. Palla a, A. Rizzi a,b, G. Rolandi a,c,
S. Roy Chowdhury a,c, A. L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy Scribano a, N. Shafiei a,b, P. Spagnolo a, R. Tenchini a, G. Tonelli a,b, N. Turini a,
A. Venturi a, P.G. Verdini a
a INFN Sezione di Pisa, Pisa, Italy
b Università di Pisa, Pisa, Italy
c Scuola Normale Superiore di Pisa, Pisa, Italy
F. Cavallari a, M. Cipriani a,b, D. Del Re a,b, E. Di Marco a, M. Diemoz a, E. Longo a,b, P. Meridiani a,
G. Organtini a,b, F. Pandolfia, R. Paramatti a,b, C. Quaranta a,b, S. Rahatlou a,b, C. Rovelli a,
F. Santanastasio a,b, L. Soffia,b, R. Tramontano a,b
a INFN Sezione di Roma, Rome, Italy
b Sapienza Università di Roma, Rome, Italy K. Androsov a, P. Azzurri a, G. Bagliesi a, V. Bertacchi a,c, L. Bianchini a, T. Boccali a, R. Castaldi a,
M.A. Ciocci a,b, R. Dell’Orso a, M.R. Di Domenico a,b, S. Donato a, L. Giannini a,c, A. Giassi a, M.T. Grippo a,
F. Ligabue a,c, E. Manca a,c, G. Mandorli a,c, A. Messineo a,b, F. Palla a, A. Rizzi a,b, G. Rolandi a,c,
S. Roy Chowdhury a,c, A. Scribano a, N. Shafiei a,b, P. Spagnolo a, R. Tenchini a, G. Tonelli a,b, N. Turini a,
A. Venturi a, P.G. Verdini a
a INFN Sezione di Pisa, Pisa, Italy
b U i
ità di Pi
Pi
It l F. Cavallari a, M. Cipriani a,b, D. Del Re a,b, E. Di Marco a, M. Diemoz a, E. Longo a,b, P. Meridiani a,
G. Organtini a,b, F. Pandolfia, R. Paramatti a,b, C. Quaranta a,b, S. Rahatlou a,b, C. Rovelli a,
F. Santanastasio a,b, L. Soffia,b, R. Tramontano a,b
a INFN Sezione di Roma, Rome, Italy 14 Physics Letters B 816 (2021) 136253 The CMS Collaboration N. Amapane a,b, R. Arcidiacono a,c, S. Argiro a,b, M. Arneodo a,c, N. Bartosik a, R. Bellan a,b, A. Bellora a,b,
C. Biino a, A. Cappati a,b, N. Cartiglia a, S. Cometti a, M. Costa a,b, R. Covarelli a,b, N. Demaria a, B. Kiani a,b,
F. Legger a, C. Mariotti a, S. Maselli a, E. Migliore a,b, V. Monaco a,b, E. Monteil a,b, M. Monteno a,
M.M. Obertino a,b, G. Ortona a, L. Pacher a,b, N. Pastrone a, M. Pelliccioni a, G.L. Pinna Angioni a,b,
M. Ruspa a,c, R. Salvatico a,b, F. Siviero a,b, V. Sola a, A. Solano a,b, D. Soldi a,b, A. Staiano a, D. J. Goh, A. Gurtu H.S. Kim, Y. Kim
Sejong University, Seoul, Republic of Korea J. Almond, J.H. Bhyun, J. Choi, S. Jeon, J. Kim, J.S. Kim, S. Ko, H. Kwon, H
B.H. Oh, M. Oh, S.B. Oh, B.C. Radburn-Smith, H. Seo, U.K. Yang, I. Yoon
Seoul National University, Seoul, Republic of Korea H. Kim, D.H. Moon B. Francois, T.J. Kim, J. Park
Hanyang University, Seoul, Republic of Korea S. Cho, S. Choi, Y. Go, S. Ha, B. Hong, K. Lee, K.S. Lee, J. Lim, J. Park, S.K. Park, Y. Roh, J. Yoo
Korea University, Seoul, Republic of Korea L. Benussi, S. Bianco, D. Piccolo
INFN Laboratori Nazionali di Frascati, Frascati, Italy Trocino a,b
a INFN Sezione di Torino, Torino, Italy
b Università di Torino, Torino, Italy
c Università del Piemonte Orientale, Novara, Italy y
b Università di Torino, Torino, Italy c Università del Piemonte Orientale, Novara, Italy S. Belforte a, V. Candelise a,b, M. Casarsa a, F. Cossutti a, A. Da Rold a,b, G. Della Ricca a,b, F. Vazzoler a,b
a INFN Sezione di Trieste, Trieste, Italy
b Università di Trieste, Trieste, Italy a INFN Sezione di Trieste, Trieste, Italy
b Università di Trieste, Trieste, Italy S. Dogra, C. Huh, B. Kim, D.H. Kim, G.N. Kim, J. Lee, S.W. Lee, C.S. Moon, Y.D. Oh, S.I. Pak, S. Sekmen,
D.C. Son, Y.C. Yang
Kyungpook National University, Daegu, Republic of Korea Kyungpook National University, Daegu, Republic of Korea Y. Maghrbi 15 The CMS Collaboration
Physics Letters B 816 (202
W.A.T. Wan Abdullah, M.N. Yusli, Z. Zolkapli
National Centre for Particle Physics, Universiti Malaya, Kuala Lumpur, Malaysia
J.F. Benitez, A. Castaneda Hernandez, J.A. Murillo Quijada, L. Valencia Palomo
Universidad de Sonora (UNISON), Hermosillo, Mexico
H. Castilla-Valdez, E. De La Cruz-Burelo, I. Heredia-De La Cruz 40, R. Lopez-Fernandez,
A. Sanchez-Hernandez
Centro de Investigacion y de Estudios Avanzados del IPN, Mexico City, Mexico
S. Carrillo Moreno, C. Oropeza Barrera, M. Ramirez-Garcia, F. Vazquez Valencia
Universidad Iberoamericana, Mexico City, Mexico
J. Eysermans, I. Pedraza, H.A. Salazar Ibarguen, C. Uribe Estrada
Benemerita Universidad Autonoma de Puebla, Puebla, Mexico
A. Morelos Pineda
Universidad Autónoma de San Luis Potosí, San Luis Potosí, Mexico
J. Mijuskovic 3, N. Raicevic
University of Montenegro, Podgorica, Montenegro
D. Krofcheck
University of Auckland, Auckland, New Zealand
S. Bheesette, P.H. Butler
University of Canterbury, Christchurch, New Zealand
A. Ahmad, M. Ahmad, M.I. Asghar, M.I.M. Awan, Q. Hassan, H.R. Hoorani, W.A. Khan, M.A. Shah,
M. Shoaib, M. Waqas
National Centre for Physics, Quaid-I-Azam University, Islamabad, Pakistan
V. Avati, L. Grzanka, M. Malawski
AGH University of Science and Technology Faculty of Computer Science, Electronics and Telecommunications, Krakow, Poland
H. Bialkowska, M. Bluj, B. Boimska, T. Frueboes, M. Górski, M. Kazana, M. Szleper, P. Traczyk, P. Zale
National Centre for Nuclear Research, Swierk, Poland
K. Bunkowski, A. Byszuk 41, K. Doroba, A. Kalinowski, M. Konecki, J. Krolikowski, M. Olszewski,
M. Walczak
Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Warsaw, Poland
M. Araujo, P. Bargassa, D. Bastos, A. Di Francesco, P. Faccioli, B. Galinhas, M. Gallinaro, J. Hollar,
N. Leonardo, T. Niknejad, J. Seixas, K. Shchelina, O. Toldaiev, J. Varela
Laboratório de Instrumentação e Física Experimental de Partículas, Lisboa, Portugal
A. Baginyan, A. Golunov, I. Golutvin, I. Gorbunov, V. Karjavine, I. Kashunin, A. Lanev, A. Malakhov,
V. Matveev 42,43, V.V. Mitsyn, P. Moisenz, V. Palichik, V. Perelygin, S. Shmatov, O. Teryaev, V. Trofim
N. Voytishin, B.S. Yuldashev 44, A. Zarubin, V. Zhiltsov The CMS Collaboration Physics Letters B 816 (2021) 136253 W.A.T. Wan Abdullah, M.N. Yusli, Z. Zolkapli
National Centre for Particle Physics, Universiti Malaya, Kuala Lumpur, Malaysia W.A.T. Wan Abdullah, M.N. Yusli, Z. Zolkapli
National Centre for Particle Physics, Universiti Malaya, Kuala Lumpur, Malaysia W.A.T. Wan Abdullah, M.N. Yusli, Z. Zolkapli
National Centre for Particle Physics, Universiti Malaya, Kuala Lumpur, Malaysia J.F. Benitez, A. Castaneda Hernandez, J.A. Murillo Quijada, L. Valencia Palomo
Universidad de Sonora (UNISON), Hermosillo, Mexico A. Ahmad, M. Ahmad, M.I. Asghar, M.I.M. Awan, Q. Hassan, H.R. Hoorani, W.A. Khan, M.A. Shah,
M. Shoaib, M. Waqas
National Centre for Physics, Quaid-I-Azam University, Islamabad, Pakistan
V. Avati, L. Grzanka, M. Malawski
AGH University of Science and Technology Faculty of Computer Science, Electronics and Telecommunications, Krakow, Poland H. Bialkowska, M. Bluj, B. Boimska, T. Frueboes, M. Górski, M. Kazana, M. Szleper, P. Traczyk, P. Zalewski
National Centre for Nuclear Research, Swierk, Poland K. Bunkowski, A. Byszuk 41, K. Doroba, A. Kalinowski, M. Konecki, J. Krolikowski, M. Olszewski,
M. Walczak
Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Warsaw, Poland M. Araujo, P. Bargassa, D. Bastos, A. Di Francesco, P. Faccioli, B. Galinhas, M. Gallinaro, J. Hollar,
N. Leonardo, T. Niknejad, J. Seixas, K. Shchelina, O. Toldaiev, J. Varela
Laboratório de Instrumentação e Física Experimental de Partículas, Lisboa, Portugal A. Baginyan, A. Golunov, I. Golutvin, I. Gorbunov, V. Karjavine, I. Kashunin, A. Lanev, A. Malakhov,
V. Matveev 42,43, V.V. Mitsyn, P. Moisenz, V. Palichik, V. Perelygin, S. Shmatov, O. Teryaev, V. Trofimov,
N. Voytishin, B.S. Yuldashev 44, A. Zarubin, V. Zhiltsov
Joint Institute for Nuclear Research, Dubna, Russia 16 Physics Letters B 816 (2021) 136253 The CMS Collaboration G. Gavrilov, V. Golovtcov, Y. Ivanov, V. Kim 45, E. Kuznetsova 46, V. Murzin, V. Oreshkin, I. Smirnov,
D. Sosnov, V. Sulimov, L. Uvarov, S. Volkov, A. Vorobyev
Petersburg Nuclear Physics Institute, Gatchina (St. Petersburg), Russia Yu. Andreev, A. Dermenev, S. Gninenko, N. Golubev, A. Karneyeu, M. Kirsanov, N. Krasnikov,
A. Pashenkov, G. Pivovarov, D. Tlisov, A. Toropin
Institute for Nuclear Research, Moscow, Russia V. Epshteyn, V. Gavrilov, N. Lychkovskaya, A. Nikitenko 47, V. Popov, I. Pozdnyakov, G. Safronov,
A. Spiridonov, A. Stepennov, M. Toms, E. Vlasov, A. Zhokin
Institute for Theoretical and Experimental Physics named by A.I. Alikhanov of NRC ‘Kurchatov Institute’, Moscow, Russia T. Aushev Moscow Institute of Physics and Technology, Moscow, Russia O. Bychkova, R. Chistov 48, M. Danilov 48, A. Oskin, P. Parygin, S. Polikarpov 48
National Research Nuclear University ‘Moscow Engineering Physics Institute’ (MEPhI), Moscow, Russia V. Andreev, M. Azarkin, I. Dremin, M. Kirakosyan, A. Terkulov
P.N. Lebedev Physical Institute, Moscow, Russia A. Belyaev, E. Boos, A. Demiyanov, A. Ershov, A. Gribushin, O. Kodolova, V. Korotkikh, I. Lokhtin,
S. Obraztsov, S. Petrushanko, V. Savrin, A. Snigirev, I. Vardanyan
Skobeltsyn Institute of Nuclear Physics, Lomonosov Moscow State University, Moscow, Russia I. Azhgirey, I. Bayshev, S. Bitioukov, V. Kachanov, A. Kalinin, D. Konstantinov, V. Petrov, R. Ryutin,
A. Sobol, S. Troshin, N. Tyurin, A. Uzunian, A. Volkov
Institute for High Energy Physics of National Research Centre ‘Kurchatov Institute’, Protvino, Russia A. Babaev, A. Iuzhakov, V. Okhotnikov
National Research Tomsk Polytechnic University, Tomsk, Russia W.G.D. Dharmaratna, K. Liyanage, N. Perera, N. Wickramage
University of Ruhuna, Department of Physics, Matara, Sri Lanka University of Ruhuna, Department of Physics, Matara, Sri Lanka T.K. Aarrestad, D. Abbaneo, B. Akgun, E. Auffray, G. Auzinger, J. Baechler, P. Baillon, A.H. Ball, D. Barney,
J. Bendavid, M. Bianco, A. Bocci, P. Bortignon, E. Bossini, E. Brondolin, T. Camporesi, G. Cerminara,
L. Cristella, D. d’Enterria, A. Dabrowski, N. Daci, V. Daponte, A. David, A. De Roeck, M. Deile, R. Di Maria
M. Dobson, M. Dünser, N. Dupont, A. Elliott-Peisert, N. Emriskova, F. Fallavollita 53, D. Fasanella,
S. Fiorendi, G. Franzoni, J. Fulcher, W. Funk, S. Giani, D. Gigi, K. Gill, F. Glege, L. Gouskos, M. Gruchala,
M. Guilbaud, D. Gulhan, J. Hegeman, C. Heidegger, Y. Iiyama, V. Innocente, T. James, P. Janot, J. Kaspar,
J. Kieseler, M. Komm, N. Kratochwil, C. Lange, P. Lecoq, K. Long, C. Lourenço, L. Malgeri, M. Mannelli,
A. Massironi, F. Meijers, S. Mersi, E. Meschi, F. Moortgat, M. Mulders, J. Ngadiuba, J. Niedziela,
S. Orfanelli, L. Orsini, F. Pantaleo 17, L. Pape, E. Perez, M. Peruzzi, A. Petrilli, G. Petrucciani, A. Pfeiffer,
M. Pierini, F.M. Pitters, D. Rabady, A. Racz, M. Rovere, H. Sakulin, J. Salfeld-Nebgen, S. Scarfi, C. Schäfer,
C. Schwick, M. Selvaggi, A. Sharma, P. Silva, W. Snoeys, P. Sphicas 54, J. Steggemann, S. Summers,
V.R. Tavolaro, D. Treille, A. Tsirou, G.P. Van Onsem, A. Vartak, M. Verzetti, K.A. Wozniak, W.D. Zeuner
CERN, European Organization for Nuclear Research, Geneva, Switzerland L. Caminada 55, K. Deiters, W. Erdmann, R. Horisberger, Q. Ingram, H.C. Kaestli, D. Kotlinski,
U. Langenegger, T. Rohe
Paul Scherrer Institut, Villigen, Switzerland M. Backhaus, P. Berger, A. Calandri, N. Chernyavskaya, G. Dissertori, M. Dittmar, M. Donegà, C. Dorfer,
T. Gadek, T.A. Gómez Espinosa, C. Grab, D. Hits, W. Lustermann, A.-M. Lyon, R.A. Manzoni,
M.T. Meinhard, F. Micheli, P. Musella, F. Nessi-Tedaldi, F. Pauss, V. Perovic, G. Perrin, L. Perrozzi,
S. Pigazzini, M.G. Ratti, M. Reichmann, C. Reissel, T. Reitenspiess, B. Ristic, D. Ruini, D.A. Sanz Becerra,
M. Schönenberger, L. Shchutska, V. Stampf, M.L. Vesterbacka Olsson, R. Wallny, D.H. Zhu
ETH Zurich – Institute for Particle Physics and Astrophysics (IPA), Zurich, Switzerland C. Amsler 56, C. Botta, D. Brzhechko, M.F. Canelli, A. De Cosa, R. Del Burgo, J.K. Heikkilä, M. Huwiler,
B. Kilminster, S. Leontsinis, A. Macchiolo, V.M. Mikuni, I. Neutelings, G. Rauco, P. Robmann, K. Schweiger,
Y. Takahashi, S. Wertz
Universität Zürich, Zurich, Switzerland C. Adloff 57, C.M. Kuo, W. Lin, A. Roy, T. Sarkar 32, S.S. V. Borchsh, V. Ivanchenko, E. Tcherniaev
Tomsk State University, Tomsk, Russia 17 Physics Letters B 816 (2021) 136253 The CMS Collaboration B. Alvarez Gonzalez, J. Cuevas, C. Erice, J. Fernandez Menendez, S. Folgueras, I. Gonzalez Caballero,
E. Palencia Cortezon, C. Ramón Álvarez, V. Rodríguez Bouza, S. Sanchez Cruz
Universidad de Oviedo, Instituto Universitario de Ciencias y Tecnologías Espaciales de Asturias (ICTEA), Oviedo, Spain I.J. Cabrillo, A. Calderon, B. Chazin Quero, J. Duarte Campderros, M. Fernandez, P.J. Fernández Manteca,
A. García Alonso, G. Gomez, C. Martinez Rivero, P. Martinez Ruiz del Arbol, F. Matorras, J. Piedra Gomez,
C. Prieels, F. Ricci-Tam, T. Rodrigo, A. Ruiz-Jimeno, L. Russo 51, L. Scodellaro, I. Vila, J.M. Vizan Garcia
Instituto de Física de Cantabria (IFCA), CSIC-Universidad de Cantabria, Santander, Spain M.K. Jayananda, B. Kailasapathy 52, D.U.J. Sonnadara, D.D.C. Wickramarathna
University of Colombo, Colombo, Sri Lanka W.G.D. Dharmaratna, K. Liyanage, N. Perera, N. Wickramage
University of Ruhuna, Department of Physics, Matara, Sri Lanka W.G.D. Dharmaratna, K. Liyanage, N. Perera, N. Wickramage
University of Ruhuna, Department of Physics, Matara, Sri Lanka Yu
National Central University, Chung-Li, Taiwan L. Ceard, P. Chang, Y. Chao, K.F. Chen, P.H. Chen, W.-S. Hou, Y.y. Li, R.-S. Lu, E. Paganis, A. Psallidas,
A. Steen
National Taiwan University (NTU), Taipei, Taiwan 18 18 Physics Letters B 816 (2021) 136253 The CMS Collaboration B. Asavapibhop, C. Asawatangtrakuldee, N. Srimanobhas
Chulalongkorn University, Faculty of Science, Department of Physics, Bangkok, Thailand B. Asavapibhop, C. Asawatangtrakuldee, N. Srimanobhas
Chulalongkorn University, Faculty of Science, Department of Physics, Bangkok, Thailand A. Bat, F. Boran, S. Damarseckin 58, Z.S. Demiroglu, F. Dolek, C. Dozen 59, I. Dumanoglu 60, E. Eskut,
G. Gokbulut, Y. Guler, E. Gurpinar Guler 61, I. Hos 62, C. Isik, E.E. Kangal 63, O. Kara, A. Kayis Topaksu,
U. Kiminsu, G. Onengut, K. Ozdemir 64, A. Polatoz, A.E. Simsek, B. Tali 65, U.G. Tok, S. Turkcapar,
I.S. Zorbakir, C. Zorbilmez
Çukurova University, Physics Department, Science and Art Faculty, Adana, Turkey B. Isildak 66, G. Karapinar 67, K. Ocalan 68, M. Yalvac 69
Middle East Technical University, Physics Department, Ankara, Turkey I.O. Atakisi, E. Gülmez, M. Kaya 70, O. Kaya 71, Ö. Özçelik, S. Tekten 72, E.A. Yetkin 73
Bogazici University, Istanbul, Turkey A. Cakir, K. Cankocak 60, Y. Komurcu, S. Sen 74
Istanbul Technical University, Istanbul, Turkey F. Aydogmus Sen, S. Cerci 65, B. Kaynak, S. Ozkorucuklu, D. Sunar Cerci 65
Istanbul University, Istanbul, Turkey L. Levchuk E. Bhal, S. Bologna, J.J. Brooke, D. Burns 75, E. Clement, D. Cussans, H. Flacher, J. Goldstein, G.P. Heath,
H.F. Heath, L. Kreczko, B. Krikler, S. Paramesvaran, T. Sakuma, S. Seif El Nasr-Storey, V.J. Smith, J. Taylor,
A. Titterton
University of Bristol, Bristol, United Kingdom K.W. Bell, A. Belyaev 76, C. Brew, R.M. Brown, D.J.A. Cockerill, K.V. Ellis, K. Harder, S. Harper, J. Linacre,
K. Manolopoulos, D.M. Newbold, E. Olaiya, D. Petyt, T. Reis, T. Schuh, C.H. Shepherd-Themistocleous,
A. Thea, I.R. Tomalin, T. Williams
Rutherford Appleton Laboratory, Didcot, United Kingdom R. Bainbridge, P. Bloch, S. Bonomally, J. Borg, S. Breeze, O. Buchmuller, A. Bundock, V. Cepaitis,
G.S. Chahal 77, D. Colling, P. Dauncey, G. Davies, M. Della Negra, P. Everaerts, G. Fedi, G. Hall, G. Iles,
J. Langford, L. Lyons, A.-M. Magnan, S. Malik, A. Martelli, V. Milosevic, A. Morton, J. Nash 78, V. Palladino,
M. Pesaresi, D.M. Raymond, A. Richards, A. Rose, E. Scott, C. Seez, A. Shtipliyski, M. Stoye, A. Tapper,
K. Uchida, T. Virdee 17, N. Wardle, S.N. Webb, D. Winterbottom, A.G. Zecchinelli, S.C. Zenz
Imperial College, London, United Kingdom J.E. Cole, P.R. Hobson, A. Khan, P. Kyberd, C.K. Mackay, I.D. Reid, L. Teodorescu, S. Zahid
Brunel University, Uxbridge, United Kingdom A. Brinkerhoff, K. Call, B. Caraway, J. Dittmann, K. Hatakeyama, C. Madrid, B. McMaster, N. Pastika,
C. Smith
Baylor University, Waco, USA R. Bartek, A. Dominguez, R. Uniyal, A.M. Vargas Hernandez
Catholic University of America, Washington, DC, USA 19 Physics Letters B 816 (2021) 136253 The CMS Collaboration A. Buccilli, O. Charaf, S.I. Cooper, S.V. Gleyzer, C. Henderson, P. Rumerio, C. West A. Buccilli, O. Charaf, S.I. Cooper, S.V. Gleyzer, C. Henderson, P. Rumerio, C. West
The University of Alabama, Tuscaloosa, USA A. Albert, D. Arcaro, Z. Demiragli, D. Gastler, C. Richardson, J. Rohlf, D. Sperka, D. Spitzbart, I. Suarez,
D. Zou
Boston University, Boston, USA G. Benelli, B. Burkle, X. Coubez 18, D. Cutts, Y.t. Duh, M. Hadley, U. Heintz, J.M. Hogan 79, K.H.M. Kwok,
E. Laird, G. Landsberg, K.T. Lau, J. Lee, M. Narain, S. Sagir 80, R. Syarif, E. Usai, W.Y. Wong, D. Yu, W. Zhang
Brown University, Providence, USA R. Band, C. Brainerd, R. Breedon, M. Calderon De La Barca Sanchez, M. Chertok, J. Conway, R. Conway,
P.T. Cox, R. Erbacher, C. Flores, G. Funk, F. Jensen, W. Ko †, O. Kukral, R. Lander, M. Mulhearn, D. Pellett,
J. Pilot, M. L. Levchuk Shi, D. Taylor, K. Tos, M. Tripathi, Z. Wang, Y. Yao, F. Zhang
University of California, Davis, Davis, USA M. Bachtis, C. Bravo, R. Cousins, A. Dasgupta, A. Florent, D. Hamilton, J. Hauser, M. Ignatenko, T. Lam,
N. Mccoll, W.A. Nash, S. Regnard, D. Saltzberg, C. Schnaible, B. Stone, V. Valuev
University of California, Los Angeles, USA K. Burt, Y. Chen, R. Clare, J.W. Gary, S.M.A. Ghiasi Shirazi, G. Hanson, G. Karapostoli, O.R. Long,
N. Manganelli, M. Olmedo Negrete, M.I. Paneva, W. Si, S. Wimpenny, Y. Zhang
University of California, Riverside, Riverside, USA J.G. Branson, P. Chang, S. Cittolin, S. Cooperstein, N. Deelen, M. Derdzinski, J. Duarte, R. Gerosa,
D. Gilbert, B. Hashemi, D. Klein, V. Krutelyov, J. Letts, M. Masciovecchio, S. May, S. Padhi, M. Pieri,
V. Sharma, M. Tadel, F. Würthwein, A. Yagil
University of California, San Diego, La Jolla, USA N. Amin, R. Bhandari, C. Campagnari, M. Citron, A. Dorsett, V. Dutta, J. Incandela, B. Marsh, H. Mei,
A. Ovcharova, H. Qu, J. Richman, U. Sarica, D. Stuart, S. Wang
University of California, Santa Barbara – Department of Physics, Santa Barbara, USA D. Anderson, A. Bornheim, O. Cerri, I. Dutta, J.M. Lawhorn, N. Lu, J. Mao, H.B. Newman, T.Q. Nguyen,
J. Pata, M. Spiropulu, J.R. Vlimant, S. Xie, Z. Zhang, R.Y. Zhu
California Institute of Technology, Pasadena, USA J. Alison, M.B. Andrews, T. Ferguson, T. Mudholkar, M. Paulini, M. Sun, I. Vorobiev, M. Weinberg
Carnegie Mellon University, Pittsburgh, USA J.P. Cumalat, W.T. Ford, E. MacDonald, T. Mulholland, R. Patel, A. Perloff, K. Stenson, K.A. Ulmer,
S.R. Wagner
University of Colorado Boulder, Boulder, USA J. Alexander, Y. Cheng, J. Chu, A. Datta, A. Frankenthal, K. Mcdermott, J. Monroy, J.R. Patterson, D. Quach,
A. Ryd, W. Sun, S.M. Tan, Z. Tao, J. Thom, P. Wittich, M. Zientek
Cornell University, Ithaca, USA S. Abdullin, M. Albrow, M. Alyari, G. Apollinari, A. Apresyan, A. Apyan, S. Banerjee, L.A.T. Bauerdick,
A. Beretvas, D. Berry, J. Berryhill, P.C. Bhat, K. Burkett, J.N. Butler, A. Canepa, G.B. Cerati, H.W.K. Cheung, S. Abdullin, M. Albrow, M. Alyari, G. Apollinari, A. Apresyan, A. Apyan, S. Banerjee, L.A.T. Bauerdick,
A. Beretvas, D. Berry, J. Berryhill, P.C. Bhat, K. Burkett, J.N. Butler, A. Canepa, G.B. Cerati, H.W.K. Cheung,
F. Chlebana, M. Cremonesi, V.D. Elvira, J. Freeman, Z. Gecse, E. Gottschalk, L. Gray, D. L. Levchuk Green,
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R. Lipton, M. Liu, T. Liu, J. Lykken, K. Maeshima, J.M. Marraffino, D. Mason, P. McBride, P. Merkel,
S. Mrenna, S. Nahn, V. O’Dell, V. Papadimitriou, K. Pedro, C. Pena 81, O. Prokofyev, F. Ravera,
A. Reinsvold Hall, L. Ristori, B. Schneider, E. Sexton-Kennedy, N. Smith, A. Soha, W.J. Spalding, L. Spiegel,
S. Stoynev, J. Strait, L. Taylor, S. Tkaczyk, N.V. Tran, L. Uplegger, E.W. Vaandering, M. Wang, H.A. Weber,
A. Woodard
Fermi National Accelerator Laboratory, Batavia, USA D. Acosta, P. Avery, D. Bourilkov, L. Cadamuro, V. Cherepanov, F. Errico, R.D. Field, D. Guerrero,
B.M. Joshi, M. Kim, J. Konigsberg, A. Korytov, K.H. Lo, K. Matchev, N. Menendez, G. Mitselmakher,
D. Rosenzweig, K. Shi, J. Wang, S. Wang, X. Zuo
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USA E. Adams, A. Baden, O. Baron, A. Belloni, S.C. Eno, Y. Feng, N.J. Hadley, S. Jabeen, G.Y. Jeng, R.G. Kellogg,
T. Koeth, A.C. Mignerey, S. Nabili, M. Seidel, A. Skuja, S.C. Tonwar, L. Wang, K. Wong
University of Maryland, College Park, USA Y.R. Joshi Florida International University, Miami, USA Florida International University, Miami, USA Florida International University, Miami, USA T. Adams, A. Askew, D. Diaz, R. Habibullah, S. Hagopian, V. Hagopian, K.F. Johnson, R. Khurana,
T. Kolberg, G. Martinez, H. Prosper, C. Schiber, R. Yohay, J. Zhang
Florida State University, Tallahassee, USA M.M. Baarmand, S. Butalla, T. Elkafrawy 13, M. Hohlmann, D. Noonan, M. Rahmani, M. Saunders,
F. Yumiceva
Florida Institute of Technology, Melbourne, USA M.R. Adams, L. Apanasevich, H. Becerril Gonzalez, R.R. Betts, R. Cavanaugh, X. Chen, S. Dittmer,
O. Evdokimov, C.E. Gerber, D.A. Hangal, D.J. Hofman, V. Kumar, C. Mills, G. Oh, T. Roy, M.B. Tonjes,
N. Varelas, J. Viinikainen, H. Wang, X. Wang, Z. Wu
University of Illinois at Chicago (UIC), Chicago, USA M. Alhusseini, B. Bilki 61, K. Dilsiz 82, S. Durgut, R.P. Gandrajula, M. Haytmyradov, V. Khristenko,
O.K. Köseyan, J.-P. Merlo, A. Mestvirishvili 83, A. Moeller, J. Nachtman, H. Ogul 84, Y. Onel, F. Ozok 85,
A. Penzo, C. Snyder, E. Tiras, J. Wetzel, K. Yi 86
The University of Iowa, Iowa City, USA O. Amram, B. Blumenfeld, L. Corcodilos, M. Eminizer, A.V. Gritsan, S. Kyriacou, P. Maksimovic,
C. Mantilla, J. Roskes, M. Swartz, T.Á. Vámi
Johns Hopkins University, Baltimore, USA C. Baldenegro Barrera, P. Baringer, A. Bean, S. Boren, A. Bylinkin, T. Isidori, S. Khalil, J. King, G. Krintiras,
A. Kropivnitskaya, C. Lindsey, W. Mcbrayer, N. Minafra, M. Murray, C. Rogan, C. Royon, S. Sanders,
E. Schmitz, J.D. Tapia Takaki, Q. Wang, J. Williams, G. Wilson
The University of Kansas, Lawrence, USA S. Duric, A. Ivanov, K. Kaadze, D. Kim, Y. Maravin, D.R. Mendis, T. Mitchell, A. Modak, A. Mohammadi
Kansas State University, Manhattan, USA
F. Rebassoo, D. Wright S. Duric, A. Ivanov, K. Kaadze, D. Kim, Y. Maravin, D.R. Mendis, T. Mitchell, A. Modak, A. Mohammadi
Kansas State University, Manhattan, USA F. Rebassoo, D. Wright
Lawrence Livermore National Laboratory, Livermore, USA E. Adams, A. Baden, O. Baron, A. Belloni, S.C. Eno, Y. Feng, N.J. Hadley, S. Jabeen, G.Y. Jeng, R.G. Kellogg,
T. Koeth, A.C. Mignerey, S. Nabili, M. Seidel, A. Skuja, S.C. Tonwar, L. Wang, K. Wong
University of Maryland, College Park, USA 21 Physics Letters B 816 (2021) 136253 The CMS Collaboration D. Abercrombie, B. Allen, R. Bi, S. Brandt, W. Busza, I.A. Cali, Y. Chen, M. D’Alfonso, G. Gomez Ceballos,
M. Goncharov, P. Harris, D. Hsu, M. Hu, M. Klute, D. Kovalskyi, J. Krupa, Y.-J. Lee, P.D. Luckey, B. Maier,
A.C. Marini, C. Mcginn, C. Florida International University, Miami, USA Mironov, S. Narayanan, X. Niu, C. Paus, D. Rankin, C. Roland, G. Roland, Z. Shi,
G.S.F. Stephans, K. Sumorok, K. Tatar, D. Velicanu, J. Wang, T.W. Wang, B. Wyslouch
Massachusetts Institute of Technology, Cambridge, USA R.M. Chatterjee, A. Evans, S. Guts †, P. Hansen, J. Hiltbrand, Sh. Jain, M. Krohn, Y. Kubota, Z. Lesko,
J. Mans, M. Revering, R. Rusack, R. Saradhy, N. Schroeder, N. Strobbe, M.A. Wadud
University of Minnesota, Minneapolis, USA J.G. Acosta, S. Oliveros
University of Mississippi, Oxford, USA K. Bloom, S. Chauhan, D.R. Claes, C. Fangmeier, L. Finco, F. Golf, J.R. González Fernández, I. Kravchenko,
J.E. Siado, G.R. Snow †, B. Stieger, W. Tabb
University of Nebraska-Lincoln, Lincoln, USA G. Agarwal, C. Harrington, I. Iashvili, A. Kharchilava, C. McLean, D. Nguyen, A. Parker, J. Pekkanen,
S. Rappoccio, B. Roozbahani
State University of New York at Buffalo, Buffalo, USA G. Alverson, E. Barberis, C. Freer, Y. Haddad, A. Hortiangtham, G. Madigan, B. Marzocchi, D.M. Morse,
V. Nguyen, T. Orimoto, L. Skinnari, A. Tishelman-Charny, T. Wamorkar, B. Wang, A. Wisecarver, D. Wood
Northeastern University, Boston, USA S. Bhattacharya, J. Bueghly, Z. Chen, A. Gilbert, T. Gunter, K.A. Hahn, N. Odell, M.H. Schmitt, K. Sung,
M. Velasco
Northwestern University, Evanston, USA R. Bucci, N. Dev, R. Goldouzian, M. Hildreth, K. Hurtado Anampa, C. Jessop, D.J. Karmgard, K. Lannon,
W. Li, N. Loukas, N. Marinelli, I. Mcalister, F. Meng, K. Mohrman, Y. Musienko 42, R. Ruchti, P. Siddireddy,
S. Taroni, M. Wayne, A. Wightman, M. Wolf, L. Zygala
University of Notre Dame, Notre Dame, USA J. Alimena, B. Bylsma, B. Cardwell, L.S. Durkin, B. Francis, C. Hill, W. Ji, A. Lefeld, B.L. Winer, B.R. Yates
The Ohio State University, Columbus, USA G. Dezoort, P. Elmer, N. Haubrich, S. Higginbotham, A. Kalogeropoulos, G. Kopp, S. Kwan, D. Lange,
M.T. Lucchini, J. Luo, D. Marlow, K. Mei, I. Ojalvo, J. Olsen, C. Palmer, P. Piroué, D. Stickland, C. Tully
Princeton University, Princeton, USA S. Malik, S. Norberg
University of Puerto Rico, Mayaguez, USA V.E. Barnes, R. Chawla, S. Das, L. Gutay, M. Jones, A.W. Jung, B. Mahakud, G. Negro, N. Neumeister,
C.C. Peng, S. Piperov, H. Qiu, J.F. Schulte, N. Trevisani, F. Wang, R. Xiao, W. Xie
Purdue University, West Lafayette, USA T. Cheng, J. Dolen, N. Parashar
Purdue University Northwest, Hammond, USA 22 Physics Letters B 816 (2021) 136253 The CMS Collaboration A. Baty, S. Dildick, K.M. Ecklund, S. Freed, F.J.M. Florida International University, Miami, USA Geurts, M. Kilpatrick, A. Kumar, W. Li, B.P. Padley,
R. Redjimi, J. Roberts †, J. Rorie, W. Shi, A.G. Stahl Leiton, Z. Tu, A. Zhang
Rice University, Houston, USA A. Bodek, P. de Barbaro, R. Demina, J.L. Dulemba, C. Fallon, T. Ferbel, M. Galanti, A. Garcia-Bellido,
O. Hindrichs, A. Khukhunaishvili, E. Ranken, R. Taus
University of Rochester, Rochester, USA B. Chiarito, J.P. Chou, A. Gandrakota, Y. Gershtein, E. Halkiadakis, A. Hart, M. Heindl, E. Hughes,
S. Kaplan, O. Karacheban 21, I. Laflotte, A. Lath, R. Montalvo, K. Nash, M. Osherson, S. Salur, S. Schnetzer,
S. Somalwar, R. Stone, S. Thomas
Rutgers, The State University of New Jersey, Piscataway, USA University of Wisconsin – Madison, Madison, WI, USA University of Wisconsin – Madison, Madison, WI, USA † Deceased. 1 Also at Vienna University of Technology, Vienna, Austria. 2 Also at Université Libre de Bruxelles, Bruxelles, Belgium. 3 Also at IRFU, CEA, Université Paris-Saclay, Gif-sur-Yvette, France. 4 Also at Universidade Estadual de Campinas, Campinas, Brazil. 5 Also at Federal University of Rio Grande do Sul, Porto Alegre, Brazil. 6 Also at UFMS, Nova Andradina, Brazil. 7 Also at Universidade Federal de Pelotas, Pelotas, Brazil. 8 Also at University of Chinese Academy of Sciences, Beijing, China. 9 Also at Institute for Theoretical and Experimental Physics named by A.I. Alikhan
10 Also at Joint Institute for Nuclear Research, Dubna, Russia. 11 Also at Helwan University, Cairo, Egypt. 12 Now at Zewail City of Science and Technology, Zewail, Egypt. 13 Also at Ain Shams University, Cairo, Egypt. 14 Also at Purdue University, West Lafayette, USA. † Deceased. 1 Also at Vienna University of Technology, Vienna, Austria. 2 Also at Université Libre de Bruxelles, Bruxelles, Belgium. 3 Also at IRFU, CEA, Université Paris-Saclay, Gif-sur-Yvette, France. 4 Also at Universidade Estadual de Campinas, Campinas, Brazil. 5 Also at Federal University of Rio Grande do Sul, Porto Alegre, Brazil. 6 Also at UFMS, Nova Andradina, Brazil. 7 Also at Universidade Federal de Pelotas, Pelotas, Brazil. 8 Also at University of Chinese Academy of Sciences, Beijing, China. 9 Also at Institute for Theoretical and Experimental Physics named by A.I. Alikhanov of NRC ‘Kurchatov Institute’, Moscow,
10 Also at Joint Institute for Nuclear Research, Dubna, Russia. 11 Also at Helwan University, Cairo, Egypt. 12 Now at Zewail City of Science and Technology, Zewail, Egypt. 13 Also at Ain Shams University, Cairo, Egypt. 14 Also at Purdue University, West Lafayette, USA. † Deceased. 1 Also at Vienna University of Technology, Vienna, Austria. 1 Also at Vienna University of Technology, Vienna, Austria. 2 Also at Université Libre de Bruxelles, Bruxelles, Belgium. 2 Also at Université Libre de Bruxelles, Bruxelles, Belgium. 3 Also at IRFU, CEA, Université Paris-Saclay, Gif-sur-Yvette, France. 4 Also at Universidade Estadual de Campinas, Campinas, Brazil. 5 Also at Federal University of Rio Grande do Sul, Porto Alegre, Brazil. 5 Also at Federal University of Rio Grande do Sul, Porto Alegre, Brazil. 6 Also at UFMS, Nova Andradina, Brazil. 7 Also at Universidade Federal de Pelotas, Pelotas, Brazil. 8 Also at University of Chinese Academy of Sciences, Beijing, China. P.E. Karchin, N. Poudyal, J. Sturdy, P. Thapa
Wayne State University, Detroit, USA K. Black, T. Bose, J. Buchanan, C. Caillol, S. Dasu, I. De Bruyn, L. Dodd, C. Galloni, H. He, M. Herndon,
A. Hervé, U. Hussain, A. Lanaro, A. Loeliger, R. Loveless, J. Madhusudanan Sreekala, A. Mallampalli,
D. Pinna, T. Ruggles, A. Savin, V. Shang, V. Sharma, W.H. Smith, D. Teague, S. Trembath-reichert,
W. Vetens K. Black, T. Bose, J. Buchanan, C. Caillol, S. Dasu, I. De Bruyn, L. Dodd, C. Galloni, H. He, M. Herndon,
A. Hervé, U. Hussain, A. Lanaro, A. Loeliger, R. Loveless, J. Madhusudanan Sreekala, A. Mallampalli,
D. Pinna, T. Ruggles, A. Savin, V. Shang, V. Sharma, W.H. Smith, D. Teague, S. Trembath-reichert,
W. Vetens K. Black, T. Bose, J. Buchanan, C. Caillol, S. Dasu, I. De Bruyn, L. Dodd, C. Galloni, H. He, M. Herndon,
A. Hervé, U. Hussain, A. Lanaro, A. Loeliger, R. Loveless, J. Madhusudanan Sreekala, A. Mallampalli,
D. Pinna, T. Ruggles, A. Savin, V. Shang, V. Sharma, W.H. Smith, D. Teague, S. Trembath-reichert,
W. Vetens University of Wisconsin – Madison, Madison, WI, USA H. Acharya, A.G. Delannoy, S. Spanier
University of Tennessee, Knoxville, USA H. Acharya, A.G. Delannoy, S. Spanier
University of Tennessee, Knoxville, USA O. Bouhali 87, M. Dalchenko, A. Delgado, R. Eusebi, J. Gilmore, T. Huang, T. Kamon 88, H. Kim, S. Luo,
S. Malhotra, D. Marley, R. Mueller, D. Overton, L. Perniè, D. Rathjens, A. Safonov
Texas A&M University, College Station, USA N. Akchurin, J. Damgov, V. Hegde, S. Kunori, K. Lamichhane, S.W. Lee, T. Mengke, S. Muthumuni,
T. Peltola, S. Undleeb, I. Volobouev, Z. Wang, A. Whitbeck
Texas Tech University, Lubbock, USA E. Appelt, S. Greene, A. Gurrola, R. Janjam, W. Johns, C. Maguire, A. Melo, H. Ni, K. Padeken, F. Romeo,
P. Sheldon, S. Tuo, J. Velkovska, M. Verweij
Vanderbilt University, Nashville, USA L. Ang, M.W. Arenton, B. Cox, G. Cummings, J. Hakala, R. Hirosky, M. Joyce, A. Ledovskoy, C. Neu,
B. Tannenwald, Y. Wang, E. Wolfe, F. Xia
University of Virginia, Charlottesville, USA P.E. Karchin, N. Poudyal, J. Sturdy, P. Thapa
Wayne State University, Detroit, USA Also at Institute for Theoretical and Experimental Physics named by A.I. Alikhanov of NRC ‘Kurchatov Institute’, Moscow, Russia.
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i University of Wisconsin – Madison, Madison, WI, USA Petersburg, Russia. 45 Also at St. Petersburg State Polytechnical University, St. Petersburg, Russia. 46 Also at University of Florida, Gainesville, USA. 47 Also at Imperial College, London, United Kingdom. 48 Also at P.N. Lebedev Physical Institute, Moscow, Russia. 49 Also at Budker Institute of Nuclear Physics, Novosibirsk, Russia. 49 Also at Budker Institute of Nuclear Physics, Novosibirsk, Russia. 50 Also at Faculty of Physics, University of Belgrade, Belgrade, Serbia. 50 Also at Faculty of Physics, University of Belgrade, Belgrade, Serbia. 51 Also at Università degli Studi di Siena, Siena, Italy. 52 Also at Trincomalee Campus, Eastern University, Sri Lanka, Nilaveli, Sri Lanka. 52 Also at Trincomalee Campus, Eastern University, Sri Lanka, Nilaveli, Sri Lanka. 52 Also at Trincomalee Campus, Eastern University, Sri Lank 53 Also at INFN Sezione di Pavia a, Università di Pavia b, Pavia, Italy, Pavia, Italy. 53 Also at INFN Sezione di Pavia a, Università di Pavia b, Pavia, Italy, Pavia, Italy. 54 Also at National and Kapodistrian University of Athens, Athens, Greece. 54 Also at National and Kapodistrian University of Athens, Athens, Greece. 55 Also at Universität Zürich, Zurich, Switzerland. 56 Also at Stefan Meyer Institute for Subatomic Physics, Vienna, Austria, Vienna, Austria 56 Also at Stefan Meyer Institute for Subatomic Physics, Vienna, Austria, Vienna, Austria oire d’Annecy-le-Vieux de Physique des Particules, IN2P3-CNRS, An 57 Also at Laboratoire d’Annecy-le-Vieux de Physique des Particules, IN2P3-CNRS, Annecy-le-Vieux, France. 57 Also at Laboratoire d’Annecy-le-Vieux de Physique des Particules, IN2P3-CNRS, Annecy-l 58 Also at ¸Sırnak University, Sirnak, Turkey. 59 Also at Department of Physics, Tsinghua University, Beijing, China, Beijing, Ch 59 Also at Department of Physics, Tsinghua University, Beijing, China, Beijing, China. 60 Also at Near East University, Research Center of Experimental Health Science, Nicosia, Turk 60 Also at Near East University, Research Center of Experimental Health Science, Nicosia, Turkey. 60 Also at Near East University, Research Center of Experimental 61 Also at Beykent University, Istanbul, Turkey, Istanbul, Turkey. 61 Also at Beykent University, Istanbul, Turkey, Istanbul, Turkey. 62 Also at Istanbul Aydin University, Application and Research Center for Advanced Studies (App. & Res. Cent. for Advanced Studies), Istanbul, Turkey. 63 Also at Mersin University, Mersin, Turkey. 62 Also at Istanbul Aydin University, Application and Research Cen 62 Also at Istanbul Aydin University, Application and Research Center for Advanced Studies (App. & Res. Cent. for Advanced Studies), Istanbul, Turkey. 63 63 Also at Mersin University, Mersin, Turkey. University of Wisconsin – Madison, Madison, WI, USA 27 Al
IIT Bh b
Bh b
I di
Bh b
I di 26 Also at MTA-ELTE Lendület CMS Particle and Nuclear Physics Group, Eötvös Loránd University, Budapest, Hunga 26 Also at MTA-ELTE Lendület CMS Particle and Nuclear Physics Group, Eötvös Loránd University, Budapest, Hungary, Budapest, Hun 26 Also at MTA-ELTE Lendület CMS Particle and Nuclear Physics Group, E 27 Also at IIT Bhubaneswar, Bhubaneswar, India, Bhubaneswar, India. 27 Also at IIT Bhubaneswar, Bhubaneswar, India, Bhubaneswar, India. 28 Also at Institute of Physics, Bhubaneswar, India. 29 Also at G.H.G. Khalsa College, Punjab, India. 30 Also at Shoolini University, Solan, India. 31 Also at University of Hyderabad, Hyderabad, India. 32 Also at University of Visva-Bharati, Santiniketan, India. 33 Also at Indian Institute of Technology (IIT), Mumbai, India. 34 Also at Deutsches Elektronen-Synchrotron, Hamburg, Germany. 35 Also at Department of Physics, University of Science and Technology of Mazandaran, Behshahr, Iran. 35 Also at Department of Physics, University of Science and Technology of Mazandaran, Behshahr, Iran. 6 Now at INFN Sezione di Bari a, Università di Bari b, Politecnico d 37 Also at Italian National Agency for New Technologies, Energy and Sustainable Economic Development, Bologna, Italy. 37 Also at Italian National Agency for New Technologies, Energy and Sustainable Economic Development, Bologna, Italy. 37 Also at Italian National Agency for New Technologies, Energy and Sustainable Econom 38 Also at Centro Siciliano di Fisica Nucleare e di Struttura Della Materia, Catania, Italy. 38 Also at Centro Siciliano di Fisica Nucleare e di Struttura Della Materia, Catania, Italy. 38 Also at Centro Siciliano di Fisica Nucleare e di Struttura Della 39 Also at Riga Technical University, Riga, Latvia, Riga, Latvia. 40 Also at Consejo Nacional de Ciencia y Tecnología, Mexico City 40 Also at Consejo Nacional de Ciencia y Tecnología, Mexico City, Mexico. 41 Also at Warsaw University of Technology, Institute of Electronic Systems, Warsaw, Poland. 41 Also at Warsaw University of Technology, Institute of Electronic Systems, Warsaw, Poland. 41 Also at Warsaw University of Technology, Institute of Ele 42 Also at Institute for Nuclear Research, Moscow, Russia. 43 Now at National Research Nuclear University ‘Moscow Engineering Physics Institute’ (MEPhI), Moscow, Russia. 43 Now at National Research Nuclear University ‘Moscow Engineering Physics Institute’ (MEPhI), Moscow 43 Now at National Research Nuclear University ‘Moscow Engin 4 Also at Institute of Nuclear Physics of the Uzbekistan Academy o 45 Also at St. Petersburg State Polytechnical University, St. University of Wisconsin – Madison, Madison, WI, USA 9 Also at Institute for Theoretical and Experimental Physics named by A.I. Alikhanov of NRC ‘Kurchatov Institute’, Moscow, Rus
10 9 Also at Institute for Theoretical and Experimental Physics named by A.I. Alikhanov of NRC ‘Kurch 9 Also at Institute for Theoretical and Experimental Physics name 10 Also at Joint Institute for Nuclear Research, Dubna, Russia. 11 Also at Helwan University, Cairo, Egypt. 12 Now at Zewail City of Science and Technology, Zewail, Egypt. 13 Also at Ain Shams University, Cairo, Egypt. 14 Also at Purdue University, West Lafayette, USA. 23 Physics Letters B 816 (2021) 136253 The CMS Collaboration 15 Also at Université de Haute Alsace, Mulhouse, France. 15 Also at Université de Haute Alsace, Mulhouse, France. 16 Also at Erzincan Binali Yildirim University, Erzincan, Turkey. 16 Also at Erzincan Binali Yildirim University, Erzincan, Turkey. 17 Also at CERN, European Organization for Nuclear Research, Ge 18 Also at RWTH Aachen University, III. Physikalisches Institut A, Aachen, Germany. 18 Also at RWTH Aachen University, III. Physikalisches Institut A, Aachen, Germany. 18 Also at RWTH Aachen University, III. Physikalisches In 19 Also at University of Hamburg, Hamburg, Germany. y
g
g
y
20 Also at Department of Physics, Isfahan University of Technology, Isfahan, Iran, Isfahan, Iran. 20 Also at Department of Physics, Isfahan University of Technology, Isfahan, Iran, Isfahan, Iran. 20 Also at Department of Physics, Isfahan University of Te 1 Also at Brandenburg University of Technology, Cottbus, Germany g
y
gy
y
22 Also at Skobeltsyn Institute of Nuclear Physics, Lomonosov Moscow State University, Moscow, Russia. 22 Also at Skobeltsyn Institute of Nuclear Physics, Lomonosov Moscow State University, Moscow, Russia 2 Also at Skobeltsyn Institute of Nuclear Physics, Lomonosov Mosc 23 Also at Institute of Physics, University of Debrecen, Debrecen, Hungary, Debrecen, Hungary. 23 Also at Institute of Physics, University of Debrecen, Debrecen, Hungary, Debrecen, Hungary. 3 Also at Institute of Physics, University of Debrecen, Debrecen, H 24 Also at Physics Department, Faculty of Science, Assiut University, Assiut, Egypt. 4 Also at Physics Department, Faculty of Science, Assiut University 5 Also at Institute of Nuclear Research ATOMKI, Debrecen, Hungar 26 Also at MTA-ELTE Lendület CMS Particle and Nuclear Physics Group, Eötvös Loránd University, Budapest, Hungary, Budapest, Hungary. 86 Also at Nanjing Normal University Department of Physics, Nanjing, China. 87 Also at Texas A&M University at Qatar, Doha, Qatar. 88 Also at Kyungpook National University, Daegu, Korea, Daegu, Republic of Korea. The CMS Collaboration 86 Also at Nanjing Normal University Department of Physics, Nanjing, China.
87 Also at Texas A&M University at Qatar, Doha, Qatar.
88 Also at Kyungpook National University, Daegu, Korea, Daegu, Republic of Korea. University of Wisconsin – Madison, Madison, WI, USA 64 Also at Piri Reis University, Istanbul, Turkey. 65 Also at Adiyaman University, Adiyaman, Turkey. 65 Also at Adiyaman University, Adiyaman, Turkey. 66 Also at Ozyegin University, Istanbul, Turkey. 67 Also at Izmir Institute of Technology, Izmir, Turke 67 Also at Izmir Institute of Technology, Izmir, Turkey. 68 Also at Necmettin Erbakan University, Konya, Turkey. 68 Also at Necmettin Erbakan University, Konya, Turkey. 69 Also at Bozok Universitetesi Rektörlügü, Yozgat, Turkey. 69 Also at Bozok Universitetesi Rektörlügü, Yozgat, Turkey. 70 Also at Marmara University, Istanbul, Turkey. 71 Also at Milli Savunma University, Istanbul, Turk 72 Also at Kafkas University, Kars, Turkey. 73 Also at Istanbul Bilgi University, Istanbul, Turkey. 73 Also at Istanbul Bilgi University, Istanbul, Turkey. 74 Also at Hacettepe University, Ankara, Turkey. 75 Also at Vrije Universiteit Brussel, Brussel, Belgium. 75 Also at Vrije Universiteit Brussel, Brussel, Belgium. 76 Also at School of Physics and Astronomy, University of Southampton, Southampton, United Kingdom. 76 Also at School of Physics and Astronomy, University o 76 Also at School of Physics and Astronomy, University of Southampton, Southampton, United Kingdo 77 Also at IPPP Durham University, Durham, United Kingdom. 77 Also at IPPP Durham University, Durham, United Kingdom. 78 Also at Monash University, Faculty of Science, Clayton, Austral 78 Also at Monash University, Faculty of Science, Clayton, Austr 79 Also at Bethel University, St. Paul, Minneapolis, USA, St. Paul, USA. 79 Also at Bethel University, St. Paul, Minneapolis, USA, St. Paul, USA. 80 Also at Karamano˘glu Mehmetbey University, Karaman, Turkey. 80 Also at Karamano˘glu Mehmetbey University, Karaman, Turkey. 81 Also at California Institute of Technology, Pasadena, USA. 81 Also at California Institute of Technology, Pasadena, USA. 82 Also at Bingol University, Bingol, Turkey. 83 Also at Georgian Technical University, Tbilisi, Georgia. 83 Also at Georgian Technical University, Tbilisi, Georgia. 84 Also at Sinop University, Sinop, Turkey. 85 Also at Mimar Sinan University, Istanbul, Istanbul, Turkey. 85 Also at Mimar Sinan University, Istanbul, Istanbul, Turkey. 24 Physics Letters B 816 (2021) 136253 25
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Which Will Be the Right One? The Beginnings of Fairytale Genre in Czech Animated Film after 1948
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1) Pokud bychom se nezaměřovali pouze na období státního filmového monopolu, pak by historie domácí fil-
mové pohádky či obecněji filmu pro děti začínala už před rokem 1945, kdy se objevují první individuální
pokusy o tento typ tvorby (hrané tituly Oldřicha Kmínka nebo animované snímky Hermíny Týrlové). Ná-
sledující text se však věnuje vývoji filmu pro děti právě v podmínkách zestátněné československé kinemato-
grafie. gi
2) K počátkům filmu pro děti v Československu viz Lukáš Skupa, Film pro děti mezi vědou, uměním a průmy-
slem. Počátky žánru dětského filmu v české kinematografii 1945 až 1955. In: Pavel Skopal (ed.), Naplánova-
ná kinematografie. Český filmový průmysl 1945 až 1960. Praha 2012, s. 166–174. Lukáš Skupa
(Národní filmový archiv) Lukáš Skupa
(Národní filmový archiv) 1) Pokud bychom se nezaměřovali pouze na období státního filmového monopolu, pak by historie domácí fil-
mové pohádky či obecněji filmu pro děti začínala už před rokem 1945, kdy se objevují první individuální
pokusy o tento typ tvorby (hrané tituly Oldřicha Kmínka nebo animované snímky Hermíny Týrlové). Ná-
sledující text se však věnuje vývoji filmu pro děti právě v podmínkách zestátněné československé kinemato-
grafie.
2) K počátkům filmu pro děti v Československu viz Lukáš Skupa, Film pro děti mezi vědou, uměním a průmy-
slem. Počátky žánru dětského filmu v české kinematografii 1945 až 1955. In: Pavel Skopal (ed.), Naplánova-
ná kinematografie. Český filmový průmysl 1945 až 1960. Praha 2012, s. 166–174. ČLÁNKY K TÉMATU ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) 9 Počátky žánru pohádky v českém animovaném filmu po roce 1948 Historie české fi lmové pohádky v zestátněné kinematografi i obvykle začíná v roce 1952
u Pyšné princezny.1) V tomtéž roce však měla premiéru ještě jiná pohádka — Jablůňka
se zlatými jablky vyprávějící o zvířátkách, kterým pionýři dopomohli k vypěstování
chutných jablek. Snímek v režii Eduarda Hofmana byl považován za dosud nejambicióz-
nější projekt studia Bratři v triku — zásadní nejen svou celkovou poetikou, ale i průkop-
nický z technicko-realizačního hlediska. Šlo totiž o vůbec první československý kreslený
fi lm střední metráže. Po svém uvedení získal kromě ceny na MFF v Karlových Varech také
status vzorového díla domácí fi lmové tvorby pro děti. i
Kdysi tak významný snímek, kterým vyvrcholil dlouhodobější a poměrně intenzivní
žánrový vývoj, zná dnes jen málokdo. A je na tom cosi příznačného. Dějiny poválečné čes-
koslovenské animace stále zůstávají vyprávěním o fenomenálním úspěchu odstartovaném
zestátněním kinematografi e v roce 1945 a zejména pak příběhem úspěchů Jiřího Trnky,
Karla Zemana a Hermíny Týrlové. Paralelně s jejich fi lmy však vznikalo i mnoho dalších,
z nichž některé upadly v zapomnění, jiné se v historických přehledech sotva mihnou jako
příklady toho, jak se animátoři po únoru 1948 proti své vůli podřídili diktátu centralizo-
vané dramaturgie a z donucení změnili styl tvorby. Z většiny jde právě o fi lmy pro děti
a především o pohádky.i V tomto textu se pokusím přiblížit jednu z kapitol utváření žánru fi lmu pro děti, jemuž
byla v zestátněné kinematografi i přisouzena mimořádná pozice.2) Zatímco hrané fi lmy Lukáš Skupa: Která bude ta pravá? 10 10 pro děti vznikaly v první poválečné dekádě spíš ojediněle, v oblasti animace se tvorba pro
děti dostala přímo do čela plánu a probíhaly zde živelné diskuze o její podobě. Za krátkou
dobu se právě ve studiích animovaného fi lmu vyprofi lovaly základy tohoto žánru, jehož
výzkum představuje několikanásobnou výzvu — od minimálního zpracování daného té-
matu přes stále nedořešené otázky výzkumu žánrů v socialistickém fi lmovém systému až
po skutečnost, že kategorie „dětského fi lmu“ v sobě zahrnuje další žánrové druhy. i
Jinou badatelskou výzvu, jež se dotýká i tohoto textu, představují silné vazby domácí-
ho fi lmu pro děti s literaturou. Také mnoho kreslených a loutkových snímků čerpalo z kla-
sických nebo moderních děl převážně české literatury, ke spolupráci na animovaných fi l-
mech byli navíc pravidelně vyzýváni spisovatelé a ilustrátoři dětských publikací. Svým
zaměřením se tedy text vztahuje k problematice adaptace. 3) Více na toto téma viz Lukáš Skupa, Ze zámku do podzámčí. Kontexty prvních československých filmových
pohádek. Cinepur 19, 2012, č. 79, s. 85–88.
4) Například v roce 1951 bylo z devadesáti hodnocených námětů k dalšímu zpracování přijato devět. Zároveň
prošlo lekturou 513 pohádek, z nichž ke zpracování bylo přijato pět a do zálohy tři. Zápis konference výrob-
ních složek Čs. státního filmu a poradního sboru pro výrobu při 21. Svazu ROH, Praha, 22. 1. 1952. Národ-
ní filmový archiv (NFA), f. Ústřední ředitelství Československého státního filmu (ÚŘ ČSF), k. R10/AI/
2P/9K. 3) Více na toto téma viz Lukáš Skupa, Ze zámku do podzámčí. Kontexty prvních československých filmových
pohádek. Cinepur 19, 2012, č. 79, s. 85–88. 5) Zpráva o první celostátní konferenci Československého státního filmu, Praha, listopad 1948. NFA, f. ÚŘ
ČSF, k. R10/AII/2P/9K.
6) Srov. např. Náš kreslený film. Kino 3, 1948, č. 35, s. 651; Otakar Mrkvička, Pod lupou. Výboje českých filmo-
vých kreslířů stále ještě nestrefují hřebíček na hlavičku. Kino 4, 1949, č. 8, s. 100–101; Nový český kreslený
film pro nejmenší. Kino 5, 1950, č. 23, s. 536–537.
7) Plnění úkolů vyplývajících z usnesení předsednictva ÚV KSČ, Praha, 9. 10. 1950. NFA, f. ÚŘ ČSF, k. R5/AII/
2P/6K.
8) Zápis o poradě vedoucích pracovníků úseků ČSF konané dne 27. června 1952 ve Filmovém klubu v Praze.
NFA, f. ÚŘ ČSF, k. R9/BI/4P/4K. Počátky žánru pohádky v českém animovaném filmu po roce 1948 Mohl by se vydat nejjednoduš-
ší cestou srovnávání literárních předloh s hotovými díly, což by však mnoho překvapivých
závěrů nepřineslo. Potvrdilo by se jen to, co lze tušit od začátku — že v době rozmáhající-
ho se socialistického realismu a budovatelského dramatu docházelo k tendenčním úpra-
vám či akcentování některých příhodných motivů adaptovaných děl.3) Objevnější zjištění může na bídnout průzkum dramaturgie, jejíž orgány sehrály zásad-
ní roli v rozhodování, co a jak adaptovat, a zároveň vyjednávaly spolupráci s literáty nebo
výtvarníky. V této studii se soustředím na nejzákladnější otázky, jimiž se dramaturgie za-
bývala ve vztahu k žánru animované pohádky a jejím literárním předobrazům: jak sama
zacházela s kategorií pohádky, jak přijímala či odmítala některé její typy ustavené v literár-
ní oblasti a jaké pohádky nakonec uznala za vlastní ideální vzor a proč.l Počátky pohádkového žánru budu refl ektovat v přibližném rozmezí let 1948 až 1953,
kdy se domácí animovaná tvorba dočasně přeorientovala téměř výhradně na dětské pub-
likum. Neopomenu ani projekty, jejichž výroba z různých příčin ztroskotala, protože pro
pochopení priorit dramaturgie se zdají stejně důležité jako snímky realizované. Pro něko-
lik sledovaných let je typický citelný pokles natáčených fi lmů a naopak vysoký počet zamí-
taných námětů i neuskutečněných projektů, jež někdy postoupily i do pokročilejší fáze
literárních příprav.4) Vědomě se vzdávám detailnější analýzy či interpretace konkrétních
děl — domnívám se, že k rozboru těchto adaptací by bylo lepší přistoupit teprve po zma-
pování strategií a nároků dramaturgie, které musely nutně ovlivňovat, jaké fi lmy a v jaké
podobě vůbec vzniknou. Po roce 1948 fungovala studia kresleného a loutkového fi lmu nejprve jako součást
Krátkého fi lmu (dále KF) s vlastní dramaturgií podřízenou Ústřední dramaturgii krátko-
metrážního fi lmu, jež posuzovala náměty a scénáře. Dramaturgii a produkci však usměr-
ňovaly i vyšší nadřízené orgány — Ústřední dramaturgie, Filmová rada nebo instituce
zodpovědné za oblast výchovy, tedy Výzkumný ústav pedagogický či ministerstva školství
a informací, která se taktéž mohla vyjadřovat ke scénářům i hotovým snímkům. Proto ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) ILUMINACE Ročník 32, 2020, č. 4 (120) 11 bude třeba brát v úvahu i postoje těchto činitelů, kteří obrat v dramaturgii zařídili jako
„mávnutím kouzelného proutku“. V praxi se však přechod na tvorbu pro děti poněkud
komplikoval. Tvůrci se de facto „za pochodu“ museli vypořádat s několika zásadními kon-
cepčními i praktickými úkoly, jejichž vyřešení podmiňovalo vznik úspěšných děl nové
dramaturgie. Zpráva o první celostátní konferenci Československého státního filmu, Praha, listopad 1948. NFA, f. ÚŘ
ČSF, k. R10/AII/2P/9K.ii A od teď už jen pro děti…
Úkoly nové dramaturgie animované tvorby Požadavek natáčet fi lmy pro děti skutečně přišel „shora“, a vcelku náhle, a tvůrce zastihl
povětšinou nepřipravené. Na první celopodnikové konferenci Československého státního
fi lmu, která se konala v listopadu 1948, se řešil také stav animované tvorby a krušné chví-
le tu zažívali především autoři kreslených fi lmů. Jejich snímky — jakkoli technicky vyspě-
lé — prý zatím nenašly správná témata a nechávaly publikum chladným a lhostejným
v porovnání „s životností a živými reakcemi diváků na lidské loutkové fi lmy Karla Zema-
na a Hermíny Týrlové“. Na stejné konferenci si animátoři zároveň vyslechli úkoly pro ná-
sledující období — natáčet fi lmy pro děti a vydat se směrem, jaký vytyčil Zeman se sérií
agitek o Panu Prokoukovi.5) Kritické připomínky se začaly množit také v tisku — podle
těchto ohlasů se především kreslená animace ocitla na scestí a fi lmy jako Šváb, O milio-
náři, který ukradl slunce nebo Vzducholoď a láska trpěly nesrozumitelností a za-
ostáváním obsahu za formou.6) Od této kritiky se odvíjela brzká reorganizace dramaturgie studií animované tvorby. Koncem roku 1949 vznikl kolektiv dětského, kresleného a loutkového fi lmu pod vedením
Jiřího Trnky, fungující jako součást výroby uměleckých fi lmů. Svou činnost zahájily také
Ústřední dramaturgie kresleného a loutkového fi lmu a scenáristické oddělení kresleného
fi lmu. Vzápětí se uskutečnila sebekritická diskuze o dosavadních výsledcích tvorby a byl
vypracován nový tematický plán se stěžejním cílem ideové převýchovy dětí a výchovy ob-
čanů nového socialistického typu.7)i Změna dramaturgie byla ofi ciálně zdůvodňována tím, že animovaná forma má nejblíž
právě dětem a že jde o nejlepší prostředek, jak je umělecky oslovit a vychovávat. K pro-
měně však existovaly i praktičtější důvody — podle vedoucího Studia kresleného fi lmu
Eduar da Hofmana se jednalo také o reakci na permanentní nedostatek fi lmových progra-
mů pro děti v distribuci.8) V Československu zatím nefungovala soustavná produkce těch-
to fi lmů a nikterak rozsáhlá nebyla ani zahraniční nabídka sestávající hlavně z titulů sovět-
ských. Studia animovaných fi lmů v Praze, Zlíně a Brně se tak přeorientovala primárně na Lukáš Skupa: Která bude ta pravá? 12 12 tvorbu pro děti, natáčet však měla i snímky s aktuální tematikou (politické satiry a agitky)
a populárně-instrukční fi lmy pro mládež.9) i
Nejzákladnější z úkolů, s nímž se musela nová dramaturgie na přelomu 40. a 50. let vy-
pořádat, spočíval v zajištění autorů vhodných námětů a scénářů. 9) Plnění úkolů vyplývajících z usnesení předsednictva ÚV KSČ, Praha, 9. 10. 1950. NFA, f. ÚŘ ČSF, k. R5/AII/
2P/6K.
10) Zápis ze schůze Ústřední dramaturgie konané ve čtvrtek dne 8. prosince 1949. NFA, f. ÚŘ ČSF, k. R14/BI/
1P/3K.
11) Srov. např. dopis od Ericha Švabíka pro Vladimíra Václavíka, Praha, 3. 11. 1948. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/4K.
12) Záznam o 9. schůzi Kolektivního vedení dětského, kresleného a loutkového filmu, Praha, 11. 12. 1953. NFA,
f. ÚŘ ČSF, k. R5/AII/2P/7K.
13) Zápis ze schůze Ústřední dramaturgie konané ve čtvrtek dne 8. prosince 1949. NFA, f. ÚŘ ČSF, k. R14/BI/
1P/3K.
14) Zápis o 1. schůzi Ústřední dramaturgie, Praha, 14. ledna 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
15) Zápis ze schůze ÚD, Praha, 9. 9. 1949; Zápis ze schůze Ústřední dramaturgie, Praha, 23. 9. 1949. NFA, f. ÚŘ
ČSF, k. R13/AII/2P/5K.
16) Zápis ze schůze Ústřední dramaturgie konané ve čtvrtek dne 8. prosince 1949. NFA, f. ÚŘ ČSF, k. R14/BI/
1P/3K.
17) Výtah ze zápisu o 17. schůzi scenáristického oddělení kresleného filmu, Praha, 2. 5. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K. 9) Plnění úkolů vyplývajících z usnesení předsednictva ÚV KSČ, Praha, 9. 10. 1950. NFA, f. ÚŘ ČSF, k. R5/AII/
2P/6K. )
p
g
15) Zápis ze schůze ÚD, Praha, 9. 9. 1949; Zápis ze schůze Ústřední dramaturgie, Praha, 23. 9. 1949. NFA, f. ÚŘ
ČSF, k. R13/AII/2P/5K. ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) mitelně, nařídila ho přepracovat a 70 metrů natočeného materiálu vyhodit. V rozhodnutí
ukončit Mannovu režijní kariéru navíc sehrály roli i jeho předešlé „nezdary“ — Basa tvr-
dí muziku, Šváb a Ivánku náš.18) Tváří v tvář novým úkolům se však měli polepšit i ně-
kteří z nejoslavovanějších. Jiří Trnka měl své poetické fi lmy učinit přístupnější publiku19)
a Hermíně Týrlové bylo vyčítáno, že „dělá formálně stále totéž“ a že se nemůže pořád
schovávat do „ideového bezvětří“.20) Oběma byly ve sledovaném období některé projekty
podsouvány, zatímco jiných se museli vzdát.21) mitelně, nařídila ho přepracovat a 70 metrů natočeného materiálu vyhodit. V rozhodnutí
ukončit Mannovu režijní kariéru navíc sehrály roli i jeho předešlé „nezdary“ — Basa tvr-
dí muziku, Šváb a Ivánku náš.18) Tváří v tvář novým úkolům se však měli polepšit i ně-
kteří z nejoslavovanějších. Jiří Trnka měl své poetické fi lmy učinit přístupnější publiku19)
a Hermíně Týrlové bylo vyčítáno, že „dělá formálně stále totéž“ a že se nemůže pořád
schovávat do „ideového bezvětří“.20) Oběma byly ve sledovaném období některé projekty
podsouvány, zatímco jiných se museli vzdát.21) Stejně tak někteří z výtvarníků vyhovovali nárokům dramaturgie více, jiní méně. Ilus-
trace knih pro děti, odkud se často rekrutovali výtvarníci animovaných fi lmů, po roce
1948 obecně směřovaly k popisnosti svázané socialistickým realismem.22) Jakákoli výraz-
nější stylizace reality narážela na kritiku, což se dotklo například Zdeňka Seydla, který se
jako výtvarník uplatnil i na zmiňovaných fi lmech Šváb a Basa tvrdí muziku. Ondřej Se-
kora, pasovaný po roce 1948 do pozice vzorového autora dětské knihy a ilustrace, podro-
bil zdrcující kritice Seydlovo výtvarné provedení Říkadel Josefa Kainara, vybočující ze zá-
vazného realistického kánonu: „[…] obrázek za obrázkem v knize působí jako rána pěstí. Jsou tu nestvůrné huby s odpornými detaily. Postavy se zvráceným požitkem znetvořené. Úplný výsměch dětské touze po poznání. Radost z hrubého vyjadřování. […] je vidno, jak
je Seydl svou duší ne na hony, ale na nekonečnost vzdálen dětskému světu.“23)ii V roce 1950 fi gurovali v seznamu výtvarníků animovaných fi lmů Jiří Trnka, Kamil
Lhoták, Zdeněk Seydl, František Freiwillig, Jaroslav Kándl, Zdeněk Miler, Karel Štrebl,
Ondřej Sekora, Josef Lada, Václav Sivko, Adolf Zábranský nebo Václav Zykmund.24) V pra-
xi se však tato sestava výrazně zúžila, a to nejen z důvodů výše naznačených. Seydl a Lho-
ták nenašli v tvorbě pro děti uplatnění a ke spolupráci s animovaným fi lmem se vrátili až
koncem 50. 21) Trnkův nerealizovaný projekt Děvčátko se sirkami byl označen za „zpozdilou andersenovštinu“, „příliš ma-
loměšťácký a nicneříkající“. Zápis ze schůze Ústřední dramaturgie, Praha, 6. 5. 1949. NFA, f. ÚŘ ČSF, k. R13/
AII/2P/5K. Jiný autorův námět Mořská panenka a chlapeček se zdál „silně abstraktní“, „nesrozumitelný pro
děti“ a neměl vzniknout už proto, aby se Trnkův talent využil pro důležitější úkoly. Zápis ze schůze Ústřed-
ní dramaturgie, Praha, 22. 4. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. Týrlové byla vnucována realizace
zakázkové agitky Tři fáze pana Mráze, aby mohla dál pokračovat v natáčení filmů pro děti. Srov. Zápis
o 14. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 16. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K. Přestože se Týrlová tomuto úkolu nakonec vyhnula, později se zřejmě právě s ohledem na nutnost
„zvyšování ideovosti“ chopila projektu Pohádka o drakovi. 26) Zápis schůze Ústřední dramaturgie, Praha, 23. 9. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. p
25) Zápis ze schůze Ústřední dramaturgie, Praha, 29. 4. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. p
y
ýi
19) Zápis o 1. schůzi Ústřední dramaturgie, Praha, 14. 1. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.i 20) Zápis o 15. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 23. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K. 18) Překročené rozpočty kreslených filmů, Praha, 14. 11. 1950. NFA, f. ÚŘ ČSF, k. R11/BII/5P/9K. y
p
p
j
22) Pavel Ryška – Jan Šrámek, Pionýři a roboti. Československá ilustrace a vizuální kultura 1950–1970. Praha –
Brno: Paseka – Fakulta výtvarných umění VUT 2016, s. 14–15. ý
ý
23) Ondřej Sekora, Nad několika ilustrovanými knihami. Štěpnice 2, 1949, č. 6–7, s. 95. j
ý
p
24) Seznam tvůrčích pracovníků, Praha, 1. 3. 1950. NFA, f. ÚŘ ČSF, k. R11/BI/5P/7K. )
p
y
ýi
,
,
,
,
19) Zápis o 1. schůzi Ústřední dramaturgie, Praha, 14. 1. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
20) Zápis o 15. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 23. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K A od teď už jen pro děti…
Úkoly nové dramaturgie animované tvorby Především ty ke kresle-
ným fi lmům nebyly podle názoru Ústřední dramaturgie dobře napsané a uchylovaly se
více k lyrickému vyprávění.10) Studiím chyběli kvalitní scenáristé i látky,11) což měla napra-
vit spolupráce s předními domácími spisovateli dětské literatury. Těm se však spolupraco-
vat příliš nechtělo už proto, že fi lm vnímali jako riziko velké práce bez jistoty odpovídají-
cího honoráře a realizace příslušné látky. Když už námět či scénář schválila studiová
dramaturgie, nemusel se zamlouvat členům Ústřední dramaturgie nebo Filmové rady,
čímž se další vývoj projektu komplikoval nebo úplně zastavil. Tento problém se nedařilo
dlouhodoběji uspokojivě vyřešit. Jak konstatoval Jiří Trnka v prosinci 1953, dramaturgie
sice hledá aktuální a umělecky hodnotné náměty, ale výsledky nebudou lepší, dokud tyto
náměty nezačnou psát autoři dětské literatury.12) Druhý velký úkol se týkal nalezení správné tematiky a způsobu vyprávění. Už v roce
1949 vznikly převážně podle původních námětů první „novodobé“ animované fi lmy pro
děti jako Lenora, A B C nebo Ivánku náš. Členové Ústřední dramaturgie je ovšem zhod-
notili jako samoúčelné,13) nevhodné nebo nesrozumitelné.14) Vyčítali jim nedostatek
ideovosti a odvahy, lidskosti či humoru, ale třeba i absenci komentáře nebo dialogu.15) Jiří
Trnka na adresu těchto fi lmů kriticky poznamenal, že „za poslední dva roky se dělaly jen
samé legrácky“ nebo že předkládané náměty jsou stále „viditelné kopie amerických grote-
sek“.16) V rámci interních hodnocení vzali zástupci scenáristického oddělení kresleného
fi lmu z dřívější tvorby „na milost“ pouze snímek Zasadil dědek řepu. Ačkoli i jemu vy-
tkli ideovou nevyváženost a formalistický závěr, ocenili na něm českost, zemitost a osobi-
tost.17) S otázkou, o čem a jak by měly fi lmy pro děti vyprávět, souviselo i angažmá konkrét-
ních režisérů a výtvarníků. Například Karel Mann byl po dokončení snímku Závistivá
zvířátka přeřazen z pozice režiséra do jiných funkcí (později působil jako kreslíř nebo
zvukař). V průběhu realizace Ústřední dramaturgie KF usoudila, že fi lm vyznívá nesrozu- 14) Zápis o 1. schůzi Ústřední dramaturgie, Praha, 14. ledna 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. p
g
15) Zápis ze schůze ÚD, Praha, 9. 9. 1949; Zápis ze schůze Ústřední dramaturgie, Praha, 23. 9. 1949. NFA, f. ÚŘ
ČSF, k. R13/AII/2P/5K. 16) Zápis ze schůze Ústřední dramaturgie konané ve čtvrtek dne 8. prosince 1949. NFA, f. ÚŘ ČSF, k. R14/BI/
1P/3K. 17) Výtah ze zápisu o 17. schůzi scenáristického oddělení kresleného filmu, Praha, 2. 5. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K. ČLÁNKY K TÉMATU
13 18) Překročené rozpočty kreslených filmů, Praha, 14. 11. 1950. NFA, f. ÚŘ ČSF, k. R11/BII/5P/9K.
19) Zápis o 1. schůzi Ústřední dramaturgie, Praha, 14. 1. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
20) Zápis o 15. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 23. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/ 27) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 22. 1. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K.
28) Usnesení ze dne 5. 6. 1950 a ze dne 12. 6. 1950, Praha, 1. 10. 1950. NFA, f. ÚŘ ČSF, k. R18/AI/6P/2K.
29) Vystihuje to například shrnutí diskuze po projekcích sovětského kombinovaného filmu Zlatý klíček a Pi-
nocchia od Walta Disneyho. Zatímco sovětský snímek byl v diskuzi vyhodnocen jako „lidský a pravdivý“,
zdůrazňující skutečnost i v loutkových pasážích, Disneyho film „odlidšťuje i reálnou skutečnost“. Zápis
o 15. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 23. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K.
30) Úkoly vyplývající z usnesení předsednictva ÚV KSČ pro oblast animované tvorby dokonce přímo obsahují
pokyn promítat si na schůzích sovětské filmy a učit se z nich. Srov. Plnění úkolů vyplývajících z usnesení
předsednictva ÚV KSČ, Praha, 9. 10. 1950. NFA, f. ÚŘ ČSF, k. R5/AII/2P/6K. 30) Úkoly vyplývající z usnesení předsednictva ÚV KSČ pro oblast animované tvorby dokonce přímo obsahují
pokyn promítat si na schůzích sovětské filmy a učit se z nich. Srov. Plnění úkolů vyplývajících z usnesení
předsednictva ÚV KSČ, Praha, 9. 10. 1950. NFA, f. ÚŘ ČSF, k. R5/AII/2P/6K. 27) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 22. 1. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K.
ÚŘ Č 36) Zápis konference výrobních složek Čs. státního filmu a poradního sboru pro výrobu při 21. Svazu ROH,
Praha, 22. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/2P/9K. Navýšení rozpočtu snímku Hrnečku, vař! tak bylo
vykládáno nikoli jako pochybení, ale jako „nová cesta při prohlubování uměleckého mistrovství v linii so-
cialistického realismu podle vzoru kreslených sovětských filmů. Realitě pohybu a bohatosti obrazu je věno-
vána větší péče, než tomu bylo dosud“. Definitivní zvýšený rozpočet barevného kresleného filmu Hrnečku,
vař!, Praha, 1. 11. 1952. NFA, f. ÚŘ ČSF, k. R12/AI/4P/2K. p
g
35) Kolektivní vedení Ústřední dramaturgie například doporučilo brněnskému studiu, aby se při přípravě lout-
kového snímku O medvídkovi a zajíčkovi poučilo u sovětského filmu Šedé pírko, dodržujícího „ve zvířecí
fabuli přísně všechny zákony přírody“. Scénosled loutkového filmu O medvídkovi a zajíčkovi, Praha, 20. 6.
1950. NFA, f. ÚŘ ČSF, k. R14/BII/2P/5K. 32) Zápis o 4. schůzi Ústřední dramaturgie, Praha, 4. 2. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. 31) Vývoj kresleného a loutkového filmu od resoluce předsednictva ÚV KSČ z dubna 1950 do konce roku 1953.
NFA, f. ÚŘ ČSF, k. R19/AI/5P/1K. 33) Zápis ze schůze Ústřední dramaturgie, Praha, 29. 4. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K. 31) Vývoj kresleného a loutkového filmu od resoluce předsednictva ÚV KSČ z dubna 1950 do konce roku 1953.
NFA, f. ÚŘ ČSF, k. R19/AI/5P/1K.
32) Zápis o 4. schůzi Ústřední dramaturgie, Praha, 4. 2. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
33) Zápis ze schůze Ústřední dramaturgie, Praha, 29. 4. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
34) Zápis ze schůze Ústřední dramaturgie, Praha, 11. 2. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
35) Kolektivní vedení Ústřední dramaturgie například doporučilo brněnskému studiu, aby se při přípravě lout-
kového snímku O medvídkovi a zajíčkovi poučilo u sovětského filmu Šedé pírko, dodržujícího „ve zvířecí
fabuli přísně všechny zákony přírody“. Scénosled loutkového filmu O medvídkovi a zajíčkovi, Praha, 20. 6.
1950. NFA, f. ÚŘ ČSF, k. R14/BII/2P/5K.
36) Zápis konference výrobních složek Čs. státního filmu a poradního sboru pro výrobu při 21. Svazu ROH,
Praha, 22. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/2P/9K. Navýšení rozpočtu snímku Hrnečku, vař! tak bylo
vykládáno nikoli jako pochybení, ale jako „nová cesta při prohlubování uměleckého mistrovství v linii so-
cialistického realismu podle vzoru kreslených sovětských filmů. Realitě pohybu a bohatosti obrazu je věno-
vána větší péče, než tomu bylo dosud“. Definitivní zvýšený rozpočet barevného kresleného filmu Hrnečku,
vař!, Praha, 1. 11. 1952. NFA, f. ÚŘ ČSF, k. R12/AI/4P/2K. ČLÁNKY K TÉMATU let. Kándl se ve sledovaném období podílel pouze na jediném fi lmu Kombajn,
což navíc u členů Ústřední dramaturgie vzbuzovalo obavy ze schematičnosti a inklinace
ke stylu americké grotesky.25) Zykmund působil v brněnském studiu loutkového fi lmu,
které se zaměřovalo na publikum nejmladšího věku, avšak podle hodnocení dramaturgie
nebyl výtvarníkem pro děti.26) Neuspokojivá výtvarná stránka jeho počinů, označovaných Lukáš Skupa: Která bude ta pravá? 14 Obr. 1 — Výtvarník František
Freiwillig (vlevo) a režisér Eduard
Hofman nad výtvarnými návrhy
pro snímek Jablůňka se zlatými
jablky (reprint z magazínu
Czechoslovak Film 5, 1952, č. 11,
s. 5) Obr. 1 — Výtvarník František
Freiwillig (vlevo) a režisér Eduard
Hofman nad výtvarnými návrhy
pro snímek Jablůňka se zlatými
jablky (reprint z magazínu
Czechoslovak Film 5, 1952, č. 11,
s. 5) Obr. 1 — Výtvarník František
Freiwillig (vlevo) a režisér Eduard
Hofman nad výtvarnými návrhy
pro snímek Jablůňka se zlatými
jablky (reprint z magazínu
Czechoslovak Film 5, 1952, č. 11,
s. 5) jako „neradostné a nevkusné“ nebo „dětská demagogie“, dokonce přispěla k rozpuštění
brněnské skupiny v roce 1952.27) Dramaturgie naopak chtěla spolupracovat se Zábranským, Sekorou a Ladou, kteří se
však k fi lmové tvorbě dostávali spíš výjimečně kvůli vlastní časové vytíženosti. Sekoru lá-
kali i do Kolektivního vedení kresleného a loutkového fi lmu, což však odmítl pro svou za-
neprázdněnost ve Státním nakladatelství dětské knihy, kde působil v letech 1949–1952
jako vedoucí Knižnice pro nejmenší.28) Z hlediska počtu realizovaných snímků i z hledis-
ka charakteru jejich výtvarného stylu se tak z celé skupiny nejvíce osvědčili Miler a Frei-
willig.i Třetí klíčový úkol nové dramaturgie lze shrnout jako hledání národní specifi čnosti. Hlavně v případě kreslených fi lmů se opakovaně navracel požadavek „národní osobitosti“,
českého nebo i slovanského stylu animace. O přesnou defi nici takového stylu se zřejmě ni-
kdo nepokusil, ale o jeho náplni leccos prozradí neustálé vymezování se vůči „konkuren-
ci“. Členové dramaturgie si pravidelně promítali zahraniční animované fi lmy, přičemž
z následných diskuzí většinou vyplynul kontrast sovětské a americké animace.29) Zatímco
k sovětským fi lmům měli domácí animátoři vzhlížet jako ke vzoru,30) inspirace americkou ILUMINACE Ročník 32, 2020, č. 4 (120) ČLÁNKY K TÉMATU animací spojovanou s Waltem Disneym byla nežádoucí. ČLÁNKY K TÉMATU Podle interního zhodnocení vý-
voje animace v letech 1950 až 1953 dosáhl domácí kreslený fi lm vynikajících výsledků
proto, že se odpoutal od vlivů „západního, bezideového a standardizovaného disneyov-
ského fi lmu“ a poučil se u fi lmu sovětského.31) animací spojovanou s Waltem Disneym byla nežádoucí. Podle interního zhodnocení vý-
voje animace v letech 1950 až 1953 dosáhl domácí kreslený fi lm vynikajících výsledků
proto, že se odpoutal od vlivů „západního, bezideového a standardizovaného disneyov-
ského fi lmu“ a poučil se u fi lmu sovětského.31) ii
Přestože stopy „národního stylu“ směřovaly jednoznačně do Sovětského svazu, vliv
Disneyho silně zakořenil nejen ve vědomí tvůrců, ale i těch, kteří zasedali ve schvalovacích
sborech. Dokládají to třeba útržky z porady Ústřední dramaturgie nad nerealizovaným
námětem Jak se liška vytančila: Z filmu bude americká groteska. Má to disneyovský švih. Musíme najít švih český. Z filmu bude americká groteska. Má to disneyovský švih. Musíme najít švih český. Můžeme sice diváka ošidit českou hudbou, ale to nechceme. […] Chceme kreslený
film počeštit. Bojujeme o to, ale obecenstvo chce disneyovky. Myslím, že je povin-
nost dramaturgie podpořit český humor, český typ. Chceme lidi vychovávat. […]
Neumím si představit českého pidimužíka. Je to čistý Disneyův film. Mám dojem, že
neumíme hledat cestu.32) Komise by svolila s natáčením, jedině pokud by se našel výtvarník, který by snímek
„zčeštil“. Český styl se tak prolínal i do výtvarného zpracování, o němž však dochované
materiály neprozrazují zdaleka tolik jako o literárních přípravách. Jde spíš jen o zmínky
typu „udržet to na české základně“33) nebo „pozor na typy, aby byly české“34) bez přesnější
specifi kace. Autoři byli nicméně vybízeni k realismu a pravdivosti animace prostředí a fi -
gur, v čemž jako vzor fungovaly právě sovětské fi lmy.35) Příklad Jablůňky se zlatými ja-
blky později ukáže, že hledání národní specifi čnosti se překrývalo s cestou k realistické-
mu animovanému fi lmu, která podle Hofmana měla vést od realistické myšlenky přes
realistický pohyb až po prohloubení veškeré práce a zpomalení výrobního tempa.36) Lukáš Skupa: Která bude ta pravá? 16 40) Výtah ze zápisu o 2. schůzi scenáristického oddělení, Praha, 17. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K.
V animovaných pohádkách ve sledovaných letech královské prostředí spíš absentuje. Pokud se v některém
námětu vyskytlo, zpravidla se řešilo jeho vykreslení — třeba u nerealizovaného projektu Rozesmějte princez-
nu bylo dle námitky království „líčeno tak, jako by bylo, proč po něm toužit“. Výtah ze zápisu o 18. schůzi
scenáristického oddělení, Praha, 15. 11. 1950. NFA, f. ÚŘ ČSF, k. R5/BI/1P/4K. 37) Více k diskuzím o poválečné literatuře pro děti a mládež viz Pavel Janoušek a kol., Dějiny české literatury
1945–1989, I. 1945–1948. Praha: Academia 2007, s. 321–330.
38) Literární věda používá pro tuto kategorii pojmy „parodická pohádka“ nebo „antipohádka“ a zároveň ji na-
hlíží jako podtyp pohádky autorské (či moderní), kterou však filmová dramaturgie nahlížela jako samostat-
nou kategorii. Srov. Dagmar Mocná – Josef Peterka a kol., Encyklopedie literárních žánrů. Praha: Paseka
2004, s. 472–474.
39) Srov. např. Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k.
R14/BI/2P/3K.
40) Výtah ze zápisu o 2. schůzi scenáristického oddělení, Praha, 17. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K.
V animovaných pohádkách ve sledovaných letech královské prostředí spíš absentuje. Pokud se v některém
námětu vyskytlo, zpravidla se řešilo jeho vykreslení — třeba u nerealizovaného projektu Rozesmějte princez-
nu bylo dle námitky království „líčeno tak, jako by bylo, proč po něm toužit“. Výtah ze zápisu o 18. schůzi
scenáristického oddělení, Praha, 15. 11. 1950. NFA, f. ÚŘ ČSF, k. R5/BI/1P/4K. 39) Srov. např. Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k.
R14/BI/2P/3K. 38) Literární věda používá pro tuto kategorii pojmy „parodická pohádka“ nebo „antipohádka“ a zároveň ji na-
hlíží jako podtyp pohádky autorské (či moderní), kterou však filmová dramaturgie nahlížela jako samostat-
nou kategorii. Srov. Dagmar Mocná – Josef Peterka a kol., Encyklopedie literárních žánrů. Praha: Paseka
2004, s. 472–474.i j
p
yp p
yi
g
j
nou kategorii. Srov. Dagmar Mocná – Josef Peterka a kol., Encyklopedie literárních žánrů. Praha: Paseka
2004, s. 472–474.
39) Srov. např. Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k.
R14/BI/2P/3K.
)
h
h
k h
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h
f ÚŘ Č
k 41) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/3K.
42) Schůze Ústřední dramaturgie krátkometrážního úseku, Praha, 23. 6. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/
2P/5K.
43) Zápis o 18. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 21. 5. 1951. NFA, f. ÚŘ ČSF, k.R9/AI/
4P/8K.
44) Zápis ze schůze Ústřední dramaturgie, Praha, 22. 4. 1949. NFA, f. ÚŘ ČSF, k. R13/AII/2P/5K.
45) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 29. 1. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K.
46) Výtah ze zápisu o 12. schůzi scenáristického oddělení kresleného filmu, Praha, 27. 3. 1951. NFA, f. ÚŘ ČSF,
k. R11/BII/5P/9K. Základní typologie:
pohádka klasická, přehodnocená a moderní Z předchozích odstavců vyplývá, že řízený rozvoj animovaného fi lmu pro děti neurčovaly
ani tak přesné defi nice požadavků na tento žánr, jako spíš některá vágní doporučení a pře-
devším formulace „co je špatně“ a „jak by to nemělo vypadat“. I proto nijak nepřekvapí, že
dramaturgie se při svém hledání správné koncepce fi lmu pro děti uchýlila k domácí litera-
tuře, v níž mohla čerpat z dlouhodobějších zkušeností, hlubších tradic i jasných aktuál-
ních postojů. Literatura pro děti a mládež se těsně po roce 1945 stala klíčovou složkou
tzv. výchovy uměním a ocitla se pod drobnohledem státních a společenských institucí. Stě-
žejním tématem bylo zamítnutí komerční, zábavně orientované literatury, spojované s mi-
nulostí, která měla jako nežádoucí „škvár“ zmizet z veřejných knihoven i domácností.37) j
j ý
Ideální možnost zvládnutí všech dříve popsaných úkolů spatřovala dramaturgie v lite-
rárně osvědčeném žánru pohádky. Po roce 1945 ho formovaly publikace výborů klasic-
kých pohádek Boženy Němcové nebo Karla Jaromíra Erbena, vydání pohádek nového
typu, jež uzpůsobovaly tradiční pohádkové postavy a motivy nové společensko-politické
situaci a vsazovaly je do realistického rámce, a také autorské pohádky, které se od tradic
chtěly zcela odtrhnout a sloužit aktuálním potřebám kulturní politiky. Všechny tyto dru-
hy se dostávaly do hledáčku dramaturgie animované tvorby, jež ke každému z nich zaují-
mala odlišné stanovisko. O žánru přitom uvažovala ve třech kategoriích, téměř souhlas-
ných s uvedenými literárními variantami: klasická (národní) pohádka, přehodnocená
pohádka (jinak též upravená pohádka nebo pohádka naruby)38) a moderní (autorská) po-
hádka.39) Postoj ke klasickým pohádkám, jež tvoří nejpočetnější skupinu tehdy realizovaných
fi lmů pro děti, se řešil napříč uměleckými obory. V roce 1952 uspořádalo Státní naklada-
telství dětské knihy poradu, na které se debatovalo o výběru pohádek Němcové a Erbena
využitelných pro jakoukoli formu umění pro mládež. Na řadu přišlo mimo jiné i téma,
nakolik děti zásobovat pohádkami s nadpřirozenými bytostmi (například čerty) nebo po-
hádkami s hodnými králi. V této záležitosti převládlo mínění, že pokaždé záleží na
charakteru a funkci dané postavy v pohádkovém vyprávění.40) Nikoli náhodou se i drama-
turgie animovaného fi lmu obracela nejčastěji k předlohám Erbena a Němcové — z reali-
zovaných to byly například Bajaja, Hrnečku, vař!, O kohoutkovi a slepičce nebo ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) Čert a Káča, z nerealizovaných Tři zlaté vlasy děda Vševěda, Chytrá horákyně nebo Jak
Jaromil ke štěstí přišel. Základní typologie:
pohádka klasická, přehodnocená a moderní p
Z kánonu klasických pohádek dostávaly přednost ty, které pasovaly na teze z tematic-
kých plánů: úcta k hrdinovi a hrdinství, nenávist k „človíčkovství“ (k pouhé existenci bez
velkého cíle), láska k práci ve všech podobách, nenávist k lenochům, kladný vztah ke ko-
lektivu, nenávist k sobectví, soucit s trpícími (zvláště těmi, kteří trpí v kapitalistickém sys-
tému) či nenávist ke kapitalismu.41) Výčet z roku 1952 sám o sobě napovídá, že zastřešují-
cím kritériem výběru byl potenciál „starých pohádek“ promlouvat k současnosti. Například Trnka musel Ústřední dramaturgii vysvětlovat, proč si od Němcové, jež napsa-
la i jiné pohádky s aktuálnějším vyzněním, vybral právě Bajaju, který je nevhodný pro
děti, založený na „zastřené mythice oidipovského komplexu [… a] odtažitý od toho,
k čemu chceme lidi dostat“.42) Podobné námitky vyvstaly i během diskuzí o adaptacích ji-
ných pohádek Němcové — Silný Ctibor43) a O dvanácti měsíčkách,44) z nichž druhá zmíně-
ná nakonec vznikla v roce 1960. Obr. 2–3 — Problém aktualizace pohádky Hrnečku, vař!: statkářští manželé měli podle dramaturgie připomínat
nikoli „kulaky“, ale „nenažrané sobce“ Obr. 2–3 — Problém aktualizace pohádky Hrnečku, vař!: statkářští manželé měli podle dramaturgie připomínat
nikoli „kulaky“, ale „nenažrané sobce“ Zatímco aktuálnost jakožto přidanou hodnotu klasických pohádek dramaturgie vítala,
k aktualizaci přistupovala velmi citlivě až rezervovaně — skutečně zásadním revizím pů-
vodního vyprávění z pozice „nové doby“ se spíš bránila. Například aktuální společenské
zasazení Erbenovy pohádky Hrnečku, vař! zpochybňovala jako schematické s tím, že
snímek by měl zpodobňovat obecné lidské typy a nevzbuzovat analogie se sociální proble-
matikou tehdejšího venkova.45) Konkrétně šlo o postavy statkářských manželů, kteří titul-
ní hrdince ukradli kouzelný krajáček a kteří měli být vykresleni jednoduše jako „žrouti“.46) Lukáš Skupa: Která bude ta pravá? 18 K tomuto motivu se členové dramaturgie vraceli opakovaně a dobrali se jednohlasného
stanoviska, že v protikladu zde proti sobě stojí nikoli „kulak a vykořisťovaný podruh“, ale
„nenažraný sobec a dobrý člověk“.47) I přes snahy korigovat a otupit aktuálně vyznívající
reference (tematický plán přiblížil poslání snímku jako „učit děti nenávidět sobectví a ne-
nažranost a budit v nich víru v sílu účinné lásky člověka k člověku“48)) výsledný fi lm k ta-
kovému čtení silně svádí, což ostatně stvrzují i ohlasy v dobovém tisku.49) Klasické pohádky však do značné míry sloužily „jen“ jako východisko z nouze. Před-
nostně měly vznikat fi lmy svázané s aktuálními hospodářskými, politickými a sociálními
problémy,50) avšak použitelných látek tohoto typu bylo poskrovnu. 47) Zápis o 12. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 2. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K. Ke stejné záležitosti vyjádřili totožný názor také „pracovníci ve výchově mládeže uměním“. Srov. Vý-
tah ze zápisu o schůzi scenáristického oddělení, Praha, 17. 4. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K. 47) Zápis o 12. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 2. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K. Ke stejné záležitosti vyjádřili totožný názor také „pracovníci ve výchově mládeže uměním“. Srov. Vý-
tah ze zápisu o schůzi scenáristického oddělení, Praha, 17. 4. 1952. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K.
48) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/3K.
49) Např. Pohádka „Hrnečku, vař!“ v kresleném filmu. Filmové informace 2, 1951, č. 38, s. 17.
50) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 11. 12. 1950. NFA, f. ÚŘ ČSF,
k. R5/BI/1P/4K.
51) K představitelům této žánrové varianty patřil i Josef Lada se svými sbírkami Pohádky naruby (1939) nebo
Nezbedné pohádky (1946). Více o vývoji pohádky po roce 1945 viz P. Janoušek a kol., c. d., s. 332–335.
52) Výtah o 23. schůzi scenáristického oddělení kresleného filmu, Praha, 20. 6. 1951. NFA, f. ÚŘ ČSF, k. R11/
BII/5P/9K.
53) Výtah ze zápisu o 22. schůzi scenáristického oddělení kresleného filmu, Praha, 14. 5. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K.
54) Výtah ze zápisu o 15. schůzi scenáristického oddělení kresleného filmu, Praha, 11. 4. 1951. NFA, f. ÚŘ ČSF,
k. R11/BII/5P/9K. 54) Výtah ze zápisu o 15. schůzi scenáristického oddělení kresleného filmu, Praha, 11. 4. 1951. NFA, f. ÚŘ ČSF,
k. R11/BII/5P/9K. p
y
ý
j p
y p
52) Výtah o 23. schůzi scenáristického oddělení kresleného filmu, Praha, 20. 6. 1951. NFA, f. ÚŘ ČSF, k. R11/
BII/5P/9K. 53) Výtah ze zápisu o 22. schůzi scenáristického oddělení kresleného filmu, Praha, 14. 5. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K.i p
48) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/3K. i
50) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 11. 12. 1950. NFA, f. ÚŘ ČSF,
k. R5/BI/1P/4K. k. R5/BI/1P/4K.
51) K představitelům této žánrové varianty patřil i Josef Lada se svými sbírkami Pohádky naruby (1939) nebo
Nezbedné pohádky (1946). Více o vývoji pohádky po roce 1945 viz P. Janoušek a kol., c. d., s. 332–335. ČLÁNKY K TÉMATU ČLÁNKY K TÉMATU 19 ho současníka, který sice stejně miluje prostotu, moudrost a poezii lidového
vyprávění, ale který má jiný poměr k silám, kouzlům a sociálním vztahům světa sta-
rých pohádek […]. Zůstává otázka, zda úmysl autorů zobrazit básnickým příměrem
mírový boj pokrokového lidstva nezjednodušuje naši skutečnost.55) Jediným realizovaným fi lmem tohoto typu je ve sledovaném období Pohádka o dra-
kovi podle předlohy Jiřího Marka. Námět, v němž se po staletích probuzený drak přemě-
ňuje v lokomotivu, podle explikace režisérky Hermíny Týrlové „[…] staví staré pohádky
na hlavu. Přetváří zlý a nebezpečný charakter draka na dobrý a využije jeho síly k užitku
a prospěchu lidí“.56) Výlučnost projektu dokládá i jeho hodnocení před zahájením natáče-
ní — píše se v něm o náročném úkolu z hlediska dramaturgického, výrazového i technic-
kého s tím, že modernizaci tradičních pohádkových motivů se ještě v žádném z umělec-
kých oborů nepodařilo úspěšně zvládnout. Také Filmová rada vynesla při projednávání
scénáře jasné stanovisko k těmto „pohádkám naruby“: buď mají tvůrci točit klasické po-
hádky v původní podobě, nebo se pokusit o pohádky zcela nové — s novou, jim vlastní
motivikou.57) V tomto smyslu našla dramaturgie ideál v moderní autorské pohádce, jež se v literární
oblasti zesíleně prosazovala po založení Státního nakladatelství dětské knihy v roce 1949. Snažila se o umělecko-naučné uchopení dobových událostí, změn či kampaní (brigádnic-
ká činnost, sběr surovin či potírání záškodnictví) a měla přesvědčovat o prospěšnosti pro-
bíhajících společensko-politických změn. K autorům těchto knih, z nichž mnohé vynika-
ly silnou schematičností v duchu budovatelské poetiky, patřili Ondřej Sekora (O zlém
brouku bramborouku), Julie Brožová-Malá (O neústupné hoře), František Čečetka (O vláč-
ku Kolejáčku a jiné pohádky) nebo Bohumila Sílová (Třicet koní Vénu honí).58)i O některých z nich se uvažovalo i pro fi lmovou adaptaci. Například knihu Sílové vy-
právějící o napraveném dělníkovi, který v továrně zanedbával ošetřování své brusky, měla
jako loutkový snímek natočit brněnská skupina. Její vedoucí Václav Zykmund nicméně
předem zapochyboval, že by děti zajímal fi lm v hlavní roli s personifi kovanou bruskou,
a z realizace sešlo.59) Ačkoli ne všechny moderní pohádky byly shledány za vhodné pro
zfi lmování, daly se jimi lépe a snadněji naplňovat stanovené cíle výchovy a převýchovy,
a to i s eventuálním přidáváním dalších potřebných motivů, aniž by v celku vyprávění pů-
sobily neústrojně či rušivě. V porovnání s klasickými pohádkami sem mohla bez zábran
pronikat dramaturgií preferovaná současná tematika. 55) O Lojzkovi Krajánkovi — literární scénář Vl. Kabelíka a H. Blocha, Praha, 25. 6. 1953. NFA, f. ÚŘ ČSF,
k. R19/AI/5P/1K. 56) Pohádka o zkroceném draku. Podle Jiřího Marka upravila H. Týrlová. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K.
57) Rozbor plnění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K.
58) Srov. Pavel Janoušek a kol., Dějiny české literatury 1945–1989, II. 1948–1958. Praha: Academia 2007, s. 438.
59) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 4. 1. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K.
60) Na konferenci zaznělo, že Sekora dokázal v Pohádce o stromech a větru přiblížit nejen souvislost mezi světem
rostlin a počasím, ale i proces přetváření přírody a jeho společenské příčiny. O Pohádce o jablůňce se zlatými Základní typologie:
pohádka klasická, přehodnocená a moderní Někteří autoři se ve
svých námětech pokoušeli přenášet klasické pohádkové motivy do soudobého prostředí,
s čímž však u fi lmové dramaturgie neuspěli. V literatuře tento trend, představující jakýsi
mezistupeň mezi pohádkou klasickou a moderní, zastupoval například Jiří Marek. V roce
1947 publikoval sbírku Veselé pohádky vzhůru nohama, ve které se tradiční pohádkové by-
tosti (čert, drak či víla) ocitají v realitě, v níž se přeměňují a nakonec jí slouží k užitku.51)
K podobným projektům se dramaturgie animovaného fi lmu stavěla striktně negativně se
stále stejnými argumenty. Scenáristické oddělení zamítlo námět Jaroslava Šikla Práce nad zlato pro jeho schema-
tičnost a umělecké i ideové nedostatky, kdy „[…] je na starou pohádku neorganicky na-
roubována sociální problematika v dnešním pohledu.“52) Stejně tak odsoudilo zpracování
Pohádky o Honzovi podle námětu Václava Hrubína, které násilně vsazovalo pohádkového
hrdinu do aktuálních souvislostí.53) Schematický a nepřesvědčivý byl i další nepřijatý ná-
mět Pohádka o dvou zemích Ludvíka Tomana a Karla Friedricha: „Prostředky staré pohád-
ky se tu aktualizují bez vnitřní pravdivosti“.54) Jeden z konkrétních důvodů nelibosti vůči
těmto projektům poodhaluje posudek na scénář Vladimíra Kabelíka a Henryka Blocha
O Lojzkovi Krajánkovi: V přenášení tradičních pohádkových vlastností do příběhu se soudobou tematikou
je nepochybně skryto jisté nebezpečí, protože tento postup zkresluje a ochuzuje pro-
jevy dnešního života a porušuje příčinné souvislosti mezi okruhem představ člově-
ka žijícího na určitém překonaném stupni společenského vývoje a prožitkem naše- ILUMINACE Ročník 32, 2020, č. 4 (120) ČLÁNKY K TÉMATU 6) Pohádka o zkroceném draku. Podle Jiřího Marka upravila H. Týrlová. NFA, f. ÚŘ ČSF, k. R10/AII/4P/2K.
7) Rozbor plnění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K. ČLÁNKY K TÉMATU V animované tvorbě vyvrcholila tendence moderní pohádky snímky Pohádka
o stromech a větru a především Jablůňka se zlatými jablky podle předloh Ondřeje
Sekory a Karla Černého, o nichž se ještě v roce 1955 na celostátní konferenci o literatuře Lukáš Skupa: Která bude ta pravá? 20 pro děti mluvilo ve smyslu příkladného zvládnutí žánru.60) Filmovou Pohádku o stro-
mech a větru, na níž se Sekora podílel také jako výtvarník, označovala dramaturgie již
během realizace za „velký pokrok naší práce v tomto oboru“.61) Posléze byl snímek vyhod-
nocen jako nejlepší kreslený fi lm roku 195162) a režisér Václav Bedřich za něj získal umě-
leckou prémii ministra informací a osvěty.63) Jablůňka se zlatými jablky obdržela kro-
mě ceny na MFF v Karlových Varech64) i uměleckou prémii ministra informací a osvěty
pro režiséra Eduarda Hofmana.65) Oba tituly vyšly také na diafi lmech a jejich scénáře měly
být rozesílány potenciálním autorům námětů jako vzory, kudy by se měla ubírat jejich in-
vence.66) jablky bylo řečeno, že personifikace zvířat a rostlin zde mají hluboký humanistický smysl, v němž autor na-
vázal na klasické dědictví domácí pohádkové tvorby, naplňované novým, socialistickým obsahem. Srov.
Nejlepší síly spisovatelů literatuře pro mládež. Celostátní konference o literatuře pro děti. Literární noviny 4,
1955, č. 43, s. 6.i 61) Zápis o 14. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 16. 4. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K.i 64) Snímek získal cenu s odůvodněním, že „[…] umělecky svěžím a srozumitelným způsobem sděluje i nej-
menším divákům svou zdravou a prospěšnou myšlenku o potřebě a blahodárnosti lidské práce“. Rozbor pl-
nění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K. 62) Zápis konference výrobních složek Čs. státního filmu a poradního sboru pro výrobu při 21. Svazu ROH,
Praha, 22. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AI/2P/9K. p
65) Státní ceny Klementa Gottwalda r. 1956, Praha, 25. 8. 1955. NFA, f. ÚŘ ČSF, k. R10/AI/2P/1K.i ,
,
,
63) Udělení státních cen v oboru filmu a uměleckých prémií za některé filmy, Praha, 10. 5. 1952. NFA, f. ÚŘ ČSF,
k. R11/BI/5P/7K. Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu Po pohádkovém žánru vyžadoval, aby „neplodnou
a škodlivou fantasii primitiva a folklorního člověka“, jež děti odvracela od skutečnosti, na-
hradila „plodná a tvořivá fantasie dnešního pracujícího člověka budujícího socialismus“.69)
Tematický plán prezentoval chystaný fi lm právě skrze tyto rysy, které odpovídaly požado-
vaným vlastnostem, k nimž měla být vychovávána nová mladá generace: láska k přírodě
a práci, radostný a kladný vztah ke kolektivu, pomoc druhým, slabším a nešťastným, chá-
pání práce jako tvůrčí moci vytvářející kladné hodnoty nebo opovrhování lenochy, kteří
uhýbají před společnými úkoly.70) gie v mnoha směrech. V základním ideovém východisku se tento projekt shodoval s náro-
ky na „moderní socialistickou pohádku“, které formuloval například Josef Šlajer v kritické
revue tvorby pro mládež Štěpnice. Po pohádkovém žánru vyžadoval, aby „neplodnou
a škodlivou fantasii primitiva a folklorního člověka“, jež děti odvracela od skutečnosti, na-
hradila „plodná a tvořivá fantasie dnešního pracujícího člověka budujícího socialismus“.69)
Tematický plán prezentoval chystaný fi lm právě skrze tyto rysy, které odpovídaly požado-
vaným vlastnostem, k nimž měla být vychovávána nová mladá generace: láska k přírodě
a práci, radostný a kladný vztah ke kolektivu, pomoc druhým, slabším a nešťastným, chá-
pání práce jako tvůrčí moci vytvářející kladné hodnoty nebo opovrhování lenochy, kteří
uhýbají před společnými úkoly.70) Projekt navíc sliboval příkladné ztvárnění dalších tezí, jež ve výchově a výuce dětí
a mládeže patřily mezi obzvlášť protežované a jež mohla vhodnou formou opět tlumočit
moderní pohádka — slovy Josefa Šlajera odhalit „nesčíslné možnosti dalšího ovládnutí
přírodních sil a živlů pracujícím člověkem, vyzbrojeným všemi prostředky současné tech-
niky“.71) Podle tematického plánu měla i Jablůňka se zlatými jablky vyvolávat v divá-
cích hrdý pocit člověka jako „vládce a aktivního a vědomého přetvořitele přírody“ nebo
přispět k „boji“ proti oživování přírody v mystické, vševládnoucí podobě.72) Zvířátka, která s planou jabloní sama nic nezmůžou, se ze všeho nejdříve vypraví pro
radu k zemi, slunci, měsíci, větru a vodě. Ani personifi kované živly si však s problémem
nedovedou poradit, a tak se zvířátka vrací s nepořízenou. Vzápětí se od datla dozví, že je-
diný, kdo jim poskytne účinnou pomoc, je člověk. V tomto momentě dochází k narušení
stavu „vnitřně soběstačné“ pohádkové fantazie z přírodní říše. V duchu „moderní sociali-
stické pohádky“ do vyprávění zvenčí proniká racionální prvek, který musí zvítězit — pro-
blém vyřeší člověk reálným, praktickým a navíc i poučným postupem. Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu Na rozdíl od adaptací některých klasických pohádek se příprava a realizace Jablůňky se
zlatými jablky obešly bez závažnějších zádrhelů nebo zdlouhavých debat o úpravách. Adaptace námětu Karla Černého byla projednávána od počátku roku 1951 a scenáristicky
ji zajišťoval Josef Menzel — jiný spisovatel dětských knih a tou dobou pravidelný spolu-
pracovník studií animované tvorby. Hned rozšířenou fi lmovou povídku Ústřední drama-
turgie chválila za snahu o využití dialogu, považovaného za nezbytnou součást animova-
né tvorby pro děti, a také za to, že projekt výborně zapadal do „vzestupné tematické linie“
kresleného fi lmu: „Je v tom život a není to schematické rozvedení anekdoty.“67) I na poz-
dějších poradách dramaturgie zaznívaly pochvaly literárnímu scénáři či ukázkám nasní-
mané animace.68) Příběh o lesních zvířátkách, kterým z jadérka vyroste jabloň, jež však začne plodit
chutná jablka až poté, co stromek naroubují pionýři, vyhovoval potřebám nové dramatur- 62) Zápis konference výrobních složek Čs. státního filmu a poradního sboru pro výrobu při 21. Svazu ROH,
Praha, 22. 1. 1952. NFA, f. ÚŘ ČSF, k. R10/AI/2P/9K.ii 63) Udělení státních cen v oboru filmu a uměleckých prémií za některé filmy, Praha, 10. 5. 1952. NFA, f. ÚŘ ČSF,
k. R11/BI/5P/7K. 64) Snímek získal cenu s odůvodněním, že „[…] umělecky svěžím a srozumitelným způsobem sděluje i nej-
menším divákům svou zdravou a prospěšnou myšlenku o potřebě a blahodárnosti lidské práce“. Rozbor pl-
nění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K. p
65) Státní ceny Klementa Gottwalda r. 1956, Praha, 25. 8. 1955. NFA, f. ÚŘ ČSF, k. R10/AI/2P/1K 66) Zápis ze 17. schůze Dramaturgie kresleného a loutkového filmu, Praha, 7. 5. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/
4P/8K.i 67) Zápis o schůzi Ústřední dramaturgie kresleného a loutkového filmu, Praha, 29. 1. 1951. NFA, f. ÚŘ ČSF,
k. R12/AI/3P/5K.i 68) Srov. např. Výtah ze zápisu o schůzi scenáristického oddělení kresleného a loutkového filmu, Praha, 13. 2. 1951. NFA, f. ÚŘ ČSF, k. R9/AI/4P/8K. Zápis o schůzi Dramaturgie kresleného a loutkového filmu, Praha,
12. 2. 1951. NFA, f. ÚŘ ČSF, k. R18/BI/2P/3K. ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) gie v mnoha směrech. V základním ideovém východisku se tento projekt shodoval s náro-
ky na „moderní socialistickou pohádku“, které formuloval například Josef Šlajer v kritické
revue tvorby pro mládež Štěpnice. 73) Námět O neústupné hoře zamítlo scenáristické oddělení mimo jiné proto, že jeho děj se odehrává v „prostře-
dí pro nás netypickém“ (v poušti). Výtah ze zápisu o 13. schůzi scenáristického oddělení kresleného filmu,
Praha, 28. 3. 1951. NFA, f. ÚŘ ČSF, k. R11/BII/5P/9K. Jiné náměty nevyhovovaly celkovou prezentací příro-
dy — například Mramorová tetka svým „mytizujícím pojetím přírody jako vševládné moci nad člověkem“.
Výtah ze zápisu o 19. schůzi scenáristického oddělení kresleného filmu, Praha, 10. 5. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K. i
71) J. Šlajer, c. d., s. 53–54. Státní pedagogické nakladatelství zahrnovalo toto téma i do učebnic pro základní
školy. Například čítanka pro pátý ročník obsahovala samostatný oddíl „Člověk poznává a přetváří přírodu“,
v němž byly kromě sovětských autorů otištěny i texty Josefa Věromíra Plevy nebo Jiřího Marka. Srov. Fran-
tišek V. Kříž – Rudolf Puchýř, Čítanka pro 5. postupný ročník všeobecně vzdělávacích škol. Praha: Státní pe-
dagogické nakladatelství 1955, s. 201–227.
72) Kulturně politické úkoly kresleného filmu a thematický plán pro rok 1952 NFA f ÚŘ ČSF k R14/BI/ 69) Josef Šlajer, Literatura pro děti a mládež jako prostředek socialistické výchovy. Štěpnice 2, 1949, č. 4–5, s.
70) Aprobace kresl. filmu Jablůňka, Praha, 18. 7. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/1P/4K. 69) Josef Šlajer, Literatura pro děti a mládež jako prostředek socialistické výchovy. Štěpnice 2, 1949, č. 4–5, s. 53. 69) Josef Šlajer, Literatura pro děti a mládež jako prostředek socialistické výchovy. Štěpnice 2, 1949, č. 4–5, s. 53.
70) A
b
k
l fil
J blůňk
P
h
18 7 1952 NFA f ÚŘ ČSF k R14/AI/1P/4K 70) Aprobace kresl. filmu Jablůňka, Praha, 18. 7. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/1P/4K. 69) Josef Šlajer, Literatura pro děti a mládež jako prostředek socialistické výchovy. Štěpnice 2, 1949, č. 4–5, s. 53.
70) Aprobace kresl. filmu Jablůňka, Praha, 18. 7. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/1P/4K.
71) J. Šlajer, c. d., s. 53–54. Státní pedagogické nakladatelství zahrnovalo toto téma i do učebnic pro základní
školy. Například čítanka pro pátý ročník obsahovala samostatný oddíl „Člověk poznává a přetváří přírodu“,
v němž byly kromě sovětských autorů otištěny i texty Josefa Věromíra Plevy nebo Jiřího Marka. Srov. Fran-
tišek V. Kříž – Rudolf Puchýř, Čítanka pro 5. postupný ročník všeobecně vzdělávacích škol. Praha: Státní pe-
dagogické nakladatelství 1955, s. 201–227.
72) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/3K.
73) Námět O neústupné hoře zamítlo scenáristické oddělení mimo jiné proto, že jeho děj se odehrává v „prostře-
dí pro nás netypickém“ (v poušti). Výtah ze zápisu o 13. schůzi scenáristického oddělení kresleného filmu,
Praha, 28. 3. 1951. NFA, f. ÚŘ ČSF, k. R11/BII/5P/9K. Jiné náměty nevyhovovaly celkovou prezentací příro-
dy — například Mramorová tetka svým „mytizujícím pojetím přírody jako vševládné moci nad člověkem“.
Výtah ze zápisu o 19. schůzi scenáristického oddělení kresleného filmu, Praha, 10. 5. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K. g g
72) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/B
2P/3K. 9) Josef Šlajer, Literatura pro děti a mládež jako prostředek socialistické výchovy. Štěpnice 2, 1949, č. 4–5, s. 53.
0) Aprobace kresl. filmu Jablůňka, Praha, 18. 7. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/1P/4K. Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu K obdobně antro-
pocentrickému rozuzlení dospívá také Pohádka o stromech a větru, kde děti mladými
větrolamy osázejí vyprahlou krajinu, kde předtím vzrostlé stromy vykácel „ziskuchtivý
boháč“. Námětů z přírodní oblasti prošlo scenáristickým oddělením víc, ale na rozdíl od
těchto dvou nezapadaly do stanoveného pojetí přírody a vztahu člověka k ní.73)l Scénář Jablůňky však výrazněji refl ektoval ještě jeden velmi důležitý a aktuální motiv,
jenž se v námětu nevyskytoval a stal se jedinou zásadnější dějovou změnou provedenou
během scenáristického vývoje. V původní verzi planou jabloň narouboval sadař. V již po- Lukáš Skupa: Která bude ta pravá? 22 Obr. 4–5 — Člověk jako přetvořitel přírody v Pohádce o stromech a větru: děti osází vyprahlou krajinu
mladými větrolamy, které předtím vykácel „ziskuchtivý boháč“ Obr. 4–5 — Člověk jako přetvořitel přírody v Pohádce o stromech a větru: děti osází vyprahlou krajinu
mladými větrolamy, které předtím vykácel „ziskuchtivý boháč“ kročilejší fázi prací na scénáři a výtvarných návrzích ovšem z Filmové rady přišel pokyn,
aby tuto postavu nahradila skupina pionýrů-mičurinců.74) Nikoli náhodou — v tvorbě cí-
lené na dětské publikum se po roce 1948 intenzivně řešila otázka reprezentace pionýrů
v hraných, animovaných i dokumentárních fi lmech. Šlo přitom o téma dosti citlivé, neboť
pionýrské hnutí v organizované podobě bylo tehdy v Československu teprve v počátcích. Ústřední dramaturgie a Filmová rada projednávaly v těchto letech vcelku mnoho „pionýr-
ských látek“, jejichž realizaci zamítaly s odvoláním na schematičnost či nevěrohodnost
zpracování.75) Zástupci schvalovacích sborů autory podněcovali k přirozenějšímu a nenásilnějšímu
pojednání pionýrské činnosti, k čemuž využili právě Jablůňku se zlatými jablky. Po-
dobně jako v Pohádce o stromech a větru sem pionýrský motiv pronikl v rozsahem
vedlejší, avšak významově důležité dějové peripetii. Aniž by byly využity dialogy, účinku-
jí v obou fi lmech pionýrské kolektivy jako veselé skupiny, které do vývoje vyprávění zasa-
hují až v jeho závěru a jedině svými užitečnými skutky. Tím se zároveň defi nitivně stvrzu-
je rozhodující role člověka, jak ji stanovil tematický plán. Úspěšnost „nenuceného“
zakomponování pionýrského motivu si mohli tvůrci alespoň v případě Pohádky o stro-
mech a větru ověřit v diskuzi s publikem v rámci Dětského fi lmového festivalu v Gott-
waldově. Mladí diváci si povšimli, že ne všechny děti měly kolem krku uvázané pionýrské
šátky, což však podle zprávy z této diskuze pokládali za důkaz, že se pionýři neizolují od
ostatní mládeže, ale že ji vedou.76) Jablůňka se zlatými jablky však v reprezentaci pionýrské činnosti zašla podstatně
dál. 75) Například v záznamu diskuze o námětu Jana Kloboučníka Kouzelný šátek lze mezi řádky dokonce vyčíst
obavy, že takový snímek by mohl zafungovat kontraproduktivně, odrazovat od vstupu do pionýrské organi-
zace. Srov. Zápis ze 4. schůze ÚD, Praha, 26. 1. 1950. NFA, f. ÚŘ ČSF, k. R14/BI/4P/1K. 74) Výtah ze zápisu o 13. schůzi scenáristického oddělení kresleného filmu, Praha, 28. 3. 1951. NFA, f. ÚŘ ČSF,
k. R11/BII/5P/9K. 77) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/
2P/3K. p
76) Politické zhodnocení „Dětského filmového festivalu“ v Gottwaldově 1951. NFA, f. ÚŘ ČSF, k. R14/BI/
4P/8K. 76) Politické zhodnocení „Dětského filmového festivalu v Gottwaldově 1951. NFA, f. ÚŘ ČSF, k. R14/BI/
4P/8K.
77) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952 NFA f ÚŘ ČSF k R14/BI/ i
4P/8K.
77) Kulturně-politické úkoly kresleného filmu a thematický plán pro rok 1952. NFA, f. ÚŘ ČSF, k. R14/BI/ Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu Pionýrský motiv autoři provázali s poučným záměrem ve vztahu k přeměně a zužitko-
vání divoké přírody — fi lm měl děti informovat o způsobu a smyslu štěpování stromů
a přiblížit jim tzv. mičurinské pěstitelské metody.77) Od okamžiku, kdy se na scéně objeví ILUMINACE Ročník 32, 2020, č. 4 (120) ČLÁNKY K TÉMATU ILUMINACE Ročník 32, 2020, č. 4 (120) 23 skupina pionýrů, ustupuje pohádková linie vyprávění do pozadí a fi lm postupně nabývá
podoby až nenarativní instruktáže. Závěrečné části dominují převážně detailní záběry ná-
zorného postupu štěpování ovocného stromu a poté i jeho důsledků a přínosu. Právě o zá-
věru členové dramaturgie více diskutovali a zavrhli možnost, že by se u vzrostlého stromu
namísto pionýrů zjevili již svazáci. Uspěl naopak návrh, že „[…] nakonec je třeba ukázat
krásnou jabloň v závěrečné apotheoze“.78) Ve fi nále se tak snímek do pohádkové roviny vý-
razněji nevrací. Nekončí záběry zvířátek pochutnávajících si na sladkých jablkách, ale sta-
tickým pohledem na větve stromku bohatě obsypané velkými zářivými plody. skupina pionýrů, ustupuje pohádková linie vyprávění do pozadí a fi lm postupně nabývá
podoby až nenarativní instruktáže. Závěrečné části dominují převážně detailní záběry ná-
zorného postupu štěpování ovocného stromu a poté i jeho důsledků a přínosu. Právě o zá-
věru členové dramaturgie více diskutovali a zavrhli možnost, že by se u vzrostlého stromu
namísto pionýrů zjevili již svazáci. Uspěl naopak návrh, že „[…] nakonec je třeba ukázat
krásnou jabloň v závěrečné apotheoze“.78) Ve fi nále se tak snímek do pohádkové roviny vý-
razněji nevrací. Nekončí záběry zvířátek pochutnávajících si na sladkých jablkách, ale sta-
tickým pohledem na větve stromku bohatě obsypané velkými zářivými plody. Snímek ve výsledku balancuje na pomezí pohádkového vyprávění a věcného didaktic-
kého sdělení. Zasahuje tím do další kategorie, jež měla mít vedle pohádky v animované
tvorbě pro děti po roce 1948 důležité místo — do populárně-naučného žánru, který se
ovšem ve sledovaném období nestačil v praxi srovnatelným způsobem rozvinout. Jen
o rok později vznikl populárně-naučný fi lm O makovém koláči, který ve výkladovém
stylu podobném mičurinskému motivu z Jablůňky ukazoval malým divákům celý pra-
covní proces předcházející upečení koláče. Snímek v režii Zdeňka Milera ofi ciálně zahajo-
val plánovanou sérii „fi lmových leporel o práci“, ale neměl dalších následovníků. i
Naznačená žánrová hybridnost se viditelně otiskla i do výtvarné koncepce Jablůňky,
pečlivě promýšlené již od výběru výtvarníka — Hofman sliboval co nejdokonalejší návr- Obr. 78) Zápis o 16. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 30. 4. 1951. NFA, f. ÚŘ ČSF, k. R12/
AI/3P/5K. Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu 6–9 — Jablůňka se zlatými jablky: od fi kční pohádky o lesních zvířátkách k populárně-naučné
instruktáži o způsobu a smyslu štěpování ovocných stromů Obr. 6–9 — Jablůňka se zlatými jablky: od fi kční pohádky o lesních zvířátkách k populárně-naučné
instruktáži o způsobu a smyslu štěpování ovocných stromů Obr. 6–9 — Jablůňka se zlatými jablky: od fi kční pohádky o lesních zvířátkách k populárně-naučné
instruktáži o způsobu a smyslu štěpování ovocných stromů 78) Zápis o 16. schůzi Dramaturgie kresleného a loutkového filmu, Praha, 30. 4. 1951. NFA, f. ÚŘ ČSF, k. R12/
AI/3P/5K. Lukáš Skupa: Která bude ta pravá? 24 hy, i kdyby se tím měla zdržet realizace fi lmu.79) Roztroušené zmínky přitom naznačují,
že „nejdokonalejšími“ se rozuměly návrhy co nejvíce realistické. Konkurzu se zúčastnil
Václav Pátek, kterému dramaturgie doporučila držet se ve skicách věrně námětu a zejmé-
na skutečné přírody.80) Jeho „málo jemné a citlivé“ návrhy však neschválila a Pátek další
práci na fi lmu sám vzdal.81) Návrhy dvou dalších nejmenovaných výtvarníků shledala
dramaturgie příliš groteskními a závislými na Disneyho a Sekorově vzoru.82) Za nejvhod-
nějšího kandidáta pak zvolila již osvědčeného Františka Freiwilliga, kterého jako výtvar-
níka snímku schválila i Filmová rada s poznámkou, aby pro zvířata použil „lehčího kolo-
ritu“.83) hy, i kdyby se tím měla zdržet realizace fi lmu.79) Roztroušené zmínky přitom naznačují,
že „nejdokonalejšími“ se rozuměly návrhy co nejvíce realistické. Konkurzu se zúčastnil
Václav Pátek, kterému dramaturgie doporučila držet se ve skicách věrně námětu a zejmé-
na skutečné přírody.80) Jeho „málo jemné a citlivé“ návrhy však neschválila a Pátek další
práci na fi lmu sám vzdal.81) Návrhy dvou dalších nejmenovaných výtvarníků shledala
dramaturgie příliš groteskními a závislými na Disneyho a Sekorově vzoru.82) Za nejvhod-
nějšího kandidáta pak zvolila již osvědčeného Františka Freiwilliga, kterého jako výtvar-
níka snímku schválila i Filmová rada s poznámkou, aby pro zvířata použil „lehčího kolo-
ritu“.83) Freiwillig pojal výtvarnou stránku v duchu hyperrealistické animace, v níž se snažil za-
chovávat reálné vzezření, vlastnosti, zvyky či očekávanou povahu skutečných zvířat a kte-
rou ještě povýšilo hlasové ztvárnění jednotlivých zvířecích postav. 79) Výtah ze zápisu o schůzi Dramaturgie kresleného a loutkového filmu, Praha, 19. 2. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K.
80) Výtah ze zápisu o schůzi Dramaturgie kresleného a loutkového filmu, Praha, 12. 2. 1951. NFA, f. ÚŘ ČSF,
k. R18/BI/2P/3K.
81) Zpráva o kontrole rozboru a zhodnocení plnění výrobního plánu za duben 1951. NFA, f. ÚŘ ČSF, k. R18/
AII/2P/2K.
82) Zápis o schůzi Dramaturgie kresleného a loutkového filmu, Praha, 19. 2. 1951. NFA, f. ÚŘ ČSF, k. R12/AI/
3P/5K.
83) Výtah ze zápisu o 13. schůzi scenáristického oddělení kresleného filmu, Praha, 28. 3. 1951. NFA, f. ÚŘ ČSF,
k. R11/BII/5P/9K.
84) Podobně doslovný realismus rozvinul Freiwillig také v již zmíněném snímku O makovém koláči.
85) Rozbor plnění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K. Jablůňka se zlatými jablky
Jablůňka se zlatými jablky:
Po stopách zapomenutého ideálu Pionýrský motiv štěpo-
vání však od zbytku fi lmu nápadně oddělil odlišnou stylizací, usilující o co nejvěrnější
nápodobu reality (záběry pracovního postupu roubování větví stromu, květů opylovaných
včelou či kreslený „časosběrný záznam“ zrání jablek — viz spodní dva obrázky na přede-
šlé straně).84) Zdá se, že právě toto ztvárnění odpovídalo i představám o „českém stylu“:
podle rozboru plnění plánu studií kresleného a loutkového fi lmu roku 1952 dílo „čerpá
z poctivé realistické základny a mluví svébytně českou řečí v pojetí a hře fi gur i v krajin-
ném prostředí“.85) Ve stejném hodnocení byla Jablůňka označena zároveň za první zdaři-
lé dílo socialistického realismu v domácím kresleném fi lmu. ý
g
j
85) Rozbor plnění plánu v r. 1952, Praha, 23. 2. 1952. NFA, f. ÚŘ ČSF, k. R14/AI/6P/7K. 79) Výtah ze zápisu o schůzi Dramaturgie kresleného a loutkového filmu, Praha, 19. 2. 1951. NFA, f. ÚŘ ČSF,
k. R9/AI/4P/8K. 84) Podobně doslovný realismus rozvinul Freiwillig také v již zmíněném snímku O makovém koláči. Závěr:
Od animovaného k hranému filmu pro děti? Jablůňka se zlatými jablky představovala průsečík většiny nároků kladených na ani-
movanou tvorbu pro děti těsně po roce 1948. Film vznikl podle současného námětu, kte-
rý ideálně zapadal do tematických plánů a na jehož adaptaci se podíleli autoři dětské lite-
ratury. Aktuální tematiku navíc posílili s ohledem na potřebu propagace pionýrského
hnutí. Snímek naplňoval i kýžený požadavek realistického zpracování a srozumitelnosti,
kterou podpořilo užití dialogu, ale třeba i názorné oddělení smyšlené pohádkové linie pří-
běhu od pasáží s didaktickým posláním. Film se tím vzdaloval kritizované samoúčelnosti ILUMINACE Ročník 32, 2020, č. 4 (120) ČLÁNKY K TÉMATU 25 25 některých jeho předchůdců a naopak přibližoval populárně-naučné tendenci, jež v tema-
tických plánech animované produkce patřila k dalším upřednostňovaným žánrům.86) některých jeho předchůdců a naopak přibližoval populárně-naučné tendenci, jež v tema-
tických plánech animované produkce patřila k dalším upřednostňovaným žánrům.86) Ačkoli z dnešního pohledu lze Hofmanův snímek ocenit zejména jako kvintesenci teh-
dejšího ideálu pohádkového fi lmu pro děti, v praxi se jím animovaná tvorba dostala do
slepé uličky. Přestože měla Jablůňka zafungovat coby vzor, žádný podobně koncipovaný
projekt na ni nenavázal. Za několik let už fi lm ani v různých ofi ciálních proslovech nebo
historických shrnutích animace nefi guroval jako klíčové dílo. Pozdější texty tento snímek
zmiňují buď jen velmi okrajově, nebo ho hodnotí přinejmenším shovívavě, ostatně stejně
jako celou vývojovou fázi, během níž se animovaná tvorba na několik let obrátila k dětské-
mu publiku.87) Tato výhradní orientace začala slábnout přibližně od poloviny 50. let —
oproti předešlému období měly získat více prostoru i fi lmy pro dospělé diváky.88) i
Přesto je tato krátká etapa důležitá už proto, že dovoluje nahlédnout do prvopočátků
českého fi lmu pro děti, kdy domácí studia teprve prozkoumávala možnosti, jak by tvorba
pro nejmenší publikum měla vůbec vypadat. Tato pozapomenutá kapitola navíc potvrzu-
je propustnost hranic mezi tradičně oddělovanými sférami hraného a animovaného fi lmu. Ukázalo se, že rozvoj animovaných pohádek se do velké míry odvíjel od všeobecného ne-
dostatku tohoto typu hraných snímků. Kreslené a loutkové fi lmy tak alespoň částečně za-
plňovaly mezeru na trhu v době, kdy hraná produkce prozatím nedokázala uspokojivě re-
agovat na sílící poptávku po fi lmech pro děti. Následující vývoj žánru naznačuje, že hraná a animovaná tvorba sdílely nejen náměto-
vé okruhy, ale i refl exi žánrových specifi k. 86) O animovaných populárně-naučných filmech viz také Lucie Česálková, Atomy věčnosti. Český krátký film
30. až 50. let. Praha: NFA 2014.
87) Srov. např. Marie Benešová, Kapitoly z dějin českého animovaného filmu. Praha: ČFÚ 1979, s. 93–99; Jan Poš,
Čs. animovaný film (9), Bratři v triku 1949–1957. Zpravodaj československého filmu 13, 1987, č. 9, s. 36–42.
88) Podle tematického plánu Studia kresleného a loutkového filmu na rok 1956 je cílem animované tvorby bo-
hatší a rozmanitější rejstřík — mimo jiné satirické filmy, ve větší míře než dosud grotesky, moderní pohád-
ky a bajky, ale také zahraniční spolupráce. Srov. Thematický plán na rok 1956, Praha, září 1955. NFA, f. ÚŘ
ČSF, k. R14/AI/6P/6K.
89) Zpráva pro ředitelství filmové tvorby, Praha, 1. 7. 1954. NFA, f. ÚŘ ČSF, k. R9/AI/5P/4K. Závěr:
Od animovaného k hranému filmu pro děti? Samostatné Studio dětského, kresleného a lout-
kového fi lmu, tedy první specializovaná platforma orientovaná na tento žánr, vzniklo až
v roce 1954. Jeho základní plány a cíle přitom obsahovaly tytéž teze, jež tvořily páteř dra-
maturgie animovaných fi lmů v předchozích letech: aby náměty „[…] ať ze současnosti,
nebo minulosti, či z prostředí pohádkového [byly] zpracovány umělecky tak, aby živě
a přesvědčivě hovořily k dnešnímu mladému divákovi, aby pomáhaly překonávat v myšle-
ní a cítění lidí přežitky buržoazní ideologie a úpadkového umění, aby […] pomáhaly ve
výchově nového člověka“.89) Stejně jako v animovaném fi lmu i v hrané tvorbě hledala dramaturgie těsné spoje
s dětskou literaturou a jejími autory či deklarovala snahu o národní specifi čnost nebo rea-
listické zpracování. Vrací se také poznatek o formování žánru skrze motiv popírání — na-
místo defi nic, jak by hrané fi lmy pro děti měly vypadat, se častěji rozebíralo, jak by vypa-
dat neměly. S tím přímo souvisejí i spory, připomínající přípravu a vznik prvních
animovaných pohádek — schvalování několika verzí scénáře Pyšné princezny provázely
sáhodlouhé diskuze, jak co nejcitlivěji aktualizovat předlohu Boženy Němcové Potrestaná Lukáš Skupa: Která bude ta pravá? 6 26 pýcha. Podobný průběh jako u animovaných projektů mívaly debaty o převážně nerealizo-
vaných námětech hraných fi lmů podle přehodnocených a moderních pohádek. Konkrét-
ní rysy, jež si žánr animované pohádky při své vývojové cestě osvojoval, tedy v kategorii
fi lmu pro děti hlouběji zakořenily a vynořovaly se i v příštích letech. Text vznikl v návaznosti na výzkumný projekt Historie českého fi lmu pro děti 1945–1992, realizovaný
v rámci Národního fi lmového archivu a podpořený Státním fondem kinematografi e. Citované fi lmy: A B C (Eduard Hofman, 1949), Bajaja (Jiří Trnka, 1950), Basa tvrdí muziku (Karel Mann, 1948),
Čert a Káča (Václav Bedřich, 1955), Hrnečku, vař! (Václav Bedřich, 1953), Ivánku náš (Karel Mann,
1949), Jablůňka se zlatými jablky (Eduard Hofman, 1952), Kombajn (Josef Kluge, 1951), Lenora
(Eduard Hofman, 1949), O kohoutkovi a slepičce (Zdeněk Miler, 1953), O makovém koláči (Zdeněk
Miler, 1953), O milionáři, který ukradl slunce (Zdeněk Miler, 1948), Pinocchio (Hamilton Luske, Ben
Sharpsteen, 1940), Pohádka o drakovi (Hermína Týrlová, 1953), Pohádka o stromech a větru (Václav
Bedřich, 1951), Pyšná princezna (Bořivoj Zeman, 1952), Šedé pírko (Seraja šejka; Leonid Amalrik,
Vladimir Polkovnikov, 1948), Šváb (Karel Mann, 1948), Vzducholoď a láska (Jiří Brdečka, 1948), Za-
sadil dědek řepu (Jiří Trnka, 1945), Závistivá zvířátka (Karel Mann, 1950), Zlatý klíček (Zolotoj
ključik; Alexandr Ptuško, 1939). Lukáš Skupa p
Absolvent Ústavu fi lmu a audiovizuální kultury Filozofi cké fakulty Masarykovy univerzity, fi lmový
historik, pedagog, publicista. Své texty a studie publikuje ve fi lmově zaměřených magazínech a pub-
likacích, na některých z nich se podílel i jako editor (Ostře sledované vlaky, Tajemství hradu v Kar-
patech). V roce 2016 vydal v Národním fi lmovém archivu svoji první monografi i Vadí — nevadí. Česká fi lmová cenzura v 60. letech, aktuálně připravuje knihu o dějinách českého fi lmu pro děti v le-
tech 1945 až 1992. THEMED ARTICLES ILUMINACE Volume 32, 2020, No. 4 (120) Which Will Be the Right One?
Th e Beginnings of Fairytale Genre in Czech Animated Film aft er 1948 Lukáš Skupa Th e brief period from 1948 to 1953 can be considered as the formative years of fairytale genre in
Czech animated fi lm. Th is study focuses on the dramaturgy of animated fi lm studios, which were
obliged to reorient their production almost exclusively to children‘s audience. For this purpose the
conncetions between children’s fi lms and children‘s literature were developing aft er 1948. Scriptwrit-
ers as well as artists were recruited from the literary area and some of literary fairy tales were adapt-
ed as animated fi lms. Especially the type of so called modern fairy tale was determined as the best
option for fi lm adaptation with regard to the goals of education and reeducation of children — mod-
ern fairy tales could served for articulation of actual needs of socialist cultural policy. Th e Apple-Tree
with the Golden Fruit — Eduard Hofman‘s fi lm from 1952 which tells the story about animals, to
whom a group of Pioneers help to grow tasty apples — became the exemplary work of this tenden-
cy. Th is fi lm combined fairytale narration with didactic popular-educational message and represent
the intersection of most demands placed on Czech animated fi lms for children aft er 1948. It was
marked as the fi rst succesful work of socialistic realism made in Czech animated fi lm studios. klíčová slova: československá zestátněná kinematografi e, český animovaný fi lm, fi lm pro děti,
fi lmová adaptace, fi lmová pohádka keywords: Czechoslovak nationalized cinema, Czech animated fi lm, children‘s fi lm, fi lm
adaptation, fairytale fi lm
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Effect of various mulches on soil physico—Chemical properties and tree growth (Sophora japonica) in urban tree pits
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PloS one
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cc-by
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OPEN ACCESS Citation: Qu B, Liu Y, Sun X, Li S, Wang X, Xiong
K, et al. (2019) Effect of various mulches on soil
physico—Chemical properties and tree growth
(Sophora japonica) in urban tree pits. PLoS ONE 14
(2): e0210777. https://doi.org/10.1371/journal. pone.0210777 Editor: Arun Jyoti Nath, Assam University, INDIA
Received: December 14, 2017
Accepted: January 2, 2019
Published: February 6, 2019
Copyright: © 2019 Qu et al. This is an open access
article distributed under the terms of the Creative
Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in
any medium, provided the original author and
source are credited. Editor: Arun Jyoti Nath, Assam University, INDIA
Received: December 14, 2017
Accepted: January 2, 2019
Published: February 6, 2019 Copyright: © 2019 Qu et al. This is an open access
article distributed under the terms of the Creative
Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in
any medium, provided the original author and
source are credited. Effect of various mulches on soil physico—
Chemical properties and tree growth (Sophora
japonica) in urban tree pits Bingpeng QuID1,2☯, Yuanxin Liu1☯, Xiangyang Sun1*, Suyan Li1*, Xinyu Wang1,
Kaiyi Xiong1, Binghui Yun1, Hua Zhang1
1 Forestry College, Beijing Forestry University, Beijing, China, 2 Tianjin LVYIN Landscape and Ecology
Construction Co., Ltd, Tianjin, China Bingpeng QuID1,2☯, Yuanxin Liu1☯, Xiangyang Sun1*, Suyan Li1*, Xinyu Wang1,
Kaiyi Xiong1, Binghui Yun1, Hua Zhang1 Kaiyi Xiong , Binghui Yun , Hua Zhang
1 Forestry College, Beijing Forestry University, Beijing, China, 2 Tianjin LVYIN Landscape and Ecology
Construction Co., Ltd, Tianjin, China 1 Forestry College, Beijing Forestry University, Beijing, China, 2 Tianjin LVYIN Landscape and Ecolog
Construction Co., Ltd, Tianjin, China 1 Forestry College, Beijing Forestry University, Beijing, China, 2 Tianjin LVYIN Landscape and Ecology
Construction Co., Ltd, Tianjin, China a1111111111
a1111111111
a1111111111
a1111111111
a1111111111 ☯These authors contributed equally to this work. * sunxy@bjfu.edu.cn(XS); lisuyan@bjfu.edu.cn(SL) RESEARCH ARTICLE Abstract Mulching is a widely employed soil management practice. The mulches used have variable
effects on the soil properties and plant growth. In China, mulches are used to cover bare soil
at a few places in landscapes, where most of the soil is uncovered, especially in tree pits. As
to improve our understanding on the effect of various mulches on soil properties and tree
growth after two years of the treatment justifying its implication in soil fertility and tree growth. A comparison study was conducted to determine the effects of inorganic (cobblestone—CB;
water permeable brick—WPB), organic (pine bark—PB; green waste compost—GWC), and
living (turf grass—TG) mulches on soil physical and chemical properties at three different
depths (0–10 cm, 10–20 cm, and 20–40cm), and on tree growth (Sophora japonica) in urban
tree pits. Soil moisture was measured once a month in 2015.The soil samples were collected
from the tree pits two years after mulching and used to evaluate the physical and chemical
properties. Further, trunk diameter and tree height were determined once a year. During the
most months, all types of mulches significantly affected the moisture content of the soil at all
the depths analyzed. In July and August, however, the moisture content of PB and TG treated
soil decreased when compared with that of unmulched bare soil. Two years after mulching,
the bulk density of the soil treated with PB, GWC, and TG was significantly affected at10–20
cm, with GWC exhibiting a relatively better effect. The treatments with PB, GWC, and TG
also improved the total porosity, macroporosity, and microporosity of the soil at lower depths. Further, WPB worsened the bulk density and porosity of the soil, elevating the pH at lower
depths. The organic matter, total N, mineral N, available P, and available K contents of the
soil at lower depths increased when mulched with organic material. Turf grass significantly
increased only the total N and available K at 0–10 and 10–20 cm. There was no significant
difference in the soil properties among the treatments at 20–40 cm. Furthermore, the trunk
diameter and tree height were not affected by the mulches two years after mulching. In con-
clusion, organic mulches especially GWC, not only increased soil fertility significantly but
improved soil physical characters (0–10 cm depth) comparing to other mulches, are suitable
to cover bare soil in urban tree pits. Introduction to declare. This does not alter our adherence to
PLOS ONE policies on sharing data and materials. to declare. This does not alter our adherence to
PLOS ONE policies on sharing data and materials. Mulches have been widely used in agriculture lands, orchards, forests, and landscapes in many
parts of the world [1]. Generally, mulches reduce competition from weeds, maintain soil
temperature, and reduce evaporation from soil [2]. They protect the soil from wind-, water-,
and traffic-induced erosion. Further, fugitive dust from soil is suppressed by mulching [3]. Mulches also improve soil properties by improving moisture retention capacity, releasing dif-
ferent nutrients, and enhancing biological activities [4]. Therefore, with the improved soil
properties, plants grow better [5, 6]. Mulches are broadly classified into three main groups: organic, inorganic, and living
mulches. Organic mulches are derived from organic substances, such as agricultural wastes
(straw and rice husks), waste from wood industries (sawdust and barks), and green wastes
(leaves and wood chips) [1]. Inorganic mulches include polyethylene film, gravel, bricks,
and cobblestones. Living mulches include clover, Manila grass, ryegrass, dwarf lily turf, and
other types of grasses [7]. Each type of mulch has a particular set of characteristics. The
choice of mulch depends on the type of soil, climate, and nutritional requirements of plants
[8]. In China, gravel (cobblestone) is the inorganic mulch widely used in landscapes [9].It
reduces water evaporation, moderates soil temperature, and suppresses weeds [10, 11]. Fur-
ther, barks are used as the major organic mulch in landscapes. The barks not only maintain
soil moisture, reduce soil temperature fluctuations, release nutrients into the soil, but also
help enrich landscape types [12, 13]. However, one of the major concerns in using organic
mulch is its potential to reduce the mineral nitrogen content in the soil [14]. Various stud-
ies have reported that decaying organisms under organic mulches require immobilized
mineral nitrogen [15]. The living mulches can enhance soil quality and conserve soil mois-
ture content in landscapes. However, a few studies have demonstrated that the living
mulches also compete with the plants for nutrients and water, thus limiting plant growth
[16]. Mulches have been widely used in gardens and green belts along the roadside in developed
countries [17–19]. They can improve plant growth in landscapes by enhancing soil quality,
such as conserving soil moisture and increasing soil nutrients [20]. Mulches, soil and tree growth Data Availability Statement: All relevant data are
within the paper. Funding: This work was supported by the
Municipal Project for Science and Technology of
Beijing (Z161100001116061 “Research and
demonstration of key technologies for improving
soil quality of greenbelt and forest land in Beijing”,
and Z161100001316004). Competing interests: BQ is affiliated with Tianjin
LVYIN Landscape and Ecology Construction Co.,
Ltd. There are no patents, products in development
or marketed products associated with this research 1 / 12 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Mulches, soil and tree growth Table 1. Physical properties of the soil at different depths in the urban tree pits before the experiment. Depth
Bulk density (g.cm-3)
Total porosity (%)
Macroporosity (%)
Microporosity (%)
0–10 cm
1.45±0.03a
45.15±2.13a
36.84±2.69a
8.31±1.80a
10–20 cm
1.50±0.05a
42.08±1.55a
35.09±0.79a
6.99±0.79a
20–40 cm
1.52±0.06a
41.54±1.91a
34.12±0.62a
7.42±0.62a
Values with different letters in the same column indicate significant differences between treatments (p<0.05, n = 3). Data are means ± standard deviation. Table 1. Physical properties of the soil at different depths in the urban tree pits before the experiment. Table 1. Physical properties of the soil at different depths in the urban tree p Values with different letters in the same column indicate significant differences between treatments (p<0.05, n = 3). Data are means ± standard deviation. Values with different letters in the same column indicate significant differences between treatments (p<0.05, n = 3). Dat https://doi.org/10.1371/journal.pone.0210777.t001 Material and methods
Site description The present study was conducted at Zhixin Road, Haidian District, Beijing, China (39˚ The present study was conducted at Zhixin Road, Haidian District, Beijing, China (39˚
59040@N, 116˚22018@E; altitude 43.5 m). The region has a typical temperate and monsoon cli-
mate with average annual temperature and rainfall of 12.5˚C and 628.9 mm, respectively. The
tree species in the tree pits along the roadside included S. japonica L., Populus tomentosa Carr.,
Platycladus orientalis L., and Ginkgo biloba L. The tree pits with S. japonica located north of the road were chosen for the present study. The age of the trees were 12–15 years, with average height and trunk diameter of 12.2 m and
26.7 cm, respectively. The inter-tree spacing was 4 m and area of each tree pit was 1.44 m2 (1.2
m × 1.2 m). The soil of the tree pits was classified as brown soil (19.5% clay, 65.1% silt, and
15.4% sand), which was cleared manually before the study. The physical and chemical proper-
ties of the soil (0–10 cm, 10–20 cm, and 20–40 cm depth) in the tree pits were determined
before mulching (Tables 1 and 2). Introduction In China, mulches are used
to cover bare soil only at a few places in landscapes, where most of the soil is uncovered, espe-
cially in tree pits [21]. The tree-pit soil is an important source of nutrients for the growth of plants [22]. However,
the soil in urban tree pits in China is associated with many problems, such as a low degree of
porosity and high soil pH and density, which limit tree growth to some extent [23]. In Beijing,
the capital of China, the pH of urban soils might be elevated due to limestone inadvertently
being introduced through discarded construction material and rubbles [24]. And particulate
matter 2.5 (PM2.5) has become one of the serious environmental problems in Beijing. The soil
dust originating from the bare soil—of the tree pits and edge of flower beds—is one of the
sources of PM2.5 [25]. Therefore, covering the bare soil in tree pits with mulch not only
improves soil properties but also inhibits the generation of soil dust from tree pits. Under-
standing the effect of various mulches on pit soil properties and tree growth is vital for apply-
ing appropriately. Few studies have evaluated the effect of various mulches on the soil quality
and plant growth in urban tree pits, especially in China. The aim of the present study was to
determine the effects of three types of mulches (organic, inorganic, and living) on the physical
and chemical properties of soil, to identify the effects on the growth of street tree (Sophora
japonica), and to provide preliminary suggestion for choosing suitable mulches based on our
research. 2 / 12 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Soil sampling and analysis The soil moisture content was measured at monthly interval during 2015. Three random soil
samples at depths 0–10, 10–20, and 20–40 cm, respectively, were collected. The individual soil
samples were then placed in sterile plastic sealing bags and transported to the laboratory,
where they were oven-dried at 105˚C for 8 h and used for further analysis. Two-years after mulching, the soil was sampled from the tree pits on June 26, 2016. Three
soil samples per tree pit at depths 0–10, 10–20, and 20–40, respectively, were collected in 100
cm3 volumetric containers and used to evaluate the physical properties of soil. Further, six ran-
dom soil samples at depths 0–10, 10–20, and 20–40 cm were collected to evaluate the chemical
properties of soil. All the soil samples were placed in sterile plastic sealing bags and transported
to the laboratory in car refrigerators. The soil sample used to evaluate the chemical properties was divided into three subsamples. The first subsample was stored at 4˚C to analyze the moisture content. The second subsample
was air-dried in a soil drying room and ground. The sample was then passed through a 2-mm
sieve to remove pebbles, construction wastes, fine roots, and other foreign materials before the
analyses. The third subsample was obtained by passing a portion of the second subsample
through a 0.149-mm sieve. The bulk density of the sampled soil was measured as the mass of oven-dried soil. The total
porosity was assessed by measuring soil saturation (total volume of water-filled soil pores);
microporosity was assessed using tension table and water column of 6 × 10−3 MPa; macropor-
osity was calculated as the difference between the total porosity and microporosity. All evalua-
tions were performed according to the methodologies described by Embrapa (1997) [28].The
second subsample of soil was used to determine the mineral nitrogen (N) content and pH. The
mineral N was determined by alkali-hydrolytic diffusion method [29]. The soil pH was deter-
mined using a pH meter at a soil to water ratio of 2:5. The third subsample was used to esti-
mate organic matter, available phosphorus (P), available potassium (K), and total N. The
organic matter was measured by sulfuric acid-potassium dichromate wet oxidation, followed
by titration with ferrous sulfate according to the procedure of Walkley-Black [30]. The avail-
able P in the soil was measured by the Olsen method. Experimental design Six treatments were established for the present study: (1) unmulched bare soil (CK); (2) cob-
blestones (CB) (inorganic mulch) of thickness < 2 cm and diameter 1–3 cm; (3) water perme-
able bricks (WPB) (inorganic mulch)of size 15 cm × 15 cm × 6 cm; (4) pine barks (PB)
(organic mulch), wood processing residues of Pinus tabuliformis Carr., of thickness < 1 cm
and length < 6 cm (PB); (5) green waste compost (GWC) (organic mulch) from green waste. Green waste was obtained from municipal curbside collection, cut into pieces (0.5–1 cm parti-
cle size), and then subjected to fermentation. The moisture content of the raw material was
adjusted to 60% (w/w) and carbon to nitrogen ratio was adjusted between 25 and 30 for opti-
mal microbial activity during fermentation [26]; (6) turf grass (TG) (living mulch); Ophiopo-
gon japonicus var.nana,a herb, has been widely used in landscaping in Northern China. It was
chosen to be planted as turf grass in the present study, the height of the aerial part was 5–6 cm. Each tree pit was laid with different mulch (up to 6 cm depth). A circle of diameter 6–10 cm
larger than the trunk diameter was created around the base of the trunk without any mulch. This was done to avoid the limiting effect of mulch on tree growth. The turf grass O. japonicus Table 2. Chemical properties of the soil at different depths in the urban tree pits before the experiment. Depth
pH
Organic matter (g.kg-1)
Total N (g.kg-1)
Mineral N (mg.kg-1)
Available P (mg.kg-1)
Available K (mg.kg-1)
0–10 cm
8.32±0.31a
15.70±2.48a
1.15±0.05a
8.63±0.80ab
14.32±0.97a
89.15±3.60a
10–20 cm
8.28±0.41a
11.02±1.04a
1.02±0.11b
11.5±2.24a
13.79±0.99ab
88.09±6.56a
20–40 cm
8.66±0.42a
12.98±1.74a
0.94±0.10b
7.35±0.78b
12.76±1.74b
92.70±3.85a
Values with different letters in the same column indicate significant differences between treatments (p<0.05, n = 3). Data are means ± standard deviation. htt
//d i
/10 1371/j
l
0210777 t002 emical properties of the soil at different depths in the urban tree pits before the experiment. ent depths in the urban tree pits before the experiment. Table 2. Chemical properties of the soil at different depths in the urban tree pits before the experiment. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 3 / 12 Mulches, soil and tree growth was transplanted at a spacing of 10 cm. The experiment was a randomized complete block
design with three replicates. Experimental design Studies have suggested that soil properties and tree growth could be determined to compare
different mulching effect under the same conditions (in the botanic garden or at the institute,
other environmental factors had no differences) after one-year’s study period [14, 27]. Consid-
ering the slow growth of the trees in the tree pits, two-years’ study period was used during this
experiment. The study commenced on July 5, 2014 and finished at June 26, 2016. After laying
the mulches, the tree pits were watered adequately. During the study period, anthropogenic
activities, daily landscape maintenance by the government authorities (such as watering the
trees every 3–7 days), and application of fertilizer were avoided. Soil sampling and analysis The available K in the soil was deter-
mined using a flame photometer after ammonium acetate extraction. Effect of different mulches on the moisture content of soil in the tree pits The moisture content of the soil treated with different mulches was measured at depths 0–10,
10–20, and 20–40 cm from January to December 2015. The results indicated that at different
depths, different mulches exhibited different effects on the soil moisture content. Further, dur-
ing most months, the mulches significantly affected the moisture content of the soil at all the
depths analyzed. Fig 1 represents the rainfall in Beijing in 2015. The study site also received
similar amount of rain, with a relatively high rainfall in summer (July and August). Conse-
quently, all the treatments exhibited relatively high moisture content in July and August. At 0–10 cm, during most months, the mulches increased the soil moisture content, except
in July and August (Fig 2). Further, the effect of inorganic mulches (CB, WPB) on the soil
moisture content was stronger than that of the organic mulches (PB, GWC) and living mulch
(TG). In July and August, CB and WPB increased the soil moisture content. This might be
associated with decreased rate of evaporation from the soil due to CB and WPB. However, the
effect of other mulch types (PB, GWC and TG) on soil moisture content was lesser than that of
CK. This might be due to the following reasons: (1) the organic mulches (PB, GWC) perhaps
increased the rate of evaporation, and they absorbed water from the soil at tropical tempera-
tures and (2) the TG and plants competed for water when the transpiration rate increased in
plants due to hot weather. Similar results were observed at depths 10–20 and 20–40 cm, how-
ever the variations were significantly high. The moisture content of the soil treated with different mulches was measured at depths 0–10,
10–20, and 20–40 cm from January to December 2015. The results indicated that at different
depths, different mulches exhibited different effects on the soil moisture content. Further, dur-
ing most months, the mulches significantly affected the moisture content of the soil at all the
depths analyzed. Fig 1 represents the rainfall in Beijing in 2015. The study site also received
similar amount of rain, with a relatively high rainfall in summer (July and August). Conse-
quently, all the treatments exhibited relatively high moisture content in July and August. At 0–10 cm, during most months, the mulches increased the soil moisture content, except
in July and August (Fig 2). Tree growth The trunk diameter and tree height were determined once a year. The first measurement was
recorded on June 20, 2015, with subsequent measurement on the day of soil sampling. The
height of the tree was measured from the soil surface to the highest point of the tree crown
using a Blume-Leiss altimeter. The diameter of the tree trunk was measured 15 cm above the
soil surface using a tree caliper. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 4 / 12 Mulches, soil and tree growth Statistical analysis All data were analyzed using the SPSS software package (version 20.0) (IBM Corporation,
Armonk, New York). The data were subjected to one-way of variance (ANOVA) and means
were separated by the least significant difference test at P < 0.05. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Effect of different mulches on the moisture content of soil in the tree pits Further, the effect of inorganic mulches (CB, WPB) on the soil
moisture content was stronger than that of the organic mulches (PB, GWC) and living mulch
(TG). In July and August, CB and WPB increased the soil moisture content. This might be
associated with decreased rate of evaporation from the soil due to CB and WPB. However, the
effect of other mulch types (PB, GWC and TG) on soil moisture content was lesser than that of
CK. This might be due to the following reasons: (1) the organic mulches (PB, GWC) perhaps
increased the rate of evaporation, and they absorbed water from the soil at tropical tempera-
tures and (2) the TG and plants competed for water when the transpiration rate increased in
plants due to hot weather. Similar results were observed at depths 10–20 and 20–40 cm, how-
ever the variations were significantly high. Fig 1. Monthly rainfall in Beijing, 2015. https://doi.org/10.1371/journal.pone.0210777.g001
PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019
5 / 12 Fig 1. Monthly rainfall in Beijing, 2015. https://doi.org/10.1371/journal.pone.0210777.g001 https://doi.org/10.1371/journal.pone.0210777.g001 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 5 / 12 Mulches, soil and tree growth Fig 2. Monthly soil moisture content at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks
(WPB), pine barks (PB), green waste compost (GWC), and turf grass (TG) in 2015. https://doi.org/10.1371/journal.pone.0210777.g002 Fig 2. Monthly soil moisture content at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks
(WPB), pine barks (PB), green waste compost (GWC), and turf grass (TG) in 2015. Fi 2 M
hl
il
i
h
diff
d
h i
h
b
i
d
i h b
il (CK)
bbl
(CB)
bl b i k Fig 2. Monthly soil moisture content at three different depths in the urban tree pits treated with bare soil (CK), cobble
(WPB), pine barks (PB), green waste compost (GWC), and turf grass (TG) in 2015. https://doi.org/10.1371/journal.pone.0210777.g002 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 6 / 12 Mulches, soil and tree growth Earlier studies have suggested that mulches, including plastic, gravel, barks, wood chips,
and grass can retain the moisture content of soil by reducing the rate of evaporation [9, 31–
33]. Effect of different mulches on the moisture content of soil in the tree pits Several studies have revealed that living mulches can increase the moisture content of soil
[27, 34], but because of the increasing rate of transpiration in plants, the opposite results were
observed in the present study for living mulches in July and August. Effect of different mulches on the physical properties of soil in the tree pits There was no significant difference in the bulk density of the soil treated with different
mulches when compared with that of CK, except for PB, GWC, and TG treated soil at10–20
cm (Table 3). The treatments with PB, GWC, and TG significantly affected the bulk density at
10–20 cm when compared with that of CK, with GWC exhibiting a relatively strong effect. Some studies have shown that mulches can improve the bulk density of soil, but not signifi-
cantly [27, 35]. At all the depths analyzed, WPB treated soil exhibited relatively low total poros-
ity, macroporosity, and microporosity at the depths 0–10 and 10–20 cm. Relatively high total
porosity, macroporosity, and microporosity were observed in GWC treated soil at 0–20 cm. Furthermore, at depths 0–10 and 10–20 cm, TG treated soil exhibited relatively high total
porosity and macroporosity. These results might be due to many reasons: (1) when it rains,
organic mulches (PB, GWC) might release organic matter creating a favorable condition for
microorganism. Therefore, the porosity and bulk density of the soil at lower depths is im-
proved [36]; (2)the root activity of TG—a living mulch—might have played an important role
in improving the physical properties of the soil [27]; and (3) the use of high-density WPB as a
mulch to cover bare soil can make the soil compact. Values with different letters in the same column indicate significant differences between treatments (p<0.05, n = 3). Data ar https://doi.org/10.1371/journal.pone.0210777.t003 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 https://doi.org/10.1371/journal.pone.0210777.t003 Effect of different mulches on the chemical properties of soil in the tree pits The chemical properties of the soil two-years after mulching have been represented in Fig 3. The pH of the soil (Fig 3A) treated with WPB was significantly higher than that of CK. The pH Table 3. Soil physical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC) and turf grass (TG) two-years after mulching. Depth
Treatment
Bulk density (g.cm-3)
Total porosity (%)
Macroporosity (%)
Microporosity (%)
0–10 cm
CK
1.47±0.13ab
44.28±1.72b
36.47±1.54bc
7.81±0.38bc
CB
1.51±0.03a
45.13±1.16b
35.97±1.18bc
9.16±0.30ab
WPB
1.55±0.06a
40.64±1.29c
33.46±0.70c
7.18±0.65c
PB
1.39±0.03ab
47.21±1.25ab
37.84±2.37ab
9.37±1.41ab
GWC
1.34±0.04a
49.96±1.22a
39.67±1.63ab
10.29±0.45a
TG
1.39±0.07ab
48.31±1.81a
40.96±2.26a
7.35±0.45c
10–20 cm
CK
1.52±0.05a
43.64±1.10bc
35.41±1.14b
8.23±0.34a
CB
1.48±0.03ab
43.71±1.09bc
36.77±1.52ab
6.94±0.70a
WPB
1.55±0.06a
41.27±1.76c
35.09±1.04b
6.18±1.64a
PB
1.42±0.03bc
44.54±1.47bc
37.69±2.22ab
6.85±2.51a
GWC
1.36±0.07c
48.72±1.87a
40.06±1.13a
8.66±0.78a
TG
1.42±0.05bc
46.80±2.39ab
37.86±2.45ab
8.94±0.45a
20–40 cm
CK
1.50±0.09a
41.92±0.83a
35.02±1.02a
6.90±0.35a
CB
1.52±0.03a
42.98±1.14a
35.84±1.83a
7.14±0.72a
WPB
1.51±0.04a
41.76±1.87a
35.25±1.39a
6.51±0.85a
PB
1.54±0.07a
42.76±1.15a
35.24±1.13a
7.52±0.60a
GWC
1.55±0.07a
41.69±1.51a
35.18±1.24a
6.51±0.48a
TG
1.53±0.07a
43.40±1.93a
35.98±1.50a
7.42±0.68a fferent depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
d turf grass (TG) two-years after mulching. Table 3. Soil physical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC) and turf grass (TG) two-years after mulching. Soil physical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), w
PB), green waste compost (GWC) and turf grass (TG) two-years after mulching. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 7 / 12 Fig 3. Soil chemical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC), and turf grass (TG) two years after mulching. Different letters above the error bars indicate significant differences between
treatments (p<0.05, n = 3). The error bars indicate standard deviation. https://doi org/10 1371/journal pone 0210777 g003
Mulches, soil and tree growth Mulches, soil and tree growth Fig 3. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Effect of different mulches on the chemical properties of soil in the tree pits Soil chemical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC), and turf grass (TG) two years after mulching. Different letters above the error bars indicate significant differences between
treatments (p<0.05, n = 3). The error bars indicate standard deviation. Fig 3. Soil chemical properties at three different depths in the urban tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC), and turf grass (TG) two years after mulching. Different letters above the error bars indicate significant differences between
treatments (p<0.05, n = 3). The error bars indicate standard deviation. https://doi.org/10.1371/journal.pone.0210777.g003 https://doi.org/10.1371/journal.pone.0210777.g003 of soil at depths 0–10 and 10–20 cm were 7.8% and 6.6% higher than that of CK. The relatively
high pH observed was probably due to leaching of basic cations from WPB that are made of
concrete. There was no significant difference in the soil pH among other treatments. Billeaud
et al. [37] reported that mulches, such as wood chips, pine barks, and cypress, can significantly
decrease the pH of fine sandy loam soil two-years after mulching. Pickering et al. [36] observed
that mulching with cocoa shells, wood chips, garden compost, and horse manure can signifi-
cantly increase the pH of sandy loam soil after one year. Qian et al. [7] reported that there was
no significant difference in the soil pH between no mulch and living mulch treatments in the
sandy loam soil of an apple orchard three-years after the treatment. The results of the present
study were synonymous with these reports. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 8 / 12 Mulches, soil and tree growth The organic matter in GWC and PB treated soils was significantly high at the depths 0–10
and 10–20 cm. Further, the organic matter in GWC treated soil was significantly higher than
that of PB treated soil. Earlier studies have shown that organic mulches increase the soil
organic matter by increasing biological activity in the soil [38, 39]. Similar results were
observed in the present study. However, TG did not significantly affect the soil organic matter
in the present study. This result contradicted earlier studies, which reported that grass signifi-
cantly increased soil organic matter due to increased photosynthesis [27, 40]. Effect of different mulches on the chemical properties of soil in the tree pits The total N content of the soil treated with GWC and TG was significantly higher at the
depths 0–10 and 10–20 cm when compared with that of CK. In GWC treated soil, significant
increase of 37.1% and 26.1% were recorded at the depths of 0–10 and 10–20 cm, respectively. Further, PB and CB treatments also increased the total N content of the soil at 0–10 cm, how-
ever, the increase was not significant. The results revealed that the soil treated with organic
mulches—GWC and PB—exhibited higher mineral N content when compared with that of
CK, with GWC treated soil exhibiting a relatively high mineral N content. There was no signif-
icant difference among the treatments at the depths 10–20 and 20–40 cm. The available P content was significantly affected only by the organic mulches—GWC and
PB—at 0–10 cm. Further, GWC treatment showed a significant increase in the available P con-
tent at 10–20 cm. Although not significant, PB treatment also increased the available P content
at 10–20 cm. Furthermore, GWC and PB treatment also significantly increased the available K
content at 0–10 cm. Treatment with TG significantly increased the available K content at the
depths 0–10 and 10–20 cm. It was evident from the results that the differences in the mineral N, available P, and avail-
able K contents between treatments were similar to that of the organic matter. The favorable
change in the available nutrients due to mulching with organic materials can be attributed to
the increased biological activity in the soil. Thus, resulting in the mineralization of organic
matter leading to increased nutrient content [41]. The treatment with GWC significantly
increased not only the soil nutrients, but also total N. This might be because GWC was rich in
nutrients. Therefore, the layer between GWC and soil supported a large number soil microbe
population, which increased the enzyme activity in the region. In the present study, the treatment with TG increased the total N content of the soil at lower
depths two-years after mulching. This might be due to the activity of TG roots providing suitable
condition for the growth of microorganisms that release N via organic matter decomposition [7]. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Conclusions The inorganic mulches (CB and WPB), organic mulches (GWC and PB) and living mulch
(TG) exhibited positive effect by elevating the soil moisture content. However, the moisture
content of soil applying GWC, PB, and TG decreased in July and August when compared with
that of CK. PB, GWC, and TG improved the soil bulk density, total porosity, macroporosity,
and microporosity of the soil to some extent at lower depths, whilst WPB worsened those fac-
tors. In addition, by comparing with other treatments, GWC significantly increased the level
of all the nutrients. There was no significant difference in the soil properties among the treat-
ments at 20–40 cm. Furthermore, the trunk diameter and tree height of S. japonica were not
affected by the mulches. Organic mulches, especially GWC, seem to be better than other
mulches analyzed to cover bare soil in the urban tree pits. Effect of different mulches on the trunk diameter and height of the tree The ANOVA revealed that the trunk diameter and height of S. japonica was not affected by dif-
ferent mulches. None of the treatments increased the trunk diameter and tree height signifi-
cantly (Fig 4). Several studies have reported that mulching had no effect on tree growth. Ferrini et al. [42] observed that pine bark as mulch did not significantly affect the trunk diame-
ter and height of ornamental trees. Iles et al. [43] demonstrated that organic mulches had no
effect on the trunk diameter and height of red maple trees two-years after mulching. However,
studies have also reported that mulching, especially with organic mulches, can improve growth
and yield of plants [5, 44, 45]. The results observed in the present study might be attributed to
the soil layer, where the tree roots were growing, not being affected by mulching. This can be
validated by the physical and chemical properties of the soil, which did not differ significantly
among the treatments at 20–40 cm, except for the soil moisture content. The ANOVA revealed that the trunk diameter and height of S. japonica was not affected by dif-
ferent mulches. None of the treatments increased the trunk diameter and tree height signifi-
cantly (Fig 4). Several studies have reported that mulching had no effect on tree growth. Furthermore, factors, such as high average age of the trees, low inter-tree spacing distance,
and lack of fertilizer, might have limited tree growth to some extent in urban tree pits. There-
fore, further studies are necessary in this direction. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 9 / 12 Mulches, soil and tree growth Fig 4. Annual increase in trunk diameter and tree height of trees in tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC) and turf grass (TG). Different letters above the error bars indicate significant differences between treatments (p<0.05, n = 3). The error bars indicate standard deviation. Fig 4. Annual increase in trunk diameter and tree height of trees in tree pits treated with bare soil (CK), cobblestones (CB), water permeable bricks (WPB), pine
barks (PB), green waste compost (GWC) and turf grass (TG). Different letters above the error bars indicate significant differences between treatments (p<0.05, n = 3). The error bars indicate standard deviation. https://doi.org/10.1371/journal.pone.0210777.g004 Acknowledgments We thank Ms. Lu Zhang at Beijing Forestry University for her advice in this research. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 References 1. Kader MA, Senge M, Mojid MA, Ito K. Recent advances in mulching materials and methods for modify-
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ment and development in ornamental prairie plantings. Plos One. 2017; 12 (2): e0171533. https://doi. org/10.1371/journal.pone.0171533 PMID: 28166269 12. Koski R, Jacobi WR. Tree pathogen survival in chipped wood mulch. J Arboric. 2004; 30(3): 165–171. 13. Cook HF, Valdes GSB, Lee HC. Mulch effects on rainfall interception, soil physical characteristics and
temperature under Zea mays L. Soil Till Res. 2006; 91 (1–2): 227–235. 14. Writing – review & editing: Bingpeng Qu, Yuanxin Liu. Writing – review & editing: Bingpeng Qu, Yuanxin Liu. Author Contributions Conceptualization: Bingpeng Qu, Xiangyang Sun. Data curation: Bingpeng Qu, Binghui Yun. Formal analysis: Bingpeng Qu. Funding acquisition: Bingpeng Qu, Suyan Li. Investigation: Bingpeng Qu. Methodology: Bingpeng Qu, Suyan Li, Xinyu Wang. Project administration: Bingpeng Qu, Suyan Li. Resources: Bingpeng Qu, Hua Zhang. Software: Bingpeng Qu, Xinyu Wang, Kaiyi Xiong. Conceptualization: Bingpeng Qu, Xiangyang Sun. Conceptualization: Bingpeng Qu, Xiangyang Sun. 10 / 12 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
February 6, 2019 Mulches, soil and tree growth Supervision: Bingpeng Qu, Suyan Li. Validation: Bingpeng Qu. Visualization: Bingpeng Qu. Writing – original draft: Bingpeng Qu. Writing – review & editing: Bingpeng Qu, Yuanxin Liu. Supervision: Bingpeng Qu, Suyan Li. Validation: Bingpeng Qu. Visualization: Bingpeng Qu. Writing – original draft: Bingpeng Qu. Writing – review & editing: Bingpeng Q Writing – original draft: Bingpeng Qu. PLOS ONE | https://doi.org/10.1371/journal.pone.0210777
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Flame red maple trees. J Arboric. 1999; 25: 163–167. 44. Liedgens M, Soldati A, Stamp P. Interactions of maize and Italian ryegrass in a living mulch system: (1)
Shoot growth and rooting patterns. Plant Soil. 2004; 262 (1–2): 191–203. 45. Jafari M, Haghighi JAP, Zare H. Mulching impact on plant growth and production of rainfed fig orchards
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February 6, 2019
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Disparities in cervical and breast cancer mortality in Brazil
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Rev Saúde Pública 2014;48(3):459-467 Rev Saúde Pública 2014;48(3):459-467 DOI:10.1590/S0034-8910.2014048005214 Original Articles Disparities in cervical and
breast cancer mortality in Brazil Vania Reis GirianelliI
Carmen Justina GamarraII
Gulnar Azevedo e SilvaIII ABSTRACT OBJECTIVE: To analyze cervical and breast cancer mortality in Brazil
according to socioeconomic and welfare indicators. METHODS: Data on breast and cervical cancer mortality covering a 30-year
period (1980-2010) were analyzed. The data were obtained from the National
Mortality Database, population data from the Brazilian Institute of Geography
and Statistics database, and socioeconomic and welfare information from the
Institute of Applied Economic Research. Moving averages were calculated,
disaggregated by capital city and municipality. The annual percent change
in mortality rates was estimated by segmented linear regression using the
joinpoint method. Pearson’s correlation coefficients were conducted between
average mortality rate at the end of the three-year period and selected
indicators in the state capital and each Brazilian state. RESULTS: There was a decline in cervical cancer mortality rates throughout
the period studied, except in municipalities outside of the capitals in the North
and Northeast. There was a decrease in breast cancer mortality in the capitals
from the end of the 1990s onwards. Favorable socioeconomic indicators were
inversely correlated with cervical cancer mortality. A strong direct correlation
was found with favorable indicators and an inverse correlation with fertility
rate and breast cancer mortality in inner cities. I Plataforma de Pesquisa Clínica. Instituto
Nacional de Infectologia Evandro Chagas. Fundação Oswaldo Cruz. Rio de Janeiro,
RJ, Brasil I Plataforma de Pesquisa Clínica. Instituto
Nacional de Infectologia Evandro Chagas. Fundação Oswaldo Cruz. Rio de Janeiro,
RJ, Brasil I Plataforma de Pesquisa Clínica. Instituto
Nacional de Infectologia Evandro Chagas. Fundação Oswaldo Cruz. Rio de Janeiro,
RJ, Brasil CONCLUSIONS: There is an ongoing dynamic process of increased risk of
cervical and breast cancer and attenuation of mortality because of increased,
albeit unequal, access to and provision of screening, diagnosis and treatment. II Instituto Latino-americano de Ciências da
Vida e da Natureza. Universidade Federal
da Integração Latino-americana. Foz do
Iguaçu, PR, Brasil II Instituto Latino-americano de Ciências da
Vida e da Natureza. Universidade Federal
da Integração Latino-americana. Foz do
Iguaçu, PR, Brasil DESCRIPTORS: Breast Neoplasms, mortality. Uterine Cervical
Neoplasms, mortality. Health Services Accessibility. Time Series Studies. III Departamento de Epidemiologia. Instituto
de Medicina Social. Universidade do Estado
do Rio de Janeiro. Rio de Janeiro, RJ, Brasil RESUMO OBJETIVO: Analisar a evolução da mortalidade por câncer do colo uterino
e de mama no Brasil, segundo indicadores socioeconômicos e assistenciais. MÉTODOS: Foram analisados dados agregados de 30 anos (1980-
2010) de mortalidade por câncer de mama e colo uterino. Os dados de
óbitos foram extraídos do Sistema de Informações sobre Mortalidade,
os denominadores populacionais, do Instituto Brasileiro de Geografia e
Estatística, e os indicadores socioeconômicos e assistenciais do Instituto
de Pesquisa Econômica e Aplicada. Foram calculadas as médias móveis
desagregadas por capitais e municípios do interior dos estados. O
percentual de mudança anual das taxas foi estimado a partir da regressão
linear segmentada por joinpoint. Foi feita correlação de Pearson entre as
taxas médias trienais do final do período e os indicadores selecionados
das capitais e de cada estado brasileiro. RESULTADOS: Houve queda da mortalidade por câncer do colo uterino em
todo o período, exceto em municípios das regiões Norte e Nordeste fora das
capitais. Houve declínio na mortalidade por câncer de mama nas capitais a
partir do final da década de 1990. Os indicadores socioeconômicos positivos
correlacionaram-se inversamente com a mortalidade de câncer do colo uterino. Observou-se forte correlação direta entre indicadores positivos e inversa com
a taxa de fecundidade e a mortalidade por câncer de mama nos municípios do
interior dos estados. CONCLUSÕES: Encontra-se em curso um mecanismo dinâmico entre aumento
de risco por câncer de mama e do colo uterino com atenuação da mortalidade
em função da expansão de oferta e acesso ao rastreamento, diagnóstico e
tratamento, porém de forma desigual. DESCRITORES: Neoplasias da Mama, mortalidade. Neoplasias do Colo
do Útero, mortalidade. Acesso aos Serviços de Saúde. Estudos de Séries
Temporais.. Correspondence: Correspondence:
Vania Reis Girianelli
Av. Brasil, 4365 Manguinhos
21040-360 Rio de Janeiro, RJ, Brasil
E-mail: vania.girianelli@ipec.fiocruz.br Article available from: www.scielo.br/rsp 460 Disparities in cancer mortality in females
Girianelli VR et al RESUMO a Ministério da Saúde, Departamento de Informática do Sistema Único de Saúde. Informações das estatísticas de mortalidade e demográficas
[cited 2013 Jan]. Available from: www2.datasus.gov.br/DATASUS/index.php?area=02 INTRODUCTION Using
these data to project expected number of new cases in
this country, more than 50,000 women would be diag-
nosed with breast cancer annually, and around 20,000
with cervical cancer. Analysis of temporal trends in mortality between 1980
and 2006 indicate that there are differentiated patterns to
these cancers in Brazil, with a fall in cervical cancer and
an increase in breast cancer.4 When data for state capi-
tals and other municipalities are disaggregated, these
trends are shown to have different inclinations. The
decrease in cervical cancer in women in the Southeast,
South and Central-West is clear. However, in the North
and Northeast, such a fall is only apparent in the state
capitals and rates in residents in the interior show statis-
tically significant increases. Correction for breast and cervical cancer deaths was
conducted by proportionally redistributing 50.0% of
deaths classified as “undefined” (codes: 780-799 of the
ICD-9 and R00-R99 of the ICD-10), using the World
Health Organization (WHO) proportional redistribution
methodology.c Correction factors were calculated for
each five-year-period, age and sex for capital and inte-
rior of each state. Cervical cancer deaths were corrected
by redistributing all deaths classified as non-specified
uterine cancer (codes 179 of the ICD-9 and C55 of the
ICD-10), maintaining the same proportion of deaths
from cervical and uterine cancer.9 During the period, increases in mortality from breast
cancer were verified in all five regions of Brazil, although
a declining trend in the coefficients was observed in state
capitals from the end of the 1990s onwards. The great challenge for middle and low income coun-
tries is to ensure strategies that enable early diagnosis
of these two types of cancer and to reduce trends in
mortality. It would be possible to reduce cervical cancer
incidence if all women with premalignant lesions had
access to appropriate treatment. Average breast and cervical cancer mortality rates
(to the order of three) were calculated and standard-
ized, corrected for age group using the direct method,
taking the world population standard as a reference.d
The time trend was evaluated using joinpoint regres-
sion, version 3.4.3. Year of death was the independent
variable and mortality rates the dependent variable,
according to the following groupings: large regions,
state capitals and other municipalities (interior). INTRODUCTION decreased number of pre-clinical cases detected through
screening, which began over 20 years ago.12 Breast and cervical cancer are the most common malignant
neoplasms in women worldwide. Breast cancer, together
with lung and colorectal cancer, are among the most
common types in high income countries, while uterine
cancer exceeds other types in low income countries.6 The introduction of cervical cancer screening in devel-
oped countries proved that this measure significantly
reduces incidence of, and mortality from, this disease
and prolongs patient survival. However, this has not
been observed in low income countries where access
to primary and specialized care is limited.16 Increased incidence of breast cancer in various countries
can be partially explained by demographic and lifestyle
changes affecting reproductive factors, such as delayed
first pregnancy, having fewer children and shorter duration
of breastfeeding.17 Over the last few decades, mortality
has declined in developed countries such as the USA, the
United Kingdom, France and Australia. The fall in inci-
dence in the USA from the year 2000 onwards is attributed
to the reduction in hormone replacement therapy and the In Brazil, the Sistema de Informações sobre Mortalidade
(SIM – National Mortality Database) recorded 12,705
deaths from breast cancer and 4,986 from cervical
cancer in 2010, together accounting for 21.4% of
cancer deaths in that year.a The indicators would be 461 Rev Saúde Pública 2014;48(3):459-467 Between 1980 and 1995, the deaths included were
those classified using the Manual of the International
Statistical Classification of Diseases, Injuries, and
Causes of death (ICD-9); and those occurring between
1996 and 2010, classified using the International
Statistical Classification of Diseases and Related
Health Problems (ICD-10). Breast cancer and
cervical cancer correspond to codes: 174(ICD-9) and
C50(ICD-10), 180(ICD-9) and C53(ICD-10), respec-
tively, reported to the SIM and classified under these
codes after correction. Between 1980 and 1995, the deaths included were
those classified using the Manual of the International
Statistical Classification of Diseases, Injuries, and
Causes of death (ICD-9); and those occurring between
1996 and 2010, classified using the International
Statistical Classification of Diseases and Related
Health Problems (ICD-10). Breast cancer and
cervical cancer correspond to codes: 174(ICD-9) and
C50(ICD-10), 180(ICD-9) and C53(ICD-10), respec-
tively, reported to the SIM and classified under these
codes after correction. greater than those officially released if corrections
referring to “undefined” causes and cases classified
as “non-specified area of uterus” were made. b Instituto de Pesquisa Econômica Aplicada. Indicadores sociais [cited 2013 Jan]. Available from: www.ipeadata.gov.br
c Mathers CD, Bernard C, Iburg KM, Inoue M, Fat DM, Shibuya K, Stein C, Tomijima N, Xu H. Global Burden of Disease in 2002: data
sources, methods and results. Global Programme on Evidence for Health Policy Discussion Paper No. 54 World Health Organization
December 2003 (revised February 2004).
d Segi M. Cancer mortality for selected sites in 24 countries (1950-57). Sendai: Tohoku University School of Public Health; 1960. INTRODUCTION The
models were adjusted assuming a different number of
joinpoints, from zero (trend represented by one single
segment of the line) to three; considering changes in
the rates over time. Annual percentage changes (APC)
were calculated for each type of cancer, period and
geographic area. It is essential that trends in incidence and mortality are
monitored in order to evaluate the results of screening
strategies which have been shown to be effective in
other countries. The aim of this study was to analyze the evolution of
breast and cervical cancer mortality in state capitals and
other Brazilian municipalities, according to socioeco-
nomic and health care indicators. The adjusted and corrected mean rates for the last
three-year-period (2008 to 2010) were correlated
with socioeconomic and health care indicators avail-
able for 2000 in the IPEA for state capitals and states. The indicators were classed as positive or negative. The positive corresponded to better living condi-
tions: number of doctors/1,000 inhabitants, except
for Teresina, PI, where this indicator was not avail-
able; proportion of individuals aged ≥ 25 with at
least 11 years of schooling; proportion of individ-
uals living in households with electricity; human
development index (HDI); proportion of individuals
living in households with plumbing. The negative The adjusted and corrected mean rates for the last
three-year-period (2008 to 2010) were correlated
with socioeconomic and health care indicators avail-
able for 2000 in the IPEA for state capitals and states. METHODS This is a study of breast and cervical cancer mortality
in state capitals and other municipalities in the five
Brazilian regions, using aggregated time series data
from a 30-year-period (1980-2010). The data on
deaths were obtained from the SIM, population data
from Brazilian Institute of Geography and Statistics
(IBGE) demographic censusesa and socioeconomic
and health care indicators from the Institute of Applied
Economic Research (IPEA).b Population data between
the censuses were estimated using linear interpolation. The indicators were classed as positive or negative. The positive corresponded to better living condi-
tions: number of doctors/1,000 inhabitants, except
for Teresina, PI, where this indicator was not avail-
able; proportion of individuals aged ≥ 25 with at
least 11 years of schooling; proportion of individ-
uals living in households with electricity; human
development index (HDI); proportion of individuals
living in households with plumbing. The negative 462 Disparities in cancer mortality in females
Girianelli VR et al cancer increased evenly in all municipalities. This
profile differed in the Northeast, where breast cancer
mortality rates exceeded those of cervical cancer at the
end of the period, with a marked increase in women
living in the interior. Cervical cancer mortality coef-
ficients in the South and Southeast were lower than
those of breast cancer and there was a clear trend of
decline in the female population as a whole. A decline
in deaths from breast cancer was noted in the state
capitals of the South from the mid-1990s onwards,
and in the Southeast from the end of that decade. In
the Central-West, throughout the period, there was an
increase in breast cancer mortality and a fall in cervical
cancer mortality. The speed of the advance of breast
cancer among women living outside the state capitals
was considerable. indicators corresponded to worse conditions: propor-
tion of population aged ≥ 25 who are illiterate;
fertility rate; proportion of individuals living below
the poverty line, i.e., on under half a minimum wage
per capita; mortality rate in children < 5/1,000 live
births; percentage of households headed by women,
without partner and with children < 15. Pearson’s correlation coefficients were calculated
between the three-year average rates (2008-2010) in
the Federal District and the state capitals and other
municipalities in each state and the indicators in the
capitals and states, respectively, using the R statistic
program (version 2.7.1). RESULTS In the 1980s, the magnitude of breast cancer mortality
in Brazilian women was similar to that of cervical
cancer. However, breast cancer always weighed
more heavily in state capitals (Figure 1). Patterns in
mortality rates for the two types of cancer in Brazil
differed over the 30 years: a clear fall in cervical
cancer, more pronounced in state capitals, and an
increase in breast cancer. From the end of the 1990s,
number of deaths from this cancer began to decrease
in women living in the state capitals. In the 1980s, the magnitude of breast cancer mortality
in Brazilian women was similar to that of cervical
cancer. However, breast cancer always weighed
more heavily in state capitals (Figure 1). Patterns in
mortality rates for the two types of cancer in Brazil
differed over the 30 years: a clear fall in cervical
cancer, more pronounced in state capitals, and an
increase in breast cancer. From the end of the 1990s,
number of deaths from this cancer began to decrease
in women living in the state capitals. The patterns of breast cancer mortality trends differed
between the regions and between the state capitals and
other municipalities (Table 2). There was an overall
increase in rates in the first observation period, with
the exception of in the interior of the North. Mortality
rates for this cancer continued to grow in the interior
of the North and Northeast and in the state capitals in
the North. However, the rates began to fall in other
regions, both in the state capitals and outside of them. The greatest growth was observed in the North, in
municipalities in the interior, between 1999 and 2010
(APC = 5.0; 95%CI 3.3;6.8) and in the interior of the
Northeast (APC = 4.9; 95%CI 4.4;5.5). The greatest
annual drop was observed between 1997 and 2002 in
the Southeast (APC = -2.8; 95%CI -3.7;-1.9). The same comparison with data disaggregated by
region shows a difference in magnitude and trends of
mortality for these two types of cancer (Figure 2). In
the North, cervical cancer mortality was higher than
that of breast cancer throughout the period, and there
was a fall in deaths among women living in the state
capitals and an increase in those in the interior. Breast a per 100,000 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corrected (see Methods). METHODS Values between 0.70 and 1
were deemed strong correlation, moderate 0.30 to 0.69
and weak 0 to 0.29. The research was approved by the Research Ethics
Committee of the Instituto de Medicina Social,
Universidade do Estado do Rio de Janeiro (Process
CAAE 0027.0.259.000-09, 8/12/2009), and followed
ethical standards. Regression analysis using joinpoint showed a signifi-
cantly statistic drop in mortality rates from cervical
cancer in the state capitals and other municipalities in
the Southeast, South and Central-West (Table 1). In
the North and Northeast, there was a fall in the state
capitals and an increase in the interior. The largest
drop was observed in state capitals in the Northeast
(APC -3.3; 95%CI -3.5;-3.1). RESULTS Mortality indicators behaved differently for cervical
cancer in the state capitals: of the five positive indica-
tors, three were negatively correlated, and of the five
negatives, three correlated positively. The indicators
analyzed do not show correlation with cervical cancer
mortality rates in the interior, with the exception of
percentage of individuals living below the poverty line
(r = 0.46; p = 0.017). Correlation analysis was conducted between some
socioeconomic and health care positive and negative
indicators (doctors/1,000 inhabitants) and mortality
rates in residents in the state capitals or other munici-
palities in the states. A moderate direct relationship
between three positive indicators and an inverse rela-
tionship, also moderate, were found with breast cancer. In the interior, there were strong correlations with
almost all the positive indicators and, of the negative
indicators, there was a strong correlation with fertility
(r = -0.92; p < 0.0001) and the percentage of female
head of household, single and with children under 15
(r = -0.82; p < 0.0001), with moderate correlation with
the other three (Table 3). RESULTS Cervical cancer
Breast cancer
Other municipalities
Brazil
Capitals Breast cancer Breast cancer Breast cancer Breast cancer Cervical cancer Other municipalities
Brazil
Capitals a per 100,000 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corrected (see Methods). Figure 1. Breast and cervical cancer mortalitya. Brazil, overall and by state capitals and other municipalities, 1980 to 2010. Figure 1. Breast and cervical cancer mortalitya. Brazil, overall and by state capitals and other municipalities, 1980 to 2010. 463 Rev Saúde Pública 2014;48(3):459-467 Figure 2. Breast and cervical cancer mortality.a Brazil, overall and by state capitals and other municipalities, 1980 to 2010. a per 100,000 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corrected (see Methods). Other municipalities
Brazil
North
Cervical cancer
Breast cancer
Northeast
Cervical cancer
Breast cancer
Southeast
Cervical cancer
Breast cancer
South
Cervical cancer
Breast cancer
Central-West
Cervical cancer
Breast cancer
Capitals Breast cancer Breast cancer Breast cancer Breast cancer Cervical cancer Breast cancer Breast cancer Central-West
Cervical cancer Breast cancer Brazil
Capitals a per 100,000 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corrected (see Methods). a per 100,000 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corre 0 inhabitants, adjusted for the population of the demographic census – Brazil 2010 and corrected (see Methods). Figure 2. Breast and cervical cancer mortality.a Brazil, overall and by state capitals and other municipalities, 1980 to 2010. 464 Disparities in cancer mortality in females
Girianelli VR et al Table 1. Joinpoint regression analysis of cervical cancer mortality trends. State capitals and other municipalities in the Brazilian
regions, 1980 to 2010. Region
Municipalities
Trend 1
Years
APC
95%CI
North
Capitals
1980 to 2010
-1.8
-2.0;-1.6
Other municipalities
1980 to 2010
1.7
1.2;2.2
Northeast
Capitals
1980 to 2010
-3.3
-3.5;-3.1
Other municipalities
1980 to 2010
1.0
0.8;1.3
Southeast
Capitals
1980 to 2010
-1.9
-2.2;-1.7
Other municipalities
1980 to 2010
-2.2
-2.5;-2.0
South
Capitals
1980 to 2010
-2.7
-3.1;-2.2
Other municipalities
1980 to 2010
-1.8
-2.1;-1.4
Central-West
Capitals
1980 to 2010
-3.1
-3.4;-2.8
Other municipalities
1980 to 2010
-1.1
-1.6;-0.6
Brazil
Capitals
1980 to 2010
-2.2
-2.4;-2.0
Other municipalities
1980 to 2010
-1.0
-1.2;-0.9
APC: annual percentage change. npoint regression analysis of cervical cancer mortality trends. State capitals and other municipalities in the Brazilian
80 to 2010 APC: annual percentage change. DISCUSSION The results suggest there is a dynamic mechanism
between determining risk exposures in the appear-
ance of breast and cervical cancer. At the same time, Table 2. Joinpoint regression analysis of breast cancer mortality trends. State capitals and other municipalities in the Brazilian
regions, 1980 to 2010. Region
Municipalities
Trend 1
Trend 2
Trend 3
Years
APC
95%CI
Years
APC
95%CI
Years
APC
95%CI
North
Capitals
1980 to 1984 -2.1 -7.3;3.4 1984 to 2010
1.0
0.6;1.3
–
–
–
Other municipalities 1980 to 1999
3.1
2.4;3.9 1999 to 2010
5.0
3.3;6.8
–
–
–
Northeast
Capitals
1980 to 1992
0.4
-0.3;1.1 1992 to 2010 -0.5 -0.8;-0.1
–
–
–
Other municipalities 1980 to 1997
1.5
1.2;1.9 1997 to 2010
4.9
4.4;5.5
–
–
–
Southeast
Capitals
1980 to 1997
0.9
0.8;1.0 1997 to 2002 -2.8 -3.7;-1.9 2002 to 2010 -0.6 -0.9;-0.2
Other municipalities 1980 to 2010
0.6
0.4;0.8
–
–
–
–
–
–
South
Capitals
1980 to 1991
1.6
1.1;2.2 1991 to 2010 -1.3 -1.5;-1.1
–
–
–
Other municipalities 1980 to 1993
1.9
1.6;2.2 1993 to 2010 -0.2
-0.4;0.0
–
–
–
Central-West
Capitals
1980 to 1996
1.4
1.0;1.8 1996 to 2010 -0.9 -1.4;-0.4
–
–
–
Other municipalities 1980 to 2010
2.4
2.0;2.8
–
–
–
–
–
–
Brazil
Capitals
1980 to 1997
0.7
0.6;0.8 1997 to 2003 -2.6 -3.2;-1.9 2003 to 2010 0.0
-0.4;0.5
Other municipalities 1980 to 2010
1.1
1.0;1.2
–
–
–
–
–
–
APC: annual percentage change gression analysis of breast cancer mortality trends. State capitals and other municipalities in the Brazilian
10 465 Rev Saúde Pública 2014;48(3):459-467 Table 3. Correlation between breast and cervical cancer mortality rates and selected socioeconomic and health care indicators. Capitals and other municipalities in Brazil, 2008 to 2010. DISCUSSION Breast cancer
Cervical cancer
Indicators
Capitals
Other municipalities
Capitals
Other municipalities
Correlation
p
Correlation
p
Correlation
p
Correlation
p
Positive
Human development index (HDI)
0.32
0.0990
0.58
0.0020
-0.38
0.0502
-0.38
0.0585
Doctors (per thousand inhabitants)a
0.61
0.0008
0.81
< 0.0001
-0.51
0.0084
-0.29
0.1581
% individuals aged ≤ 25 with more
than 11 years of schooling
0.51
0.0063
0.78
< 0.0001
-0.57
0.0018
-0.37
0.0636
% individuals in households with
electricity
0.36
0.0635
0.71
< 0.0001
-0.17
0.3828
-0.36
0.0745
% individuals in households with
plumbing
0.53
0.0047
0.82
< 0.0001
-0.40
0.0404
-0.36
0.0689
Negative
% population aged ≤ 25 who are
illiterate
-0.22
0.2764
-0.47
0.0155
0.18
0.3588
0.27
0.1893
Fertility rate (%)
-0.59
0.0013
-0.92
< 0.0001
0.52
0.0056
0.35
0.0754
% individuals living below the
poverty line
-0.26
0.1878
-0.62
0.0007
0.45
0.0198
0.46
0.0173
Mortality rate in children < 5 per
1,000 LB
-0.16
0.4283
-0.40
0.0429
0.36
0.0613
0.39
0.0508
% female head of household,
single and with children aged ≤ 15
-0.44
0.0223
-0.82
< 0.0001
0.33
0.0895
0.24
0.2421
LB: live births
a Except for Teresina, PI, as the indicator was not available. a Except for Teresina, PI, as the indicator was not available. due to access to diagnostic and treatment measures,
more effective at the initial stages of the disease. mortality is attenuated due to better access to diagnostic
and treatment measures. The fall in cervical cancer
mortality occurred in women all over the Southeast
and South, the most developed regions of the country,
whereas for women in the North and Northeast it only
occurred in the state capitals. Likewise, deaths from
breast cancer began to decrease in women living in
the state capitals in the South and Southeast from the
mid-1990s onwards. Brazilian studies have shown overall rates of HPV infec-
tion varying between 13.7% and 54.3%.3 The number of
women infected is growing in all regions of the country,
probably accompanying early onset of sexual activity.e This is a malignant neoplasm that can be prevented by
screening for intraepithelial neoplastic lesions. Thus, the
fall in mortality that began in the more developed regions
of Brazil, moving thence to the state capitals in other
regions, indicates certain success in screening interven-
tions. The increased availability of the Papanicolaou test
may have partially reversed the risk of death in women
with access to preventative measures. e Ministério da Saúde. Pesquisa Nacional Demografia e Saúde da Criança e da Mulher (PNDS-2006). Brasília (DF); 2008.
f Instituto Nacional do Câncer. Viva Mulher. Câncer do Colo do Útero: informações técnico-gerenciais e ações desenvolvidas. Rio de Janeiro:
INCA; 2002. g Lago TG. Políticas nacionais de rastreamento do câncer do colo uterino no Brasil: análise do período 1998 a 2002 [doctoral the
Campinas: Instituto de Filosofia e Ciências Humanas da UNICAMP; 2004. e Ministério da Saúde. Pesquisa Nacional Demografia e Saúde da Criança e da Mulher (PNDS-2006). Brasília (DF); 2008.
f Instituto Nacional do Câncer. Viva Mulher. Câncer do Colo do Útero: informações técnico-gerenciais e ações desenvolvidas. Rio de Janeiro
INCA; 2002.
g Lago TG. Políticas nacionais de rastreamento do câncer do colo uterino no Brasil: análise do período 1998 a 2002 [doctoral thesis].
Campinas: Instituto de Filosofia e Ciências Humanas da UNICAMP; 2004. DISCUSSION Although some breast cancer screening proposals were
included in the intensification stage of the Viva Mulher
program in 2002, it was in 2004 that the Ministry of
Health launched the breast cancer control consensus,
recommending that women aged between 50 and 69
have a mammogram every two years.j Data from household surveys confirm an increase in
preventative examinations that became part of national
policy for controlling cancer over the last decade. Data
from the Pesquisa Nacional de Amostras de Domicílio
(PNAD – National Household Survey) estimated 82.6%
coverage of preventative gynecological examinations
in women aged 25 to 59 in 2003.h This estimate was
87.1% in 2008, an increase across all income classes,
although of a more marked nature among the poorest. PNAD datag from 2003 and 2008 confirm increased
numbers of mammograms in women in the Ministry of
Health target age group (50 to 69 years old), especially
among those on higher incomes and with higher levels
of education and who have health insurance. Although
the percentage of women between 50 and 69 who
reported having a mammogram increased from 54.6%
in 2003 to 71.5% in 2008, among those with household
income > 5 minimum wages, the increase was from
76.6% to 80.7%. These percentages were 20.2% in 2003
and 29.1% in households on an income of < 25.0%
minimum wage in 2008, demonstrating considerable
inequality in access by socioeconomic level. Residents
in metropolitan areas are three times more likely to have
the examination than those in other areas.15 Correlation analysis confirms what has been shown
in the literature.4,6,9 Cervical cancer mortality coeffi-
cients in the state capitals are inversely correlated with
indicators of better socioeconomic status and directly
correlated with negative indicators. This indicates that
it is more difficult for women living in state capitals to
have cervical cancer screening tests done. In the inte-
rior, only the percentage of individuals living below
the poverty line was related to increased mortality
from this type of cancer. Are we faced with a situation
in which women, outside of urban centers, who do not
undergo cervical cancer screening are living in worse
conditions? Is inequality in access concentrated in those
who live in extreme poverty, while barriers also affect
others on low incomes in the capitals? DISCUSSION Declining cervical cancer mortality may reflect the
protection provided by the Pap smear test. This was
not observed in the interior of the North and Northeast. Breast cancer mortality increased, partly, due to
increased incidence of the disease. This may be
explained by the increasing adoption of urban life-
styles among women, which directly affects the degree
of exposure to reproductive factors. The large annual
increase in mortality rates in the North and Northeast
suggests that changes in reproductive and sexual
behavior, initiated in large urban centers, were rapidly
adopted by women in other municipalities, even those
in low income areas. On the other hand, the reversal in
mortality rates that began in the capitals was possible Actions to control cervical cancer began in the 1980s. They intensified between 1999 and 2002, period in
which two campaigns of the Viva Mulher program were
conducted,f leading to a substantial increase in provi-
sion of the Papanicolaou test in all regions of Brazil.g However, a fall in mortality from this cancer had been
detected in several states even before these initiative 466 Disparities in cancer mortality in females
Girianelli VR et al Mortality accompanied this increase, which occurred
homogeneously in women in different regions of the
country. Lifestyle changes appear to be reflected in
reproductive factors and rapidly affect women in the
interior. Decreased breast cancer deaths in the South
and Southeast may show the results of better access to
mammography and specialist treatment services, bene-
fitting women diagnosed when the disease is in its early
stages. In Brazil, this phenomenon is more recent than
the Pap smear examination. commenced. Decreases in cervical cancer deaths had
been recorded in the municipality of Sao Paulo between
1980 and 1999,8 between 1984 and 20032 and between
1987 and 1998 in the municipalities of Campinas,
Piracicaba and Sao Joao da Boa Vista,18 all in the
state of Sao Paulo. Similar declines were identified in
Paraná5 and in Minas Gerais between 1980 and 2005.1
Actions to organize screening for the disease at a state
and local level, in the case of Sao Paulo and Paraná,
may have encouraged the start of the fall in mortality
in the South and Southeast. h Instituto Brasileiro de Geografia e Estatística. Pesquisa Nacional por Amostra de Domicílios: um panorama da saúde no Brasil: acesso e
utilização dos serviços, condições de saúde e fatores de risco e proteção à saúde - PNAD 2008. Rio de Janeiro; 2010.
i Minamiguchi MM. Segunda transição demográfica: o que se pode dizer da situação brasileira? [dissertação de mestrado]. Rio de Janeiro:
Escola Nacional de Ciências Estatísticas; 2011.
j Instituto Nacional de Câncer, Coordenação de Prevenção e Vigilância. Controle do câncer de mama no Brasil: documento de consenso. Rio
de Janeiro; 2004 [cited 2012 Dec]. Available from: http://www1.inca.gov.br/publicacoes/Consensointegra.pdf DISCUSSION The study was based on SIM data, which could pose
a significant limitation due to regional differences in
the coverage, completeness and quality of the informa-
tion.7 However, previous studies indicate that cancer
deaths are better recorded, although some of them may
be included in the undefined causes categories, which
have decreased markedly in this country.13 The deci-
sion to correct 50.0% of undefined deaths, together
with moving averages and joinpoint regression meant
that the trends in cancer mortality coefficients could
be approximated. Breast cancer mortality rates rose in Brazil according
to increased positive socioeconomic indicators and
decreased as negative indicators, such as fertility rates,
increased. This relationship appears better in the inte-
rior, suggesting that rates outside of the capitals were
not attenuated by women diagnosed with the disease
receiving treatment. The effect of screening on reducing breast cancer
mortality is lower than that which occurred with
cervical cancer. Recent studies indicate that the impact
of mammography screening on mortality is less than
hoped10 and that the large reduction in mortality in
developed countries is attributable to recent advances
in treatment.11 This reality is compatible with countries The main breast cancer risk factors are related to sexual
and reproductive history.14 The fall in mortality in devel-
oped countries is attributed to improved survival due to
awareness, early detection and improved treatment.11
The demographic transition taking place in Brazil, with
falling birth rates and having children later,i weighs
significantly in increased incidence of the disease. 467 Rev Saúde Pública 2014;48(3):459-467 Urgent measures are needed to guarantee that all
women have access to the best diagnosis and treat-
ment available. Urgent measures are needed to guarantee that all
women have access to the best diagnosis and treat-
ment available. in which it is not so difficult to access cancer diagnosis
and treatment services. The obstacles are enormous
for Brazil’s socioeconomically deprived population. The authors declare that there is no conflict of interest. REFERENCES 9. Gamarra CJ, Valente JG, Silva GA. Correction for
reported cervical cancer mortality data in Brazil,
1996-2005. Rev Saude Publica. 2010;44(4):629-38. DOI:10.1590/S0034-89102010000400006 1. Alves CMM, Guerra MR, Bastos RR. Tendência
de mortalidade por câncer de colo de útero
para o Estado de Minas Gerais, Brasil, 1980-
2005. Cad Saude Publica. 2009;25(8):1693-700. DOI:10.1590/S0102-311X2009000800005 10. Independent UK Panel on Breast Cancer Screening. The benefits and harms of breast cancer screening: an
independent review. Lancet. 2012;380(9855):1778-86. DOI:10.1016/S0140-6736(12)61611-0 2. Antunes JLF, Wünsch-Filho V. The effect of
performing corrections on reported uterine
cancer mortality data in the city of São Paulo. Braz J Med Biol Res. 2006;39(8):1091-9. DOI:10.1590/S0100-879X2006000800012 11. Jatoi I, Miller AB. Why is breast-cancer mortality
declining? Lancet Oncol. 2003;4(4):251-4. DOI:10.1016/S1470-2045(03)01037 3. Ayres ARG, Azevedo e Silva G. Prevalência de
infecção do colo do útero pelo HPV no Brasil: revisão
sistemática. Rev Saude Publica. 2010;44(5):963-74. DOI:10.1590/S0034-89102010000500023 12. Jemal A, Center MM, DeSantis C, Ward EM. Global
patterns of cancer and mortality rates and trends. Cancer
Epidemiol Biomarkers Prev. 2010;19(8):1893-907. DOI:10.1158/1055-9965.EPI-10-0437 4. Azevedo e Silva G, Gamarra CJ, Girianelli VR,
Valente JG. Tendência da mortalidade por câncer
nas capitais e interior do Brasil entre 1980 e
2006. Rev Saude Publica. 2011;45(6):1009-18. DOI:10.1590/S0034-89102011005000076 13. Jorge MHPM, Gotlieb SLD, Laurenti R. O sistema de
informações sobre mortalidade: problemas e propostas
para o seu enfrentamento. I - Mortes por causas
naturais. Rev Bras Epidemiol. 2002;5(2):197-211. DOI:10.1590/S1415-790X2002000200007 5. Bleggi Torres LF, Werner B, Totsugui J, Collaço
LM, Araújo, Huçulak M, et al. Cervical cancer
screening program of Paraná: cost-effective model
in a developing country. Diagn Cytopathol. 2003;29(1):49-54. DOI:10.1002/dc.10269 14. Kobayashi S, Sugiura H, Ando Y, Shiraki N, Yanagi
T, Yamashita H, et al. Reproductive history and
breast cancer risk. Breast Cancer. 2012;19(4):302-8. DOI:10.1007/s12282-012-0384-8 6. Bray F, Jemal A, Grey N, Ferlay J, Forman D. Global cancer transitions according to the Human
Development Index (2008-2030): a population-
based study. Lancet Oncol. 2012;13(8):790-801. DOI:10.1016/S1470-2045(12)70211-5 15. Oliveira EXG, Pinheiro RS, Melo ECP, Carvalho MS.. Condicionantes socioeconômicos e geográficos
do acesso à mamografia no Brasil, 2003-2008. Cienc Saude Coletiva. 2011;16(9):3649-64. DOI:10.1590/S1413-81232011001000002 7. Felix JD, Zandonade E, Amorim MHC, Castro
DS. Avaliação da completude das variáveis
epidemiológicas do Sistema de Informação sobre
Mortalidade em mulheres com óbitos por câncer
de mama na Região Sudeste: Brasil (1998 a
2007). Cienc Saude Coletiva. 2012;17(4):945-53. DOI:10.1590/S1413-81232012000400016 16. Pecorelli S, Favalli G, Zigliani L, Odicino F. Cancer in
women. Int J Gynaecol Obstet. 2003;82(3):369-79. DOI:10.1016/S0020-7292(03)00225 17. Porter PL. REFERENCES Global trends in breast cancer incidence and
mortality. Salud Publica Mex. 2009;51(Suppl 2):s141-6. DOI:10.1590/S0036-36342009000800003 17. Porter PL. Global trends in breast cancer incidence and
mortality. Salud Publica Mex. 2009;51(Suppl 2):s141-6. DOI:10.1590/S0036-36342009000800003 8. Fonseca LAM, Ramacciotti AS, Eluf Neto J. Tendência da mortalidade por câncer do
útero no Município de São Paulo entre 1980 e
1999. Cad Saude Publica. 2004;20(1):136-42. DOI:10.1590/S0102-311X2004000100029 18. Zeferino LC. O desafio de reduzir a
mortalidade por câncer do colo do útero. Rev Bras Ginecol Obstet. 2008;30(5):213-5. DOI:10.1590/S0100-72032008000500001 18. Zeferino LC. O desafio de reduzir a
mortalidade por câncer do colo do útero. Rev Bras Ginecol Obstet. 2008;30(5):213-5. DOI:10.1590/S0100-72032008000500001 The authors declare that there is no conflict of interest.
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Remedying Contact Dermatitis in Broiler Chickens with Novel Flooring Treatments
|
Animals
| 2,020
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cc-by
| 14,658
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animals Access to clean litter prevented and remedied contact dermatitis, and a
oach may be commercially feasible. matitis in Broiler Chickens
atments
, Alexa Johnson 1, Victoria Marshall 1, An Garmyn 3
Virginia Polytechnic Institute and State University,
u (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
isheries and Food (ILVO), 9090 Melle, Belgium;
rsity, 9820 Merelbeke, Belgium; an.garmyn@ugent.be
-231-4735
ber 2020; Published: 28 September 2020
overarching term for inflamed or necrotic lesions after
r chickens commonly experience these lesions due to
mmonia within litter. This study aimed to find methods to
hocks, and breast. Furthermore, the impact of treatments
ht was investigated. We applied novel flooring treatments
mats containing povidone-iodine, which we compared
ositive control (clean litter). Weekly measurements on a
impact of both flooring treatments and age (weeks) on
tations, the novel flooring treatments did not prevent or
ve control, consisting of replacing litter every four days,
imited to no contact dermatitis at week seven of age. matitis (FPD), hock burns, and breast dermatitis) is a
pain and behavioral restrictions. Once lesions develop,
for the affected flock. Our objective was to evaluate
l by providing preventative and remedial treatments
, and gait impairment. Broilers (n = 546) were housed
treatments (four) included used litter (NEG), new pine
lled with 1% povidone-iodine solution (MAT), and the
T). Flooring treatments were provided from day one
day 29 (remedial approach; REM). Contact dermatitis,
kly (seven birds/pen). Results showed a treatment effect
Overall, the POS treatment resulted in the best welfare
worst contact dermatitis was found in the MAT and SLAT
rmatitis, opposite to expectations. Weights were lower
tments with povidone-iodine were deemed ineffective
itter prevented and remedied contact dermatitis, and a
feasible. Received: 19 August 2020; Accepted: 23 September 2020; Published: 28 September 2020 Simple Summary: Contact dermatitis is an overarching term for inflamed or necrotic lesions after
contact with an allergen or irritant. Broiler chickens commonly experience these lesions due to
prolonged contact with moisture, feces, and ammonia within litter. This study aimed to find methods to
prevent and remedy lesions on broilers’ feet, hocks, and breast. Furthermore, the impact of treatments
on plumage cleanliness, gait, and body weight was investigated. We applied novel flooring treatments
consisting of plastic slats and disinfectant mats containing povidone-iodine, which we compared
to the industry control (used litter) and a positive control (clean litter). Weekly measurements on a
sample of birds in each pen showed us the impact of both flooring treatments and age (weeks) on
animal welfare outcomes. Contrary to expectations, the novel flooring treatments did not prevent or
remedy contact dermatitis. In fact, the positive control, consisting of replacing litter every four days,
resulted in the best welfare condition, with limited to no contact dermatitis at week seven of age. Abstract: Contact dermatitis (footpad dermatitis (FPD), hock burns, and breast dermatitis) is a
welfare issue for broiler chickens, causing pain and behavioral restrictions. Once lesions develop,
often nothing is done to remedy the issue for the affected flock. Our objective was to evaluate
novel flooring treatments at the flock level by providing preventative and remedial treatments
against contact dermatitis, plumage soiling, and gait impairment. Broilers (n = 546) were housed
in 42 pens, with 13 birds/pen. The flooring treatments (four) included used litter (NEG), new pine
shavings replaced regularly (POS), a mat filled with 1% povidone-iodine solution (MAT), and the
iodine mat placed on a slatted floor (SLAT). Flooring treatments were provided from day one
of age (preventative approach; PREV) or day 29 (remedial approach; REM). Contact dermatitis,
soiling, gait, and weight were recorded weekly (seven birds/pen). Results showed a treatment effect
for all measures, dependent on bird age. Overall, the POS treatment resulted in the best welfare
outcomes (FPD, hock burns, and gait). The worst contact dermatitis was found in the MAT and SLAT
groups. NEG birds showed little contact dermatitis, opposite to expectations. Weights were lower
for PREV-POS in week seven only. The treatments with povidone-iodine were deemed ineffective
against contact dermatitis. Access to clean litter prevented and remedied contact dermatitis, and a
comparable approach may be commercially feasible. Remedying Contact Dermatitis in Broiler Chickens
with Novel Flooring Treatments Nathan Freeman 1, Frank A. M. Tuyttens 2,3
, Alexa Johnson 1, Victoria Marshall 1, An Garmy
and Leonie Jacobs 1,* 1
Department of Animal and Poultry Sciences, Virginia Polytechnic Institute and State University,
Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
2
Flanders Research Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium;
frank.tuyttens@ilvo.vlaanderen.be
3
Faculty of Veterinary Medicine, Ghent University, 9820 Merelbeke, Belgium; an.garmyn@ugent.be
*
Correspondence: jacobsl@vt.edu; Tel.: +1-540-231-4735 1
Department of Animal and Poultry Sciences, Virginia Polytechnic Institute and State University,
Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.) 1
Department of Animal and Poultry Sciences, Virginia Polytechnic Institute and State University,
Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
2
Flanders Research Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium; 1
Department of Animal and Poultry Sciences, Virginia Polytechnic Institute and State University,
Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
2
Flanders Research Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium;
frank.tuyttens@ilvo.vlaanderen.be p
y
g
y
y
Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
2
Flanders Research Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium;
frank.tuyttens@ilvo.vlaanderen.be Blacksburg, VA 24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
2
Flanders Research Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium;
frank.tuyttens@ilvo.vlaanderen.be 3
Faculty of Veterinary Medicine, Ghent University, 9820 Merelbeke, Belgium; an.garmyn@ugent.be
*
Correspondence: jacobsl@vt.edu; Tel.: +1-540-231-4735 y
y
y
g
g
y
g
*
Correspondence: jacobsl@vt.edu; Tel.: +1-540-231-4735 animals animals animals ng Contact Dermatitis in Broiler Chickens
el Flooring Treatments
, Frank A. M. Tuyttens 2,3
, Alexa Johnson 1, Victoria Marshall 1, An Garmyn 3
s 1,*
Animal and Poultry Sciences, Virginia Polytechnic Institute and State University,
24060, USA; nathanf1@vt.edu (N.F.); alexaj3@vt.edu (A.J.); victm97@vt.edu (V.M.)
rch Institute for Agriculture, Fisheries and Food (ILVO), 9090 Melle, Belgium;
@ilvo.vlaanderen.be
rinary Medicine, Ghent University, 9820 Merelbeke, Belgium; an.garmyn@ugent.be
e: jacobsl@vt.edu; Tel.: +1-540-231-4735
st 2020; Accepted: 23 September 2020; Published: 28 September 2020
y: Contact dermatitis is an overarching term for inflamed or necrotic lesions after
allergen or irritant. Broiler chickens commonly experience these lesions due to
t with moisture, feces, and ammonia within litter. This study aimed to find methods to
dy lesions on broilers’ feet, hocks, and breast. Furthermore, the impact of treatments
liness, gait, and body weight was investigated. We applied novel flooring treatments
stic slats and disinfectant mats containing povidone-iodine, which we compared
ontrol (used litter) and a positive control (clean litter). Weekly measurements on a
n each pen showed us the impact of both flooring treatments and age (weeks) on
utcomes. Contrary to expectations, the novel flooring treatments did not prevent or
ermatitis. In fact, the positive control, consisting of replacing litter every four days,
st welfare condition, with limited to no contact dermatitis at week seven of age. ct dermatitis (footpad dermatitis (FPD), hock burns, and breast dermatitis) is a
broiler chickens, causing pain and behavioral restrictions. Once lesions develop,
done to remedy the issue for the affected flock. Our objective was to evaluate
eatments at the flock level by providing preventative and remedial treatments
ermatitis, plumage soiling, and gait impairment. Broilers (n = 546) were housed
13 birds/pen. The flooring treatments (four) included used litter (NEG), new pine
d regularly (POS), a mat filled with 1% povidone-iodine solution (MAT), and the
ed on a slatted floor (SLAT). Flooring treatments were provided from day one
tive approach; PREV) or day 29 (remedial approach; REM). Contact dermatitis,
weight were recorded weekly (seven birds/pen). Results showed a treatment effect
dependent on bird age. Overall, the POS treatment resulted in the best welfare
hock burns, and gait). The worst contact dermatitis was found in the MAT and SLAT
ds showed little contact dermatitis, opposite to expectations. Weights were lower
week seven only. The treatments with povidone-iodine were deemed ineffective
ermatitis. animals animals 1. Introduction Contact dermatitis is an inflammation and irritation of the skin due to contact with an irritant
or allergen. A common type of contact dermatitis in broiler chickens is footpad dermatitis (FPD),
with necrotic lesions on the plantar surface of the central footpad [1]. When these lesions are
left untreated, and environmental conditions remain the same or deteriorate, lesions will worsen,
eventually encompassing the entire footpad, including the toes [2]. The lesions can be associated with
bacterial infections, especially with Staphylococcus aureus and Escherichia coli (E. coli) that can be present
in the litter and on the skin [3]. FPD is a common condition found in commercial broiler chickens. Approximately 50% of heavy broilers (3.6 to 3.8 kg) reared on commercial farms in the Southeastern
United States (US) present with some degree of FPD [4]. Furthermore, studies in Europe showed FPD
in 58% of the assessed commercial broilers [5]. Besides on the feet, similar types of contact dermatitis
can occur on hocks (hock burns) and the abdomen (breast dermatitis) [6]. The most important risk factor for the development of FPD is the litter condition [7]. The litter
moisture and ammonia concentration from built-up fecal material can burn and weaken the dermis of
the footpad [8], with an increased severity of FPD resulting from the prolonged exposure of feet to wet
litter. Moisture causes the outer layer of the dermis to soften, posing a risk of microbial contamination,
leading to necrosis [9]. Broilers reared on wet litter had 43% of the feet plantar surface affected with
FPD compared to 0.2% to 0.3% in broilers reared on dry litter [10]. Risk factors impacting the litter
condition and, in turn, FPD include bird sex and size [11], nutrition [12], bedding material [7,13],
stocking density [14], and seasonality [6]. Contrary to Europe, where litter is replaced after every flock, the US broiler and broiler breeder
industry reuses litter for successive flocks [13,15]. This could be another factor impacting the litter
condition and, in turn, poses a risk for contact dermatitis when compared to fresh litter. Chicken feet
(paws) can be a highly profitable by-product for the industry, and poor footpad conditions due to FPD
reduces the product quality, ultimately resulting in rejections and loss of revenue. The US revenue of
chicken feet may vary depending on the trade market, yet was estimated at 270 million USD from
export to countries in Asia in 2018 [16]. Received: 19 August 2020; Accepted: 23 September 2020; Published: 28 September 2020 Keywords: animal welfare; footpad dermatitis; poultry; plumage cleanliness; remedial treatmen Animals 2020, 10, 1761; doi:10.3390/ani10101761 www.mdpi.com/journal/animals Animals 2020, 10, 1761 2 of 14 1. Introduction Moreover, FPD has an overall negative impact on broiler
productivity. Untreated FPD lesions can lead to lower body weight gain, feed intake, and water intake,
resulting in overall lower carcass weights [17]. FPD lesion scores were positively correlated with
condemnation rates and negatively correlated with the live weights and leg meat yields [18]. Therefore,
the broiler industry has started to take more interest in preventative and remedial approaches, but there
are few feasible treatment options available. FPD affects bird welfare, especially when lesions are severe and painful [19,20], with behavioral
restrictions as a consequence [17]. Chickens have nociceptors in the scale skin of the shanks and feet;
therefore, it is considered that broilers feel pain in the presence of severe lesions [19]. Furthermore,
FPD can be a gateway for potential bacterial infections and the increased prevalence of lameness [4]. Nearly 47% of heavy broilers (3.6 to 3.8 kg) showed some degree of gait impairment [4]. Broilers with
FPD lesions had significantly increased gait impairments in comparison to broilers without lesions [17]. Latency-to-lie test outcomes, indicative of leg weakness, were strongly related to FPD and lameness [21]. Severe gait impairment can be a reason for euthanasia and, thus, an additional loss of revenue. Furthermore, following the EU legislation on fitness for transport, birds with gait impairment would be
deemed unfit to travel, resulting in a serious financial loss [22]. Birds affected by FPD and/or lameness
can reduce activity, increasing the contact of feet, hocks, and breast with litter material, increasing the
susceptibility to abdominal dermatitis and plumage soiling [4,6] and further worsening the risk of feet
and hock dermatitis. Even though a number of risk factors have been identified, the industry rarely takes remedial
actions once lesions have developed in a flock. In a number of European countries, a monitoring and
penalization system has been developed, where high flock scores at slaughter result in mandatory
changes in the management of subsequent flocks, including lowering the stocking density. However,
during the production round, little is done to remedy lesions once they have developed. When lesions 3 of 14 Animals 2020, 10, 1761 are small or moderately sized, intervention in terms of replacing the litter could prevent aggravation
and result in healing before birds reach the slaughter age [10]. Changing the litter can be unpractical in
a commercial system; thus, other remedial interventions need to be studied. 2. Materials and Methods This experiment was carried out between March and May 2019 and was approved by the
Institutional Animal Care and Use Committee (IACUC) of Virginia Tech (protocol 18-246). 1. Introduction Some possible currently
applied interventions could include adapting ventilation, thinning the flock, and resolve leaking
drinkers. The use of a partially slatted floor was hypothesized to result in good footpad conditions
when provided from day one until slaughter [23], although another study found no effect of elevated
platform access on FPD scores [24]. The addition of such flooring could potentially remediate lesions,
although a remedial approach has not yet been assessed. Additionally, the topical treatment of
lesions with an antiseptic could reduce or reverse lesion developments [25]. Yet, in a commercial
setting, individual treatment is not feasible. Thus, in addition to litter management, a potential
practical antiseptic treatment could be applied. Povidone-iodine is an antiseptic that is bactericidal,
fungicidal, tuberculocidal, viricidal, and sporicidal. Iodine can effectively kill yeasts, Staphylococcus sp.,
and E. coli [26], with the latter two associated with severe FPD lesions, as birds with FPD may develop
more severe lesions due to bacterial infections [3]. Therefore, exposure to the antiseptic could be
expected to limit the development of lesions. Thus, the objective of this study was to evaluate novel
flooring treatments at the flock level by providing preventative and remedial treatments to prevent or
reduce contact dermatitis severity, focusing on FPD, in addition to gait impairment and plumage soiling,
in broiler chickens. We hypothesized that (1) access to povidone-iodine would limit the development
of contact dermatitis lesions and that (2) access to slatted flooring would limit lesion developments
further (by reducing the time spent in direct contact with the litter). In addition, the preventative
approach was hypothesized to be optimal, limiting lesion development. The remedial approach was
hypothesized to result in the healing of lesions that developed prior to exposure to the antiseptic and
slatted flooring. 2.2. Treatments The experiment consisted of an incomplete factorial design, with two timing treatments
(preventative and remedial) and four flooring treatments. The flooring treatments included a negative
control, a positive control, and two novel flooring treatments with disinfectant mats containing a
povidone-iodine solution (Table 1). Treatments were randomly allocated over six blocks of pens,
resulting in 7 flooring × timing treatments combinations with 6 replicates. Table 1. Pen-based flooring treatments provided at two different times during production
(preventative or remedial). Prior to day 29 treatment, REM pens were kept under NEG conditions with
used litter. Table 1. Pen-based flooring treatments provided at two different times during production
(preventative or remedial). Prior to day 29 treatment, REM pens were kept under NEG conditions with
used litter. Treatment
Timing
Treatment
(Starting Day)
Flooring
Treatment
Description
NEG
N/A (day 1)
Negative control
Birds were housed on used litter from a previous
flock from day 1 to day 49, which models the
industry standard. PREV-POS
Preventative
(day 1)
Positive control
Birds were provided with new pine shavings at a
depth of 6 cm; the shavings were replaced every
four days. REM-POS
Remedial
(day 29)
PREV-MAT
Preventative
(day 1)
Mat with iodine
solution
Birds were provided a disinfectant mat with 3 L of
a 1% povidone-iodine solution; the mats were
removed from the pen, cleaned, and refilled with
disinfectant solution every four days. REM-MAT
Remedial
(day 29)
PREV-SLAT
Preventative
(day 1)
Mat with iodine
solution and
slatted floor
Birds were provided a disinfectant mat with 3 L of
a 1% povidone-iodine solution and plastic slatted
floor; the mats were removed from the pen,
cleaned, and refilled with disinfectant solution
every four days; excess litter and fecal matter was
removed from the slatted floor as needed. Table 1. Pen-based flooring treatments provided at two different times during production
(preventative or remedial). Prior to day 29 treatment, REM pens were kept under NEG conditions with
used litter. Timing was either preventative (PREV), meaning birds were exposed to the flooring treatments
from the start of the experiment (at day 1 of age), or remedial (REM), meaning birds were kept in
conditions identical to the negative control (NEG) up until day 29 of age (REM) when they received
one of three other flooring treatments (positive control, mat, or mat with slatted flooring). 2.1. Animal Housing Forthisexperiment, wecomparedtheimpactofflooringtreatmentsoncontactdermatitisdevelopment
in a commercial broiler chicken strain. One-day-old male broiler chicks (n = 546 Hubbard × Ross) were
housed in 42 identical pens of 1.25 m2 (10.4 chicks/m2 at placement; stocking density of 35.8 kg/m2 on
day 49) in one climate-controlled research facility until slaughter age of 49 days. Upon arrival from the
commercial hatchery (transported for approximately 6 h), chicks were randomly allocated to a pen with
13 birds per pen. Each pen contained a nipple drinker line with three nipples, a feed pan, and pine
shavings. A heat lamp was provided during the first 7 days, resulting in continuous lighting during that
period. Thereafter, a 18L:6D lighting schedule was applied, with a dark period between 12 a.m. and
6 a.m. A feed flat with starter feed was provided on top of the shavings for the first 7 days to allow
for easy access to feed. House temperature was gradually decreased from 32 ◦C on day 1 to 21 ◦C on
day 49. At the hatchery, birds were vaccinated for Marek’s disease. The birds had ad libitum access
to a commercial diet formulated for each life stage (i.e., starter from day 0–16, grower from day 16–27,
and finisher from day 27–49). The feed was formulated to contain a high percentage of soybean meal
(starter 37% soybean, 57% corn; grower 31.8% soybean, 61% corn; and finisher 29.2% soybean, 63.3% corn)
and met nutritional requirements [27]. Soybean meal contains oligosaccharides, stachyose, and raffinose,
which are indigestible for monogastrics [28]. Stachyose and raffinose cause a higher concentration of
solutes in the lumen of the large intestine, resulting in decreased water absorption. Thus, broilers fed
higher levels of soybean meal will have more viscous excreta, aiding in the development of contact
dermatitis lesions. 4 of 14 Animals 2020, 10, 1761 2.2. Treatments The NEG treatment consisted of housing birds on used litter (19.1% moisture content) that was
collected from a previous broiler flock, to model an industry standard. Litter was collected from
the experimental pens in the same facility and piled in the center hallway. The used litter was
mixed manually and returned to the pens the next day to ensure an equal distribution of litter at
a depth of approximately 6 cm. The NEG flooring treatment did not receive a timing treatment. The positive control treatment (POS) was provided as a preventative or remedial treatment, with new
pine shavings provided on day 1 (PREV) or day 29 (REM) at a depth of 6 cm, and shavings completely
replaced every four days. Pine wood shavings are commonly used as litter for broiler chickens in
the United States (US). All treatments received litter at a depth of 6 cm, and PREV-POS is the only
treatment group that did not receive used litter at any timepoint. The novel flooring treatment with
a disinfectant mat (MAT; Figure 1a consisted of a 60 × 70 cm mat (disinfection mat product number
802010, Agri-Pro Enterprises of Iowa, Inc. Iowa Falls, IA, USA) placed in the back middle of the pen
under the drinker line. The rationale for the location of the mats was two-fold: we wanted to ensure
all birds in the pen were exposed to the treatment (all birds have to drink), and we wanted to place the
mats in a location with a high moisture content. The mat was filled with 3 L of a 1% povidone-iodine
solution (diluted with tap water; 050AB Povidone Iodine Solution 10%, Vi-Jon Inc., Saint Louis, MO, 5 of 14 Animals 2020, 10, 1761 USA) and provided on day 1 (PREV) or day 29 (REM). Every four days, the mats were removed from
the pen, cleaned, and refilled with the disinfectant solution. The second novel flooring treatment
entailed the provision of the MAT treatment (mat plus disinfectant solution), with the additional
provision of a black plastic slatted floor (SLAT; Figure 1b) (60 × 120 cm, DURA-SLAT® Black Poultry
& Kennel Flooring, Southwest Agri-Plastics Inc. Addison, TX, USA), placed on top of the litter but
not elevated from the ground, as commonly done for laying hens and broiler breeders [24]. 2.2. Treatments The mat
was placed on top of the slatted floor, and both were placed in the back middle of the pen under the
drinker, provided on day 1 (PREV) or day 29 (REM). The slatted flooring was removed as needed to
remove excess litter but was not cleaned. Animals 2020, 10, x
5 of 15
solution and
slatted floor
the mats were removed from the pen, cleaned, and
refilled with disinfectant solution every four days;
excess litter and fecal matter was removed from the
slatted floor as needed. Figure 1. Top view of a mat filled with 1% povidone-iodine solution (MAT) (a) and the iodine mat
placed on a slatted floor (SLAT) (b) pen-based flooring treatments. The pens contained a hanging
drinker line (light grey), a metal feeder (dark grey circles), litter (white), a disinfection mat (dark grey
rectangle border and patterned fill), and for the SLAT treatment, a plastic slatted floor (rectangle with
black and white grid pattern). Illustration is not to scale. Figure 1. Top view of a mat filled with 1% povidone-iodine solution (MAT) (a) and the iodine mat
placed on a slatted floor (SLAT) (b) pen-based flooring treatments. The pens contained a hanging
drinker line (light grey), a metal feeder (dark grey circles), litter (white), a disinfection mat (dark grey
rectangle border and patterned fill), and for the SLAT treatment, a plastic slatted floor (rectangle with
black and white grid pattern). Illustration is not to scale. Figure 1. Top view of a mat filled with 1% povidone-iodine solution (MAT) (a) and the iodine mat
placed on a slatted floor (SLAT) (b) pen-based flooring treatments. The pens contained a hanging
drinker line (light grey), a metal feeder (dark grey circles), litter (white), a disinfection mat (dark grey
rectangle border and patterned fill), and for the SLAT treatment, a plastic slatted floor (rectangle with
black and white grid pattern). Illustration is not to scale. Figure 1. Top view of a mat filled with 1% povidone-iodine solution (MAT) (a) and the iodine mat
placed on a slatted floor (SLAT) (b) pen-based flooring treatments. The pens contained a hanging
drinker line (light grey), a metal feeder (dark grey circles), litter (white), a disinfection mat (dark grey
rectangle border and patterned fill), and for the SLAT treatment, a plastic slatted floor (rectangle with
black and white grid pattern). Illustration is not to scale. 2.3. Measurements
2.3. Measurements At the start of the trial, seven sentinel birds from each pen (n = 294 birds) were randomly selected
and ring-banded for identification. Only the ring-banded birds were used for measurements each
week. Measurements were performed at the start of the trial and then repeated weekly. Measurements included FPD, hock burn, breast dermatitis, plumage soiling, gait, and body weight
(Table 2). Each measurement was performed by one observer, eliminating potential interobserver
differences, with the exception of body weight, which was recorded by one out of nine people
involved Observers could not be blinded for treatments as measurements were performed in or near
At the start of the trial, seven sentinel birds from each pen (n = 294 birds) were randomly selected
and ring-banded for identification. Only the ring-banded birds were used for measurements each week. Measurements were performed at the start of the trial and then repeated weekly. Measurements included
FPD, hock burn, breast dermatitis, plumage soiling, gait, and body weight (Table 2). Each measurement
was performed by one observer, eliminating potential interobserver differences, with the exception of
body weight, which was recorded by one out of nine people involved. Observers could not be blinded
for treatments, as measurements were performed in or near the home pens. home pens. Table 2 Measurements (unit) and age of birds on sampling days All categorical scoring systems were
Table 2. Measurements (unit) and age of birds on sampling days. All categorical scoring systems were
transformed to a continuous score using a tagged visual analog scale. home pens. Table 2 Measurements (unit) and age of birds on sampling days All categorical scoring systems were
Table 2. Measurements (unit) and age of birds on sampling days. All categorical scoring systems were
transformed to a continuous score using a tagged visual analog scale. Table 2. Measurements (unit) and age of birds on sampling days. All categorical scoring systems wer
transformed to a continuous score using a tagged visual analog scale. 2.3. Measurements
2.3. Measurements For example,
gait (0-2 ordinal scale) was scored on a 0–100 continuous scale, where 0 through 33 represented a score
0, 34 through 66 represented a score 1, and 67–100 a score 2. For body weight recording, birds were
weighed individually on tabletop scales (8 kg × 0.0001 kg, Adam Equipment Inc, Oxford, CT, USA). FPD was scored for both feet using the tVAS, tagged with the welfare quality 0 to 4 categorical
scale (i.e., score 0 = no lesions, score 1 = small area (≤10%) superficial discoloration, score 2 = large
area (≥10%) superficial discoloration, score 3 = deep lesion and ulceration (up to 50%), and score
4 = deep lesion and ulceration (>50%)) [29]. Hock burns were scored for both hocks using the tVAS,
tagged with a 0 to 4 categorical scale (i.e., score 0 = no visible discoloration, score 1 = small size
(<20%) discoloration, score 2 = small size (20–40%) discoloration, score 3 = moderate size (40–60%)
dark discoloration, and score 4 = large size (>60%) dark discoloration) [29]. Breast dermatitis was
scored by assessing the breast for inflammation using a tVAS tagged with a 0 to 3 categorical scale
(i.e., score 0 = skin without lesion, inflammation, or erythema; score 1 = generalized erythema (up to
25% of ventral body area); score 2 = different degrees of erythema (from 25% to 50% of the ventral
body area); and score 3 = large area inflamed (more than 50% of ventral body area), brown spots,
or breast blisters) [30]. Soiling of the abdominal feathers was scored using the tVAS, tagged with a 0 to
3 categorical scale (i.e., score 0 = absence, score 1 = light, score 2 = moderate, and score 3 = severe) [31]. Lameness was scored by observing the gait pattern of a bird walking a distance of 1.5 m using a tVAS
with a categorical scale from 0 to 2 (i.e., score 0 = no impairment, score 1 = obvious impairment,
walk with a clear limp or awkward gait, and score 2 = severe impairment, may shuffle on shanks or
hocks with assistance of wings) [31]. In addition to the animal-based welfare measures, litter samples were collected from each pen
every week to assess the moisture content. The samples were weighed, dried for 24 h at 100 ◦C,
and then reweighed. 2.3. Measurements
2.3. Measurements Measurement
Sampling Day
Contact dermatitis
Footpad dermatitis 1 (0–4 score)
11, 18, 25, 32, 39, 46
Hock burn 1 (0–4 score)
11, 18, 25, 32, 39, 46
Breast dermatitis 2 (0–3 score)
9, 16, 23, 30, 37, 44
Plumage soiling 2 (0–3 score)
9, 16, 23, 30, 37, 44
Gait 3 (0–2 score)
12, 19, 26, 33, 40, 47
Body weight (kg)
1, 8, 15, 22, 29, 36, 43, 49
1 Adapted from [29], 2 adapted from [30], and 3 adapted from [31]. Table 2. Measurements (unit) and age of birds on sampling days. All categorical scoring systems wer
transformed to a continuous score using a tagged visual analog scale. Measurement
Sampling Day
Contact dermatitis
Footpad dermatitis 1 (0–4 score)
11, 18, 25, 32, 39, 46
Hock burn 1 (0–4 score)
11, 18, 25, 32, 39, 46
Breast dermatitis 2 (0–3 score)
9, 16, 23, 30, 37, 44
Plumage soiling 2 (0–3 score)
9, 16, 23, 30, 37, 44
Gait 3 (0–2 score)
12, 19, 26, 33, 40, 47
Body weight (kg)
1, 8, 15, 22, 29, 36, 43, 49
1 Adapted from [29], 2 adapted from [30], and 3 adapted from [31]. p
[
]
p
[
]
p
[
]
With the exception of weights, all measurements were scored on a tagged visual analog scale
(tVAS) of 10 cm labeled with descriptors representing an increasing degree of the welfare concern
(FPD, hock burn, breast dermatitis, plumage soiling, and gait/lameness) [32]; thus, higher scores
represented a more severe presence of a certain condition. The tVAS ranged from 0 to 100 for all
measurements, allowing for precision and sensitivity to the score while having equal or superior
interobserver reliability compared to ordinal scales [32] Ordinal scales as described in Table 2 were
With the exception of weights, all measurements were scored on a tagged visual analog scale
(tVAS) of 10 cm labeled with descriptors representing an increasing degree of the welfare concern
(FPD, hock burn, breast dermatitis, plumage soiling, and gait/lameness) [32]; thus, higher scores
represented a more severe presence of a certain condition. The tVAS ranged from 0 to 100 for all
measurements, allowing for precision and sensitivity to the score while having equal or superior
interobserver reliability compared to ordinal scales [32]. Ordinal scales as described in Table 2 were 6 of 14 Animals 2020, 10, 1761 transformed to the continuous scale, including the categorical scores as descriptors. 2.3. Measurements
2.3. Measurements The difference between the weights determined the moisture loss, which was
expressed as a percentage. 2.4. Statistical Analysis Treatments were randomly allocated over six blocks, with an incomplete factorial design of three
flooring treatments × two timing treatments, plus a negative control treatment, resulting in seven
treatments (NEG, PREV-MAT, REM-MAT, PREV-SLAT, REM-SLAT, PREV-POS, and REM-POS). Distribution of residuals were determined through visual assessment of QQ plots for the output
variables FPD score (average of two feet), hock burn score (average of two hocks), breast dermatitis
score, plumage cleanliness, gait score, and body weight. All outcomes were considered to be normally
distributed. Data were analyzed using mixed models with block (n = 6) as a random factor and week
(n = 8) and bird ID (n = 294) as repeated factors. The treatments (n = 7), week (bird age n = 8), and their
interactions were included as factors. Tukey-Kramer means separation was used to identify pairwise
differences at p < 0.05. Due to a human error, birds in 4 pens received incorrect treatments from day
13 through day 16 of age. Data analysis omitting the 4 pens from the dataset revealed that this error
did not affect the impact of the independent variables on the dependent variable modeled means. Therefore, authors decided to not omit the 4 pens from the dataset. 3.1. FPD
e Treatment, age, and their interaction affected the FPD scores (p < 0.001; Table 3). In the preventative
approach, PREV-POS showed the best (lowest) FPD scores compared to the other treatments (p ≤0.002;
Figure 2a). The most severe (high) FPD scores were found in PREV-MAT and PREV-SLAT compared to
the best (lowest) FPD scores in NEG and PREV-POS (pairwise comparisons between flooring treatments
p ≤0.002; Figure 2a). Similar results were found in the remedial approach (Figure 2b), with REM-MAT
and REM-SLAT showing higher FPD scores compared to NEG and REM-POS (pairwise comparisons
between flooring treatments p < 0.001). In the remedial approach, REM-POS did not differ from the
NEG outcomes. Within the PREV-POS and REM-POS treatments, scores decreased with age (p < 0.05;
Figure 2). Within PREV-MAT, PREV-SLAT, REM-MAT, and REM-SLAT, scores generally increased as
the birds aged (p < 0.05; Figure 2), whilst scores in the NEG treatment were not affected by age. we found an interaction effect between the treatment and week (Table 3), pairwise differences
between the groups were not significant within the preventative or remedial applications (Figure 7). Furthermore, limited effects of the flooring treatments were found within a week, resulting in
comparable body weights between the treatments, with the exception of week seven, where PREV-
POS showed lower weights compared to NEG, PREV-MAT, and PREV-SLAT (Figure 7a). 3.7. Litter Moisture
The litter moisture differed over time, with an increasing moisture in the litter as the birds aged
from (LS MEAN ± SEM) 9.8% ± 0.9% in week two to 32.6% ± 0.9% in week eight (p < 0.001). It tended
to differ between treatments (p = 0.095), with 22.8% litter moisture in the NEG, 23.4% in REM-POS,
24.3% in REM-MAT, 24.8% in PREV-SLAT, 25.5% in PREV-POS, 25.8% in REM-SLAT, and 27.0% in
PREV-MAT (SEM = 1.1%). Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 20-point increase). 3.1. FPD
e The arrows in (b) indicate the
timing applications of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–c) differ significantly at p < 0.05. 3.1. FPD
e The arrows in (b)
indicate the timing applications of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–c) differ significantly at p < 0.05. Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 20-point increase). The arrows in (b) indicate the
timing applications of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–c) differ significantly at p < 0.05. Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 20-point increase). The arrows in (b)
indicate the timing applications of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–c) differ significantly at p < 0.05. Figure 2. Mean footpad dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 20-point increase). 3. Results The experimental treatments resulted in differences in FPD, hock burns, breast dermatitis scores,
plumage cleanliness, gait, and body weights (Table 3). 7 of 14 Animals 2020, 10, 1761 Table 3. LS MEANS (and SEM) of the welfare outcomes for all flooring × timing treatment combinations
(trt). p-values of the main and interactive effects are given for all welfare outcomes. (trt). p values of the main and interactive effects are given for all welfare outcomes. Welfare
Outcome
NEG
PREV-
POS
PREV-
MAT
PREV-
SLAT
REM-
POS
REM-
MAT
REM-
SLAT
SEM
p-Value
Trt
Week
(Age)
Trt ×
Week
Footpad
dermatitis 1
15.4 d
3.2 f
47.4 b
52.5 a
10.8 e
31.5 c
33.5 c
0.9
<0.001
<0.001
<0.001
Hock burns 1
6.5 c
1.4 d
12.4 b
14.3 a,b
3.4 d
12.4 b
16.1 a
0.6
<0.001
<0.001
<0.001
Breast
dermatitis 1
9.8
10.1
13.7
14.0
13.8
10.0
13.6
0.6
0.008 *
<0.001
0.001
Plumage
cleanliness 1
13.7 b,c
12.9 c
18.8 a,b
19.2 a,b
14.7 a,b,c
17.0 a,b,c
19.8 a
1.2
0.001
<0.001
<0.001
Gait 1
11.2 b
8.0 c
13.9 a
12.8 a,b
11.2 b
12.7 a,b
13.4 a,b
0.6
<0.001
<0.001
0.004
Body weight (g)
1681.4 a
1564.8 c
1620.9 b
1603.4 b,c
1695.1 a
1596.2 b,c
1710.8 a
13.1
0.038
<0.001
<0.001
1 Expressed as a tagged visual analog scale (tVAS) score between 0–100, with 0 representing the best possible
outcome and 100 the worst possible outcome. Means in a row with different superscripts (a–f) differ significantly
at p < 0.05. * No pairwise differences between treatment groups after Tukey-Kramer adjustment. Animals 2020, 10, x
8 of 15
3.5. Gait
Treatment, age, and their interaction affected the gait scores (respectively, p < 0.001, p < 0.001,
and p = 0.004; Table 3). In the preventative approach, PREV-POS resulted in better (lower) gait scores
compared to NEG, PREV-MAT, and PREV-SLAT (p ≤ 0.002; Figure 6a). Furthermore, PREV-MAT
showed worse (higher) gait scores compared to the NEG (p = 0.035; Figure 6a). Within the remedial
approach, no pairwise differences between flooring treatments were found (Figure 6b). Within all
treatment groups, the gait scores worsened (increased) with age (p < 0.05; Figure 6). 3.6. Body Weight 3.3. Breast Dermatitis
provision of
flooring trea Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Treatment, age, and their interaction affected the breast dermatitis scores (respectively, p = 0.008,
p < 0.001, and p = 0.001; Table 3). However, no pairwise differences were found between flooring
treatments within the preventative or remedial approaches. Within all treatment groups, the breast
dermatitis scores increased with age (p < 0.05; Figure 4). flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mate with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 20-point increase). The arrows (b) indicate
the timing applications of the remedial treatments. Means in a flooring*timing treatment combination
with different superscripts (a–c) differ significantly at p < 0.05. provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. 3.2. Hock Burns Mean hock dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Animals 2020, 10, x
9 of 15 igu e
(
)
(a) p
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mate with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 20-point increase). The arrows (b) indicate
the timing applications of the remedial treatments. Means in a flooring*timing treatment combination
with different superscripts (a–c) differ significantly at p < 0.05. provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mate with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 20-point increase). The arrows (b) indicate the timing
applications of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–c) differ significantly at p < 0.05. Within the PREV and REM-POS treatment groups, the hock burn scores remained low over time,
showing no age effect. Within all other treatment groups, the hock burn scores increased with age
(p < 0.05; Figure 3). Figure 3 Mea
ho k de
atiti
o e
( a
ea
± SEM) of bi d
that e ei ed (a)
e e tati e 3.3. Breast Dermatitis
provision of
flooring trea Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b) indicate the
timing application of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–d) differ significantly at p < 0.05. provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Figure 4. 3.2. Hock Burns Treatment, age, and their interaction affected the hock burn scores (p < 0.001; Table 3). In the
preventative approach PREV-POS showed the best (lowest) hock burn scores compared to the other
treatments (p < 0.001; Figure 3a). The most severe (high) hock burn scores were found in PREV-MAT
and PREV-SLAT compared to NEG and PREV-POS (pairwise comparisons between flooring treatments 8 of 14 Animals 2020, 10, 1761 p < 0.001; Figure 3a). Similar results were found in the remedial approach, with REM-POS showing
the lowest scores compared to the other treatment groups (Figure 3b). In addition, REM-MAT and
REM-SLAT showed higher hock burn scores compared to NEG and REM-POS (pairwise comparisons
between flooring treatments p < 0.001; Figure 3b). Animals 2020, 10, x
9 of 15 Figure 3. Mean hock dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mate with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 20-point increase). The arrows (b) indicate
the timing applications of the remedial treatments. Means in a flooring*timing treatment combination
with different superscripts (a–c) differ significantly at p < 0.05. Figure 3. Mean hock dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mate with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 20-point increase). The arrows (b) indicate the timing
applications of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–c) differ significantly at p < 0.05. Animals 2020, 10, x
9 of 15 Figure 3. Mean hock dermatitis scores (raw means ± SEM) of birds that received (a) preventative
Figure 3. 3.5. Gait Treatment, age, and their interaction affected the gait scores (respectively, p < 0.001, p < 0.001,
and p = 0.004; Table 3). In the preventative approach, PREV-POS resulted in better (lower) gait scores
compared to NEG, PREV-MAT, and PREV-SLAT (p ≤0.002; Figure 6a). Furthermore, PREV-MAT
showed worse (higher) gait scores compared to the NEG (p = 0.035; Figure 6a). Within the remedial
approach, no pairwise differences between flooring treatments were found (Figure 6b). Within all
treatment groups, the gait scores worsened (increased) with age (p < 0.05; Figure 6). Animals 2020, 10, x
10 of 15
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. (a)
(b)
Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue
scale (tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are
indicated by the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing
application of remedial treatments. Means in a flooring*timing treatment combination with different
superscripts (a–e) differ significantly at p < 0.05. Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue scale
(tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are indicated by
the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing application of
remedial treatments. 3.3. Breast Dermatitis
provision of
flooring trea Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. g
p
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b) indicate the
timing application of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–d) differ significantly at p < 0.05. 9 of 14 Animals 2020, 10, 1761 3.4. Plumage Cleanliness (Soiling)
Fi
4 M
b
t d Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative Fi
5 M
l
ili
(
± SEM)
f bi d th t
i
d ( )
t ti
Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring p
g
(
) ( )
p
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b) indicate the
timing application of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–d) differ significantly at p < 0.05. 3.4. Plumage Cleanliness (Soiling)
Fi
4 M
b
t d Treatment, age, and their interaction affected the plumage soiling scores (p ≤0.001; Table 3). In the preventative application, PREV-POS had lower scores (less soiling) compared to PREV-MAT
and PREV-SLAT (p ≤0.030) but did not differ from the NEG (Figure 5a). In the remedial approach,
REM-SLAT showed higher scores (more soiling) compared to the NEG treatment (p = 0.024; Figure 5b). No other pairwise differences were found within the preventative or remedial approaches. For all
treatments, the plumage soiling scores increased (more soiling) over time (p < 0.05; Figure 5). Figure 4. Mean breast dermatitis scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
flooring treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat
(MAT), or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–
100 tagged analog scale (tVAS), with increasing scores representing a worsened condition (the “tags”
on the VAS are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b)
indicate the timing application of the remedial treatments. Means in a flooring*timing treatment
combination with different superscripts (a–d) differ significantly at p < 0.05. Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the
Figure 5. Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative
provision of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT),
or iodine mat with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged
analog scale (tVAS), with increasing scores representing a worsened condition (the “tags” on the VAS
are indicated by the horizontal gridlines at each 25-point increase). The arrows in (b) indicate the
timing application of the remedial treatments. Means in a flooring × timing treatment combination
with different superscripts (a–d) differ significantly at p < 0.05. Figure 5 Mean plumage soiling scores (raw means ± SEM) of birds that received (a) preventative Fi
5 M
l
ili
(
± SEM)
f bi d th t
i
d ( )
t ti
Figure 5. 3.5. Gait Means in a flooring × timing treatment combination with different superscripts
(a–e) differ significantly at p < 0.05. (b) (a) (b) (b) (a) Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
fl
i
t
t
t (PREV) (b)
bi d th t
i
d
di l
i i
f th
fl
i
t
t
t
Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue
scale (tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are
indicated by the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing
application of remedial treatments. Means in a flooring*timing treatment combination with different
superscripts (a–e) differ significantly at p < 0.05. Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue scale
(tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are indicated by
the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing application of
remedial treatments. Means in a flooring × timing treatment combination with different superscripts
(a–e) differ significantly at p < 0.05. Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. 3.6. Body Weight
Figure 6. M Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or
iodine mat with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the
timing application of the remedial treatments. Means in a flooring*timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the timing
application of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or
iodine mat with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the
timing application of the remedial treatments. Means in a flooring*timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. g
(
) ( )
p
g
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the timing
application of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. 4. Discussion
3.7. Litter Moisture Contact dermatitis, and especially FPD, is a prevalent welfare issue for broiler chickens, with
potential economic consequences. Common commercial practices do not include any type of
treatment of these lesions, which could be costly and labor-intensive if birds would need to be treated
individually. Therefore, this study aimed to identify effective preventative and remedial flooring
treatments at the flock level to reduce contact dermatitis in broilers. The remedial flooring treatments
were compared to preventative applications of the same treatments. To our knowledge, this is the
The litter moisture differed over time, with an increasing moisture in the litter as the birds aged
from (LS MEAN ± SEM) 9.8% ± 0.9% in week two to 32.6% ± 0.9% in week eight (p < 0.001). It tended
to differ between treatments (p = 0.095), with 22.8% litter moisture in the NEG, 23.4% in REM-POS,
24.3% in REM-MAT, 24.8% in PREV-SLAT, 25.5% in PREV-POS, 25.8% in REM-SLAT, and 27.0% in
PREV-MAT (SEM = 1.1%). 3.5. Gait Scores are on a 0–100 tagged analogue
scale (tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are
indicated by the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing
application of remedial treatments. Means in a flooring*timing treatment combination with different
superscripts (a–e) differ significantly at p < 0.05. Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue scale
(tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are indicated by
the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing application of
remedial treatments. Means in a flooring × timing treatment combination with different superscripts
(a–e) differ significantly at p < 0.05. 10 of 14 10 of 14 Animals 2020, 10, 1761 3.6. Body Weight
Figure 6. M The body weights significantly increased each week in all treatment groups (Figure 7). Although we
found an interaction effect between the treatment and week (Table 3), pairwise differences between the
groups were not significant within the preventative or remedial applications (Figure 7). Furthermore,
limited effects of the flooring treatments were found within a week, resulting in comparable body
weights between the treatments, with the exception of week seven, where PREV-POS showed lower
weights compared to NEG, PREV-MAT, and PREV-SLAT (Figure 7a). Figure 6. Mean gait scores (raw means ± SEM) of birds that received (a) preventative provision of the
flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 7 of age. Scores are on a 0–100 tagged analogue
scale (tVAS) with increasing scores representing a worsened condition (the ‘tags’ on the VAS are
indicated by the horizontal gridlines at each 33-point increase). The arrows in (b) indicate timing
application of remedial treatments. Means in a flooring*timing treatment combination with different
superscripts (a–e) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or
iodine mat with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the
timing application of the remedial treatments. Means in a flooring*timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the timing
application of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. 3.6. Body Weight
Figure 6. M Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or
iodine mat with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the
timing application of the remedial treatments. Means in a flooring*timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the timing
application of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring
treatments (REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or
iodine mat with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the
timing application of the remedial treatments. Means in a flooring*timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. Mean body weights (kg) (raw means ± SEM) of birds that received (a) preventative provision
of the flooring treatments (PREV) (b) or birds that received remedial provision of the flooring treatments
(REM): negative control (NEG (presented twice)), clean litter (POS), iodine mat (MAT), or iodine mat
with slatted floor (SLAT) during weeks 2 through 8 of age. The arrow in (b) indicates the timing
application of the remedial treatments. Means in a flooring × timing treatment combination with
different superscripts (a–b) differ significantly at p < 0.05. Figure 7. 4. Discussion The slats used
in this study are commonly used for broiler breeders, thus were deemed appropriate for the broilers. Another contributing factor for differences in lesions between the flooring treatments is the
chemical and microbial compositions of the litter or on the footpads due to the antiseptic effects
of povidone-iodine in SLAT and MAT and due to replacement of the litter in the POS treatment. However, chemical and microbial compositions were not measured in this study. Even though MAT
and SLAT resulted in the highest FPD scores compared to the other treatments, the lesions were
superficial. A remedial (healing) effect could only be detected if there were a sufficient number of
birds that were (badly) affected by dermatitis. It seems at least questionable whether this was the
case in this experiment, so the power for detecting a remedying effect may have been low. Indeed,
when considering the mean FPD scores for the REM treatments prior to applying the REM treatments
in week four, the scores were low (REM-POS 12, REM-MAT 25, and REM-SLAT 27 out of 100), making it
less likely for a remedial effect to be detected. Application of the flooring treatments on a commercial
scale, where mats and slatted flooring take up a smaller proportion of the space, may be beneficial,
yet the current data do not support this. In-line with previous findings, clean and dry litter effectively prevents [17] and remedies FPD and
hock lesions [10,35]. Hock burn scores were consistently low within the POS treatments, the lowest
compared to all the other treatment groups. For FPD, both PREV-POS and REM-POS showed a
decrease in scores from mean scores of 13 and 19 (out of 100) in week two to 0 and 2 in week seven,
which reflected a nearly perfect footpad condition. Both PREV and REM approaches showed a similar
pattern for this treatment, where the lesions almost completely healed. The POS treatment showed the
greatest reduction in lesion scores compared to any of the other treatments. This treatment shows great
promise for application in the broiler chicken industry, albeit with modifications. In the trial, all the
litter was replaced in the POS treatments, which is not feasible on a commercial farm. Yet, top-dressing
with fresh litter, combined with removing and replacing the worst parts of the litter, has the potential
to remedy FPD lesions, although scientific findings are lacking. 4. Discussion Contact dermatitis, and especially FPD, is a prevalent welfare issue for broiler chickens,
with potential economic consequences. Common commercial practices do not include any type
of treatment of these lesions, which could be costly and labor-intensive if birds would need to be treated
individually. Therefore, this study aimed to identify effective preventative and remedial flooring
treatments at the flock level to reduce contact dermatitis in broilers. The remedial flooring treatments
were compared to preventative applications of the same treatments. To our knowledge, this is the first
study assessing an antiseptic (in combination with slatted flooring) as a potential remedy for contact
dermatitis, plumage soiling, and lameness. The flooring treatments applied in this study had an impact on the welfare outcomes, including FPD,
hock burns, gait (depending on week of age), plumage cleanliness, and breast dermatitis. Contrary to
the hypothesis, however, providing access to an antiseptic solution (povidone-iodine) either from
day one or day 29, resulted in more severe FPD lesions as compared to both the negative control
(model of the industry standard) and the positive control. The litter moisture hardly differed between
the treatments (means ranged from 22% to 27%), so this could not explain the large disparity in lesion
scores between the treatment groups. Yet, an inherent part of the MAT and SLAT treatments was
exposing birds to a liquid (the antiseptic solution), which could cause increased lesion scores due
to softening of the dermis [9], potentially loosening the scales and resulting in an acceleration of 11 of 14 Animals 2020, 10, 1761 lesion development [33]. Furthermore, the mat and slatted floorings took up a large proportion of
the pens (25–55% of the floor space). This could contribute to the induction of FPD lesions, as birds
were potentially exposed to the moist surface of the mat for prolonged periods of time. Reducing the
amount of solution in the mats could have a limited lesion development, yet a priori, we theorized
that the liquid would be beneficial, and thus, the mats needed be saturated for the birds to come into
contact with the antiseptic solution. Cengiz et al. [34] reported low FPD severity in broilers kept on plastic wire floors. In contrast,
our findings suggest that the slatted floor contributed to FPD lesion developments when exposed from
day one but not when exposed from day 29 when we compared MAT to SLAT (Table 3). 4. Discussion The NEG treatment resulted in stable and relatively low FPD scores as the birds aged, which is
opposite to the expectation. Exposure to previously used litter was used to model a US commercial
standard, and following that exposure, we expected poor footpad quality, as found in the commercial
setting [4]. However, the NEG treatment was associated with higher FPD scores compared to the
positive control but only when the litter was replaced regularly from day one, as the NEG scores did
not differ from REM-POS. The lack of severe lesions in NEG birds was likely due to low litter moisture
in the NEG treatment group, similar to findings with turkeys, where dry litter (under 30% moisture)
resulted in low FPD scores (<1 on a 0 to 5 scale), regardless of bird age [36]. Another difference
compared to the commercial standards was that the used litter came from a single flock, rather than
multiple, with the latter approach being linked to more severe FPD lesions with each successive use [13]. In our case, this could have resulted in better-quality litter compared to commercial standards. For FPD and gait, longer exposure to the flooring treatments (day one compared to day 29; Table 3)
resulted in increased severity of these indicators, although the effect was less pronounced for the gait
scores. PREV-POS was the best approach to maintain low gait scores compared to all other flooring ×
timing treatments. For all flooring × timing treatment combinations, the gait scores were relatively
low compared to commercial farms [4]; within our study, obvious impairments were only observed 12 of 14 Animals 2020, 10, 1761 in week seven of age (eight birds originating from all treatments; data not shown). In part, this is a
result of euthanasia decisions. Severe lameness was a reason to euthanize birds, which impacted the
mean gait scores in all the treatments. Out of 294 sentinel birds, 23 died or were euthanized, of which
seven were for leg issues when birds were six or seven weeks old (one from a NEG pen, one from a
PREV-MAT pen, four from three REM-MAT pens, and one from a REM-POS pen). Breast dermatitis and plumage soiling scores showed limited responses to the treatments. Compared to birds at a Brazilian slaughter plant [31], the prevalence of breast dermatitis was
low in our study. 4. Discussion They recorded 72% of birds with a categorical score of 1 or higher (tVAS > 25) at
four weeks of age [31], whilst all birds in our study showed a score between 0 and 1 (tVAS < 25) in
weeks one through six of age. For all treatments, the mean plumage soiling scores were low, as 0 on the
ordinal scale (no soiling) was scored most frequently (71% of the scores; data not shown). These low
scores are not in-line with a previous study under comparable stocking density conditions, where 99%
of birds showed slight soiling irrespective of the litter treatment [24]. PREV-POS resulted in lower
(better) soiling scores compared to PREV-MAT and PREV-SLAT, suggesting that the best strategy to
avoid soiling is early access to clean litter that is regularly replaced. The welfare indicators generally worsened with the bird age. The positive association between
FPD, hock burns, breast dermatitis, plumage soiling, and gait with age was in-line with the expectations
and findings in the literature: FPD [17,35,37,38], hock burns [17], plumage soiling [17], and gait [17,38]. Bird weights were recorded as an indicator of productivity. The treatments had a minimal
effect on the body weights, with the only difference found in week seven of age, where PREV-POS The welfare indicators generally worsened with the bird age. The positive association between
FPD, hock burns, breast dermatitis, plumage soiling, and gait with age was in-line with the expectations
and findings in the literature: FPD [17,35,37,38], hock burns [17], plumage soiling [17], and gait [17,38]. The welfare indicators generally worsened with the bird age. The positive association between
FPD, hock burns, breast dermatitis, plumage soiling, and gait with age was in-line with the expectations
and findings in the literature: FPD [17,35,37,38], hock burns [17], plumage soiling [17], and gait [17,38]. Bird weights were recorded as an indicator of productivity. The treatments had a minimal
effect on the body weights, with the only difference found in week seven of age, where PREV-POS
showed lower body weights compared to NEG and the other PREV treatments. In week eight,
this difference was no longer observed. Based on observations of a subsample of pens, birds in the POS
treatments generally spent more time being active compared to birds in the other flooring × treatment
combinations, which could have contributed to the weight difference. 4. Discussion The birds were active for 48%
(PREV-POS), 43% (REM-POS), 41% (NEG), 39% (REM-MAT), 34% (REM-SLAT), 33% (PREV-MAT),
and 32% (PREV-SLAT) of the time they were observed [39]. Due to the limited impact of treatments
on weights, we can cautiously conclude that applying these treatments does not negatively impact
the productivity, regardless of the lesion scores. This contradicts the previously reported negative
relationship of productivity with litter deterioration and FPD at day 37 of age [17]. Author Contributions: Conceptualization, F.A.M.T. and L.J.; methodology, F.A.M.T., L.J., A.G., and N.F.;
formal analysis, L.J.; investigation, N.F., V.M., A.J., and L.J.; resources, L.J. and F.T.; data curation, L.J.;
writing—original draft preparation, N.F. and L.J.; writing—review and editing, F.A.M.T. and A.G.; visualization,
L.J.; supervision, L.J.; and project administration, L.J. All authors have read and agreed to the published version of
the manuscript. Acknowledgments: We would like to thank Gracie Anderson, Carley Elliot, Caroline Flood, Kylie Hericks,
Shawnna Melick, and Sarina Williams for their valuable help during the experiment. Thanks to VT SAIG assistance
with the formal analysis by Justin Loda and Minh Le. We owe many thanks to the late Dale Shumate for his
valuable help as the farm manager. Conflicts of Interest: The authors declare no conflict of interest. References [CrossRef]
11
Bilgili S F ; Alley M A ; Hess J B ; Nagaraj M Influence of Age and Sex on Footpad Quality and Yield in 10. Martland, M.F. Ulcerative dermatitis dm broiler chickens: The effects of wet litter. Avian Pathol. 1985, 14,
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china-may-presage-new-trading-era (accessed on 22 June 2020). 17. De Jong, I.C.; Gunnink, H.; van Harn, J. Wet litter not only induces footpad dermatitis but also reduces
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contact dermatitis, focusing on footpad dermatitis in broiler chickens. The exposure to mats with
antiseptics, either combined with slatted flooring or on its own, did not prove an appropriate method to
prevent or reduce contact dermatitis, plumage soiling, or lameness in the current experiment. The most
promising approach to both prevent and remedy these welfare concerns was access to clean litter. In the
current study, fresh litter was replaced regularly, which is not feasible in the industry. Yet, removing
and replacing litter in certain spots on-farm, top-dressing with fresh litter, and replacing used litter in
between flocks seems worth investigating in order to remedy contact dermatitis, especially footpad
lesions in commercial flocks. Author Contributions: Conceptualization, F.A.M.T. and L.J.; methodology, F.A.M.T., L.J., A.G., and N.F.;
formal analysis, L.J.; investigation, N.F., V.M., A.J., and L.J.; resources, L.J. and F.T.; data curation, L.J.;
writing—original draft preparation, N.F. and L.J.; writing—review and editing, F.A.M.T. and A.G.; visualization,
L.J.; supervision, L.J.; and project administration, L.J. All authors have read and agreed to the published version of
the manuscript. Funding: We would like to thank the Flanders Institute for Agricultural and Fisheries Research for financial sup Acknowledgments: We would like to thank Gracie Anderson, Carley Elliot, Caroline Flood, Kylie Hericks,
Shawnna Melick, and Sarina Williams for their valuable help during the experiment. Thanks to VT SAIG assistance
with the formal analysis by Justin Loda and Minh Le. We owe many thanks to the late Dale Shumate for his
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on the welfare of fast-growing broilers with an extended fattening period. Arch. Anim. Breed. 2018, 61, 9–16. [CrossRef] 24. Kaukonen, E.; Norring, M.; Valros, A. Evaluating the effects of bedding materials and elevated platforms on
contact dermatitis and plumage cleanliness of commercial broilers and on litter conditions in broiler houses. Br. Poult. Sci. 2017, 58, 480–489. [CrossRef] 25. Doneley, R.J.T.; Smith, B.A.; Gibson, J.S. Use of a Vascular Access Port for Antibiotic Administration in the
Treatment of Pododermatitis in a Chicken. J. Avian Med. Surg. 2015, 29, 130–135. [CrossRef] [PubMed] 26. Zamora, J.L. Chemical and microbiologic characteristics and toxicity of povidone-iodine solutions. Am. J. Surg. 1986, 151, 400–406. [CrossRef] 27. National Research Council. Nutrient Requirements of Poultry, 9th ed.; National Academies Press: Washington,
DC, USA, 1994. 28. Youssef, I.M.I.; Beineke, A.; Rohn, K.; Kamphues, J. Effects of high dietary levels of soybean meal and its
constituents (potassium, oligosaccharides) on foot pad dermatitis in growing turkeys housed on dry and wet
litter. Arch. Anim. Nutr. 2011, 65, 148–162. [CrossRef] 29. Welfare Quality®Network. Welfare Quality®Assessment Protocol for Poultry (Broilers, Laying Hens); Welfare
Quality®Network: Lelystad, The Netherlands, 2009. 30. Souza, A.P.O.; Soriano, V.S.; Schnaider, M.A.; Rucinque, D.S.; Molento, C.F.M. Development and refinement
of three animal-based broiler chicken welfare indicators. Anim. Welf. 2018, 27, 263–274. [CrossRef] 31. Webster, A.B.; Fairchild, B.D.; Cummings, T.S.; Stayer, P.A. Validation of a Three-Point Gait-Scoring System
for Field Assessment of Walking Ability of Commercial Broilers. J. Appl. Poult. Res. 2008, 17, 529–539. [CrossRef] 32. Tuyttens, F.A.M.; de Graaf, S.; Heerkens, J.L.T.; Jacobs, L.; Nalon, E.; Ott, S.; Stadig, L.M.; van Laer, E.;
Ampe, B. © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
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foot pad scores were minimized when litter moisture was less than 30%. Poult. Sci. 2011, 90, 1170–1178. [CrossRef] 37. Baéza, E.; Arnould, C.; Jlali, M.; Chartrin, P.; Gigaud, V.; Mercerand, F.; Durand, C.; Méteau, K.;
le Bihan-Duval, E.; Berri, C. Influence of increasing slaughter age of chickens on meat quality, welfare, and
technical and economic results. J. Anim. Sci. 2012, 90, 2003–2013. [CrossRef] [PubMed] 38. Bassler, W.; Arnould, C.; Butterworth, A.; Colin, L.; de Jong, I.C.; Ferrante, V.; Ferrari, P.; Haslam, S.;
Wemelsfelder, F.; Blokhuis, H.J. Potential risk factors associated with contact dermatitis, lameness, negative
emotional state, and fear of humans in broiler chicken flocks. Poult. Sci. 2013, 92, 2811–2826. [CrossRef] 39. Jacobs, L. Behavioral Observations of Broiler Chickens Housed With Novel Flooring Treatments. Virginia Polytechnic
Institute and State University: Blacksburg, VA, USA, 2019; Unpublished work. © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
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Research article Research article Biogeosciences, 19, 1377–1394, 2022
https://doi.org/10.5194/bg-19-1377-2022
© Author(s) 2022. This work is distributed under
the Creative Commons Attribution 4.0 License. MODIS Vegetation Continuous Fields tree cover needs
calibrating in tropical savannas Kelley (doukel@ceh.ac.uk) Correspondence: Rahayu Adzhar (adzharrb@miamioh.edu) and Douglas I. Kelley (doukel@c Received: 8 December 2020 – Discussion started: 2 February 2021
Revised: 12 January 2022
Accepted: 22 January 2022
Published: 7 March 2022 Received: 8 December 2020 – Discussion started: 2 February 2021
Revised: 12 January 2022 – Accepted: 22 January 2022 – Published: 7 March 2022 Revised: 12 January 2022 – Accepted: 22 January 2022 – Published: 7 March 2022 Abstract. The Moderate Resolution Imaging Spectrora-
diometer Vegetation Continuous Fields (MODIS VCF) Earth
observation product is widely used to estimate forest cover
changes and to parameterize vegetation and Earth system
models and as a reference for validation or calibration where
field data are limited. However, although limited independent
validations of MODIS VCF have shown that MODIS VCF’s
accuracy decreases when estimating tree cover in sparsely
vegetated areas such as tropical savannas, no study has yet
assessed the impact this may have on the VCF-based tree
cover data used by many in their research. Using tropical
forest and savanna inventory data collected by the Tropical
Biomes in Transition (TROBIT) project, we produce a series of calibration scenarios that take into account (i) the spatial
disparity between the in situ plot size and the MODIS VCF
pixel and (ii) the trees’ spatial distribution within in situ plots. To identify if a disparity also exists in products trained using
VCF, we used a similar approach to evaluate the finer-scale
Landsat Tree Canopy Cover (TCC) product. For MODIS
VCF, we then applied our calibrations to areas identified as
forest or savanna in the International Geosphere-Biosphere
Programme (IGBP) land cover mapping product. All IGBP
classes identified as “savanna” show substantial increases in
cover after calibration, indicating that the most recent version
of MODIS VCF consistently underestimates woody cover in
tropical savannas. We also found that these biases are prop- MODIS Vegetation Continuous Fields tree cover needs
calibrating in tropical savannas Rahayu Adzhar1,2,3, Douglas I. Kelley1, Ning Dong3,4, Charles George1, Mireia Torello Raventos5,
Elmar Veenendaal6, Ted R. Feldpausch7, Oliver L. Phillips8, Simon L. Lewis8,9, Bonaventure Sonké10,
Herman Taedoumg11, Beatriz Schwantes Marimon12, Tomas Domingues13, Luzmila Arroyo14, Gloria Djagbletey15,
Gustavo Saiz16, and France Gerard1 Adzhar1,2,3, Douglas I. Kelley1, Ning Dong3,4, Charles George1, Mireia Torello Raventos5,
6
7
8
8 9
1 Rahayu Adzhar
, Douglas I. Kelley , Ning Dong
, Charles George , Mireia Torello Raventos ,
Elmar Veenendaal6, Ted R. Feldpausch7, Oliver L. Phillips8, Simon L. MODIS Vegetation Continuous Fields tree cover needs
calibrating in tropical savannas Lewis8,9, Bonaventure Sonké10,
Herman Taedoumg11, Beatriz Schwantes Marimon12, Tomas Domingues13, Luzmila Arroyo14, Gloria Djagbletey15,
Gustavo Saiz16, and France Gerard1 Herman Taedoumg11, Beatriz Schwantes Marimon12, Tomas Domingues13, Luzmila Arroyo14, Gloria Djagbletey15,
Gustavo Saiz16, and France Gerard1 1UK Centre for Ecology & Hydrology, Wallingford, Oxfordshire, UK 1UK Centre for Ecology & Hydrology, Wallingford, Oxfordshire, UK UK Centre for Ecology & Hydrology, Wallingford, Oxfordshire, UK UK Centre for Ecology & Hydrology, Wallingford, Oxfordshire, UK
2Department of Ecology, Evolution, and Environmental Biology, Miami University, Oxford, Ohio, USA
3 gy
y
gy
g
t of Ecology, Evolution, and Environmental Biology, Miami University, Oxford, Ohio, USA p
gy
gy
3Department of Life Sciences, Imperial College London, Berkshire, UK Department of Life Sciences, Imperial College London, Berkshire, UK p
p
g
4Department of Biological Sciences, Macquarie University, North Ryde, Australia
5 4Department of Biological Sciences, Macquarie University, North Ryde, Australia 5School of Earth and Environmental Science, James Cook University, Cairns, Australia 6Plant Ecology and Nature Conservation, Wageningen University & Research, Wageningen, the Netherlands
7College of Life and Environmental Sciences, University of Exeter, Exeter, UK 6Plant Ecology and Nature Conservation, Wageningen University & Research, Wageningen, the Netherlands
7College of Life and Environmental Sciences, University of Exeter, Exeter, UK 8School of Geography, University of Leeds, Leeds, UK g p y
y
9Department of Geography, University College London, London, UK 9Department of Geography, University College London, London, UK 10Plant Systematics and Ecology Laboratory, Department of Biology, 10Plant Systematics and Ecology Laboratory, Department of Biology, Higher Teachers’ Training College, 10Plant Systematics and Ecology Laboratory, Department of Biology, Higher Teachers’ Training College,
University of Yaoundé, Yaoundé, Cameroon 10Plant Systematics and Ecology Laboratory, D University of Yaoundé, Yaoundé, Cameroon 11Consultative Group on International Agricultural Research (CGIAR), Bioversity International, Yaoundé, Cameroon
12 11Consultative Group on International Agricultural Research (CGIAR), Bioversity International, Yaoundé, Cameroon
12Departamento de Ciências Biológicas, Mato Grosso State University, Mato Grosso, Brazil 11Consultative Group on International Agricultural Research (CGIAR), Bioversity International, Y
12 11Consultative Group on International Agricu 12Departamento de Ciências Biológicas, Mato Grosso State University, Mato Grosso, Brazil 13Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto,
University of São Paulo, São Paulo, Brazil 13Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto,
University of São Paulo, São Paulo, Brazil Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, 14Departamento de Biología, Universidad Autónoma Gabriel René Moreno, Santa Cruz, Bolivia
15 Departamento de Biología, Universidad Autónoma Gabriel René Moreno, Santa Cruz, Bolivia 15Forest and Climate Change, Forestry Research Institute of Ghana, Kumasi, Ghana
16Facultad de Ciencias Universidad Católica de la Santísima Concepción Concepción Chile espondence: Rahayu Adzhar (adzharrb@miamioh.edu) and Douglas I. 1
Introduction Tree cover values derived from Earth observation (EO) data
form a fundamental part of ecological research. They are
used to estimate forest cover change, biomass, and carbon
stocks (Bastin et al., 2019; Giriraj et al., 2017; Saatchi et
al., 2011; Song et al., 2014) and to help identify key areas
for conservation efforts (Miles et al., 2006) and as a basis
for climatic and vegetation modelling and model evaluation
(Brovkin et al., 2013; Burton et al., 2019; Kelley et al., 2013). All this research, in turn, plays a vital role in informing lo-
cal, regional, and global environmental policies (Harris et al.,
2012). As such, an EO product’s accuracy is important to
consider, as any errors in the initial tree cover estimate can
be further compounded in downstream work. Only a handful of EO products provide global maps of
percent tree cover or forest and shrub cover distributions
(Bartholomé and Belward, 2005; Bicheron et al., 2008), and
fewer still provide information stretching over at least a
decade (Friedl et al., 2002; Hansen et al., 2003; Sexton et
al., 2013; DiMiceli, 2017). Of these, one of the products
most widely used in ecological modelling is the Moderate
Resolution Imaging Spectroradiometer Vegetation Continu-
ous Fields (MODIS VCF) product (DiMiceli, 2017). MODIS
VCF is a yearly product that provides percent tree cover glob-
ally at a spatial resolution of 250 m. The most recent iteration
(Collection 6) is available for the years 2000 through to 2020. Its quantitative measure of woody cover is recorded annually
and is described as a percentage of ground cover, making it
particularly suited for use in evaluating dynamic global mod-
els (Lasslop et al., 2018; Rabin et al., 2017), as a proxy for in
situ data that are harder to collect (Kelley et al., 2019), and to
help define parameters for calculating global tree restoration
potential (Bastin et al., 2019). MODIS VCF is also used to
train alternative products, such as the newer finer-scale Land-
sat Tree Canopy Cover (TCC) product (Sexton et al., 2013). Tropical savannas have woody covers that fall within the
range particularly affected by the reported MODIS VCF er-
rors. A large proportion of these savannas can be found in
tropical developing countries (Boval and Dixon, 2012) and
are predicted to be home to half of the world’s population by
2050 (Archer et al., 2020). Tropical savannas are therefore
highly vulnerable to anthropogenic change. R. Adzhar et al.: VCF needs calibrating in tropical savannas 1378 R. Adzhar et al.: VCF needs calibrating in tropical savannas few and limited to sites within a biome (Montesano et al.,
2009), a region (Hansen et al., 2005; White et al., 2005), or
within a country (Gao et al., 2014; Sexton et al., 2013). The
MODIS VCF product evaluated was the most recent collec-
tion available at the time (i.e. Collection 3: Hansen et al.,
2005; White et al., 2005; Collection 4: Montesano et al.,
2009; Collection 5: Gao et al., 2015; Sexton et al., 2013). To
our knowledge, no such independent validation experiment
has yet been conducted on Collection 6, which produces tree
cover estimates in the same manner as Collection 5 but with
improvements made to the upstream inputs to enhance its ac-
curacy (DiMiceli, 2017). Likewise, validation of the finer-
scale TCC product has been limited to its penultimate ver-
sion and to the taiga–tundra circumpolar region (Montesano
et al., 2016). agated in the finer-scale Landsat TCC. Our scenarios sug-
gest that MODIS VCF accuracy can vary substantially, with
tree cover underestimation ranging from 0 % to 29 %. Mod-
els that use MODIS VCF as their benchmark could therefore
be underestimating the carbon uptake in forest–savanna ar-
eas and misrepresenting forest–savanna dynamics. Because
of the limited in situ plot number, our results are designed
to be used as an indicator of where the product is potentially
more or less reliable. Until more in situ data are available
to produce more accurate calibrations, we recommend cau-
tion when using uncalibrated MODIS VCF data in tropical
savannas. ,
)
The validations found that MODIS VCF may be less suit-
able for estimating tree cover in sparsely vegetated areas. Huang and Siegert (2006) noted that MODIS VCF classi-
fied large areas of land as “bare” where their land cover clas-
sification system identified it as sparsely vegetated. Mon-
tesano et al. (2009) found that MODIS VCF data (Collec-
tion 4) overestimated cover in areas of low tree cover in
taiga–tundra transition zones. Sexton et al. (2013) found that
the Collection 5 product overestimated cover in areas of low
cover (below 20 %) and underestimated in areas of higher
tree cover, while Gao et al. (2015) found that MODIS VCF
can only partially discriminate between tropical forest and
non-forest, struggling in areas that have greater heterogene-
ity. Similarly to MODIS VCF (Montesano et al., 2009), Mon-
tesano et al. R. Adzhar et al.: VCF needs calibrating in tropical savannas (2016) revealed an overestimation of the taiga–
tundra low tree covers in the finer-scale Landsat TCC, sug-
gesting that using VCF as training has propagated these over-
estimations into the higher-resolution product. What is clear
from the history of these validation and comparison experi-
ments is that MODIS VCF has accuracy issues in areas with
low woody-vegetation cover, which has implications when
its tree cover estimates are treated as accurately representa-
tive of real-world conditions. Failure to account for VCF’s
difficulty in estimating low woody covers can, therefore, lead
to miscalibrated models and estimations that do not reflect
real-world conditions. This, in turn, has knock-on effects on
environmental policymaking, conservation efforts, and future
ecological research, especially in areas with vegetation cover
types that are most prone to error. Published by Copernicus Publications on behalf of the European Geosciences Union. Published by Copernicus Publications on behalf of the European Geosciences Union. 2.2
Converting in situ canopy area index to MODIS
VCF–Landsat TCC percent tree cover We used the MODIS VCF Collection 6 product (250 m
spatial resolution, DiMiceli, 2017) with tree cover values
averaged across the years 2006 through to 2009 to reflect the
range of the field data collection period. MODIS VCF was
downloaded using the MODIS R package (Hijmans, 2017)
in R 3.5.2 (R Core Team, 2018). We used the 2005 and 2010
30 m Landsat TCC version 4 product (https://lcluc.umd.edu/
metadata/global-30m-landsat-tree-canopy-version-4,
last
access: 31 October 2021) and worked with the 2005 and 2010
average values. The product was downloaded manually from
https://e4ftl01.cr.usgs.gov/MEASURES/GFCC30TC.003/
(last access: 31 October 2021). CAI is defined as the sum of the projected areas of individ-
ual tree crowns divided by the ground area. In the TRO-
BIT project (Torello-Raventos et al., 2013; Veenendaal et
al., 2015), plot-wide CAI is made up of the sum of the
upper-stratum, mid-stratum, and subordinate-stratum crown
areas. Membership to a stratum is determined by the tree’s
dbh (diameter at breast height; upper stratum: dbh > 10 cm,
mid stratum: 2.5 cm < dbh < 10 cm, and subordinate stratum:
dbh < 2.5 cm; height > 1.5 m). About 50 trees per stratum
per plot were measured to derive plot-specific allometric
relations between stem diameter and crown area (Supple-
ment B of Torello-Raventos et al., 2013). These were then
applied to the whole plot to establish plot-level CAI. For the
allometric relationships, tree crowns were treated as circles,
and the individual tree projected crown area was determined
using the average of crown radii measured along the four car-
dinal points (i.e. from the centre of the stem to the distance
furthest from the stem). (
)
The in situ field data were sourced from the “Trop-
ical Biomes in Transition” project (TROBIT) (https://
www.forestplots.net, last access: 31 October 2021, Torello-
Raventos et al., 2013) and accessed via the https://www. forestplots.net database (Lopez-Gonzalez et al., 2011, 2009). The data we used include the corner locations and the canopy
area index (CAI) values for 17 forest and 31 savanna sites
distributed across Australia, Brazil, Bolivia, Cameroon, and
Ghana (Fig. 1 and Table A1 in the Appendix; Fig. 2 in
Torello-Raventos et al., 2013). R. Adzhar et al.: VCF needs calibrating in tropical savannas The classification of the TROBIT sites as either “forest” or
“savanna” is based on the parameters described in Torello-
Raventos et al. (2013) and Veenendaal et al. (2015). A sa-
vanna is a natural land cover that is not a forest, bare ground,
or body of water. Forest is defined as woody vegetation with
an average tree height of or exceeding 6 m and a canopy area
index (CAI) value of at least 0.3 for open forests and 0.7 for
forests. In addition, floristic differences (i.e. dominance of
savanna species) are used to differentiate forests from taller-
growing savannas that have similar CAIs and tree heights
(see Fig. 9 of Torello-Raventos et al., 2013). monitor the state, dynamics, and woody cover trends of trop-
ical savannas is a vital part of understanding how and why sa-
vannas are changing in the tropics (Harris et al., 2012; Miles
et al., 2006), while also improving modelled climate projec-
tions and vegetation dynamics for this complex biome. In this study, we evaluate MODIS VCF Collection 6 in
tropical savannas and forest areas by comparing VCF’s tree
cover percentage to corresponding field data. Similarly, we
evaluate Landsat TCC (version 4) to explore if, when VCF
is used as training, VCF biases are propagated. We then, for
MODIS VCF, characterize the observed bias in woody cov-
ers across both savanna and forest ecosystems and apply our
calibration across the tropics to highlight the regions most
likely affected by these inaccuracies. We finish by discussing
the implications the uncovered biases may have on tropical
vegetation and terrestrial biogeochemical modelling. There is some ambiguity in how savannas and “grass-
lands” are defined. Some modelling-based research treat the
two biomes as different (Whitley et al., 2017), while studies
based on plant functional traits group them together (Solo-
fondranohatra et al., 2018; White et al., 2000). As there is
some concern that MODIS VCF will struggle to pick up
woody cover in areas with really sparse vegetation, in this
paper we decided to treat grasslands as part of the savanna
domain. 1
Introduction In the face of a
growing population, land fragmentation, and changing cli-
mate, a savanna’s ability to maintain robust ecosystem func-
tions is directly linked to the amount of woody cover present
(Sankaran et al., 2006). As a result, the ability to accurately As the VCF product has progressed from Collection 1 to
its current Collection 6, several validations using in situ field
data or higher-resolution remotely sensed data as a refer-
ence measurement have been carried out. These have been Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 1379 2.2
Converting in situ canopy area index to MODIS
VCF–Landsat TCC percent tree cover The TROBIT field campaigns
were carried out over a 3-year period, from 2006 to 2009,
and the field plots used in this study are 1 ha in size except
for BFI-01 (0.5 ha), BFI-02 (0.5 ha), BFI-03 (0.5 ha), CTC-
01 (0.93 ha), and VCR-01 (0.6 ha). CAI values do not account for within-site tree canopy dis-
tribution patterns and the overlap between individual tree
canopies. We account for this by converting each CAI value
into a probability distribution function incorporating the fol-
lowing two extreme scenarios: “enforced overlap”, where the
location probability of individual canopies increases linearly
from 0 to 1 across a site, and “unenforced overlap”, where
individual canopies follow a uniform random distribution
pattern and canopy overlap is not purposefully introduced
(Fig. 2). We repeated this 1000 times per CAI measurement
to determine the probability distribution of expected CAI for
each field plot. All the sites fall within the tropics, that is, within 23.5◦
north and south of the Equator and were selected in re-
gions where savannas and forests were in close proximity
and within ecotones or “zones of tension”. As such, the sites
sampled show a large variation in physiognomy and edaphic
and climatic conditions (Table S1 in the Supplement; Vee-
nendaal et al., 2015). Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 1380
R. Adzhar et al.: VCF needs calibrating in tropical savannas
Figure 1. Location of sampling sites in Africa, Australia, and South America from the TROBIT project (based on Fig. 2 of Torello-Raventos
et al., 2013) shown in MODIS VCF (DiMiceli, 2017). Of the 63 field sites, only the 48 sites with available GPS coordinates were selected
(https://www.forestplots.net). R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1380 Figure 1. Location of sampling sites in Africa, Australia, and South America from the TROBIT project (based on Fig. 2 of Torello-Raventos
et al., 2013) shown in MODIS VCF (DiMiceli, 2017). Of the 63 field sites, only the 48 sites with available GPS coordinates were selected
(https://www.forestplots.net). Figure 2. Visual representation of the effects of enforcing overlap
within a (100 m × 100 m) TROBIT site with a given canopy area
index (CAI). (a) Overlap is not enforced, and individual crowns
follow a uniform random distribution. 2.2
Converting in situ canopy area index to MODIS
VCF–Landsat TCC percent tree cover (b) Overlap is enforced by
linearly increasing the probability of a canopy being located more
on one side of the site (i.e. here the right side of the site) than the
other. This results in tree canopies “overlapping” to a greater extent,
which affects how accurately CAI represents actual canopy cover. Unlike CAI, which is the fraction of ground covered by
tree crowns, the percent tree cover value from MODIS VCF
(and so Landsat TCC) is defined as “the portion of the sky-
light orthogonal to the surface which is intercepted by trees”
(Hansen et al., 2002). To make MODIS VCF and Landsat
TCC comparable to tree cover derived from TROBIT plot
CAIs, we divided these product values by 0.8 as suggested
by Hansen et al. (2002). This is also the standard approach in
most modelling studies using VCF (e.g. Lasslop et al., 2020;
Kelley et al., 2013; Burton et al., 2019). The 0.8 value can
be thought of as a gap correction factor (GCF) that accounts
for within-canopy gaps. Although the GCF has been shown
to vary with vegetation type (Lloyd et al., 2008; 0.34–0.60)
and crown cover (Tang et al., 2019b; 0.70–0.96), we opted
to use 0.8, as we found that it yielded more conservative re-
sults compared to a variable GCF. It also avoided introducing
additional parameters into our analysis. Figure 2. Visual representation of the effects of enforcing overlap
within a (100 m × 100 m) TROBIT site with a given canopy area
index (CAI). (a) Overlap is not enforced, and individual crowns
follow a uniform random distribution. (b) Overlap is enforced by
linearly increasing the probability of a canopy being located more
on one side of the site (i.e. here the right side of the site) than the
other. This results in tree canopies “overlapping” to a greater extent,
which affects how accurately CAI represents actual canopy cover. Next, to account for the difference in size between the
MODIS VCF pixel (250 m × 250 m) and the smaller field
plot size (100 m × 100 m), we calculated the possible percent
tree cover an area the size of a TROBIT field plot could have,
given the MODIS VCF percent tree cover for a MODIS-sized
pixel. 2.4
Mapping MODIS VCF uncertainty across the
tropics Figure 3. (a, c) Example of the effects of unenforced and enforced
clumping in a 250 m × 250 m MODIS VCF pixel with 50 % tree
cover. Clumping all the cover (green) to one side of the pixel (c)
affects the average canopy cover value of a 100 m × 100 m sized
TROBIT site (black boxes). (b, d) Example of the effects of unen-
forced and enforced clumping on 30 m × 30 m Landsat TCC pixels
with a mix of tree cover (green) and non-tree cover (brown). White
dotted lines are TCC pixel boundaries. Clumping all the cover to
one side of the pixel (d) affects the average canopy cover value of a
100 m × 100 m sized TROBIT site (black boxes). We evaluated the impact of the MODIS VCF biases inferred
from these regression equations across the tropics by in-
verting our calculation of MODIS VCF bias (Fig. A1) as
follows: first, the inverse (i.e. solving for C) of Eq. (1)
was applied to MODIS VCF values after conversion to a
100 m × 100 m pixel size grid (matching the field site area);
then this calibrated value was translated back to the origi-
nal 250 m × 250 m VCF pixel size. As the inverse of Eq. (1)
has no analytical solution, we found the rounded percent
value of C that minimizes the absolute difference between
the left- and right-hand side of the equation. For computa-
tional feasibility, we constructed maps of the tropics with
calibrated MODIS VCF values (Fig. A2) by randomly sam-
pling 5 iterations from each of our 10 optimization chains
(50 in total) and masking out pixels with cover types not con-
sidered forest or savanna in the 500 m MODIS Land Cover
Type (MCD12Q1 – Collection 6) (Sulla-Menashe and Friedl,
2018). 2.2
Converting in situ canopy area index to MODIS
VCF–Landsat TCC percent tree cover Equation (1) allowed us to assume nor-
mally distributed model errors, thus describing our condi-
tional probability of observations for a given parameter com-
bination by a normal distribution (Gelman et al., 2013). Each
combination was run over 10 chains, with 1000 warm-up it-
erations and 10 000 sampling iterations. Optimization was
performed using the rstan 2.19.2 (Stan Development Team,
2019) package in R 3.5.2 (R Core Team, 2018). Our op-
timization accounts for potential errors in TROBIT cover,
which includes those caused by the allometric construction
of the CAI, provided that the errors are unbiased and re-
main roughly consistent across sites (Gelman et al., 2013). As the TROBIT plots have relatively small total errors asso-
ciated with the allometric relationships (Table B1 in Torello-
Raventos et al., 2013), systematic errors are unlikely to affect
our results. to assume an increasing relationship between MODIS VCF–
Landsat TCC and C. Equation (1) allowed us to assume nor-
mally distributed model errors, thus describing our condi-
tional probability of observations for a given parameter com-
bination by a normal distribution (Gelman et al., 2013). Each
combination was run over 10 chains, with 1000 warm-up it-
erations and 10 000 sampling iterations. Optimization was
performed using the rstan 2.19.2 (Stan Development Team,
2019) package in R 3.5.2 (R Core Team, 2018). Our op-
timization accounts for potential errors in TROBIT cover,
which includes those caused by the allometric construction
of the CAI, provided that the errors are unbiased and re-
main roughly consistent across sites (Gelman et al., 2013). As the TROBIT plots have relatively small total errors asso-
ciated with the allometric relationships (Table B1 in Torello-
Raventos et al., 2013), systematic errors are unlikely to affect
our results. 2.2
Converting in situ canopy area index to MODIS
VCF–Landsat TCC percent tree cover This was done for two extreme scenarios: “enforced
clumping,” where all the tree cover for the given MODIS
VCF value is forcibly “clumped” on one side of the pixel,
or “unenforced clumping”, where “clumping” is not en-
forced and tree cover is distributed randomly within the pixel
(Fig. 3). The clumping scenarios introduce possible varia-
tions in percent cover due to the area and location mismatch
between a TROBIT field plot and a MODIS pixel. A proba-
bility distribution was generated for each MODIS VCF pixel
by calculating percent tree cover values for 1000 samples
(100 m × 100 m) randomly placed within the 250 m × 250 m
MODIS VCF pixel. field site size by summing the percent tree cover within the
TCC pixel part found inside the TROBIT field site and then
dividing the sum by the TROBIT site area. As TROBIT
site orientation was not recorded, we randomized the angle
between the TROBIT site and TCC pixel grid for each of
the 1000 samples when generating the probability distribu-
tion. Enforced clumping was performed as per MODIS VCF
(Fig. 3), with the direction of clumping randomized. For Landsat TCC, where the Landsat TCC pixels
(30 m × 30 m) are smaller than the TROBIT field sites, we
calculated a TCC percent tree cover to match the TROBIT https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 1381 R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas Figure 3. (a, c) Example of the effects of unenforced and enforced
clumping in a 250 m × 250 m MODIS VCF pixel with 50 % tree
cover. Clumping all the cover (green) to one side of the pixel (c)
affects the average canopy cover value of a 100 m × 100 m sized
TROBIT site (black boxes). (b, d) Example of the effects of unen-
forced and enforced clumping on 30 m × 30 m Landsat TCC pixels
with a mix of tree cover (green) and non-tree cover (brown). White
dotted lines are TCC pixel boundaries. Clumping all the cover to
one side of the pixel (d) affects the average canopy cover value of a
100 m × 100 m sized TROBIT site (black boxes). to assume an increasing relationship between MODIS VCF–
Landsat TCC and C. 2.3
Calculating uncertainty under different
clumping–overlap scenarios We compared both MODIS VCF and Landsat TCC with
TROBIT under four different scenarios: (1) unenforced over-
lap and clumping, (2) enforced overlap and unenforced
clumping, (3) unenforced overlap and enforced clumping,
and (4) enforced overlap and clumping. Comparisons were
conducted by fitting the following logit function: We then used the MCD12Q1 product to identify the ar-
eas of forest and savanna across the tropics in the MODIS
VCF product. MCD12Q1 is widely used by the global land
surface modelling community (e.g. Sellar et al., 2019; Wilt-
shire et al., 2021) and describes land cover in terms of 17
global land cover classes as per the International Geosphere-
Biosphere Programme (IGBP, Table 3 in Sulla-Menashe and
Friedl, 2018). The product is based on the same spectro-
radiometer (MODIS) and temporal resolution as the VCF
product. Referring to the definition of savanna of Veenendaal
et al. (2015), the following land cover classes were chosen
to represent savanna: closed shrublands, open shrublands,
woody savanna, savanna, and grassland; meanwhile forest
encompasses the following: evergreen needleleaf forests, ev-
ergreen broadleaf forests, deciduous needleleaf forests, de-
ciduous broadleaf forests, and mixed forests. We subset logit(Pixel) = C0 + 1 × log(Cτ1/(1 −Cτ2)),
(1) (1) where C0, 1, τ1, and τ2 are optimized parameters and
Pixel and C are the MODIS VCF–Landsat TCC pixel (post-
conversion as described in Sect. 2.2) and TROBIT site prob-
ability distributions, respectively. This is similar to a stan-
dard linear regression of logit-transformed data, account-
ing for maximum and minimum bounds of 0 %–100 % tree
cover, with τ1 and τ2 allowing for a non-symmetric trans-
formation of tree cover. To account for the probability den-
sity of each point, we inferred the parameters in Eq. (1) us-
ing a total least-squares Bayesian inference technique using a
Metropolis–Hastings Markov chain Monte Carlo step. Priors
were uninformed but physically bounded (i.e. 1, τ1, τ2 > 0) where C0, 1, τ1, and τ2 are optimized parameters and
Pixel and C are the MODIS VCF–Landsat TCC pixel (post-
conversion as described in Sect. 2.2) and TROBIT site prob-
ability distributions, respectively. This is similar to a stan-
dard linear regression of logit-transformed data, account-
ing for maximum and minimum bounds of 0 %–100 % tree
cover, with τ1 and τ2 allowing for a non-symmetric trans-
formation of tree cover. 3.1
Comparing MODIS VCF and Landsat TCC to tree
cover from TROBIT field sites The distribution of tree cover change after calibrating
against field data are similar across the four scenarios
(Fig. A2), and the regions where all four scenarios agree on
the direction of change (positive and negative) are substan-
tial (Fig. 6). However, there are some differences caused by
the uncertainty introduced by different extents of overlap and
clumping. While we see a significant increase in tree cover
across all clumping–overlap combinations in many regions
of tropical savannas and grasslands (Pennington et al., 2018),
such as in the forest–savanna mosaics that surround Congo-
lian rainforests, we do not see the same pattern in the Cerrado
of Brazil. This is likely because the African forest–savanna
regions fall within the range of MODIS VCF values that con-
sistently undergo a positive calibration (∼30 %–50 %, see
Fig. A1), while the Cerrado of Brazil does not. MODIS VCF underestimates tree cover within the 19 %
to 81 % range across all four combinations of enforced–
unenforced overlap and clumping (black line, Fig. 4). Be-
low 12 %, MODIS VCF tree cover values do not significantly
disagree with TROBIT field data and may instead be overes-
timating tree cover (50 % confidence, dashed line, Fig. 4). A similar pattern is seen when tree cover exceeds 84 %:
MODIS VCF does not differ significantly from TROBIT
when there is enforced overlap (i.e. when tree canopies are
clustering towards one side increasing the degree of canopy
overlap, Fig. 4 right) but may underestimate tree cover when
overlap is not enforced (i.e. tree canopies are spaced ran-
domly within the site, Fig. 4 left). There is a clear difference in how accurately MODIS VCF
estimates tree cover in forested areas (in green, Fig. 4) as
opposed to areas identified as savannas (in orange, Fig. 4). In savanna sites, MODIS VCF significantly and consistently
underestimates tree cover regardless of the amount of over-
lap and clumping. Significant underestimation (at 95 % con-
fidence) occurs when in situ tree cover exceeds 19 %–21 %
(without enforced clumping) or 11 %–12 % (with enforced
clumping). In forest sites, MODIS VCF does not show the
same pattern of systematic underestimation. Divergence does
occur at high covers, depending on the enforcement of over-
lap or clumping. R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas MCD12Q1 to the tropical zone between 30◦north and south
of the Equator and took the median class for the 2006 to 2009
period, matching the field data collection period. covers below 75 %–80 % (without enforced overlap) or be-
low 52 %–60 % (with enforced overlap). In forest sites (green
line, Fig. 5) there is no systematic difference. For a more detailed land-cover-specific evaluation, we
extracted the calibrated 250 m MODIS VCF pixel values
for each corresponding 500 m MCD12Q1 pixel to construct
land-cover-specific MODIS VCF tree cover frequency distri-
butions (Fig. A3). Our tree cover calibration by cover type
for the four clumping–overlap regression combinations was
then calculated by multiplying each cover type MODIS VCF
frequency distribution (Fig. A3) with curves representing the
median, 5 %, and 95 % confidence lines of the calibration
equation ensembles. 3
Results 3.1
Comparing MODIS VCF and Landsat TCC to tree
cover from TROBIT field sites 3.2
Global estimates of post-calibration change in
tropical tree cover We assessed the impact of VCF’s underestimation of inter-
mediate tree covers across the tropics (IGBP forest and sa-
vanna land cover classes), using a calibration based on the
combined forest and savanna sites (black curve, Fig. 4) in-
stead of using the savanna-only sites for a savanna-specific
calibration (orange curve, Fig. 4). This is because there were
few TROBIT sites representing savanna with MODIS VCF
tree cover values exceeding 40 %, and global land cover
maps disagree on the distribution of savannas within the
forest–savanna ecotone (Herold et al., 2008). 2.3
Calculating uncertainty under different
clumping–overlap scenarios To account for the probability den-
sity of each point, we inferred the parameters in Eq. (1) us-
ing a total least-squares Bayesian inference technique using a
Metropolis–Hastings Markov chain Monte Carlo step. Priors
were uninformed but physically bounded (i.e. 1, τ1, τ2 > 0) https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 1382 R. Adzhar et al.: VCF needs calibrating in tropical savannas The vertical error bars represent uncertainty introduced by clumping; the horizontal error bars represent the uncertainty introduced by
overlap. are most useful in identifying areas where MODIS VCF es-
timates may be more or less reliable. nas aligns with where the calibration for maximum overlap
and clumping is the largest (i.e. at about 20 % tree cover; see
Fig. A3), while the peak in cover distribution for woody sa-
vannas (26 %–67 %, Fig. A3) aligns with the cover range that
undergoes the greatest change (Fig. 7) in the other clumping
and overlap scenarios. R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1383 Figure 4. MODIS VCF percent tree cover versus percent tree cover from TROBIT field data, taking into account uncertainties associated
with tree cover spatial distributions within a MODIS pixel and/or field site. The four combinations are the following: (1) no overlap and no
clumping, where tree canopies are randomly distributed within both the pixel and site; (2) no overlap and maximum clumping, where tree
canopies are clustered in one area of the pixel and randomly distributed throughout the field site; (3) with overlap and no clumping, where
tree canopies are randomly distributed within the pixel but overlap substantially within the field site; and (4) with overlap and maximum
clumping, where tree canopies are clustered to one side within the pixel and overlap substantially within the site. The bolded dashed line in
black shows the 1 : 1 relationship. The solid lines represent the median of the respective regressions (green for forest, orange for savanna,
and black for forest and savanna combined), and the thin lines represent the 5 % and 95 % confidence interval of their respective regression
lines. The vertical error bars represent uncertainty introduced by clumping; the horizontal error bars represent the uncertainty introduced by
overlap. Figure 4. MODIS VCF percent tree cover versus percent tree cover from TROBIT field data, taking into account uncertainties associated
with tree cover spatial distributions within a MODIS pixel and/or field site. The four combinations are the following: (1) no overlap and no
clumping, where tree canopies are randomly distributed within both the pixel and site; (2) no overlap and maximum clumping, where tree
canopies are clustered in one area of the pixel and randomly distributed throughout the field site; (3) with overlap and no clumping, where
tree canopies are randomly distributed within the pixel but overlap substantially within the field site; and (4) with overlap and maximum
clumping, where tree canopies are clustered to one side within the pixel and overlap substantially within the site. The bolded dashed line in
black shows the 1 : 1 relationship. The solid lines represent the median of the respective regressions (green for forest, orange for savanna,
and black for forest and savanna combined), and the thin lines represent the 5 % and 95 % confidence interval of their respective regression
lines. 3.1
Comparing MODIS VCF and Landsat TCC to tree
cover from TROBIT field sites MODIS VCF underestimates tree cover
where tree cover exceeds 84 % (at the 95 % confidence inter-
val) when neither overlap nor clumping is enforced and over-
estimates where tree cover exceeds 78 % (at 5 % confidence
interval) when both overlap and clumping are enforced. Using field plots over a limited geographic extent creates
uncertainty that may be unaccounted for in our analysis when
our calibration is broadly applied across the highly variable
tropical-forest–savanna ecotone. By multiplying the uncer-
tainty range of our calibrations with the geographical dis-
tance to the closest sampled TROBIT site, we identified pri-
ority regions for further field surveying (Fig. 6 bottom). We
found Southeast Asia, Central America, and Mexico are ar-
eas where additional in situ observations would greatly help
improve confidence. Field data from the northwestern region
of South America, the southeast of the African continent, and
Madagascar would also help. As our calibrations were based on a limited number of sites
in a limited number of regions, it is important to note that
the maps shown in Figs. 6 and A2 are not definitive. For in-
stance, we found a significant tree cover decrease in the Sahel
post-calibration in multiple scenarios, which runs counter to
the results of Brandt et al. (2020), who found that tree cover
was underestimated in this region. This disparity may be due
to our lack of field sites in these more arid regions, further
highlighting the importance of more in situ data for more
accurate and precise calibration. Therefore, our calibrations Similar patterns can be observed with Landsat TCC (black
line, Fig. 5). There is a significant underestimation of tree
cover in the lower cover ranges up to 59 % when there is
enforced overlap and up to 82 % when overlap is not en-
forced. In savanna sites (orange line, Fig. 5) the underesti-
mation (at 95 % confidence) is significant and consistent for Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1384 R. Adzhar et al.: VCF needs calibrating in tropical savannas g
p
Figure 5. Landsat TCC percent tree cover versus percent tree cover from TROBIT field data, taking into account uncertainties associated
with tree cover spatial distributions within a TCC pixel and/or field site. The four combinations are the following: (1) no overlap and no
clumping, where tree canopies are randomly distributed within both the pixel and site; (2) no overlap and maximum clumping, where tree
canopies are clustered in one area of the pixel and randomly distributed throughout the field site; (3) with overlap and no clumping, where
tree canopies are randomly distributed within the pixel but overlap substantially within the field site; and (4) with overlap and maximum
clumping, where tree canopies are clustered to one side within the pixel and overlap substantially within the site. The bolded dashed line in
black shows the 1 : 1 relationship. The solid lines represent the median of the respective regressions (green for forest, orange for savanna,
and black for forest and savanna combined), and the thin lines represent the 5 % and 95 % confidence interval of their respective regression
lines. The vertical error bars represent uncertainty introduced by clumping; the horizontal error bars represent the uncertainty introduced by
overlap. Figure 5. Landsat TCC percent tree cover versus percent tree cover from TROBIT field data, taking into account uncertainties associated
with tree cover spatial distributions within a TCC pixel and/or field site. The four combinations are the following: (1) no overlap and no
clumping, where tree canopies are randomly distributed within both the pixel and site; (2) no overlap and maximum clumping, where tree
canopies are clustered in one area of the pixel and randomly distributed throughout the field site; (3) with overlap and no clumping, where
tree canopies are randomly distributed within the pixel but overlap substantially within the field site; and (4) with overlap and maximum
clumping, where tree canopies are clustered to one side within the pixel and overlap substantially within the site. The bolded dashed line in
black shows the 1 : 1 relationship. The solid lines represent the median of the respective regressions (green for forest, orange for savanna,
and black for forest and savanna combined), and the thin lines represent the 5 % and 95 % confidence interval of their respective regression
lines. R. Adzhar et al.: VCF needs calibrating in tropical savannas The vertical error bars represent uncertainty introduced by clumping; the horizontal error bars represent the uncertainty introduced by
overlap. 3.3
Post-calibration change in tree cover within
different vegetation classes in tropical ecosystems When looking at our calibration in more detail, we see that
MODIS VCF significantly underestimates tree cover in all
the IGBP land cover classes that we considered, regardless
of overlap or clumping (95 % confidence interval) (Fig. 7). The most substantial and significant underestimation is in the
classes “woody savannas” and savannas. The underestima-
tion is the largest in woody savannas, except when clumping
and overlap are enforced (in purple, Fig. 7). This is because
the peak in the tree cover frequency distribution for savan- “Open shrublands” only show a small underestimation of
tree cover, despite the woody cover definition (10 %–60 %)
matching the range where MODIS VCF most underestimates
tree cover (26 %–67 % cover). The discrepancy may be be-
cause the majority of the open-shrublands class commission
error is with the grasslands class (see Table S6 in Sulla-
Menashe et al., 2019). The MODIS VCF tree cover in areas
classified as open shrublands is therefore likely to be lower https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 R. Adzhar et al.: VCF needs calibrating in tropical savannas (b) Uncertainty range
of the post-calibration change in tree cover, calculated as the 90th
percentile (maximum of the four scenarios in Fig. A2) minus the
10th percentile (minimum of the four scenarios in Fig. A2). (c) Ge-
ographic distance to the closest TROBIT site sampled. (d) Regions
coloured to denote priority for field surveying to constrain map un-
certainty (based on multiplying the uncertainty range of each pixel
with the pixel’s geographical distance to the closest TROBIT site
sampled). Figure 6. (a) The post-calibration change in tree cover that is sta-
tistically significant (95 % interval) in the same direction (positive
or negative calibration leading to an increase or decrease in tree
cover, respectively) across all four scenarios. (b) Uncertainty range
of the post-calibration change in tree cover, calculated as the 90th
percentile (maximum of the four scenarios in Fig. A2) minus the
10th percentile (minimum of the four scenarios in Fig. A2). (c) Ge-
ographic distance to the closest TROBIT site sampled. (d) Regions
coloured to denote priority for field surveying to constrain map un-
certainty (based on multiplying the uncertainty range of each pixel
with the pixel’s geographical distance to the closest TROBIT site
sampled). We also found discrepancies between the tree cover val-
ues derived from MODIS VCF and the corresponding class
definition of the MCD12Q1 product (Fig. A3) which again
suggests that the 5 m height threshold may not always ap-
ply in MODIS VCF. For example, MODIS VCF recorded
tree cover in the open-shrublands and “closed-shrublands”
classes of the MCD12Q1 product (Fig. A3), even though the
height range for these classes is 1–2 m. For the savanna class,
MODIS VCF yields a percent tree cover range that matches
closely with the savanna class definition (between 10 % and
30 %), despite the differing tree thresholds for MODIS VCF
and IGBP (5 m minimum for MODIS VCF and 2 m mini-
mum for IGBP). These discrepancies suggest one of the fol-
lowing three things: open/closed shrublands and savannas
contain trees taller than 5 m, MODIS VCF is distinguishing
trees below the 5 m threshold, or some combination of both. flects the TROBIT savanna plots. This is evidenced by work
done by Veenendaal et al. (2015), where TROBIT plots were
tested for complete spatial randomness, and only minor indi-
cations of overlap were found. 4
Discussion than the IGBP definition would suggest (see Fig. A3), result-
ing in calibrations that are more conservative. We found significant increases in tree cover for forests in
every calibration scenario, though net change is not signifi-
cant (95 % confidence) when overlap is enforced. This can be
explained by the presence of both negative and positive cal-
ibrations in the higher ranges of tree cover when overlap is
enforced. Similarly, the net change is insignificant across all
clumping and overlap scenarios for the IGBP classes match-
ing the lower ranges of tree cover (grasslands, closed shrub-
lands, and open shrublands). While MODIS VCF is a powerful and accessible tool to map
tree cover, our field-data-based calibrations indicate that the
latest MODIS VCF Collection 6 is missing a lot of woody
cover, even when uncertainty introduced by site canopy over-
lap and clumping within the MODIS VCF pixel are ac-
counted for. The Landsat TCC product, which may be viewed
as an alternative with a higher spatial resolution, behaves in
a similar manner. Our map (Fig. 6 top) highlights that this
potential underestimation of woody cover is mainly occur-
ring in tropical savannas. Moreover, the highest underestima-
tion in the savanna classes occurs when there is no enforced
overlap (Fig. A2) (i.e. when there is a uniform random dis-
tribution of trees) which is the scenario that most likely re- Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1385 R. Adzhar et al.: VCF needs calibrating in tropical savannas clumping and overlap or 0 %–21 % for when either clumping
or overlap are enforced (5 %–95 % confidence). An overestimation at the lower end of the cover (< 20 %)
(Hansen et al., 2002; Sexton et al., 2013) and underesti-
mation in the lower to middle range of cover (20 %–60 %)
have been identified in validations of previous MODIS VCF
collections (Gross et al., 2018; Yang and Crews, 2019) and
Landsat TCC versions (Montesano et al., 2016). According
to its definition, MODIS VCF only maps trees that are 5 m
or taller (Hansen et al., 2003), while the TROBIT CAI in-
cludes all trees with a minimum dbh of 2.5 cm, as well as
trees with a height exceeding 1.5 m when dbh < 2.5 cm. This
could explain our observed underestimation in the lower tree
cover ranges for both MODIS VCF and Landsat TCC. In fact
Montesano et al. (2016) showed an improved match between
Landsat TCC and their lidar-derived tree cover reference data
when reducing the height threshold from 5 to 2 m. However,
because of how our field reference CAI is derived, we were
not able to conclusively link the 5 m threshold to our ob-
served underestimation. On the other hand, when looking at the relationship be-
tween TROBIT’s upper-stratum canopy height and the dif-
ference between TROBIT and VCF, we would have expected
an increasing underestimation in the lower height ranges. In-
stead we found a low R2 and a mixture of under- and overes-
timations in heights between 0 and 10 m (Fig. A4). This sug-
gests that the inclusion of trees below 5 m height in the TRO-
BIT inventory does not fully explain the observed underesti-
mation. However, as the relationship between upper canopy
heights and the subordinate strata composition (and canopy
cover thereof) varies widely depending on factors including
ecosystem type and altitude (Rutten et al., 2015), more re-
search needs to be done with in situ height data. Figure 6. (a) The post-calibration change in tree cover that is sta-
tistically significant (95 % interval) in the same direction (positive
or negative calibration leading to an increase or decrease in tree
cover, respectively) across all four scenarios. R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1386 R. Adzhar et al.: VCF needs calibrating in tropical savannas
ange in tree cover post-calibration (clockwise: no enforced clumping or overlap (black), enforced clumping and no
e), no enforced clumping and enforced overlap (red), and enforced clumping and overlap (pink)) in the forest supercat-
vanna classes. The palest tone indicates positive change; the mid tone indicates negative change; and the darkest tone
Error bars denote the 5 %–95 % confidence interval; if the error bar extends past the x axis, the post-calibration change
nificant. Figure 7. Percent change in tree cover post-calibration (clockwise: no enforced clumping or overlap (black), enforced clumping and no
enforced overlap (blue), no enforced clumping and enforced overlap (red), and enforced clumping and overlap (pink)) in the forest supercat-
egory and the five savanna classes. The palest tone indicates positive change; the mid tone indicates negative change; and the darkest tone
indicates net change. Error bars denote the 5 %–95 % confidence interval; if the error bar extends past the x axis, the post-calibration change
is not considered significant. the training data (Gerard et al., 2017) and using a CART
(classification and regression tree) model (Hanan et al., 2013)
are inherent and still present within this version of MODIS
VCF. Similar results for MODIS VCF and Landsat TCC also
suggest that by training TCC with VCF tree cover these bi-
ases have been propagated into the finer-scale product. There
is a risk of bias propagation when MODIS VCF (or related
products) is used as a single source for benchmarking mod-
els (e.g. Brandt et al., 2017; Lasslop et al., 2020; Burton et
al., 2019; Kelley et al., 2019, 2021). To avoid this, studies
should try use multiple data sources (e.g. for woody cover
possibly the GEDI – Global Ecosystem Dynamics Investiga-
tion – lidar-based canopy cover product, Tang et al., 2019a)
whether for model calibration/validation (e.g. Sellar et al.,
2019; Wiltshire et al., 2021; Burton et al., 2021) or hypothe-
sis testing (e.g. Taylor et al., 2012). open shrublands, woody savannas, and savannas. The ma-
jority of the open-shrublands class commission error is with
the grasslands class, and there is confusion to a lesser ex-
tent between open shrublands, woody savannas, and savan-
nas. R. Adzhar et al.: VCF needs calibrating in tropical savannas Woody savannas, as an exam-
ple, may have their tree cover underestimated by up to 32 %
(95 % confidence) when neither clumping nor overlap is en-
forced (grey tones, Fig. 7). If our results are representative of
the tropics, then overall, MODIS VCF may be underestimat-
ing tropical tree cover by between 7 %–29 % for unenforced Another explanation for the discrepancy between the
IGBP class definitions and those estimated through MODIS
VCF could be the between-class omission and commission
errors (Fig. 5 in the present paper and Table S6 in Sulla-
Menashe et al., 2019). For example, the accuracy for closed
shrublands is particularly low. It is mainly confused with https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 R. Adzhar et al.: VCF needs calibrating in tropical savannas As this tree cover
is likely to have been underestimated substantially, the de-
rived available land space for replanting may be less than
projected, with the restoration potential being overestimated. However, our results also indicate an underestimated tree
cover in woodier savannas and forests. Accounting for this,
the restoration potential could actually be greater than an-
ticipated because the carrying capacity of a unit of land
could be greater than previously thought. Calibration could
also result in a more uniform cover distribution across re-
gions, producing a more gradual transition between low-
cover savannas and high-cover forests. This could have im-
plications for work that, for example, uses MODIS VCF to
study forest–savanna dynamics and bi-stability (Lasslop et
al., 2018; Wuyts et al., 2017; Xu et al., 2016). Finally, factors such as cloud cover, landscape heterogene-
ity, phenology, vegetation type, and soil type affect the ac-
curacy of remotely sensed products like MODIS VCF and
Landsat TCC (Hansen et al., 2003; Huete et al., 1997; Smith
et al., 2002). Data characterizing these at the plot level would
help identify potential confounding factors affecting perfor-
mance and so help further constrain uncertainties. Alternatively, comparing MODIS VCF to other land cover
maps or higher-resolution remotely sensed data are recom-
mended (Gross et al., 2018; Lary and Lait, 2006; Monte-
sano et al., 2016), though without a large-scale effort to re-
calibrate MODIS VCF and products trained using VCF like
Landsat TCC, the question of how appropriate MODIS VCF
is for use in both forests and savannas in the tropics will
remain. By highlighting the extent to which MODIS VCF
struggles to estimate tree cover in tropical forests and savan-
nas, we hope to inform the future use of this product and
improve its useability. y
To ensure the appropriate use of both products, we suggest
that where field data are available, the products should be cal-
ibrated for use in the target region. However, calibrating on
a large scale using field data as a reference presents several
challenges. Firstly, different in situ measurement techniques
tend to measure different types of tree cover (e.g. Fiala et al.,
2006; Korhonen et al., 2006; Rautiainen et al., 2005), and
each will require a specific conversion method to enable di-
rect comparison with MODIS VCF or Landsat TCC. For ex-
ample, the comparison of Montesano et al. R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas We suggest that while MODIS VCF and landsat TCC give
a good overview of tree cover on a global scale, both should
be used cautiously in savanna regions. Special care should
be taken in savannas, a biome that has long been noted as be-
ing challenging for EO products to characterize, as solitary
trees in the landscape tend to be missed by global tree cover
products (Jung et al., 2006; Brandt et al., 2020). The poor
performance of MODIS VCF and Landsat TCC in savannas
in particular (Gaughan et al., 2013; Gross et al., 2018; Kumar
et al., 2019) emphasizes the importance of continuous inde-
pendent validation and re-calibration of these products. The
ecosystem functions of savannas can vary drastically with
just a slight difference in tree cover (Gaughan et al., 2013),
and even slight errors may create issues in how we interpret
the state and dynamics of the biome, which in turn affects
how the land is managed. and show no systematic bias and are therefore not expected
to significantly change our results. However, our results in
Fig. 6 have been extrapolated from a limited number of field
sites, with somewhat limited distribution across the tropics. While our uncertainty map gives a good idea of areas of con-
cern, for a more robust description of MODIS VCF’s accu-
racy, we would need substantially more widely distributed in
situ sites to represent the tropics. Field data collection re-
mains costly and labour intensive, and adopting protocols
that ensure standardization and data sharing via information
platforms would facilitate large-scale validation exercises. In
the case of validating woody cover products, including direct
tree cover measurements such as crown area would also help. However, the reality for much of pre-existing data is that
they are not standardized, and field data (whether standard-
ized or not) are not designed to be directly comparable to
remote-sensing-derived variables. Here, our technique offers
a potential solution to this. By accounting for uncertainties
that arise from differences between the available in situ data
and the remotely sensed variable, we are able to use in situ
data, which may initially seem unsuitable, to carry out a more
conservative evaluation. Work on forest restoration potential would also be im-
pacted. Bastin et al. (2019), for example, used MODIS VCF
to estimate tree cover in agricultural land. R. Adzhar et al.: VCF needs calibrating in tropical savannas (2016) did not
acknowledge VCF’s and thus TCC’s “within canopy gaps”,
which may explain their observed underestimation in cov-
ers above 80 %. In our case, to account for gaps between
tree crowns, we applied the 0.8 “gap correction factor” to
the CAI. However, the GCF and resulting tree cover could
vary widely on a plot-by-plot basis (Lloyd et al., 2008). With
further in situ data that describe tropical vegetation-type-
specific GCF variation, we may be able to incorporate site-
specific GCFs into our analysis. R. Adzhar et al.: VCF needs calibrating in tropical savannas Also, the “cropland/natural vegetation mosaics” class is
often mapped as closed shrublands, woody savannas, savan-
nas, or grasslands. More work needs to be done to evaluate how effective both
MODIS VCF, Landsat TCC, and MCD12Q1 are at imple-
menting the height thresholds in their respective “tree” def-
initions, as this may have implications when MODIS VCF,
Landsat TCC, and MCD12Q1 are used for global model cal-
ibration or validation. Overall, our results suggest that the biases found in the
previous collections may have persisted in Collection 6, de-
spite reported improvement in accuracy (DiMiceli et al.,
2017). This indicates that the biases introduced by binning Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 1387 Appendix A Appendix A Table A1. Site names, locations, canopy area index values, MODIS VCF percent tree cover values, cover type, and TROBIT site descriptions
of the 48 TROBIT project plots used in this study. TROBIT site descriptions are based on Table S1 of Veenendaal et al. (2015). of the 48 TROBIT project plots used in this study. TROBIT site descriptions are based on Table S1 of Veenendaal et al. (2015). 5
Conclusion We found that MODIS VCF significantly underestimates tree
cover in tropical forests and savannas, even when within-site
and pixel–site variation are accounted for in calibration. We
also found that using MODIS VCF for training likely prop-
agates these biases, even in the finer-scale Landsat TCC. As MODIS VCF is a product that is commonly used in
a wide variety of ecological research including vegetation
modelling, estimating restoration potential, and identifying
forest–savanna bimodality, we stress that more independent
work on validating and re-calibrating is required before its
tree cover estimates can be relied upon in the tropics. There is also the uncertainty associated with the field data
collection. In our case, the site-specific CAI standard errors
(Supplement B in Torello-Raventos et al., 2013) are small https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas 1388 Appendix A Adzhar et al.: VCF needs calibrating in tropical savannas Appendix A Site
Country
Latitude
Longitude
MODIS
Canopy
Average
Cover
TROBIT site
name
VCF tree
area
upper-
type
description
cover
index
stratum
(%)
height (m)
ALC-01
Brazil
−2.53
−54.91
12.5
0.32
6.56
Savanna
Savanna woodland
ALF-01
Brazil
−9.6
−55.94
77
2.31
37.02
Forest
Tall forest
ALF-02
Brazil
−9.58
−55.92
76
2.65
41.32
Forest
Tall forest
ASU-01
Ghana
7.14
−2.45
41.33
2.54
45.27
Forest
Tall forest
BBI-01
Burkina Faso
12.73
−1.17
1.33
0.52
12.53
Savanna
Savanna woodland
BBI-02
Burkina Faso
12.73
−1.16
1.5
0.99
13.6
Savanna
Savanna woodland
BDA-01
Burkina Faso
10.94
−3.15
6.17
0.3
14.53
Savanna
Shrub-rich savanna woodland
BDA-02
Burkina Faso
10.94
−3.15
4.5
0.18
14.47
Savanna
Shrub-rich savanna woodland
BFI-01
Ghana
7.71
−1.69
15
1.22
29.67
Savanna
Tall closed woodland
BFI-02
Ghana
7.71
−1.69
12.83
1.08
28.2
Savanna
Tall savanna woodland
BFI-03
Ghana
7.71
−1.7
25.83
2.54
45.07
Savanna
Tall savanna woodland
CTC-01
Australia
−16.1
145.45
72.67
2.35
40.37
Forest
Tall forest
DCR-01
Australia
−17.02
145.58
21.67
1.67
27.19
Savanna
Tall savanna woodland
DCR-02
Australia
−17.03
145.6
65.67
0.71
22.51
Savanna
Tall savanna woodland
EKP-01
Australia
−18.07
145.99
43.5
0.74
28.13
Savanna
Tall savanna woodland
FLO-01
Brazil
−12.81
−51.85
65.67
2.4
28.21
Forest
Forest
FMS-01
Australia
−18.09
144.84
7.67
0.32
20.03
Savanna
Shrub-rich savanna woodland
FMS-02
Australia
−18.11
144.82
44.17
1.21
16.69
Forest
Stunted shrub-rich forest
HOM-01
Mali
15.34
−1.47
0.5
0.05
3.87
Savanna
Savanna grassland
HOM-02
Mali
15.33
−1.55
0.83
0.16
6.13
Savanna
Savanna grassland
IBG-01
Brazil
−15.95
−47.87
20.83
0.22
7.48
Savanna
Scrub savanna
IBG-02
Brazil
−15.95
−47.87
20
0.02
6.29
Savanna
Scrub savanna
IBG-03
Brazil
−15.93
−47.87
20.5
0.12
8.01
Savanna
Scrub savanna
IBG-04
Brazil
−15.94
−47.86
27.17
0.77
12.65
Savanna
Savanna woodland
KBL-01
Australia
−17.77
145.54
75
1.69
39.5
Forest
Tall forest
KBL-02
Australia
−17.85
145.53
61.17
0.81
29.2
Savanna
Tall savanna woodland
KBL-03
Australia
−17.69
145.53
79.5
3
36.62
Forest
Tall forest
KCR-01
Australia
−17.11
145.6
78.83
2.44
42.37
Forest
Tall forest
LFB-03
Bolivia
−14.6
−60.85
28.17
0.39
9.93
Savanna
Shrub-rich savanna woodland
MDJ-01
Cameroon
6.17
12.83
42
3.24
45
Forest
Tall forest
MDJ-02
Cameroon
6.16
12.82
18.67
0.44
16.13
Savanna
Long-grass savanna
MDJ-03
Cameroon
5.98
12.87
64.67
2.97
36.53
Forest
Stunted shrub-rich forest
MDJ-04
Cameroon
6
12.87
15
0.37
18.93
Savanna
Long-grass savanna
MDJ-05
Cameroon
5.98
12.87
70.33
2.85
21.27
Forest
Stunted shrub-rich forest
MDJ-06
Cameroon
6
12.89
20.5
0.68
15.27
Savanna
Long-grass savanna
MDJ-07
Cameroon
6.01
12.89
57.33
1.75
42.67
Forest
Tall forest
MDJ-08
Cameroon
6.21
12.75
15
0.48
18
Savanna
Long-grass savanna
MLE-01
Ghana
9.3
−1.86
10
0.34
14.67
Savanna
Savanna woodland
NXV-02
Brazil
−14.7
−52.35
20.83
1.82
15.76
Savanna
Tall closed woodland
RSC-01
Australia
−20.16
146.54
28
1.15
13.14
Forest
Stunted forest
SMT-01
Brazil
−12.82
−51.77
36.67
1.55
14.37
Savanna
Savanna woodland
SMT-02
Brazil
−12.82
−51.77
41.5
1.44
14.64
Savanna
Savanna woodland
SMT-03
Brazil
−12.83
−51.77
19.33
0.53
11.19
Savanna
Savanna woodland
TUC-01
Bolivia
−18.52
−60.81
50.33
1.29
14.9
Forest
Stunted forest
TUC-02
Bolivia
−18.53
−60.63
21.67
0.81
12.05
Savanna
Shrub-rich woodland
TUC-03
Bolivia
−18.19
−60.86
10.83
0.37
14.11
Savanna
Savanna woodland
VCR-01
Brazil
−14.83
−52.16
69.5
2.81
28.94
Forest
Tall forest
VCR-02
Brazil
−14.83
−52.17
69.67
2.74
30.93
Forest
Forest https://doi.org/10.5194/bg-19-1377-2022 Biogeosciences, 19, 1377–1394, 2022 R. R. Adzhar et al.: VCF needs calibrating in tropical savannas 1389 VC
g
p
Figure A1. The calibration curves developed for MODIS VCF based on four pixel–site mismatch scenarios (no clumping and no overlap,
enforced clumping and no overlap, no clumping and enforced overlap, and enforced clumping and enforced overlap). The dashed line
signifies the “ideal” 1 : 1 relationship wherein calibrated MODIS VCF is unchanged from the original MODIS VCF values. The shaded
regions represent the 5 % to 95 % confidence intervals for the respective calibrated MODIS VCF values. Figure A1. The calibration curves developed for MODIS VCF based on four pixel–site mismatch scenarios (no clumping and no overlap,
enforced clumping and no overlap, no clumping and enforced overlap, and enforced clumping and enforced overlap). The dashed line
signifies the “ideal” 1 : 1 relationship wherein calibrated MODIS VCF is unchanged from the original MODIS VCF values. The shaded
regions represent the 5 % to 95 % confidence intervals for the respective calibrated MODIS VCF values. Figure A2. The change in tree cover post-calibration for all four scenarios. Black dots indicate areas where the post-calibration values have
a 95 % certainty of being positive (increasing cover) or negative (decreasing cover) calibrations. These uncertainty maps are indicators of
areas where MODIS VCF estimates may be more or less reliable and cannot be used as definitive calibrations due to the limited number of
field sites used as reference. Figure A2. The change in tree cover post-calibration for all four scenarios. Black dots indicate areas where the post-calibration values have
a 95 % certainty of being positive (increasing cover) or negative (decreasing cover) calibrations. These uncertainty maps are indicators of
areas where MODIS VCF estimates may be more or less reliable and cannot be used as definitive calibrations due to the limited number of
field sites used as reference. Biogeosciences, 19, 1377–1394, 2022 https://doi.org/10.5194/bg-19-1377-2022 https://doi.org/10.5194/bg-19-1377-2022 1390 R. Adzhar et al.: VCF needs calibrating in tropical savannas R. Adzhar et al.: VCF needs calibrating in tropical savannas the National Council for Scientific and Technological Development
(CNPq, grant no. 301153/2018-3). the National Council for Scientific and Technological Development
(CNPq, grant no. 301153/2018-3). Code and data availability. The code and data used to support the
findings of this study are archived at https://github.com/douglask3/
VCF_vs_sites with revision number fdda3ff (Adzhar et al., 2022). Upon being granted access, the data are available at https://www. forestplots.net and can be found using the site names we have listed
in Table A1 (Torello-Raventos et al., 2013). Review statement. This paper was edited by Alexandra Konings
and reviewed by three anonymous referees. Review statement. This paper was edited by Alexandra Konings
and reviewed by three anonymous referees. MODIS VCF is accessible via the MODIS R package
(http://CRAN.R-project.org/package=raster;
Hijmans,
2017). The product itself should be attributed to DiMiceli (2017,
https://doi.org/10.5067/MODIS/MOD44B.006). References TCC
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are
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English
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Evaluating the Efficacy of Seattle-PAP for the Respiratory Support of Premature Neonates: Study Protocol for a Randomized Controlled Trial
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Research Square (Research Square)
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cc-by
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© The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Abstract Background: At birth, the majority of neonates born at <30 weeks of gestation require respiratory support to
facilitate transition and ensure adequate gas exchange. Although the optimal approach to the initial respiratory
management is uncertain, the American Academy of Pediatrics endorses noninvasive respiratory support with nasal
continuous positive airway pressure (nCPAP) for premature neonates with respiratory insufficiency. Despite evidence for
its use, nCPAP failure, requiring intubation and mechanical ventilation, is common. Recently, investigators have
described a novel method to deliver bubble nCPAP, termed Seattle-PAP. While preclinical and pilot studies
are encouraging regarding the potential value of Seattle-PAP, a large trial is needed to compare Seattle-PAP
directly with the current standard of care for bubble nCPAP (Fisher & Paykel CPAP or FP-CPAP). Methods/design: We designed a multicenter, non-blinded, randomized controlled trial that will enroll 230
premature infants (220/7 to 296/7 weeks of gestation). Infants will be randomized to receive Seattle-PAP or FP-
CPAP. The primary outcome is respiratory failure requiring intubation and mechanical ventilation. Secondary
outcomes include measures of short- and long-term respiratory morbidity and cost-effectiveness. Discussion: This trial will assess whether Seattle-PAP is more efficacious and cost-effective than FP-CPAP in
real-world practice among premature neonates. Trial registration: ClinicalTrials.gov, NCT03085329. Registered on 21 March 2017. Keywords: Neonate, Continuous positive airway pressure (CPAP), Preterm Infant, Infant, Costs, Costs analysis,
Randomized controlled trial Keywords: Neonate, Continuous positive airway pressure (CPAP), Preterm Infant, Infant, Costs, Costs analysis,
Randomized controlled trial capacity are critical tasks in a newborn’s adaptation to
the extrauterine environment. Beginning with their ini-
tial breath after birth, term infants clear fluid from their
lungs and establish functional residual capacity to facili-
tate gas exchange [2]. In contrast, among many preterm
neonates, surfactant deficiency, immature epithelial so-
dium channels, instability of the thorax, respiratory fa-
tigue, and apnea contribute to respiratory insufficiency,
often resulting in respiratory failure and the need for re-
spiratory support [3–5]. Although the best approach to
the initial respiratory management of these infants is Evaluating the efficacy of Seattle-PAP for
the respiratory support of premature
neonates: study protocol for a randomized
controlled trial Carl H. Backes1,2,3*, Jennifer L. Notestine1, Jane M. Lamp4, Jeanne C. Balough1, Allison M. Notestine1,
Crystal M. Alfred2, Juli M. Kern5, Michael R. Stenger2, Brian K. Rivera1, Mohannad Moallem2, Randy R. Miller5,6,
Apurwa Naik7, Jennifer N. Cooper2,8,9, Christopher R. Howard10, Stephen E. Welty10,11, Noah H. Hillman12,
John A. F. Zupancic13,14, Larissa I. Stanberry15, Thomas N. Hansen10 and Charles V. Smith10 STUDY PROTOCOL Open Access Backes et al. Trials (2019) 20:63
https://doi.org/10.1186/s13063-018-3166-6 Backes et al. Trials (2019) 20:63
https://doi.org/10.1186/s13063-018-3166-6 STUDY PROTOCOL
Open Access
Evaluating the efficacy of Seattle-PAP for
the respiratory support of premature
neonates: study protocol for a randomized
controlled trial
Carl H. Backes1,2,3*, Jennifer L. Notestine1, Jane M. Lamp4, Jeanne C. Balough1, Allison M. Notestine1,
Crystal M. Alfred2, Juli M. Kern5, Michael R. Stenger2, Brian K. Rivera1, Mohannad Moallem2, Randy R. Miller5,6,
Apurwa Naik7, Jennifer N. Cooper2,8,9, Christopher R. Howard10, Stephen E. Welty10,11, Noah H. Hillman12,
John A. F. Zupancic13,14, Larissa I. Stanberry15, Thomas N. Hansen10 and Charles V. Smith10 * Correspondence: Carl.Backes@nationwidechildrens.org;
carl.backesjr@nationwidechildrens.org
1Center for Perinatal Research, The Research Institute at Nationwide
Children’s Hospital, 700 Children’s Drive, Columbus, OH 43205, USA
2Department of Pediatrics, The Ohio State University Wexner Medical Center,
Columbus, OH, USA
Full list of author information is available at the end of the article Background Neonates face significant challenges in the successful
transition from fetal life [1, 2]. Aeration of liquid-filled
lungs
and
the
establishment of
functional
residual © The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Backes et al. Trials (2019) 20:63 Backes et al. Trials (2019) 20:63 Page 2 of 10 In general, bubble nCPAP delivery systems consist of
six primary components (Fig. 1): (1) sources of air, oxy-
gen, and other breathing gases, (2) a blender to mix
these gases together, (3) a gas heater and humidifier, (4)
inspiratory and expiratory tubing (breathing circuit), (5)
a patient interface (e.g., bi-nasal prongs or a nasal mask),
and (6) a pressure generator (e.g., bubbler device). The
design of the bubbler device can take many forms, but
they generally comprise a tube with its distal end sub-
merged in a body of water. The airway pressure provided
by the system is determined largely by the depth of the
distal end of the bubbler tube below the surface of the
water (5 cm below surface = 5 cm H2O). In conventional
bubble nCPAP systems, the bubbler tube points straight
down (perpendicular to the surface of the water), which
we define as 0°. With Seattle-PAP, the vertical section of
the bubbler tube is connected to a horizontal section,
which is connected to an upward sloping section, which
we optimized at 135°. uncertain, evidence that even brief exposures to tracheal
intubation and mechanical ventilation cause lung injury
have led to interest among health-care providers in the
use of noninvasive respiratory support devices, primarily
nasal continuous positive airway pressure (nCPAP) [6]. The American Academy of Pediatrics has endorsed
the use of nCPAP among premature neonates with
respiratory distress [7]. This recommendation is based
on data indicating lower rates of bronchopulmonary
dysplasia (BPD) in infants treated with nCPAP than
are observed with more invasive modes of ventilatory
support [8–10]. Background However, nCPAP failure, requiring in-
tubation and mechanical ventilation in preterm in-
fants, is common, with failure rates exceeding 50% in
large clinical trials [8–10]. In developed countries,
CPAP failure is associated with greater morbidity; in
developing countries, CPAP failure is associated with
greater mortality [11, 12]. More recently, bubble nCPAP has re-emerged as a po-
tential strategy to address high nCPAP failure rates [13]. Although there are limited data, neonates on bubble
nCPAP had a lower incidence of respiratory failure (tra-
cheal intubation and mechanical ventilation) than did in-
fants supported on ventilator-derived CPAP [12, 14]. Despite evidence that bubble nCPAP may be advanta-
geous for preterm infants, the optimal delivery for new-
borns with respiratory distress is unknown. To address
high failure rates associated with CPAP among preterm
neonates, investigations of novel strategies to deliver more
effective bubble nCPAP are warranted [15]. One potential
strategy is the use of Seattle-PAP [12, 13, 16, 17]. p
Preclinical evidence demonstrated that this 135° modi-
fication provides fluctuations in airway pressures, includ-
ing lower-frequency pressure oscillations than those
created by conventional systems [17, 18]. Moreover, a re-
cent study showed that prematurely born neonates
(mean gestational age [GA] of 29 weeks) with minimal
parenchymal lung disease exerted less effort in spontan-
eous breathing with a Seattle-PAP than a conventional
bubble nCPAP (Fisher & Paykel CPAP or FP-CPAP)
[13]. While the results of preliminary studies are encour-
aging, safety and efficacy data among smaller, more im-
mature preterm infants are lacking. Fig. 1 Schematic showing the primary components of a bubble nCPAP circuit. The differences in conventional and Seattle-PAP bubbler devices
are shown. All other elements of the CPAP circuit are identical between the groups Fig. 1 Schematic showing the primary components of a bubble nCPAP circuit. The differences in conventional and Seattle-PAP bubbler devices
are shown. All other elements of the CPAP circuit are identical between the groups Backes et al. Trials (2019) 20:63 Backes et al. Trials (2019) 20:63 Page 3 of 10 Page 3 of 10 If the Seattle-PAP arm proves superior to conventional
bubble nCPAP, the likely benefits would be large, with
clear applications in both the developed world and in low-
and middle-income countries. However, to recommend
Seattle-PAP over the current standard of care for bubble
nCPAP (FP-CPAP), a large comparative trial is needed. Aims The primary objective of this study is to compare the
rates of respiratory failure from 72 h post-delivery to 32
weeks’ postmenstrual age in neonates born at 220/7 to
296/7 weeks’ GA who receive either Seattle-PAP or con-
ventional bubble nCPAP (FP-CPAP). Background Additionally, since respiratory failure is associated with
higher daily costs related to more intensive monitoring
and personnel requirements [19], Seattle-PAP may result
in lower overall treatment costs if it prevents respiratory
failure. Similarly, since infants failing CPAP stay in hospital
longer than those successfully supported on CPAP, the
total costs should be lower if use of Seattle-PAP is associ-
ated with a lower incidence of CPAP failure than FP-CPAP
[20]. To estimate the cost-effectiveness of Seattle-PAP, a
formal economic evaluation ancillary to the proposed ran-
domized controlled trial (RCT) is necessary [21]. using FP-CPAP as a non-invasive respiratory support
modality. Participating sites did not use Seattle-PAP
prior to study commencement. All participating sites administer an exogenous surfac-
tant if the infant requires tracheal intubation for respira-
tory distress. One NICU (Mt. Carmel St. Ann’s Hospital)
has some experience with less invasive surfactant adminis-
tration, in which infants on bubble nCPAP are given a sur-
factant via thin catheters directly placed into the trachea
while they are breathing spontaneously, thereby avoiding
the need for intubation for surfactant administration [22]. All participating centers have 24-h on-site neonatal staff. The Institutional Review Board (IRB) of NCH ap-
proved the trial (16–00678). The IRB at Mt. Carmel St. Ann’s Hospital granted separate approval for the trial
(170817–2). Of note, prior to study commencement, the
Seattle-PAP device received clearance from the U.S. Food and Drug Administration via the 510(k) pathway
(K131502; 11 October 2013). The present report describes the rationale and design for
an ongoing RCT that aims to compare the effectiveness of
Seattle-PAP versus conventional FP-CPAP in the preven-
tion of respiratory failure of premature neonates born at
220/7–296/7 weeks’ GA. This study protocol was written in
accordance with the Standard Protocol Items: Recommen-
dations for Interventional Trials (SPIRIT) guidelines. The
SPIRIT Checklist has been included in Additional file 1. Inclusion and exclusion criteria
The inclusion criteria are as follows: The inclusion criteria are as follows: 1. parental consent and/or legal guardian consent
given to participate in this research study 2. preterm infants delivered at 22 to 30 weeks of
completed gestation by best obstetrical estimate
3. weight <1500 g at birth Methods/design
Study settings g
2. Infants with known genetic anomalies (e.g., trisomy 21). This is a multicenter non-blinded RCT in premature ne-
onates born at 220/7 to 296/7 weeks’ GA. The trial is on-
going at five hospitals within the Neonatal Research
Network (NRN) of the Nationwide Children’s Hospital
(NCH): NCH Main Campus, Ohio State University
Wexner Medical Center, OhioHealth Riverside Method-
ist Hospital, Mt. Carmel St. Ann’s Hospital, and Ohio-
Health Grant Medical Center. NCH is a Level IV
referral facility covering Central Ohio and parts of
West Virginia and Kentucky. All neonatal intensive
care units (NICUs) in NCH-NRN routinely care for
premature
neonates
with
respiratory
insufficiency, 3. Infants born to mothers who are unable to give
informed consent. 3. Infants born to mothers who are unable to give
informed consent. Hypothesis We are testing the hypothesis that premature neonates
supported by Seattle-PAP will have lower rates of re-
spiratory failure (need for tracheal intubation and mech-
anical ventilation) than will neonates supported with
conventional bubble CPAP (FP-CPAP). g
g
4. able to be weaned from mechanical ventilator
support within 72 h of birth The exclusion criteria are as follows: The exclusion criteria are as follows: 1. Infants with known major congenital cardiac (e.g.,
transposition of the great arteries), pulmonary
(e.g., pulmonary and/or tracheal hypoplasia), or
physiological (e.g., anencephaly, omphalocele,
congenital diaphragmatic hernia) anomalies. For
the purposes of this definition, common preterm
cardiac issues such as patent ductus arteriosus and
patent foramen ovale or atrial septal defect will not
be grounds for exclusion. Recruitment A member of the study team will approach parents of
potentially eligible infants with threatened preterm deliv-
ery between 220/7 and 296/7 weeks’ GA to offer study
participation and to obtain written informed consent. Consent will be obtained by team members who have
been trained in obtaining consent for clinical trials and
who are familiar with the trial protocol. Whenever Backes et al. Trials (2019) 20:63 Backes et al. Trials (2019) 20:63 Page 4 of 10 Page 4 of 10 between the groups is the bubbler, which generates the
pressure for the circuit (Fig. 1). possible, consent will be obtained by someone not dir-
ectly involved in the clinical care of the infant. If it is
not possible to obtain consent during the antenatal
period, then as soon as possible following the birth, a
member of the study team will approach the infant’s par-
ents or legal guardians to obtain consent (up to 72 h
post-delivery). Parents or guardians of infants who are
not yet eligible but are likely to become eligible (e.g., in-
fants requiring mechanical ventilation who are likely to
transition from mechanical ventilation to bubble nCPAP
prior to 72 h) will also be approached. Fig. 2 is the
schedule of enrollment, interventions, and assessments. Recruitment began in March 2017. Blinding Blinding of the allocated treatment is not feasible, as the
mode of respiratory support is apparent to health-care
professionals and families. However, statisticians per-
forming the data analyses will be blinded to group as-
signment. All enrolled neonates will be given clinical
care according to established guidelines, including wean-
ing, extubation, and reintubation [23]. Moreover, the pri-
mary outcome (respiratory failure, as defined below) is
based on an objective set of criteria applied to both
groups. Clinical and respiratory guidelines Enrolled eligible infants are randomly assigned to one of
two treatment arms (Seattle-PAP vs. FP-CPAP) (Fig. 3). The allocation sequence is generated using an online,
computer-generated
randomizer
(https://sealedenvelo-
pe.com), with a block size of 6, stratified by GA (27–296/
7 and 22–27 weeks’ GA). We did not stratify by site, as
the treatment of premature neonates is based on shared
guidelines throughout the NCH-NRN [23]. Multiple ges-
tations (twins or triplets) are assigned to the same treat-
ment arm. Because the duration and type of respiratory support are
critical end points, we have taken a number of steps to
ensure that respiratory support is applied similarly to
both groups: (1) NCH-NRN follows a respiratory algo-
rithm for care (see Additional file 2: Figure S1) and (2)
the thresholds for achieving the primary end point of re-
spiratory failure are clearly defined (see below). In
addition to standardized respiratory support, all aspects
of neonatal management and treatment will be in ac-
cordance with local guidelines [23]. Internal audits to in-
vestigate maintenance and adherence to the guidelines
are conducted on a routine basis. Intervention Treatment failure is defined as any of the following: The allocated treatment is applied immediately after
randomization. Infants whose condition cannot be main-
tained with the assigned method of noninvasive respira-
tory
support
will
be
intubated
and
the
originally
assigned intervention resumed after extubation. In the
NCH-NRN,
bubble nCPAP
is
given
through
nasal
prongs or a mask with an initial pressure of 5 to 6
cm H2O. As described above, the primary difference 1. Tracheal intubation within 72 h for surfactant
administration after initiation of bubble nCPAP and
then not extubated by 72 h. 2. Tracheal intubation of the infant or support with
biphasic CPAP (SiPAP) in the NICU after 72 h and
up to 32 weeks’ GA. As adjudicated by an Study Period
Consent /
Enrollment
Study
Qualification
Randomization
/ Study
Allocation
Data
Collection
Close-Out /
Study
Conclusion
Timepoint
-t
t0
t1
tend-1
tend
Enrollment
Eligibility Screen
X
Informed Consent
X
Study Qualification
X
Allocation
X
CPAP Type
Seattle-PAP
X
Fisher-Paykel
X
Assessments
Demographics
X
Clinical data
X
X
X
X
Primary outcome
X
X
Secondary outcomes
X
X
X
Economic outcomes
X
Fig. 2 Schedule of enrollment, interventions, and assessments during the study Page 5 of 10 Backes et al. Trials Backes et al. Trials (2019) 20:63 et al. Trials (2019) 20:63 Fig. 3 Study flow chart. bn-CPAP bubble nasal continuous positive airway pressure, CGA corrected gestational age Fig. 3 Study flow chart. bn-CPAP bubble nasal continuous positive airway pressure, CGA corrected gestational age independent party, tracheal intubation for non-
respiratory issues after 72 h and up to 32 weeks’ GA
(e.g., surgery for retinopathy of prematurity) will
not be considered a treatment failure. independent party, tracheal intubation for non-
respiratory issues after 72 h and up to 32 weeks’ GA
(e.g., surgery for retinopathy of prematurity) will
not be considered a treatment failure. proportion of patients experiencing respiratory failure in
the control arm (FP-CPAP) will be 50%. Based on pre-
clinical work, we estimate the failure rate in the
Seattle-PAP arm will be 30% [13, 17]. Accounting for
two interim analyses, 200 neonates are needed with a
two-tailed type I error rate of 0.05 and a power of 80%
[25]. Multiples will be randomized as a set to the same
study arm, requiring an inflation of the estimate by 1.12
to allow for the design effect due to clustering [26, 27]. Study conclusion Patient participation in the study will conclude at 36
weeks’ corrected GA or at discharge, whichever comes
first. Outcomes Except
for
data
related
to
the
screening
log,
all
remaining clinical data will be obtained from the elec-
tronic medical records (Additional file 4: Table S2). The primary outcome is treatment failure (as defined
above). We will capture important secondary outcomes,
including short- and longer-term respiratory morbidities
(Additional file 3: Table S1). Routinely available costs of inpatient stay will be
sourced from the hospital costing units. Direct medical
resource utilization will be ascertained through collec-
tion of itemized billing records and UB-04 forms, a uni-
form billing statement recommended by the National
Uniform Billing Committee and utilized for reporting of
hospital expenditures by third-party payers including the
Centers for Medicare and Medicaid Services (CMS). We
will convert hospital-reported charges to costs by apply-
ing the appropriate CMS cost center specific ratio of
costs to charges. Total hospital costs will be the sum
of the product of the number of days in each cost
category and the calculated per diem costs. Physician
professional fees for the initial hospitalization will be
based on CMS reimbursement levels for each day of
stay and non-bundled procedures. With a birth to hospital discharge time horizon, a
cost-effectiveness analysis from the perspective of the
health-care system was planned a priori and will be
conducted alongside the RCT. From a health-care
sector perspective, cost-effectiveness analyses include
formal health sector (medical) costs, such as those
paid by a third party such as government or a private
insurer [24]. Intervention Thus, the calculated sample size is 115 infants per treat-
ment arm. 3. The infant cannot sustain SpO2 of at least 90%,
despite noninvasive respiratory support of up to
and including 8 cm H2O bubble nCPAP, and FiO2
greater than 0.40 for more than 1 h. Sample size Using data from other recent large neonatal randomized
trials with similar populations, including the SUPPORT
trial and the COIN trial [9, 10], as well as contemporary
data
within
the
NCH-NRN,
we
estimate
that
the Page 6 of 10 Page 6 of 10 Page 6 of 10 Backes et al. Trials (2019) 20:63 Backes et al. Trials (2019) 20:63 Backes et al. Trials Data management, processing, monitoring, and security Data management, processing, monitoring, and security
The data generated in this study will be appropriately
documented and checked for validity and accuracy. The
data will be entered into a database, then the data will
be matched and checked for validity and accuracy by a
second person before being endorsed for analysis. A rec-
ord of all discrepancies and resolutions will be kept by
the study coordinator (JLN). Outlier data will be investi-
gated. Data will be primarily managed using REDCap
(Research Electronic Data Capture) [28]. REDCap is a
secure web-based application designed to support data
capture in clinical studies. All data with identifiers will
be stored on firewall-protected secure servers. Study
monitoring visits are performed by the sponsor and their
representatives after the enrollment of 50, 100, 150, and
200 infants, and upon the closure of the study. y
Data and safety monitoring board The projected duration of the study is 3 years, including
2½ years for subject recruitment. We have assembled a data and safety monitoring board
(DSMB) to protect study subjects and monitor the overall
conduct of the trial. Prior to study commencement, the
four-person DSMB agreed to the following activities: (1)
review the protocol and amendments; (2) participate in
the development, finalization, and approval of the DSMB
charter; (3) recommend discontinuation of the trial for
safety reasons; (4) recommend changes to the study proto-
col (amendments) for safety reasons; (5) evaluate emer-
gent safety information, evaluate any risk, and identify any
potential safety concerns for study patients; (6) request
additional data not included in reports, if deemed neces-
sary for effective safety monitoring; (7) communicate
DSMB findings and recommendations (to stop, continue,
or modify the study) to the site principal investigator (PI;
CHB); and (8) provide written minutes following sched-
uled and ad hoc meetings. Statistical analysis
Clinical outcomes
l
ll b Analyses will be performed using Statistical Analysis
Software (SAS) Enterprise Guide version 7.15 (SAS Insti-
tute Inc., Cary, NC, USA) using an as randomized
(intention-to-treat) principle to compare the primary
outcome between treatment arms. Continuous data will
be expressed as means with standard deviations or as
medians with ranges, whereas categorical variables will
be expressed with frequencies and proportions. The pri-
mary outcome, and other binary outcomes occurring in
at least 5% of patients, will be analyzed using Pearson
chi-squared tests. Less common binary outcomes will be
compared using Fisher’s exact tests. Risk differences and Training of clinical staff Comprehensive education and training were undertaken
to ensure technical proficiency and compliance with the
intervention and study protocol at all sites. The site PI
(CHB), study coordinator (JLN), and nursing leadership
on the study (JML and JCB) ran a boot camp at the start
of the study for all participating sites, with mandatory at-
tendance
for
physicians,
respiratory
therapists,
and
nursing staff to accomplish the following: (1) review differ-
ences between the Seattle-PAP and the conventional
FP-CPAP devices; (2) identify areas for device trouble-
shooting; (3) demarcate study goals and objectives; and (4)
review safety protocols, procedures, and guidelines for
clinical and respiratory care. This effort was intended to
optimize reproducibility and consistency. All centers re-
ceived detailed written instruction on study protocol. The
PI and study coordinator were available 24 h each day to
answer any questions or concerns. Adverse events and their relationship to study, sever-
ity, time of experience, expectation, actions taken to re-
solve the event, and final outcome will be recorded. All
serious adverse events (SAEs) will be sent within 48 h to
the DSMB and local IRB. A SAE for this study is any un-
toward medical occurrence that is believed by the inves-
tigators to be causally related to the study intervention
and results in any of the following: a life-threatening
condition (that is, immediate risk of death), severe or
permanent disability, or prolonged hospitalization. SAEs
occurring after a subject is discontinued from the study
will not be reported, unless the investigators feel that the
event may have been caused by the study device. All
SAEs will be followed until a satisfactory resolution is
achieved or until the health-care provider responsible
for the care of the participant deems the event to be
chronic or the patient to be stable. All expected and un-
expected SAEs, whether or not they are attributable to
the study intervention, will be reviewed by the site PI
(CHB) to determine if there is a reasonable suspected
causal relationship to the intervention. If the relationship
is reasonable, SAEs will be reported to DSMB and local
IRB for consideration. Discussion Previous investigators have shown that tracheal intub-
ation and mechanical ventilation are associated with
ventilator-induced lung injury and airway inflammation,
which contribute to long-term adverse respiratory con-
sequences [33, 34]. Walsh et al. reported that the dur-
ation of tracheal intubation and mechanical ventilation
among premature neonates is associated with an in-
creased risk of death or survival with neurocognitive
impairment
[35]. These
observations
have
led
health-care providers to pursue less invasive forms of
respiratory support for premature neonates, including
nCPAP [9, 10]. Study progress The trial began recruiting in March 2017. It is expected
that recruitment for the study will be completed in
2019. Interim analysis Interim analyses were conducted by the DSMB following
the enrollment of 25 and 110 infants, respectively. The
analyses compared the two groups with respect to effi-
cacy, safety, and futility. A Haybittle–Peto stopping
guideline was set at P < 0.001 for each interim analysis. The interim analyses were completed in June 2017 and
January 2018. Based on these analyses, and on safety re-
views conducted to date, the DSMB recommended that
the trial continue without modification. Page 7 of 10 Backes et al. Trials (2019) 20:63 Page 7 of 10 Backes et al. Trials (2019) 20:63 the number needed to treat, along with their 95% New-
combe hybrid score confidence intervals will be calcu-
lated,
and
risk
ratios,
and
their
95%
Koopman
asymptotic score confidence intervals, will also be calcu-
lated. Normally distributed, continuous outcomes will be
compared using Student’s t-test, whereas the Wilcoxon
rank-sum test will be used to compare continuous out-
comes with skewed distributions. from the study data set [30, 31]. For each of the 1000 sam-
ples, we will calculate the mean cost, mean effect, and in-
cremental cost-effectiveness ratio. The results of this
analysis will be presented using an incremental cost-effect-
iveness plot and cost-effectiveness acceptability curves
[32]. Parameter
uncertainty,
for
variables
such
as
price weights, will be estimated using a deterministic
sensitivity analysis, in which the results are generated
again after varying the parameter through its plausible
range [30]. In
additional
analyses,
log
binomial
regression
models will be used to evaluate heterogeneity in the
effect of treatment with Seattle-PAP on the primary
outcome. Treatment effect heterogeneity will be ex-
plored across several clinical characteristics that are
known to be associated with CPAP treatment failure:
GA, birth weight, and exposure to antenatal cortico-
steroids [15]. Treatment effect heterogeneity will be
tested by evaluating the significance of interactions
between the factors of interest and treatment arm in
log
binomial
regression
models
that
include
the
factor, treatment arm, and their interaction. Effect es-
timates within subgroups defined by these factors will
be expressed as risk ratios with maximum likelihood-
based 95% confidence limits. Regression models will
be estimated using generalized estimating equations
to account for the inherent correlation expected with
multiples [27]. Dissemination of results The
results
of
the
trial
will
be
published
in
a
peer-reviewed journal with open access. To facilitate shar-
ing of the clinical research data, the data from the present
study will be made available at the global data sharing en-
terprise Vivli (https://vivli.org/resources). The Vivli plat-
form provides an independent data repository, search
engine, and a secure cloud-based analytics platform. Economic outcomes Recognizing the need for a more efficient bubble nCPAP
system among preterm infants [15], investigators de-
signed and developed an alternative device, termed
Seattle-PAP [12, 13, 17, 18]. In preclinical studies with
juvenile rabbits lavaged to produce pulmonary surfactant
deficiency [12, 13, 17, 18], Seattle-PAP provides oscil-
lations in airway pressures at lower frequencies than
are observed with other devices [17, 18]. The fre-
quencies of airway pressure oscillations generated by
Seattle-PAP are lower than those generated by con-
ventional bubble nCPAP [17, 18]. While conventional
bubble CPAP provides stabilization and distension of
small airways and alveolar spaces, the range of fluctu-
ations in airway pressure provided by Seattle-PAP
may improve recruitment of and ventilation to the
low ventilation-perfusion compartments of the lung
that contribute to hypoxemia [12]. would be feasible. Finally, blinding of the interventions,
which would have been preferable from a study design
perspective, was not feasible, due to the complexity of
building and operating a system that disguised which de-
vice was providing respiratory support to the infant, yet
allowed the clinical care team to monitor effective en-
gagement of the infant with the pressure-generating
system. The results of the present study will inform the
design of larger multicenter trials that investigate
more fully the benefits and risks of bubble nCPAP
in low- and middle-income countries. Since CPAP
failure in developing countries may result in neo-
natal death, potential use in these countries would
be even more attractive [11, 14]. While previous in-
vestigators have described barriers to successful use
of bubble CPAP in such settings (fixation devices are
bulky and cover much of the infant’s face; interfer-
ence with parental interaction and feeding; trauma
to the nasal skin or septum; and need for nursing
vigilance to ensure an adequate seal), others have
shown that bubble nCPAP can be applied effectively
in resource-limited situations [14, 37, 38]. These observations led to a recent study that com-
pared Seattle-PAP and conventional bubble nCPAP
(FP-CPAP) among premature infants with an average
GA of 29 weeks. The authors observed, among 40 neo-
nates, reduced breathing effort with Seattle-PAP than
with FP-CPAP; however, the study was not designed to
detect important clinical differences between the two de-
vices [13]. To recommend Seattle-PAP over the current
standard of care (bubble nCPAP), a larger comparative
trial is needed. Economic outcomes To inform whether it is cost-effective to incorporate
FP-CPAP or Seattle-PAP into the existing health system,
a decision analysis will be constructed based on the pri-
mary outcome and associated hospital costs. Univariate
and probabilistic sensitivity analyses will be conducted
to test the impact of uncertainty in the data. We will
first compare the mean patient-level costs for each treat-
ment arm, without consideration of effectiveness. Be-
cause of the anticipated skewed nature of the cost data,
we will model the logarithm of the mean costs directly,
using a generalized linear model with a logarithmic link
function and gamma distribution [24, 29]. In addition to
treatment assignment, covariates will be entered into
this model to account for any differences in baseline
prognostic indicators (e.g., exposure to antenatal cortico-
steroids) that are evident despite randomization. The
model will take into account clustering among twins. nCPAP is an established alternative to intubation
and mechanical ventilation for premature infants with
respiratory distress [7]. Although meta-analytic data
show that early adoption of nCPAP was associated
with lower risk of BPD than was observed with intub-
ation and ventilation, a high proportion of neonates
are not adequately supported on nCPAP and require
tracheal intubation and mechanical ventilation (CPAP
failure) [8–10]. In fact, the data suggest that 34 to
83% of premature neonates maintained on nCPAP in
the delivery room require tracheal intubation and
mechanical ventilation [8–10]. We will then determine the simultaneous outcome of
cost and effect, or value for money, expressed as the incre-
mental cost-effectiveness ratio, which is calculated as the
difference in mean cost per patient in the Seattle-PAP and
FP-CPAP groups divided by the difference in the mean ef-
fect between the two groups. We will determine the statis-
tical uncertainty in the joint distribution of costs and
effects using nonparametric bootstrapping, in which we
will draw 1000 samples of 220 infants, with replacement, More recently, bubble nCPAP has re-emerged as a po-
tential strategy to address high nCPAP failure rates [13]. Although neonates on bubble nCPAP had lower inci-
dence of respiratory failure (tracheal intubation and Backes et al. Trials (2019) 20:63 Backes et al. Trials (2019) 20:63 Page 8 of 10 Page 8 of 10 mechanical ventilation) than did infants supported on
ventilator-derived CPAP, the optimal delivery of nCPAP
to premature neonates remains unknown [12, 14, 15]. Economic outcomes Thus, the present study is designed to
compare the incidence of respiratory failure over the
first weeks of life between the two groups. While the
present study is not designed to determine differences in
longer-term clinical outcomes (BPD), prespecified sec-
ondary analyses are planned. The
airway
pressure
oscillations
generated with
Seattle-PAP, including the lower frequency of pres-
sure oscillations, are modestly different from those
generated with conventional bubble nCPAP [17, 18]. Although preclinical and small clinical studies con-
ducted to date have not indicated there is a greater
risk of adverse events associated with Seattle-PAP
than with conventional bubble nCPAP [13, 17, 18],
the present trial will provide an opportunity to ob-
serve any unanticipated SAEs. Use of CPAP among preterm infants is associated with
reduced hospital stay [16] and with savings exceeding
$10,000 for every six neonates treated with CPAP [39]. Given the interrelatedness of health, availability of re-
sources, and the constraints on health-care budgets, as
well as the substantial resource utilization by preterm
neonates, improving health-care status is likely to have
important cost implications; thus, we will perform a for-
mal economic evaluation ancillary to the proposed RCT
with the goal of informing the practice of effective and
efficient health care. We acknowledge a number of challenges in the design
of the trial. First, the potential benefits and risks of
Seattle-PAP
versus
conventional
bubble
nCPAP
(FP-CPAP) may not be uniform across infants born at all
GAs. To that end, we determined a priori to perform a
subgroup analysis on infants born after <27 weeks versus
those born after 27 to 30 weeks of gestation. Second, our
recruitment window provides enrollment up to 72 h of
life, irrespective of previous respiratory support (mech-
anical ventilation or CPAP) and likely over a wide range
of acutely impaired lung function. We recognize that
this potential heterogeneity increases the risk of type II
errors [36]. Third, some infants allocated to Seattle-PAP
may
receive
a
brief
period
of
FP-CPAP
prior
to
randomization, which conceivably could affect the inter-
pretation of the results. We attempted to restrict the im-
pact of this by excluding from the trial infants who have
received more than 72 h of FP-CPAP, which we felt to be
the shortest window in which seeking parental consent Potential impact Seattle-PAP is a promising new bubble CPAP delivery
system that may reduce the need for tracheal intub-
ation and mechanical ventilation among preterm neo-
nates. The Seattle-PAP trial will provide relevant and
timely evidence on the efficacy and safety of the
Seattle-PAP device compared to conventional bubble
CPAP
(FP-CPAP)
in
this
vulnerable
subgroup
of
patients. Page 9 of 10 Backes et al. Trials (2019) 20:63 Page 9 of 10 Backes et al. Trials Backes et al. Trials (2019) 20:63 (2019) 20:63 References 1. Malkar MB, Gardner WP, Mandy GT, Stenger MR, Nelin LD, Shepherd EG,
Welty SE. Respiratory severity score on day of life 30 is predictive of
mortality and the length of mechanical ventilation in premature infants
with protracted ventilation. Pediatr Pulmonol. 2015;50:363–9. 1. Malkar MB, Gardner WP, Mandy GT, Stenger MR, Nelin LD, Shepherd EG,
Welty SE. Respiratory severity score on day of life 30 is predictive of
mortality and the length of mechanical ventilation in premature infants
with protracted ventilation. Pediatr Pulmonol. 2015;50:363–9. Author details
1 1Center for Perinatal Research, The Research Institute at Nationwide
Children’s Hospital, 700 Children’s Drive, Columbus, OH 43205, USA. 2Department of Pediatrics, The Ohio State University Wexner Medical Center,
Columbus, OH, USA. 3The Heart Center, Nationwide Children’s Hospital,
Columbus, OH, USA. 4OhioHealth Research Institute, Riverside Methodist
Hospital, Columbus, OH, USA. 5Pediatrix Medical Group of Ohio, Columbus,
OH, USA. 6Mt. Carmel St. Ann’s Hospital, Westerville, OH, USA. 7OhioHealth,
Grant Medical Center, Columbus, OH, USA. 8Center for Surgical Outcomes,
The Research at Nationwide Children’s Hospital, Columbus, OH, USA. 9Division of Epidemiology, The Ohio State University College of Public
Health, Columbus, OH, USA. 10Seattle Children’s Research Institute, Seattle
Children’s Hospital, Seattle, WA, USA. 11Seattle Children’s Neonatology
Program, CHI Franciscan Health, Tacoma, WA, USA. 12SSM Health, Cardinal
Glennon Children’s Hospital, St. Louis University, St. Louis, MO, USA. 13Division of Newborn Medicine Harvard Medical School Boston MA USA Additional file 2: Figure S1. NCH-NRN ventilation weaning protocol. (JPG 1579 kb) Additional file 2: Figure S1. NCH-NRN ventilation weaning protocol. (JPG 1579 kb) Additional file 3: Table S1. Baseline characteristics and secondary
outcomes of the Seattle-PAP trial [40–45]. (DOCX 21 kb) Additional file 3: Table S1. Baseline characteristics and secondary
outcomes of the Seattle-PAP trial [40–45]. (DOCX 21 kb) Additional file 4: Table S2. Additional data collected during the study
period. (DOC 43 kb) Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published
maps and institutional affiliations. Additional file 1: Standard Protocol Items: Recommendations for
Interventional Trials (SPIRIT) 2013 Checklist: recommended items to
address in a clinical trial protocol and related documents. (DOC 120 kb)
Additional file 2: Figure S1. NCH-NRN ventilation weaning protocol. (JPG 1579 kb)
Additional file 3: Table S1. Baseline characteristics and secondary
outcomes of the Seattle-PAP trial [40–45]. (DOCX 21 kb)
Additional file 4: Table S2. Additional data collected during the study
period. (DOC 43 kb) Additional file 1: Standard Protocol Items: Recommendations for
Interventional Trials (SPIRIT) 2013 Checklist: recommended items to
address in a clinical trial protocol and related documents. (DOC 120 kb) Consent for publication
Not applicable. 10. Support Study Group of the Eunice Kennedy Shriver NICHD Neonatal
Research Network, Finer NN, Carlo WA, Walsh MC, Rich W, Gantz MG,
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Physiol (1985). 1987;62:170–4. The study sponsor is the Seattle Children’s Research Institute, Seattle, WA. 5. Hillman NH, Kemp MW, Miura Y, Kallapur SG, Jobe AH. Sustained inflation at
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treated fetal lambs. PLoS One. 2014;9:e113473. Funding
l f
d Trial funding was provided by the Bill & Melinda Gates Foundation
(www.gatesfoundation.org). The foundation played no role in the design of
the study or the preparation of the manuscript. Further, it has no intent to
play any role in the collection, analysis, and interpretation of the data when
the study is complete. Received: 1 November 2018 Accepted: 24 December 2018 Authors’ contributions All authors agree to be held accountable for all aspects of the work reported
herein and to ensure that questions related to the accuracy and integrity of
the work are appropriately investigated and resolved. CHB, NHH, JAZ, LIS,
TNH, and CVS designed or conceived the study and this protocol. CHB, JLN,
JML, JCB, AMN, CMA, BKR, JNC, and LIS acquired, analyzed, or interpreted the
data. All authors drafted, revised, and prepared the manuscript, and gave
final approval for the final submitted manuscript. 6. Subramaniam P, Ho JJ, Davis PG. Prophylactic nasal continuous positive
airway pressure for preventing morbidity and mortality in very preterm
infants. Cochrane Database Syst Rev. 2016;(6). Article No.: CD001243. https://
doi.org/10.1002/14651858.CD001243.pub3. 7. American Academy of Pediatrics: Committee on Fetus and Newborn. Respiratory support in preterm infants at birth. Pediatrics. 2014;133:171–4. 8. Dunn MS, Kaempf J, de Klerk A, de Klerk R, Reilly M, Howard D, Ferrelli K,
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and other hospital staff in the cooperating NCH-NRN hospitals for their efforts. The authors wish to acknowledge Edward Shepherd MD, Thomas Bartman MD,
and the Cure-Me BPD team at NCH, who designed and implemented
the respiratory treatment algorithm for premature neonates. 14Department of Neonatology, Beth Israel Deaconess Medical Center, Boston
MA, USA. 15Minneapolis Heart Institute Foundation, Minneapolis, MN, USA. 14Department of Neonatology, Beth Israel Deaconess Medical Center, Boston
MA, USA. 15Minneapolis Heart Institute Foundation, Minneapolis, MN, USA. y
MA, USA. 15Minneapolis Heart Institute Foundation, Minneapolis, MN, USA. Availability of data and materials The results of the trial will be published in a journal with an open-access
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f The IRB of NCH approved the trial (16–00678). The IRB at Mt. Carmel St. Ann’s
Hospital granted separate approval for the trial (170817–2). The IRB of NCH approved the trial (16–00678). The IRB at Mt. Carmel St. Ann’s
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several regions or countries, including Europe (2303373), Korea (10–1,540,948),
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